@Article{IPB-2503, author = {Gasperini, D. and Howe, G. A. and}, title = {{Phytohormones in a universe of regulatory metabolites: lessons from jasmonate}}, year = {2024}, journal = {Plant Physiol.}, doi = {10.1093/plphys/kiae045}, url = {https://doi.org/10.1093/plphys/kiae045}, abstract = {Small-molecule phytohormones exert control over plant growth, development, and stress responses by coordinating the patterns of gene expression within and between cells. Increasing evidence indicates that currently recognized plant hormones are part of a larger group of regulatory metabolites that have acquired signaling properties during the evolution of land plants. This rich assortment of chemical signals reflects the tremendous diversity of plant secondary metabolism, which offers evolutionary solutions to the daunting challenges of sessility and other unique aspects of plant biology. A major gap in our current understanding of plant regulatory metabolites is the lack of insight into the direct targets of these compounds. Here, we illustrate the blurred distinction between classical phytohormones and other bioactive metabolites by highlighting the major scientific advances that transformed the view of jasmonate from an interesting floral scent to a potent transcriptional regulator. Lessons from jasmonate research generally apply to other phytohormones and thus may help provide a broad understanding of regulatory metabolite–protein interactions. In providing a framework that links small-molecule diversity to transcriptional plasticity, we hope to stimulate future research to explore the evolution, functions, and mechanisms of perception of a broad range of plant regulatory metabolites.} } @Article{IPB-212, author = {Wasternack, C. and Hause, B. and}, title = {{BFP1: One of 700 Arabidopsis F-box proteins mediates degradation of JA oxidases to promote plant immunity}}, year = {2024}, pages = {375-376}, journal = {Mol. Plant}, doi = {10.1016/j.molp.2024.02.008}, url = {https://doi.org/10.1016/j.molp.2024.02.008}, volume = {17}, } @Article{IPB-203, author = {Khalil, S. and Strah, R. and Lodovici, A. and Vojta, P. and Berardinis, F. D. and Ziegler, J. and Novak, M. P. and Zanin, L. and Tomasi, N. and Forneck, A. and Griesser, M. and}, title = {{The activation of iron deficiency responses of grapevine rootstocks is dependent to the availability of the nitrogen forms}}, year = {2024}, pages = {218}, journal = {BMC Plant Biol.}, doi = {10.1186/s12870-024-04906-y}, url = {https://doi.org/10.1186/s12870-024-04906-y}, volume = {24}, abstract = {Background  In viticulture, iron (Fe) chlorosis is a common abiotic stress that impairs plant development and leads to yield and quality losses. Under low availability of the metal, the applied N form (nitrate and ammonium) can play a role in promoting or mitigating Fe deficiency stresses. However, the processes involved are not clear in grapevine. Therefore, the aim of this study was to investigate the response of two grapevine rootstocks to the interaction between N forms and Fe uptake. This process was evaluated in a hydroponic experiment using two ungrafted grapevine rootstocks Fercal (Vitis berlandieri x V. vinifera) tolerant to deficiency induced Fe chlorosis and Couderc 3309 (V. riparia x V. rupestris) susceptible to deficiency induced Fe chlorosis. Results  The results could differentiate Fe deficiency effects, N-forms effects, and rootstock effects. Interveinal chlorosis of young leaves appeared earlier on 3309 C from the second week of treatment with NO3−/NH4+ (1:0)/-Fe, while Fercal leaves showed less severe symptoms after four weeks of treatment, corresponding to decreased chlorophyll concentrations lowered by 75% in 3309 C and 57% in Fercal. Ferric chelate reductase (FCR) activity was by trend enhanced under Fe deficiency in Fercal with both N combinations, whereas 3309 C showed an increase in FCR activity under Fe deficiency only with NO3−/NH4+ (1:1) treatment. With the transcriptome analysis, Gene Ontology (GO) revealed multiple biological processes and molecular functions that were significantly regulated in grapevine rootstocks under Fe-deficient conditions, with more genes regulated in Fercal responses, especially when both forms of N were supplied. Furthermore, the expression of genes involved in the auxin and abscisic acid metabolic pathways was markedly increased by the equal supply of both forms of N under Fe deficiency conditions. In addition, changes in the expression of genes related to Fe uptake, regulation, and transport reflected the different responses of the two grapevine rootstocks to different N forms. Conclusions  Results show a clear contribution of N forms to the response of the two grapevine rootstocks under Fe deficiency, highlighting the importance of providing both N forms (nitrate and ammonium) in an appropriate ratio in order to ease the rootstock responses to Fe deficiency. } } @Article{IPB-195, author = {Clúa, J. and Montpetit, J. and Jimenez-Sandoval, P. and Naumann, C. and Santiago, J. and Poirier, Y. and}, title = {{A CYBDOM protein impacts iron homeostasis and primary root growth under phosphate deficiency in Arabidopsis}}, year = {2024}, pages = {423}, journal = {Nat. Commun.}, doi = {10.1038/s41467-023-43911-x}, url = {https://doi.org/10.1038/s41467-023-43911-x}, volume = {15}, abstract = {Arabidopsis primary root growth response to phosphate (Pi) deficiency is mainly controlled by changes in apoplastic iron (Fe). Upon Pi deficiency, apoplastic Fe deposition in the root apical meristem activates pathways leading to the arrest of meristem maintenance and inhibition of cell elongation. Here, we report that a member of the uncharacterized cytochrome b561 and DOMON domain (CYBDOM) protein family, named CRR, promotes iron reduction in an ascorbate-dependent manner and controls apoplastic iron deposition. Under low Pi, the crr mutant shows an enhanced reduction of primary root growth associated with increased apoplastic Fe in the root meristem and a reduction in meristematic cell division. Conversely, CRR overexpression abolishes apoplastic Fe deposition rendering primary root growth insensitive to low Pi. The crr single mutant and crr hyp1 double mutant, harboring a null allele in another member of the CYDOM family, shows increased tolerance to high-Fe stress upon germination and seedling growth. Conversely, CRR overexpression is associated with increased uptake and translocation of Fe to the shoot and results in plants highly sensitive to Fe excess. Our results identify a ferric reductase implicated in Fe homeostasis and developmental responses to abiotic stress, and reveal a biological role for CYBDOM proteins in plants.} } @Article{IPB-231, author = {Dahiya, P. and Bürstenbinder, K. and}, title = {{The making of a ring: Assembly and regulation of microtubule-associated proteins during preprophase band formation and division plane set-up}}, year = {2023}, pages = {102366}, journal = {Curr. Opin. Plant Biol.}, doi = {10.1016/j.pbi.2023.102366}, url = {https://doi.org/10.1016/j.pbi.2023.102366}, volume = {73}, abstract = {The preprophase band (PPB) is a transient cytokinetic structure that marks the future division plane at the onset of mitosis. The PPB forms a dense cortical ring of mainly microtubules, actin filaments, endoplasmic reticulum, and associated proteins that encircles the nucleus of mitotic cells. After PPB disassembly, the positional information is preserved by the cortical division zone (CDZ). The formation of the PPB and its contribution to timely CDZ set-up involves activities of functionally distinct microtubule-associated proteins (MAPs) that interact physically and genetically to support robust division plane orientation in plants. Recent studies identified two types of plant-specific MAPs as key regulators of PPB formation, the TON1 RECRUITMENT MOTIF (TRM) and IQ67 DOMAIN (IQD) families. Both families share hallmarks of disordered scaffold proteins. Interactions of IQDs and TRMs with multiple binding partners, including the microtubule severing KATANIN1, may provide a molecular framework to coordinate PPB formation, maturation, and disassembly.} } @Article{IPB-226, author = {Bürstenbinder, K. and Schwarzerová, K. and}, title = {{European Plant Cytoskeletal Club meeting: A vital platform for advancing plant cytoskeleton research}}, year = {2023}, pages = {397-399}, journal = {Cytoskeleton}, doi = {10.1002/cm.21780}, url = {https://doi.org/10.1002/cm.21780}, volume = {80}, abstract = {This contribution reports on a meeting of plant cytoskeleton scientists-the European Plant Cytoskeletal Club 2023 conference.} } @Article{IPB-225, author = {Blatt-Janmaat, K. and Neumann, S. and Schmidt, F. and Ziegler, J. and Qu, Y. and Peters, K. and}, title = {{Impact of in vitro phytohormone treatments on the metabolome of the leafy liverwort Radula complanata (L.) Dumort}}, year = {2023}, pages = {17}, journal = {Metabolomics}, doi = {10.1007/s11306-023-01979-y}, url = {https://doi.org/10.1007/s11306-023-01979-y}, volume = {19}, abstract = {Introduction Liverworts are a group of non-vascular plants that possess unique metabolism not found in other plants. Many liverwort metabolites have interesting structural and biochemical characteristics, however the fluctuations of these metabolites in response to stressors is largely unknown. Objectives To investigate the metabolic stress-response of the leafy liverwort Radula complanata. Methods Five phytohormones were applied exogenously to in vitro cultured R. complanata and an untargeted metabolomic analysis was conducted. Compound classification and identification was performed with CANOPUS and SIRIUS while statistical analyses including PCA, ANOVA, and variable selection using BORUTA were conducted to identify metabolic shifts.Results It was found that R. complanata was predominantly composed of carboxylic acids and derivatives, followed by benzene and substituted derivatives, fatty acyls, organooxygen compounds, prenol lipids, and flavonoids. The PCA revealed that samples grouped based on the type of hormone applied, and the variable selection using BORUTA (Random Forest) revealed 71 identified and/or classified features that fluctuated with phytohormone application. The stress-response treatments largely reduced the production of the selected primary metabolites while the growth treatments resulted in increased production of these compounds. 4-(3-Methyl-2-butenyl)-5-phenethylbenzene-1,3-diol was identified as a biomarker for the growth treatments while GDP-hexose was identified as a biomarker for the stress-response treatments. Conclusion Exogenous phytohormone application caused clear metabolic shifts in Radula complanata that deviate from the responses of vascular plants. Further identification of the selected metabolite features can reveal metabolic biomarkers unique to liverworts and provide more insight into liverwort stress responses. } } @Article{IPB-224, author = {Blatt-Janmaat, K. L. and Neumann, S. and Ziegler, J. and Peters, K. and}, title = {{Host tree and geography induce metabolic shifts in the epiphytic liverwort Radula complanata}}, year = {2023}, pages = {571}, journal = {Plants}, doi = {10.3390/plants12030571}, url = {https://doi.org/10.3390/plants12030571}, volume = {12}, abstract = {Bryophytes are prolific producers of unique, specialized metabolites that are not found in other plants. As many of these unique natural products are potentially interesting, for example, pharmacological use, variations in the production regarding ecological or environmental conditions have not often been investigated. Here, we investigate metabolic shifts in the epiphytic Radula complanata L. (Dumort) with regard to different environmental conditions and the type of phorophyte (host tree). Plant material was harvested from three different locations in Sweden, Germany, and Canada and subjected to untargeted liquid chromatography high-resolution mass-spectrometry (UPLC/ESI-QTOF-MS) and data-dependent acquisition (DDA-MS). Using multivariate statistics, variable selection methods, in silico compound identification, and compound classification, a large amount of variation (39%) in the metabolite profiles was attributed to the type of host tree and 25% to differences in environmental conditions. We identified 55 compounds to vary significantly depending on the host tree (36 on the family level) and 23 compounds to characterize R. complanata in different environments. Taken together, we found metabolic shifts mainly in primary metabolites that were associated with the drought response to different humidity levels. The metabolic shifts were highly specific to the host tree, including mostly specialized metabolites suggesting high levels of ecological interaction. As R. complanata is a widely distributed generalist species, we found it to flexibly adapt its metabolome according to different conditions. We found metabolic composition to also mirror the constitution of the habitat, which makes it interesting for conservation measures.} } @Article{IPB-222, author = {Bao, Z. and Guo, Y. and Deng, Y. and Zang, J. and Zhang, J. and Deng, Y. and Ouyang, B. and Qu, X. and Bürstenbinder, K. and Wang, P. and}, title = {{Microtubule-associated protein SlMAP70 interacts with IQ67-domain protein SlIQD21a to regulate fruit shape in tomato}}, year = {2023}, pages = {4266-4283}, journal = {Plant Cell}, doi = {10.1093/plcell/koad231}, url = {https://doi.org/10.1093/plcell/koad231}, volume = {35}, abstract = {Tomato (Solanum lycopersicum) fruit shape is related to microtubule organization and the activity of microtubule-associated proteins (MAPs). However, insights into the mechanism of fruit shape formation from a cell biology perspective remain limited. Analysis of the tissue expression profiles of different microtubule regulators revealed that functionally distinct classes of MAPs, including members of the plant-specific MICROTUBULE-ASSOCIATED PROTEIN 70 (MAP70) and IQ67 DOMAIN (IQD, also named SUN in tomato) families, are differentially expressed during fruit development. SlMAP70-1–3 and SlIQD21a are highly expressed during fruit initiation, which relates to the dramatic microtubule pattern rearrangements throughout this developmental stage of tomato fruits. Transgenic tomato lines overexpressing SlMAP70-1 or SlIQD21a produced elongated fruits with reduced cell circularity and microtubule anisotropy, while their loss-of-function mutants showed the opposite phenotype, harboring flatter fruits. Fruits were further elongated in plants coexpressing both SlMAP70-1 and SlIQD21a. We demonstrated that SlMAP70s and SlIQD21a physically interact and that the elongated fruit phenotype is likely due to microtubule stabilization induced by the SlMAP70–SlIQD21a interaction. Together, our results identify SlMAP70 proteins and SlIQD21a as important regulators of fruit elongation and demonstrate that manipulating microtubule function during early fruit development provides an effective approach to alter fruit shape.} } @Article{IPB-220, author = {Aryal, B. and Xia, J. and Hu, Z. and Stumpe, M. and Tsering, T. and Liu, J. and Huynh, J. and Fukao, Y. and Glöckner, N. and Huang, H.-Y. and Sancho-Andrés, G. and Pakula, K. and Ziegler, J. and Gorzolka, K. and Zwiewka, M. and Nodzynski, T. and Harter, K. and Sánchez-Rodríguez, C. and Jasiński, M. and Rosahl, S. and Geisler, M. M. and}, title = {{An LRR receptor kinase controls ABC transporter substrate preferences during plant growth-defense decisions}}, year = {2023}, pages = {2008-2023}, journal = {Curr. Biol.}, doi = {10.1016/j.cub.2023.04.029}, url = {https://doi.org/10.1016/j.cub.2023.04.029}, volume = {33}, abstract = {The exporter of the auxin precursor indole-3-butyric acid (IBA), ABCG36/PDR8/PEN3, from the model plant Arabidopsis has recently been proposed to also function in the transport of the phytoalexin camalexin. Based on these bonafide substrates, it has been suggested that ABCG36 functions at the interface between growth and defense. Here, we provide evidence that ABCG36 catalyzes the direct, ATP-dependent export of camalexin across the plasma membrane. We identify the leucine-rich repeat receptor kinase, QIAN SHOU KINASE1 (QSK1), as a functional kinase that physically interacts with and phosphorylates ABCG36. Phosphorylation of ABCG36 by QSK1 unilaterally represses IBA export, allowing camalexin export by ABCG36 conferring pathogen resistance. As a consequence, phospho-dead mutants of ABCG36, as well as qsk1 and abcg36 alleles, are hypersensitive to infection with the root pathogen Fusarium oxysporum, caused by elevated fungal progression. Our findings indicate a direct regulatory circuit between a receptor kinase and an ABC transporter that functions to control transporter substrate preference during plant growth and defense balance decisions.} } @Article{IPB-217, author = {Abukhalaf, M. and Proksch, C. and Thieme, D. and Ziegler, J. and Hoehenwarter, W. and}, title = {{Changing turn-over rates regulate abundance of tryptophan, GS biosynthesis, IAA transport and photosynthesis proteins in Arabidopsis growth defense transitions}}, year = {2023}, pages = {249}, journal = {BMC Biol.}, doi = {10.1186/s12915-023-01739-3}, url = {https://doi.org/10.1186/s12915-023-01739-3}, volume = {21}, abstract = {Background Shifts in dynamic equilibria of the abundance of cellular molecules in plant-pathogen interactions need further exploration. We induced PTI in optimally growing Arabidopsis thaliana seedlings for 16 h, returning them to growth conditions for another 16 h. Methods Turn-over and abundance of 99 flg22 responding proteins were measured chronologically using a stable heavy nitrogen isotope partial labeling strategy and targeted liquid chromatography coupled to mass spectrometry (PRM LC–MS). These experiments were complemented by measurements of mRNA and phytohormone levels. Results Changes in synthesis and degradation rate constants (Ks and Kd) regulated tryptophane and glucosinolate, IAA transport, and photosynthesis-associated protein (PAP) homeostasis in growth/PTI transitions independently of mRNA levels. Ks values increased after elicitation while protein and mRNA levels became uncorrelated. mRNA returned to pre-elicitation levels, yet protein abundance remained at PTI levels even 16 h after media exchange, indicating protein levels were robust and unresponsive to transition back to growth. The abundance of 23 PAPs including FERREDOXIN-NADP( +)-OXIDOREDUCTASE (FNR1) decreased 16 h after PAMP exposure, their depletion was nearly abolished in the myc234 mutant. FNR1 Kd increased as mRNA levels decreased early in PTI, its Ks decreased in prolonged PTI. FNR1 Kd was lower in myc234, mRNA levels decreased as in wild type. Conclusions Protein Kd and Ks values change in response to flg22 exposure and constitute an additional layer of protein abundance regulation in growth defense transitions next to changes in mRNA levels. Our results suggest photosystem remodeling in PTI to direct electron flow away from the photosynthetic carbon reaction towards ROS production as an active defense mechanism controlled post-transcriptionally and by MYC2 and homologs. Target proteins accumulated later and PAP and auxin/IAA depletion was repressed in myc234 indicating a positive effect of the transcription factors in the establishment of PTI. } } @Article{IPB-290, author = {Richter, C. and Grafahrend-Belau, E. and Ziegler, J. and Raorane, M. L. and Junker, B. H. and}, title = {{Improved 13C metabolic flux analysis in Escherichia coli metabolism: application of a high-resolution MS (GC–EI–QTOF) for comprehensive assessment of MS/MS fragments}}, year = {2023}, pages = {kuad039}, journal = {Journal of Industrial Microbiology and Biotechnology}, doi = {10.1093/jimb/kuad039}, url = {https://doi.org/10.1093/jimb/kuad039}, volume = {50}, abstract = {Gas chromatography–tandem mass spectrometry with electron ionization (GC–EI–MS/MS) provides rich information on stable-isotope labeling for 13C-metabolic flux analysis (13C-MFA). To pave the way for the routine application of tandem MS data for metabolic flux quantification, we aimed to compile a comprehensive library of GC–EI–MS/MS fragments of tert-butyldimethylsilyl (TBDMS) derivatized proteinogenic amino acids. First, we established an analytical workflow that combines high-resolution gas chromatography-quadrupole time-of-flight mass spectrometry and fully 13C-labeled biomass to identify and structurally elucidate tandem MS amino acid fragments. Application of the high-mass accuracy MS procedure resulted into the identification of 129 validated precursor–product ion pairs of 13 amino acids with 30 fragments being accepted for 13C-MFA. The practical benefit of the novel tandem MS data was demonstrated by a proof–of–concept study, which confirmed the importance of the compiled library for high-resolution 13C-MFA. } } @Article{IPB-278, author = {Picchianti, L. and Sanchez de Medina Hernandez, V. and Zhan, N. and Irwin, N. A. and Groh, R. and Stephani, M. and Hornegger, H. and Beveridge, R. and Sawa‐Makarska, J. and Lendl, T. and Grujic, N. and Naumann, C. and Martens, S. and Richards, T. A. and Clausen, T. and Ramundo, S. and Karagöz, G. E. and Dagdas, Y. and}, title = {{Shuffled ATG8 interacting motifs form an ancestral bridge between UFMylation and autophagy}}, year = {2023}, pages = {e112053}, journal = {EMBO J.}, doi = {10.15252/embj.2022112053}, url = {https://doi.org/10.15252/embj.2022112053}, volume = {42}, abstract = {UFMylation involves the covalent modification of substrate proteins with UFM1 (Ubiquitin-fold modifier 1) and is important for maintaining ER homeostasis. Stalled translation triggers the UFMylation of ER-bound ribosomes and activates C53-mediated autophagy to clear toxic polypeptides. C53 contains noncanonical shuffled ATG8-interacting motifs (sAIMs) that are essential for ATG8 interaction and autophagy initiation. However, the mechanistic basis of sAIM-mediated ATG8 interaction remains unknown. Here, we show that C53 and sAIMs are conserved across eukaryotes but secondarily lost in fungi and various algal lineages. Biochemical assays showed that the unicellular alga Chlamydomonas reinhardtii has a functional UFMylation pathway, refuting the assumption that UFMylation is linked to multicellularity. Comparative structural analyses revealed that both UFM1 and ATG8 bind sAIMs in C53, but in a distinct way. Conversion of sAIMs into canonical AIMs impaired binding of C53 to UFM1, while strengthening ATG8 binding. Increased ATG8 binding led to the autoactivation of the C53 pathway and sensitization of Arabidopsis thaliana to ER stress. Altogether, our findings reveal an ancestral role of sAIMs in UFMylation-dependent fine-tuning of C53-mediated autophagy activation.} } @Article{IPB-274, author = {Paponov, M. and Ziegler, J. and Paponov, I. A. and}, title = {{Light exposure of roots in aeroponics enhances the accumulation of phytochemicals in aboveground parts of the medicinal plants Artemisia annua and Hypericum perforatum}}, year = {2023}, pages = {1079656}, journal = {Front. Plant Sci.}, doi = {10.3389/fpls.2023.1079656}, url = {https://doi.org/10.3389/fpls.2023.1079656}, volume = {14}, abstract = {Light acts as a trigger to enhance the accumulation of secondary compounds in the aboveground part of plants; however, whether a similar triggering effect occurs in roots is unclear. Using an aeroponic setup, we investigated the effect of long-term exposure of roots to LED lighting of different wavelengths on the growth and phytochemical composition of two high-value medicinal plants, Artemisia annua and Hypericum perforatum. In A. annua, root exposure to white, blue, and red light enhanced the accumulation of artemisinin in the shoots by 2.3-, 2.5-, and 1.9-fold, respectively. In H. perforatum, root exposure to white, blue, red, and green light enhanced the accumulation of coumaroylquinic acid in leaves by 89, 65, 84, and 74%, respectively. Root lighting also increased flavonol concentrations. In contrast to its effects in the shoots, root illumination did not change phytochemical composition in the roots or root exudates. Thus, root illumination induces a systemic response, resulting in modulation of the phytochemical composition in distal tissues remote from the light exposure site.} } @Article{IPB-273, author = {Paponov, M. and Flate, J. and Ziegler, J. and Lillo, C. and Paponov, I. A. and}, title = {{Heterogeneous nutrient supply modulates root exudation and accumulation of medicinally valuable compounds in Artemisia annua and Hypericum perforatum}}, year = {2023}, pages = {1174151}, journal = {Front. Plant Sci.}, doi = {10.3389/fpls.2023.1174151}, url = {https://doi.org/10.3389/fpls.2023.1174151}, volume = {14}, abstract = {Plants have evolved complex mechanisms to adapt to nutrient-deficient environments, including stimulating lateral root proliferation into local soil patches with high nutrient content in response to heterogeneous nutrient distribution. Despite the widespread occurrence of this phenomenon in soil, the effect of heterogeneous nutrient distribution on the accumulation of secondary compounds in plant biomass and their exudation by roots remains largely unknown. This study aims to fill this critical knowledge gap by investigating how deficiency and unequal distributions of nitrogen (N), phosphorus (P), and iron (Fe) affect plant growth and accumulation of the antimalarial drug artemisinin (AN) in leaves and roots of Artemisia annua, as well as AN exudation by roots. Heterogeneous N and P supplies strongly increased root exudation of AN in half of a split-root system exposed to nutrient deficiency. By contrast, exposure to a homogeneous nitrate and phosphate deficiency did not modulate root exudation of AN. This indicates that a combination of local and systemic signals, reflecting low and high nutritional statuses, respectively, were required to enhance AN exudation. This exudation response was independent of the regulation of root hair formation, which was predominantly modulated by the local signal. In contrast to the heterogeneous supply of N and P, heterogeneous Fe supply did not modulate AN root exudation but increased AN accumulation in locally Fe-deficient roots. No modulation of nutrient supply significantly changed the accumulation of AN in A. annua leaves. The impact of a heterogeneous nitrate supply on growth and phytochemical composition was also investigated in Hypericum perforatum plants. Unlike in A. annue, the uneven N supply did not significantly influence the exudation of secondary compounds in the roots of H. perforatum. However, it did enhance the accumulation of several biologically active compounds, such as hypericin, catechin, and rutin isomers, in the leaves of H. perforatum. We propose that the capacity of plants to induce the accumulation and/or differential exudation of secondary compounds under heterogeneous nutrient supply is both species- and compound-specific. The ability to differentially exude AN may contribute to A. annua’s adaptation to nutrient disturbances and modulate allelopathic and symbiotic interactions in the rhizosphere.} } @Article{IPB-265, author = {Nietzschmann, L. and Smolka, U. and Perino, E. H. B. and Gorzolka, K. and Stamm, G. and Marillonnet, S. and Bürstenbinder, K. and Rosahl, S. and}, title = {{The secreted PAMP-induced peptide StPIP1\_1 activates immune responses in potato}}, year = {2023}, pages = {20534}, journal = {Sci. Rep.}, doi = {10.1038/s41598-023-47648-x}, url = {https://doi.org/10.1038/s41598-023-47648-x}, volume = {13}, abstract = {Treatment of potato plants with the pathogen-associated molecular pattern Pep-13 leads to the activation of more than 1200 genes. One of these, StPIP1\_1, encodes a protein of 76 amino acids with sequence homology to PAMP-induced secreted peptides (PIPs) from Arabidopsis thaliana. Expression of StPIP1\_1 is also induced in response to infection with Phytophthora infestans, the causal agent of late blight disease. Apoplastic localization of StPIP1\_1-mCherry fusion proteins is dependent on the presence of the predicted signal peptide. A synthetic peptide corresponding to the last 13 amino acids of StPIP1\_1 elicits the expression of the StPIP1\_1 gene itself, as well as that of pathogenesis related genes. The oxidative burst induced by exogenously applied StPIP1\_1 peptide in potato leaf disks is dependent on functional StSERK3A/B, suggesting that StPIP1\_1 perception occurs via a receptor complex involving the co-receptor StSERK3A/B. Moreover, StPIP1\_1 induces expression of FRK1 in Arabidopsis in an RLK7-dependent manner. Expression of an RLK from potato with high sequence homology to AtRLK7 is induced by StPIP1\_1, by Pep-13 and in response to infection with P. infestans. These observations are consistent with the hypothesis that, upon secretion, StPIP1\_1 acts as an endogenous peptide required for amplification of the defense response.} } @Article{IPB-264, author = {Montpetit, J. and Clúa, J. and Hsieh, Y.-F. and Vogiatzaki, E. and Müller, J. and Abel, S. and Strasser, R. and Poirier, Y. and}, title = {{Endoplasmic reticulum calnexins participate in the primary root growth response to phosphate deficiency}}, year = {2023}, pages = {1719-1733}, journal = {Plant Physiol.}, doi = {10.1093/plphys/kiac595}, url = {https://doi.org/10.1093/plphys/kiac595}, volume = {191}, abstract = {Accumulation of incompletely folded proteins in the endoplasmic reticulum (ER) leads to ER stress, activates ER protein degradation pathways, and upregulates genes involved in protein folding. This process is known as the unfolded protein response (UPR). The role of ER protein folding in plant responses to nutrient deficiencies is unclear. We analyzed Arabidopsis (Arabidopsis thaliana) mutants affected in ER protein quality control and established that both CALNEXIN (CNX) genes function in the primary root’s response to phosphate (Pi) deficiency. CNX1 and CNX2 are homologous ER lectins promoting protein folding of N-glycosylated proteins via the recognition of the GlcMan9GlcNAc2 glycan. Growth of cnx1-1 and cnx2-2 single mutants was similar to that of the wild type under high and low Pi conditions, but the cnx1-1 cnx2-2 double mutant showed decreased primary root growth under low Pi conditions due to reduced meristematic cell division. This phenotype was specific to Pi deficiency; the double mutant responded normally to osmotic and salt stress. Expression of CNX2 mutated in amino acids involved in binding the GlcMan9GlcNAc2 glycan failed to complement the cnx1-1 cnx2-2 mutant. The root growth phenotype was Fe dependent and was associated with root apoplastic Fe accumulation. Two genes involved in Fe-dependent inhibition of primary root growth under Pi deficiency, the ferroxidase LOW PHOSPHATE 1 (LPR1) and P5-type ATPase PLEIOTROPIC DRUG RESISTANCE 2 (PDR2) were epistatic to CNX1/CNX2. Overexpressing PDR2 failed to complement the cnx1-1 cnx2-2 root phenotype. The cnx1-1 cnx2-2 mutant showed no evidence of UPR activation, indicating a limited effect on ER protein folding. CNX might process a set of N-glycosylated proteins specifically involved in the response to Pi deficiency.} } @Article{IPB-263, author = {Mik, V. and Pospíšil, T. and Brunoni, F. and Grúz, J. and Nožková, V. and Wasternack, C. and Miersch, O. and Strnad, M. and Floková, K. and Novák, O. and Široká, J. and}, title = {{Synthetic and analytical routes to the L-amino acid conjugates of cis-OPDA and their identification and quantification in plants}}, year = {2023}, pages = {113855}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2023.113855}, url = {https://doi.org/10.1016/j.phytochem.2023.113855}, volume = {215}, abstract = {Cis-(+)-12-oxophytodienoic acid (cis-(+)-OPDA) is a bioactive jasmonate, a precursor of jasmonic acid, which also displays signaling activity on its own. Modulation of cis-(+)-OPDA actions may be carried out via biotransformation leading to metabolites of various functions. This work introduces a methodology for the synthesis of racemic cis-OPDA conjugates with amino acids (OPDA-aa) and their deuterium-labeled analogs, which enables the unambiguous identification and accurate quantification of these compounds in plants. We have developed a highly sensitive liquid chromatography-tandem mass spectrometry-based method for the reliable determination of seven OPDA-aa (OPDA-Alanine, OPDA-Aspartate, OPDA-Glutamate, OPDA-Glycine, OPDA-Isoleucine, OPDA-Phenylalanine, and OPDA-Valine) from minute amount of plant material. The extraction from 10 mg of fresh plant tissue by 10% aqueous methanol followed by single-step sample clean-up on hydrophilic–lipophilic balanced columns prior to final analysis was optimized. The method was validated in terms of accuracy and precision, and the method parameters such as process efficiency, recovery and matrix effects were evaluated. In mechanically wounded 30-day-old Arabidopsis thaliana leaves, five endogenous (+)-OPDA-aa were identified and their endogenous levels were estimated. The time-course accumulation revealed a peak 60 min after the wounding, roughly corresponding to the accumulation of cis-(+)-OPDA. Our synthetic and analytical methodologies will support studies on cis-(+)-OPDA conjugation with amino acids and research into the biological significance of these metabolites in plants.} } @Article{IPB-261, author = {Meena, S. K. and Heidecker, M. and Engelmann, S. and Jaber, A. and de Vries, T. and Triller, S. and Baumann‐Kaschig, K. and Abel, S. and Behrens, S. and Gago-Zachert, S. and}, title = {{Altered expression levels of long noncoding natural antisense transcripts overlapping the UGT73C6 gene affect rosette size in Arabidopsis thaliana}}, year = {2023}, pages = {460-477}, journal = {Plant J.}, doi = {10.1111/tpj.16058}, url = {https://doi.org/10.1111/tpj.16058}, volume = {113}, abstract = {Natural antisense long noncoding RNAs (lncNATs) are involved in the regulation of gene expression in plants, modulating different relevant developmental processes and responses to various stimuli. We have identified and characterized two lncNATs (NAT1UGT73C6 and NAT2UGT73C6, collectively NATsUGT73C6) from Arabidopsis thaliana that are transcribed from gene fully overlapping UGT73C6, a member of the UGT73C subfamily of genes encoding UDP-glycosyltransferases (UGTs). Expression of both NATsUGT73C6 is developmentally controlled and occurs independently of the transcription of UGT73C6 in cis. Downregulation of NATsUGT73C6 levels through artificial microRNAs results in a reduction of the rosette area, while constitutive overexpression of NAT1UGT73C6 or NAT2UGT73C6 leads to the opposite phenotype, an increase in rosette size. This activity of NATsUGT73C6 relies on its RNA sequence, and, although modulation of UGT73C6 in cis cannot be excluded, the observed phenotypes are not a consequence of the regulation of UGT73C6 in trans. The NATsUGT73C6 levels were shown to affect cell proliferation and thus individual leaf size. Consistent with this concept, our data suggest that the NATsUGT73C6 influence the expression levels of key transcription factors involved in regulating leaf growth by modulating cell proliferation. These findings thus reveal an additional regulatory layer on the process of leaf growth.} } @Article{IPB-241, author = {Haq, I. U. and Krukiewicz, K. and Tayyab, H. and Khan, I. and Khan, M. and Yahya, G. and Cavalu, S. and}, title = {{Molecular understanding of ACE-2 and HLA-conferred differential susceptibility to COVID-19: Host-directed insights opening new windows in COVID-19 therapeutics}}, year = {2023}, pages = {2645}, journal = {Journal of Clinical Medicine}, doi = {10.3390/jcm12072645}, url = {https://doi.org/10.3390/jcm12072645}, volume = {12}, abstract = {The genetic variants of HLAs (human leukocyte antigens) play a crucial role in the virus–host interaction and pathology of COVID-19. The genetic variants of HLAs not only influence T cell immune responses but also B cell immune responses by presenting a variety of peptide fragments of invading pathogens. Peptide cocktail vaccines produced by using various conserved HLA-A2 epitopes provoke substantial specific CD8+ T cell responses in experimental animals. The HLA profiles vary among individuals and trigger different T cell-mediated immune responses in COVID-19 infections. Those with HLA-C*01 and HLA-B*44 are highly susceptible to the disease. However, HLA-A*02:01, HLA-DR*03:01, and HLA-Cw*15:02 alleles show resistance to SARS infection. Understanding the genetic association of HLA with COVID-19 susceptibility and severity is important because it can help in studying the transmission of COVID-19 and its physiopathogenesis. The HLA-C*01 and B*44 allele pathways can be studied to gain insight into disease transmission and physiopathogenesis. Therefore, integrating HLA testing is suggested in the ongoing pandemic, which will help in the rapid identification of highly susceptible populations worldwide and possibly acclimate vaccine development. Therefore, understanding the correlation between HLA and SARS-CoV-2 is critical in opening new insights into COVID-19 therapeutics, based on previous studies conducted.} } @Article{IPB-408, author = {Yang, B. and Stamm, G. and Bürstenbinder, K. and Voiniciuc, C. and}, title = {{Microtubule‐associated IQD9 orchestrates cellulose patterning in seed mucilage}}, year = {2022}, pages = {1096-1110}, journal = {New Phytol.}, doi = {10.1111/nph.18188}, url = {https://doi.org/10.1111/nph.18188}, volume = {235}, abstract = {Arabidopsis seeds release large capsules of mucilaginous polysaccharides, which are shaped by an intricate network of cellulosic microfibrils. Cellulose synthase complexes are guided by the microtubule cytoskeleton, but it is unclear which proteins mediate this process in the seed coat epidermis. Using reverse genetics, we identified IQ67 DOMAIN 9 (IQD9) and KINESIN LIGHT CHAIN-RELATED 1 (KLCR1) as two highly expressed genes during seed development and comprehensively characterized their roles in cell wall polysaccharide biosynthesis. Mutations in IQD9 as well as in KLCR1 lead to compact mucilage capsules with aberrant cellulose distribution, which can be rescued by transgene complementation. IQD9 physically interacts with KLCR1 and localizes to cortical MTs to maintain their organization in SCE cells. IQD9 as well as a previously identified TONNEAU1 (TON1) RECRUITING MOTIF 4 (TRM4) protein act to maintain cellulose synthase velocity. Our results demonstrate that IQD9, KLCR1 and TRM4 are MT-associated proteins that are required for seed mucilage architecture. This study provides the first direct evidence that members of the IQD, KLCR and TRM families have overlapping roles in cell wall biosynthesis. Therefore, SCE cells provide an attractive system to further decipher the complex genetic regulation of polarized cellulose deposition.} } @Article{IPB-382, author = {Prasad, A. and Breithaupt, C. and Nguyen, D.-A. and Lilie, H. and Ziegler, J. and Stubbs, M. T. and}, title = {{Mechanism of chorismate dehydratase MqnA, the first enzyme of the futalosine pathway, proceeds via substrate-assisted catalysis}}, year = {2022}, pages = {102601}, journal = {J. Biol. Chem.}, doi = {10.1016/j.jbc.2022.102601}, url = {https://doi.org/10.1016/j.jbc.2022.102601}, volume = {298}, abstract = {MqnA, the only chorismate dehydratase known so far, catalyzes the initial step in the biosynthesis of menaquinone via the futalosine pathway. Details of the MqnA reaction mechanism remain unclear. Here, we present crystal structures of Streptomyces coelicolor MqnA and its active site mutants in complex with chorismate and the product 3-enolpyruvyl-benzoate, produced during heterologous expression in Escherichia coli. Together with activity studies, our data are in line with dehydration proceeding via substrate assisted catalysis, with the enol pyruvyl group of chorismate acting as catalytic base. Surprisingly, structures of the mutant Asn17Asp with copurified ligand suggest that the enzyme converts to a hydrolase by serendipitous positioning of the carboxyl group. All complex structures presented here exhibit a closed Venus flytrap fold, with the enzyme exploiting the characteristic ligand binding properties of the fold for specific substrate binding and catalysis. The conformational rearrangements that facilitate complete burial of substrate/product, with accompanying topological changes to the enzyme surface, could foster substrate channeling within the biosynthetic pathway.} } @Article{IPB-376, author = {Naumann, C. and Heisters, M. and Brandt, W. and Janitza, P. and Alfs, C. and Tang, N. and Toto Nienguesso, A. and Ziegler, J. and Imre, R. and Mechtler, K. and Dagdas, Y. and Hoehenwarter, W. and Sawers, G. and Quint, M. and Abel, S. and}, title = {{Bacterial-type ferroxidase tunes iron-dependent phosphate sensing during Arabidopsis root development}}, year = {2022}, pages = {2189-2205}, journal = {Curr. Biol.}, doi = {10.1016/j.cub.2022.04.005}, url = {https://doi.org/10.1016/j.cub.2022.04.005}, volume = {32}, abstract = {Access to inorganic phosphate (Pi), a principal intermediate of energy and nucleotide metabolism, profoundly affects cellular activities and plant performance. In most soils, antagonistic Pi-metal interactions restrict Pi bioavailability, which guides local root development to maximize Pi interception. Growing root tips scout the essential but immobile mineral nutrient; however, the mechanisms monitoring external Pi sta-tus are unknown. Here, we show that Arabidopsis LOW PHOSPHATE ROOT 1 (LPR1), one key determinant of Fe-dependent Pi sensing in root meristems, encodes a novel ferroxidase of high substrate specificity and affinity (apparent KM ∼2 μmM Fe2+). LPR1 typifies an ancient, Fe-oxidizing multicopper protein family that evolved early upon bacterial land colonization. The ancestor of streptophyte algae and embryophytes (land plants) acquired LPR1-type ferroxidase from soil bacteria via horizontal gene transfer, a hypothesis supported by phylogenomics, homology modeling, and biochemistry. Our molecular and kinetic data on LPR1 regulation indicate that Pi-dependent Fe substrate availability determines LPR1 activity and function. Guided by the metabolic lifestyle of extant sister bacterial genera, we propose that Arabidopsis LPR1 monitors subtle concentration differentials of external Fe availability as a Pi-dependent cue to adjust root meristem maintenance via Fe redox signaling and cell wall modification. We further hypothesize that the acquisition of bacterial LPR1-type ferroxidase by embryophyte progenitors facilitated the evolution of local Pi sensing and acquisition during plant terrestrialization.} } @Article{IPB-404, author = {Wasternack, C. and}, title = {{Deciphering the oxylipin signatures of necrotrophic infection in plants. A commentary on: Differential modulation of the lipoxygenase cascade during typical and latent Pectobacterium atrosepticum infections}}, year = {2022}, pages = {i-iii}, journal = {Ann. Bot.}, doi = {10.1093/aob/mcab142}, url = {https://doi.org/10.1093/aob/mcab142}, volume = {129}, } @Article{IPB-397, author = {Terrile, M. C. and Tebez, N. M. and Colman, S. L. and Mateos, J. L. and Morato-López, E. and Sánchez-López, N. and Izquierdo-Álvarez, A. and Marina, A. and Calderón Villalobos, L. I. A. and Estelle, M. and Martínez-Ruiz, A. and Fiol, D. F. and Casalongué, C. A. and Iglesias, M. J. and}, title = {{S-Nitrosation of E3 ubiquitin ligase complex components regulates hormonal signalings in Arabidopsis}}, year = {2022}, pages = {794582}, journal = {Front. Plant Sci.}, doi = {10.3389/fpls.2021.794582}, url = {https://doi.org/10.3389/fpls.2021.794582}, volume = {12}, abstract = {E3 ubiquitin ligases mediate the last step of the ubiquitination pathway in the ubiquitin-proteasome system (UPS). By targeting transcriptional regulators for their turnover, E3s play a crucial role in every aspect of plant biology. In plants, SKP1/CULLIN1/F-BOX PROTEIN (SCF)-type E3 ubiquitin ligases are essential for the perception and signaling of several key hormones including auxins and jasmonates (JAs). F-box proteins, TRANSPORT INHIBITOR RESPONSE 1 (TIR1) and CORONATINE INSENSITIVE 1 (COI1), bind directly transcriptional repressors AUXIN/INDOLE-3-ACETIC ACID (AUX/IAA) and JASMONATE ZIM-DOMAIN (JAZ) in auxin- and JAs-depending manner, respectively, which permits the perception of the hormones and transcriptional activation of signaling pathways. Redox modification of proteins mainly by S-nitrosation of cysteines (Cys) residues via nitric oxide (NO) has emerged as a valued regulatory mechanism in physiological processes requiring its rapid and versatile integration. Previously, we demonstrated that TIR1 and Arabidopsis thaliana SKP1 (ASK1) are targets of S-nitrosation, and these NO-dependent posttranslational modifications enhance protein-protein interactions and positively regulate SCFTIR1 complex assembly and expression of auxin response genes. In this work, we confirmed S-nitrosation of Cys140 in TIR1, which was associated in planta to auxin-dependent developmental and stress-associated responses. In addition, we provide evidence on the modulation of the SCFCOI1 complex by different S-nitrosation events. We demonstrated that S-nitrosation of ASK1 Cys118 enhanced ASK1-COI1 protein-protein interaction. Overexpression of non-nitrosable ask1 mutant protein impaired the activation of JA-responsive genes mediated by SCFCOI1 illustrating the functional relevance of this redox-mediated regulation in planta. In silico analysis positions COI1 as a promising S-nitrosation target, and demonstrated that plants treated with methyl JA (MeJA) or S-nitrosocysteine (NO-Cys, S-nitrosation agent) develop shared responses at a genome-wide level. The regulation of SCF components involved in hormonal perception by S-nitrosation may represent a key strategy to determine the precise time and site-dependent activation of each hormonal signaling pathway and highlights NO as a pivotal molecular player in these scenarios.} } @Article{IPB-396, author = {Tang, N. and Naumann, C. and}, title = {{Native protein purification of ferroxidase LPR1 from leaf extracts of a transgenic Arabidopsis thaliana line}}, year = {2022}, pages = {101733}, journal = {STAR Protocols}, doi = {10.1016/j.xpro.2022.101733}, url = {https://doi.org/10.1016/j.xpro.2022.101733}, volume = {3}, abstract = {(LOW PHOSPHATE ROOT 1), a bacterial-type plant ferroxidase, is crucial for local root phosphate (Pi) sensing. Here, we present a detailed protocol for native (tag-free) protein purification of LPR1 from leaf extracts by differential ammonium sulfate precipitation, size exclusion, and cation exchange chromatography of a transgenic Arabidopsis thaliana line overexpressing LPR1. We outline steps for LPR1 purification tracking via immune blot analysis and ferroxidase activity assay. The protocol yields highly pure and active LPR1 protein for biochemical analysis. For complete details on the use and execution of this protocol, please refer to Naumann et al. (2022) } } @Article{IPB-521, author = {Vaddepalli, P. and de Zeeuw, T. and Strauss, S. and Bürstenbinder, K. and Liao, C.-Y. and Ramalho, J. J. and Smith, R. S. and Weijers, D. and}, title = {{Auxin-dependent control of cytoskeleton and cell shape regulates division orientation in the Arabidopsis embryo}}, year = {2021}, pages = {4946-4955}, journal = {Curr. Biol.}, doi = {10.1016/j.cub.2021.09.019}, url = {https://doi.org/10.1016/j.cub.2021.09.019}, volume = {31}, abstract = {Premitotic control of cell division orientation is critical for plant development, as cell walls prevent extensive cell remodeling or migration. While many divisions are proliferative and add cells to existing tissues, some divisions are formative and generate new tissue layers or growth axes. Such formative divisions are often asymmetric in nature, producing daughters with different fates. We have previously shown that, in the Arabidopsis thaliana embryo, developmental asymmetry is correlated with geometric asymmetry, creating daughter cells of unequal volume. Such divisions are generated by division planes that deviate from a default “minimal surface area” rule. Inhibition of auxin response leads to reversal to this default, yet the mechanisms underlying division plane choice in the embryo have been unclear. Here, we show that auxin-dependent division plane control involves alterations in cell geometry, but not in cell polarity axis or nuclear position. Through transcriptome profiling, we find that auxin regulates genes controlling cell wall and cytoskeleton properties. We confirm the involvement of microtubule (MT)-binding proteins in embryo division control. Organization of both MT and actin cytoskeleton depends on auxin response, and genetically controlled MT or actin depolymerization in embryos leads to disruption of asymmetric divisions, including reversion to the default. Our work shows how auxin-dependent control of MT and actin cytoskeleton properties interacts with cell geometry to generate asymmetric divisions during the earliest steps in plant development.Graphical abstract} } @Article{IPB-531, author = {Ziegler, J. and Bochnia, M. and Zeyner, A. and}, title = {{Aminosäurennachweis in geringsten ProbenmengenBestimmung von Hypoglycin A}}, year = {2021}, journal = {Wiley Analytical Science}, url = {https://analyticalscience.wiley.com/do/10.1002/was.000600226/full/}, } @Article{IPB-527, author = {Zang, J. and Klemm, S. and Pain, C. and Duckney, P. and Bao, Z. and Stamm, G. and Kriechbaumer, V. and Bürstenbinder, K. and Hussey, P. J. and Wang, P. and}, title = {{A novel plant actin-microtubule bridging complex regulates cytoskeletal and ER structure at ER-PM contact sites}}, year = {2021}, pages = {1251-1260}, journal = {Curr. Biol.}, doi = {10.1016/j.cub.2020.12.009}, url = {https://doi.org/10.1016/j.cub.2020.12.009}, volume = {31}, abstract = {In plants, the cortical endoplasmic reticulum (ER) network is connected to the plasma membrane (PM) through the ER-PM contact sites (EPCSs), whose structures are maintained by EPCS resident proteins and the cytoskeleton.1-7 Strong co-alignment between EPCSs and the cytoskeleton is observed in plants,1,8 but little is known of how the cytoskeleton is maintained and regulated at the EPCS. Here, we have used a yeast-two-hybrid screen and subsequent in vivo interaction studies in plants by fluorescence resonance energy transfer (FRET)-fluorescence lifetime imaging microscopy (FLIM) analysis to identify two microtubule binding proteins, KLCR1 (kinesin-light-chain-related protein 1) and IQD2 (IQ67-domain 2), that interact with the actin binding protein NET3C and form a component of plant EPCS that mediates the link between the actin and microtubule networks. The NET3C-KLCR1-IQD2 module, acting as an actin-microtubule bridging complex, has a direct influence on ER morphology and EPCS structure. Their loss-of-function mutants, net3a/NET3C RNAi, klcr1, or iqd2, exhibit defects in pavement cell morphology, which we suggest is linked to the disorganization of both actin filaments and microtubules. In conclusion, our results reveal a novel cytoskeletal-associated complex, which is essential for the maintenance and organization of cytoskeletal structure and ER morphology at the EPCS and for normal plant cell morphogenesis.} } @Article{IPB-483, author = {Mielke, S. and Zimmer, M. and Meena, M. K. and Dreos, R. and Stellmach, H. and Hause, B. and Voiniciuc, C. and Gasperini, D. and}, title = {{Jasmonate biosynthesis arising from altered cell walls is prompted by turgor-driven mechanical compression}}, year = {2021}, pages = {eabf0356}, journal = {Sci. Adv.}, doi = {10.1126/sciadv.abf0356}, url = {https://advances.sciencemag.org/content/7/7/eabf0356}, volume = {7}, abstract = {Despite the vital roles of jasmonoyl-isoleucine (JA-Ile) in governing plant growth and environmental acclimation, it remains unclear what intracellular processes lead to its induction. Here, we provide compelling genetic evidence that mechanical and osmotic regulation of turgor pressure represents a key elicitor of JA-Ile biosynthesis. After identifying cell wall mutant alleles in KORRIGAN1 (KOR1) with elevated JA-Ile in seedling roots, we found that ectopic JA-Ile resulted from cell nonautonomous signals deriving from enlarged cortex cells compressing inner tissues and stimulating JA-Ile production. Restoring cortex cell size by cell type–specific KOR1 complementation, by isolating a genetic kor1 suppressor, and by lowering turgor pressure with hyperosmotic treatments abolished JA-Ile signaling. Conversely, hypoosmotic treatment activated JA-Ile signaling in wild-type plants. Furthermore, constitutive JA-Ile levels guided mutant roots toward greater water availability. Collectively, these findings enhance our understanding on JA-Ile biosynthesis initiation and reveal a previously undescribed role of JA-Ile in orchestrating environmental resilience.} } @Article{IPB-482, author = {Mercier, C. and Roux, B. and Havé, M. and Le Poder, L. and Duong, N. and David, P. and Leonhardt, N. and Blanchard, L. and Naumann, C. and Abel, S. and Cuyas, L. and Pluchon, S. and Laurent, N. and Desnos, T. and}, title = {{Root responses to aluminium and iron stresses require the SIZ1 SUMO ligase to modulate the STOP1 transcription factor}}, year = {2021}, pages = {1507-1521}, journal = {Plant J.}, doi = {10.1111/tpj.15525}, url = {https://doi.org/10.1111/tpj.15525}, volume = {108}, abstract = {STOP1, an Arabidopsis transcription factor favouring root growth tolerance against Al toxicity, acts in the response to iron under low Pi (-Pi). Previous studies have shown that Al and Fe regulate the stability and accumulation of STOP1 in roots, and that the STOP1 protein is sumoylated by an unknown E3 ligase. Here, using a forward genetics suppressor screen, we identified the E3 SUMO (small ubiquitin-like modifier) ligase SIZ1 as a modulator of STOP1 signalling. Mutations in SIZ1 increase the expression of ALMT1 (a direct target of STOP1) and root growth responses to Al and Fe stress in a STOP1-dependent manner. Moreover, loss-of-function mutations in SIZ1 enhance the abundance of STOP1 in the root tip. However, no sumoylated STOP1 protein was detected by western blot analysis in our sumoylation assay in E. coli, suggesting the presence of a more sophisticated mechanism. We conclude that the sumo ligase SIZ1 negatively regulates STOP1 signalling, at least in part by modulating STOP1 protein in the root tip. Our results will allow a better understanding of this signalling pathway. } } @Article{IPB-481, author = {Menna, A. and Dora, S. and Sancho-Andrés, G. and Kashyap, A. and Meena, M. K. and Sklodowski, K. and Gasperini, D. and Coll, N. S. and Sánchez-Rodríguez, C. and}, title = {{A primary cell wall cellulose-dependent defense mechanism against vascular pathogens revealed by time-resolved dual transcriptomics}}, year = {2021}, pages = {161}, journal = {BMC Biol.}, doi = {10.1186/s12915-021-01100-6}, url = {https://doi.org/10.1186/s12915-021-01100-6}, volume = {19}, abstract = {Background: Cell walls (CWs) are protein-rich polysaccharide matrices essential for plant growth and environmental acclimation. The CW constitutes the first physical barrier as well as a primary source of nutrients for microbes interacting with plants, such as the vascular pathogen Fusarium oxysporum (Fo). Fo colonizes roots, advancing through the plant primary CWs towards the vasculature, where it grows causing devastation in many crops. The pathogenicity of Fo and other vascular microbes relies on their capacity to reach and colonize the xylem. However, little is known about the root-microbe interaction before the pathogen reaches the vasculature and the role of the plant CW during this process. Results: Using the pathosystem Arabidopsis-Fo5176, we show dynamic transcriptional changes in both fungus and root during their interaction. One of the earliest plant responses to Fo5176 was the downregulation of primary CW synthesis genes. We observed enhanced resistance to Fo5176 in Arabidopsis mutants impaired in primary CW cellulose synthesis. We confirmed that Arabidopsis roots deposit lignin in response to Fo5176 infection, but we show that lignin-deficient mutants were as susceptible as wildtype plants to Fo5176. Genetic impairment of jasmonic acid biosynthesis and signaling did not alter Arabidopsis response to Fo5176, whereas impairment of ethylene signaling did increase vasculature colonization by Fo5176. Abolishing ethylene signaling attenuated the observed resistance while maintaining the dwarfism observed in primary CW cellulose-deficient mutants. Conclusions: Our study provides significant insights on the dynamic root-vascular pathogen interaction at the transcriptome level and the vital role of primary CW cellulose during defense response to these pathogens. These findings represent an essential resource for the generation of plant resistance to Fo that can be transferred to other vascular pathosystems.Keywords: Arabidopsis, Fusarium oxysporum, Ralstonia solanacearum, plant-pathogen interaction, dual-time coursetranscriptomics, cellulose, ethylene, defense response} } @Article{IPB-480, author = {Meena, S. and Wagner, C. and Caggegi, L. and Baumann-Kaschig, K. and Ried, M. K. and}, title = {{A user-friendly protocol for the cultivation and successful crossing of Lotus japonicus}}, year = {2021}, journal = {Bio Protoc.}, doi = {10.21769/p1464}, abstract = {This is a detailed and user-friendly protocol for the cultivation and successful crossing of Lotus japonicus (L. japonicus) e.g. for the generation of higher order mutants, based on methods previously reported (Grant et al., 1962; Handberg and Stougaards, 1992; Jiang and Gresshoff, 1997; Pajuelo and Stougaard, 2005).} } @Article{IPB-469, author = {Kumari, P. and Dahiya, P. and Livanos, P. and Zergiebel, L. and Kölling, M. and Poeschl, Y. and Stamm, G. and Hermann, A. and Abel, S. and Müller, S. and Bürstenbinder, K. and}, title = {{IQ67 DOMAIN proteins facilitate preprophase band formation and division-plane orientation}}, year = {2021}, pages = {739-747}, journal = {Nat. Plants}, doi = {10.1038/s41477-021-00923-z}, url = {https://doi.org/10.1038/s41477-021-00923-z}, volume = {7}, abstract = {Spatiotemporal control of cell division is essential for the growth and development of multicellular organisms. In plant cells, proper cell plate insertion during cytokinesis relies on the premitotic establishment of the division plane at the cell cortex. Two plant-specific cytoskeleton arrays, the preprophase band (PPB) and the phragmoplast, play important roles in division-plane orientation and cell plate formation, respectively1. Microtubule organization and dynamics and their communication with membranes at the cortex and cell plate are coordinated by multiple, mostly distinct microtubule-associated proteins2. How division-plane selection and establishment are linked, however, is still unknown. Here, we report members of the Arabidopsis IQ67 DOMAIN (IQD) family3 as microtubule-targeted proteins that localize to the PPB and phragmoplast and additionally reside at the cell plate and a polarized cortical region including the cortical division zone (CDZ). IQDs physically interact with PHRAGMOPLAST ORIENTING KINESIN (POK) proteins4,5 and PLECKSTRIN HOMOLOGY GTPase ACTIVATING (PHGAP) proteins6, which are core components of the CDZ1. The loss of IQD function impairs PPB formation and affects CDZ recruitment of POKs and PHGAPs, resulting in division-plane positioning defects. We propose that IQDs act as cellular scaffolds that facilitate PPB formation and CDZ set-up during symmetric cell division.} } @Article{IPB-466, author = {Klionsky, D. J. and Abdel-Aziz, A. K. and Abdelfatah, S. and Abdellatif, M. and Abdoli, A. and Abel, S. and Naumann, C. and et al., . and}, title = {{Guidelines for the use and interpretation of assays for monitoring autophagy (4th edition)}}, year = {2021}, pages = {1-382}, journal = {Autophagy}, doi = {10.1080/15548627.2020.1797280}, volume = {17}, abstract = {In 2008, we published the first set of guidelines for standardizing research in autophagy. Since then, this topic has received increasing attention, and many scientists have entered the field. Our knowledge base and relevant new technologies have also been expanding. Thus, it is important to formulate on a regular basis  updated  guidelines  for  monitoring  autophagy  in  different  organisms.  Despite  numerous  reviews, there  continues  to  be  confusion  regarding  acceptable  methods  to  evaluate  autophagy,  especially  in multicellular  eukaryotes.  Here,  we  present  a  set  of  guidelines  for  investigators  to  select  and  interpret methods  to  examine  autophagy  and  related  processes,  and  for  reviewers  to  provide  realistic  and reasonable  critiques  of  reports  that  are  focused  on  these  processes.  These  guidelines  are  not  meant to be a dogmatic set of rules, because the appropriateness of any assay largely depends on the question being  asked  and  the  system  being  used.  Moreover,  no  individual  assay  is  perfect  for  every situation, calling  for  the  use  of  multiple  techniques  to  properly  monitor  autophagy  in  each  experimental  setting. Finally,  several  core  components  of  the  autophagy  machinery  have  been  implicated  in  distinct  auto-phagic  processes  (canonical  and  noncanonical  autophagy),  implying  that  genetic  approaches  to  block autophagy  should  rely  on  targeting  two  or  more  autophagy-related  genes  that  ideally  participate  in distinct steps of the pathway. Along similar lines, because multiple proteins involved in autophagy also regulate other cellular pathways including apoptosis, not all of them can be used as a specific marker for bona fide autophagic responses. Here, we critically discuss current methods of assessing autophagy and the information they can, or cannot, provide. Our ultimate goal is to encourage intellectual and technical innovation  in  the  field.} } @Article{IPB-464, author = {Kahsay, B. N. and Ziegler, J. and Imming, P. and Gebre-Mariam, T. and Neubert, R. H. H. and Moeller, L. and}, title = {{Free amino acid contents of selected Ethiopian plant and fungi species: a search for alternative natural free amino acid sources for cosmeceutical applications}}, year = {2021}, pages = {1105-1122}, journal = {Amino Acids}, doi = {10.1007/s00726-021-03008-5}, url = {https://link.springer.com/article/10.1007/s00726-021-03008-5}, volume = {53}, abstract = {Free amino acids (FAAs), the major constituents of the natural moisturizing factor (NMF), are very important for maintaining the moisture balance of human skin and their deficiency results in dry skin conditions. There is a great interest in the identification and use of nature-based sources of these molecules for such cosmeceutical applications. The objective of the present study was, therefore, to investigate the FAA contents of selected Ethiopian plant and fungi species; and select the best sources so as to use them for the stated purpose. About 59 different plant species and oyster mushroom were included in the study and the concentrations of 27 FAAs were analyzed. Each sample was collected, lyophilized, extracted using aqueous solvent, derivatized with Fluorenylmethoxycarbonyl chloride (Fmoc-Cl) prior to solid-phase extraction and quantified using Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC-ESI–MS/MS) system. All the 27 FAAs were detected in most of the samples. The dominant FAAs that are part of the NMF were found at sufficiently high concentration in the mushroom and some of the plants. This indicates that FAAs that could be included in the preparations for the management of dry skin condition can be obtained from a single natural resource and the use of these resources for the specified purpose have both economic and therapeutic advantage in addition to fulfilling customer needs.} } @Article{IPB-455, author = {Hou, S. and Thiergart, T. and Vannier, N. and Mesny, F. and Ziegler, J. and Pickel, B. and Hacquard, S. and}, title = {{A microbiota–root–shoot circuit favours Arabidopsis growth over defence under suboptimal light}}, year = {2021}, pages = {1078–1092}, journal = {Nat. Plants}, doi = {10.1038/s41477-021-00956-4}, url = {https://www.nature.com/articles/s41477-021-00956-4}, volume = {7}, abstract = {AbstractBidirectional root–shoot signalling is probably key in orchestrating stress responses and ensuring plant survival. Here, we show that Arabidopsis thaliana responses to microbial root commensals and light are interconnected along a microbiota–root–shoot axis. Microbiota and light manipulation experiments in a gnotobiotic plant system reveal that low photosynthetically active radiation perceived by leaves induces long-distance modulation of root bacterial communities but not fungal or oomycete communities. Reciprocally, microbial commensals alleviate plant growth deficiency under low photosynthetically active radiation. This growth rescue was associated with reduced microbiota-induced aboveground defence responses and altered resistance to foliar pathogens compared with the control light condition. Inspection of a set of A. thaliana mutants reveals that this microbiota- and light-dependent growth–defence trade-off is directly explained by belowground bacterial community composition and requires the host transcriptional regulator MYC2. Our work indicates that aboveground stress responses in plants can be modulated by signals from microbial root commensals.} } @Article{IPB-435, author = {Chutia, R. and Scharfenberg, S. and Neumann, S. and Abel, S. and Ziegler, J. and}, title = {{Modulation of phosphate deficiency-induced metabolic changes by iron availability in Arabidopsis thaliana}}, year = {2021}, pages = {7609}, journal = {Int. J. Mol. Sci.}, doi = {10.3390/ijms22147609}, url = {https://doi.org/10.3390/ijms22147609}, volume = {22}, abstract = {Concurrent suboptimal supply of several nutrients requires the coordination of nutrient-specific transcriptional, phenotypic, and metabolic changes in plants in order to optimize growth and development in most agricultural and natural ecosystems. Phosphate (Pi) and iron (Fe) deficiency induce overlapping but mostly opposing transcriptional and root growth responses in Arabidopsis thaliana. On the metabolite level, Pi deficiency negatively modulates Fe deficiency-induced coumarin accumulation, which is controlled by Fe as well as Pi deficiency response regulators. Here, we report the impact of Fe availability on seedling growth under Pi limiting conditions and on Pi deficiency-induced accumulation of amino acids and organic acids, which play important roles in Pi use efficiency. Fe deficiency in Pi replete conditions hardly changed growth and metabolite profiles in roots and shoots of Arabidopsis thaliana, but partially rescued growth under conditions of Pi starvation and severely modulated Pi deficiency-induced metabolic adjustments. Analysis of T-DNA insertion lines revealed the concerted coordination of metabolic profiles by regulators of Fe (FIT, bHLH104, BRUTUS, PYE) as well as of Pi (SPX1, PHR1, PHL1, bHLH32) starvation responses. The results show the interdependency of Pi and Fe availability and the interplay between Pi and Fe starvation signaling on the generation of plant metabolite profiles.} } @Article{IPB-431, author = {Bochnia, M. and Ziegler, J. and Glatter, M. and Zeyner, A. and}, title = {{Hypoglycin A in cow’s milk—A pilot study}}, year = {2021}, pages = {381}, journal = {Toxins}, doi = {10.3390/toxins13060381}, url = {https://www.mdpi.com/2072-6651/13/6/381}, volume = {13}, abstract = {Hypoglycin A (HGA) originating from soapberry fruits (litchi, and ackee) seeds or seedlings from the sycamore maple (SM) tree (related to Sapindaceae) may cause Jamaican vomiting sickness in humans and atypical myopathy in horses and ruminants. A possible transfer into dairy cow’s milk cannot be ruled out since the literature has revealed HGA in the milk of mares and in the offal of captured deer following HGA intoxication. From a study, carried out for another purpose, bulk raw milk samples from four randomly selected dairy farms were available. The cows were pastured in the daytime. A sycamore maple tree was found on the pasture of farm No. 1 only. Bulk milk from the individual tank or milk filling station was sampled in parallels and analyzed for HGA by LC-ESI-MS/MS. Measurable concentrations of HGA occurred only in milk from farm No. 1 and amounted to 120 and 489 nmol/L. Despite low and very variable HGA concentrations, the results indicate that the ingested toxin, once eaten, is transferred into the milk. However, it is unknown how much HGA the individual cow ingested during grazing and what amount was transferred into the bulk milk samples. As a prerequisite for a possible future safety assessment, carry-over studies are needed. Furthermore, the toxins’ stability during milk processing should also be investigated as well.} } @Article{IPB-425, author = {Bao, Z. and Xu, Z. and Zang, J. and Bürstenbinder, K. and Wang, P. and}, title = {{The Morphological Diversity of Plant Organs: Manipulating the Organization of Microtubules May Do the Trick}}, year = {2021}, pages = {649626}, journal = {Front Cell Dev Biol}, doi = {10.3389/fcell.2021.649626}, url = {https://doi.org/10.3389/fcell.2021.649626}, volume = {9}, } @Article{IPB-542, author = {Dallery, J.-F. and Zimmer, M. and Halder, V. and Suliman, M. and Pigné, S. and Le Goff, G. and Gianniou, D. D. and Trougakos, I. P. and Ouazzani, J. and Gasperini, D. and O’Connell, R. J. and}, title = {{Inhibition of jasmonate-mediated plant defences by the fungal metabolite higginsianin B}}, year = {2020}, pages = {2910-2921}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/eraa061}, volume = {71}, abstract = {Infection of Arabidopsis thaliana by the ascomycete fungus Colletotrichum higginsianum is characterised by an early symptomless biotrophic phase followed by a destructive necrotrophic phase. The fungal genome contains 77 secondary metabolism-related biosynthetic gene clusters (BGCs), and their expression during the infection process is tightly regulated. Deleting CclA, a chromatin regulator involved in repression of some BGCs through H3K4 trimethylation, allowed overproduction of 3 families of terpenoids and isolation of 12 different molecules. These natural products were tested in combination with methyl jasmonate (MeJA), an elicitor of jasmonate responses, for their capacity to alter defence gene induction in Arabidopsis. Higginsianin B inhibited MeJA-triggered expression of the defence reporter VSP1p:GUS, suggesting it may block bioactive JA-Ile synthesis or signalling in planta. Using the JA-Ile sensor Jas9-VENUS, we found that higginsianin B, but not three other structurally-related molecules, suppressed JA-Ile signalling by preventing degradation of JAZ proteins, the repressors of JA responses. Higginsianin B likely blocks the 26S proteasome-dependent degradation of JAZ proteins because it inhibited chymotrypsin- and caspase-like protease activities. The inhibition of target degradation by higginsianin B also extended to auxin signalling, as higginsianin B treatment reduced IAA-dependent expression of DR5p:GUS. Overall, our data indicate that specific fungal secondary metabolites can act similarly to protein effectors to subvert plant immune and developmental responses.} } @Article{IPB-536, author = {Bassal, M. and Abukhalaf, M. and Majovsky, P. and Thieme, D. and Herr, T. and Ayash, M. and Tabassum, N. and Al Shweiki, M. R. and Proksch, C. and Hmedat, A. and Ziegler, J. and Lee, J. and Neumann, S. and Hoehenwarter, W. and}, title = {{Reshaping of the Arabidopsis thaliana Proteome Landscape and Co-regulation of Proteins in Development and Immunity}}, year = {2020}, pages = {1709-1732}, journal = {Mol. Plant}, doi = {10.1016/j.molp.2020.09.024}, volume = {13}, abstract = {Proteome remodeling is a fundamental adaptive response, and proteins in complexes and functionally related proteins are often co-expressed. Using a deep sampling strategy we define core proteomes of Arabidopsis thaliana tissues with around 10 000 proteins per tissue, and absolutely quantify (copy numbers per cell) nearly 16 000 proteins throughout the plant lifecycle. A proteome-wide survey of global post-translational modification revealed amino acid exchanges pointing to potential conservation of translational infidelity in eukaryotes. Correlation analysis of protein abundance uncovered potentially new tissue- and age-specific roles of entire signaling modules regulating transcription in photosynthesis, seed development, and senescence and abscission. Among others, the data suggest a potential function of RD26 and other NAC transcription factors in seed development related to desiccation tolerance as well as a possible function of cysteine-rich receptor-like kinases (CRKs) as ROS sensors in senescence. All of the components of ribosome biogenesis factor (RBF) complexes were found to be co-expressed in a tissue- and age-specific manner, indicating functional promiscuity in the assembly of these less-studied protein complexes in Arabidopsis. Furthermore, we characterized detailed proteome remodeling in basal immunity by treating Arabidopsis seeldings with flg22. Through simultaneously monitoring phytohormone and transcript changes upon flg22 treatment, we obtained strong evidence of suppression of jasmonate (JA) and JA-isoleucine (JA-Ile) levels by deconjugation and hydroxylation by IAA-ALA RESISTANT3 (IAR3) and JASMONATE-INDUCED OXYGENASE 2 (JOX2), respectively, under the control of JASMONATE INSENSITIVE 1 (MYC2), suggesting an unrecognized role of a new JA regulatory switch in pattern-triggered immunity. Taken together, the datasets generated in this study present extensive coverage of the Arabidopsis proteome in various biological scenarios, providing a rich resource available to the whole plant science community.} } @Article{IPB-533, author = {Anwer, M. U. and Davis, A. and Davis, S. J. and Quint, M. and}, title = {{Photoperiod sensing of the circadian clock is controlled by EARLY FLOWERING 3 and GIGANTEA}}, year = {2020}, pages = {1397-1410}, journal = {Plant J.}, doi = {10.1111/tpj.14604}, volume = {101}, abstract = {ELF3 and GI are two important components of the Arabidopsis circadian clock. They are not only essential for the oscillator function but are also pivotal in mediating light inputs to the oscillator. Lack of either results in a defective oscillator causing severely compromised output pathways, such as photoperiodic flowering and hypocotyl elongation. Although single loss of function mutants of ELF3 and GI have been well‐studied, their genetic interaction remains unclear. We generated an elf3 gi double mutant to study their genetic relationship in clock‐controlled growth and phase transition phenotypes. We found that ELF3 and GI repress growth differentially during the night and the day, respectively. Circadian clock assays revealed that ELF3 and GI are essential Zeitnehmers that enable the oscillator to synchronize the endogenous cellular mechanisms to external environmental signals. In their absence, the circadian oscillator fails to synchronize to the light‐dark cycles even under diurnal conditions. Consequently, clock‐mediated photoperiod‐responsive growth and development are completely lost in plants lacking both genes, suggesting that ELF3 and GI together convey photoperiod sensing to the central oscillator. Since ELF3 and GI are conserved across flowering plants and represent important breeding and domestication targets, our data highlight the possibility of developing photoperiod‐insensitive crops by adjusting the allelic combination of these two key genes.} } @Article{IPB-582, author = {Orosa, B. and Üstün, S. and Calderón Villalobos, L. I. A. and Genschik, P. and Gibbs, D. and Holdsworth, M. J. and Isono, E. and Lois, M. and Trujillo, M. and Sadanandom, A. and}, title = {{Plant proteostasis – shaping the proteome: a research community aiming to understand molecular mechanisms that control protein abundance}}, year = {2020}, pages = {1028-1033}, journal = {New Phytol.}, doi = {10.1111/nph.16664}, volume = {227}, } @Article{IPB-578, author = {Niemeyer, M. and Moreno Castillo, E. and Ihling, C. H. and Iacobucci, C. and Wilde, V. and Hellmuth, A. and Hoehenwarter, W. and Samodelov, S. L. and Zurbriggen, M. D. and Kastritis, P. L. and Sinz, A. and Calderón Villalobos, L. I. A. and}, title = {{Flexibility of intrinsically disordered degrons in AUX/IAA proteins reinforces auxin co-receptor assemblies}}, year = {2020}, pages = {2277}, journal = {Nat. Commun.}, doi = {10.1038/s41467-020-16147-2}, volume = {11}, abstract = {Cullin RING-type E3 ubiquitin ligases SCFTIR1/AFB1-5 and their AUX/IAA targets perceive the phytohormone auxin. The F-box protein TIR1 binds a surface-exposed degron in AUX/IAAs promoting their ubiquitylation and rapid auxin-regulated proteasomal degradation. Here, by adopting biochemical, structural proteomics and in vivo approaches we unveil how flexibility in AUX/IAAs and regions in TIR1 affect their conformational ensemble allowing surface accessibility of degrons. We resolve TIR1·auxin·IAA7 and TIR1·auxin·IAA12 complex topology, and show that flexible intrinsically disordered regions (IDRs) in the degron’s vicinity, cooperatively position AUX/IAAs on TIR1. We identify essential residues at the TIR1 N- and C-termini, which provide non-native interaction interfaces with IDRs and the folded PB1 domain of AUX/IAAs. We thereby establish a role for IDRs in modulating auxin receptor assemblies. By securing AUX/IAAs on two opposite surfaces of TIR1, IDR diversity supports locally tailored positioning for targeted ubiquitylation, and might provide conformational flexibility for a multiplicity of functional states.} } @Article{IPB-565, author = {Kuhn, J. H. and Adkins, S. and Alioto, D. and Gago-Zachert, S. and et al., . and}, title = {{2020 taxonomic update for phylum Negarnaviricota (Riboviria: Orthornavirae), including the large orders Bunyavirales and Mononegavirales}}, year = {2020}, pages = {3023–3072}, journal = {Arch. Virol.}, doi = {10.1007/s00705-020-04731-2}, url = {https://doi.org/10.1007/s00705-020-04731-2}, volume = {165}, abstract = {In March 2020, following the annual International Committee on Taxonomy of Viruses (ICTV) ratification vote on newly proposed taxa, the phylum Negarnaviricota was amended and emended. At the genus rank, 20 new genera were added, two were deleted, one was moved, and three were renamed. At the species rank, 160 species were added, four were deleted, ten were moved and renamed, and 30 species were renamed. This article presents the updated taxonomy of Negarnaviricota as now accepted by the ICTV.} } @Article{IPB-618, author = {Wasternack, C. and}, title = {{Sulfation switch in the shade}}, year = {2020}, pages = {186-187}, journal = {Nat. Plants}, doi = {10.1038/s41477-020-0620-8}, volume = {6}, abstract = {Plants adjust the balance between growth and defence using photoreceptors and jasmonates. Levels of active jasmonates are reduced in a phytochrome B-dependent manner by upregulation of a 12-hydroxyjasmonate sulfotransferase, leading to increase in shade avoidance and decrease in defence.} } @Article{IPB-617, author = {Wasternack, C. and}, title = {{Determination of sex by jasmonate}}, year = {2020}, pages = {162-164}, journal = {J. Integr. Plant Biol.}, doi = {10.1111/jipb.12840}, volume = {62}, } @Article{IPB-606, author = {Stephani, M. and Picchianti, L. and Gajic, A. and Beveridge, R. and Skarwan, E. and Sanchez de Medina Hernandez, V. and Mohseni, A. and Clavel, M. and Zeng, Y. and Naumann, C. and Matuszkiewicz, M. and Turco, E. and Loefke, C. and Li, B. and Durnberger, G. and Schutzbier, M. and Chen, H. T. and Abdrakhmanov, A. and Savova, A. and Chia, K.-S. and Djamei, A. and Schaffner, I. and Abel, S. and Jiang, L. and Mechtler, K. and Ikeda, F. and Martens, S. and Clausen, T. and Dagdas, Y. and}, title = {{A cross-kingdom conserved ER-phagy receptor maintains endoplasmic reticulum homeostasis during stress}}, year = {2020}, pages = {e58396}, journal = {eLife}, doi = {10.7554/elife.58396}, url = {https://elifesciences.org/articles/58396}, volume = {9}, abstract = {Eukaryotes have evolved various quality control mechanisms to promote proteostasis in the endoplasmic reticulum (ER). Selective removal of certain ER domains via autophagy (termed as ER-phagy) has emerged as a major quality control mechanism. However, the degree to which ER-phagy is employed by other branches of ER-quality control remains largely elusive. Here, we identify a cytosolic protein, C53, that is specifically recruited to autophagosomes during ER-stress, in both plant and mammalian cells. C53 interacts with ATG8 via a distinct binding epitope, featuring a shuffled ATG8 interacting motif (sAIM). C53 senses proteotoxic stress in the ER lumen by forming a tripartite receptor complex with the ER-associated ufmylation ligase UFL1 and its membrane adaptor DDRGK1. The C53/UFL1/DDRGK1 receptor complex is activated by stalled ribosomes and induces the degradation of internal or passenger proteins in the ER. Consistently, the C53 receptor complex and ufmylation mutants are highly susceptible to ER stress. Thus, C53 forms an ancient quality control pathway that bridges selective autophagy with ribosome-associated quality control in the ER.} } @Article{IPB-597, author = {Serra, P. and Carbonell, A. and Navarro, B. and Gago-Zachert, S. and Li, S. and Di Serio, F. and Flores, R. and}, title = {{Symptomatic plant viroid infections in phytopathogenic fungi: A request for a critical reassessment}}, year = {2020}, pages = {10126-10128}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.1922249117}, volume = {117}, } @Article{IPB-2495, author = {Ried, M. K. and Wild, R. and Zhu, J. and Broger, L. and Harmel, R. K. and Hothorn, L. A. and Fiedler, D. and Hothorn, M. and}, title = {{Inositol pyrophosphates promote the interaction of SPX domains with the coiled-coil motif of PHR transcription factors to regulate plant phosphate homeostasis}}, year = {2019}, journal = {bioRxiv}, doi = {10.1101/2019.12.13.875393}, abstract = {Phosphorus is an essential nutrient taken up by organisms in the form of inorganic phosphate (Pi). Eukaryotes have evolved sophisticated Pi sensing and signalling cascades, enabling them to maintain cellular Pi concentrations. Pi homeostasis is regulated by inositol pyrophosphate signalling molecules (PP-InsPs), which are sensed by SPX-domain containing proteins. In plants, PP-InsP bound SPX receptors inactivate Myb coiled-coil (MYB-CC) Pi starvation response transcription factors (PHRs) by an unknown mechanism. Here we report that a InsP8 – SPX complex targets the plant-unique CC domain of PHRs. Crystal structures of the CC domain reveal an unusual four-stranded anti-parallel arrangement. Interface mutations in the CC domain yield monomeric PHR1, which is no longer able to bind DNA with high affinity. Mutation of conserved basic residues located at the surface of the CC domain disrupt interaction with the SPX receptor in vitro and in planta, resulting in constitutive Pi starvation responses. Together, our findings suggest that InsP8 regulates plant Pi homeostasis by controlling the oligomeric state and hence the promoter binding capability of PHRs via their SPX receptors.} } @Article{IPB-722, author = {Ronzan, M. and Piacentini, D. and Fattorini, L. and Federica, D. R. and Caboni, E. and Eiche, E. and Ziegler, J. and Hause, B. and Riemann, M. and Betti, C. and Altamura, M. M. and Falasca, G. and}, title = {{Auxin-jasmonate crosstalk in Oryza sativa L. root system formation after cadmium and/or arsenic exposure}}, year = {2019}, pages = {59-69}, journal = {Environ. Exp. Bot.}, doi = {10.1016/j.envexpbot.2019.05.013}, volume = {165}, abstract = {Soil pollutants may affect root growth through interactions among phytohormones like auxin and jasmonates. Rice is frequently grown in paddy fields contaminated by cadmium and arsenic, but the effects of these pollutants on jasmonates/auxin crosstalk during adventitious and lateral roots formation are widely unknown. Therefore, seedlings of Oryza sativa cv. Nihonmasari and of the jasmonate-biosynthetic mutant coleoptile photomorphogenesis2 were exposed to cadmium and/or arsenic, and/or jasmonic acid methyl ester, and then analysed through morphological, histochemical, biochemical and molecular approaches.In both genotypes, arsenic and cadmium accumulated in roots more than shoots. In the roots, arsenic levels were more than twice higher than cadmium levels, either when arsenic was applied alone, or combined with cadmium. Pollutants reduced lateral root density in the wild -type in every treatment condition, but jasmonic acid methyl ester increased it when combined with each pollutant. Interestingly, exposure to cadmium and/or arsenic did not change lateral root density in the mutant. The transcript levels of OsASA2 and OsYUCCA2, auxin biosynthetic genes, increased in the wild-type and mutant roots when pollutants and jasmonic acid methyl ester were applied alone. Auxin (indole-3-acetic acid) levels transiently increased in the roots with cadmium and/or arsenic in the wild-type more than in the mutant. Arsenic and cadmium, when applied alone, induced fluctuations in bioactive jasmonate contents in wild-type roots, but not in the mutant. Auxin distribution was evaluated in roots of OsDR5::GUS seedlings exposed or not to jasmonic acid methyl ester added or not with cadmium and/or arsenic. The DR5::GUS signal in lateral roots was reduced by arsenic, cadmium, and jasmonic acid methyl ester. Lipid peroxidation, evaluated as malondialdehyde levels, was higher in the mutant than in the wild-type, and increased particularly in As presence, in both genotypes.Altogether, the results show that an auxin/jasmonate interaction affects rice root system development in the presence of cadmium and/or arsenic, even if exogenous jasmonic acid methyl ester only slightly mitigates pollutants toxicity.} } @Article{IPB-720, author = {Ried, M. K. and Banhara, A. and Hwu, F.-Y. and Binder, A. and Gust, A. A. and Höfle, C. and Hückelhoven, R. and Nürnberger, T. and Parniske, M. and}, title = {{A set of Arabidopsis genes involved in the accommodation of the downy mildew pathogen Hyaloperonospora arabidopsidis}}, year = {2019}, pages = {e1007747}, journal = {PLOS Pathog.}, doi = {10.1371/journal.ppat.1007747}, volume = {15}, abstract = {The intracellular accommodation structures formed by plant cells to host arbuscular mycorrhiza fungi and biotrophic hyphal pathogens are cytologically similar. Therefore we investigated whether these interactions build on an overlapping genetic framework. In legumes, the malectin-like domain leucine-rich repeat receptor kinase SYMRK, the cation channel POLLUX and members of the nuclear pore NUP107-160 subcomplex are essential for symbiotic signal transduction and arbuscular mycorrhiza development. We identified members of these three groups in Arabidopsis thaliana and explored their impact on the interaction with the oomycete downy mildew pathogen Hyaloperonospora arabidopsidis (Hpa). We report that mutations in the corresponding genes reduced the reproductive success of Hpa as determined by sporangiophore and spore counts. We discovered that a developmental transition of haustorial shape occurred significantly earlier and at higher frequency in the mutants. Analysis of the multiplication of extracellular bacterial pathogens, Hpa-induced cell death or callose accumulation, as well as Hpa- or flg22-induced defence marker gene expression, did not reveal any traces of constitutive or exacerbated defence responses. These findings point towards an overlap between the plant genetic toolboxes involved in the interaction with biotrophic intracellular hyphal symbionts and pathogens in terms of the gene families involved.} } @Article{IPB-702, author = {Naumann, C. and Müller, J. and Sakhonwasee, S. and Wieghaus, A. and Hause, G. and Heisters, M. and Bürstenbinder, K. and Abel, S. and}, title = {{The Local Phosphate Deficiency Response Activates Endoplasmic Reticulum Stress-Dependent Autophagy}}, year = {2019}, pages = {460-476}, journal = {Plant Physiol.}, doi = {10.1104/pp.18.01379}, volume = {179}, abstract = {Inorganic phosphate (Pi) is often a limiting plant nutrient. In members of the Brassicaceae family, such as Arabidopsis (Arabidopsis thaliana), Pi deprivation reshapes root system architecture to favor topsoil foraging. It does so by inhibiting primary root extension and stimulating lateral root formation. Root growth inhibition from phosphate (Pi) deficiency is triggered by iron-stimulated, apoplastic reactive oxygen species generation and cell wall modifications, which impair cell-to-cell communication and meristem maintenance. These processes require LOW PHOSPHATE RESPONSE1 (LPR1), a cell wall-targeted ferroxidase, and PHOSPHATE DEFICIENCY RESPONSE2 (PDR2), the single endoplasmic reticulum (ER)-resident P5-type ATPase (AtP5A), which is thought to control LPR1 secretion or activity. Autophagy is a conserved process involving the vacuolar degradation of cellular components. While the function of autophagy is well established under nutrient starvation (C, N, or S), it remains to be explored under Pi deprivation. Because AtP5A/PDR2 likely functions in the ER stress response, we analyzed the effect of Pi limitation on autophagy. Our comparative study of mutants defective in the local Pi deficiency response, ER stress response, and autophagy demonstrated that ER stress-dependent autophagy is rapidly activated as part of the developmental root response to Pi limitation and requires the genetic PDR2-LPR1 module. We conclude that Pi-dependent activation of autophagy in the root apex is a consequence of local Pi sensing and the associated ER stress response, rather than a means for systemic recycling of the macronutrient.} } @Article{IPB-699, author = {Mitra, D. and Klemm, S. and Kumari, P. and Quegwer, J. and Möller, B. and Poeschl, Y. and Pflug, P. and Stamm, G. and Abel, S. and Bürstenbinder, K. and}, title = {{Microtubule-associated protein IQ67 DOMAIN5 regulates morphogenesis of leaf pavement cells in Arabidopsis thaliana}}, year = {2019}, pages = {529-543}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/ery395}, volume = {70}, abstract = {Plant microtubules form a highly dynamic intracellular network with important roles for regulating cell division, cell proliferation and cell morphology. Its organization and dynamics are coordinated by various microtubule-associated proteins (MAPs) that integrate environmental and developmental stimuli to fine-tune and adjust cytoskeletal arrays. IQ67 DOMAIN (IQD) proteins recently emerged as a class of plant-specific MAPs with largely unknown functions. Here, using a reverse genetics approach, we characterize Arabidopsis IQD5 in terms of its expression domains, subcellular localization and biological functions. We show that IQD5 is expressed mostly in vegetative tissues, where it localizes to cortical microtubule arrays. Our phenotypic analysis of iqd5 loss-of-function lines reveals functions of IQD5 in pavement cell (PC) shape morphogenesis. Histochemical analysis of cell wall composition further suggests reduced rates of cellulose deposition in anticlinal cell walls, which correlate with reduced anisotropic expansion. Lastly, we demonstrate IQD5-dependent recruitment of calmodulin calcium sensors to cortical microtubule arrays and provide first evidence for important roles of calcium in regulation of PC morphogenesis. Our work thus identifies IQD5 as a novel player in PC shape regulation, and, for the first time, links calcium signaling to developmental processes that regulate anisotropic growth in PCs.} } @Article{IPB-698, author = {Mielke, S. and Gasperini, D. and}, title = {{Interplay between Plant Cell Walls and Jasmonate Production}}, year = {2019}, pages = {2629-2637}, journal = {Plant Cell Physiol.}, doi = {10.1093/pcp/pcz119}, volume = {60}, abstract = {Plant cell walls are sophisticated carbohydrate-rich structures representing the immediate contact surface with the extracellular environment, often serving as the first barrier against biotic and abiotic stresses. Notably, a variety of perturbations in plant cell walls result in upregulated jasmonate (JA) production, a phytohormone with essential roles in defense and growth responses. Hence, cell wall-derived signals can initiate intracellular JA-mediated responses and the elucidation of the underlying signaling pathways could provide novel insights into cell wall maintenance and remodeling, as well as advance our understanding on how is JA biosynthesis initiated. This Mini Review will describe current knowledge about cell wall-derived damage signals and their effects on JA biosynthesis, as well as provide future perspectives.} } @Article{IPB-749, author = {Wasternack, C. and Hause, B. and}, title = {{The missing link in jasmonic acid biosynthesis}}, year = {2019}, pages = {776-777}, journal = {Nat. Plants}, doi = {10.1038/s41477-019-0492-y}, volume = {5}, abstract = {Jasmonic acid biosynthesis starts in chloroplasts and is finalized in peroxisomes. The required export of a crucial intermediate out of the chloroplast is now shown to be mediated by a protein from the outer envelope called JASSY.} } @Article{IPB-748, author = {Wasternack, C. and}, title = {{Termination in Jasmonate Signaling by MYC2 and MTBs}}, year = {2019}, pages = {667-669}, journal = {Trends Plant Sci.}, doi = {10.1016/j.tplants.2019.06.001}, volume = {24}, abstract = {Jasmonic acid (JA) signaling can be switched off by metabolism of JA. The master regulator MYC2, interacting with MED25, has been shown to be deactivated by the bHLH transcription factors MTB1, MTB2, and MTB3. An autoregulatory negative feedback loop has been proposed for this termination in JA signaling.} } @Article{IPB-747, author = {Wasternack, C. and}, title = {{New Light on Local and Systemic Wound Signaling}}, year = {2019}, pages = {102-105}, journal = {Trends Plant Sci.}, doi = {10.1016/j.tplants.2018.11.009}, volume = {24}, abstract = {Electric signaling and Ca2\+ waves were discussed to occur in systemic wound responses. Two new overlapping scenarios were identified: (i) membrane depolarization in two special cell types followed by an increase in systemic cytoplasmic Ca2\+ concentration ([Ca2\+]cyt), and (ii) glutamate sensed by GLUTAMATE RECEPTOR LIKE proteins and followed by Ca2\+-based defense in distal leaves.} } @Article{IPB-746, author = {Wasternack, C. and Strnad, M. and}, title = {{Jasmonates are signals in the biosynthesis of secondary metabolites — Pathways, transcription factors and applied aspects — A brief review}}, year = {2019}, pages = {1-11}, journal = {New Biotechnol.}, doi = {10.1016/j.nbt.2017.09.007}, volume = {48}, abstract = {Jasmonates (JAs) are signals in plant stress responses and development. One of the first observed and prominent responses to JAs is the induction of biosynthesis of different groups of secondary compounds. Among them are nicotine, isoquinolines, glucosinolates, anthocyanins, benzophenanthridine alkaloids, artemisinin, and terpenoid indole alkaloids (TIAs), such as vinblastine. This brief review describes modes of action of JAs in the biosynthesis of anthocyanins, nicotine, TIAs, glucosinolates and artemisinin. After introducing JA biosynthesis, the central role of the SCFCOI1-JAZ co-receptor complex in JA perception and MYB-type and MYC-type transcription factors is described. Brief comments are provided on primary metabolites as precursors of secondary compounds. Pathways for the biosynthesis of anthocyanin, nicotine, TIAs, glucosinolates and artemisinin are described with an emphasis on JA-dependent transcription factors, which activate or repress the expression of essential genes encoding enzymes in the biosynthesis of these secondary compounds. Applied aspects are discussed using the biotechnological formation of artemisinin as an example of JA-induced biosynthesis of secondary compounds in plant cell factories.} } @Article{IPB-745, author = {Wasternack, C. and Hause, B. and Abel, S. and}, title = {{Benno Parthier (1932–2019)}}, year = {2019}, pages = {519-520}, journal = {Plant Mol. Biol.}, doi = {10.1007/s11103-019-00927-6}, volume = {101}, } @Article{IPB-731, author = {Schulze, A. and Zimmer, M. and Mielke, S. and Stellmach, H. and Melnyk, C. W. and Hause, B. and Gasperini, D. and}, title = {{Wound-Induced Shoot-to-Root Relocation of JA-Ile Precursors Coordinates Arabidopsis Growth}}, year = {2019}, pages = {1383-1394}, journal = {Mol. Plant}, doi = {10.1016/j.molp.2019.05.013}, volume = {12}, abstract = {Multicellular organisms rely on the movement of signaling molecules across cells, tissues, and organs to communicate among distal sites. In plants, localized leaf damage activates jasmonic acid (JA)-dependent transcriptional reprogramming in both harmed and unharmed tissues. Although it has been indicated that JA species can translocate from damaged into distal sites, the identity of the mobile compound(s), the tissues through which they translocate, and the effect of their relocation remain unknown. Here, we found that following shoot wounding, the relocation of endogenous jasmonates through the phloem is essential to initiate JA signaling and stunt growth in unharmed roots of Arabidopsis thaliana. By employing grafting experiments and hormone profiling, we uncovered that the hormone precursor cis-12-oxo-phytodienoic acid (OPDA) and its derivatives, but not the bioactive JA-Ile conjugate, translocate from wounded shoots into undamaged roots. Upon root relocation, the mobile precursors cooperatively regulated JA responses through their conversion into JA-Ile and JA signaling activation. Collectively, our findings demonstrate the existence of long-distance translocation of endogenous OPDA and its derivatives, which serve as mobile molecules to coordinate shoot-to-root responses, and highlight the importance of a controlled redistribution of hormone precursors among organs during plant stress acclimation.} } @Article{IPB-686, author = {Kölling, M. and Kumari, P. and Bürstenbinder, K. and}, title = {{Calcium- and calmodulin-regulated microtubule-associated proteins as signal-integration hubs at the plasma membrane–cytoskeleton nexus}}, year = {2019}, pages = {387-396}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/ery397}, volume = {70}, abstract = {Plant growth and development are a genetically predetermined series of events but can change dramatically in response to environmental stimuli, involving perpetual pattern formation and reprogramming of development. The rate of growth is determined by cell division and subsequent cell expansion, which are restricted and controlled by the cell wall–plasma membrane–cytoskeleton continuum, and are coordinated by intricate networks that facilitate intra- and intercellular communication. An essential role in cellular signaling is played by calcium ions, which act as universal second messengers that transduce, integrate, and multiply incoming signals during numerous plant growth processes, in part by regulation of the microtubule cytoskeleton. In this review, we highlight recent advances in the understanding of calcium-mediated regulation of microtubule-associated proteins, their function at the microtubule cytoskeleton, and their potential role as hubs in crosstalk with other signaling pathways.} } @Article{IPB-684, author = {Hussain, H. and Ziegler, J. and Mrestani, Y. and Neubert, R. H. H. and}, title = {{Studies of the Corneocytary Pathway Across the Stratum Corneum. Part I: Diffusion of Amino Acids Into the Isolated Corneocytes}}, year = {2019}, pages = {340-344}, journal = {Pharmazie}, doi = {10.1691/ph.2019.8098}, volume = {74}, abstract = {Amino acids (AAs), important constituents of natural moisturizing factors (NMFs) of the skin are decreased in diseased conditions such as psoriasis and atopic dermatitis. No study so far investigated the uptake of AAs into isolated corneocytes (COR). The present study was performed using 19 AAs, including taurine (TAU), to measure their amount diffused into the COR and binding of these AAs to keratin. Incubation of alanine, aspartic acid, asparagine, glutamine, glutamic acid, histidine, proline, serine and TAU with the isolated COR showed uptake after 24 h of 51.6, 95.4, 98.6, 94.1, 95.6, 90.1, 94.6, 72.9 and 57.8 %, respectively, into the COR but no binding with keratin. Uptake of TAU was validated by time dependent in-vitro diffusion models \'without COR and \'with COR\'. The time dependent curve fitting showed that in in-vitro diffusion model \'without COR\' there was no change in the total concentration of TAU until 72 hours, while in diffusion model \'with COR\' the total conc. decreased to 37.8 % after 72 hours. The Pearson\'s correlation coefficient \'r\' between the conc. curves of both in-vitro diffusion models was -0.54 that was an evidence of significant amount of TAU uptake by the COR. AAs as part of the NMFs have a great potential to be diffused into the COR. This property of the AAs can be employed in further dermatological research on diseased or aged skin conditions with NMFs deficiency.} } @Article{IPB-683, author = {Hussain, H. and Ziegler, J. and Hause, G. and Wohlrab, J. and Neubert, R. H. and}, title = {{Quantitative Analysis of Free Amino Acids and Urea Derived from Isolated Corneocytes of Healthy Young, Healthy Aged, and Diseased Skin}}, year = {2019}, pages = {94-100}, journal = {Skin Pharmacol. Physiol.}, doi = {10.1159/000495992}, volume = {32}, abstract = {Background/Aims: Free amino acids (FAAs) and urea, present inside the corneocytes, can be important indicators of skin condition. However, due to the lack of a standard extraction protocol for FAAs from corneocytes, conflicting research results have been reported. Therefore, the purpose of this study was (1) to standardize the extraction protocol and (2) to investigate FAA profiles in healthy young and healthy old volunteers, as well as in psoriasis and atopic dermatitis patients. Methods: Skin samples were collected from four groups (healthy young, healthy old, and psoriasis and atopic dermatitis patients) with 5 volunteers per group. Corneocytes were isolated and examined microscopically. FAAs and urea were extracted from the isolated corneocytes, and their amounts were quantified using LC-ESI/MS/MS (after derivatization with Fmoc-Cl) and colorimetric methods, respectively. Results: The micrographs of the corneocytes showed no morphological features attributable to age or disease conditions. The highest and lowest concentrations of total FAAs and urea were observed in the healthy old group and the healthy young group, respectively. Unlike the other FAAs and urea, citrulline was found at a higher level in the healthy young group than in the disease groups. Conclusion: This study suggests that the levels of FAAs and urea in the skin are affected by age and skin conditions (healthy/diseased). However, further studies are needed to show the effects of different skin conditions on the levels of FAAs and urea.} } @Article{IPB-668, author = {Girardin, A. and Wang, T. and Ding, Y. and Keller, J. and Buendia, L. and Gaston, M. and Ribeyre, C. and Gasciolli, V. and Auriac, M.-C. and Vernié, T. and Bendahmane, A. and Ried, M. K. and Parniske, M. and Morel, P. and Vandenbussche, M. and Schorderet, M. and Reinhardt, D. and Delaux, P.-M. and Bono, J.-J. and Lefebvre, B. and}, title = {{LCO Receptors Involved in Arbuscular Mycorrhiza Are Functional for Rhizobia Perception in Legumes}}, year = {2019}, pages = {4249-4259.e5}, journal = {Curr. Biol.}, doi = {10.1016/j.cub.2019.11.038}, volume = {29}, abstract = {Bacterial lipo-chitooligosaccharides (LCOs) are key mediators of the nitrogen-fixing root nodule symbiosis (RNS) in legumes. The isolation of LCOs from arbuscular mycorrhizal fungi suggested that LCOs are also signaling molecules in arbuscular mycorrhiza (AM). However, the corresponding plant receptors have remained uncharacterized. Here we show that petunia and tomato mutants in the LysM receptor-like kinases LYK10 are impaired in AM formation. Petunia and tomato LYK10 proteins have a high affinity for LCOs (Kd in the nM range) comparable to that previously reported for a legume LCO receptor essential for the RNS. Interestingly, the tomato and petunia LYK10 promoters, when introduced into a legume, were active in nodules similarly to the promoter of the legume orthologous gene. Moreover, tomato and petunia LYK10 coding sequences restored nodulation in legumes mutated in their orthologs. This combination of genetic and biochemical data clearly pinpoints Solanaceous LYK10 as part of an ancestral LCO perception system involved in AM establishment, which has been directly recruited during evolution of the RNS in legumes.} } @Article{IPB-667, author = {Gago-Zachert, S. and Schuck, J. and Weinholdt, C. and Knoblich, M. and Pantaleo, V. and Grosse, I. and Gursinsky, T. and Behrens, S.-E. and}, title = {{Highly efficacious antiviral protection of plants by small interfering RNAs identified in vitro}}, year = {2019}, pages = {9343-9357}, journal = {Nucleic Acids Res.}, doi = {10.1093/nar/gkz678}, volume = {47}, abstract = {In response to a viral infection, the plant’s RNA silencing machinery processes viral RNAs into a huge number of small interfering RNAs (siRNAs). However, a very few of these siRNAs actually interfere with viral replication. A reliable approach to identify these immunologically effective siRNAs (esiRNAs) and to define the characteristics underlying their activity has not been available so far. Here, we develop a novel screening approach that enables a rapid functional identification of antiviral esiRNAs. Tests on the efficacy of such identified esiRNAs of a model virus achieved a virtual full protection of plants against a massive subsequent infection in transient applications. We find that the functionality of esiRNAs depends crucially on two properties: the binding affinity to Argonaute proteins and the ability to access the target RNA. The ability to rapidly identify functional esiRNAs could be of great benefit for all RNA silencing-based plant protection measures against viruses and other pathogens.} } @Article{IPB-640, author = {Chutia, R. and Abel, S. and Ziegler, J. and}, title = {{Iron and Phosphate Deficiency Regulators Concertedly Control Coumarin Profiles in Arabidopsis thaliana Roots During Iron, Phosphate, and Combined Deficiencies}}, year = {2019}, pages = {113}, journal = {Front. Plant Sci.}, doi = {10.3389/fpls.2019.00113}, volume = {10}, abstract = {Plants face varying nutrient conditions, to which they have to adapt to. Adaptive responses are nutrient-specific and strategies to ensure supply and homeostasis for one nutrient might be opposite to another one, as shown for phosphate (Pi) and iron (Fe) deficiency responses, where many genes are regulated in an opposing manner. This was also observed on the metabolite levels. Whereas root and exudate levels of catechol-type coumarins, phenylpropanoid-derived 2-benzopyranones, which facilitate Fe acquisition, are elevated after Fe deficiency, they are decreased after Pi deficiency. Exposing plants to combined Pi and Fe deficiency showed that the generation of coumarin profiles in Arabidopsis thaliana roots by Pi deficiency considerably depends on the availability of Fe. Similarly, the effect of Fe deficiency on coumarin profiles is different at low compared to high Pi availability. These findings suggest a fine-tuning of coumarin profiles, which depends on Fe and Pi availability. T-DNA insertion lines exhibiting aberrant expression of genes involved in the regulation of Pi starvation responses (PHO1, PHR1, bHLH32, PHL1, SPX1) and Fe starvation responses (BRUTUS, PYE, bHLH104, FIT) were used to analyze the regulation of the generation of coumarin profiles in Arabidopsis thaliana roots by Pi, Fe, and combined Pi and Fe deficiency. The analysis revealed a role of several Fe-deficiency response regulators in the regulation of Fe and of Pi deficiency-induced coumarin profiles as well as for Pi deficiency response regulators in the regulation of Pi and of Fe deficiency-induced coumarin profiles. Additionally, the regulation of Fe deficiency-induced coumarin profiles by Fe deficiency response regulators is influenced by Pi availability. Conversely, regulation of Pi deficiency-induced coumarin profiles by Pi deficiency response regulators is modified by Fe availability.} } @Article{IPB-635, author = {Bochnia, M. and Sander, J. and Ziegler, J. and Terhardt, M. and Sander, S. and Janzen, N. and Cavalleri, J.-M. V. and Zuraw, A. and Wensch-Dorendorf, M. and Zeyner, A. and}, title = {{Detection of MCPG metabolites in horses with atypical myopathy}}, year = {2019}, pages = {e0211698}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0211698}, volume = {14}, abstract = {Atypical myopathy (AM) in horses is caused by ingestion of seeds of the Acer species (Sapindaceae family). Methylenecyclopropylacetyl-CoA (MCPA-CoA), derived from hypoglycin A (HGA), is currently the only active toxin in Acer pseudoplatanus or Acer negundo seeds related to AM outbreaks. However, seeds or arils of various Sapindaceae (e.g., ackee, lychee, mamoncillo, longan fruit) also contain methylenecyclopropylglycine (MCPG), which is a structural analogue of HGA that can cause hypoglycaemic encephalopathy in humans. The active poison formed from MCPG is methylenecyclopropylformyl-CoA (MCPF-CoA). MCPF-CoA and MCPA-CoA strongly inhibit enzymes that participate in β-oxidation and energy production from fat. The aim of our study was to investigate if MCPG is involved in Acer seed poisoning in horses. MCPG, as well as glycine and carnitine conjugates (MCPF-glycine, MCPF-carnitine), were quantified using high-performance liquid chromatography-tandem mass spectrometry of serum and urine from horses that had ingested Acer pseudoplatanus seeds and developed typical AM symptoms. The results were compared to those of healthy control horses. For comparison, HGA and its glycine and carnitine derivatives were also measured. Additionally, to assess the degree of enzyme inhibition of β-oxidation, several acyl glycines and acyl carnitines were included in the analysis. In addition to HGA and the specific toxic metabolites (MCPA-carnitine and MCPA-glycine), MCPG, MCPF-glycine and MCPF-carnitine were detected in the serum and urine of affected horses. Strong inhibition of β-oxidation was demonstrated by elevated concentrations of all acyl glycines and carnitines, but the highest correlations were observed between MCPF-carnitine and isobutyryl-carnitine (r \= 0.93) as well as between MCPA- (and MCPF-) glycine and valeryl-glycine with r \= 0.96 (and r \= 0.87). As shown here, for biochemical analysis of atypical myopathy of horses, it is necessary to take MCPG and the corresponding metabolites into consideration.} } @Article{IPB-633, author = {Berens, M. L. and Wolinska, K. W. and Spaepen, S. and Ziegler, J. and Nobori, T. and Nair, A. and Krüler, V. and Winkelmüller, T. M. and Wang, Y. and Mine, A. and Becker, D. and Garrido-Oter, R. and Schulze-Lefert, P. and Tsuda, K. and}, title = {{Balancing trade-offs between biotic and abiotic stress responses through leaf age-dependent variation in stress hormone cross-talk}}, year = {2019}, pages = {2364-2373}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.1817233116}, volume = {116}, abstract = {In nature, plants must respond to multiple stresses simultaneously, which likely demands cross-talk between stress-response pathways to minimize fitness costs. Here we provide genetic evidence that biotic and abiotic stress responses are differentially prioritized in Arabidopsis thaliana leaves of different ages to maintain growth and reproduction under combined biotic and abiotic stresses. Abiotic stresses, such as high salinity and drought, blunted immune responses in older rosette leaves through the phytohormone abscisic acid signaling, whereas this antagonistic effect was blocked in younger rosette leaves by PBS3, a signaling component of the defense phytohormone salicylic acid. Plants lacking PBS3 exhibited enhanced abiotic stress tolerance at the cost of decreased fitness under combined biotic and abiotic stresses. Together with this role, PBS3 is also indispensable for the establishment of salt stress- and leaf age-dependent phyllosphere bacterial communities. Collectively, our work reveals a mechanism that balances trade-offs upon conflicting stresses at the organism level and identifies a genetic intersection among plant immunity, leaf microbiota, and abiotic stress tolerance.} } @Article{IPB-632, author = {Bellstaedt, J. and Trenner, J. and Lippmann, R. and Poeschl, Y. and Zhang, X. and Friml, J. and Quint, M. and Delker, C. and}, title = {{A Mobile Auxin Signal Connects Temperature Sensing in Cotyledons with Growth Responses in Hypocotyls}}, year = {2019}, pages = {757-766}, journal = {Plant Physiol.}, doi = {10.1104/pp.18.01377}, volume = {180}, abstract = {Plants have a remarkable capacity to adjust their growth and development to elevated ambient temperatures. Increased elongation growth of roots, hypocotyls, and petioles in warm temperatures are hallmarks of seedling thermomorphogenesis. In the last decade, significant progress has been made to identify the molecular signaling components regulating these growth responses. Increased ambient temperature utilizes diverse components of the light sensing and signal transduction network to trigger growth adjustments. However, it remains unknown whether temperature sensing and responses are universal processes that occur uniformly in all plant organs. Alternatively, temperature sensing may be confined to specific tissues or organs, which would require a systemic signal that mediates responses in distal parts of the plant. Here, we show that Arabidopsis (Arabidopsis thaliana) seedlings show organ-specific transcriptome responses to elevated temperatures and that thermomorphogenesis involves both autonomous and organ-interdependent temperature sensing and signaling. Seedling roots can sense and respond to temperature in a shoot-independent manner, whereas shoot temperature responses require both local and systemic processes. The induction of cell elongation in hypocotyls requires temperature sensing in cotyledons, followed by the generation of a mobile auxin signal. Subsequently, auxin travels to the hypocotyl, where it triggers local brassinosteroid-induced cell elongation in seedling stems, which depends upon a distinct, permissive temperature sensor in the hypocotyl.} } @Article{IPB-851, author = {Wasternack, C. and Feussner, I. and}, title = {{The Oxylipin Pathways: Biochemistry and Function}}, year = {2018}, pages = {363-386}, journal = {Annu. Rev. Plant Biol.}, doi = {10.1146/annurev-arplant-042817-040440}, volume = {69}, abstract = {Plant oxylipins form a constantly growing group of signaling molecules that comprise oxygenated fatty acids and metabolites derived therefrom. In the last decade, the understanding of biosynthesis, metabolism, and action of oxylipins, especially jasmonates, has dramatically improved. Additional mechanistic insights into the action of enzymes and insights into signaling pathways have been deepened for jasmonates. For other oxylipins, such as the hydroxy fatty acids, individual signaling properties and cross talk between different oxylipins or even with additional phytohormones have recently been described. This review summarizes recent understanding of the biosynthesis, regulation, and function of oxylipins.} } @Article{IPB-850, author = {Wasternack, C. and Strnad, M. and}, title = {{Jasmonates: News on Occurrence, Biosynthesis, Metabolism and Action of an Ancient Group of Signaling Compounds}}, year = {2018}, pages = {2539}, journal = {Int. J. Mol. Sci.}, doi = {10.3390/ijms19092539}, volume = {19}, abstract = {Jasmonic acid (JA) and its related derivatives are ubiquitously occurring compounds of land plants acting in numerous stress responses and development. Recent studies on evolution of JA and other oxylipins indicated conserved biosynthesis. JA formation is initiated by oxygenation of α-linolenic acid (α-LeA, 18:3) or 16:3 fatty acid of chloroplast membranes leading to 12-oxo-phytodienoic acid (OPDA) as intermediate compound, but in Marchantiapolymorpha and Physcomitrellapatens, OPDA and some of its derivatives are final products active in a conserved signaling pathway. JA formation and its metabolic conversion take place in chloroplasts, peroxisomes and cytosol, respectively. Metabolites of JA are formed in 12 different pathways leading to active, inactive and partially active compounds. The isoleucine conjugate of JA (JA-Ile) is the ligand of the receptor component COI1 in vascular plants, whereas in the bryophyte M. polymorpha COI1 perceives an OPDA derivative indicating its functionally conserved activity. JA-induced gene expressions in the numerous biotic and abiotic stress responses and development are initiated in a well-studied complex regulation by homeostasis of transcription factors functioning as repressors and activators.} } @Article{IPB-849, author = {Wasternack, C. and Hause, B. and}, title = {{A Bypass in Jasmonate Biosynthesis – the OPR3-independent Formation}}, year = {2018}, pages = {276-279}, journal = {Trends Plant Sci.}, doi = {10.1016/j.tplants.2018.02.011}, volume = {23}, abstract = {For the first time in 25 years, a new pathway for biosynthesis of jasmonic acid (JA) has been identified. JA production takes place via 12-oxo-phytodienoic acid (OPDA) including reduction by OPDA reductases (OPRs). A loss-of-function allele, opr3-3, revealed an OPR3-independent pathway converting OPDA to JA.} } @Article{IPB-816, author = {Nishiyama, T. and Sakayama, H. and de Vries, J. and Buschmann, H. and Saint-Marcoux, D. and Ullrich, K. K. and Haas, F. B. and Vanderstraeten, L. and Becker, D. and Lang, D. and Vosolsobě, S. and Rombauts, S. and Wilhelmsson, P. K. and Janitza, P. and Kern, R. and Heyl, A. and Rümpler, F. and Calderón Villalobos, L. I. A. and Clay, J. M. and Skokan, R. and Toyoda, A. and Suzuki, Y. and Kagoshima, H. and Schijlen, E. and Tajeshwar, N. and Catarino, B. and Hetherington, A. J. and Saltykova, A. and Bonnot, C. and Breuninger, H. and Symeonidi, A. and Radhakrishnan, G. V. and Van Nieuwerburgh, F. and Deforce, D. and Chang, C. and Karol, K. G. and Hedrich, R. and Ulvskov, P. and Glöckner, G. and Delwiche, C. F. and Petrášek, J. and Van de Peer, Y. and Friml, J. and Beilby, M. and Dolan, L. and Kohara, Y. and Sugano, S. and Fujiyama, A. and Delaux, P.-M. and Quint, M. and Theißen, G. and Hagemann, M. and Harholt, J. and Dunand, C. and Zachgo, S. and Langdale, J. and Maumus, F. and Van Der Straeten, D. and Gould, S. B. and Rensing, S. A. and}, title = {{The Chara Genome: Secondary Complexity and Implications for Plant Terrestrialization}}, year = {2018}, pages = {448-464.e24}, journal = {Cell}, doi = {10.1016/j.cell.2018.06.033}, volume = {174}, abstract = {Land plants evolved from charophytic algae, among which Charophyceae possess the most complex body plans. We present the genome of Chara braunii; comparison of the genome to those of land plants identified evolutionary novelties for plant terrestrialization and land plant heritage genes. C. braunii employs unique xylan synthases for cell wall biosynthesis, a phragmoplast (cell separation) mechanism similar to that of land plants, and many phytohormones. C. braunii plastids are controlled via land-plant-like retrograde signaling, and transcriptional regulation is more elaborate than in other algae. The morphological complexity of this organism may result from expanded gene families, with three cases of particular note: genes effecting tolerance to reactive oxygen species (ROS), LysM receptor-like kinases, and transcription factors (TFs). Transcriptomic analysis of sexual reproductive structures reveals intricate control by TFs, activity of the ROS gene network, and the ancestral use of plant-like storage and stress protection proteins in the zygote.} } @Article{IPB-798, author = {Krägeloh, T. and Cavalleri, J. M. V. and Ziegler, J. and Sander, J. and Terhardt, M. and Breves, G. and Cehak, A. and}, title = {{Identification of hypoglycin A binding adsorbents as potential preventive measures in co-grazers of atypical myopathy affected horses}}, year = {2018}, pages = {220-227}, journal = {Equine Vet. J.}, doi = {10.1111/evj.12723}, volume = {50}, abstract = {BackgroundIntestinal absorption of hypoglycin A (HGA) and its metabolism are considered major prerequisites for atypical myopathy (AM). The increasing incidence and the high mortality rate of AM urgently necessitate new therapeutic and/or preventative approaches.ObjectivesTo identify a substance for oral administration capable of binding HGA in the intestinal lumen and effectively reducing the intestinal absorption of the toxin.Study designExperimental in vitro study.MethodsSubstances commonly used in equine practice (activated charcoal composition, di‐tri‐octahedral smectite, mineral oil and activated charcoal) were tested for their binding capacity for HGA using an in vitro incubation method. The substance most effective in binding HGA was subsequently tested for its potential to reduce intestinal HGA absorption. Jejunal tissues of 6 horses were incubated in Ussing chambers to determine mucosal uptake, tissue accumulation, and serosal release of HGA in the presence and absence of the target substance. Potential intestinal metabolism in methylenecyclopropyl acetic acid (MCPA)‐conjugates was investigated by analysing their concentrations in samples from the Ussing chambers.ResultsActivated charcoal composition and activated charcoal were identified as potent HGA binding substances with dose and pH dependent binding capacity. There was no evidence of intestinal HGA metabolism.Main limitationsBinding capacity of adsorbents was tested in vitro using aqueous solutions, and in vivo factors such as transit time and composition of intestinal content, may affect adsorption capacity after oral administration.ConclusionsFor the first time, this study identifies substances capable of reducing HGA intestinal absorption. This might have major implications as a preventive measure in cograzers of AM affected horses but also in horses at an early stage of intoxication.} } @Article{IPB-796, author = {Jung, J.-Y. and Ried, M. K. and Hothorn, M. and Poirier, Y. and}, title = {{Control of plant phosphate homeostasis by inositol pyrophosphates and the SPX domain}}, year = {2018}, pages = {156-162}, journal = {Curr. Opin. Biotech.}, doi = {10.1016/j.copbio.2017.08.012}, volume = {49}, abstract = {Proteins containing a SPX domain are involved in phosphate (Pi) homeostasis, including Pi transport and adaptation to Pi deficiency. The SPX domain harbors a basic surface binding Pi at low affinity and inositol pyrophosphates (PP-InsPs) at high affinity. Genetic and biochemical studies revealed that PP-InsPs serve as ligands for the SPX domain. Residues in the PHO1 SPX domain involved in PP-InsPs binding are critical for its Pi export activity, and the interaction between SPX proteins and the PHR1 transcription factor, which results in PHR1 inactivation, is promoted by PP-InsPs. Changes in PP-InsPs levels in response to Pi deficiency may thus contribute to the adaptation of plants to stress via the modulation of the activity of SPX-containing proteins and their interactors. Modulating PP-InsP levels or the affinity/specificity of the SPX domain for PP-InsP could potentially be used to engineer crops to maintain high yield under reduced Pi fertilizer input.} } @Article{IPB-795, author = {Jablonická, V. and Ziegler, J. and Vatehová, Z. and Lišková, D. and Heilmann, I. and Obložinský, M. and Heilmann, M. and}, title = {{Inhibition of phospholipases influences the metabolism of wound-induced benzylisoquinoline alkaloids in Papaver somniferum L.}}, year = {2018}, pages = {1-8}, journal = {J. Plant Physiol.}, doi = {10.1016/j.jplph.2018.01.007}, volume = {223}, abstract = {Benzylisoquinoline alkaloids (BIAs) are important secondary plant metabolites and include medicinally relevant drugs, such as morphine or codeine. As the de novo synthesis of BIA backbones is (still) unfeasible, to date the opium poppy plant Papaver somniferum L. represents the main source of BIAs. The formation of BIAs is induced in poppy plants by stress conditions, such as wounding or salt treatment; however, the details about regulatory processes controlling BIA formation in opium poppy are not well studied. Environmental stresses, such as wounding or salinization, are transduced in plants by phospholipid-based signaling pathways, which involve different classes of phospholipases. Here we investigate whether pharmacological inhibition of phospholipase A2 (PLA2, inhibited by aristolochic acid (AA)) or phospholipase D (PLD; inhibited by 5-fluoro-2-indolyl des-chlorohalopemide (FIPI)) in poppy plants influences wound-induced BIA accumulation and the expression of key biosynthetic genes. We show that inhibition of PLA2 results in increased morphinan biosynthesis concomitant with reduced production of BIAs of the papaverine branch, whereas inhibition of PLD results in increased production of BIAs of the noscapine branch. The data suggest that phospholipid-dependent signaling pathways contribute to the activation of morphine biosynthesis at the expense of the production of other BIAs in poppy plants. A better understanding of the effectors and the principles of regulation of alkaloid biosynthesis might be the basis for the future genetic modification of opium poppy to optimize BIA production.} } @Article{IPB-794, author = {Iglesias, M. J. and Terrile, M. C. and Correa-Aragunde, N. and Colman, S. L. and Izquierdo-Álvarez, A. and Fiol, D. F. and París, R. and Sánchez-López, N. and Marina, A. and Calderón Villalobos, L. I. A. and Estelle, M. and Lamattina, L. and Martínez-Ruiz, A. and Casalongué, C. A. and}, title = {{Regulation of SCFTIR1/AFBs E3 ligase assembly by S-nitrosylation of Arabidopsis SKP1-like1 impacts on auxin signaling}}, year = {2018}, pages = {200-210}, journal = {Redox Biol.}, doi = {10.1016/j.redox.2018.07.003}, volume = {18}, abstract = {The F-box proteins (FBPs) TIR1/AFBs are the substrate recognition subunits of SKP1–cullin–F-box (SCF) ubiquitin ligase complexes and together with Aux/IAAs form the auxin co-receptor. Although tremendous knowledge on auxin perception and signaling has been gained in the last years, SCFTIR1/AFBs complex assembly and stabilization are emerging as new layers of regulation. Here, we investigated how nitric oxide (NO), through S-nitrosylation of ASK1 is involved in SCFTIR1/AFBs assembly. We demonstrate that ASK1 is S-nitrosylated and S-glutathionylated in cysteine (Cys) 37 and Cys118 residues in vitro. Both, in vitro and in vivo protein-protein interaction assays show that NO enhances ASK1 binding to CUL1 and TIR1/AFB2, required for SCFTIR1/AFB2 assembly. In addition, we demonstrate that Cys37 and Cys118 are essential residues for proper activation of auxin signaling pathway in planta. Phylogenetic analysis revealed that Cys37 residue is only conserved in SKP proteins in Angiosperms, suggesting that S-nitrosylation on Cys37 could represent an evolutionary adaption for SKP1 function in flowering plants. Collectively, these findings indicate that multiple events of redox modifications might be part of a fine-tuning regulation of SCFTIR1/AFBs for proper auxin signal transduction.} } @Article{IPB-785, author = {García, M. L. and Bó, E. D. and da Graça, J. V. and Gago-Zachert, S. and Hammond, J. and Moreno, P. and Natsuaki, T. and Pallás, V. and Navarro, J. A. and Reyes, C. A. and Luna, G. R. and Sasaya, T. and Tzanetakis, I. E. and Vaira, A. M. and Verbeek, M. and ICTV Report Consortium, . and}, title = {{Corrigendum: ICTV Virus Taxonomy Profile: Ophioviridae}}, year = {2018}, pages = {949-949}, journal = {J. Gen. Virol.}, doi = {10.1099/jgv.0.001093}, volume = {99}, } @Article{IPB-784, author = {Gantner, J. and Ordon, J. and Ilse, T. and Kretschmer, C. and Gruetzner, R. and Löfke, C. and Dagdas, Y. and Bürstenbinder, K. and Marillonnet, S. and Stuttmann, J. and}, title = {{Peripheral infrastructure vectors and an extended set of plant parts for the Modular Cloning system}}, year = {2018}, pages = {e0197185}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0197185}, volume = {13}, abstract = {Standardized DNA assembly strategies facilitate the generation of multigene constructs from collections of building blocks in plant synthetic biology. A common syntax for hierarchical DNA assembly following the Golden Gate principle employing Type IIs restriction endonucleases was recently developed, and underlies the Modular Cloning and GoldenBraid systems. In these systems, transcriptional units and/or multigene constructs are assembled from libraries of standardized building blocks, also referred to as phytobricks, in several hierarchical levels and by iterative Golden Gate reactions. Here, a toolkit containing further modules for the novel DNA assembly standards was developed. Intended for use with Modular Cloning, most modules are also compatible with GoldenBraid. Firstly, a collection of approximately 80 additional phytobricks is provided, comprising e.g. modules for inducible expression systems, promoters or epitope tags. Furthermore, DNA modules were developed for connecting Modular Cloning and Gateway cloning, either for toggling between systems or for standardized Gateway destination vector assembly. Finally, first instances of a “peripheral infrastructure” around Modular Cloning are presented: While available toolkits are designed for the assembly of plant transformation constructs, vectors were created to also use coding sequence-containing phytobricks directly in yeast two hybrid interaction or bacterial infection assays. The presented material will further enhance versatility of hierarchical DNA assembly strategies.} } @Article{IPB-760, author = {Bochnia, M. and Scheidemann, W. and Ziegler, J. and Sander, J. and Vollstedt, S. and Glatter, M. and Janzen, N. and Terhardt, M. and Zeyner, A. and}, title = {{Predictive value of hypoglycin A and methylencyclopropylacetic acid conjugates in a horse with atypical myopathy in comparison to its cograzing partners}}, year = {2018}, pages = {24-28}, journal = {Equine Vet. Educ.}, doi = {10.1111/eve.12596}, volume = {30}, abstract = {Hypoglycin A (HGA) was detected in blood and urine of a horse suffering from atypical myopathy (AM; Day 2, serum, 8290 μg/l; urine: Day 1, 574, Day 2, 742 μg/l) and in its cograzing partners with a high variability (46–1570 μg/l serum). Over the period of disease, the level of the toxic metabolites (methylencyclopropylacetic acid [MCPA]‐conjugates) increased in body fluids of the AM horse (MCPA‐carnitine: Day 2, 0.246, Day 3, 0.581 μmol/l serum; MCPA‐carnitine: Day 2, 0.621, Day 3, 0.884 μmol/mmol creatinine in urine) and HGA decreased rapidly (Day 3, 2430 μg/l serum). In cograzing horses MCPA‐conjugates were not detected. HGA in seeds ranged from 268 to 367 μg/g. Although HGA was present in body fluids of healthy cograzing horses, MCPA‐conjugates were not detectable, in contrast to the AM horse. Therefore, increasing concentrations of MCPA‐conjugates are supposed to be linked with the onset of AM and both parameters seem to indicate the clinical stage of disease. However, detection of HGA in body fluids of cograzing horses might be a promising step in preventing the disease.} } @Article{IPB-757, author = {Bagchi, R. and Melnyk, C. W. and Christ, G. and Winkler, M. and Kirchsteiner, K. and Salehin, M. and Mergner, J. and Niemeyer, M. and Schwechheimer, C. and Calderón Villalobos, L. I. A. and Estelle, M. and}, title = {{The Arabidopsis ALF4 protein is a regulator of SCF E3 ligases}}, year = {2018}, pages = {255-268}, journal = {EMBO J.}, doi = {10.15252/embj.201797159}, volume = {37}, abstract = {The cullin‐RING E3 ligases (CRLs) regulate diverse cellular processes in all eukaryotes. CRL activity is controlled by several proteins or protein complexes, including NEDD8, CAND1, and the CSN. Recently, a mammalian protein called Glomulin (GLMN) was shown to inhibit CRLs by binding to the RING BOX (RBX1) subunit and preventing binding to the ubiquitin‐conjugating enzyme. Here, we show that Arabidopsis ABERRANT LATERAL ROOT FORMATION4 (ALF4) is an ortholog of GLMN. The alf4 mutant exhibits a phenotype that suggests defects in plant hormone response. We show that ALF4 binds to RBX1 and inhibits the activity of SCFTIR1, an E3 ligase responsible for degradation of the Aux/IAA transcriptional repressors. In vivo, the alf4 mutation destabilizes the CUL1 subunit of the SCF. Reduced CUL1 levels are associated with increased levels of the Aux/IAA proteins as well as the DELLA repressors, substrate of SCFSLY1. We propose that the alf4 phenotype is partly due to increased levels of the Aux/IAA and DELLA proteins.} } @Article{IPB-793, author = {Ibañez, C. and Delker, C. and Martinez, C. and Bürstenbinder, K. and Janitza, P. and Lippmann, R. and Ludwig, W. and Sun, H. and James, G. V. and Klecker, M. and Grossjohann, A. and Schneeberger, K. and Prat, S. and Quint, M. and}, title = {{Brassinosteroids Dominate Hormonal Regulation of Plant Thermomorphogenesis via BZR1}}, year = {2018}, pages = {303-310.e3}, journal = {Curr. Biol.}, doi = {10.1016/j.cub.2017.11.077}, volume = {28}, abstract = {Thermomorphogenesis is defined as the suite of morphological changes that together are likely to contribute to adaptive growth acclimation to usually elevated ambient temperature [1, 2]. While many details of warmth-induced signal transduction are still elusive, parallels to light signaling recently became obvious (reviewed in [3]). It involves photoreceptors that can also sense changes in ambient temperature [3, 4, 5] and act, for example, by repressing protein activity of the central integrator of temperature information PHYTOCHROME-INTERACTING FACTOR 4 (PIF4 [6]). In addition, PIF4 transcript accumulation is tightly controlled by the evening complex member EARLY FLOWERING 3 [7, 8]. According to the current understanding, PIF4 activates growth-promoting genes directly but also via inducing auxin biosynthesis and signaling, resulting in cell elongation. Based on a mutagenesis screen in the model plant Arabidopsis thaliana for mutants with defects in temperature-induced hypocotyl elongation, we show here that both PIF4 and auxin function depend on brassinosteroids. Genetic and pharmacological analyses place brassinosteroids downstream of PIF4 and auxin. We found that brassinosteroids act via the transcription factor BRASSINAZOLE RESISTANT 1 (BZR1), which accumulates in the nucleus at high temperature, where it induces expression of growth-promoting genes. Furthermore, we show that at elevated temperature BZR1 binds to the promoter of PIF4, inducing its expression. These findings suggest that BZR1 functions in an amplifying feedforward loop involved in PIF4 activation. Although numerous negative regulators of PIF4 have been described, we identify BZR1 here as a true temperature-dependent positive regulator of PIF4, acting as a major growth coordinator.} } @Article{IPB-884, author = {García, M. L. and Bó, E. D. and da Graça, J. V. and Gago-Zachert, S. and Hammond, J. and Moreno, P. and Natsuaki, T. and Pallás, V. and Navarro, J. A. and Reyes, C. A. and Luna, G. R. and Sasaya, T. and Tzanetakis, I. E. and Vaira, A. M. and Verbeek, M. and ICTV Report Consortium, . and}, title = {{ICTV Virus Taxonomy Profile: Ophioviridae}}, year = {2017}, pages = {1161-1162}, journal = {J. Gen. Virol.}, doi = {10.1099/jgv.0.000836}, volume = {98}, abstract = {The Ophioviridae is a family of filamentous plant viruses, with single-stranded negative, and possibly ambisense, RNA genomes of 11.3–12.5 kb divided into 3–4 segments, each encapsidated separately. Virions are naked filamentous nucleocapsids, forming kinked circles of at least two different contour lengths. The sole genus, Ophiovirus, includes seven species. Four ophioviruses are soil-transmitted and their natural hosts include trees, shrubs, vegetables and bulbous or corm-forming ornamentals, both monocots and dicots. This is a summary of the International Committee on Taxonomy of Viruses (ICTV) Report on the taxonomy of the Ophioviridae, which is available at http://www.ictv.global/report/ophioviridae.} } @Article{IPB-868, author = {Bürstenbinder, K. and Möller, B. and Plötner, R. and Stamm, G. and Hause, G. and Mitra, D. and Abel, S. and}, title = {{The IQD Family of Calmodulin-Binding Proteins Links Calcium Signaling to Microtubules, Membrane Subdomains, and the Nucleus}}, year = {2017}, pages = {1692-1708}, journal = {Plant Physiol.}, doi = {10.1104/pp.16.01743}, volume = {173}, abstract = {Calcium (Ca2\+) signaling and dynamic reorganization of the cytoskeleton are essential processes for the coordination and control of plant cell shape and cell growth. Calmodulin (CaM) and closely related calmodulin-like (CML) polypeptides are principal sensors of Ca2\+ signals. CaM/CMLs decode and relay information encrypted by the second messenger via differential interactions with a wide spectrum of targets to modulate their diverse biochemical activities. The plant-specific IQ67 DOMAIN (IQD) family emerged as possibly the largest class of CaM-interacting proteins with undefined molecular functions and biological roles. Here, we show that the 33 members of the IQD family in Arabidopsis (Arabidopsis thaliana) differentially localize, using green fluorescent protein (GFP)-tagged proteins, to multiple and distinct subcellular sites, including microtubule (MT) arrays, plasma membrane subdomains, and nuclear compartments. Intriguingly, the various IQD-specific localization patterns coincide with the subcellular patterns of IQD-dependent recruitment of CaM, suggesting that the diverse IQD members sequester Ca2\+-CaM signaling modules to specific subcellular sites for precise regulation of Ca2\+-dependent processes. Because MT localization is a hallmark of most IQD family members, we quantitatively analyzed GFP-labeled MT arrays in Nicotiana benthamiana cells transiently expressing GFP-IQD fusions and observed IQD-specific MT patterns, which point to a role of IQDs in MT organization and dynamics. Indeed, stable overexpression of select IQD proteins in Arabidopsis altered cellular MT orientation, cell shape, and organ morphology. Because IQDs share biochemical properties with scaffold proteins, we propose that IQD families provide an assortment of platform proteins for integrating CaM-dependent Ca2\+ signaling at multiple cellular sites to regulate cell function, shape, and growth.} } @Article{IPB-867, author = {Bürstenbinder, K. and Mitra, D. and Quegwer, J. and}, title = {{Functions of IQD proteins as hubs in cellular calcium and auxin signaling: A toolbox for shape formation and tissue-specification in plants?}}, year = {2017}, pages = {e1331198}, journal = {Plant Signal Behav.}, doi = {10.1080/15592324.2017.1331198}, volume = {12}, abstract = {Calcium (Ca2\+) ions play pivotal roles as second messengers in intracellular signal transduction, and coordinate many biological processes. Changes in intracellular Ca2\+ levels are perceived by Ca2\+ sensors such as calmodulin (CaM) and CaM-like (CML) proteins, which transduce Ca2\+ signals into cellular responses by regulation of diverse target proteins. Insights into molecular functions of CaM targets are thus essential to understand the molecular and cellular basis of Ca2\+ signaling. During the last decade, IQ67-domain (IQD) proteins emerged as the largest class of CaM targets in plants with mostly unknown functions. In the March issue of Plant Physiology, we presented the first comprehensive characterization of the 33-membered IQD family in Arabidopsis thaliana. We showed, by analysis of the subcellular localization of translational green fluorescent protein (GFP) fusion proteins, that most IQD members label microtubules (MTs), and additionally often localize to the cell nucleus or to membranes, where they recruit CaM Ca2\+ sensors. Important functions at MTs are supported by altered MT organization and plant growth in IQD gain-of-function lines. Because IQD proteins share structural hallmarks of scaffold proteins, we propose roles of IQDs in the assembly of macromolecular complexes to orchestrate Ca2\+ CaM signaling from membranes to the nucleus. Interestingly, expression of several IQDs is regulated by auxin, which suggests functions of IQDs as hubs in cellular auxin and calcium signaling to regulate plant growth and development.} } @Article{IPB-862, author = {Balzergue, C. and Dartevelle, T. and Godon, C. and Laugier, E. and Meisrimler, C. and Teulon, J.-M. and Creff, A. and Bissler, M. and Brouchoud, C. and Hagège, A. and Müller, J. and Chiarenza, S. and Javot, H. and Becuwe-Linka, N. and David, P. and Péret, B. and Delannoy, E. and Thibaud, M.-C. and Armengaud, J. and Abel, S. and Pellequer, J.-L. and Nussaume, L. and Desnos, T. and}, title = {{Low phosphate activates STOP1-ALMT1 to rapidly inhibit root cell elongation}}, year = {2017}, pages = {15300}, journal = {Nat. Commun.}, doi = {10.1038/ncomms15300}, volume = {8}, abstract = {Environmental cues profoundly modulate cell proliferation and cell elongation to inform and direct plant growth and development. External phosphate (Pi) limitation inhibits primary root growth in many plant species. However, the underlying Pi sensory mechanisms are unknown. Here we genetically uncouple two Pi sensing pathways in the root apex of Arabidopsis thaliana. First, the rapid inhibition of cell elongation in the transition zone is controlled by transcription factor STOP1, by its direct target, ALMT1, encoding a malate channel, and by ferroxidase LPR1, which together mediate Fe and peroxidase-dependent cell wall stiffening. Second, during the subsequent slow inhibition of cell proliferation in the apical meristem, which is mediated by LPR1-dependent, but largely STOP1–ALMT1-independent, Fe and callose accumulate in the stem cell niche, leading to meristem reduction. Our work uncovers STOP1 and ALMT1 as a signalling pathway of low Pi availability and exuded malate as an unexpected apoplastic inhibitor of root cell wall expansion.} } @Article{IPB-914, author = {Möller, B. and Poeschl, Y. and Plötner, R. and Bürstenbinder, K. and}, title = {{PaCeQuant: A Tool for High-Throughput Quantification of Pavement Cell Shape Characteristics}}, year = {2017}, pages = {998-1017}, journal = {Plant Physiol.}, doi = {10.1104/pp.17.00961}, volume = {175}, abstract = {Pavement cells (PCs) are the most frequently occurring cell type in the leaf epidermis and play important roles in leaf growth and function. In many plant species, PCs form highly complex jigsaw-puzzle-shaped cells with interlocking lobes. Understanding of their development is of high interest for plant science research because of their importance for leaf growth and hence for plant fitness and crop yield. Studies of PC development, however, are limited, because robust methods are lacking that enable automatic segmentation and quantification of PC shape parameters suitable to reflect their cellular complexity. Here, we present our new ImageJ-based tool, PaCeQuant, which provides a fully automatic image analysis workflow for PC shape quantification. PaCeQuant automatically detects cell boundaries of PCs from confocal input images and enables manual correction of automatic segmentation results or direct import of manually segmented cells. PaCeQuant simultaneously extracts 27 shape features that include global, contour-based, skeleton-based, and PC-specific object descriptors. In addition, we included a method for classification and analysis of lobes at two-cell junctions and three-cell junctions, respectively. We provide an R script for graphical visualization and statistical analysis. We validated PaCeQuant by extensive comparative analysis to manual segmentation and existing quantification tools and demonstrated its usability to analyze PC shape characteristics during development and between different genotypes. PaCeQuant thus provides a platform for robust, efficient, and reproducible quantitative analysis of PC shape characteristics that can easily be applied to study PC development in large data sets.} } @Article{IPB-908, author = {López-Carrasco, A. and Ballesteros, C. and Sentandreu, V. and Delgado, S. and Gago-Zachert, S. and Flores, R. and Sanjuán, R. and}, title = {{Different rates of spontaneous mutation of chloroplastic and nuclear viroids as determined by high-fidelity ultra-deep sequencing}}, year = {2017}, pages = {e1006547}, journal = {PLOS Pathog.}, doi = {10.1371/journal.ppat.1006547}, volume = {13}, abstract = {Mutation rates vary by orders of magnitude across biological systems, being higher for simpler genomes. The simplest known genomes correspond to viroids, subviral plant replicons constituted by circular non-coding RNAs of few hundred bases. Previous work has revealed an extremely high mutation rate for chrysanthemum chlorotic mottle viroid, a chloroplast-replicating viroid. However, whether this is a general feature of viroids remains unclear. Here, we have used high-fidelity ultra-deep sequencing to determine the mutation rate in a common host (eggplant) of two viroids, each representative of one family: the chloroplastic eggplant latent viroid (ELVd, Avsunviroidae) and the nuclear potato spindle tuber viroid (PSTVd, Pospiviroidae). This revealed higher mutation frequencies in ELVd than in PSTVd, as well as marked differences in the types of mutations produced. Rates of spontaneous mutation, quantified in vivo using the lethal mutation method, ranged from 1/1000 to 1/800 for ELVd and from 1/7000 to 1/3800 for PSTVd depending on sequencing run. These results suggest that extremely high mutability is a common feature of chloroplastic viroids, whereas the mutation rates of PSTVd and potentially other nuclear viroids appear significantly lower and closer to those of some RNA viruses.} } @Article{IPB-906, author = {Liu, S. and Ziegler, J. and Zeier, J. and Birkenbihl, R. P. and Somssich, I. E. and}, title = {{Botrytis cinerea B05.10 promotes disease development in Arabidopsis by suppressing WRKY33-mediated host immunity}}, year = {2017}, pages = {2189-2206}, journal = {Plant Cell Environ.}, doi = {10.1111/pce.13022}, volume = {40}, abstract = {The large WRKY transcription factor family is mainly involved in regulating plant immune responses. Arabidopsis WRKY33 is a key transcriptional regulator of hormonal and metabolic processes towards Botrytis cinerea strain 2100 infection and is essential for resistance. In contrast to B. cinerea strain 2100, the strain B05.10 is virulent on wild‐type (WT) Col‐0 Arabidopsis plants highlighting the genetic diversity within this pathogen species. We analysed how early WRKY33‐dependent responses are affected upon infection with strain B05.10 and found that most of these responses were strongly dampened during this interaction. Ectopic expression of WRKY33 resulted in complete resistance towards this strain indicating that virulence of B05.10, at least partly, depends on suppressing WRKY33 expression/protein accumulation. As a consequence, the expression levels of direct WRKY33 target genes, including those involved in the biosynthesis of camalexin, were also reduced upon infection. Concomitantly, elevated levels of the phytohormone abscisic acid (ABA) were observed. Molecular and genetic studies revealed that ABA negatively influences defence to B05.10 and effects jasmonic acid/ethylene (JA/ET) and salicylic acid (SA) levels. Susceptibility/resistance was determined by the antagonistic effect of ABA on JA, and this crosstalk required suppressing WRKY33 functions at early infection stages. This indicates that B. cinerea B05.10 promotes disease by suppressing WRKY33‐mediated host defences.} } @Article{IPB-960, author = {Ziegler, J. and Schmidt, S. and Strehmel, N. and Scheel, D. and Abel, S. and}, title = {{Arabidopsis Transporter ABCG37/PDR9 contributes primarily highly oxygenated Coumarins to Root Exudation}}, year = {2017}, pages = {3704}, journal = {Sci. Rep.}, doi = {10.1038/s41598-017-03250-6}, volume = {7}, abstract = {The chemical composition of root exudates strongly impacts the interactions of plants with microorganisms in the rhizosphere and the efficiency of nutrient acquisition. Exudation of metabolites is in part mediated by ATP-binding cassette (ABC) transporters. In order to assess the contribution of individual ABC transporters to root exudation, we performed an LC-MS based non-targeted metabolite profiling of semi-polar metabolites accumulating in root exudates of Arabidopsis thaliana plants and mutants deficient in the expression of ABCG36 (PDR8/PEN3), ABCG37 (PDR9) or both transporters. Comparison of the metabolite profiles indicated distinct roles for each ABC transporter in root exudation. Thymidine exudation could be attributed to ABCG36 function, whereas coumarin exudation was strongly reduced only in ABCG37 deficient plants. However, coumarin exudation was compromised in abcg37 mutants only with respect to certain metabolites of this substance class. The specificity of ABCG37 for individual coumarins was further verified by a targeted LC-MS based coumarin profiling method. The response to iron deficiency, which is known to strongly induce coumarin exudation, was also investigated. In either treatment, the distribution of individual coumarins between roots and exudates in the investigated genotypes suggested the involvement of ABCG37 in the exudation specifically of highly oxygenated rather than monohydroxylated coumarins.} } @Article{IPB-957, author = {Winkler, M. and Niemeyer, M. and Hellmuth, A. and Janitza, P. and Christ, G. and Samodelov, S. L. and Wilde, V. and Majovsky, P. and Trujillo, M. and Zurbriggen, M. D. and Hoehenwarter, W. and Quint, M. and Calderón Villalobos, L. I. A. and}, title = {{Variation in auxin sensing guides AUX/IAA transcriptional repressor ubiquitylation and destruction}}, year = {2017}, pages = {15706}, journal = {Nat. Commun.}, doi = {10.1038/ncomms15706}, volume = {8}, abstract = {Auxin is a small molecule morphogen that bridges SCFTIR1/AFB-AUX/IAA co-receptor interactions leading to ubiquitylation and proteasome-dependent degradation of AUX/IAA transcriptional repressors. Here, we systematically dissect auxin sensing by SCFTIR1-IAA6 and SCFTIR1-IAA19 co-receptor complexes, and assess IAA6/IAA19 ubiquitylation in vitro and IAA6/IAA19 degradation in vivo. We show that TIR1-IAA19 and TIR1-IAA6 have distinct auxin affinities that correlate with ubiquitylation and turnover dynamics of the AUX/IAA. We establish a system to track AUX/IAA ubiquitylation in IAA6 and IAA19 in vitro and show that it occurs in flexible hotspots in degron-flanking regions adorned with specific Lys residues. We propose that this signature is exploited during auxin-mediated SCFTIR1-AUX/IAA interactions. We present evidence for an evolving AUX/IAA repertoire, typified by the IAA6/IAA19 ohnologues, that discriminates the range of auxin concentrations found in plants. We postulate that the intrinsic flexibility of AUX/IAAs might bias their ubiquitylation and destruction kinetics enabling specific auxin responses.} } @Article{IPB-951, author = {Wasternack, C. and}, title = {{The Trojan horse coronatine: the COI1-JAZ2-MYC2,3,4-ANAC019,055,072 module in stomata dynamics upon bacterial infection}}, year = {2017}, pages = {972-975}, journal = {New Phytol.}, doi = {10.1111/nph.14417}, volume = {213}, abstract = {This article is a Commentary on Gimenez‐Ibanez et al., 213: 1378–1392.} } @Article{IPB-950, author = {Wasternack, C. and Song, S. and}, title = {{Jasmonates: biosynthesis, metabolism, and signaling by proteins activating and repressing transciption}}, year = {2017}, pages = {1303-1321}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/erw443}, volume = {68}, abstract = {The lipid-derived phytohormone jasmonate (JA) regulates plant growth, development, secondary metabolism, defense against insect attack and pathogen infection, and tolerance to abiotic stresses such as wounding, UV light, salt, and drought. JA was first identified in 1962, and since the 1980s many studies have analyzed the physiological functions, biosynthesis, distribution, metabolism, perception, signaling, and crosstalk of JA, greatly expanding our knowledge of the hormone’s action. In response to fluctuating environmental cues and transient endogenous signals, the occurrence of multilayered organization of biosynthesis and inactivation of JA, and activation and repression of the COI1–JAZ-based perception and signaling contributes to the fine-tuning of JA responses. This review describes the JA biosynthetic enzymes in terms of gene families, enzymatic activity, location and regulation, substrate specificity and products, the metabolic pathways in converting JA to activate or inactivate compounds, JA signaling in perception, and the co-existence of signaling activators and repressors.} } @Article{IPB-949, author = {Wasternack, C. and}, title = {{A plant\'s balance of growth and defense - revisited}}, year = {2017}, pages = {1291-1294}, journal = {New Phytol.}, doi = {10.1111/nph.14720}, volume = {215}, abstract = {This article is a Commentary on Major et al., 215: 1533–1547.} } @Article{IPB-945, author = {Trenner, J. and Poeschl, Y. and Grau, J. and Gogol-Döring, A. and Quint, M. and Delker, C. and}, title = {{Auxin-induced expression divergence between Arabidopsis species may originate within the TIR1/AFB–AUX/IAA–ARF module}}, year = {2017}, pages = {539-552}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/erw457}, volume = {68}, abstract = {Auxin is an essential regulator of plant growth and development, and auxin signaling components are conserved among land plants. Yet, a remarkable degree of natural variation in physiological and transcriptional auxin responses has been described among Arabidopsis thaliana accessions. As intraspecies comparisons offer only limited genetic variation, we here inspect the variation of auxin responses between A. thaliana and A. lyrata. This approach allowed the identification of conserved auxin response genes including novel genes with potential relevance for auxin biology. Furthermore, promoter divergences were analyzed for putative sources of variation. De novo motif discovery identified novel and variants of known elements with potential relevance for auxin responses, emphasizing the complex, and yet elusive, code of element combinations accounting for the diversity in transcriptional auxin responses. Furthermore, network analysis revealed correlations of interspecies differences in the expression of AUX/IAA gene clusters and classic auxin-related genes. We conclude that variation in general transcriptional and physiological auxin responses may originate substantially from functional or transcriptional variations in the TIR1/AFB, AUX/IAA, and ARF signaling network. In that respect, AUX/IAA gene expression divergence potentially reflects differences in the manner in which different species transduce identical auxin signals into gene expression responses.} } @Article{IPB-855, author = {Abel, S. and}, title = {{Phosphate scouting by root tips}}, year = {2017}, pages = {168-177}, journal = {Curr. Opin. Plant Biol.}, doi = {10.1016/j.pbi.2017.04.016}, volume = {39}, abstract = {Chemistry assigns phosphate (Pi) dominant roles in metabolism; however, it also renders the macronutrient a genuinely limiting factor of plant productivity. Pi bioavailability is restricted by low Pi mobility in soil and antagonized by metallic toxicities, which force roots to actively seek and selectively acquire the vital element. During the past few years, a first conceptual outline has emerged of the sensory mechanisms at root tips, which monitor external Pi and transmit the edaphic cue to inform root development. This review highlights new aspects of the Pi acquisition strategy of Arabidopsis roots, as well as a framework of local Pi sensing in the context of antagonistic interactions between Pi and its major associated metallic cations, Fe3\+ and Al3\+.} } @Article{IPB-893, author = {Ibañez, C. and Poeschl, Y. and Peterson, T. and Bellstädt, J. and Denk, K. and Gogol-Döring, A. and Quint, M. and Delker, C. and}, title = {{Ambient temperature and genotype differentially affect developmental and phenotypic plasticity in Arabidopsis thaliana}}, year = {2017}, pages = {114}, journal = {BMC Plant Biol.}, doi = {10.1186/s12870-017-1068-5}, volume = {17}, abstract = {BackgroundGlobal increase in ambient temperatures constitute a significant challenge to wild and cultivated plant species. Forward genetic analyses of individual temperature-responsive traits have resulted in the identification of several signaling and response components. However, a comprehensive knowledge about temperature sensitivity of different developmental stages and the contribution of natural variation is still scarce and fragmented at best.ResultsHere, we systematically analyze thermomorphogenesis throughout a complete life cycle in ten natural Arabidopsis thaliana accessions grown under long day conditions in four different temperatures ranging from 16 to 28 °C. We used Q10, GxE, phenotypic divergence and correlation analyses to assess temperature sensitivity and genotype effects of more than 30 morphometric and developmental traits representing five phenotype classes. We found that genotype and temperature differentially affected plant growth and development with variing strengths. Furthermore, overall correlations among phenotypic temperature responses was relatively low which seems to be caused by differential capacities for temperature adaptations of individual accessions.ConclusionGenotype-specific temperature responses may be attractive targets for future forward genetic approaches and accession-specific thermomorphogenesis maps may aid the assessment of functional relevance of known and novel regulatory components.} } @Article{IPB-1017, author = {López-Carrasco, A. and Gago-Zachert, S. and Mileti, G. and Minoia, S. and Flores, R. and Delgado, S. and}, title = {{The transcription initiation sites of eggplant latent viroid strands map within distinct motifs in their in vivo RNA conformations}}, year = {2016}, pages = {83-97}, journal = {RNA Biol.}, doi = {10.1080/15476286.2015.1119365}, volume = {13}, abstract = {Eggplant latent viroid (ELVd), like other members of family Avsunviroidae, replicates in plastids through a symmetric rolling-circle mechanism in which elongation of RNA strands is most likely catalyzed by a nuclear-encoded polymerase (NEP) translocated to plastids. Here we have addressed where NEP initiates transcription of viroid strands. Because this step is presumably directed by sequence/structural motifs, we have previously determined the conformation of the monomeric linear (\+) and (−) RNAs of ELVd resulting from hammerhead-mediated self-cleavage. In silico predictions with 3 softwares led to similar bifurcated conformations for both ELVd strands. In vitro examination by non-denaturing PAGE showed that they migrate as prominent single bands, with the ELVd (\+) RNA displaying a more compact conformation as revealed by its faster electrophoretic mobility. In vitro SHAPE analysis corroborated the ELVd conformations derived from thermodynamics-based predictions in silico. Moreover, sequence analysis of 94 full-length natural ELVd variants disclosed co-variations, and mutations converting canonical into wobble pairs or vice versa, which confirmed in vivo most of the stems predicted in silico and in vitro, and additionally helped to introduce minor structural refinements. Therefore, results from the 3 experimental approaches were essentially consistent among themselves. Application to RNA preparations from ELVd-infected tissue of RNA ligase-mediated rapid amplification of cDNA ends, combined with pretreatments to modify the 5′ ends of viroid strands, mapped the transcription initiation sites of ELVd (\+) and (−) strands in vivo at different sequence/structural motifs, in contrast with the situation previously observed in 2 other members of the family Avsunviroidae.} } @Article{IPB-1011, author = {Kowalski, A. M. and Gooding, M. and Ferrante, A. and Slafer, G. A. and Orford, S. and Gasperini, D. and Griffiths, S. and}, title = {{Agronomic assessment of the wheat semi-dwarfing gene Rht8 in contrasting nitrogen treatments and water regimes}}, year = {2016}, pages = {150-160}, journal = {Field Crops Res.}, doi = {10.1016/j.fcr.2016.02.026}, volume = {191}, abstract = {Reduced height 8 (Rht8) is the main alternative to the GA-insensitive Rht alleles in hot and dry environments where it reduces plant height without yield penalty. The potential of Rht8 in northern-European wheat breeding remains unclear, since the close linkage with the photoperiod-insensitive allele Ppd-D1a is unfavourable in the relatively cool summers. In the present study, two near-isogenic lines (NILs) contrasting for the Rht8/tall allele from Mara in a UK-adapted and photoperiod-sensitive wheat variety were evaluated in trials with varying nitrogen fertiliser (N) treatments and water regimes across sites in the UK and Spain.The Rht8 introgression was associated with a robust height reduction of 11% regardless of N treatment and water regime and the Rht8 NIL was more resistant to root-lodging at agronomically-relevant N levels than the tall NIL. In the UK with reduced solar radiation over the growing season than the site in Spain, the Rht8 NIL showed a 10% yield penalty at standard agronomic N levels due to concomitant reduction in grain number and spike number whereas grain weight and harvest index were not significantly different to the tall NIL. The yield penalty associated with the Rht8 introgression was overcome at low N and in irrigated conditions in the UK, and in the high-temperature site in Spain. Decreased spike length and constant spikelet number in the Rht8 NIL resulted in spike compaction of 15%, independent of N and water regime. The genetic interval of Rht8 overlaps with the compactum gene on 2DS, raising the possibility of the same causative gene. Further genetic dissection of these loci is required.} } @Article{IPB-1004, author = {Hoehenwarter, W. and Mönchgesang, S. and Neumann, S. and Majovsky, P. and Abel, S. and Müller, J. and}, title = {{Comparative expression profiling reveals a role of the root apoplast in local phosphate response}}, year = {2016}, pages = {106}, journal = {BMC Plant Biol.}, doi = {10.1186/s12870-016-0790-8}, volume = {16}, abstract = {BackgroundPlant adaptation to limited phosphate availability comprises a wide range of responses to conserve and remobilize internal phosphate sources and to enhance phosphate acquisition. Vigorous restructuring of root system architecture provides a developmental strategy for topsoil exploration and phosphate scavenging. Changes in external phosphate availability are locally sensed at root tips and adjust root growth by modulating cell expansion and cell division. The functionally interacting Arabidopsis genes, LOW PHOSPHATE RESPONSE 1 and 2 (LPR1/LPR2) and PHOSPHATE DEFICIENCY RESPONSE 2 (PDR2), are key components of root phosphate sensing. We recently demonstrated that the LOW PHOSPHATE RESPONSE 1 - PHOSPHATE DEFICIENCY RESPONSE 2 (LPR1-PDR2) module mediates apoplastic deposition of ferric iron (Fe3\+) in the growing root tip during phosphate limitation. Iron deposition coincides with sites of reactive oxygen species generation and triggers cell wall thickening and callose accumulation, which interfere with cell-to-cell communication and inhibit root growth.ResultsWe took advantage of the opposite phosphate-conditional root phenotype of the phosphate deficiency response 2 mutant (hypersensitive) and low phosphate response 1 and 2 double mutant (insensitive) to investigate the phosphate dependent regulation of gene and protein expression in roots using genome-wide transcriptome and proteome analysis. We observed an overrepresentation of genes and proteins that are involved in the regulation of iron homeostasis, cell wall remodeling and reactive oxygen species formation, and we highlight a number of candidate genes with a potential function in root adaptation to limited phosphate availability. Our experiments reveal that FERRIC REDUCTASE DEFECTIVE 3 mediated, apoplastic iron redistribution, but not intracellular iron uptake and iron storage, triggers phosphate-dependent root growth modulation. We further highlight expressional changes of several cell wall-modifying enzymes and provide evidence for adjustment of the pectin network at sites of iron accumulation in the root.ConclusionOur study reveals new aspects of the elaborate interplay between phosphate starvation responses and changes in iron homeostasis. The results emphasize the importance of apoplastic iron redistribution to mediate phosphate-dependent root growth adjustment and suggest an important role for citrate in phosphate-dependent apoplastic iron transport. We further demonstrate that root growth modulation correlates with an altered expression of cell wall modifying enzymes and changes in the pectin network of the phosphate-deprived root tip, supporting the hypothesis that pectins are involved in iron binding and/or phosphate mobilization.} } @Article{IPB-997, author = {Gharsallah, C. and Fakhfakh, H. and Grubb, D. and Gorsane, F. and}, title = {{Effect of salt stress on ion concentration, proline content, antioxidant enzyme activities and gene expression in tomato cultivars}}, year = {2016}, pages = {plw055}, journal = {AoB PLANTS}, doi = {10.1093/aobpla/plw055}, volume = {8}, abstract = {Salinity is a constraint limiting plant growth and productivity of crops throughout the world. Understanding the mechanism underlying plant response to salinity provides new insights into the improvement of salt tolerance-crops of importance. In the present study, we report on the responses of twenty cultivars of tomato. We have clustered genotypes into scale classes according to their response to increased NaCl levels. Three local tomato genotypes, representative of different saline scale classes, were selected for further investigation. During early (0 h, 6 h and 12 h) and later (7 days) stages of the response to salt treatment, ion concentrations (Na \+ , K \+  and Ca 2\+ ), proline content, enzyme activities (catalase, ascorbate peroxidase and guiacol peroxidase) were recorded. qPCR analysis of candidate genes WRKY (8, 31and 39), ERF (9, 16 and 80), LeNHX (1, 3 and 4) and HKT (class I) were performed. A high K \+ , Ca 2 \+ and proline accumulation as well as a decrease of Na \+  concentration-mediated salt tolerance. Concomitant with a pattern of high-antioxidant enzyme activities, tolerant genotypes also displayed differential patterns of gene expression during the response to salt stress.} } @Article{IPB-995, author = {Gasperini, D. and Acosta, I. F. and Farmer, E. E. and}, title = {{Cotyledon Wounding of Arabidopsis Seedlings}}, year = {2016}, pages = {e1712}, journal = {Bio Protoc.}, doi = {10.21769/BioProtoc.1712}, volume = {6}, abstract = {Damage to plant organs through both biotic and abiotic injury is very common in nature. Arabidopsis thaliana 5-day-old (5-do) seedlings represent an excellent system in which to study plant responses to mechanical wounding, both at the site of the damage and in distal unharmed tissues. Seedlings of wild type, transgenic or mutant lines subjected to single or repetitive cotyledon wounding can be used to quantify morphological alterations (e.g., root length, Gasperini et al., 2015), analyze the dynamics of reporter genes in vivo (Larrieu et al., 2015; Gasperini et al., 2015), follow transcriptional changes by quantitative RT-PCR (Acosta et al., 2013; Gasperini et al., 2015) or examine additional aspects of the wound response with a plethora of downstream procedures. Here we illustrate how to rapidly and reliably wound cotyledons of young seedlings, and show the behavior of two promoters driving the expression of β-glucuronidase (GUS) in entire seedlings and in the primary root meristem, following single or repetitive cotyledon wounding respectively. We describe two procedures that can be easily adapted to specific experimental needs.} } @Article{IPB-994, author = {Gago-Zachert, S. and}, title = {{Viroids, infectious long non-coding RNAs with autonomous replication}}, year = {2016}, pages = {12-24}, journal = {Virus Res.}, doi = {10.1016/j.virusres.2015.08.018}, volume = {212}, abstract = {Transcriptome deep-sequencing studies performed during the last years confirmed that the vast majority of the RNAs transcribed in higher organisms correspond to several types of non-coding RNAs including long non-coding RNAs (lncRNAs). The study of lncRNAs and the identification of their functions, is still an emerging field in plants but the characterization of some of them indicate that they play an important role in crucial regulatory processes like flowering regulation, and responses to abiotic stress and plant hormones. A second group of lncRNAs present in plants is formed by viroids, exogenous infectious subviral plant pathogens well known since many years. Viroids are composed of circular RNA genomes without protein-coding capacity and subvert enzymatic activities of their hosts to complete its own biological cycle. Different aspects of viroid biology and viroid-host interactions have been elucidated in the last years and some of them are the main topic of this review together with the analysis of the state-of-the-art about the growing field of endogenous lncRNAs in plants.} } @Article{IPB-982, author = {Drost, H.-G. and Bellstädt, J. and Ó'Maoiléidigh, D. S. and Silva, A. T. and Gabel, A. and Weinholdt, C. and Ryan, P. T. and Dekkers, B. J. W. and Bentsink, L. and Hilhorst, H. W. M. and Ligterink, W. and Wellmer, F. and Grosse, I. and Quint, M. and}, title = {{Post-embryonic Hourglass Patterns Mark Ontogenetic Transitions in Plant Development}}, year = {2016}, pages = {1158-1163}, journal = {Mol. Biol. Evol.}, doi = {10.1093/molbev/msw039}, volume = {33}, abstract = {The historic developmental hourglass concept depicts the convergence of animal embryos to a common form during the phylotypic period. Recently, it has been shown that a transcriptomic hourglass is associated with this morphological pattern, consistent with the idea of underlying selective constraints due to intense molecular interactions during body plan establishment. Although plants do not exhibit a morphological hourglass during embryogenesis, a transcriptomic hourglass has nevertheless been identified in the model plant Arabidopsis thaliana. Here, we investigated whether plant hourglass patterns are also found postembryonically. We found that the two main phase changes during the life cycle of Arabidopsis, from embryonic to vegetative and from vegetative to reproductive development, are associated with transcriptomic hourglass patterns. In contrast, flower development, a process dominated by organ formation, is not. This suggests that plant hourglass patterns are decoupled from organogenesis and body plan establishment. Instead, they may reflect general transitions through organizational checkpoints.} } @Article{IPB-979, author = {Dinesh, D. C. and Calderón Villalobos, L. I. A. and Abel, S. and}, title = {{Structural Biology of Nuclear Auxin Action}}, year = {2016}, pages = {302-316}, journal = {Trends Plant Sci.}, doi = {10.1016/j.tplants.2015.10.019}, volume = {21}, abstract = {Auxin coordinates plant development largely via hierarchical control of gene expression. During the past decades, the study of early auxin genes paired with the power of Arabidopsis genetics have unraveled key nuclear components and molecular interactions that perceive the hormone and activate primary response genes. Recent research in the realm of structural biology allowed unprecedented insight into: (i) the recognition of auxin-responsive DNA elements by auxin transcription factors; (ii) the inactivation of those auxin response factors by early auxin-inducible repressors; and (iii) the activation of target genes by auxin-triggered repressor degradation. The biophysical studies reviewed here provide an impetus for elucidating the molecular determinants of the intricate interactions between core components of the nuclear auxin response module.} } @Article{IPB-967, author = {Arnold, M. D. and Gruber, C. and Floková, K. and Miersch, O. and Strnad, M. and Novák, O. and Wasternack, C. and Hause, B. and}, title = {{The Recently Identified Isoleucine Conjugate of cis-12-Oxo-Phytodienoic Acid Is Partially Active in cis-12-Oxo-Phytodienoic Acid-Specific Gene Expression of Arabidopsis thaliana}}, year = {2016}, pages = {e0162829}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0162829}, volume = {11}, abstract = {Oxylipins of the jasmonate family are active as signals in plant responses to biotic and abiotic stresses as well as in development. Jasmonic acid (JA), its precursor cis-12-oxo-phytodienoic acid (OPDA) and the isoleucine conjugate of JA (JA-Ile) are the most prominent members. OPDA and JA-Ile have individual signalling properties in several processes and differ in their pattern of gene expression. JA-Ile, but not OPDA, is perceived by the SCFCOI1-JAZ co-receptor complex. There are, however, numerous processes and genes specifically induced by OPDA. The recently identified OPDA-Ile suggests that OPDA specific responses might be mediated upon formation of OPDA-Ile. Here, we tested OPDA-Ile-induced gene expression in wild type and JA-deficient, JA-insensitive and JA-Ile-deficient mutant background. Tests on putative conversion of OPDA-Ile during treatments revealed only negligible conversion. Expression of two OPDA-inducible genes, GRX480 and ZAT10, by OPDA-Ile could be detected in a JA-independent manner in Arabidopsis seedlings but less in flowering plants. The data suggest a bioactivity in planta of OPDA-Ile.} } @Article{IPB-990, author = {Floková, K. and Feussner, K. and Herrfurth, C. and Miersch, O. and Mik, V. and Tarkowská, D. and Strnad, M. and Feussner, I. and Wasternack, C. and Novák, O. and}, title = {{A previously undescribed jasmonate compound in flowering Arabidopsis thaliana – The identification of cis-(\+)-OPDA-Ile}}, year = {2016}, pages = {230-237}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2015.11.012}, volume = {122}, abstract = {Jasmonates (JAs) are plant hormones that integrate external stress stimuli with physiological responses. (\+)-7-iso-JA-L-Ile is the natural JA ligand of COI1, a component of a known JA receptor. The upstream JA biosynthetic precursor cis-(\+)-12-oxo-phytodienoic acid (cis-(\+)-OPDA) has been reported to act independently of COI1 as an essential signal in several stress-induced and developmental processes. Wound-induced increases in the endogenous levels of JA/JA-Ile are accompanied by two to tenfold increases in the concentration of OPDA, but its means of perception and metabolism are unknown. To screen for putative OPDA metabolites, vegetative tissues of flowering Arabidopsis thaliana were extracted with 25% aqueous methanol (v/v), purified by single-step reversed-phase polymer-based solid-phase extraction, and analyzed by high throughput mass spectrometry. This enabled the detection and quantitation of a low abundant OPDA analog of the biologically active (\+)-7-iso-JA-L-Ile in plant tissue samples. Levels of the newly identified compound and the related phytohormones JA, JA-Ile and cis-(\+)-OPDA were monitored in wounded leaves of flowering Arabidopsis lines (Col-0 and Ws) and compared to the levels observed in Arabidopsis mutants deficient in the biosynthesis of JA (dde2-2, opr3) and JA-Ile (jar1). The observed cis-(\+)-OPDA-Ile levels varied widely, raising questions concerning its role in Arabidopsis stress responses.} } @Article{IPB-1069, author = {Ziegler, J. and Schmidt, S. and Chutia, R. and Müller, J. and Böttcher, C. and Strehmel, N. and Scheel, D. and Abel, S. and}, title = {{Non-targeted profiling of semi-polar metabolites in Arabidopsis root exudates uncovers a role for coumarin secretion and lignification during the local response to phosphate limitation}}, year = {2016}, pages = {1421-1432}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/erv539}, volume = {67}, abstract = {Plants have evolved two major strategies to cope with phosphate (Pi) limitation. The systemic response, mainly comprising increased Pi uptake and metabolic adjustments for more efficient Pi use, and the local response, enabling plants to explore Pi-rich soil patches by reorganization of the root system architecture. Unlike previous reports, this study focused on root exudation controlled by the local response to Pi deficiency. To approach this, a hydroponic system separating the local and systemic responses was developed. Arabidopsis thaliana genotypes exhibiting distinct sensitivities to Pi deficiency could be clearly distinguished by their root exudate composition as determined by non-targeted reversed-phase ultraperformance liquid chromatography electrospray ionization quadrupole-time-of-flight mass spectrometry metabolite profiling. Compared with wild-type plants or insensitive low phosphate root 1 and 2 (lpr1 lpr2) double mutant plants, the hypersensitive phosphate deficiency response 2 (pdr2) mutant exhibited a reduced number of differential features in root exudates after Pi starvation, suggesting the involvement of PDR2-encoded P5-type ATPase in root exudation. Identification and analysis of coumarins revealed common and antagonistic regulatory pathways between Pi and Fe deficiency-induced coumarin secretion. The accumulation of oligolignols in root exudates after Pi deficiency was inversely correlated with Pi starvation-induced lignification at the root tips. The strongest oligolignol accumulation in root exudates was observed for the insensitive lpr1 lpr2 double mutant, which was accompanied by the absence of Pi deficiency-induced lignin deposition, suggesting a role of LPR ferroxidases in lignin polymerization during Pi starvation.} } @Article{IPB-1062, author = {Wasternack, C. and Hause, B. and}, title = {{OPDA-Ile – a new JA-Ile-independent signal?}}, year = {2016}, pages = {e1253646}, journal = {Plant Signal Behav.}, doi = {10.1080/15592324.2016.1253646}, volume = {11}, abstract = {Expression takes place for most of the jasmonic acid (JA)-induced genes in a COI1-dependent manner via perception of its conjugate JA-Ile in the SCFCOI1-JAZ co-receptor complex. There are, however, numerous genes and processes, which are preferentially induced COI1-independently by the precursor of JA, 12-oxo-phytodienoic acid (OPDA). After recent identification of the Ile-conjugate of OPDA, OPDA-Ile, biological activity of this compound could be unequivocally proven in terms of gene expression. Any interference of OPDA, JA, or JA-Ile in OPDA-Ile-induced gene expression could be excluded by using different genetic background. The data suggest individual signaling properties of OPDA-Ile. Future studies for analysis of an SCFCOI1-JAZ co-receptor-independent route of signaling are proposed.} } @Article{IPB-1061, author = {Wasternack, C. and Strnad, M. and}, title = {{Jasmonate signaling in plant stress responses and development – active and inactive compounds}}, year = {2016}, pages = {604-613}, journal = {New Biotechnol.}, doi = {10.1016/j.nbt.2015.11.001}, volume = {33}, abstract = {Jasmonates (JAs) are lipid-derived signals mediating plant responses to biotic and abiotic stresses and in plant development. Following the elucidation of each step in their biosynthesis and the important components of perception and signaling, several activators, repressors and co-repressors have been identified which contribute to fine-tuning the regulation of JA-induced gene expression. Many of the metabolic reactions in which JA participates, such as conjugation with amino acids, glucosylation, hydroxylation, carboxylation, sulfation and methylation, lead to numerous compounds with different biological activities. These metabolites may be highly active, partially active in specific processes or inactive. Hydroxylation, carboxylation and sulfation inactivate JA signaling. The precursor of JA biosynthesis, 12-oxo-phytodienoic acid (OPDA), has been identified as a JA-independent signaling compound. An increasing number of OPDA-specific processes is being identified. To conclude, the numerous JA compounds and their different modes of action allow plants to respond specifically and flexibly to alterations in the environment.} } @Article{IPB-1051, author = {Strehmel, N. and Mönchgesang, S. and Herklotz, S. and Krüger, S. and Ziegler, J. and Scheel, D. and}, title = {{Piriformospora indica Stimulates Root Metabolism of Arabidopsis thaliana}}, year = {2016}, pages = {1091}, journal = {Int. J. Mol. Sci.}, doi = {10.3390/ijms17071091}, volume = {17}, abstract = {Piriformospora indica is a root-colonizing fungus, which interacts with a variety of plants including Arabidopsis thaliana. This interaction has been considered as mutualistic leading to growth promotion of the host. So far, only indolic glucosinolates and phytohormones have been identified as key players. In a comprehensive non-targeted metabolite profiling study, we analyzed Arabidopsis thaliana’s roots, root exudates, and leaves of inoculated and non-inoculated plants by ultra performance liquid chromatography/electrospray ionization quadrupole-time-of-flight mass spectrometry (UPLC/(ESI)-QTOFMS) and gas chromatography/electron ionization quadrupole mass spectrometry (GC/EI-QMS), and identified further biomarkers. Among them, the concentration of nucleosides, dipeptides, oligolignols, and glucosinolate degradation products was affected in the exudates. In the root profiles, nearly all metabolite levels increased upon co-cultivation, like carbohydrates, organic acids, amino acids, glucosinolates, oligolignols, and flavonoids. In the leaf profiles, we detected by far less significant changes. We only observed an increased concentration of organic acids, carbohydrates, ascorbate, glucosinolates and hydroxycinnamic acids, and a decreased concentration of nitrogen-rich amino acids in inoculated plants. These findings contribute to the understanding of symbiotic interactions between plant roots and fungi of the order of Sebacinales and are a valid source for follow-up mechanistic studies, because these symbioses are particular and clearly different from interactions of roots with mycorrhizal fungi or dark septate endophytes} } @Article{IPB-1038, author = {Quint, M. and Delker, C. and Franklin, K. A. and Wigge, P. A. and Halliday, K. J. and van Zanten, M. and}, title = {{Molecular and genetic control of plant thermomorphogenesis}}, year = {2016}, pages = {15190}, journal = {Nat. Plants}, doi = {10.1038/nplants.2015.190}, volume = {2}, abstract = {Temperature is a major factor governing the distribution and seasonal behaviour of plants. Being sessile, plants are highly responsive to small differences in temperature and adjust their growth and development accordingly. The suite of morphological and architectural changes induced by high ambient temperatures, below the heat-stress range, is collectively called thermomorphogenesis. Understanding the molecular genetic circuitries underlying thermomorphogenesis is particularly relevant in the context of climate change, as this knowledge will be key to rational breeding for thermo-tolerant crop varieties. Until recently, the fundamental mechanisms of temperature perception and signalling remained unknown. Our understanding of temperature signalling is now progressing, mainly by exploiting the model plant Arabidopsis thaliana. The transcription factor PHYTOCHROME INTERACTING FACTOR 4 (PIF4) has emerged as a critical player in regulating phytohormone levels and their activity. To control thermomorphogenesis, multiple regulatory circuits are in place to modulate PIF4 levels, activity and downstream mechanisms. Thermomorphogenesis is integrally governed by various light signalling pathways, the circadian clock, epigenetic mechanisms and chromatin-level regulation. In this Review, we summarize recent progress in the field and discuss how the emerging knowledge in Arabidopsis may be transferred to relevant crop systems.} } @Article{IPB-1027, author = {Otto, M. and Naumann, C. and Brandt, W. and Wasternack, C. and Hause, B. and}, title = {{Activity Regulation by Heteromerization of Arabidopsis Allene Oxide Cyclase Family Members}}, year = {2016}, pages = {3}, journal = {Plants}, doi = {10.3390/plants5010003}, volume = {5}, abstract = {Jasmonates (JAs) are lipid-derived signals in plant stress responses and development. A crucial step in JA biosynthesis is catalyzed by allene oxide cyclase (AOC). Four genes encoding functional AOCs (AOC1, AOC2, AOC3 and AOC4) have been characterized for Arabidopsis thaliana in terms of organ- and tissue-specific expression, mutant phenotypes, promoter activities and initial in vivo protein interaction studies suggesting functional redundancy and diversification, including first hints at enzyme activity control by protein-protein interaction. Here, these analyses were extended by detailed analysis of recombinant proteins produced in Escherichia coli. Treatment of purified AOC2 with SDS at different temperatures, chemical cross-linking experiments and protein structure analysis by molecular modelling approaches were performed. Several salt bridges between monomers and a hydrophobic core within the AOC2 trimer were identified and functionally proven by site-directed mutagenesis. The data obtained showed that AOC2 acts as a trimer. Finally, AOC activity was determined in heteromers formed by pairwise combinations of the four AOC isoforms. The highest activities were found for heteromers containing AOC4 \+ AOC1 and AOC4 \+ AOC2, respectively. All data are in line with an enzyme activity control of all four AOCs by heteromerization, thereby supporting a putative fine-tuning in JA formation by various regulatory principles.} } @Article{IPB-1090, author = {Drost, H.-G. and Gabel, A. and Grosse, I. and Quint, M. and}, title = {{Evidence for Active Maintenance of Phylotranscriptomic Hourglass Patterns in Animal and Plant Embryogenesis}}, year = {2015}, pages = {1221-1231}, journal = {Mol. Biol. Evol.}, doi = {10.1093/molbev/msv012}, volume = {32}, abstract = {The developmental hourglass model has been used to describe the morphological transitions of related species throughout embryogenesis. Recently, quantifiable approaches combining transcriptomic and evolutionary information provided novel evidence for the presence of a phylotranscriptomic hourglass pattern across kingdoms. As its biological function is unknown it remains speculative whether this pattern is functional or merely represents a nonfunctional evolutionary relic. The latter would seriously hamper future experimental approaches designed to test hypotheses regarding its function. Here, we address this question by generating transcriptome divergence index (TDI) profiles across embryogenesis of Danio rerio, Drosophila melanogaster, and Arabidopsis thaliana. To enable meaningful evaluation of the resulting patterns, we develop a statistical test that specifically assesses potential hourglass patterns. Based on this objective measure we find that two of these profiles follow a statistically significant hourglass pattern with the most conserved transcriptomes in the phylotypic periods. As the TDI considers only recent evolutionary signals, this indicates that the phylotranscriptomic hourglass pattern is not a rudiment but possibly actively maintained, implicating the existence of some linked biological function associated with embryogenesis in extant species.} } @Article{IPB-1087, author = {Dinesh, D. C. and Kovermann, M. and Gopalswamy, M. and Hellmuth, A. and Calderón Villalobos, L. I. A. and Lilie, H. and Balbach, J. and Abel, S. and}, title = {{Solution structure of the PsIAA4 oligomerization domain reveals interaction modes for transcription factors in early auxin response}}, year = {2015}, pages = {6230-6235}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.1424077112}, volume = {112}, abstract = {The plant hormone auxin activates primary response genes by facilitating proteolytic removal of AUXIN/INDOLE-3-ACETIC ACID (AUX/IAA)-inducible repressors, which directly bind to transcriptional AUXIN RESPONSE FACTORS (ARF). Most AUX/IAA and ARF proteins share highly conserved C-termini mediating homotypic and heterotypic interactions within and between both protein families. The high-resolution NMR structure of C-terminal domains III and IV of the AUX/IAA protein PsIAA4 from pea (Pisum sativum) revealed a globular ubiquitin-like β-grasp fold with homologies to the Phox and Bem1p (PB1) domain. The PB1 domain of wild-type PsIAA4 features two distinct surface patches of oppositely charged amino acid residues, mediating front-to-back multimerization via electrostatic interactions. Mutations of conserved basic or acidic residues on either face suppressed PsIAA4 PB1 homo-oligomerization in vitro and confirmed directional interaction of full-length PsIAA4 in vivo (yeast two-hybrid system). Mixing of oppositely mutated PsIAA4 PB1 monomers enabled NMR mapping of the negatively charged interface of the reconstituted PsIAA4 PB1 homodimer variant, whose stoichiometry (1:1) and equilibrium binding constant (KD ∼6.4 μM) were determined by isothermal titration calorimetry. In silico protein–protein docking studies based on NMR and yeast interaction data derived a model of the PsIAA4 PB1 homodimer, which is comparable with other PB1 domain dimers, but indicated considerable differences between the homodimeric interfaces of AUX/IAA and ARF PB1 domains. Our study provides an impetus for elucidating the molecular determinants that confer specificity to complex protein–protein interaction circuits between members of the two central families of transcription factors important to the regulation of auxin-responsive gene expression.} } @Article{IPB-1084, author = {Buhtz, A. and Witzel, K. and Strehmel, N. and Ziegler, J. and Abel, S. and Grosch, R. and}, title = {{Perturbations in the Primary Metabolism of Tomato and Arabidopsis thaliana Plants Infected with the Soil-Borne Fungus Verticillium dahliae}}, year = {2015}, pages = {e0138242}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0138242}, volume = {10}, abstract = {The hemibiotrophic soil-borne fungus Verticillium dahliae is a major pathogen of a number of economically important crop species. Here, the metabolic response of both tomato and Arabidopsis thaliana to V. dahliae infection was analysed by first using non-targeted GC-MS profiling. The leaf content of both major cell wall components glucuronic acid and xylose was reduced in the presence of the pathogen in tomato but enhanced in A. thaliana. The leaf content of the two tricarboxylic acid cycle intermediates fumaric acid and succinic acid was increased in the leaf of both species, reflecting a likely higher demand for reducing equivalents required for defence responses. A prominent group of affected compounds was amino acids and based on the targeted analysis in the root, it was shown that the level of 12 and four free amino acids was enhanced by the infection in, respectively, tomato and A. thaliana, with leucine and histidine being represented in both host species. The leaf content of six free amino acids was reduced in the leaf tissue of diseased A. thaliana plants, while that of two free amino acids was raised in the tomato plants. This study emphasizes the role of primary plant metabolites in adaptive responses when the fungus has colonized the plant.} } @Article{IPB-1077, author = {Bochnia, M. and Ziegler, J. and Sander, J. and Uhlig, A. and Schaefer, S. and Vollstedt, S. and Glatter, M. and Abel, S. and Recknagel, S. and Schusser, G. F. and Wensch-Dorendorf, M. and Zeyner, A. and}, title = {{Hypoglycin A Content in Blood and Urine Discriminates Horses with Atypical Myopathy from Clinically Normal Horses Grazing on the Same Pasture}}, year = {2015}, pages = {e0136785}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0136785}, volume = {10}, abstract = {Hypoglycin A (HGA) in seeds of Acer spp. is suspected to cause seasonal pasture myopathy in North America and equine atypical myopathy (AM) in Europe, fatal diseases in horses on pasture. In previous studies, this suspicion was substantiated by the correlation of seed HGA content with the concentrations of toxic metabolites in urine and serum (MCPA-conjugates) of affected horses. However, seed sampling was conducted after rather than during an outbreak of the disease. The aim of this study was to further confirm the causality between HGA occurrence and disease outbreak by seed sampling during an outbreak and the determination of i) HGA in seeds and of ii) HGA and MCPA-conjugates in urine and serum of diseased horses. Furthermore, cograzing healthy horses, which were present on AM affected pastures, were also investigated. AM-pastures in Germany were visited to identify seeds of Acer pseudoplatanus and serum (n \= 8) as well as urine (n \= 6) from a total of 16 diseased horses were analyzed for amino acid composition by LC-ESI-MS/MS, with a special focus on the content of HGA. Additionally, the content of its toxic metabolite was measured in its conjugated form in body fluids (UPLC-MS/MS). The seeds contained 1.7–319.8 μg HGA/g seed. The content of HGA in serum of affected horses ranged from 387.8–8493.8 μg/L (controls \< 10 μg/L), and in urine from 143.8–926.4 μg/L (controls \< 10 μg/L), respectively. Healthy cograzing horses on AM-pastures showed higher serum (108.8 ± 83.76 μg/L) and urine concentrations (26.9 ± 7.39 μg/L) compared to control horses, but lower concentrations compared to diseased horses. The range of MCPA-carnitine and creatinine concentrations found in diseased horses in serum and urine were 0.17–0.65 mmol/L (controls \< 0.01), and 0.34–2.05 μmol/mmoL (controls \< 0.001), respectively. MCPA-glycine levels in urine of cograzing horses were higher compared to controls. Thus, the causal link between HGA intoxication and disease outbreak could be further substantiated, and the early detection of HGA in cograzing horses, which are clinically normal, might be a promising step in prophylaxis.} } @Article{IPB-1150, author = {Ryan, P. T. and Ó’Maoiléidigh, D. S. and Drost, H.-G. and Kwaśniewska, K. and Gabel, A. and Grosse, I. and Graciet, E. and Quint, M. and Wellmer, F. and}, title = {{Patterns of gene expression during Arabidopsis flower development from the time of initiation to maturation}}, year = {2015}, pages = {488}, journal = {BMC Genomics}, doi = {10.1186/s12864-015-1699-6}, volume = {16}, abstract = {BackgroundThe formation of flowers is one of the main model systems to elucidate the molecular mechanisms that control developmental processes in plants. Although several studies have explored gene expression during flower development in the model plant Arabidopsis thaliana on a genome-wide scale, a continuous series of expression data from the earliest floral stages until maturation has been lacking. Here, we used a floral induction system to close this information gap and to generate a reference dataset for stage-specific gene expression during flower formation.ResultsUsing a floral induction system, we collected floral buds at 14 different stages from the time of initiation until maturation. Using whole-genome microarray analysis, we identified 7,405 genes that exhibit rapid expression changes during flower development. These genes comprise many known floral regulators and we found that the expression profiles for these regulators match their known expression patterns, thus validating the dataset. We analyzed groups of co-expressed genes for over-represented cellular and developmental functions through Gene Ontology analysis and found that they could be assigned specific patterns of activities, which are in agreement with the progression of flower development. Furthermore, by mapping binding sites of floral organ identity factors onto our dataset, we were able to identify gene groups that are likely predominantly under control of these transcriptional regulators. We further found that the distribution of paralogs among groups of co-expressed genes varies considerably, with genes expressed predominantly at early and intermediate stages of flower development showing the highest proportion of such genes.ConclusionsOur results highlight and describe the dynamic expression changes undergone by a large number of genes during flower development. They further provide a comprehensive reference dataset for temporal gene expression during flower formation and we demonstrate that it can be used to integrate data from other genomics approaches such as genome-wide localization studies of transcription factor binding sites.} } @Article{IPB-1147, author = {Rekik, I. and Drira, N. and Grubb, C. D. and Elleuch, A. and}, title = {{Molecular characterization and evolution studies of a SERK like gene transcriptionally induced during somatic embryogenesis in Phoenix Dactylifera L v Deglet Nour}}, year = {2015}, pages = {323-337}, journal = {Genetika}, doi = {10.2298/GENSR1501323R}, volume = {47}, abstract = {A somatic embryogenesis receptor kinase like (SERKL) cDNA, designated PhSERKL, was isolated from date palm (Phoenix Dactylifera L) using RACE PCR. PhSERKL protein shared all the characteristic domains of the SERK family, including five leucine-rich repeats, one proline-rich region motif, a transmembrane domain, and kinase domains. Phylogenetic analyses using PHYLIP and Notung 2.7 programs suggest that the SERK proteins of some plant species resulted from relatively ancient duplication events. We predict an ancestor protein of monocots and dicots SERK using FASTML program. Somatic embryogenic cultures of date palm were established following transfer of callus cultures to medium containing 2, 4-dichlorophenoxyacetic acid. The role of PhSERKL gene during establishment of somatic embryogenesis in culture was investigated using quantitative real-time PCR. PhSERKL gene was highly expressed during embryogenic competence acquisition and globular embryo formation in culture. Overall, levels of expression of PhSERKL gene were lower in nonembryogenic tissues and organs than in embryogenic callus.} } @Article{IPB-1146, author = {Rekik, I. and Chaâbene, Z. and Grubb, C. D. and Drira, N. and Cheour, F. and Elleuch, A. and}, title = {{In silico characterization and Molecular modeling of double-strand break repair protein MRE11 from Phoenix dactylifera v deglet nour}}, year = {2015}, pages = {23}, journal = {Theor. Biol. Med. Model.}, doi = {10.1186/s12976-015-0013-2}, volume = {12}, abstract = {BackgroundDNA double-strand breaks (DSBs) are highly cytotoxic and mutagenic. MRE11 plays an essential role in repairing DNA by cleaving broken ends through its 3′ to 5′ exonuclease and single-stranded DNA endonuclease activities.MethodsThe present study aimed to in silico characterization and molecular modeling of MRE11 from Phoenix dactylifera L cv deglet nour (DnMRE11) by various bioinformatic approaches. To identify DnMRE11 cDNA, assembled contigs from our cDNA libraries were analysed using the Blast2GO2.8 program.ResultsThe DnMRE11 protein length was 726 amino acids. The results of HUMMER show that DnMRE11 is formed by three domains: the N-terminal core domain containing the nuclease and capping domains, the C-terminal half containing the DNA binding and coiled coil region. The structure of DnMRE11 is predicted using the Swiss-Model server, which contains the nuclease and capping domains. The obtained model was verified with the structure validation programs such as ProSA and QMEAN servers for reliability. Ligand binding studies using COACH indicated the interaction of DnMRE11 protein with two Mn2\+ ions and dAMP. The ConSurf server predicted that residues of the active site and Nbs binding site have high conservation scores between plant species.ConclusionsA model structure of DnMRE11 was constructed and validated with various bioinformatics programs which suggested the predicted model to be satisfactory. Further validation studies were conducted by COACH analysis for active site ligand prediction, and revealed the presence of six ligands binding sites and two ligands (2 Mn2\+ and dAMP).} } @Article{IPB-1145, author = {Raschke, A. and Ibañez, C. and Ullrich, K. K. and Anwer, M. U. and Becker, S. and Glöckner, A. and Trenner, J. and Denk, K. and Saal, B. and Sun, X. and Ni, M. and Davis, S. J. and Delker, C. and Quint, M. and}, title = {{Natural variants of ELF3 affect thermomorphogenesis by transcriptionally modulating PIF4-dependent auxin response genes}}, year = {2015}, pages = {197}, journal = {BMC Plant Biol.}, doi = {10.1186/s12870-015-0566-6}, volume = {15}, abstract = {BackgroundPerception and transduction of temperature changes result in altered growth enabling plants to adapt to increased ambient temperature. While PHYTOCHROME-INTERACTING FACTOR4 (PIF4) has been identified as a major ambient temperature signaling hub, its upstream regulation seems complex and is poorly understood. Here, we exploited natural variation for thermo-responsive growth in Arabidopsis thaliana using quantitative trait locus (QTL) analysis.ResultsWe identified GIRAFFE2.1, a major QTL explaining ~18 % of the phenotypic variation for temperature-induced hypocotyl elongation in the Bay-0 x Sha recombinant inbred line population. Transgenic complementation demonstrated that allelic variation in the circadian clock regulator EARLY FLOWERING3 (ELF3) is underlying this QTL. The source of variation could be allocated to a single nucleotide polymorphism in the ELF3 coding region, resulting in differential expression of PIF4 and its target genes, likely causing the observed natural variation in thermo-responsive growth.ConclusionsIn combination with other recent studies, this work establishes the role of ELF3 in the ambient temperature signaling network. Natural variation of ELF3-mediated gating of PIF4 expression during nightly growing periods seems to be affected by a coding sequence quantitative trait nucleotide that confers a selective advantage in certain environments. In addition, natural ELF3 alleles seem to differentially integrate temperature and photoperiod information to induce architectural changes. Thus, ELF3 emerges as an essential coordinator of growth and development in response to diverse environmental cues and implicates ELF3 as an important target of adaptation.} } @Article{IPB-1136, author = {Müller, J. and Toev, T. and Heisters, M. and Teller, J. and Moore, K. and Hause, G. and Dinesh, D. and Bürstenbinder, K. and Abel, S. and}, title = {{Iron-Dependent Callose Deposition Adjusts Root Meristem Maintenance to Phosphate Availability}}, year = {2015}, pages = {216-230}, journal = {Dev. Cell}, doi = {10.1016/j.devcel.2015.02.007}, volume = {33}, abstract = {Plant root development is informed by numerous edaphic cues. Phosphate (Pi) availability impacts the root system architecture by adjusting meristem activity. However, the sensory mechanisms monitoring external Pi status are elusive. Two functionally interacting Arabidopsis genes, LPR1 (ferroxidase) and PDR2 (P5-type ATPase), are key players in root Pi sensing, which is modified by iron (Fe) availability. We show that the LPR1-PDR2 module facilitates, upon Pi limitation, cell-specific apoplastic Fe and callose deposition in the meristem and elongation zone of primary roots. Expression of cell-wall-targeted LPR1 determines the sites of Fe accumulation as well as callose production, which interferes with symplastic communication in the stem cell niche, as demonstrated by impaired SHORT-ROOT movement. Antagonistic interactions of Pi and Fe availability control primary root growth via meristem-specific callose formation, likely triggered by LPR1-dependent redox signaling. Our results link callose-regulated cell-to-cell signaling in root meristems to the perception of an abiotic cue.} } @Article{IPB-1135, author = {Moss, B. L. and Mao, H. and Guseman, J. M. and Hinds, T. R. and Hellmuth, A. and Kovenock, M. and Noorassa, A. and Lanctot, A. and Calderón Villalobos, L. I. A. and Zheng, N. and Nemhauser, J. L. and}, title = {{Rate Motifs Tune Auxin/Indole-3-Acetic Acid Degradation Dynamics}}, year = {2015}, pages = {803-813}, journal = {Plant Physiol.}, doi = {10.1104/pp.15.00587}, volume = {169}, abstract = {Ubiquitin-mediated protein degradation is a common feature in diverse plant cell signaling pathways; however, the factors that control the dynamics of regulated protein turnover are largely unknown. One of the best-characterized families of E3 ubiquitin ligases facilitates ubiquitination of auxin (aux)/indole-3-acetic acid (IAA) repressor proteins in the presence of auxin. Rates of auxin-induced degradation vary widely within the Aux/IAA family, and sequences outside of the characterized degron (the minimum region required for auxin-induced degradation) can accelerate or decelerate degradation. We have used synthetic auxin degradation assays in yeast (Saccharomyces cerevisiae) and in plants to characterize motifs flanking the degron that contribute to tuning the dynamics of Aux/IAA degradation. The presence of these rate motifs is conserved in phylogenetically distant members of the Arabidopsis (Arabidopsis thaliana) Aux/IAA family, as well as in their putative Brassica rapa orthologs. We found that rate motifs can act by enhancing interaction between repressors and the E3, but that this is not the only mechanism of action. Phenotypes of transgenic plants expressing a deletion in a rate motif in IAA28 resembled plants expressing degron mutations, underscoring the functional relevance of Aux/IAA degradation dynamics in regulating auxin responses.} } @Article{IPB-1126, author = {Liu, S. and Kracher, B. and Ziegler, J. and Birkenbihl, R. P. and Somssich, I. E. and}, title = {{Negative regulation of ABA signaling by WRKY33 is critical for Arabidopsis immunity towards Botrytis cinerea 2100}}, year = {2015}, pages = {e07295}, journal = {eLife}, doi = {10.7554/eLife.07295}, volume = {4}, abstract = {The Arabidopsis mutant wrky33 is highly susceptible to Botrytis cinerea. We identified \>1680 Botrytis-induced WRKY33 binding sites associated with 1576 Arabidopsis genes. Transcriptional profiling defined 318 functional direct target genes at 14 hr post inoculation. Comparative analyses revealed that WRKY33 possesses dual functionality acting either as a repressor or as an activator in a promoter-context dependent manner. We confirmed known WRKY33 targets involved in hormone signaling and phytoalexin biosynthesis, but also uncovered a novel negative role of abscisic acid (ABA) in resistance towards B. cinerea 2100. The ABA biosynthesis genes NCED3 and NCED5 were identified as direct targets required for WRKY33-mediated resistance. Loss-of-WRKY33 function resulted in elevated ABA levels and genetic studies confirmed that WRKY33 acts upstream of NCED3/NCED5 to negatively regulate ABA biosynthesis. This study provides the first detailed view of the genome-wide contribution of a specific plant transcription factor in modulating the transcriptional network associated with plant immunity.} } @Article{IPB-1108, author = {Hamdi, I. and Elleuch, A. and Bessaies, N. and Grubb, C. D. and Fakhfakh, H. and}, title = {{First report of Citrus viroid V in North Africa}}, year = {2015}, pages = {87-91}, journal = {J. Gen. Plant Pathol.}, doi = {10.1007/s10327-014-0556-9}, volume = {81}, abstract = {We tested citrus samples from Tunisia using reverse transcription-polymerase chain reaction (RT-PCR), and for the first time, Citrus viroid V (CVd-V) was reported in North Africa. Fourteen of 38 tested citrus trees were infected by CVd-V including the majority of varieties grown in Tunisia. Some RT-PCR results were also supported by biological indexing. After sequencing the RT-PCR products, three new CVd-V variants were identified, showing 80–91 % nucleotide sequence identity with those reported previously. Based on phylogenetic analysis using all CVd-V sequences in GenBank, two main CVd-V groups were identified. Furthermore, construction of a genetic network of the detected haplotypes using the same sequences shows a clear geographical structuring of Tunisian CVd-V variants.} } @Article{IPB-1107, author = {Guseman, J. M. and Hellmuth, A. and Lanctot, A. and Feldman, T. P. and Moss, B. L. and Klavins, E. and Calderón Villalobos, L. I. A. and Nemhauser, J. L. and}, title = {{Auxin-induced degradation dynamics set the pace for lateral root development}}, year = {2015}, pages = {905-909}, journal = {Development}, doi = {10.1242/dev.117234}, volume = {142}, abstract = {Auxin elicits diverse cell behaviors through a simple nuclear signaling pathway initiated by degradation of Aux/IAA co-repressors. Our previous work revealed that members of the large Arabidopsis Aux/IAA family exhibit a range of degradation rates in synthetic contexts. However, it remained an unresolved issue whether differences in Aux/IAA turnover rates played a significant role in plant responses to auxin. Here, we use the well-established model of lateral root development to directly test the hypothesis that the rate of auxin-induced Aux/IAA turnover sets the pace for auxin-regulated developmental events. We did this by generating transgenic plants expressing degradation rate variants of IAA14, a crucial determinant of lateral root initiation. Progression through the well-established stages of lateral root development was strongly correlated with the engineered rates of IAA14 turnover, leading to the conclusion that Aux/IAAs are auxin-initiated timers that synchronize developmental transitions.} } @Article{IPB-1103, author = {Gasperini, D. and Chételat, A. and Acosta, I. F. and Goossens, J. and Pauwels, L. and Goossens, A. and Dreos, R. and Alfonso, E. and Farmer, E. E. and}, title = {{Multilayered Organization of Jasmonate Signalling in the Regulation of Root Growth}}, year = {2015}, pages = {e1005300}, journal = {PLOS Genet.}, doi = {10.1371/journal.pgen.1005300}, volume = {11}, abstract = {Physical damage can strongly affect plant growth, reducing the biomass of developing organs situated at a distance from wounds. These effects, previously studied in leaves, require the activation of jasmonate (JA) signalling. Using a novel assay involving repetitive cotyledon wounding in Arabidopsis seedlings, we uncovered a function of JA in suppressing cell division and elongation in roots. Regulatory JA signalling components were then manipulated to delineate their relative impacts on root growth. The new transcription factor mutant myc2-322B was isolated. In vitro transcription assays and whole-plant approaches revealed that myc2-322B is a dosage-dependent gain-of-function mutant that can amplify JA growth responses. Moreover, myc2-322B displayed extreme hypersensitivity to JA that totally suppressed root elongation. The mutation weakly reduced root growth in undamaged plants but, when the upstream negative regulator NINJA was genetically removed, myc2-322B powerfully repressed root growth through its effects on cell division and cell elongation. Furthermore, in a JA-deficient mutant background, ninja1 myc2-322B still repressed root elongation, indicating that it is possible to generate JA-responses in the absence of JA. We show that NINJA forms a broadly expressed regulatory layer that is required to inhibit JA signalling in the apex of roots grown under basal conditions. By contrast, MYC2, MYC3 and MYC4 displayed cell layer-specific localisations and MYC3 and MYC4 were expressed in mutually exclusive regions. In nature, growing roots are likely subjected to constant mechanical stress during soil penetration that could lead to JA production and subsequent detrimental effects on growth. Our data reveal how distinct negative regulatory layers, including both NINJA-dependent and -independent mechanisms, restrain JA responses to allow normal root growth. Mechanistic insights from this work underline the importance of mapping JA signalling components to specific cell types in order to understand and potentially engineer the growth reduction that follows physical damage.} } @Article{IPB-1102, author = {Gasperini, D. and Chauvin, A. and Acosta, I. F. and Kurenda, A. and Stolz, S. and Chételat, A. and Wolfender, J.-L. and Farmer, E. E. and}, title = {{Axial and Radial Oxylipin Transport}}, year = {2015}, pages = {2244-2254}, journal = {Plant Physiol.}, doi = {10.1104/pp.15.01104}, volume = {169}, abstract = {Jasmonates are oxygenated lipids (oxylipins) that control defense gene expression in response to cell damage in plants. How mobile are these potent mediators within tissues? Exploiting a series of 13-lipoxygenase (13-lox) mutants in Arabidopsis (Arabidopsis thaliana) that displays impaired jasmonic acid (JA) synthesis in specific cell types and using JA-inducible reporters, we mapped the extent of the transport of endogenous jasmonates across the plant vegetative growth phase. In seedlings, we found that jasmonate (or JA precursors) could translocate axially from wounded shoots to unwounded roots in a LOX2-dependent manner. Grafting experiments with the wild type and JA-deficient mutants confirmed shoot-to-root oxylipin transport. Next, we used rosettes to investigate radial cell-to-cell transport of jasmonates. After finding that the LOX6 protein localized to xylem contact cells was not wound inducible, we used the lox234 triple mutant to genetically isolate LOX6 as the only JA precursor-producing LOX in the plant. When a leaf of this mutant was wounded, the JA reporter gene was expressed in distal leaves. Leaf sectioning showed that JA reporter expression extended from contact cells throughout the vascular bundle and into extravascular cells, revealing a radial movement of jasmonates. Our results add a crucial element to a growing picture of how the distal wound response is regulated in rosettes, showing that both axial (shoot-to-root) and radial (cell-to-cell) transport of oxylipins plays a major role in the wound response. The strategies developed herein provide unique tools with which to identify intercellular jasmonate transport routes.} } @Article{IPB-1176, author = {Zayneb, C. and Bassem, K. and Zeineb, K. and Grubb, C. D. and Noureddine, D. and Hafedh, M. and Amine, E. and}, title = {{Physiological responses of fenugreek seedlings and plants treated with cadmium}}, year = {2015}, pages = {10679-10689}, journal = {Environ. Sci. Pollut. Res.}, doi = {10.1007/s11356-015-4270-8}, volume = {22}, abstract = {The bioaccumulation efficiency of cadmium (Cd) by fenugreek (Trigonella foenum-graecum) was examined using different concentrations of CdCl2. The germination rate was similar to control except at 10 mM Cd. However, early seedling growth was quite sensitive to the metal from the lowest Cd level. Accordingly, amylase activity was reduced substantially on treatment of seeds with 0.5, 1, and 10 mM Cd. Cadmium also affected various other plant growth parameters. Its accumulation was markedly lower in shoots as compared to roots, reducing root biomass by almost 50 %. Plants treated with 1 and 5 mM Cd presented chlorosis due to a significant reduction in chlorophyll b especially. Furthermore, at Cd concentrations greater than 0.1 mM, plants showed several signs of oxidative stress; an enhancement in root hydrogen peroxide (H2O2) level and in shoot malondialdehyde (MDA) content was observed. Conversely, antioxidant enzyme activities (superoxide dismutase (SOD), ascorbate peroxidase (APX), and catalase (CAT)) increased in various plant parts. Likewise, total phenolic and flavonoid contents reached their highest values in the 0.5 mM Cd treatment, consistent with their roles in quenching low concentrations of reactive oxygen species (ROS). Consequently, maintaining oxidant and antioxidant balance may permit fenugreek to hyperaccumulate Cd and allow it to be employed in extremely Cd polluted soils for detoxification purposes.} } @Article{IPB-1175, author = {Zayneb, C. and Lamia, K. and Olfa, E. and Naïma, J. and Grubb, C. D. and Bassem, K. and Hafedh, M. and Amine, E. and}, title = {{Morphological, Physiological and Biochemical Impact of Ink Industry Effluent on Germination of Maize (Zea mays), Barley (Hordeum vulgare) and Sorghum (Sorghum bicolor)}}, year = {2015}, pages = {687-693}, journal = {Bull. Environ. Contam. Toxicol.}, doi = {10.1007/s00128-015-1600-y}, volume = {95}, abstract = {The present study focuses on effects of untreated and treated ink industry wastewater on germination of maize, barley and sorghum. Wastewater had a high chemical oxygen demand (COD) and metal content compared to treated effluent. Germination decreased with increasing COD concentration. Speed of germination also followed the same trend, except for maize seeds exposed to untreated effluent (E), which germinated slightly faster than controls. These alterations of seedling development were mirrored by changes in soluble protein content. E exerted a positive effect on soluble protein content and maximum levels occurred after 10 days with treated effluent using coagulation/flocculation (TEc/f) process and treated effluent using combined process (coagulation/flocculation/biosorption) (TEc/f/b). Likewise, activity of α-amylase was influenced by effluent composition. Its expression depended on the species, exposure time and applied treatment. Nevertheless, current results indicated TEc/f/b had no observable toxic effects on germination and could be a beneficial alternative resource to irrigation water.} } @Article{IPB-1170, author = {Wasternack, C. and}, title = {{How Jasmonates Earned their Laurels: Past and Present}}, year = {2015}, pages = {761-794}, journal = {J. Plant Growth Regul.}, doi = {10.1007/s00344-015-9526-5}, volume = {34}, abstract = {The histories of research regarding all plant hormones are similar. Identification and structural elucidation have been followed by analyses of their biosynthesis, distributions, signaling cascades, roles in developmental or stress response programs, and crosstalk. Jasmonic acid (JA) and its derivatives comprise a group of plant hormones that were discovered recently, compared to auxin, abscisic acid, cytokinins, gibberellic acid, and ethylene. Nevertheless, there have been tremendous advances in JA research, following the general progression outlined above and parallel efforts focused on several other “new” plant hormones (brassinosteroids, salicylate, and strigolactones). This review focuses on historical aspects of the identification of jasmonates, and characterization of their biosynthesis, distribution, perception, signaling pathways, crosstalk with other hormones and roles in plant stress responses and development. The aim is to illustrate how our present knowledge on jasmonates was generated and how that influences current efforts to extend our knowledge.} } @Article{IPB-1224, author = {Jayaweera, T. and Siriwardana, C. and Dharmasiri, S. and Quint, M. and Gray, W. M. and Dharmasiri, N. and}, title = {{Alternative Splicing of Arabidopsis IBR5 Pre-mRNA Generates Two IBR5 Isoforms with Distinct and Overlapping Functions}}, year = {2014}, pages = {e102301}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0102301}, volume = {9}, abstract = {The INDOLE-3-BUTYRIC ACID RESPONSE5 (IBR5) gene encodes a dual specificity phosphatase that regulates plant auxin responses. IBR5 has been predicted to generate two transcripts through alternative splicing, but alternative splicing of IBR5 has not been confirmed experimentally. The previously characterized ibr5-1 null mutant exhibits many auxin related defects such as auxin insensitive primary root growth, defective vascular development, short stature and reduced lateral root development. However, whether all these defects are caused by the lack of phosphatase activity is not clear. Here we describe two new auxin insensitive IBR5 alleles, ibr5-4, a catalytic site mutant, and ibr5-5, a splice site mutant. Characterization of these new mutants indicates that IBR5 is post-transcriptionally regulated to generate two transcripts, AT2G04550.1 and AT2G04550.3, and consequently two IBR5 isoforms, IBR5.1 and IBR5.3. The IBR5.1 isoform exhibits phosphatase catalytic activity that is required for both proper degradation of Aux/IAA proteins and auxin-induced gene expression. These two processes are independently regulated by IBR5.1. Comparison of new mutant alleles with ibr5-1 indicates that all three mutant alleles share many phenotypes. However, each allele also confers distinct defects implicating IBR5 isoform specific functions. Some of these functions are independent of IBR5.1 catalytic activity. Additionally, analysis of these new mutant alleles suggests that IBR5 may link ABP1 and SCFTIR1/AFBs auxin signaling pathways.} } @Article{IPB-1219, author = {Grubb, C. D. and Zipp, B. J. and Kopycki, J. and Schubert, M. and Quint, M. and Lim, E.-K. and Bowles, D. J. and Pedras, M. S. C. and Abel, S. and}, title = {{Comparative analysis of Arabidopsis UGT74 glucosyltransferases reveals a special role of UGT74C1 in glucosinolate biosynthesis}}, year = {2014}, pages = {92-105}, journal = {Plant J.}, doi = {10.1111/tpj.12541}, volume = {79}, abstract = {The study of glucosinolates and their regulation has provided a powerful framework for the exploration of fundamental questions about the function, evolution, and ecological significance of plant natural products, but uncertainties about their metabolism remain. Previous work has identified one thiohydroximate S‐glucosyltransferase, UGT74B1, with an important role in the core pathway, but also made clear that this enzyme functions redundantly and cannot be the sole UDP‐glucose dependent glucosyltransferase (UGT) in glucosinolate synthesis. Here, we present the results of a nearly comprehensive in vitro activity screen of recombinant Arabidopsis Family 1 UGTs, which implicate other members of the UGT74 clade as candidate glucosinolate biosynthetic enzymes. Systematic genetic analysis of this clade indicates that UGT74C1 plays a special role in the synthesis of aliphatic glucosinolates, a conclusion strongly supported by phylogenetic and gene expression analyses. Finally, the ability of UGT74C1 to complement phenotypes and chemotypes of the ugt74b1‐2 knockout mutant and to express thiohydroximate UGT activity in planta provides conclusive evidence for UGT74C1 being an accessory enzyme in glucosinolate biosynthesis with a potential function during plant adaptation to environmental challenge.} } @Article{IPB-1213, author = {Flores, R. and Gago-Zachert, S. and Serra, P. and Sanjuán, R. and Elena, S. F. and}, title = {{Viroids: Survivors from the RNA World?}}, year = {2014}, pages = {395-414}, journal = {Annu. Rev. Microbiol.}, doi = {10.1146/annurev-micro-091313-103416}, volume = {68}, abstract = {Because RNA can be a carrier of genetic information and a biocatalyst, there is a consensus that it emerged before DNA and proteins, which eventually assumed these roles and relegated RNA to intermediate functions. If such a scenario—the so-called RNA world—existed, we might hope to find its relics in our present world. The properties of viroids that make them candidates for being survivors of the RNA world include those expected for primitive RNA replicons: (a) small size imposed by error-prone replication, (b) high G \+ C content to increase replication fidelity, (c) circular structure for assuring complete replication without genomic tags, (d) structural periodicity for modular assembly into enlarged genomes, (e) lack of protein-coding ability consistent with a ribosome-free habitat, and (f) replication mediated in some by ribozymes, the fingerprint of the RNA world. With the advent of DNA and proteins, those protoviroids lost some abilities and became the plant parasites we now know.} } @Article{IPB-1212, author = {Floková, K. and Tarkowská, D. and Miersch, O. and Strnad, M. and Wasternack, C. and Novák, O. and}, title = {{UHPLC–MS/MS based target profiling of stress-induced phytohormones}}, year = {2014}, pages = {147-157}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2014.05.015}, volume = {105}, abstract = {Stress-induced changes in phytohormone metabolite profiles have rapid effects on plant metabolic activity and growth. The jasmonates (JAs) are a group of fatty acid-derived stress response regulators with roles in numerous developmental processes. To elucidate their dual regulatory effects, which overlap with those of other important defence-signalling plant hormones such as salicylic acid (SA), abscisic acid (ABA) and indole-3-acetic acid (IAA), we have developed a highly efficient single-step clean-up procedure for their enrichment from complex plant matrices that enables their sensitive quantitative analysis using hyphenated mass spectrometry technique. The rapid extraction of minute quantities of plant material (less than 20 mg fresh weight, FW) into cold 10% methanol followed by one-step reversed-phase polymer-based solid phase extraction significantly reduced matrix effects and increased the recovery of labile JA analytes. This extraction and purification protocol was paired with a highly sensitive and validated ultra-high performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) method and used to simultaneously profile sixteen stress-induced phytohormones in minute plant material samples, including endogenous JA, several of its biosynthetic precursors and derivatives, as well as SA, ABA and IAA.} } @Article{IPB-1211, author = {Farmer, E. E. and Gasperini, D. and Acosta, I. F. and}, title = {{The squeeze cell hypothesis for the activation of jasmonate synthesis in response to wounding}}, year = {2014}, pages = {282-288}, journal = {New Phytol.}, doi = {10.1111/nph.12897}, volume = {204}, abstract = {Jasmonates are lipid mediators that control defence gene expression in response to wounding and other environmental stresses. These small molecules can accumulate at distances up to several cm from sites of damage and this is likely to involve cell‐to‐cell jasmonate transport. Also, and independently of jasmonate synthesis, transport and perception, different long‐distance wound signals that stimulate distal jasmonate synthesis are propagated at apparent speeds of several cm min–1 to tissues distal to wounds in a mechanism that involves clade 3 GLUTAMATE RECEPTOR‐LIKE (GLR) genes. A search for jasmonate synthesis enzymes that might decode these signals revealed LOX6, a lipoxygenase that is necessary for much of the rapid accumulation of jasmonic acid at sites distal to wounds. Intriguingly, the LOX6 promoter is expressed in a distinct niche of cells that are adjacent to mature xylem vessels, a location that would make these contact cells sensitive to the release of xylem water column tension upon wounding. We propose a model in which rapid axial changes in xylem hydrostatic pressure caused by wounding travel through the vasculature and lead to slower, radially dispersed pressure changes that act in a clade 3 GLR‐dependent mechanism to promote distal jasmonate synthesis.} } @Article{IPB-1205, author = {Erickson, J. L. and Ziegler, J. and Guevara, D. and Abel, S. and Klösgen, R. B. and Mathur, J. and Rothstein, S. J. and Schattat, M. H. and}, title = {{Agrobacterium-derived cytokinin influences plastid morphology and starch accumulation in Nicotiana benthamiana during transient assays}}, year = {2014}, pages = {127}, journal = {BMC Plant Biol.}, doi = {10.1186/1471-2229-14-127}, volume = {14}, abstract = {BackgroundAgrobacterium tumefaciens-based transient assays have become a common tool for answering questions related to protein localization and gene expression in a cellular context. The use of these assays assumes that the transiently transformed cells are observed under relatively authentic physiological conditions and maintain ‘normal’ sub-cellular behaviour. Although this premise is widely accepted, the question of whether cellular organization and organelle morphology is altered in Agrobacterium-infiltrated cells has not been examined in detail. The first indications of an altered sub-cellular environment came from our observation that a common laboratory strain, GV3101(pMP90), caused a drastic increase in stromule frequency. Stromules, or ‘stroma-filled-tubules’ emanate from the surface of plastids and are sensitive to a variety of biotic and abiotic stresses. Starting from this observation, the goal of our experiments was to further characterize the changes to the cell resulting from short-term bacterial infestation, and to identify the factor responsible for eliciting these changes.ResultsUsing a protocol typical of transient assays we evaluated the impact of GV3101(pMP90) infiltration on chloroplast behaviour and morphology in Nicotiana benthamiana. Our experiments confirmed that GV3101(pMP90) consistently induces stromules and alters plastid position relative to the nucleus. These effects were found to be the result of strain-dependant secretion of cytokinin and its accumulation in the plant tissue. Bacterial production of the hormone was found to be dependant on the presence of a trans-zeatin synthase gene (tzs) located on the Ti plasmid of GV3101(pMP90). Bacteria-derived cytokinins were also correlated with changes to both soluble sugar level and starch accumulation.ConclusionAlthough we have chosen to focus on how transient Agrobacterium infestation alters plastid based parameters, these changes to the morphology and position of a single organelle, combined with the measured increases in sugar and starch content, suggest global changes to cell physiology. This indicates that cells visualized during transient assays may not be as ‘normal’ as was previously assumed. Our results suggest that the impact of the bacteria can be minimized by choosing Agrobacterium strains devoid of the tzs gene, but that the alterations to sub-cellular organization and cell carbohydrate status cannot be completely avoided using this strategy.} } @Article{IPB-1199, author = {Delker, C. and Sonntag, L. and James, G. and Janitza, P. and Ibañez, C. and Ziermann, H. and Peterson, T. and Denk, K. and Mull, S. and Ziegler, J. and Davis, S. and Schneeberger, K. and Quint, M. and}, title = {{The DET1-COP1-HY5 Pathway Constitutes a Multipurpose Signaling Module Regulating Plant Photomorphogenesis and Thermomorphogenesis}}, year = {2014}, pages = {1983-1989}, journal = {Cell Rep.}, doi = {10.1016/j.celrep.2014.11.043}, volume = {9}, abstract = {Developmental plasticity enables plants to respond to elevated ambient temperatures by adapting their shoot architecture. On the cellular level, the basic-helix-loop-helix (bHLH) transcription factor PHYTOCHROME INTERACTING FACTOR 4 (PIF4) coordinates this response by activating hormonal modules that in turn regulate growth. In addition to an unknown temperature-sensing mechanism, it is currently not understood how temperature regulates PIF4 activity. Using a forward genetic approach in Arabidopsis thaliana, we present extensive genetic evidence demonstrating that the DE-ETIOLATED 1 (DET1)-CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1)-ELONGATED HYPOCOTYL 5 (HY5)-dependent photomorphogenesis pathway transcriptionally regulates PIF4 to coordinate seedling growth in response to elevated temperature. Our findings demonstrate that two of the most prevalent environmental cues, light and temperature, share a much larger set of signaling components than previously assumed. Similar to the toolbox concept in animal embryonic patterning, multipurpose signaling modules might have evolved in plants to translate various environmental stimuli into adaptational growth processes.} } @Article{IPB-1193, author = {Budiharjo, A. and Chowdhury, S. P. and Dietel, K. and Beator, B. and Dolgova, O. and Fan, B. and Bleiss, W. and Ziegler, J. and Schmid, M. and Hartmann, A. and Borriss, R. and}, title = {{Transposon Mutagenesis of the Plant-Associated Bacillus amyloliquefaciens ssp. plantarum FZB42 Revealed That the nfrA and RBAM17410 Genes Are Involved in Plant-Microbe-Interactions}}, year = {2014}, pages = {e98267}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0098267}, volume = {9}, abstract = {Bacillus amyloliquefaciens ssp. plantarum FZB42 represents the prototype of Gram-positive plant growth promoting and biocontrol bacteria. In this study, we applied transposon mutagenesis to generate a transposon library, which was screened for genes involved in multicellular behavior and biofilm formation on roots as a prerequisite of plant growth promoting activity. Transposon insertion sites were determined by rescue-cloning followed by DNA sequencing. As in B. subtilis, the global transcriptional regulator DegU was identified as an activator of genes necessary for swarming and biofilm formation, and the DegU-mutant of FZB42 was found impaired in efficient root colonization. Direct screening of 3,000 transposon insertion mutants for plant-growth-promotion revealed the gene products of nfrA and RBAM\_017140 to be essential for beneficial effects exerted by FZB42 on plants. We analyzed the performance of GFP-labeled wild-type and transposon mutants in the colonization of lettuce roots using confocal laser scanning microscopy. While the wild-type strain heavily colonized root surfaces, the nfrA mutant did not colonize lettuce roots, although it was not impaired in growth in laboratory cultures, biofilm formation and swarming motility on agar plates. The RBAM17410 gene, occurring in only a few members of the B. subtilis species complex, was directly involved in plant growth promotion. None of the mutant strains were affected in producing the plant growth hormone auxin. We hypothesize that the nfrA gene product is essential for overcoming the stress caused by plant response towards bacterial root colonization.} } @Article{IPB-1187, author = {Bosch, M. and Wright, L. P. and Gershenzon, J. and Wasternack, C. and Hause, B. and Schaller, A. and Stintzi, A. and}, title = {{Jasmonic Acid and Its Precursor 12-Oxophytodienoic Acid Control Different Aspects of Constitutive and Induced Herbivore Defenses in Tomato}}, year = {2014}, pages = {396-410}, journal = {Plant Physiol.}, doi = {10.1104/pp.114.237388}, volume = {166}, abstract = {The jasmonate family of growth regulators includes the isoleucine (Ile) conjugate of jasmonic acid (JA-Ile) and its biosynthetic precursor 12-oxophytodienoic acid (OPDA) as signaling molecules. To assess the relative contribution of JA/JA-Ile and OPDA to insect resistance in tomato (Solanum lycopersicum), we silenced the expression of OPDA reductase3 (OPR3) by RNA interference (RNAi). Consistent with a block in the biosynthetic pathway downstream of OPDA, OPR3-RNAi plants contained wild-type levels of OPDA but failed to accumulate JA or JA-Ile after wounding. JA/JA-Ile deficiency in OPR3-RNAi plants resulted in reduced trichome formation and impaired monoterpene and sesquiterpene production. The loss of these JA/JA-Ile -dependent defense traits rendered them more attractive to the specialist herbivore Manduca sexta with respect to feeding and oviposition. Oviposition preference resulted from reduced levels of repellant monoterpenes and sesquiterpenes. Feeding preference, on the other hand, was caused by increased production of cis-3-hexenal acting as a feeding stimulant for M. sexta larvae in OPR3-RNAi plants. Despite impaired constitutive defenses and increased palatability of OPR3-RNAi leaves, larval development was indistinguishable on OPR3-RNAi and wild-type plants, and was much delayed compared with development on the jasmonic acid-insensitive1 (jai1) mutant. Apparently, signaling through JAI1, the tomato ortholog of the ubiquitin ligase CORONATINE INSENSITIVE1 in Arabidopsis (Arabidopsis thaliana), is required for defense, whereas the conversion of OPDA to JA/JA-Ile is not. Comparing the signaling activities of OPDA and JA/JA-Ile, we found that OPDA can substitute for JA/JA-Ile in the local induction of defense gene expression, but the production of JA/JA-Ile is required for a systemic response.} } @Article{IPB-1181, author = {Antolín-Llovera, M. and Petutsching, E. K. and Ried, M. K. and Lipka, V. and Nürnberger, T. and Robatzek, S. and Parniske, M. and}, title = {{Knowing your friends and foes - plant receptor-like kinases as initiators of symbiosis or defence}}, year = {2014}, pages = {791-802}, journal = {New Phytol.}, doi = {10.1111/nph.13117}, volume = {204}, abstract = {The decision between defence and symbiosis signalling in plants involves alternative and modular plasma membrane‐localized receptor complexes. A critical step in their activation is ligand‐induced homo‐ or hetero‐oligomerization of leucine‐rich repeat (LRR)‐ and/or lysin motif (LysM) receptor‐like kinases (RLKs). In defence signalling, receptor complexes form upon binding of pathogen‐associated molecular patterns (PAMPs), including the bacterial flagellin‐derived peptide flg22, or chitin. Similar mechanisms are likely to operate during the perception of microbial symbiont‐derived (lipo)‐chitooligosaccharides. The structurally related chitin‐oligomer ligands chitooctaose and chitotetraose trigger defence and symbiosis signalling, respectively, and their discrimination involves closely related, if not identical, LysM‐RLKs. This illustrates the demand for and the challenges imposed on decision mechanisms that ensure appropriate signal initiation. Appropriate signalling critically depends on abundance and localization of RLKs at the cell surface. This is regulated by internalization, which also provides a mechanism for the removal of activated signalling RLKs. Abundance of the malectin‐like domain (MLD)‐LRR‐RLK Symbiosis Receptor‐like Kinase (SYMRK) is additionally controlled by cleavage of its modular ectodomain, which generates a truncated and rapidly degraded RLK fragment. This review explores LRR‐ and LysM‐mediated signalling, the involvement of MLD‐LRR‐RLKs in symbiosis and defence, and the role of endocytosis in RLK function.} } @Article{IPB-1180, author = {Antolín-Llovera, M. and Ried, M. K. and Parniske, M. and}, title = {{Cleavage of the SYMBIOSIS RECEPTOR-LIKE KINASE Ectodomain Promotes Complex Formation with Nod Factor Receptor 5}}, year = {2014}, pages = {422-427}, journal = {Curr. Biol.}, doi = {10.1016/j.cub.2013.12.053}, volume = {24}, abstract = {Plants form root symbioses with fungi and bacteria to improve their nutrient supply. SYMBIOSIS RECEPTOR-LIKE KINASE (SYMRK) is required for phosphate-acquiring arbuscular mycorrhiza, as well as for the nitrogen-fixing root nodule symbiosis of legumes [1] and actinorhizal plants [2, 3], but its precise function was completely unclear. Here we show that the extracytoplasmic region of SYMRK, which comprises three leucine-rich repeats (LRRs) and a malectin-like domain (MLD) related to a carbohydrate-binding protein from Xenopus laevis [4], is cleaved to release the MLD in the absence of symbiotic stimulation. A conserved sequence motif—GDPC—that connects the MLD to the LRRs is required for MLD release. We discovered that Nod factor receptor 5 (NFR5) [5, 6, 7, 8] forms a complex with the SYMRK version that remains after MLD release (SYMRK-ΔMLD). SYMRK-ΔMLD outcompeted full-length SYMRK for NFR5 interaction, indicating that the MLD negatively interferes with complex formation. SYMRK-ΔMLD is present at lower amounts than MLD, suggesting rapid degradation after MLD release. A deletion of the entire extracytoplasmic region increased protein abundance, suggesting that the LRR region promotes degradation. Curiously, this deletion led to excessive infection thread formation, highlighting the importance of fine-tuned regulation of SYMRK by its ectodomain.} } @Article{IPB-1252, author = {Ried, M. K. and Antolín-Llovera, M. and Parniske, M. and}, title = {{Spontaneous symbiotic reprogramming of plant roots triggered by receptor-like kinases}}, year = {2014}, pages = {e03891}, journal = {eLife}, doi = {10.7554/eLife.03891}, volume = {3}, abstract = {Symbiosis Receptor-like Kinase (SYMRK) is indispensable for the development of phosphate-acquiring arbuscular mycorrhiza (AM) as well as nitrogen-fixing root nodule symbiosis, but the mechanisms that discriminate between the two distinct symbiotic developmental fates have been enigmatic. In this study, we show that upon ectopic expression, the receptor-like kinase genes Nod Factor Receptor 1 (NFR1), NFR5, and SYMRK initiate spontaneous nodule organogenesis and nodulation-related gene expression in the absence of rhizobia. Furthermore, overexpressed NFR1 or NFR5 associated with endogenous SYMRK in roots of the legume Lotus japonicus. Epistasis tests revealed that the dominant active SYMRK allele initiates signalling independently of either the NFR1 or NFR5 gene and upstream of a set of genes required for the generation or decoding of calcium-spiking in both symbioses. Only SYMRK but not NFR overexpression triggered the expression of AM-related genes, indicating that the receptors play a key role in the decision between AM- or root nodule symbiosis-development.} } @Article{IPB-1234, author = {Maldonado-Bonilla, L. D. and Eschen-Lippold, L. and Gago-Zachert, S. and Tabassum, N. and Bauer, N. and Scheel, D. and Lee, J. and}, title = {{The Arabidopsis Tandem Zinc Finger 9 Protein Binds RNA and Mediates Pathogen-Associated Molecular Pattern-Triggered Immune Responses}}, year = {2014}, pages = {412-425}, journal = {Plant Cell Physiol.}, doi = {10.1093/pcp/pct175}, volume = {55}, abstract = {Recognition of pathogen-associated molecular patterns (PAMPs) induces multiple defense mechanisms to limit pathogen growth. Here, we show that the Arabidopsis thaliana tandem zinc finger protein 9 (TZF9) is phosphorylated by PAMP-responsive mitogen-activated protein kinases (MAPKs) and is required to trigger a full PAMP-triggered immune response. Analysis of a tzf9 mutant revealed attenuation in specific PAMP-triggered reactions such as reactive oxygen species accumulation, MAPK activation and, partially, the expression of several PAMP-responsive genes. In accordance with these weaker PAMP-triggered responses, tzf9 mutant plants exhibit enhanced susceptibility to virulent Pseudomonas syringae pv. tomato DC3000. Visualization of TZF9 localization by fusion to green fluorescent protein revealed cytoplasmic foci that co-localize with marker proteins of processing bodies (P-bodies). This localization pattern is affected by inhibitor treatments that limit mRNA availability (such as cycloheximide or actinomycin D) or block nuclear export (leptomycin B). Coupled with its ability to bind the ribohomopolymers poly(rU) and poly(rG), these results suggest involvement of TZF9 in post-transcriptional regulation, such as mRNA processing or storage pathways, to regulate plant innate immunity.} } @Article{IPB-1281, author = {Ziegler, J. and Qwegwer, J. and Schubert, M. and Erickson, J. L. and Schattat, M. and Bürstenbinder, K. and Grubb, C. D. and Abel, S. and}, title = {{Simultaneous analysis of apolar phytohormones and 1-aminocyclopropan-1-carboxylic acid by high performance liquid chromatography/electrospray negative ion tandem mass spectrometry via 9-fluorenylmethoxycarbonyl chloride derivatization}}, year = {2014}, pages = {102-109}, journal = {J. Chromatogr. A}, doi = {10.1016/j.chroma.2014.08.029}, volume = {1362}, abstract = {A strategy to detect and quantify the polar ethylene precursor 1-aminocyclopropan-1-carboxylic acid (ACC) along with the more apolar phytohormones abscisic acid (ABA), indole-3-acetic acid (IAA), jasmonic acid (JA), jasmonic acid-isoleucine conjugate (JA-Ile), 12-oxo-phytodienoic acid (OPDA), trans-zeatin, and trans-zeatin 9-riboside using a single extraction is presented. Solid phase resins commonly employed for extraction of phytohormones do not allow the recovery of ACC. We circumvent this problem by attaching an apolar group to ACC via derivatization with the amino group specific reagent 9-fluorenylmethoxycarbonyl chloride (Fmoc-Cl). Derivatization in the methanolic crude extract does not modify other phytohormones. The derivatized ACC could be purified and detected together with the more apolar phytohormones using common solid phase extraction resins and reverse phase HPLC/electrospray negative ion tandem mass spectrometry. The limit of detection was in the low nanomolar range for all phytohormones, a sensitivity sufficient to accurately determine the phytohormone levels from less than 50 mg (fresh weight) of Arabidopsis thaliana and Nicotiana benthamiana tissues. Comparison with previously published phytohormone levels and the reported changes in phytohormone levels after stress treatments confirmed the accuracy of the method.} } @Article{IPB-1280, author = {Ziegler, J. and Abel, S. and}, title = {{Analysis of amino acids by HPLC/electrospray negative ion tandem mass spectrometry using 9-fluorenylmethoxycarbonyl chloride (Fmoc-Cl) derivatization}}, year = {2014}, pages = {2799-2808}, journal = {Amino Acids}, doi = {10.1007/s00726-014-1837-5}, volume = {46}, abstract = {A new method for the determination of amino acids is presented. It combines established methods for the derivatization of primary and secondary amino groups with 9-fluorenylmethoxycarbonyl chloride (Fmoc-Cl) with the subsequent amino acid specific detection of the derivatives by LC–ESI–MS/MS using multiple reaction monitoring (MRM). The derivatization proceeds within 5 min, and the resulting amino acid derivatives can be rapidly purified from matrix by solid-phase extraction (SPE) on HR-X resin and separated by reversed-phase HPLC. The Fmoc derivatives yield several amino acid specific fragment ions which opened the possibility to select amino acid specific MRM transitions. The method was applied to all 20 proteinogenic amino acids, and the quantification was performed using l-norvaline as standard. A limit of detection as low as 1 fmol/µl with a linear range of up to 125 pmol/µl could be obtained. Intraday and interday precisions were lower than 10 % relative standard deviations for most of the amino acids. Quantification using l-norvaline as internal standard gave very similar results compared to the quantification using deuterated amino acid as internal standards. Using this protocol, it was possible to record the amino acid profiles of only a single root from Arabidopsis thaliana seedlings and to compare it with the amino acid profiles of 20 dissected root meristems (200 μm).} } @Article{IPB-1275, author = {Wasternack, C. and}, title = {{Perception, signaling and cross-talk of jasmonates and the seminal contributions of the Daoxin Xie’s lab and the Chuanyou Li’s lab}}, year = {2014}, pages = {707-718}, journal = {Plant Cell Rep.}, doi = {10.1007/s00299-014-1608-5}, volume = {33}, abstract = {Jasmonates (JAs) are lipid-derived signals in plant responses to biotic and abiotic stresses and in development. The most active JA compound is (\+)-7-iso-JA-Ile, a JA conjugate with isoleucine. Biosynthesis, metabolism and key components of perception and signal transduction have been identified and numerous JA-induced gene expression data collected. For JA-Ile perception, the SCFCOI1–JAZ co-receptor complex has been identified and crystalized. Activators such as MYC2 and repressors such as JAZs including their targets were found. Involvement of JA-Ile in response to herbivores and pathogens and in root growth inhibition is among the most studied aspects of JA-Ile signaling. There are an increasing number of examples, where JA-Ile shows cross-talk with other plant hormones. Seminal contributions in JA/JA-Ile research were given by Daoxin Xie’s lab and Chuanyou Li’s lab, both in Beijing. Here, characterization was done regarding components of the JA-Ile receptor, such as COI1 (JAI1) and SCF, regarding activators (MYCs, MYBs) and repressors (JAV1, bHLH IIId’s) of JA-regulated gene expression, as well as regarding components of auxin biosynthesis and action, such as the transcription factor PLETHORA active in the root stem cell niche. This overview reflects the work of both labs in the light of our present knowledge on biosynthesis, perception and signal transduction of JA/JA-Ile and its cross-talk to other hormones.} } @Article{IPB-1274, author = {Wasternack, C. and Hause, B. and}, title = {{Blütenduft, Abwehr, Entwicklung: Jasmonsäure - ein universelles Pflanzenhormon}}, year = {2014}, pages = {164-171}, journal = {Biologie in unserer Zeit}, doi = {10.1002/biuz.201410535}, volume = {44}, abstract = {Pflanzen müssen gegen vielfältige biotische und abiotische Umwelteinflusse eine Abwehr aufbauen. Aber gleichzeitig müssen sie wachsen und sich vermehren. Jasmonate sind neben anderen Hormonen ein zentrales Signal bei der Etablierung von Abwehrmechanismen, aber auch Signal von Entwicklungsprozessen wie Blüten‐ und Trichombildung, sowie der Hemmung von Wachstum. Biosynthese und essentielle Komponenten der Signaltransduktion von JA und seinem biologisch aktiven Konjugat JA‐Ile sind gut untersucht. Der Rezeptor ist ein Proteinkomplex, der “JA‐Ile‐Wahrnehmung” mit proteasomalem Abbau von Repressorproteinen verbindet. Dadurch können positiv agierende Transkriptionsfaktoren wirksam werden und vielfältige Genexpressionsänderungen auslösen. Dies betrifft die Bildung von Abwehrproteinen, Enzymen der JA‐Biosynthese und Sekundärstoffbildung, und Proteinen von Signalketten und Entwicklungsprozessen. Die Kenntnisse zur JA‐Ile‐Wirkung werden in Landwirtschaft und Biotechnologie genutzt.} } @Article{IPB-1273, author = {Wasternack, C. and}, title = {{Action of jasmonates in plant stress responses and development — Applied aspects}}, year = {2014}, pages = {31-39}, journal = {Biotechnol. Adv.}, doi = {10.1016/j.biotechadv.2013.09.009}, volume = {32}, abstract = {Jasmonates (JAs) are lipid-derived compounds acting as key signaling compounds in plant stress responses and development. The JA co-receptor complex and several enzymes of JA biosynthesis have been crystallized, and various JA signal transduction pathways including cross-talk to most of the plant hormones have been intensively studied. Defense to herbivores and necrotrophic pathogens are mediated by JA. Other environmental cues mediated by JA are light, seasonal and circadian rhythms, cold stress, desiccation stress, salt stress and UV stress. During development growth inhibition of roots, shoots and leaves occur by JA, whereas seed germination and flower development are partially affected by its precursor 12-oxo-phytodienoic acid (OPDA). Based on these numerous JA mediated signal transduction pathways active in plant stress responses and development, there is an increasing interest in horticultural and biotechnological applications. Intercropping, the mixed growth of two or more crops, mycorrhization of plants, establishment of induced resistance, priming of plants for enhanced insect resistance as well as pre- and post-harvest application of JA are few examples. Additional sources for horticultural improvement, where JAs might be involved, are defense against nematodes, biocontrol by plant growth promoting rhizobacteria, altered composition of rhizosphere bacterial community, sustained balance between growth and defense, and improved plant immunity in intercropping systems. Finally, biotechnological application for JA-induced production of pharmaceuticals and application of JAs as anti-cancer agents were intensively studied.} } @Article{IPB-1266, author = {Song, S. and Qi, T. and Wasternack, C. and Xie, D. and}, title = {{Jasmonate signaling and crosstalk with gibberellin and ethylene}}, year = {2014}, pages = {112-119}, journal = {Curr. Opin. Plant Biol.}, doi = {10.1016/j.pbi.2014.07.005}, volume = {21}, abstract = {The phytohormone jasmonate (JA) plays essential roles in plant growth, development and defense. In response to the JA signal, the CORONATINE INSENSITIVE 1 (COI1)-based SCF complexes recruit JASMONATE ZIM-domain (JAZ) repressors for ubiquitination and degradation, and subsequently regulate their downstream signaling components essential for various JA responses. Tremendous progress has been made in understanding the JA signaling pathway and its crosstalk with other phytohormone pathways during the past two decades. Recent studies have revealed that a variety of positive and negative regulators act as targets of JAZs to control distinctive JA responses, and that JAZs and these regulators function as crucial interfaces to mediate synergy and antagonism between JA and other phytohormones. Owing to different regulatory players in JA perception and JA signaling, a fine-tuning of JA-dependent processes in plant growth, development and defense is achieved. In this review, we will summarize the latest progresses in JA signaling and its crosstalk with gibberellin and ethylene.} } @Article{IPB-1326, author = {Kopycki, J. and Wieduwild, E. and Kohlschmidt, J. and Brandt, W. and Stepanova, A. and Alonso, J. and Pedras, M. S. and Abel, S. and Grubb, C. D. and}, title = {{Kinetic analysis of Arabidopsis glucosyltransferase UGT74B1 illustrates a general mechanism by which enzymes can escape product inhibition}}, year = {2013}, pages = {37-46}, journal = {Biochem. J.}, doi = {10.1042/BJ20121403}, volume = {450}, abstract = {Plant genomes encode numerous small molecule glycosyltransferases which modulate the solubility, activity, immunogenicity and/or reactivity of hormones, xenobiotics and natural products. The products of these enzymes can accumulate to very high concentrations, yet somehow avoid inhibiting their own biosynthesis. Glucosyltransferase UGT74B1 (UDP-glycosyltransferase 74B1) catalyses the penultimate step in the core biosynthetic pathway of glucosinolates, a group of natural products with important functions in plant defence against pests and pathogens. We found that mutation of the highly conserved Ser284 to leucine [wei9-1 (weak ethylene insensitive)] caused only very mild morphological and metabolic phenotypes, in dramatic contrast with knockout mutants, indicating that steady state glucosinolate levels are actively regulated even in unchallenged plants. Analysis of the effects of the mutation via a structural modelling approach indicated that the affected serine interacts directly with UDP-glucose, but also predicted alterations in acceptor substrate affinity and the kcat value, sparking an interest in the kinetic behaviour of the wild-type enzyme. Initial velocity and inhibition studies revealed that UGT74B1 is not inhibited by its glycoside product. Together with the effects of the missense mutation, these findings are most consistent with a partial rapid equilibrium ordered mechanism. This model explains the lack of product inhibition observed both in vitro and in vivo, illustrating a general mechanism whereby enzymes can continue to function even at very high product/precursor ratios.} } @Article{IPB-1318, author = {Huang, H. and Quint, M. and Gray, W. M. and}, title = {{The eta7/csn3-3 Auxin Response Mutant of Arabidopsis Defines a Novel Function for the CSN3 Subunit of the COP9 Signalosome}}, year = {2013}, pages = {e66578}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0066578}, volume = {8}, abstract = {The COP9 signalosome (CSN) is an eight subunit protein complex conserved in all higher eukaryotes. In Arabidopsis thaliana, the CSN regulates auxin response by removing the ubiquitin-like protein NEDD8/RUB1 from the CUL1 subunit of the SCFTIR1/AFB ubiquitin-ligase (deneddylation). Previously described null mutations in any CSN subunit result in the pleiotropic cop/det/fus phenotype and cause seedling lethality, hampering the study of CSN functions in plant development. In a genetic screen to identify enhancers of the auxin response defects conferred by the tir1-1 mutation, we identified a viable csn mutant of subunit 3 (CSN3), designated eta7/csn3-3. In addition to enhancing tir1-1 mutant phenotypes, the csn3-3 mutation alone confers several phenotypes indicative of impaired auxin signaling including auxin resistant root growth and diminished auxin responsive gene expression. Unexpectedly however, csn3-3 plants are not defective in either the CSN-mediated deneddylation of CUL1 or in SCFTIR1-mediated degradation of Aux/IAA proteins. These findings suggest that csn3-3 is an atypical csn mutant that defines a novel CSN or CSN3-specific function. Consistent with this possibility, we observe dramatic differences in double mutant interactions between csn3-3 and other auxin signaling mutants compared to another weak csn mutant, csn1-10. Lastly, unlike other csn mutants, assembly of the CSN holocomplex is unaffected in csn3-3 plants. However, we detected a small CSN3-containing protein complex that is altered in csn3-3 plants. We hypothesize that in addition to its role in the CSN as a cullin deneddylase, CSN3 functions in a distinct protein complex that is required for proper auxin signaling.} } @Article{IPB-1301, author = {Elleuch, A. and Chaâbene, Z. and Grubb, D. C. and Drira, N. and Mejdoub, H. and Khemakhem, B. and}, title = {{Morphological and biochemical behavior of fenugreek (Trigonella foenum-graecum) under copper stress}}, year = {2013}, pages = {46-53}, journal = {Ecotoxicol. Environ. Saf.}, doi = {10.1016/j.ecoenv.2013.09.028}, volume = {98}, abstract = {The effects of copper on germination and growth of fenugreek (Trigonella foenum-graecum) was investigated separately using different concentrations of CuSO4. The germination percentage and radical length had different responses to cupric ions: the root growth increased with increasing copper concentration up to 1 mM and Cu2\+ was inhibited thereafter. In contrast, the germination percentage was largely unaffected by concentrations of copper below 10 mM.The reduction in root growth may have been due to inhibition of hydrolytic enzymes such as amylase. Indeed, the average total amylolytic activity decreased from the first day of treatment with [Cu2\+] greater than 1 mM. Furthermore, copper affected various plant growth parameters. Copper accumulation was markedly higher in roots as compared to shoots. While both showed a gradual decrease in growth, this was more pronounced in roots than in leaves and in stems. Excess copper induced an increase in the rate of hydrogen peroxide (H2O2) production and lipid peroxidation in all plant parts, indicating oxidative stress. This redox stress affected leaf chlorophyll and carotenoid content which decreased in response to augmented Cu levels. Additionally, the activities of proteins involved in reactive oxygen species (ROS) detoxification were affected. Cu stress elevated the ascorbate peroxidase (APX) activity more than two times at 10 mM CuSO4. In contrast, superoxide dismutase (SOD) and catalase (CAT) levels showed only minor variations, only at 1 mM Cu2\+. Likewise, total phenol and flavonoid contents were strongly induced by low concentrations of copper, consistent with the role of these potent antioxidants in scavenging ROS such as H2O2, but returned to control levels or below at high [Cu2\+]. Taken together, these results indicate a fundamental shift in the plant response to copper toxicity at low versus high concentrations.} } @Article{IPB-1298, author = {Dekkers, B. J. and Pearce, S. and van Bolderen-Veldkamp, R. and Marshall, A. and Widera, P. and Gilbert, J. and Drost, H.-G. and Bassel, G. W. and Müller, K. and King, J. R. and Wood, A. T. and Grosse, I. and Quint, M. and Krasnogor, N. and Leubner-Metzger, G. and Holdsworth, M. J. and Bentsink, L. and}, title = {{Transcriptional Dynamics of Two Seed Compartments with Opposing Roles in Arabidopsis Seed Germination}}, year = {2013}, pages = {205-215}, journal = {Plant Physiol.}, doi = {10.1104/pp.113.223511}, volume = {163}, abstract = {Seed germination is a critical stage in the plant life cycle and the first step toward successful plant establishment. Therefore, understanding germination is of important ecological and agronomical relevance. Previous research revealed that different seed compartments (testa, endosperm, and embryo) control germination, but little is known about the underlying spatial and temporal transcriptome changes that lead to seed germination. We analyzed genome-wide expression in germinating Arabidopsis (Arabidopsis thaliana) seeds with both temporal and spatial detail and provide Web-accessible visualizations of the data reported (vseed.nottingham.ac.uk). We show the potential of this high-resolution data set for the construction of meaningful coexpression networks, which provide insight into the genetic control of germination. The data set reveals two transcriptional phases during germination that are separated by testa rupture. The first phase is marked by large transcriptome changes as the seed switches from a dry, quiescent state to a hydrated and active state. At the end of this first transcriptional phase, the number of differentially expressed genes between consecutive time points drops. This increases again at testa rupture, the start of the second transcriptional phase. Transcriptome data indicate a role for mechano-induced signaling at this stage and subsequently highlight the fates of the endosperm and radicle: senescence and growth, respectively. Finally, using a phylotranscriptomic approach, we show that expression levels of evolutionarily young genes drop during the first transcriptional phase and increase during the second phase. Evolutionarily old genes show an opposite pattern, suggesting a more conserved transcriptome prior to the completion of germination.} } @Article{IPB-1359, author = {Wasternack, C. and Hause, B. and}, title = {{Jasmonates: biosynthesis, perception, signal transduction and action in plant stress response, growth and development. An update to the 2007 review in Annals of Botany}}, year = {2013}, pages = {1021-1058}, journal = {Ann. Bot.}, doi = {10.1093/aob/mct067}, volume = {111}, abstract = {BackgroundJasmonates are important regulators in plant responses to biotic and abiotic stresses as well as in development. Synthesized from lipid-constituents, the initially formed jasmonic acid is converted to different metabolites including the conjugate with isoleucine. Important new components of jasmonate signalling including its receptor were identified, providing deeper insight into the role of jasmonate signalling pathways in stress responses and development.ScopeThe present review is an update of the review on jasmonates published in this journal in 2007. New data of the last five years are described with emphasis on metabolites of jasmonates, on jasmonate perception and signalling, on cross-talk to other plant hormones and on jasmonate signalling in response to herbivores and pathogens, in symbiotic interactions, in flower development, in root growth and in light perception.ConclusionsThe last few years have seen breakthroughs in the identification of JASMONATE ZIM DOMAIN (JAZ) proteins and their interactors such as transcription factors and co-repressors, and the crystallization of the jasmonate receptor as well as of the enzyme conjugating jasmonate to amino acids. Now, the complex nature of networks of jasmonate signalling in stress responses and development including hormone cross-talk can be addressed.} } @Article{IPB-1358, author = {Wasternack, C. and Forner, S. and Strnad, M. and Hause, B. and}, title = {{Jasmonates in flower and seed development}}, year = {2013}, pages = {79-85}, journal = {Biochimie}, doi = {10.1016/j.biochi.2012.06.005}, volume = {95}, abstract = {Jasmonates are ubiquitously occurring lipid-derived signaling compounds active in plant development and plant responses to biotic and abiotic stresses. Upon environmental stimuli jasmonates are formed and accumulate transiently. During flower and seed development, jasmonic acid (JA) and a remarkable number of different metabolites accumulate organ- and tissue specifically. The accumulation is accompanied with expression of jasmonate-inducible genes. Among these genes there are defense genes and developmentally regulated genes. The profile of jasmonate compounds in flowers and seeds covers active signaling molecules such as JA, its precursor 12-oxophytodienoic acid (OPDA) and amino acid conjugates such as JA-Ile, but also inactive signaling molecules occur such as 12-hydroxy-JA and its sulfated derivative. These latter compounds can occur at several orders of magnitude higher level than JA. Metabolic conversion of JA and JA-Ile to hydroxylated compounds seems to inactivate JA signaling, but also specific functions of jasmonates in flower and seed development were detected. In tomato OPDA is involved in embryo development. Occurrence of jasmonates, expression of JA-inducible genes and JA-dependent processes in flower and seed development will be discussed.} } @Article{IPB-1337, author = {Poeschl, Y. and Delker, C. and Trenner, J. and Ullrich, K. K. and Quint, M. and Grosse, I. and}, title = {{Optimized Probe Masking for Comparative Transcriptomics of Closely Related Species}}, year = {2013}, pages = {e78497}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0078497}, volume = {8}, abstract = {Microarrays are commonly applied to study the transcriptome of specific species. However, many available microarrays are restricted to model organisms, and the design of custom microarrays for other species is often not feasible. Hence, transcriptomics approaches of non-model organisms as well as comparative transcriptomics studies among two or more species often make use of cost-intensive RNAseq studies or, alternatively, by hybridizing transcripts of a query species to a microarray of a closely related species. When analyzing these cross-species microarray expression data, differences in the transcriptome of the query species can cause problems, such as the following: (i) lower hybridization accuracy of probes due to mismatches or deletions, (ii) probes binding multiple transcripts of different genes, and (iii) probes binding transcripts of non-orthologous genes. So far, methods for (i) exist, but these neglect (ii) and (iii). Here, we propose an approach for comparative transcriptomics addressing problems (i) to (iii), which retains only transcript-specific probes binding transcripts of orthologous genes. We apply this approach to an Arabidopsis lyrata expression data set measured on a microarray designed for Arabidopsis thaliana, and compare it to two alternative approaches, a sequence-based approach and a genomic DNA hybridization-based approach. We investigate the number of retained probe sets, and we validate the resulting expression responses by qRT-PCR. We find that the proposed approach combines the benefit of sequence-based stringency and accuracy while allowing the expression analysis of much more genes than the alternative sequence-based approach. As an added benefit, the proposed approach requires probes to detect transcripts of orthologous genes only, which provides a superior base for biological interpretation of the measured expression responses.} } @Article{IPB-1333, author = {Navarro-Quezada, A. and Schumann, N. and Quint, M. and}, title = {{Plant F-Box Protein Evolution Is Determined by Lineage-Specific Timing of Major Gene Family Expansion Waves}}, year = {2013}, pages = {e68672}, journal = {PLOS ONE}, doi = {10.1371/journal.pone.0068672}, volume = {8}, abstract = {F-box proteins (FBPs) represent one of the largest and fastest evolving gene/protein families in the plant kingdom. The FBP superfamily can be divided in several subfamilies characterized by different C-terminal protein-protein interaction domains that recruit targets for proteasomal degradation. Hence, a clear picture of their phylogeny and molecular evolution is of special interest for the general understanding of evolutionary histories of multi-domain and/or large protein families in plants. In an effort to further understand the molecular evolution of F-box family proteins, we asked whether the largest subfamily in Arabidopsis thaliana, which carries a C-terminal F-box associated domain (FBA proteins) shares evolutionary patterns and signatures of selection with other FBPs. To address this question, we applied phylogenetic and molecular evolution analyses in combination with the evaluation of transcriptional profiles. Based on the 2219 FBA proteins we de novo identified in 34 completely sequenced plant genomes, we compared their evolutionary patterns to a previously analyzed large subfamily carrying C-terminal kelch repeats. We found that these two large FBP subfamilies generally tend to evolve by massive waves of duplication, followed by sequence conservation of the F-box domain and sequence diversification of the target recruiting domain. We conclude that the earlier in evolutionary time a major wave of expansion occurred, the more pronounced these selection signatures are. As a consequence, when performing cross species comparisons among FBP subfamilies, significant differences will be observed in the selective signatures of protein-protein interaction domains. Depending on the species, the investigated subfamilies comprise up to 45% of the complete superfamily, indicating that other subfamilies possibly follow similar modes of evolution.} } @Article{IPB-1285, author = {Acosta, I. F. and Gasperini, D. and Chételat, A. and Stolz, S. and Santuari, L. and Farmer, E. E. and}, title = {{Role of NINJA in root jasmonate signaling}}, year = {2013}, pages = {15473-15478}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.1307910110}, volume = {110}, abstract = {Wound responses in plants have to be coordinated between organs so that locally reduced growth in a wounded tissue is balanced by appropriate growth elsewhere in the body. We used a JASMONATE ZIM DOMAIN 10 (JAZ10) reporter to screen for mutants affected in the organ-specific activation of jasmonate (JA) signaling in Arabidopsis thaliana seedlings. Wounding one cotyledon activated the reporter in both aerial and root tissues, and this was either disrupted or restricted to certain organs in mutant alleles of core components of the JA pathway including COI1, OPR3, and JAR1. In contrast, three other mutants showed constitutive activation of the reporter in the roots and hypocotyls of unwounded seedlings. All three lines harbored mutations in Novel Interactor of JAZ (NINJA), which encodes part of a repressor complex that negatively regulates JA signaling. These ninja mutants displayed shorter roots mimicking JA-mediated growth inhibition, and this was due to reduced cell elongation. Remarkably, this phenotype and the constitutive JAZ10 expression were still observed in backgrounds lacking the ability to synthesize JA or the key transcriptional activator MYC2. Therefore, JA-like responses can be recapitulated in specific tissues without changing a plant’s ability to make or perceive JA, and MYC2 either has no role or is not the only derepressed transcription factor in ninja mutants. Our results show that the role of NINJA in the root is to repress JA signaling and allow normal cell elongation. Furthermore, the regulation of the JA pathway differs between roots and aerial tissues at all levels, from JA biosynthesis to transcriptional activation.} } @Article{IPB-1284, author = {Abel, S. and Bürstenbinder, K. and Müller, J. and}, title = {{The emerging function of IQD proteins as scaffolds in cellular signaling and trafficking}}, year = {2013}, pages = {e24369}, journal = {Plant Signal Behav.}, doi = {10.4161/psb.24369}, volume = {8}, abstract = {Calcium (Ca2+) signaling modules are essential for adjusting plant growth and performance to environmental constraints. Differential interactions between sensors of Ca2+ dynamics and their molecular targets are at the center of the transduction process. Calmodulin (CaM) and CaM-like (CML) proteins are principal Ca2+-sensors in plants that govern the activities of numerous downstream proteins with regulatory properties. The families of IQ67-Domain (IQD) proteins are a large class of plant-specific CaM/CML-targets (e.g., 33 members in A. thaliana) which share a unique domain of multiple varied CaM retention motifs in tandem orientation. Genetic studies in Arabidopsis and tomato revealed first roles for IQD proteins related to basal defense response and plant development. Molecular, biochemical and histochemical analysis of Arabidopsis IQD1 demonstrated association with microtubules as well as targeting to the cell nucleus and nucleolus. In vivo binding to CaM and kinesin light chain-related protein-1 (KLCR1) suggests a Ca2+-regulated scaffolding function of IQD1 in kinesin motor-dependent transport of multiprotein complexes. Furthermore, because IQD1 interacts in vitro with single-stranded nucleic acids, the prospect arises that IQD1 and other IQD family members facilitate cellular RNA localization as one mechanism to control and fine-tune gene expression and protein sorting.} } @Article{IPB-1295, author = {Bürstenbinder, K. and Savchenko, T. and Müller, J. and Adamson, A. W. and Stamm, G. and Kwong, R. and Zipp, B. J. and Dinesh, D. C. and Abel, S. and}, title = {{Arabidopsis Calmodulin-binding Protein IQ67-Domain 1 Localizes to Microtubules and Interacts with Kinesin Light Chain-related Protein-1}}, year = {2013}, pages = {1871-1882}, journal = {J. Biol. Chem.}, doi = {10.1074/jbc.M112.396200}, volume = {288}, abstract = {Calcium (Ca2\+) is a key second messenger in eukaryotes and regulates diverse cellular processes, most notably via calmodulin (CaM). In Arabidopsis thaliana, IQD1 (IQ67 domain 1) is the founding member of the IQD family of putative CaM targets. The 33 predicted IQD proteins share a conserved domain of 67 amino acids that is characterized by a unique arrangement of multiple CaM recruitment motifs, including so-called IQ motifs. Whereas IQD1 has been implicated in the regulation of defense metabolism, the biochemical functions of IQD proteins remain to be elucidated. In this study we show that IQD1 binds to multiple Arabidopsis CaM and CaM-like (CML) proteins in vitro and in yeast two-hybrid interaction assays. CaM overlay assays revealed moderate affinity of IQD1 to CaM2 (Kd ∼ 0.6 μm). Deletion mapping of IQD1 demonstrated the importance of the IQ67 domain for CaM2 binding in vitro, which is corroborated by interaction of the shortest IQD member, IQD20, with Arabidopsis CaM/CMLs in yeast. A genetic screen of a cDNA library identified Arabidopsis kinesin light chain-related protein-1 (KLCR1) as an IQD1 interactor. The subcellular localization of GFP-tagged IQD1 proteins to microtubules and the cell nucleus in transiently and stably transformed plant tissues (tobacco leaves and Arabidopsis seedlings) suggests direct interaction of IQD1 and KLCR1 in planta that is supported by GFP∼IQD1-dependent recruitment of RFP∼KLCR1 and RFP∼CaM2 to microtubules. Collectively, the prospect arises that IQD1 and related proteins provide Ca2\+/CaM-regulated scaffolds for facilitating cellular transport of specific cargo along microtubular tracks via kinesin motor proteins.} } @Article{IPB-1454, author = {Wasternack, C. and Goetz, S. and Hellwege, A. and Forner, S. and Strnad, M. and Hause, B. and}, title = {{Another JA/COI1-independent role of OPDA detected in tomato embryo development}}, year = {2012}, pages = {1349-1353}, journal = {Plant Signal Behav.}, doi = {10.4161/psb.21551}, volume = {7}, abstract = {Jasmonates (JAs) are ubiquitously occurring signaling compounds in plants formed in response to biotic and abiotic stress as well as in development. (\+)-7-iso-jasmonoyl isoleucine, the bioactive JA, is involved in most JA-dependent processes mediated by the F-box protein COI1 in a proteasome-dependent manner. However, there is an increasing number of examples, where the precursor of JA biosynthesis, cis-(\+)-12-oxophytodienoic acid (OPDA) is active in a JA/COI1-independent manner. Here, we discuss those OPDA-dependent processes, thereby giving emphasis on tomato embryo development. Recent data on seed coat-generated OPDA and its role in embryo development is discussed based on biochemical and genetic evidences.} } @Article{IPB-1445, author = {Stenzel, I. and Ischebeck, T. and Quint, M. and Heilmann, I. and}, title = {{Variable regions of PI4P 5-kinases direct PtdIns(4,5)P2 toward alternative regulatory functions in tobacco pollen tubes}}, year = {2012}, pages = {114}, journal = {Front. Plant Sci.}, doi = {10.3389/fpls.2011.00114}, volume = {2}, abstract = {The apical plasma membrane of pollen tubes contains different PI4P 5-kinases that all produce phosphatidylinositol-4,5-bisphosphate [PtdIns(4,5)P2] but exert distinct cellular effects. In the present example, overexpression of Arabidopsis AtPIP5K5 or tobacco NtPIP5K6-1 caused growth defects previously attributed to increased pectin secretion. In contrast, overexpression of Arabidopsis AtPIP5K2 caused apical tip swelling implicated in altering actin fine structure in the pollen tube apex. AtPIP5K5, NtPIP5K6-1, and AtPIP5K2 share identical domain structures. Domains required for correct membrane association of the enzymes were identified by systematic deletion of N-terminal domains and subsequent expression of fluorescence-tagged enzyme truncations in tobacco pollen tubes. A variable linker region (Lin) contained in all PI4P 5-kinase isoforms of subfamily B, but not conserved in sequence, was recognized to be necessary for correct subcellular localization of AtPIP5K5, NtPIP5K6-1, and AtPIP5K2. Deletion of N-terminal domains including the Lin domain did not impair catalytic activity of recombinant AtPIP5K5, NtPIP5K6-1, or AtPIP5K2 in vitro; however, the presence of the Lin domain was necessary for in vivo effects on pollen tube growth upon overexpression of truncated enzymes. Overexpression of catalytically inactive variants of AtPIP5K5, NtPIP5K6-1, or AtPIP5K2 did not influence pollen tube growth, indicating that PtdIns(4,5)P2 production rather than structural properties of PI4P 5-kinases was relevant for the manifestation of growth phenotypes. When Lin domains were swapped between NtPIP5K6-1 and AtPIP5K2 and the chimeric enzymes overexpressed in pollen tubes, the chimeras reciprocally gained the capabilities to invoke tip swelling or secretion phenotypes, respectively. The data indicate that the Lin domain directed the enzymes into different regulatory contexts, possibly contributing to channeling of PtdIns(4,5)P2 at the interface of secretion and actin cytoskeleton.} } @Article{IPB-1444, author = {Stenzel, I. and Otto, M. and Delker, C. and Kirmse, N. and Schmidt, D. and Miersch, O. and Hause, B. and Wasternack, C. and}, title = {{ALLENE OXIDE CYCLASE (AOC) gene family members of Arabidopsis thaliana: tissue- and organ-specific promoter activities and in vivo heteromerization}}, year = {2012}, pages = {6125-6138}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/ers261}, volume = {63}, abstract = {Jasmonates are important signals in plant stress responses and plant development. An essential step in the biosynthesis of jasmonic acid (JA) is catalysed by ALLENE OXIDE CYCLASE (AOC) which establishes the naturally occurring enantiomeric structure of jasmonates. In Arabidopsis thaliana, four genes encode four functional AOC polypeptides (AOC1, AOC2, AOC3, and AOC4) raising the question of functional redundancy or diversification. Analysis of transcript accumulation revealed an organ-specific expression pattern, whereas detailed inspection of transgenic lines expressing the GUS reporter gene under the control of individual AOC promoters showed partially redundant promoter activities during development: (i) In fully developed leaves, promoter activities of AOC1, AOC2, and AOC3 appeared throughout all leaf tissue, but AOC4 promoter activity was vascular bundle-specific; (ii) only AOC3 and AOC4 showed promoter activities in roots; and (iii) partially specific promoter activities were found for AOC1 and AOC4 in flower development. In situ hybridization of flower stalks confirmed the GUS activity data. Characterization of single and double AOC loss-of-function mutants further corroborates the hypothesis of functional redundancies among individual AOCs due to a lack of phenotypes indicative of JA deficiency (e.g. male sterility). To elucidate whether redundant AOC expression might contribute to regulation on AOC activity level, protein interaction studies using bimolecular fluorescence complementation (BiFC) were performed and showed that all AOCs can interact among each other. The data suggest a putative regulatory mechanism of temporal and spatial fine-tuning in JA formation by differential expression and via possible heteromerization of the four AOCs.} } @Article{IPB-1434, author = {Reginato, M. and Abdala, G. and Miersch, O. and Ruiz, O. and Moschetti, E. and Luna, V. and}, title = {{Changes in the levels of jasmonates and free polyamines induced by Na2SO4 and NaCl in roots and leaves of the halophyte Prosopis strombulifera}}, year = {2012}, pages = {689-697}, journal = {Biologia}, doi = {10.2478/s11756-012-0052-7}, volume = {67}, abstract = {Prosopis strombulifera, a common legume in high-salinity soils of Argentina, is a useful model for elucidation of salt tolerance mechanisms and specific biochemical pathways in halophytes, since its NaCl tolerance exceeds the limit described for most halophytic plants. We analyzed the effects of the increasing concentration of two main soil salts, Na2SO4 and NaCl, on growth parameters of P. strombulifera, chlorophyll levels, and content of jasmonates (JAs) and polyamines (PAs), which are key molecules involved in stress responses. P. strombulifera showed a halophytic response (growth promotion) to NaCl, but strong growth inhibition by iso-osmotic solutions of Na2SO4. Chlorophyll levels, number of leaves and leaf area were also differentially affected. An important finding was the partial alleviation of SO42− toxicity by treatment with two-salt mixture. JAs are not directly involved in salt tolerance in this species since its levels decrease under all salt treatments. Beneficial effects of Putrescine (Put) accumulation in NaCl treated plants maybe inferred probably associated with the antioxidative defense system. Another novel finding is the accumulation of the uncommon PA cadaverine in roots under high Na2SO4, which may be related to SO42− toxicity.} } @Article{IPB-1430, author = {Quint, M. and Drost, H.-G. and Gabel, A. and Ullrich, K. K. and Bönn, M. and Grosse, I. and}, title = {{A transcriptomic hourglass in plant embryogenesis}}, year = {2012}, pages = {98-101}, journal = {Nature}, doi = {10.1038/nature11394}, volume = {490}, abstract = {Animal and plant development starts with a constituting phase called embryogenesis, which evolved independently in both lineages1. Comparative anatomy of vertebrate development—based on the Meckel-Serrès law2 and von Baer’s laws of embryology3 from the early nineteenth century—shows that embryos from various taxa appear different in early stages, converge to a similar form during mid-embryogenesis, and again diverge in later stages. This morphogenetic series is known as the embryonic ‘hourglass’4,5, and its bottleneck of high conservation in mid-embryogenesis is referred to as the phylotypic stage6. Recent analyses in zebrafish and Drosophila embryos provided convincing molecular support for the hourglass model, because during the phylotypic stage the transcriptome was dominated by ancient genes7 and global gene expression profiles were reported to be most conserved8. Although extensively explored in animals, an embryonic hourglass has not been reported in plants, which represent the second major kingdom in the tree of life that evolved embryogenesis. Here we provide phylotranscriptomic evidence for a molecular embryonic hourglass in Arabidopsis thaliana, using two complementary approaches. This is particularly significant because the possible absence of an hourglass based on morphological features in plants suggests that morphological and molecular patterns might be uncoupled. Together with the reported developmental hourglass patterns in animals, these findings indicate convergent evolution of the molecular hourglass and a conserved logic of embryogenesis across kingdoms.} } @Article{IPB-1412, author = {Janitza, P. and Ullrich, K. K. and Quint, M. and}, title = {{Toward a comprehensive phylogenetic reconstruction of the evolutionary history of mitogen-activated protein kinases in the plant kingdom}}, year = {2012}, pages = {271}, journal = {Front. Plant Sci.}, doi = {10.3389/fpls.2012.00271}, volume = {3}, abstract = {The mitogen-activated protein kinase (MAPK) pathway is a three-tier signaling cascade that transmits cellular information from the plasma membrane to the cytoplasm where it triggers downstream responses. The MAPKs represent the last step in this cascade and are activated when both tyrosine and threonine residues in a conserved TxY motif are phosphorylated by MAPK kinases, which in turn are themselves activated by phosphorylation by MAPK kinase kinases. To understand the molecular evolution of MAPKs in the plant kingdom, we systematically conducted a Hidden-Markov-Model based screen to identify MAPKs in 13 completely sequenced plant genomes. In this analysis, we included green algae, bryophytes, lycophytes, and several mono- and eudicotyledonous species covering \>800 million years of evolution. The phylogenetic relationships of the 204 identified MAPKs based on Bayesian inference facilitated the retraction of the sequence of emergence of the four major clades that are characterized by the presence of a TDY or TEY-A/TEY-B/TEY-C type kinase activation loop. We present evidence that after the split of TDY- and TEY-type MAPKs, initially the TEY-C clade emerged. This was followed by the TEY-B clade in early land plants until the TEY-A clade finally emerged in flowering plants. In addition to these well characterized clades, we identified another highly conserved clade of 45 MAPK-likes, members of which were previously described as Mak-homologous kinases. In agreement with their essential functions, molecular population genetic analysis of MAPK genes in Arabidopsis thaliana accessions reveal that purifying selection drove the evolution of the MAPK family, implying strong functional constraints on MAPK genes. Closely related MAPKs most likely subfunctionalized, a process in which differential transcriptional regulation of duplicates may be involved.} } @Article{IPB-1407, author = {Goetz, S. and Hellwege, A. and Stenzel, I. and Kutter, C. and Hauptmann, V. and Forner, S. and McCaig, B. and Hause, G. and Miersch, O. and Wasternack, C. and Hause, B. and}, title = {{Role of cis-12-Oxo-Phytodienoic Acid in Tomato Embryo Development}}, year = {2012}, pages = {1715-1727}, journal = {Plant Physiol.}, doi = {10.1104/pp.111.192658}, volume = {158}, abstract = {Oxylipins including jasmonates are signaling compounds in plant growth, development, and responses to biotic and abiotic stresses. In Arabidopsis (Arabidopsis thaliana) most mutants affected in jasmonic acid (JA) biosynthesis and signaling are male sterile, whereas the JA-insensitive tomato (Solanum lycopersicum) mutant jai1 is female sterile. The diminished seed formation in jai1 together with the ovule-specific accumulation of the JA biosynthesis enzyme allene oxide cyclase (AOC), which correlates with elevated levels of JAs, suggest a role of oxylipins in tomato flower/seed development. Here, we show that 35S::SlAOC-RNAi lines with strongly reduced AOC in ovules exhibited reduced seed set similarly to the jai1 plants. Investigation of embryo development of wild-type tomato plants showed preferential occurrence of AOC promoter activity and AOC protein accumulation in the developing seed coat and the embryo, whereas 12-oxo-phytodienoic acid (OPDA) was the dominant oxylipin occurring nearly exclusively in the seed coat tissues. The OPDA- and JA-deficient mutant spr2 was delayed in embryo development and showed an increased programmed cell death in the developing seed coat and endosperm. In contrast, the mutant acx1a, which accumulates preferentially OPDA and residual amount of JA, developed embryos similar to the wild type, suggesting a role of OPDA in embryo development. Activity of the residual amount of JA in the acx1a mutant is highly improbable since the known reproductive phenotype of the JA-insensitive mutant jai1 could be rescued by wound-induced formation of OPDA. These data suggest a role of OPDA or an OPDA-related compound for proper embryo development possibly by regulating carbohydrate supply and detoxification.} } @Article{IPB-1404, author = {Gasperini, D. and Greenland, A. and Hedden, P. and Dreos, R. and Harwood, W. and Griffiths, S. and}, title = {{Genetic and physiological analysis of Rht8 in bread wheat: an alternative source of semi-dwarfism with a reduced sensitivity to brassinosteroids}}, year = {2012}, pages = {4419-4436}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/ers138}, volume = {63}, abstract = {Over the next decade, wheat grain production must increase to meet the demand of a fast growing human population. One strategy to meet this challenge is to raise wheat productivity by optimizing plant stature. The Reduced height 8 (Rht8) semi-dwarfing gene is one of the few, together with the Green Revolution genes, to reduce stature of wheat (Triticum aestivum L.), and improve lodging resistance, without compromising grain yield. Rht8 is widely used in dry environments such as Mediterranean countries where it increases plant adaptability. With recent climate change, its use could become increasingly important even in more northern latitudes. In the present study, the characterization of Rht8 was furthered. Morphological analyses show that the semi-dwarf phenotype of Rht8 lines is due to shorter internodal segments along the wheat culm, achieved through reduced cell elongation. Physiological experiments show that the reduced cell elongation is not due to defective gibberellin biosynthesis or signalling, but possibly to a reduced sensitivity to brassinosteroids. Using a fine-resolution mapping approach and screening 3104 F2 individuals of a newly developed mapping population, the Rht8 genetic interval was reduced from 20.5 cM to 1.29 cM. Comparative genomics with model genomes confined the Rht8 syntenic intervals to 3.3 Mb of the short arm of rice chromosome 4, and to 2 Mb of Brachypodium distachyon chromosome 5. The very high resolution potential of the plant material generated is crucial for the eventual cloning of Rht8.} } @Article{IPB-1400, author = {Fellenberg, C. and Ziegler, J. and Handrick, V. and Vogt, T. and}, title = {{Polyamine homeostasis in wild type and phenolamide deficient Arabidopsis thaliana stamens}}, year = {2012}, pages = {180}, journal = {Front. Plant Sci.}, doi = {10.3389/fpls.2012.00180}, volume = {3}, abstract = {Polyamines (PAs) like putrescine, spermidine, and spermine are ubiquitous polycationic molecules that occur in all living cells and have a role in a wide variety of biological processes. High amounts of spermidine conjugated to hydroxycinnamic acids are detected in the tryphine of Arabidopsis thaliana pollen grains. Tapetum localized spermidine hydroxycinnamic acid transferase (SHT) is essential for the biosynthesis of these anther specific tris-conjugated spermidine derivatives. Sht knockout lines show a strong reduction of hydroxycinnamic acid amides (HCAAs). The effect of HCAA-deficient anthers on the level of free PAs was measured by a new sensitive and reproducible method using 9-fluorenylmethyl chloroformate (FMOC) and fluorescence detection by HPLC. PA concentrations can be accurately determined even when very limited amounts of plant material, as in the case of A. thaliana stamens, are available. Analysis of free PAs in wild type stamens compared to sht deficient mutants and transcript levels of key PA biosynthetic genes revealed a highly controlled regulation of PA homeostasis in A. thaliana anthers.} } @Article{IPB-1389, author = {Den Herder, G. and Yoshida, S. and Antolín-Llovera, M. and Ried, M. K. and Parniske, M. and}, title = {{Lotus japonicus E3 Ligase SEVEN IN ABSENTIA4 Destabilizes the Symbiosis Receptor-Like Kinase SYMRK and Negatively Regulates Rhizobial Infection}}, year = {2012}, pages = {1691-1707}, journal = {Plant Cell}, doi = {10.1105/tpc.110.082248}, volume = {24}, abstract = {The Lotus japonicus SYMBIOSIS RECEPTOR-LIKE KINASE (SYMRK) is required for symbiotic signal transduction upon stimulation of root cells by microbial signaling molecules. Here, we identified members of the SEVEN IN ABSENTIA (SINA) E3 ubiquitin-ligase family as SYMRK interactors and confirmed their predicted ubiquitin-ligase activity. In Nicotiana benthamiana leaves, SYMRK–yellow fluorescent protein was localized at the plasma membrane, and interaction with SINAs, as determined by bimolecular fluorescence complementation, was observed in small punctae at the cytosolic interface of the plasma membrane. Moreover, fluorescence-tagged SINA4 partially colocalized with SYMRK and caused SYMRK relocalization as well as disappearance of SYMRK from the plasma membrane. Neither the localization nor the abundance of Nod-factor receptor1 was altered by the presence of SINA4. SINA4 was transcriptionally upregulated during root symbiosis, and rhizobia inoculated roots ectopically expressing SINA4 showed reduced SYMRK protein levels. In accordance with a negative regulatory role in symbiosis, infection thread development was impaired upon ectopic expression of SINA4. Our results implicate SINA4 E3 ubiquitin ligase in the turnover of SYMRK and provide a conceptual mechanism for its symbiosis-appropriate spatio-temporal containment.} } @Article{IPB-1385, author = {Calderón Villalobos, L. I. A. and Lee, S. and De Oliveira, C. and Ivetac, A. and Brandt, W. and Armitage, L. and Sheard, L. B. and Tan, X. and Parry, G. and Mao, H. and Zheng, N. and Napier, R. and Kepinski, S. and Estelle, M. and}, title = {{A combinatorial TIR1/AFB–Aux/IAA co-receptor system for differential sensing of auxin}}, year = {2012}, pages = {477-485}, journal = {Nat. Chem. Biol.}, doi = {10.1038/nchembio.926}, volume = {8}, abstract = {The plant hormone auxin regulates virtually every aspect of plant growth and development. Auxin acts by binding the F-box protein transport inhibitor response 1 (TIR1) and promotes the degradation of the AUXIN/INDOLE-3-ACETIC ACID (Aux/IAA) transcriptional repressors. Here we show that efficient auxin binding requires assembly of an auxin co-receptor complex consisting of TIR1 and an Aux/IAA protein. Heterologous experiments in yeast and quantitative IAA binding assays using purified proteins showed that different combinations of TIR1 and Aux/IAA proteins form co-receptor complexes with a wide range of auxin-binding affinities. Auxin affinity seems to be largely determined by the Aux/IAA. As there are 6 TIR1/AUXIN SIGNALING F-BOX proteins (AFBs) and 29 Aux/IAA proteins in Arabidopsis thaliana, combinatorial interactions may result in many co-receptors with distinct auxin-sensing properties. We also demonstrate that the AFB5–Aux/IAA co-receptor selectively binds the auxinic herbicide picloram. This co-receptor system broadens the effective concentration range of the hormone and may contribute to the complexity of auxin response.} } @Article{IPB-1382, author = {Brandt, R. and Salla-Martret, M. and Bou-Torrent, J. and Musielak, T. and Stahl, M. and Lanz, C. and Ott, F. and Schmid, M. and Greb, T. and Schwarz, M. and Choi, S.-B. and Barton, M. K. and Reinhart, B. J. and Liu, T. and Quint, M. and Palauqui, J.-C. and Martínez-García, J. F. and Wenkel, S. and}, title = {{Genome-wide binding-site analysis of REVOLUTA reveals a link between leaf patterning and light-mediated growth responses}}, year = {2012}, pages = {31-42}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2012.05049.x}, volume = {72}, abstract = {Unlike the situation in animals, the final morphology of the plant body is highly modulated by the environment. During Arabidopsis development, intrinsic factors provide the framework for basic patterning processes. CLASS III HOMEODOMAIN LEUCINE ZIPPER (HD‐ZIPIII) transcription factors are involved in embryo, shoot and root patterning. During vegetative growth HD‐ZIPIII proteins control several polarity set‐up processes such as in leaves and the vascular system. We have identified several direct target genes of the HD‐ZIPIII transcription factor REVOLUTA (REV) using a chromatin immunoprecipitation/DNA sequencing (ChIP‐Seq) approach. This analysis revealed that REV acts upstream of auxin biosynthesis and affects directly the expression of several class II HD‐ZIP transcription factors that have been shown to act in the shade‐avoidance response pathway. We show that, as well as involvement in basic patterning, HD‐ZIPIII transcription factors have a critical role in the control of the elongation growth that is induced when plants experience shade. Leaf polarity is established by the opposed actions of HD‐ZIPIII and KANADI transcription factors. Finally, our study reveals that the module that consists of HD‐ZIPIII/KANADI transcription factors controls shade growth antagonistically and that this antagonism is manifested in the opposed regulation of shared target genes.} } @Article{IPB-1373, author = {Antolín-Llovera, M. and Ried, M. K. and Binder, A. and Parniske, M. and}, title = {{Receptor Kinase Signaling Pathways in Plant-Microbe Interactions}}, year = {2012}, pages = {451-473}, journal = {Annu. Rev. Phytopathol.}, doi = {10.1146/annurev-phyto-081211-173002}, volume = {50}, abstract = {Plant receptor-like kinases (RLKs) function in diverse signaling pathways, including the responses to microbial signals in symbiosis and defense. This versatility is achieved with a common overall structure: an extracytoplasmic domain (ectodomain) and an intracellular protein kinase domain involved in downstream signal transduction. Various surfaces of the leucine-rich repeat (LRR) ectodomain superstructure are utilized for interaction with the cognate ligand in both plant and animal receptors. RLKs with lysin-motif (LysM) ectodomains confer recognitional specificity toward N-acetylglucosamine-containing signaling molecules, such as chitin, peptidoglycan (PGN), and rhizobial nodulation factor (NF), that induce immune or symbiotic responses. Signaling downstream of RLKs does not follow a single pattern; instead, the detailed analysis of brassinosteroid (BR) signaling, innate immunity, and symbiosis revealed at least three largely nonoverlapping pathways. In this review, we focus on RLKs involved in plant-microbe interactions and contrast the signaling pathways leading to symbiosis and defense.} } @Article{IPB-1484, author = {Flores, R. and Grubb, D. and Elleuch, A. and Nohales, M.-?. and Delgado, S. and Gago, S. and}, title = {{Rolling-circle replication of viroids, viroid-like satellite RNAs and hepatitis delta virus: Variations on a theme}}, year = {2011}, pages = {200-206}, journal = {RNA Biol.}, doi = {10.4161/rna.8.2.14238}, volume = {8}, abstract = {Viroids and viroid-like satellite RNAs from plants, and the human hepatitis delta virus (HDV) RNA share some properties that include small size, circularity and replication through a rolling-circle mechanism. Replication occurs in different cell compartments (nucleus, chloroplast and membrane-associated cytoplasmatic vesicles) and has three steps: RNA polymerization, cleavage and ligation. The first step generates oligomeric RNAs that result from the reiterative transcription of the circular templates of one or both polarities, and is catalyzed by either the RNA-dependent RNA polymerase of the helper virus on which viroid-like satellite RNAs are functionally dependent, or by host DNA-dependent RNA polymerases that, remarkably, viroids and HDV redirect to transcribe RNA templates. Cleavage is mediated by host enzymes in certain viroids and viroid-like satellite RNAs, while in others and in HDV is mediated by cis-acting ribozymes of three classes. Ligation appears to be catalyzed mainly by host enzymes. Replication most likely also involves many other non-catalytic proteins of host origin and, in HDV, the single virus-encoded protein.} } @Article{IPB-1480, author = {Delker, C. and Quint, M. and}, title = {{Expression level polymorphisms: heritable traits shaping natural variation}}, year = {2011}, pages = {481-488}, journal = {Trends Plant Sci.}, doi = {10.1016/j.tplants.2011.05.009}, volume = {16}, abstract = {Natural accessions of many species harbor a wealth of genetic variation visible in a large array of phenotypes. Although expression level polymorphisms (ELPs) in several genes have been shown to contribute to variation in diverse traits, their general impact on adaptive variation has likely been underestimated. At present, ELPs have predominantly been correlated to quantitative trait loci (eQTLs) that occupy central hubs in signaling networks, which pleiotropically affect numerous traits. To increase the sensitivity of detecting minor effect eQTLs or those that act in a trait-specific manner, we emphasize the need for more systematic approaches. This requires, but is not limited to, refining experimental designs such as reduction of tissue complexity and combinatorial methods including a priori defined networks.} } @Article{IPB-1478, author = {Costa, C. T. and Strieder, M. L. and Abel, S. and Delatorre, C. A. and}, title = {{Phosphorus and nitrogen interaction: loss of QC identity in response to P or N limitation is antecipated in pdr23 mutant}}, year = {2011}, pages = {219-229}, journal = {Braz. J. Plant Physiol.}, doi = {10.1590/S1677-04202011000300006}, volume = {23}, abstract = {Changes in root architecture are an important adaptive strategy used by plants in response to limited nutrient availability to increase the odds of acquiring them. The quiescent center (QC) plays an important role by altering the meristem activity causing differentiation and therefore, inducing a determinate growth program. The arabidopsis mutant pdr23 presents primary short root in the presence of nitrate and is inefficient in the use of nucleic acids as a source of phosphorus. In this study the effect of the pdr23 mutation on the QC maintenance under low phosphorus (P) and/or nitrogen is evaluated. QC identity is maintained in wild-type in the absence of nitrate and/or phosphate if nucleic acids can be used as an alternative source of these nutrients, but not in pdr23. The mutant is not able to use nucleic acids efficiently for substitute Pi, determinate growth is observed, similar to wild-type in the total absence of P. In the absence of N pdr23 loses the expression of QC identity marker earlier than wild-type, indicating that not only the response to P is altered, but also to N. The data suggest that the mutation affects a gene involved either in the crosstalk between these nutrients or in a pathway shared by both nutrients limitation response. Moreover loss of QC identity is also observed in wild-type in the absence of N at longer limitation. Less drastic symptoms are observed in lateral roots of both genotypes.} } @Article{IPB-1476, author = {Carbonell, A. and Flores, R. and Gago, S. and}, title = {{Trans-cleaving hammerhead ribozymes with tertiary stabilizing motifs: in vitro and in vivo activity against a structured viroid RNA}}, year = {2011}, pages = {2432-2444}, journal = {Nucleic Acids Res.}, doi = {10.1093/nar/gkq1051}, volume = {39}, abstract = {Trans -cleaving hammerheads with discontinuous or extended stem I and with tertiary stabilizing motifs (TSMs) have been tested previously against short RNA substrates in vitro at low Mg 2\+ concentration. However, the potential of these ribozymes for targeting longer and structured RNAs in vitro and in vivo has not been examined. Here, we report the in vitro cleavage of short RNAs and of a 464-nt highly structured RNA from potato spindle tuber viroid (PSTVd) by hammerheads with discontinuous and extended formats at submillimolar Mg 2\+ . Under these conditions, hammerheads derived from eggplant latent viroid and peach latent mosaic viroid (PLMVd) with discontinuous and extended formats, respectively, where the most active. Furthermore, a PLMVd-derived hammerhead with natural TSMs showed activity in vivo against the same long substrate and interfered with systemic PSTVd infection, thus reinforcing the idea that this class of ribozymes has potential to control pathogenic RNA replicons.} } @Article{IPB-1468, author = {Asquini, E. and Gerdol, M. and Gasperini, D. and Igic, B. and Graziosi, G. and Pallavicini, A. and}, title = {{S-RNase-like Sequences in Styles of Coffea (Rubiaceae). Evidence for S-RNase Based Gametophytic Self-Incompatibility?}}, year = {2011}, pages = {237-249}, journal = {Trop. Plant Biol.}, doi = {10.1007/s12042-011-9085-2}, volume = {4}, abstract = {Although RNase-based self-incompatibility (SI) is suspected to operate in a wide group of plant families, it has been characterized as the molecular genetic basis of SI in only three distantly related families, Solanaceae, Plantaginaceae, and Rosaceae, all described over a decade ago. Previous studies found that gametophytic SI, controlled by a multi-allelic S-locus, operates in the coffee family (Rubiaceae). The molecular genetic basis of this mechanism remains unknown, despite the immense importance of coffee as an agricultural commodity. Here, we isolated ten sequences with features of T2-S-type RNases from two Coffea species. While three of the sequences were identified in both species and clearly do not appear to be S-locus products, our data suggest that six sequences may be S-alleles in the self-incompatible C. canephora, and one may be a relict in the self-compatible C. arabica. We demonstrate that these sequences show style-specific expression, display polymorphism in C. canephora, and cluster with S-locus products in a phylogenetic analysis that includes other plant families with RNase-based SI. Although our results are not definitive, in part because the available plant materials were limited and data patterns relatively complex, our results strongly hint that RNase-based SI mechanism operates in the Rubiaceae family.} } @Article{IPB-1467, author = {Abel, S. and}, title = {{Phosphate sensing in root development}}, year = {2011}, pages = {303-309}, journal = {Curr. Opin. Plant Biol.}, doi = {10.1016/j.pbi.2011.04.007}, volume = {14}, abstract = {Phosphate (Pi) and its anhydrides constitute major nodes in metabolism. Thus, plant performance depends directly on Pi nutrition. Inadequate Pi availability in the rhizosphere is a common challenge to plants, which activate metabolic and developmental responses to maximize Pi usage and acquisition. The sensory mechanisms that monitor environmental Pi and transmit the nutritional signal to adjust root development have increasingly come into focus. Recent transcriptomic analyses and genetic approaches have highlighted complex antagonistic interactions between external Pi and Fe bioavailability and have implicated the stem cell niche as a target of Pi sensing to regulate root meristem activity.} } @Article{IPB-1519, author = {Terrile, M. C. and París, R. and Calderón-Villalobos, L. I. A. and Iglesias, M. J. and Lamattina, L. and Estelle, M. and Casalongué, C. A. and}, title = {{Nitric oxide influences auxin signaling through S-nitrosylation of the Arabidopsis TRANSPORT INHIBITOR RESPONSE 1 auxin receptor}}, year = {2011}, pages = {492-500}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2011.04885.x}, volume = {70}, abstract = {Previous studies have demonstrated that auxin (indole‐3‐acetic acid) and nitric oxide (NO) are plant growth regulators that coordinate several plant physiological responses determining root architecture. Nonetheless, the way in which these factors interact to affect these growth and developmental processes is not well understood. The Arabidopsis thaliana F‐box proteins TRANSPORT INHIBITOR RESPONSE 1/AUXIN SIGNALING F‐BOX (TIR1/AFB) are auxin receptors that mediate degradation of AUXIN/INDOLE‐3‐ACETIC ACID (Aux/IAA) repressors to induce auxin‐regulated responses. A broad spectrum of NO‐mediated protein modifications are known in eukaryotic cells. Here, we provide evidence that NO donors increase auxin‐dependent gene expression while NO depletion blocks Aux/IAA protein degradation. NO also enhances TIR1‐Aux/IAA interaction as evidenced by pull‐down and two‐hybrid assays. In addition, we provide evidence for NO‐mediated modulation of auxin signaling through S‐nitrosylation of the TIR1 auxin receptor. S‐nitrosylation of cysteine is a redox‐based post‐translational modification that contributes to the complexity of the cellular proteome. We show that TIR1 C140 is a critical residue for TIR1–Aux/IAA interaction and TIR1 function. These results suggest that TIR1 S‐nitrosylation enhances TIR1–Aux/IAA interaction, facilitating Aux/IAA degradation and subsequently promoting activation of gene expression. Our findings underline the importance of NO in phytohormone signaling pathways.} } @Article{IPB-1517, author = {Schumann, N. and Navarro-Quezada, A. and Ullrich, K. and Kuhl, C. and Quint, M. and}, title = {{Molecular Evolution and Selection Patterns of Plant F-Box Proteins with C-Terminal Kelch Repeats}}, year = {2011}, pages = {835-850}, journal = {Plant Physiol.}, doi = {10.1104/pp.110.166579}, volume = {155}, abstract = {The F-box protein superfamily represents one of the largest families in the plant kingdom. F-box proteins phylogenetically organize into numerous subfamilies characterized by their carboxyl (C)-terminal protein-protein interaction domain. Among the largest F-box protein subfamilies in plant genomes are those with C-terminal kelch repeats. In this study, we analyzed the phylogeny and evolution of F-box kelch proteins/genes (FBKs) in seven completely sequenced land plant genomes including a bryophyte, a lycophyte, monocots, and eudicots. While absent in prokaryotes, F-box kelch proteins are widespread in eukaryotes. Nonplant eukaryotes usually contain only a single FBK gene. In land plant genomes, however, FBKs expanded dramatically. Arabidopsis thaliana, for example, contains at least 103 F-box genes with well-conserved C-terminal kelch repeats. The construction of a phylogenetic tree based on the full-length amino acid sequences of the FBKs that we identified in the seven species enabled us to classify FBK genes into unstable/stable/superstable categories. In contrast to superstable genes, which are conserved across all seven species, kelch domains of unstable genes, which are defined as lineage specific, showed strong signatures of positive selection, indicating adaptational potential. We found evidence for conserved protein features such as binding affinities toward A. thaliana SKP1-like adaptor proteins and subcellular localization among closely related FBKs. Pseudogenization seems to occur only rarely, but differential transcriptional regulation of close relatives may result in subfunctionalization.} } @Article{IPB-1499, author = {Kopycki, J. and Schmidt, J. and Abel, S. and Grubb, C. D. and}, title = {{Chemoenzymatic synthesis of diverse thiohydroximates from glucosinolate-utilizing enzymes from Helix pomatia and Caldicellulosiruptor saccharolyticus}}, year = {2011}, pages = {1039-1046}, journal = {Biotechnol. Lett.}, doi = {10.1007/s10529-011-0530-y}, volume = {33}, abstract = {Thiohydroximates comprise a diverse class of compounds important in both biological and industrial chemistry. Their syntheses are generally limited to simple alkyl and aryl compounds with few stereocenters and a narrow range of functional groups. We hypothesized that sequential action of two recombinant enzymes, a sulfatase from Helix pomatia and a β-O-glucosidase from Caldicellulosiruptor saccharolyticus, on glucosinolates would allow synthesis of thiohydroximates from a structurally broad array of abundant precursors. We report successful synthesis of thiohydroximates of varied chemical classes, including from homochiral compounds of demonstrated biological activity. The chemoenzymatic synthetic route reported here should allow access to many, if not all, of the thiohydroximate core structures of the ~200 known naturally occurring glucosinolates. The enrichment of this group for compounds with possible pharmacological potential is discussed.} } @Article{IPB-1567, author = {Ludwig-Müller, J. and Denk, K. and Cohen, J. D. and Quint, M. and}, title = {{An Inhibitor of Tryptophan-Dependent Biosynthesis of Indole-3-Acetic Acid Alters Seedling Development in Arabidopsis}}, year = {2010}, pages = {242-248}, journal = {J. Plant Growth Regul.}, doi = {10.1007/s00344-009-9128-1}, volume = {29}, abstract = {Although polar transport and the TIR1-dependent signaling pathway of the plant hormone auxin/indole-3-acetic acid (IAA) are well characterized, understanding of the biosynthetic pathway(s) leading to the production of IAA is still limited. Genetic dissection of IAA biosynthetic pathways has been complicated by the metabolic redundancy caused by the apparent existence of several parallel biosynthetic routes leading to IAA production. Valuable complementary tools for genetic as well as biochemical analysis of auxin biosynthesis would be molecular inhibitors capable of acting in vivo on specific or general components of the pathway(s), which unfortunately have been lacking. Several indole derivatives have been previously identified to inhibit tryptophan-dependent IAA biosynthesis in an in vitro system from maize endosperm. We examined the effect of one of them, 6-fluoroindole, on seedling development of Arabidopsis thaliana and tested its ability to inhibit IAA biosynthesis in feeding experiments in vivo. We demonstrated a correlation of severe developmental defects or growth retardation caused by 6-fluoroindole with significant downregulation of de novo synthesized IAA levels, derived from the stable isotope-labeled tryptophan pool, upon treatment. Hence, 6-fluoroindole shows important features of an inhibitor of tryptophan-dependent IAA biosynthesis both in vitro and in vivo and thus may find use as a promising molecular tool for the identification of novel components of the auxin biosynthetic pathway(s).} } @Article{IPB-1566, author = {Leon-Reyes, A. and Van der Does, D. and De Lange, E. S. and Delker, C. and Wasternack, C. and Van Wees, S. C. M. and Ritsema, T. and Pieterse, C. M. J. and}, title = {{Salicylate-mediated suppression of jasmonate-responsive gene expression in Arabidopsis is targeted downstream of the jasmonate biosynthesis pathway}}, year = {2010}, pages = {1423-1432}, journal = {Planta}, doi = {10.1007/s00425-010-1265-z}, volume = {232}, abstract = {Jasmonates (JAs) and salicylic acid (SA) are plant hormones that play pivotal roles in the regulation of induced defenses against microbial pathogens and insect herbivores. Their signaling pathways cross-communicate providing the plant with a regulatory potential to finely tune its defense response to the attacker(s) encountered. In Arabidopsis thaliana, SA strongly antagonizes the jasmonic acid (JA) signaling pathway, resulting in the downregulation of a large set of JA-responsive genes, including the marker genes PDF1.2 and VSP2. Induction of JA-responsive marker gene expression by different JA derivatives was equally sensitive to SA-mediated suppression. Activation of genes encoding key enzymes in the JA biosynthesis pathway, such as LOX2, AOS, AOC2, and OPR3 was also repressed by SA, suggesting that the JA biosynthesis pathway may be a target for SA-mediated antagonism. To test this, we made use of the mutant aos/dde2, which is completely blocked in its ability to produce JAs because of a mutation in the ALLENE OXIDE SYNTHASE gene. Mutant aos/dde2 plants did not express the JA-responsive marker genes PDF1.2 or VSP2 in response to infection with the necrotrophic fungus Alternaria brassicicola or the herbivorous insect Pieris rapae. Bypassing JA biosynthesis by exogenous application of methyl jasmonate (MeJA) rescued this JA-responsive phenotype in aos/dde2. Application of SA suppressed MeJA-induced PDF1.2 expression to the same level in the aos/dde2 mutant as in wild-type Col-0 plants, indicating that SA-mediated suppression of JA-responsive gene expression is targeted at a position downstream of the JA biosynthesis pathway.} } @Article{IPB-1544, author = {Delker, C. and Pöschl, Y. and Raschke, A. and Ullrich, K. and Ettingshausen, S. and Hauptmann, V. and Grosse, I. and Quint, M. and}, title = {{Natural Variation of Transcriptional Auxin Response Networks in Arabidopsis thaliana}}, year = {2010}, pages = {2184-2200}, journal = {Plant Cell}, doi = {10.1105/tpc.110.073957}, volume = {22}, abstract = {Natural variation has been observed for various traits in Arabidopsis thaliana. Here, we investigated natural variation in the context of physiological and transcriptional responses to the phytohormone auxin, a key regulator of plant development. A survey of the general extent of natural variation to auxin stimuli revealed significant physiological variation among 20 genetically diverse natural accessions. Moreover, we observed dramatic variation on the global transcriptome level after induction of auxin responses in seven accessions. Although we detect isolated cases of major-effect polymorphisms, sequencing of signaling genes revealed sequence conservation, making selective pressures that favor functionally different protein variants among accessions unlikely. However, coexpression analyses of a priori defined auxin signaling networks identified variations in the transcriptional equilibrium of signaling components. In agreement with this, cluster analyses of genome-wide expression profiles followed by analyses of a posteriori defined gene networks revealed accession-specific auxin responses. We hypothesize that quantitative distortions in the ratios of interacting signaling components contribute to the detected transcriptional variation, resulting in physiological variation of auxin responses among accessions.} } @Article{IPB-1541, author = {Calderon-Villalobos, L. I. and Tan, X. and Zheng, N. and Estelle, M. and}, title = {{Auxin Perception—Structural Insights}}, year = {2010}, pages = {a005546}, journal = {Cold Spring Harb. Perspect. Biol.}, doi = {10.1101/cshperspect.a005546}, volume = {2}, abstract = {The identity of the auxin receptor(s) and the mechanism of auxin perception has been a subject of intense interest since the discovery of auxin almost a century ago. The development of genetic approaches to the study of plant hormone signaling led to the discovery that auxin acts by promoting degradation of transcriptional repressors called Aux/IAA proteins. This process requires a ubiquitin protein ligase (E3) called SCFTIR1 and related SCF complexes. Surprisingly, auxin works by directly binding to TIR1, the F-box protein subunit of this SCF. Structural studies demonstrate that auxin acts like a “molecular glue,” to stabilize the interaction between TIR1 and the Aux/IAA substrate. These exciting results solve an old problem in plant biology and reveal new mechanisms for E3 regulation and hormone perception.} } @Article{IPB-1594, author = {Wasternack, C. and Xie, D. and}, title = {{The genuine ligand of a jasmonic acid receptor: Improved analysis of jasmonates is now required}}, year = {2010}, pages = {337-340}, journal = {Plant Signal Behav.}, doi = {10.4161/psb.5.4.11574}, volume = {5}, abstract = {Jasmonic acid (JA), its metabolites, such as the methyl ester or amino acid conjugates as well as its precursor 12-oxophytodienoic acid (OPDA) are lipid-derived signals. JA, OPDA and JA-amino acid conjugates are known to function as signals in plant stress responses and development. More recently, formation of JA-amino acid conjugates and high biological activity of JA-Isoleucine (JA-Ile) were found to be essential in JA signalling. A breakthrough was the identification of JAZ proteins which interact with the F-box protein COI1 if JA-Ile is bound. This interaction leads to proteasomal degradation of JAZs being negative regulators of JA-induced transcription. Surprisingly, a distinct stereoisomer of JA-Ile, the (\+)-7-iso-JA-Ile ((3R,7S) form) is most active. Coronatine, a bacterial phytotoxine with an identical stereochemistry at the cyclopentanone ring, has a similar bioactivity . This was explained by the recent identification of COI1 as the JA receptor and accords well with molecular modelling studies. Whereas over the last two decades JA was quantified to describe any JA dependent process, now we have to take into account a distinct stereoisomer of JA-Ile. Until recently a quantitative analysis of (\+)-7-iso-JA-Ile was missing presumable due to its equilibration to (-)-JA-Ile. Now such an analysis was achieved. These aspects will be discussed based on our new knowledge on JA perception and signalling.} } @Article{IPB-1593, author = {Wasternack, C. and Kombrink, E. and}, title = {{Jasmonates: Structural Requirements for Lipid-Derived Signals Active in Plant Stress Responses and Development}}, year = {2010}, pages = {63-77}, journal = {ACS Chem. Biol.}, doi = {10.1021/cb900269u}, volume = {5}, abstract = {Jasmonates are lipid-derived signals that mediate plant stress responses and development processes. Enzymes participating in biosynthesis of jasmonic acid (JA) (1, 2) and components of JA signaling have been extensively characterized by biochemical and molecular-genetic tools. Mutants of Arabidopsis and tomato have helped to define the pathway for synthesis of jasmonoyl-isoleucine (JA-Ile), the active form of JA, and to identify the F-box protein COI1 as central regulatory unit. However, details of the molecular mechanism of JA signaling have only recently been unraveled by the discovery of JAZ proteins that function in transcriptional repression. The emerging picture of JA perception and signaling cascade implies the SCFCOI1 complex operating as E3 ubiquitin ligase that upon binding of JA-Ile targets JAZ repressors for degradation by the 26S-proteasome pathway, thereby allowing the transcription factor MYC2 to activate gene expression. The fact that only one particular stereoisomer, (\+)-7-iso-JA-l-Ile (4), shows high biological activity suggests that epimerization between active and inactive diastereomers could be a mechanism for turning JA signaling on or off. The recent demonstration that COI1 directly binds (\+)-7-iso-JA-l-Ile (4) and thus functions as JA receptor revealed that formation of the ternary complex COI1-JA-Ile-JAZ is an ordered process. The pronounced differences in biological activity of JA stereoisomers also imply strict stereospecific control of product formation along the JA biosynthetic pathway. The pathway of JA biosynthesis has been unraveled, and most of the participating enzymes are well-characterized. For key enzymes of JA biosynthesis the crystal structures have been established, allowing insight into the mechanisms of catalysis and modes of substrate binding that lead to formation of stereospecific products.} } @Article{IPB-1584, author = {Stumpe, M. and Göbel, C. and Faltin, B. and Beike, A. K. and Hause, B. and Himmelsbach, K. and Bode, J. and Kramell, R. and Wasternack, C. and Frank, W. and Reski, R. and Feussner, I. and}, title = {{The moss Physcomitrella patens contains cyclopentenones but no jasmonates: mutations in allene oxide cyclase lead to reduced fertility and altered sporophyte morphology}}, year = {2010}, pages = {740-749}, journal = {New Phytol.}, doi = {10.1111/j.1469-8137.2010.03406.x}, volume = {188}, abstract = {Two cDNAs encoding allene oxide cyclases (PpAOC1, PpAOC2), key enzymes in the formation of jasmonic acid (JA) and its precursor (9S,13S)‐12‐oxo‐phytodienoic acid (cis‐(\+)‐OPDA), were isolated from the moss Physcomitrella patens.Recombinant PpAOC1 and PpAOC2 show substrate specificity against the allene oxide derived from 13‐hydroperoxy linolenic acid (13‐HPOTE); PpAOC2 also shows substrate specificity against the allene oxide derived from 12‐hydroperoxy arachidonic acid (12‐HPETE).In protonema and gametophores the occurrence of cis‐(\+)‐OPDA, but neither JA nor the isoleucine conjugate of JA nor that of cis‐(\+)‐OPDA was detected.Targeted knockout mutants for PpAOC1 and for PpAOC2 were generated, while double mutants could not be obtained. The ΔPpAOC1 and ΔPpAOC2 mutants showed reduced fertility, aberrant sporophyte morphology and interrupted sporogenesis.} } @Article{IPB-1583, author = {Sreenivasulu, N. and Radchuk, V. and Alawady, A. and Borisjuk, L. and Weier, D. and Staroske, N. and Fuchs, J. and Miersch, O. and Strickert, M. and Usadel, B. and Wobus, U. and Grimm, B. and Weber, H. and Weschke, W. and}, title = {{De-regulation of abscisic acid contents causes abnormal endosperm development in the barley mutant seg8}}, year = {2010}, pages = {589-603}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2010.04350.x}, volume = {64}, abstract = {Grain development of the maternal effect shrunken endosperm mutant seg8 was analysed by comprehensive molecular, biochemical and histological methods. The most obvious finding was de‐regulation of ABA levels, which were lower compared to wild‐type during the pre‐storage phase but higher during the transition from cell division/differentiation to accumulation of storage products. Ploidy levels and ABA amounts were inversely correlated in the developing endosperms of both mutant and wild‐type, suggesting an influence of ABA on cell‐cycle regulation. The low ABA levels found in seg8 grains between anthesis and beginning endosperm cellularization may result from a gene dosage effect in the syncytial endosperm that causes impaired transfer of ABA synthesized in vegetative tissues into filial grain parts. Increased ABA levels during the transition phase are accompanied by higher chlorophyll and carotenoid/xanthophyll contents. The data suggest a disturbed ABA‐releasing biosynthetic pathway. This is indicated by up‐regulation of expression of the geranylgeranyl reductase (GGR) gene, which may be induced by ABA deficiency during the pre‐storage phase. Abnormal cellularization/differentiation of the developing seg8 endosperm and reduced accumulation of starch are phenotypic characteristics that reflect these disturbances. The present study did not reveal the primary gene defect causing the seg8 phenotype, but presents new insights into the maternal/filial relationships regulating barley endosperm development.} } @Article{IPB-1578, author = {Robson, F. and Okamoto, H. and Patrick, E. and Harris, S.-R. and Wasternack, C. and Brearley, C. and Turner, J. G. and}, title = {{Jasmonate and Phytochrome A Signaling in Arabidopsis Wound and Shade Responses Are Integrated through JAZ1 Stability}}, year = {2010}, pages = {1143-1160}, journal = {Plant Cell}, doi = {10.1105/tpc.109.067728}, volume = {22}, abstract = {Jasmonate (JA) activates plant defense, promotes pollen maturation, and suppresses plant growth. An emerging theme in JA biology is its involvement in light responses; here, we examine the interdependence of the JA- and light-signaling pathways in Arabidopsis thaliana. We demonstrate that mutants deficient in JA biosynthesis and signaling are deficient in a subset of high irradiance responses in far-red (FR) light. These mutants display exaggerated shade responses to low, but not high, R/FR ratio light, suggesting a role for JA in phytochrome A (phyA) signaling. Additionally, we demonstrate that the FR light–induced expression of transcription factor genes is dependent on CORONATINE INSENSITIVE1 (COI1), a central component of JA signaling, and is suppressed by JA. phyA mutants had reduced JA-regulated growth inhibition and VSP expression and increased content of cis-(\+)-12-oxophytodienoic acid, an intermediate in JA biosynthesis. Significantly, COI1-mediated degradation of JASMONATE ZIM DOMAIN1-β-glucuronidase (JAZ1-GUS) in response to mechanical wounding and JA treatment required phyA, and ectopic expression of JAZ1-GUS resulted in exaggerated shade responses. Together, these results indicate that JA and phyA signaling are integrated through degradation of the JAZ1 protein, and both are required for plant responses to light and stress.} } @Article{IPB-1577, author = {Renovell, ?. and Gago, S. and Ruiz-Ruiz, S. and Velázquez, K. and Navarro, L. and Moreno, P. and Vives, M. C. and Guerri, J. and}, title = {{Mapping the subgenomic RNA promoter of the Citrus leaf blotch virus coat protein gene by Agrobacterium-mediated inoculation}}, year = {2010}, pages = {360-369}, journal = {Virology}, doi = {10.1016/j.virol.2010.07.034}, volume = {406}, abstract = {Citrus leaf blotch virus has a single-stranded positive-sense genomic RNA (gRNA) of 8747 nt organized in three open reading frames (ORFs). The ORF1, encoding a polyprotein involved in replication, is translated directly from the gRNA, whereas ORFs encoding the movement (MP) and coat (CP) proteins are expressed via 3\' coterminal subgenomic RNAs (sgRNAs). We characterized the minimal promoter region critical for the CP-sgRNA expression in infected cells by deletion analyses using Agrobacterium-mediated infection of Nicotiana benthamiana plants. The minimal CP-sgRNA promoter was mapped between nucleotides −67 and \+ 50 nt around the transcription start site. Surprisingly, larger deletions in the region between the CP-sgRNA transcription start site and the CP translation initiation codon resulted in increased CP-sgRNA accumulation, suggesting that this sequence could modulate the CP-sgRNA transcription. Site-specific mutational analysis of the transcription start site revealed that the \+ 1 guanylate and the \+ 2 adenylate are important for CP-sgRNA synthesis.} } @Article{IPB-1530, author = {Abel, S. and Theologis, A. and}, title = {{Odyssey of Auxin}}, year = {2010}, pages = {a004572}, journal = {Cold Spring Harb. Perspect. Biol.}, doi = {10.1101/cshperspect.a004572}, volume = {2}, abstract = {The history of plant biology is inexorably intertwined with the conception and discovery of auxin, followed by the many decades of research to comprehend its action during growth and development. Growth responses to auxin are complex and require the coordination of auxin production, transport, and perception. In this overview of past auxin research, we limit our discourse to the mechanism of auxin action. We attempt to trace the almost epic voyage from the birth of the hormonal concept in plants to the recent crystallographic studies that resolved the TIR1-auxin receptor complex, the first structural model of a plant hormone receptor. The century-long endeavor is a beautiful illustration of the power of scientific reasoning and human intuition, but it also brings to light the fact that decisive progress is made when new technologies emerge and disciplines unite.} } @Article{IPB-1618, author = {Dufour, D. and De la Peña, M. and Gago, S. and Flores, R. and Gallego, J. and}, title = {{Structure–function analysis of the ribozymes of chrysanthemum chlorotic mottle viroid: a loop–loop interaction motif conserved in most natural hammerheads}}, year = {2009}, pages = {368-381}, journal = {Nucleic Acids Res.}, doi = {10.1093/nar/gkn918}, volume = {37}, abstract = {Loop–loop tertiary interactions play a key role in the folding and catalytic activity of natural hammerhead ribozymes. Using a combination of NMR spectroscopy, site-directed mutagenesis and kinetic and infectivity analyses, we have examined the structure and function of loops 1 and 2 of the (\+) and (–) hammerheads of chrysanthemum chlorotic mottle viroid RNA. In both hammerheads, loop 1 is a heptanucleotide hairpin loop containing an exposed U at its 5′ side and an extrahelical U at its 3′-side critical for the catalytic activity of the ribozyme in vitro and for viroid infectivity in vivo , whereas loop 2 has a key opened A at its 3′-side. These structural features promote a specific loop–loop interaction motif across the major groove. The essential features of this tertiary structure element, base pairing between the 5′ U of loop 1 and the 3′ A of loop 2, and interaction of the extrahelical pyrimidine of loop 1 with loop 2, are likely shared by a significant fraction of natural hammerheads.} } @Article{IPB-1615, author = {Clarke, S. M. and Cristescu, S. M. and Miersch, O. and Harren, F. J. M. and Wasternack, C. and Mur, L. A. J. and}, title = {{Jasmonates act with salicylic acid to confer basal thermotolerance in Arabidopsis thaliana}}, year = {2009}, pages = {175-187}, journal = {New Phytol.}, doi = {10.1111/j.1469-8137.2008.02735.x}, volume = {182}, abstract = {The cpr5‐1 Arabidopsis thaliana mutant exhibits constitutive activation of salicylic acid (SA), jasmonic acid (JA) and ethylene (ET) signalling pathways and displays enhanced tolerance of heat stress (HS).cpr5‐1 crossed with jar1‐1 (a JA‐amino acid synthetase) was compromised in basal thermotolerance, as were the mutants opr3 (mutated in OPDA reductase3) and coi1‐1 (affected in an E3 ubiquitin ligase F‐box; a key JA‐signalling component). In addition, heating wild‐type Arabidopsis led to the accumulation of a range of jasmonates: JA, 12‐oxophytodienoic acid (OPDA) and a JA‐isoleucine (JA‐Ile) conjugate. Exogenous application of methyl jasmonate protected wild‐type Arabidopsis from HS.Ethylene was rapidly produced during HS, with levels being modulated by both JA and SA. By contrast, the ethylene mutant ein2‐1 conferred greater thermotolerance.These data suggest that JA acts with SA, conferring basal thermotolerance while ET may act to promote cell death.} } @Article{IPB-1667, author = {Weigelt, K. and Küster, H. and Rutten, T. and Fait, A. and Fernie, A. R. and Miersch, O. and Wasternack, C. and Emery, R. J. N. and Desel, C. and Hosein, F. and Müller, M. and Saalbach, I. and Weber, H. and}, title = {{ADP-Glucose Pyrophosphorylase-Deficient Pea Embryos Reveal Specific Transcriptional and Metabolic Changes of Carbon-Nitrogen Metabolism and Stress Responses}}, year = {2009}, pages = {395-411}, journal = {Plant Physiol.}, doi = {10.1104/pp.108.129940}, volume = {149}, abstract = {We present a comprehensive analysis of ADP-glucose pyrophosphorylase (AGP)-repressed pea (Pisum sativum) seeds using transcript and metabolite profiling to monitor the effects that reduced carbon flow into starch has on carbon-nitrogen metabolism and related pathways. Changed patterns of transcripts and metabolites suggest that AGP repression causes sugar accumulation and stimulates carbohydrate oxidation via glycolysis, tricarboxylic acid cycle, and mitochondrial respiration. Enhanced provision of precursors such as acetyl-coenzyme A and organic acids apparently support other pathways and activate amino acid and storage protein biosynthesis as well as pathways fed by cytosolic acetyl-coenzyme A, such as cysteine biosynthesis and fatty acid elongation/metabolism. As a consequence, the resulting higher nitrogen (N) demand depletes transient N storage pools, specifically asparagine and arginine, and leads to N limitation. Moreover, increased sugar accumulation appears to stimulate cytokinin-mediated cell proliferation pathways. In addition, the deregulation of starch biosynthesis resulted in indirect changes, such as increased mitochondrial metabolism and osmotic stress. The combined effect of these changes is an enhanced generation of reactive oxygen species coupled with an up-regulation of energy-dissipating, reactive oxygen species protection, and defense genes. Transcriptional activation of mitogen-activated protein kinase pathways and oxylipin synthesis indicates an additional activation of stress signaling pathways. AGP-repressed embryos contain higher levels of jasmonate derivatives; however, this increase is preferentially in nonactive forms. The results suggest that, although metabolic/osmotic alterations in iAGP pea seeds result in multiple stress responses, pea seeds have effective mechanisms to circumvent stress signaling under conditions in which excessive stress responses and/or cellular damage could prematurely initiate senescence or apoptosis.} } @Article{IPB-1666, author = {Wasternack, C. and Hause, B. and}, title = {{Emerging complexity: jasmonate-induced volatiles affect parasitoid choice}}, year = {2009}, pages = {2451-2453}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/erp197}, volume = {60}, } @Article{IPB-1664, author = {Vandenborre, G. and Miersch, O. and Hause, B. and Smagghe, G. and Wasternack, C. and Van Damme, E. J. and}, title = {{Spodoptera littoralis-Induced Lectin Expression in Tobacco}}, year = {2009}, pages = {1142-1155}, journal = {Plant Cell Physiol.}, doi = {10.1093/pcp/pcp065}, volume = {50}, abstract = {The induced defense response in plants towards herbivores is mainly regulated by jasmonates and leads to the accumulation of so-called jasmonate-induced proteins. Recently, a jasmonate (JA) inducible lectin called Nicotiana tabacum agglutinin or NICTABA was discovered in tobacco (N. tabacum cv Samsun) leaves. Tobacco plants also accumulate the lectin after insect attack by caterpillars. To study the functional role of NICTABA, the accumulation of the JA precursor 12-oxophytodienoic acid (OPDA), JA as well as different JA metabolites were analyzed in tobacco leaves after herbivory by larvae of the cotton leafworm (Spodoptera littoralis) and correlated with NICTABA accumulation. It was shown that OPDA, JA as well as its methyl ester can trigger NICTABA accumulation. However, hydroxylation of JA and its subsequent sulfation and glucosylation results in inactive compounds that have lost the capacity to induce NICTABA gene expression. The expression profile of NICTABA after caterpillar feeding was recorded in local as well as in systemic leaves, and compared to the expression of several genes encoding defense proteins, and genes encoding a tobacco systemin and the allene oxide cyclase, an enzyme in JA biosynthesis. Furthermore, the accumulation of NICTABA was quanti-fied after S. littoralis herbivory and immunofluorescence microscopy was used to study the localization of NICTABA in the tobacco leaf.} } @Article{IPB-1662, author = {Ticconi, C. A. and Lucero, R. D. and Sakhonwasee, S. and Adamson, A. W. and Creff, A. and Nussaume, L. and Desnos, T. and Abel, S. and}, title = {{ER-resident proteins PDR2 and LPR1 mediate the developmental response of root meristems to phosphate availability}}, year = {2009}, pages = {14174-14179}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.0901778106}, volume = {106}, abstract = {Inadequate availability of inorganic phosphate (Pi) in the rhizosphere is a common challenge to plants, which activate metabolic and developmental responses to maximize Pi acquisition. The sensory mechanisms that monitor environmental Pi status and regulate root growth via altered meristem activity are unknown. Here, we show that phosphate deficiency response 2 (PDR2) encodes the single P5-type ATPase of Arabidopsis thaliana. PDR2 functions in the endoplasmic reticulum (ER) and is required for proper expression of scarecrow (SCR), a key regulator of root patterning, and for stem-cell maintenance in Pi-deprived roots. We further show that the multicopper oxidase encoded by low phosphate root 1 (LPR1) is targeted to the ER and that LPR1 and PDR2 interact genetically. Because the expression domains of both genes overlap in the stem-cell niche and distal root meristem, we propose that PDR2 and LPR1 function together in an ER-resident pathway that adjusts root meristem activity to external Pi. Our data indicate that the Pi-conditional root phenotype of pdr2 is not caused by increased Fe availability in low Pi; however, Fe homeostasis modifies the developmental response of root meristems to Pi availability.} } @Article{IPB-1657, author = {Serra, P. and BANI HASHEMIAN, S. M. and PENSABENE-BELLAVIA, G. and Gago, S. and DURAN-VILA, N. and}, title = {{An artificial chimeric derivative of Citrus viroid V involves the terminal left domain in pathogenicity}}, year = {2009}, pages = {515-522}, journal = {Mol. Plant Pathol.}, doi = {10.1111/j.1364-3703.2009.00553.x}, volume = {10}, abstract = {The recently described Citrus viroid V (CVd‐V) induces, in Etrog citron, mild stunting and very small necrotic lesions and cracks, sometimes filled with gum. As Etrog citron plants co‐infected with Citrus dwarfing viroid (CDVd) and CVd‐V show synergistic interactions, these host–viroid combinations provide a convenient model to identify the pathogenicity determinant(s). The biological effects of replacing limited portions of the rod‐like structure of CVd‐V with the corresponding portions of CDVd are reported. Chimeric constructs were synthesized using a novel polymerase chain reaction‐based approach, much more flexible than those based on restriction enzymes used in previous studies. Of the seven chimeras (Ch) tested, only one (Ch5) proved to be infectious. Plants infected with Ch5 showed no symptoms and, although this novel chimera was able to replicate to relatively high titres in singly infected plants, it was rapidly displaced by either CVd‐V or CDVd in doubly infected plants. The results demonstrate that direct interaction(s) between structural elements in the viroid RNA (in this case, the terminal left domain) and as yet unidentified host factors play an important role in modulating viroid pathogenicity. This is the first pathogenic determinant mapped in species of the genus Apscaviroid.} } @Article{IPB-1655, author = {Santner, A. and Calderon-Villalobos, L. I. A. and Estelle, M. and}, title = {{Plant hormones are versatile chemical regulators of plant growth}}, year = {2009}, pages = {301-307}, journal = {Nat. Chem. Biol.}, doi = {10.1038/nchembio.165}, volume = {5}, abstract = {The plant hormones are a structurally unrelated collection of small molecules derived from various essential metabolic pathways. These compounds are important regulators of plant growth and mediate responses to both biotic and abiotic stresses. During the last ten years there have been many exciting advances in our understanding of plant hormone biology, including new discoveries in the areas of hormone biosynthesis, transport, perception and response. Receptors for many of the major hormones have now been identified, providing new opportunities to study the chemical specificity of hormone signaling. These studies also reveal a surprisingly important role for the ubiquitin-proteasome pathway in hormone signaling. In addition, recent work confirms that hormone signaling interacts at multiple levels during plant growth and development. In the future, a major challenge will be to understand how the information conveyed by these simple compounds is integrated during plant growth.} } @Article{IPB-1652, author = {Quint, M. and Barkawi, L. S. and Fan, K.-T. and Cohen, J. D. and Gray, W. M. and}, title = {{Arabidopsis IAR4 Modulates Auxin Response by Regulating Auxin Homeostasis}}, year = {2009}, pages = {748-758}, journal = {Plant Physiol.}, doi = {10.1104/pp.109.136671}, volume = {150}, abstract = {In a screen for enhancers of tir1-1 auxin resistance, we identified two novel alleles of the putative mitochondrial pyruvate dehydrogenase E1α-subunit, IAA-Alanine Resistant4 (IAR4). In addition to enhancing the auxin response defects of tir1-1, iar4 single mutants exhibit numerous auxin-related phenotypes including auxin-resistant root growth and reduced lateral root development, as well as defects in primary root growth, root hair initiation, and root hair elongation. Remarkably, all of these iar4 mutant phenotypes were rescued when endogenous indole-3-acetic acid (IAA) levels were increased by growth at high temperature or overexpression of the YUCCA1 IAA biosynthetic enzyme, suggesting that iar4 mutations may alter IAA homeostasis rather than auxin response. Consistent with this possibility, iar4 mutants exhibit increased Aux/IAA stability compared to wild type under basal conditions, but not in response to an auxin treatment. Measurements of free IAA levels detected no significant difference between iar4-3 and wild-type controls. However, we consistently observed significantly higher levels of IAA-amino acid conjugates in the iar4-3 mutant. Furthermore, using stable isotope-labeled IAA precursors, we observed a significant increase in the relative utilization of the Trp-independent IAA biosynthetic pathway in iar4-3. We therefore suggest that the auxin phenotypes of iar4 mutants are the result of altered IAA homeostasis.} } @Article{IPB-1651, author = {Pienkny, S. and Brandt, W. and Schmidt, J. and Kramell, R. and Ziegler, J. and}, title = {{Functional characterization of a novel benzylisoquinoline O-methyltransferase suggests its involvement in papaverine biosynthesis in opium poppy (Papaver somniferum L)}}, year = {2009}, pages = {56-67}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2009.03937.x}, volume = {60}, abstract = {The benzylisoquinoline alkaloids are a highly diverse group of about 2500 compounds which accumulate in a species‐specific manner. Despite the numerous compounds which could be identified, the biosynthetic pathways and the participating enzymes or cDNAs could be characterized only for a few selected members, whereas the biosynthesis of the majority of the compounds is still largely unknown. In an attempt to characterize additional biosynthetic steps at the molecular level, integration of alkaloid and transcript profiling across Papaver species was performed. This analysis showed high expression of an expressed sequence tag (EST) of unknown function only in Papaver somniferum varieties. After full‐length cloning of the open reading frame and sequence analysis, this EST could be classified as a member of the class II type O ‐methyltransferase protein family. It was related to O ‐methyltransferases from benzylisoquinoline biosynthesis, and the amino acid sequence showed 68% identical residues to norcoclaurine 6‐O ‐methyltransferase. However, rather than methylating norcoclaurine, the recombinant protein methylated norreticuline at position seven with a K m of 44 μm using S ‐adenosyl‐l ‐methionine as a cofactor. Of all substrates tested, only norreticuline was converted. Even minor changes in the benzylisoquinoline backbone were not tolerated by the enzyme. Accordingly, the enzyme was named norreticuline 7–O ‐methyltransferase (N7OMT). This enzyme represents a novel O ‐methyltransferase in benzylisoquinoline metabolism. Expression analysis showed slightly increased expression of N7OMT in P. somniferum varieties containing papaverine, suggesting its involvement in the partially unknown biosynthesis of this pharmaceutically important compound.} } @Article{IPB-1650, author = {Parry, G. and Calderon-Villalobos, L. I. and Prigge, M. and Peret, B. and Dharmasiri, S. and Itoh, H. and Lechner, E. and Gray, W. M. and Bennett, M. and Estelle, M. and}, title = {{Complex regulation of the TIR1/AFB family of auxin receptors}}, year = {2009}, pages = {22540-22545}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.0911967106}, volume = {106}, abstract = {Auxin regulates most aspects of plant growth and development. The hormone is perceived by the TIR1/AFB family of F-box proteins acting in concert with the Aux/IAA transcriptional repressors. Arabidopsis plants that lack members of the TIR1/AFB family are auxin resistant and display a variety of growth defects. However, little is known about the functional differences between individual members of the family. Phylogenetic studies reveal that the TIR1/AFB proteins are conserved across land plant lineages and fall into four clades. Three of these subgroups emerged before separation of angiosperms and gymnosperms whereas the last emerged before the monocot-eudicot split. This evolutionary history suggests that the members of each clade have distinct functions. To explore this possibility in Arabidopsis, we have analyzed a range of mutant genotypes, generated promoter swap transgenic lines, and performed in vitro binding assays between individual TIR1/AFB and Aux/IAA proteins. Our results indicate that the TIR1/AFB proteins have distinct biochemical activities and that TIR1 and AFB2 are the dominant auxin receptors in the seedling root. Further, we demonstrate that TIR1, AFB2, and AFB3, but not AFB1 exhibit significant posttranscriptional regulation. The microRNA miR393 is expressed in a pattern complementary to that of the auxin receptors and appears to regulate TIR1/AFB expression. However our data suggest that this regulation is complex. Our results suggest that differences between members of the auxin receptor family may contribute to the complexity of auxin response.} } @Article{IPB-1648, author = {Mugford, S. G. and Yoshimoto, N. and Reichelt, M. and Wirtz, M. and Hill, L. and Mugford, S. T. and Nakazato, Y. and Noji, M. and Takahashi, H. and Kramell, R. and Gigolashvili, T. and Flügge, U.-I. and Wasternack, C. and Gershenzon, J. and Hell, R. and Saito, K. and Kopriva, S. and}, title = {{Disruption of Adenosine-5′-Phosphosulfate Kinase in Arabidopsis Reduces Levels of Sulfated Secondary Metabolites}}, year = {2009}, pages = {910-927}, journal = {Plant Cell}, doi = {10.1105/tpc.109.065581}, volume = {21}, abstract = {Plants can metabolize sulfate by two pathways, which branch at the level of adenosine 5′-phosphosulfate (APS). APS can be reduced to sulfide and incorporated into Cys in the primary sulfate assimilation pathway or phosphorylated by APS kinase to 3′-phosphoadenosine 5′-phosphosulfate, which is the activated sulfate form for sulfation reactions. To assess to what extent APS kinase regulates accumulation of sulfated compounds, we analyzed the corresponding gene family in Arabidopsis thaliana. Analysis of T-DNA insertion knockout lines for each of the four isoforms did not reveal any phenotypical alterations. However, when all six combinations of double mutants were compared, the apk1 apk2 plants were significantly smaller than wild-type plants. The levels of glucosinolates, a major class of sulfated secondary metabolites, and the sulfated 12-hydroxyjasmonate were reduced approximately fivefold in apk1 apk2 plants. Although auxin levels were increased in the apk1 apk2 mutants, as is the case for most plants with compromised glucosinolate synthesis, typical high auxin phenotypes were not observed. The reduction in glucosinolates resulted in increased transcript levels for genes involved in glucosinolate biosynthesis and accumulation of desulfated precursors. It also led to great alterations in sulfur metabolism: the levels of sulfate and thiols increased in the apk1 apk2 plants. The data indicate that the APK1 and APK2 isoforms of APS kinase play a major role in the synthesis of secondary sulfated metabolites and are required for normal growth rates.} } @Article{IPB-1640, author = {Lee, C.-W. and Efetova, M. and Engelmann, J. C. and Kramell, R. and Wasternack, C. and Ludwig-Müller, J. and Hedrich, R. and Deeken, R. and}, title = {{Agrobacterium tumefaciens Promotes Tumor Induction by Modulating Pathogen Defense in Arabidopsis thaliana}}, year = {2009}, pages = {2948-2962}, journal = {Plant Cell}, doi = {10.1105/tpc.108.064576}, volume = {21}, abstract = {Agrobacterium tumefaciens causes crown gall disease by transferring and integrating bacterial DNA (T-DNA) into the plant genome. To examine the physiological changes and adaptations during Agrobacterium-induced tumor development, we compared the profiles of salicylic acid (SA), ethylene (ET), jasmonic acid (JA), and auxin (indole-3-acetic acid [IAA]) with changes in the Arabidopsis thaliana transcriptome. Our data indicate that host responses were much stronger toward the oncogenic strain C58 than to the disarmed strain GV3101 and that auxin acts as a key modulator of the Arabidopsis–Agrobacterium interaction. At initiation of infection, elevated levels of IAA and ET were associated with the induction of host genes involved in IAA, but not ET signaling. After T-DNA integration, SA as well as IAA and ET accumulated, but JA did not. This did not correlate with SA-controlled pathogenesis-related gene expression in the host, although high SA levels in mutant plants prevented tumor development, while low levels promoted it. Our data are consistent with a scenario in which ET and later on SA control virulence of agrobacteria, whereas ET and auxin stimulate neovascularization during tumor formation. We suggest that crosstalk among IAA, ET, and SA balances pathogen defense launched by the host and tumor growth initiated by agrobacteria.} } @Article{IPB-1633, author = {Hause, B. and Wasternack, C. and Strack, D. and}, title = {{Jasmonates in stress responses and development}}, year = {2009}, pages = {1483-1484}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2009.07.004}, volume = {70}, } @Article{IPB-1632, author = {Halim, V. A. and Altmann, S. and Ellinger, D. and Eschen-Lippold, L. and Miersch, O. and Scheel, D. and Rosahl, S. and}, title = {{PAMP-induced defense responses in potato require both salicylic acid and jasmonic acid}}, year = {2009}, pages = {230-242}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2008.03688.x}, volume = {57}, abstract = {To elucidate the molecular mechanisms underlying pathogen‐associated molecular pattern (PAMP)‐induced defense responses in potato (Solanum tuberosum ), the role of the signaling compounds salicylic acid (SA) and jasmonic acid (JA) was analyzed. Pep‐13, a PAMP from Phytophthora , induces the accumulation of SA, JA and hydrogen peroxide, as well as the activation of defense genes and hypersensitive‐like cell death. We have previously shown that SA is required for Pep‐13‐induced defense responses. To assess the importance of JA, RNA interference constructs targeted at the JA biosynthetic genes, allene oxide cyclase and 12‐oxophytodienoic acid reductase, were expressed in transgenic potato plants. In addition, expression of the F‐box protein COI1 was reduced by RNA interference. Plants expressing the RNA interference constructs failed to accumulate the respective transcripts in response to wounding or Pep‐13 treatment, neither did they contain significant amounts of JA after elicitation. In response to infiltration of Pep‐13, the transgenic plants exhibited a highly reduced accumulation of reactive oxygen species as well as reduced hypersensitive cell death. The ability of the JA‐deficient plants to accumulate SA suggests that SA accumulation is independent or upstream of JA accumulation. These data show that PAMP responses in potato require both SA and JA and that, in contrast to Arabidopsis, these compounds act in the same signal transduction pathway. Despite their inability to fully respond to PAMP treatment, the transgenic RNA interference plants are not altered in their basal defense against Phytophthora infestans .} } @Article{IPB-1628, author = {Gago, S. and Elena, S. F. and Flores, R. and Sanjuan, R. and}, title = {{Extremely High Mutation Rate of a Hammerhead Viroid}}, year = {2009}, pages = {1308-1308}, journal = {Science}, doi = {10.1126/science.1169202}, volume = {323}, abstract = {The mutation rates of viroids, plant pathogens with minimal non-protein-coding RNA genomes, are unknown. Their replication is mediated by host RNA polymerases and, in some cases, by hammerhead ribozymes, small self-cleaving motifs embedded in the viroid. By using the principle that the population frequency of nonviable genotypes equals the mutation rate, we screened for changes that inactivated the hammerheads of Chrysanthemum chlorotic mottle viroid. We obtained a mutation rate of 1/400 per site, the highest reported for any biological entity. Such error-prone replication can only be tolerated by extremely simple genomes such as those of viroids and, presumably, the primitive replicons of the RNA world. Our results suggest that the emergence of replication fidelity was critical for the evolution of complexity in the early history of life.} } @Article{IPB-1627, author = {Fonseca, S. and Chini, A. and Hamberg, M. and Adie, B. and Porzel, A. and Kramell, R. and Miersch, O. and Wasternack, C. and Solano, R. and}, title = {{(\+)-7-iso-Jasmonoyl-L-isoleucine is the endogenous bioactive jasmonate}}, year = {2009}, pages = {344-350}, journal = {Nat. Chem. Biol.}, doi = {10.1038/nchembio.161}, volume = {5}, abstract = {Hormone-triggered activation of the jasmonate signaling pathway in Arabidopsis thaliana requires SCFCOI1-mediated proteasome degradation of JAZ repressors. (−)-JA-L-Ile is the proposed bioactive hormone, and SCFCOI1 is its likely receptor. We found that the biological activity of (−)-JA-L-Ile is unexpectedly low compared to coronatine and the synthetic isomer (\+)-JA-L-Ile, which suggests that the stereochemical orientation of the cyclopentanone-ring side chains greatly affects receptor binding. Detailed GC-MS and HPLC analyses showed that the (−)-JA-L-Ile preparations currently used in ligand binding studies contain small amounts of the C7 epimer (\+)-7-iso-JA-L-Ile. Purification of each of these molecules demonstrated that pure (−)-JA-L-Ile is inactive and that the active hormone is (\+)-7-iso-JA-L-Ile, which is also structurally more similar to coronatine. In addition, we show that pH changes promote conversion of (\+)-7-iso-JA-L-Ile to the inactive (−)-JA-L-Ile form, thus providing a simple mechanism that can regulate hormone activity through epimerization.} } @Article{IPB-1625, author = {Flores, R. and Gas, M.-E. and Molina-Serrano, D. and Nohales, M.-?. and Carbonell, A. and Gago, S. and De la Peña, M. and Daròs, J.-A. and}, title = {{Viroid Replication: Rolling-Circles, Enzymes and Ribozymes}}, year = {2009}, pages = {317-334}, journal = {Viruses}, doi = {10.3390/v1020317}, volume = {1}, abstract = {Viroids, due to their small size and lack of protein-coding capacity, must rely essentially on their hosts for replication. Intriguingly, viroids have evolved the ability to replicate in two cellular organella, the nucleus (family Pospiviroidae) and the chloroplast (family Avsunviroidae). Viroid replication proceeds through an RNA-based rolling-circle mechanism with three steps that, with some variations, operate in both polarity strands: i) synthesis of longer-than-unit strands catalyzed by either the nuclear RNA polymerase II or a nuclear-encoded chloroplastic RNA polymerase, in both instances redirected to transcribe RNA templates, ii) cleavage to unit-length, which in the family Avsunviroidae is mediated by hammerhead ribozymes embedded in both polarity strands, while in the family Pospiviroidae the oligomeric RNAs provide the proper conformation but not the catalytic activity, and iii) circularization. The host RNA polymerases, most likely assisted by additional host proteins, start transcription from specific sites, thus implying the existence of viroid promoters. Cleavage and ligation in the family Pospiviroidae is probably catalyzed by an RNase III-like enzyme and an RNA ligase able to circularize the resulting 5’ and 3’ termini. Whether a chloroplastic RNA ligase mediates circularization in the family Avsunviroidae, or this reaction is autocatalytic, remains an open issue.} } @Article{IPB-1672, author = {Ziegler, J. and Brandt, W. and Geißler, R. and Facchini, P. J. and}, title = {{Removal of Substrate Inhibition and Increase in Maximal Velocity in the Short Chain Dehydrogenase/Reductase Salutaridine Reductase Involved in Morphine Biosynthesis}}, year = {2009}, pages = {26758-26767}, journal = {J. Biol. Chem.}, doi = {10.1074/jbc.M109.030957}, volume = {284}, abstract = {Salutaridine reductase (SalR, EC 1.1.1.248) catalyzes the stereospecific reduction of salutaridine to 7(S)-salutaridinol in the biosynthesis of morphine. It belongs to a new, plant-specific class of short-chain dehydrogenases, which are characterized by their monomeric nature and increased length compared with related enzymes. Homology modeling and substrate docking suggested that additional amino acids form a novel α-helical element, which is involved in substrate binding. Site-directed mutagenesis and subsequent studies on enzyme kinetics revealed the importance of three residues in this element for substrate binding. Further replacement of eight additional residues led to the characterization of the entire substrate binding pocket. In addition, a specific role in salutaridine binding by either hydrogen bond formation or hydrophobic interactions was assigned to each amino acid. Substrate docking also revealed an alternative mode for salutaridine binding, which could explain the strong substrate inhibition of SalR. An alternate arrangement of salutaridine in the enzyme was corroborated by the effect of various amino acid substitutions on substrate inhibition. In most cases, the complete removal of substrate inhibition was accompanied by a substantial loss in enzyme activity. However, some mutations greatly reduced substrate inhibition while maintaining or even increasing the maximal velocity. Based on these results, a double mutant of SalR was created that exhibited the complete absence of substrate inhibition and higher activity compared with wild-type SalR.} } @Article{IPB-1671, author = {Ziegler, J. and Facchini, P. J. and Geißler, R. and Schmidt, J. and Ammer, C. and Kramell, R. and Voigtländer, S. and Gesell, A. and Pienkny, S. and Brandt, W. and}, title = {{Evolution of morphine biosynthesis in opium poppy}}, year = {2009}, pages = {1696-1707}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2009.07.006}, volume = {70}, abstract = {Benzylisoquinoline alkaloids (BIAs) are a group of nitrogen-containing plant secondary metabolites comprised of an estimated 2500 identified structures. In BIA metabolism, (S)-reticuline is a key branch-point intermediate that can be directed into several alkaloid subtypes with different structural skeleton configurations. The morphinan alkaloids are one subclass of BIAs produced in only a few plant species, most notably and abundantly in the opium poppy (Papaver somniferum). Comparative transcriptome analysis of opium poppy and several other Papaver species that do not accumulate morphinan alkaloids showed that known genes encoding BIA biosynthetic enzymes are expressed at higher levels in P. somniferum. Three unknown cDNAs that are co-ordinately expressed with several BIA biosynthetic genes were identified as enzymes in the pathway. One of these enzymes, salutaridine reductase (SalR), which is specific for the production of morphinan alkaloids, was isolated and heterologously overexpressed in its active form not only from P. somniferum, but also from Papaver species that do not produce morphinan alkaloids. SalR is a member of a class of short chain dehydrogenase/reductases (SDRs) that are active as monomers and possess an extended amino acid sequence compared with classical SDRs. Homology modelling and substrate docking revealed the substrate binding site for SalR. The amino acids residues conferring salutaridine binding were compared to several members of the SDR family from different plant species, which non-specifically reduce (−)-menthone to (\+)-neomenthol. Previously, it was shown that some of these proteins are involved in plant defence. The recruitment of specific monomeric SDRs from monomeric SDRs involved in plant defence is discussed.} } @Article{IPB-1759, author = {Zhang, W. and Ito, H. and Quint, M. and Huang, H. and Noel, L. D. and Gray, W. M. and}, title = {{Genetic analysis of CAND1-CUL1 interactions in Arabidopsis supports a role for CAND1-mediated cycling of the SCFTIR1 complex}}, year = {2008}, pages = {8470-8475}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.0804144105}, volume = {105}, abstract = {SKP1-Cullin1-F-box protein (SCF) ubiquitin-ligases regulate numerous aspects of eukaryotic growth and development. Cullin-Associated and Neddylation-Dissociated (CAND1) modulates SCF function through its interactions with the CUL1 subunit. Although biochemical studies with human CAND1 suggested that CAND1 plays a negative regulatory role by sequestering CUL1 and preventing SCF complex assembly, genetic studies in Arabidopsis have shown that cand1 mutants exhibit reduced SCF activity, demonstrating that CAND1 is required for optimal SCF function in vivo. Together, these genetic and biochemical studies have suggested a model of CAND1-mediated cycles of SCF complex assembly and disassembly. Here, using the SCFTIR1 complex of the Arabidopsis auxin response pathway, we test the SCF cycling model with Arabidopsis mutant derivatives of CAND1 and CUL1 that have opposing effects on the CAND1–CUL1 interaction. We find that the disruption of the CAND1–CUL1 interaction results in an increased abundance of assembled SCFTIR1 complex. In contrast, stabilization of the CAND1–CUL1 interaction diminishes SCFTIR1 complex abundance. The fact that both decreased and increased CAND1–CUL1 interactions result in reduced SCFTIR1 activity in vivo strongly supports the hypothesis that CAND1-mediated cycling is required for optimal SCF function.} } @Article{IPB-1752, author = {Wasternack, C. and Feussner, I. and}, title = {{Multifunctional Enzymes in Oxylipin Metabolism}}, year = {2008}, pages = {2373-2375}, journal = {ChemBioChem}, doi = {10.1002/cbic.200800582}, volume = {9}, abstract = {For the first time a member of the CYP74 enzyme subfamily (9‐AOS) from tomato has been shown by chemical and analytical approaches to catalyze multiple reactions. These multifunctional properties of 9‐AOS from the oxylipin‐forming lipoxygenase (LOX) pathway raise several new questions on lipid‐derived signaling.} } @Article{IPB-1739, author = {Stenzel, I. and Hause, B. and Proels, R. and Miersch, O. and Oka, M. and Roitsch, T. and Wasternack, C. and}, title = {{The AOC promoter of tomato is regulated by developmental and environmental stimuli}}, year = {2008}, pages = {1859-1869}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2008.03.007}, volume = {69}, abstract = {The allene oxide cyclase (AOC) catalyzes the formation of cis-(\+)-12-oxophytodienoic acid, an intermediate in jasmonate biosynthesis and is encoded by a single copy gene in tomato. The full length AOC promoter isolated by genome walk contains 3600 bp. Transgenic tomato lines carrying a 1000 bp promoter fragment and the full length promoter, respectively, in front of the β-glucuronidase (GUS)-encoding uidA gene and several tobacco lines carrying the full length tomato AOC promoter before GUS were used to record organ- and tissue-specific promoter activities during development and in response to various stimuli. High promoter activities corresponding to immunocytochemically detected occurrence of the AOC protein were found in seeds and young seedlings and were confined to the root tip, hypocotyl and cotyledons of 3-d-old seedlings. In 10-d-old seedlings promoter activity appeared preferentially in the elongation zone. Fully developed tomato leaves were free of AOC promoter activity, but showed high activity upon wounding locally and systemically or upon treatment with JA, systemin or glucose. Tomato flowers showed high AOC promoter activities in ovules, sepals, anthers and pollen. Most of the promoter activity patterns found in tomato with the 1000 bp promoter fragment were also detected with the full length tomato AOC promoter in tobacco during development or in response to various stimuli. The data support a spatial and temporal regulation of JA biosynthesis during development and in response to environmental stimuli.} } @Article{IPB-1736, author = {Serra, P. and Gago, S. and Duran-Vila, N. and}, title = {{A single nucleotide change in Hop stunt viroid modulates citrus cachexia symptoms}}, year = {2008}, pages = {130-134}, journal = {Virus Res.}, doi = {10.1016/j.virusres.2008.08.003}, volume = {138}, abstract = {Cachexia disease of citrus is caused by Hop stunt viroid (HSVd). In citrus, pathogenic and non-pathogenic strains differ by a “cachexia expression motif” of five to six nucleotides located in the variable domain of the proposed rod-like secondary structure. Here, site-directed mutants were generated to investigate if all these nucleotides were required for infectivity and/or symptom expression. Specifically an artificial cachexia inducing mutant M0 was generated by introducing the six nucleotides changes of the “cachexia expression motif” into a non-pathogenic sequence variant and M0 was used as a template to systematically restore some of the introduced changes. The resulting mutants in which specific changes introduced to generate M0, were restored presented a variety of responses: (i) M1, obtained by introducing two insertions forming a base-pair, was infectious but non-pathogenic; (ii) M2, obtained by introducing an insertion and restoring a substitution, presented low infectivity and the resulting progeny reverted to M0; (iii) M3, obtained by restoring a single substitution in the lower strand of the viroid secondary structure, was infectious but induced only mild cachexia symptoms; (iv) M4, obtained by restoring a single susbtitution in the upper strand of the viroid secondary structure, was non-infectious. These results confirm that the “cachexia expression motif” plays a major role in inciting cachexia symptoms, and that subtle changes within this motif affect symptom severity and may even suppress symptom expression.} } @Article{IPB-1731, author = {Schilling, S. and Wasternack, C. and Demuth, H.-U. and}, title = {{Glutaminyl cyclases from animals and plants: a case of functionally convergent protein evolution}}, year = {2008}, journal = {Biol. Chem.}, doi = {10.1515/BC.2008.111}, volume = {389}, abstract = {Several mammalian peptide hormones and proteins from plant and animal origin contain an N-terminal pyroglutamic acid (pGlu) residue. Frequently, the moiety is important in exerting biological function in either mediating interaction with receptors or stabilizing against N-terminal degradation. Glutaminyl cyclases (QCs) were isolated from different plants and animals catalyzing pGlu formation. The recent resolution of the 3D structures of Carica papaya and human QCs clearly supports different evolutionary origins of the proteins, which is also reflected by different enzymatic mechanisms. The broad substrate specificity is revealed by the heterogeneity of physiological substrates of plant and animal QCs, including cytokines, matrix proteins and pathogenesis-related proteins. Moreover, recent evidence also suggests human QC as a catalyst of pGlu formation at the N-terminus of amyloid peptides, which contribute to Alzheimer\'s disease. Obviously, owing to its biophysical properties, the function of pGlu in plant and animal proteins is very similar in terms of stabilizing or mediating protein and peptide structure. It is possible that the requirement for catalysis of pGlu formation under physiological conditions may have triggered separate evolution of QCs in plants and animals.} } @Article{IPB-1696, author = {Floß, D. S. and Hause, B. and Lange, P. R. and Küster, H. and Strack, D. and Walter, M. H. and}, title = {{Knock-down of the MEP pathway isogene 1-deoxy-d-xylulose 5-phosphate synthase 2 inhibits formation of arbuscular mycorrhiza-induced apocarotenoids, and abolishes normal expression of mycorrhiza-specific plant marker genes}}, year = {2008}, pages = {86-100}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2008.03575.x}, volume = {56}, abstract = {The first step of the plastidial methylerythritol phosphate (MEP) pathway is catalyzed by two isoforms of 1‐deoxy‐d‐ xylulose 5‐phosphate synthase (DXS1 and DXS2). In Medicago truncatula , MtDXS1 and MtDXS2 genes exhibit completely different expression patterns. Most prominently, colonization by arbuscular mycorrhizal (AM) fungi induces the accumulation of certain apocarotenoids (cyclohexenone and mycorradicin derivatives) correlated with the expression of MtDXS2 but not of MtDXS1. To prove a distinct function of DXS2, a selective RNAi approach on MtDXS2 expression was performed in transgenic hairy roots of M. truncatula. Repression of MtDXS2 consistently led to reduced transcript levels in mycorrhizal roots, and to a concomitant reduction of AM‐induced apocarotenoid accumulation. The transcript levels of MtDXS1 remained unaltered in RNAi plants, and no phenotypical changes in non‐AM plants were observed. Late stages of the AM symbiosis were adversely affected, but only upon strong repression with residual MtDXS2‐1 transcript levels remaining below approximately 10%. This condition resulted in a strong decrease in the transcript levels of MtPT4 , an AM‐specific plant phosphate transporter gene, and in a multitude of other AM‐induced plant marker genes, as shown by transcriptome analysis. This was accompanied by an increased proportion of degenerating and dead arbuscules at the expense of mature ones. The data reveal a requirement for DXS2‐dependent MEP pathway‐based isoprenoid products to sustain mycorrhizal functionality at later stages of the symbiosis. They further validate the concept of a distinct role for DXS2 in secondary metabolism, and offer a novel tool to selectively manipulate the levels of secondary isoprenoids by targeting their precursor supply.} } @Article{IPB-1695, author = {Fellenberg, C. and Milkowski, C. and Hause, B. and Lange, P.-R. and Böttcher, C. and Schmidt, J. and Vogt, T. and}, title = {{Tapetum-specific location of a cation-dependent O-methyltransferase in Arabidopsis thaliana}}, year = {2008}, pages = {132-145}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2008.03576.x}, volume = {56}, abstract = {Cation‐ and S ‐adenosyl‐l ‐methionine (AdoMet)‐dependent plant natural product methyltransferases are referred to as CCoAOMTs because of their preferred substrate, caffeoyl coenzyme A (CCoA). The enzymes are encoded by a small family of genes, some of which with a proven role in lignin monomer biosynthesis. In Arabidopsis thaliana individual members of this gene family are temporally and spatially regulated. The gene At1g67990 is specifically expressed in flower buds, and is not detected in any other organ, such as roots, leaves or stems. Several lines of evidence indicate that the At1g67990 transcript is located in the flower buds, whereas the corresponding CCoAOMT‐like protein, termed AtTSM1, is located exclusively in the tapetum of developing stamen. Flowers of At1g67990 RNAi‐suppressed plants are characterized by a distinct flower chemotype with severely reduced levels of the N  ′,N  ′′‐ bis‐(5‐hydroxyferuloyl)‐N  ′′′‐sinapoylspermidine compensated for by N1 ,N5 ,N10 ‐tris‐(5‐hydroxyferuloyl)spermidine derivative, which is characterized by the lack of a single methyl group in the sinapoyl moiety. This severe change is consistent with the observed product profile of AtTSM1 for aromatic phenylpropanoids. Heterologous expression of the recombinant protein shows the highest activity towards a series of caffeic acid esters, but 5‐hydroxyferuloyl spermidine conjugates are also accepted substrates. The in vitro substrate specificity and the in vivo RNAi‐mediated suppression data of the corresponding gene suggest a role of this cation‐dependent CCoAOMT‐like protein in the stamen/pollen development of A. thaliana .} } @Article{IPB-1691, author = {Delker, C. and Raschke, A. and Quint, M. and}, title = {{Auxin dynamics: the dazzling complexity of a small molecule’s message}}, year = {2008}, pages = {929-941}, journal = {Planta}, doi = {10.1007/s00425-008-0710-8}, volume = {227}, abstract = {The phytohormone auxin is a potent regulator of plant development. Since its discovery in the beginning of the twentieth century many aspects of auxin biology have been extensively studied, ranging from biosynthesis and metabolism to the elucidation of molecular components of downstream signaling. With the identification of the F-box protein TIR1 as an auxin receptor a major breakthrough in understanding auxin signaling has been achieved and recent modeling approaches have shed light on the putative mechanisms underlying the establishment of auxin gradients and maxima essential for many auxin-regulated processes. Here, we review these and other recent advances in unraveling the entanglement of biosynthesis, polar transport and cellular signaling events that allow small auxinic molecules to facilitate their complex regulatory action.} } @Article{IPB-1688, author = {Carbonell, A. and Martínez de Alba, A.-E. and Flores, R. and Gago, S. and}, title = {{Double-stranded RNA interferes in a sequence-specific manner with the infection of representative members of the two viroid families}}, year = {2008}, pages = {44-53}, journal = {Virology}, doi = {10.1016/j.virol.2007.09.031}, volume = {371}, abstract = {Infection by viroids, non-protein-coding circular RNAs, occurs with the accumulation of 21–24 nt viroid-derived small RNAs (vd-sRNAs) with characteristic properties of small interfering RNAs (siRNAs) associated to RNA silencing. The vd-sRNAs most likely derive from dicer-like (DCL) enzymes acting on viroid-specific dsRNA, the key elicitor of RNA silencing, or on the highly structured genomic RNA. Previously, viral dsRNAs delivered mechanically or agroinoculated have been shown to interfere with virus infection in a sequence-specific manner. Here, we report similar results with members of the two families of nuclear- and chloroplast-replicating viroids. Moreover, homologous vd-sRNAs co-delivered mechanically also interfered with one of the viroids examined. The interference was sequence-specific, temperature-dependent and, in some cases, also dependent on the dose of the co-inoculated dsRNA or vd-sRNAs. The sequence-specific nature of these effects suggests the involvement of the RNA induced silencing complex (RISC), which provides sequence specificity to RNA silencing machinery. Therefore, viroid titer in natural infections might be regulated by the concerted action of DCL and RISC. Viroids could have evolved their secondary structure as a compromise between resistance to DCL and RISC, which act preferentially against RNAs with compact and relaxed secondary structures, respectively. In addition, compartmentation, association with proteins or active replication might also help viroids to elude their host RNA silencing machinery.} } @Article{IPB-1686, author = {Brüx, A. and Liu, T.-Y. and Krebs, M. and Stierhof, Y.-D. and Lohmann, J. U. and Miersch, O. and Wasternack, C. and Schumacher, K. and}, title = {{Reduced V-ATPase Activity in the trans-Golgi Network Causes Oxylipin-Dependent Hypocotyl Growth Inhibition in Arabidopsis}}, year = {2008}, pages = {1088-1100}, journal = {Plant Cell}, doi = {10.1105/tpc.108.058362}, volume = {20}, abstract = {Regulated cell expansion allows plants to adapt their morphogenesis to prevailing environmental conditions. Cell expansion is driven by turgor pressure created by osmotic water uptake and is restricted by the extensibility of the cell wall, which in turn is regulated by the synthesis, incorporation, and cross-linking of new cell wall components. The vacuolar H\+-ATPase (V-ATPase) could provide a way to coordinately regulate turgor pressure and cell wall synthesis, as it energizes the secondary active transport of solutes across the tonoplast and also has an important function in the trans-Golgi network (TGN), which affects synthesis and trafficking of cell wall components. We have previously shown that det3, a mutant with reduced V-ATPase activity, has a severe defect in cell expansion. However, it was not clear if this is caused by a defect in turgor pressure or in cell wall synthesis. Here, we show that inhibition of the tonoplast-localized V-ATPase subunit isoform VHA-a3 does not impair cell expansion. By contrast, inhibition of the TGN-localized isoform VHA-a1 is sufficient to restrict cell expansion. Furthermore, we provide evidence that the reduced hypocotyl cell expansion in det3 is conditional and due to active, hormone-mediated growth inhibition caused by a cell wall defect.} } @Article{IPB-1725, author = {Raffaele, S. and Vailleau, F. and Léger, A. and Joubès, J. and Miersch, O. and Huard, C. and Blée, E. and Mongrand, S. and Domergue, F. and Roby, D. and}, title = {{A MYB Transcription Factor Regulates Very-Long-Chain Fatty Acid Biosynthesis for Activation of the Hypersensitive Cell Death Response in Arabidopsis}}, year = {2008}, pages = {752-767}, journal = {Plant Cell}, doi = {10.1105/tpc.107.054858}, volume = {20}, abstract = {Plant immune responses to pathogen attack include the hypersensitive response (HR), a form of programmed cell death occurring at invasion sites. We previously reported on Arabidopsis thaliana MYB30, a transcription factor that acts as a positive regulator of a cell death pathway conditioning the HR. Here, we show by microarray analyses of Arabidopsis plants misexpressing MYB30 that the genes encoding the four enzymes forming the acyl-coA elongase complex are putative MYB30 targets. The acyl-coA elongase complex synthesizes very-long-chain fatty acids (VLCFAs), and the accumulation of extracellular VLCFA-derived metabolites (leaf epidermal wax components) was affected in MYB30 knockout mutant and overexpressing lines. In the same lines, a lipid extraction procedure allowing high recovery of sphingolipids revealed changes in VLCFA contents that were amplified in response to inoculation. Finally, the exacerbated HR phenotype of MYB30-overexpressing lines was altered by the loss of function of the acyl-ACP thioesterase FATB, which causes severe defects in the supply of fatty acids for VLCFA biosynthesis. Based on these findings, we propose a model in which MYB30 modulates HR via VLCFAs by themselves, or VLCFA derivatives, as cell death messengers in plants.} } @Article{IPB-1718, author = {Miersch, O. and Neumerkel, J. and Dippe, M. and Stenzel, I. and Wasternack, C. and}, title = {{Hydroxylated jasmonates are commonly occurring metabolites of jasmonic acid and contribute to a partial switch-off in jasmonate signaling}}, year = {2008}, pages = {114-127}, journal = {New Phytol.}, doi = {10.1111/j.1469-8137.2007.02252.x}, volume = {177}, abstract = {In potato 12‐hydroxyjasmonic acid (12‐OH‐JA) is a tuber‐inducing compound. Here, it is demonstrated that 12‐OH‐JA, as well as its sulfated and glucosylated derivatives, are constituents of various organs of many plant species. All accumulate differentially and usually to much higher concentrations than jasmonic acid (JA).In wounded tomato leaves, 12‐OH‐JA and its sulfated, as well as glucosylated, derivative accumulate after JA, and their diminished accumulation in wounded leaves of the JA‐deficient mutants spr2 and acx1 and also a JA‐deficient 35S::AOCantisense line suggest their JA‐dependent formation.To elucidate how signaling properties of JA/JAME (jasmonic acid methyl ester) are affected by hydroxylation and sulfation, germination and root growth were recorded in the presence of the different jasmonates, indicating that 12‐OH‐JA and 12‐hydroxyjasmonic acid sulfate (12‐HSO4‐JA) were not bioactive. Expression analyses for 29 genes showed that expression of wound‐inducible genes such as those coding for PROTEINASE INHIBITOR2, POLYPHENOL OXIDASE, THREONINE DEAMINASE or ARGINASE was induced by JAME and less induced or even down‐regulated by 12‐OH‐JA and 12‐HSO4‐JA. Almost all genes coding for enzymes in JA biosynthesis were up‐regulated by JAME but down‐regulated by 12‐OH‐JA and 12‐HSO4‐JA.The data suggest that wound‐induced metabolic conversion of JA/JAME into 12‐OH‐JA alters expression pattern of genes including a switch off in JA signaling for a subset of genes.} } @Article{IPB-1715, author = {Lange, P. R. and Geserick, C. and Tischendorf, G. and Zrenner, R. and}, title = {{Functions of Chloroplastic Adenylate Kinases in Arabidopsis}}, year = {2008}, pages = {492-504}, journal = {Plant Physiol.}, doi = {10.1104/pp.107.114702}, volume = {146}, abstract = {Adenosine monophosphate kinase (AMK; adenylate kinase) catalyses the reversible formation of ADP by the transfer of one phosphate group from ATP to AMP, thus equilibrating adenylates. The Arabidopsis (Arabidopsis thaliana) genome contains 10 genes with an adenylate/cytidylate kinase signature; seven of these are identified as putative adenylate kinases. Encoded proteins of at least two members of this Arabidopsis adenylate kinase gene family are targeted to plastids. However, when the individual genes are disrupted, the phenotypes of both mutants are strikingly different. Although absence of AMK2 causes only 30% reduction of total adenylate kinase activity in leaves, there is loss of chloroplast integrity leading to small, pale-looking plantlets from embryo to seedling development. In contrast, no phenotype for disruption of the second plastid adenylate kinase was found. From this analysis, we conclude that AMK2 is the major activity for equilibration of adenylates and de novo synthesis of ADP in the plastid stroma.} } @Article{IPB-1708, author = {Kienow, L. and Schneider, K. and Bartsch, M. and Stuible, H.-P. and Weng, H. and Miersch, O. and Wasternack, C. and Kombrink, E. and}, title = {{Jasmonates meet fatty acids: functional analysis of a new acyl-coenzyme A synthetase family from Arabidopsis thaliana}}, year = {2008}, pages = {403-419}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/erm325}, volume = {59}, abstract = {Arabidopsis thaliana contains a large number of genes encoding carboxylic acid-activating enzymes, including long-chain fatty acyl-CoA synthetase (LACS), 4-coumarate:CoA ligases (4CL), and proteins closely related to 4CLs with unknown activities. The function of these 4CL-like proteins was systematically explored by applying an extensive substrate screen, and it was uncovered that activation of fatty acids is the common feature of all active members of this protein family, thereby defining a new group of fatty acyl-CoA synthetase, which is distinct from the known LACS family. Significantly, four family members also displayed activity towards different biosynthetic precursors of jasmonic acid (JA), including 12-oxo-phytodienoic acid (OPDA), dinor-OPDA, 3-oxo-2(2′-[Z]-pentenyl)cyclopentane-1-octanoic acid (OPC-8), and OPC-6. Detailed analysis of in vitro properties uncovered significant differences in substrate specificity for individual enzymes, but only one protein (At1g20510) showed OPC-8:CoA ligase activity. Its in vivo function was analysed by transcript and jasmonate profiling of Arabidopsis insertion mutants for the gene. OPC-8:CoA ligase expression was activated in response to wounding or infection in the wild type but was undetectable in the mutants, which also exhibited OPC-8 accumulation and reduced levels of JA. In addition, the developmental, tissue- and cell-type specific expression pattern of the gene, and regulatory properties of its promoter were monitored by analysing promoter::GUS reporter lines. Collectively, the results demonstrate that OPC-8:CoA ligase catalyses an essential step in JA biosynthesis by initiating the β-oxidative chain shortening of the carboxylic acid side chain of its precursors, and, in accordance with this function, the protein is localized in peroxisomes.} } @Article{IPB-1705, author = {Jindaprasert, A. and Springob, K. and Schmidt, J. and De-Eknamkul, W. and Kutchan, T. M. and}, title = {{Pyrone polyketides synthesized by a type III polyketide synthase from Drosophyllum lusitanicum}}, year = {2008}, pages = {3043-3053}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2008.03.013}, volume = {69}, abstract = {To isolate cDNAs involved in the biosynthesis of acetate-derived naphthoquinones in Drosophyllum lusitanicum, an expressed sequence tag analysis was performed. RNA from callus cultures was used to create a cDNA library from which 2004 expressed sequence tags were generated. One cDNA with similarity to known type III polyketide synthases was isolated as full-length sequence and termed DluHKS. The translated polypeptide sequence of DluHKS showed 51–67% identity with other plant type III PKSs. Recombinant DluHKS expressed in Escherichia coli accepted acetyl-coenzyme A (CoA) as starter and carried out sequential decarboxylative condensations with malonyl-CoA yielding α-pyrones from three to six acetate units. However, naphthalenes, the expected products, were not isolated. Since the main compound produced by DluHKS is a hexaketide α-pyrone, and the naphthoquinones in D. lusitanicum are composed of six acetate units, we propose that the enzyme provides the backbone of these secondary metabolites. An involvement of accessory proteins in this biosynthetic pathway is discussed.} } @Article{IPB-1704, author = {Iglesias, N. G. and Gago-Zachert, S. P. and Robledo, G. and Costa, N. and Plata, M. I. and Vera, O. and Grau, O. and Semorile, L. C. and}, title = {{Population structure of Citrus tristeza virus from field Argentinean isolates}}, year = {2008}, pages = {199-207}, journal = {Virus Genes}, doi = {10.1007/s11262-007-0169-x}, volume = {36}, abstract = {We studied the genetic variability of three genomic regions (p23, p25 and p27 genes) from 11 field Citrus tristeza virus isolates from the two main citrus growing areas of Argentina, a country where the most efficient vector of the virus, Toxoptera citricida, is present for decades. The pathogenicity of the isolates was determinated by biological indexing, single-strand conformation polymorphism analysis showed that most isolates contained high intra-isolate variability. Divergent sequence variants were detected in some isolates, suggesting re-infections of the field plants. Phylogenetic analysis of the predominant sequence variants of each isolate revealed similar grouping of isolates for genes p25 and p27. The analysis of p23 showed two groups contained the severe isolates. Our results showed a high intra-isolate sequence variability suggesting that re-infections could contribute to the observed variability and that the host can play an important role in the selection of the sequence variants present in these isolates.} } @Article{IPB-1699, author = {Gao, X. and Stumpe, M. and Feussner, I. and Kolomiets, M. and}, title = {{A novel plastidial lipoxygenase of maize (Zea mays) ZmLOX6 encodes for a fatty acid hydroperoxide lyase and is uniquely regulated by phytohormones and pathogen infection}}, year = {2008}, pages = {491-503}, journal = {Planta}, doi = {10.1007/s00425-007-0634-8}, volume = {227}, abstract = {Lipoxygenases (LOXs) are members of a large enzyme family that catalyze oxygenation of free polyunsaturated fatty acids into diverse hydroperoxide compounds, collectively called oxylipins. Although LOXs have been well studied in dicot species, reports of the genes encoding these enzymes are scarce for monocots, especially maize. Herein, we reported the cloning, characterization and molecular functional analysis of a novel maize LOX gene, ZmLOX6. The ZmLOX6 nucleotide sequence encodes a deduced translation product of 892 amino acids. Phylogenetic analysis showed that ZmLOX6 is distantly related to previously reported 9- or 13-LOXs from maize and other plant species, including rice and Arabidopsis. Although sequence prediction suggested cytoplasmic localization of this protein, ZmLOX6 protein has been reportedly isolated from mesophyll cell chloroplasts, emphasizing the unique features of this protein. Plastidial localization was confirmed by chloroplast uptake experiments with the in vitro translated protein. Analysis of recombinant protein revealed that ZmLOX6 has lost fatty acid hydroperoxide forming activity but 13-LOX-derived fatty acid hydroperoxides were cleaved into odd-chain ω-oxo fatty acids and as yet not identified C5-compound. In line with its reported abundance in mesophyll cells, ZmLOX6 was predominantly expressed in leaf tissue. Northern blot analysis demonstrated that ZmLOX6 was induced by jasmonic acid, but repressed by abscisic acid, salicylic acid and ethylene and was not responsive to wounding or insects. Further, this gene was strongly induced by the fungal pathogen Cochliobolus carbonum during compatible interactions, suggesting that ZmLOX6 may contribute to susceptibility to this pathogen. The potential involvement of ZmLOX6 in maize interactions with pathogens is discussed.} } @Article{IPB-1761, author = {Abel, S. and}, title = {{Auxin Is Surfacing}}, year = {2007}, pages = {380-384}, journal = {ACS Chem. Biol.}, doi = {10.1021/cb7001158}, volume = {2}, abstract = {Indole-3-acetic acid (IAA or auxin) is essential throughout the life cycle of a plant. It controls diverse cellular processes, including gene expression. The hormone is perceived by a ubiquitin protein ligase (E3) and triggers the rapid destruction of repressors, called Aux/IAA proteins. The first structural model of a plant hormone receptor illustrates how auxin promotes Aux/IAA substrate recruitment by extending the hydrophobic protein-interaction surface. This work establishes a novel mechanism of E3 regulation by small molecules and promises a novel strategy for the treatment of human disorders associated with defective ubiquitin-dependent proteolysis.} } @Article{IPB-1791, author = {Guranowski, A. and Miersch, O. and Staswick, P. E. and Suza, W. and Wasternack, C. and}, title = {{Substrate specificity and products of side-reactions catalyzed by jasmonate:amino acid synthetase (JAR1)}}, year = {2007}, pages = {815-820}, journal = {FEBS Lett.}, doi = {10.1016/j.febslet.2007.01.049}, volume = {581}, abstract = {Jasmonate:amino acid synthetase (JAR1) is involved in the function of jasmonic acid (JA) as a plant hormone. It catalyzes the synthesis of several JA‐amido conjugates, the most important of which appears to be JA‐Ile. Structurally, JAR1 is a member of the firefly luciferase superfamily that comprises enzymes that adenylate various organic acids. This study analyzed the substrate specificity of recombinant JAR1 and determined whether it catalyzes the synthesis of mono‐ and dinucleoside polyphosphates, which are side‐reaction products of many enzymes forming acyl ∼ adenylates. Among different oxylipins tested as mixed stereoisomers for substrate activity with JAR1, the highest rate of conversion to Ile‐conjugates was observed for (±)‐JA and 9,10‐dihydro‐JA, while the rate of conjugation with 12‐hydroxy‐JA and OPC‐4 (3‐oxo‐2‐(2Z ‐pentenyl)cyclopentane‐1‐butyric acid) was only about 1–2% that for (±)‐JA. Of the two stereoisomers of JA, (−)‐JA and (\+)‐JA, rate of synthesis of the former was about 100‐fold faster than for (\+)‐JA. Finally, we have demonstrated that (1) in the presence of ATP, Mg2\+, (−)‐JA and tripolyphosphate the ligase produces adenosine 5′‐tetraphosphate (p4A); (2) addition of isoleucine to that mixture halts the p4A synthesis; (3) the enzyme produces neither diadenosine triphosphate (Ap3A) nor diadenosine tetraphosphate (Ap4A) and (4) Ap4A cannot substitute ATP as a source of adenylate in the complete reaction that yields JA‐Ile.} } @Article{IPB-1783, author = {Flores, R. and Navarro, B. and Gago, S. and De la Peña, M. and}, title = {{Chrysanthemum Chlorotic Mottle Viroid: a System for Reverse Genetics in the Family Avsunviroidae (Hammerhead Viroids)}}, year = {2007}, pages = {27-32}, journal = {Plant Viruses}, url = {http://www.globalsciencebooks.info/Online/GSBOnline/OnlinePV_1_1.html}, volume = {1}, abstract = {Viroids are small single-stranded circular RNAs able to infect plants. Chrysanthemum chlorotic mottle was one of the first viroid diseases reported, but identification and characterization of the causing RNA was delayed by its low accumulation in vivo. Chrysanthemum chlorotic mottle viroid (CChMVd) (398-401 nt) adopts a branched conformation instead of the rod-like secondary structure characteristic of most viroids. The natural sequence variability and the effects of artificial mutants support that the branched conformation is physiologically relevant and additionally stabilized by a kissing-loop interaction critical for RNA in vitro folding and in vivo viability. CChMVd shares structural similarities with peach latent mosaic viroid, with which forms the genus Pelamoviroid within the family Avsunviroidae. CChMVd adopts hammerhead structures that catalyze self-cleavage of the oligomeric strands of both polarities resulting from replication through a symmetric rolling-circle mechanism. The two CChMVd hammerheads display peculiarities: the plus has an extra A close to the central conserved core, and the minus an unsually long helix II. There are non-symptomatic strains (CChMVd-NS) that protect against challenge inoculation with severe strains (CChMVd-S). Introduction by site-directed mutagenesis of one of the CChMVd-NS specific mutations (UUUC?GAAA) is sufficient to change the symptomatic phenotype into non-symptomatic without altering the viroid titer. This pathogenicity determinant maps at a tetraloop of the CChMVd branched conformation. Co-inoculations with typical CChMVd-S and -NS variants showed that the infected plants remain symptomless only when the latter was in more than a 100-fold excess, indicating the higher fitness of the S variant. RNA silencing could mediate the observed cross-protection.} } @Article{IPB-1782, author = {Fester, T. and Lohse, S. and Halfmann, K. and}, title = {{“Chromoplast” development in arbuscular mycorrhizal roots}}, year = {2007}, pages = {92-100}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2006.09.034}, volume = {68}, abstract = {The accumulation of apocarotenoids in arbuscular mycorrhizal (AM) roots suggests a dramatic reorganization of the plastids responsible for the biosynthesis of these compounds. This review describes the cytological and biochemical characterization of this phenomenon. The results presented suggest that plastids are key organelles for the establishment of the symbiotic interface of the AM symbiosis. In addition, a complex interplay of various plant cell components during the different functional phases of this interface is suggested. Arbuscule degradation appears to be of particular interest, as it correlates with the formation of the most extensive plastid structures and with apocarotenoid accumulation.} } @Article{IPB-1780, author = {Eschen-Lippold, L. and Rothe, G. and Stumpe, M. and Göbel, C. and Feussner, I. and Rosahl, S. and}, title = {{Reduction of divinyl ether-containing polyunsaturated fatty acids in transgenic potato plants}}, year = {2007}, pages = {797-801}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2006.12.010}, volume = {68}, abstract = {Oxygenated polyunsaturated fatty acids synthesized via the lipoxygenase pathway play a role in plant responses to pathogen attack. In solanaceous plants, the preferential stimulation of the 9-lipoxygenase pathway in response to pathogen infection leads to the formation of the divinyl ether-containing polyunsaturated fatty acids colneleic and colnelenic acid, as well as hydroxy and trihydroxy polyunsaturated fatty acids. To functionally assess the role of divinyl ethers, transgenic potato plants were generated which express an RNA interference construct directed against the pathogen-inducible 9-divinyl ether synthase. Efficient reduction of 9-divinyl ether synthase transcript accumulation correlated with reduced levels of colneleic and colnelenic acid. However, in response to infection with virulent Phytophthora infestans, the causal agent of late blight disease, no significant differences in pathogen biomass could be detected suggesting that the levels of antimicrobial divinyl ethers are not critical for defense against Phytophthora infestans in a compatible interaction.} } @Article{IPB-1779, author = {Dorka, R. and Miersch, O. and Wasternack, C. and Weik, P. and}, title = {{Chronobiological phenomena and seasonal changes in jasmonate levels during the course of the year and under constant conditions in mistletoe (Viscum album L.)}}, year = {2007}, pages = {15}, journal = {Phytomedicine}, doi = {10.1016/j.phymed.2007.07.014}, volume = {14}, } @Article{IPB-1775, author = {Delker, C. and Zolman, B. K. and Miersch, O. and Wasternack, C. and}, title = {{Jasmonate biosynthesis in Arabidopsis thaliana requires peroxisomal β-oxidation enzymes – Additional proof by properties of pex6 and aim1}}, year = {2007}, pages = {1642-1650}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2007.04.024}, volume = {68}, abstract = {Jasmonic acid (JA) is an important regulator of plant development and stress responses. Several enzymes involved in the biosynthesis of JA from α-linolenic acid have been characterized. The final biosynthesis steps are the β-oxidation of 12-oxo-phytoenoic acid. We analyzed JA biosynthesis in the Arabidopsis mutants pex6, affected in peroxisome biogenesis, and aim1, disrupted in fatty acid β-oxidation. Upon wounding, these mutants exhibit reduced JA levels compared to wild type. pex6 accumulated the precursor OPDA. Feeding experiments with deuterated OPDA substantiate this accumulation pattern, suggesting the mutants are impaired in the β-oxidation of JA biosynthesis at different steps. Decreased expression of JA-responsive genes, such as VSP1, VSP2, AtJRG21 and LOX2, following wounding in the mutants compared to the wild type reflects the reduced JA levels of the mutants. By use of these additional mutants in combination with feeding experiments, the necessity of functional peroxisomes for JA-biosynthesis is confirmed. Furthermore an essential function of one of the two multifunctional proteins of fatty acid β-oxidation (AIM1) for wound-induced JA formation is demonstrated for the first time. These data confirm that JA biosynthesis occurs via peroxisomal fatty acid β-oxidation machinery.} } @Article{IPB-1771, author = {Calderon-Villalobos, L. I. A. and Nill, C. and Marrocco, K. and Kretsch, T. and Schwechheimer, C. and}, title = {{The evolutionarily conserved Arabidopsis thaliana F-box protein AtFBP7 is required for efficient translation during temperature stress}}, year = {2007}, pages = {106-116}, journal = {Gene}, doi = {10.1016/j.gene.2006.11.016}, volume = {392}, abstract = {In eukaryotes, E3 ubiquitin ligases (E3s) mediate the ubiquitylation of proteins that are destined for degradation by the ubiquitin–proteasome system. In SKP1/CDC53/F-box protein (SCF)-type E3 complexes, the interchangeable F-box protein confers specificity to the E3 ligase through direct physical interactions with the degradation substrate. The vast majority of the approximately 700 F-box proteins from the plant model organism Arabidopsis thaliana remain to be characterized. Here, we investigate the previously uncharacterized and evolutionarily conserved Arabidopsis F-box protein 7 (AtFBP7), which is encoded by a unique gene in Arabidopsis (At1g21760). Several apparent fbp7 loss-of-function alleles do not have an obvious phenotype. AtFBP7 is ubiquitously expressed and its expression is induced after cold and heat stress. When following up on a reported co-purification of the eukaryotic elongation factor-2 (eEF-2) with YLR097c, the apparent budding yeast orthologue of AtFBP7, we discovered a general defect in protein biosynthesis after cold and heat stress in fbp7 mutants. Thus, our findings suggest that AtFBP7 is required for protein synthesis during temperature stress.} } @Article{IPB-1766, author = {Biondi, E. and Branciamore, S. and Fusi, L. and Gago, S. and Gallori, E. and}, title = {{Catalytic activity of hammerhead ribozymes in a clay mineral environment: Implications for the RNA world}}, year = {2007}, pages = {10-18}, journal = {Gene}, doi = {10.1016/j.gene.2006.09.002}, volume = {389}, abstract = {The hypothesized RNA-based world would have required the presence of a protected environment in which RNA, or an RNA-like molecule, could originate and express its biological activity.Recent studies have indicated that RNA molecules adsorbed/bound on clay minerals are able to persist in the presence of degrading agents, to interact with surrounding molecules, and to transmit the information contained in their nucleotide sequences.In this study, we assessed the ability of RNA molecules with catalytic activity to perform a specific reaction in a mineral environment. For this purpose, we investigated the self-cleavage reaction of the hammerhead ribozyme of the Avocado Sun Blotch Viroid (ASBVd), both in the monomeric and in dimeric forms. The monomeric transcript was tightly bound on the clay mineral montmorillonite to form a stable complex, while the behaviour of the dimeric transcript was studied in the presence of the clay particles in the reaction mixture.The results indicated that the hammerhead ribozyme was still active when the monomeric transcript was adsorbed on the clay surface, even though its efficiency was reduced to about 20% of that in solution. Moreover, the self-cleavage of clay-adsorbed molecule was significantly enhanced (∼ four times) by the presence of the 5′ reaction product.The self-cleavage reaction of the dimeric transcript in the presence of montmorillonite indicated that the mineral particles protected the RNA molecules against aspecific degradation and increased the rate of cleavage kinetics by about one order of magnitude.These findings corroborate the hypothesis that clay-rich environments would have been a good habitat in which RNA or RNA-like molecules could originate, accumulate and undergo Darwinian evolutionary processes, leading to the first living cells on Earth.} } @Article{IPB-1839, author = {Wasternack, C. and}, title = {{Jasmonates: An Update on Biosynthesis, Signal Transduction and Action in Plant Stress Response, Growth and Development}}, year = {2007}, pages = {681-697}, journal = {Ann. Bot.}, doi = {10.1093/aob/mcm079}, volume = {100}, abstract = {BackgroundJasmonates are ubiquitously occurring lipid-derived compounds with signal functions in plant responses to abiotic and biotic stresses, as well as in plant growth and development. Jasmonic acid and its various metabolites are members of the oxylipin family. Many of them alter gene expression positively or negatively in a regulatory network with synergistic and antagonistic effects in relation to other plant hormones such as salicylate, auxin, ethylene and abscisic acid.ScopeThis review summarizes biosynthesis and signal transduction of jasmonates with emphasis on new findings in relation to enzymes, their crystal structure, new compounds detected in the oxylipin and jasmonate families, and newly found functions.ConclusionsCrystal structure of enzymes in jasmonate biosynthesis, increasing number of jasmonate metabolites and newly identified components of the jasmonate signal-transduction pathway, including specifically acting transcription factors, have led to new insights into jasmonate action, but its receptor(s) is/are still missing, in contrast to all other plant hormones.} } @Article{IPB-1838, author = {Wasternack, C. and}, title = {{Clarence A. “Bud” Ryan (29.09.1931–07.10.2007)}}, year = {2007}, pages = {709-709}, journal = {Plant Mol. Biol.}, doi = {10.1007/s11103-007-9254-8}, volume = {65}, } @Article{IPB-1836, author = {Vigliocco, A. and Alemano, S. and Miersch, O. and Alvarez, D. and Abdala, G. and}, title = {{Endogenous jasmonates in dry and imbibed sunflower seeds from plants grown at different soil moisture contents}}, year = {2007}, pages = {91-98}, journal = {Seed Sci. Res.}, doi = {10.1017/S0960258507708371}, volume = {17}, abstract = {In this study, we characterized two sunflower (Helianthus annuus L.) lines with differential sensitivity to drought, the sensitive line B59 and the tolerant line B71. Using both lines, we compared the content of endogenous jasmonates (JAs) in dry and imbibed seeds from plants grown under irrigation and drought. Jasmonic acid (JA), 12-oxo-phytodienoic acid (OPDA), 11-hydroxyjasmonate (11-OH-JA) and 12-hydroxyjasmonate (12-OH-JA) were detected in dry and imbibed sunflower seeds. Seeds from plants grown under drought had a lower content of total JAs and exhibited higher germination percentages than seeds from irrigated plants, demonstrating that environmental conditions have a strong influence on the progeny. OPDA and 12-OH-JA were the main compounds found in dry seeds of both lines. Imbibed seeds showed an enhanced amount of total JAs with respect to dry seeds produced by plants grown in both soil moisture conditions. Imbibition triggered a dramatic OPDA increase in the embryo, suggesting a role of this compound in germination. We conclude that JAs patterns vary during sunflower germination and that the environmental conditions experienced by the mother plant modify the hormonal content of the seed progeny.} } @Article{IPB-1829, author = {ten Hoopen, P. and Hunger, A. and Muller, A. and Hause, B. and Kramell, R. and Wasternack, C. and Rosahl, S. and Conrad, U. and}, title = {{Immunomodulation of jasmonate to manipulate the wound response}}, year = {2007}, pages = {2525-2535}, journal = {J. Exp. Bot.}, doi = {10.1093/jxb/erm122}, volume = {58}, abstract = {Jasmonates are signals in plant stress responses and development. The exact mode of their action is still controversial. To modulate jasmonate levels intracellularly as well as compartment-specifically, transgenic Nicotiana tabacum plants expressing single-chain antibodies selected against the naturally occurring (3R,7R)-enantiomer of jasmonic acid (JA) were created in the cytosol and the endoplasmic reticulum. Consequently, the expression of anti-JA antibodies in planta caused JA-deficient phenotypes such as insensitivity of germinating transgenic seedlings towards methyl jasmonate and the loss of wound-induced gene expression. Results presented here suggest an essential role for cytosolic JA in the wound response of tobacco plants. The findings support the view that substrate availability takes part in regulating JA biosynthesis upon wounding. Moreover, high JA levels observed in immunomodulated plants in response to wounding suggest that tobacco plants are able to perceive a reduced level of physiologically active JA and attempt to compensate for this by increased JA accumulation.} } @Article{IPB-1827, author = {Tan, X. and Calderon-Villalobos, L. I. A. and Sharon, M. and Zheng, C. and Robinson, C. V. and Estelle, M. and Zheng, N. and}, title = {{Mechanism of auxin perception by the TIR1 ubiquitin ligase}}, year = {2007}, pages = {640-645}, journal = {Nature}, doi = {10.1038/nature05731}, volume = {446}, abstract = {Auxin is a pivotal plant hormone that controls many aspects of plant growth and development. Perceived by a small family of F-box proteins including transport inhibitor response 1 (TIR1), auxin regulates gene expression by promoting SCF ubiquitin-ligase-catalysed degradation of the Aux/IAA transcription repressors, but how the TIR1 F-box protein senses and becomes activated by auxin remains unclear. Here we present the crystal structures of the Arabidopsis TIR1–ASK1 complex, free and in complexes with three different auxin compounds and an Aux/IAA substrate peptide. These structures show that the leucine-rich repeat domain of TIR1 contains an unexpected inositol hexakisphosphate co-factor and recognizes auxin and the Aux/IAA polypeptide substrate through a single surface pocket. Anchored to the base of the TIR1 pocket, auxin binds to a partially promiscuous site, which can also accommodate various auxin analogues. Docked on top of auxin, the Aux/IAA substrate peptide occupies the rest of the TIR1 pocket and completely encloses the hormone-binding site. By filling in a hydrophobic cavity at the protein interface, auxin enhances the TIR1–substrate interactions by acting as a ‘molecular glue’. Our results establish the first structural model of a plant hormone receptor.} } @Article{IPB-1823, author = {Schwager, K. M. and Calderon-Villalobos, L. I. A. and Dohmann, E. M. and Willige, B. C. and Knierer, S. and Nill, C. and Schwechheimer, C. and}, title = {{Characterization of the VIER F-BOX PROTEINE Genes from Arabidopsis Reveals Their Importance for Plant Growth and Development}}, year = {2007}, pages = {1163-1178}, journal = {Plant Cell}, doi = {10.1105/tpc.105.040675}, volume = {19}, abstract = {E3 ubiquitin ligases (E3s) target proteins for degradation by the 26S proteasome. In SKP1/CDC53/F-box protein–type E3s, substrate specificity is conferred by the interchangeable F-box protein subunit. The vast majority of the 694 F-box proteins encoded by the Arabidopsis thaliana genome remain to be understood. We characterize the VIER F-BOX PROTEINE (VFB; German for FOUR F-BOX PROTEINS) genes from Arabidopsis that belong to subfamily C of the Arabidopsis F-box protein superfamily. This subfamily also includes the F-box proteins TRANSPORT INHIBITOR RESPONSE1 (TIR1)/AUXIN SIGNALING F-BOX (AFB) proteins and EIN3 BINDING F-BOX proteins, which regulate auxin and ethylene responses, respectively. We show that loss of VFB function causes delayed plant growth and reduced lateral root formation. We find that the expression of a number of auxin-responsive genes and the activity of DR5:β-glucuronidase, a reporter for auxin reponse, are reduced in the vfb mutants. This finding correlates with an increase in the abundance of an AUXIN/INDOLE-3-ACETIC ACID repressor. However, we also find that auxin responses are not affected in the vfb mutants and that a representative VFB family member, VFB2, cannot functionally complement the tir1-1 mutant. We therefore exclude the possibility that VFBs are functional orthologs of TIR1/AFB proteins.} } @Article{IPB-1820, author = {Schilling, S. and Stenzel, I. and von Bohlen, A. and Wermann, M. and Schulz, K. and Demuth, H.-U. and Wasternack, C. and}, title = {{Isolation and characterization of the glutaminyl cyclases from Solanum tuberosum and Arabidopsis thaliana: implications for physiological functions}}, year = {2007}, pages = {145-153}, journal = {Biol. Chem.}, doi = {10.1515/BC.2007.016}, volume = {388}, abstract = {Glutaminyl cyclases (QCs) catalyze the formation of pyroglutamic acid at the N-terminus of several peptides and proteins. On the basis of the amino acid sequence of Carica papaya QC, we identified cDNAs of the putative counterparts from Solanum tuberosum and Arabidopsis thaliana. Upon expression of the corresponding cDNAs from both plants via the secretory pathway of Pichia pastoris, two active QC proteins were isolated. The specificity of the purified proteins was assessed using various substrates with different amino acid composition and length. Highest specificities were observed with substrates possessing large hydrophobic residues adjacent to the N-terminal glutamine and for fluorogenic dipeptide surrogates. However, compared to Carica papaya QC, the specificity constants were approximately one order of magnitude lower for most of the QC substrates analyzed. The QCs also catalyzed the conversion of N-terminal glutamic acid to pyroglutamic acid, but with approximately 105- to 106-fold lower specificity. The ubiquitous distribution of plant QCs prompted a search for potential substrates in plants. Based on database entries, numerous proteins, e.g., pathogenesis-related proteins, were found that carry a pyroglutamate residue at the N-terminus, suggesting QC involvement. The putative relevance of QCs and pyroglutamic acid for plant defense reactions is discussed.} } @Article{IPB-1812, author = {Pedranzani, H. and Sierra-de-Grado, R. and Vigliocco, A. and Miersch, O. and Abdala, G. and}, title = {{Cold and water stresses produce changes in endogenous jasmonates in two populations of Pinus pinaster Ait}}, year = {2007}, pages = {111-116}, journal = {Plant Growth Regul.}, doi = {10.1007/s10725-007-9166-2}, volume = {52}, abstract = {There is considerable evidence suggesting that jasmonates (JAs) play a role in plant resistance against abiotic stress. It is well known that in Angiosperms JAs are involved in the defense response, however there is little information about their role in Gymnosperms. Our proposal was to study the involvement of JAs in Pinus pinaster Ait. reaction to cold and water stress, and to compare the response of two populations of different provenances (Gredos and Bajo Tiétar) to these stresses. We detected 12-oxo-phytodienoic acid (OPDA), jasmonic acid (JA), and the hydroxylates 11-hydroxyjasmonate and 12-hydroxyjasmonate in foliage and shoots of P. pinaster plants. The response of the Gredos population to cold stress differed from that of Bajo Tiétar. Gredos plants showed a lower JA-basal level than Bajo Tiétar; under cold stress JA increased twofold at 72 h, while it decreased in Bajo Tiétar plants. The hydroxylates slightly increased in both populations due to cold stress treatment. Under water stress, plants from Gredos showed a remarkable JA-increase; thus the JA-response was much more prominent under water stress than under cold stress. In contrast, no change was found in JA-level in Bajo Tiétar plants under water stress. The level of JA-precursor, OPDA, was very low in control plants from Gredos and Bajo Tiétar. Under water stress OPDA increased only in plants from Bajo Tiétar. Therefore, we inform here of a different JAs-accumulation pattern after the stress treatment in P. pinaster from two provenances, and suggest a possible correlation with adaptations to diverse ecological conditions.} } @Article{IPB-1800, author = {Lannoo, N. and Vandenborre, G. and Miersch, O. and Smagghe, G. and Wasternack, C. and Peumans, W. J. and Van Damme, E. J. M. and}, title = {{The Jasmonate-Induced Expression of the Nicotiana tabacum Leaf Lectin}}, year = {2007}, pages = {1207-1218}, journal = {Plant Cell Physiol.}, doi = {10.1093/pcp/pcm090}, volume = {48}, abstract = {Previous experiments with tobacco (Nicotiana tabacum L. cv Samsun NN) plants revealed that jasmonic acid methyl ester (JAME) induces the expression of a cytoplasmic/nuclear lectin in leaf cells and provided the first evidence that jasmonates affect the expression of carbohydrate-binding proteins in plant cells. To corroborate the induced accumulation of relatively large amounts of a cytoplasmic/nuclear lectin, a detailed study was performed on the induction of the lectin in both intact tobacco plants and excised leaves. Experiments with different stress factors demonstrated that the lectin is exclusively induced by exogeneously applied jasmonic acid and JAME, and to a lesser extent by insect herbivory. The lectin concentration depends on leaf age and the position of the tissue in the leaf. JAME acts systemically in intact plants but very locally in excised leaves. Kinetic analyses indicated that the lectin is synthesized within 12 h exposure time to JAME, reaching a maximum after 60 h. After removal of JAME, the lectin progressively disappears from the leaf tissue. The JAME-induced accumulation of an abundant nuclear/cytoplasmic lectin is discussed in view of the possible role of this lectin in the plant.} } @Article{IPB-1927, author = {Ziegler, J. and Voigtländer, S. and Schmidt, J. and Kramell, R. and Miersch, O. and Ammer, C. and Gesell, A. and Kutchan, T. M. and}, title = {{Comparative transcript and alkaloid profiling in Papaver species identifies a short chain dehydrogenase/reductase involved in morphine biosynthesis}}, year = {2006}, pages = {177-192}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2006.02860.x}, volume = {48}, abstract = {Plants of the order Ranunculales, especially members of the species Papaver , accumulate a large variety of benzylisoquinoline alkaloids with about 2500 structures, but only the opium poppy (Papaver somniferum ) and Papaver setigerum are able to produce the analgesic and narcotic morphine and the antitussive codeine. In this study, we investigated the molecular basis for this exceptional biosynthetic capability by comparison of alkaloid profiles with gene expression profiles between 16 different Papaver species. Out of 2000 expressed sequence tags obtained from P. somniferum , 69 show increased expression in morphinan alkaloid‐containing species. One of these cDNAs, exhibiting an expression pattern very similar to previously isolated cDNAs coding for enzymes in benzylisoquinoline biosynthesis, showed the highest amino acid identity to reductases in menthol biosynthesis. After overexpression, the protein encoded by this cDNA reduced the keto group of salutaridine yielding salutaridinol, an intermediate in morphine biosynthesis. The stereoisomer 7‐epi ‐salutaridinol was not formed. Based on its similarities to a previously purified protein from P. somniferum with respect to the high substrate specificity, molecular mass and kinetic data, the recombinant protein was identified as salutaridine reductase (SalR; EC 1.1.1.248). Unlike codeinone reductase, an enzyme acting later in the pathway that catalyses the reduction of a keto group and which belongs to the family of the aldo‐keto reductases, the cDNA identified in this study as SalR belongs to the family of short chain dehydrogenases/reductases and is related to reductases in monoterpene metabolism.} } @Article{IPB-1918, author = {Wasternack, C. and Stenzel, I. and Hause, B. and Hause, G. and Kutter, C. and Maucher, H. and Neumerkel, J. and Feussner, I. and Miersch, O. and}, title = {{The wound response in tomato – Role of jasmonic acid}}, year = {2006}, pages = {297-306}, journal = {J. Plant Physiol.}, doi = {10.1016/j.jplph.2005.10.014}, volume = {163}, abstract = {Plants respond to mechanical wounding or herbivore attack with a complex scenario of sequential, antagonistic or synergistic action of different signals leading to defense gene expression. Tomato plants were used as a model system since the peptide systemin and the lipid-derived jasmonic acid (JA) were recognized as essential signals in wound-induced gene expression. In this review recent data are discussed with emphasis on wound-signaling in tomato. The following aspects are covered: (i) systemin signaling, (ii) JA biosynthesis and action, (iii) orchestration of various signals such as JA, H2O2, NO, and salicylate, (iv) local and systemic response, and (v) amplification in wound signaling. The common occurrence of JA biosynthesis and systemin generation in the vascular bundles suggest JA as the systemic signal. Grafting experiments with JA-deficient, JA-insensitive and systemin-insensitive mutants strongly support this assumption.} } @Article{IPB-1917, author = {Wasternack, C. and}, title = {{Jasmonates - Biosynthesis, Signal Transduction and Action (Annals of Botany Lecture)}}, year = {2006}, pages = {11}, journal = {Jap. Soc. Chem. Regul Plants, Abstr.}, doi = {10.18978/jscrpanb.41.0_11}, volume = {41}, } @Article{IPB-1906, author = {Sreenivasulu, N. and Radchuk, V. and Strickert, M. and Miersch, O. and Weschke, W. and Wobus, U. and}, title = {{Gene expression patterns reveal tissue-specific signaling networks controlling programmed cell death and ABA- regulated maturation in developing barley seeds}}, year = {2006}, pages = {310-327}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2006.02789.x}, volume = {47}, abstract = {Gene expression patterns covering over 10 000 seed‐expressed sequences were analyzed by macroarray technology in maternal tissue (mainly pericarp) and filial endosperm and embryo during barley seed development from anthesis until late maturation. Defined sets of genes showing distinct expression patterns characterized both tissue type and major developmental phases. The analysis focused on regulatory networks involved in programmed cell death (PCD) and abscisic acid (ABA)‐mediated maturation. These processes were similar in the different tissues, but typically involved the expression of alternative members of a common gene family. The analysis of co‐expressed gene sets and the identification of cis regulatory elements in orthologous rice gene ‘promoter’ regions suggest that PCD in the pericarp is mediated by distinct classes of proteases and is under the hormonal control of both jasmonic acid (JA) and ethylene via ethylene‐responsive element binding protein (EREBP) transcription factors (TFs). On the other hand, PCD in endosperm apparently involves only the ethylene pathway, but employs distinct gene family members from those active in the pericarp, and a different set of proteases and TFs. JA biosynthetic genes are hardly activated. Accordingly, JA levels are high in the pericarp but low in the endosperm during middle and late developmental stages. Similarly, genes acting in the deduced ABA biosynthetic pathway and signaling network differ between endosperm and embryo. ABA in the endosperm appears to exert an influence over storage product synthesis via SNF1 kinase. In the embryo, ABA seems to influence the acquisition of desiccation tolerance via ABA response element binding factors, but the data also suggest the existence of an ABA‐independent but interactive pathway acting via the dehydration‐responsive element binding (DREB) 2A TF.} } @Article{IPB-1903, author = {Sharma, V. K. and Monostori, T. and Göbel, C. and Hänsch, R. and Bittner, F. and Wasternack, C. and Feussner, I. and Mendel, R. R. and Hause, B. and Schulze, J. and}, title = {{Transgenic barley plants overexpressing a 13-lipoxygenase to modify oxylipin signature}}, year = {2006}, pages = {264-276}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2005.11.009}, volume = {67}, abstract = {Three chimeric gene constructs were designed comprising the full length cDNA of a lipoxygenase (LOX) from barley (LOX2:Hv:1) including its chloroplast targeting sequence (cTP) under control of either (1) CaMV35S- or (2) polyubiquitin-1-promoter, whereas the third plasmid contains 35S promoter and the cDNA without cTP. Transgenic barley plants overexpressing LOX2:Hv:1 were generated by biolistics of scutella from immature embryos. Transformation frequency for 35S::LOX with or without cTP was in a range known for barley particle bombardment, whereas for Ubi::cTP-LOX no transgenic plants were detected. In general, a high number of green plantlets selected on bialaphos became yellow and finally died either in vitro or after potting. All transgenic plants obtained were phenotypically indistinguishable from wild type plants and all of them set seeds. The corresponding protein (LOX-100) in transgenic T0 and T1 plants accumulated constitutively to similar levels as in the jasmonic acid methyl ester (JAME)-treated wild type plants. Moreover, LOX-100 was clearly detectable immunocytochemically within the chloroplasts of untreated T0 plants containing the LOX-100-cDNA with the chloroplast target sequence. In contrast, an exclusive localization of LOX-100 in the cytoplasm was detectable when the target sequence was removed. In comparison to sorbitol-treated wild type leaves, analysis of oxylipin profiles in T2 progenies showed higher levels of jasmonic acid (JA) for those lines that displayed elevated levels of LOX-100 in the chloroplasts and for those lines that harboured LOX-100 in the cytoplasm, respectively. The studies demonstrate for the first time the constitutive overexpression of a cDNA coding for a 13-LOX in a monocotyledonous species and indicate a link between the occurrence of LOX-100 and senescence.} } @Article{IPB-1891, author = {Quint, M. and Gray, W. M. and}, title = {{Auxin signaling}}, year = {2006}, pages = {448-453}, journal = {Curr. Opin. Plant Biol.}, doi = {10.1016/j.pbi.2006.07.006}, volume = {9}, abstract = {Auxin regulates a host of plant developmental and physiological processes, including embryogenesis, vascular differentiation, organogenesis, tropic growth, and root and shoot architecture. Genetic and biochemical studies carried out over the past decade have revealed that much of this regulation involves the SCFTIR1/AFB-mediated proteolysis of the Aux/IAA family of transcriptional regulators. With the recent finding that the TRANSPORT INHIBITOR RESPONSE1 (TIR1)/AUXIN SIGNALING F-BOX (AFB) proteins also function as auxin receptors, a potentially complete, and surprisingly simple, signaling pathway from perception to transcriptional response is now before us. However, understanding how this seemingly simple pathway controls the myriad of specific auxin responses remains a daunting challenge, and compelling evidence exists for SCFTIR1/AFB-independent auxin signaling pathways.} } @Article{IPB-1888, author = {Mur, L. A. and Kenton, P. and Atzorn, R. and Miersch, O. and Wasternack, C. and}, title = {{The Outcomes of Concentration-Specific Interactions between Salicylate and Jasmonate Signaling Include Synergy, Antagonism, and Oxidative Stress Leading to Cell Death}}, year = {2006}, pages = {249-262}, journal = {Plant Physiol.}, doi = {10.1104/pp.105.072348}, volume = {140}, abstract = {Salicylic acid (SA) has been proposed to antagonize jasmonic acid (JA) biosynthesis and signaling. We report, however, that in salicylate hydroxylase-expressing tobacco (Nicotiana tabacum) plants, where SA levels were reduced, JA levels were not elevated during a hypersensitive response elicited by Pseudomonas syringae pv phaseolicola. The effects of cotreatment with various concentrations of SA and JA were assessed in tobacco and Arabidopsis (Arabidopsis thaliana). These suggested that there was a transient synergistic enhancement in the expression of genes associated with either JA (PDF1.2 [defensin] and Thi1.2 [thionin]) or SA (PR1 [PR1a-β-glucuronidase in tobacco]) signaling when both signals were applied at low (typically 10–100 μm) concentrations. Antagonism was observed at more prolonged treatment times or at higher concentrations. Similar results were also observed when adding the JA precursor, α-linolenic acid with SA. Synergic effects on gene expression and plant stress were NPR1- and COI1-dependent, SA- and JA-signaling components, respectively. Electrolyte leakage and Evans blue staining indicated that application of higher concentrations of SA \+ JA induced plant stress or death and elicited the generation of apoplastic reactive oxygen species. This was indicated by enhancement of hydrogen peroxide-responsive AoPR10-β-glucuronidase expression, suppression of plant stress/death using catalase, and direct hydrogen peroxide measurements. Our data suggests that the outcomes of JA-SA interactions could be tailored to pathogen/pest attack by the relative concentration of each hormone.} } @Article{IPB-1869, author = {Grubb, C. D. and Abel, S. and}, title = {{Glucosinolate metabolism and its control}}, year = {2006}, pages = {89-100}, journal = {Trends Plant Sci.}, doi = {10.1016/j.tplants.2005.12.006}, volume = {11}, abstract = {Glucosinolates and their associated degradation products have long been recognized for their distinctive benefits to human nutrition and plant defense. Because most of the structural genes of glucosinolate metabolism have been identified and functionally characterized in Arabidopsis thaliana, current research increasingly focuses on questions related to the regulation of glucosinolate synthesis, distribution and degradation as well as to the feasibility of engineering customized glucosinolate profiles. Here, we highlight recent progress in glucosinolate research, with particular emphasis on the biosynthetic pathway and its metabolic relationships to auxin homeostasis. We further discuss emerging insight into the signaling networks and regulatory proteins that control glucosinolate accumulation during plant development and in response to environmental challenge.} } @Article{IPB-1863, author = {Ederli, L. and Morettini, R. and Borgogni, A. and Wasternack, C. and Miersch, O. and Reale, L. and Ferranti, F. and Tosti, N. and Pasqualini, S. and}, title = {{Interaction between Nitric Oxide and Ethylene in the Induction of Alternative Oxidase in Ozone-Treated Tobacco Plants}}, year = {2006}, pages = {595-608}, journal = {Plant Physiol.}, doi = {10.1104/pp.106.085472}, volume = {142}, abstract = {The higher plant mitochondrial electron transport chain contains, in addition to the cytochrome chain, an alternative pathway that terminates with a single homodimeric protein, the alternative oxidase (AOX). We recorded temporary inhibition of cytochrome capacity respiration and activation of AOX pathway capacity in tobacco plants (Nicotiana tabacum L. cv BelW3) fumigated with ozone (O3). The AOX1a gene was used as a molecular probe to investigate its regulation by signal molecules such as hydrogen peroxide, nitric oxide (NO), ethylene (ET), salicylic acid, and jasmonic acid (JA), all of them reported to be involved in the O3 response. Fumigation leads to accumulation of hydrogen peroxide in mitochondria and early accumulation of NO in leaf tissues. Although ET accumulation was high in leaf tissues 5 h after the start of O3 fumigation, it declined during the recovery period. There were no differences in the JA and 12-oxo-phytodienoic acid levels of treated and untreated plants. NO, JA, and ET induced AOX1a mRNA accumulation. Using pharmacological inhibition of ET and NO, we demonstrate that both NO- and ET-dependent pathways are required for O3-induced up-regulation of AOX1a. However, only NO is indispensable for the activation of AOX1a gene expression.} } @Article{IPB-1860, author = {Delker, C. and Stenzel, I. and Hause, B. and Miersch, O. and Feussner, I. and Wasternack, C. and}, title = {{Jasmonate Biosynthesis in Arabidopsis thaliana - Enzymes, Products, Regulation}}, year = {2006}, pages = {297-306}, journal = {Plant Biol.}, doi = {10.1055/s-2006-923935}, volume = {8}, abstract = {Among the plant hormones jasmonic acid and related derivatives are known to mediate stress responses and several developmental processes. Biosynthesis, regulation, and metabolism of jasmonic acid in Arabidopsis thaliana are reviewed, including properties of mutants of jasmonate biosynthesis. The individual signalling properties of several jasmonates are described.} } @Article{IPB-1859, author = {De Nardi, B. and Dreos, R. and Del Terra, L. and Martellossi, C. and Asquini, E. and Tornincasa, P. and Gasperini, D. and Pacchioni, B. and Rathinavelu, R. and Pallavicini, A. and Graziosi, G. and}, title = {{Differential responses of Coffea arabica L. leaves and roots to chemically induced systemic acquired resistance}}, year = {2006}, pages = {1594-1605}, journal = {Genome}, doi = {10.1139/g06-125}, volume = {49}, abstract = {Coffea arabica is susceptible to several pests and diseases, some of which affect the leaves and roots. Systemic acquired resistance (SAR) is the main defence mechanism activated in plants in response to pathogen attack. Here, we report the effects of benzo(1,2,3)thiadiazole-7-carbothioic acid-s-methyl ester (BTH), a SAR chemical inducer, on the expression profile of C. arabica. Two cDNA libraries were constructed from the mRNA isolated from leaves and embryonic roots to create 1587 nonredundant expressed sequence tags (ESTs). We developed a cDNA microarray containing 1506 ESTs from the leaves and embryonic roots, and 48 NBS-LRR (nucleotide-binding site leucine-rich repeat) gene fragments derived from 2 specific genomic libraries. Competitive hybridization between untreated and BTH-treated leaves resulted in 55 genes that were significantly overexpressed and 16 genes that were significantly underexpressed. In the roots, 37 and 42 genes were over and underexpressed, respectively. A general shift in metabolism from housekeeping to defence occurred in the leaves and roots after BTH treatment. We observed a systemic increase in pathogenesis-related protein synthesis, in the oxidative burst, and in the cell wall strengthening processes. Moreover, responses in the roots and leaves varied significantly.} } @Article{IPB-1853, author = {Carbonell, A. and De la Peña, M. and Flores, R. and Gago, S. and}, title = {{Effects of the trinucleotide preceding the self-cleavage site on eggplant latent viroid hammerheads: differences in co- and post-transcriptional self-cleavage may explain the lack of trinucleotide AUC in most natural hammerheads}}, year = {2006}, pages = {5613-5622}, journal = {Nucleic Acids Res.}, doi = {10.1093/nar/gkl717}, volume = {34}, abstract = {Eggplant latent viroid (ELVd) can form stable hammerhead structures in its (\+) and (−) strands. These ribozymes have the longest helices I reported in natural hammerheads, with that of the ELVd (\+) hammerhead being particularly stable (5/7 bp are G-C). Moreover, the trinucleotide preceding the self-cleavage site of this hammerhead is AUA, which together with GUA also found in some natural hammerheads, deviate from the GUC present in most natural hammerheads including the ELVd (−) hammerhead. When the AUA trinucleotide preceding the self-cleavage site of the ELVd (\+) hammerhead was substituted by GUA and GUC, as well as by AUC (essentially absent in natural hammerheads), the values of the self-cleavage rate constants at low magnesium of the purified hammerheads were: ELVd-(\+)-AUC≈ELVd-(\+)-GUC\>ELVd-(\+)-GUA\> ELVd-(\+)-AUA. However, the ELVd-(\+)-AUC hammerhead was the catalytically less efficient during in vitro transcription, most likely because of the transient adoption of catalytically-inactive metastable structures. These results suggest that natural hammerheads have been evolutionary selected to function co-transcriptionally, and provide a model explaining the lack of trinucleotide AUC preceding the self-cleavage site of most natural hammerheads. Comparisons with other natural hammerheads showed that the ELVd-(\+)-GUC and ELVd-(\+)-AUC hammerheads are the catalytically most active in a post-transcriptional context with low magnesium.} } @Article{IPB-1852, author = {Calderon-Villalobos, L. I. A. and Kuhnle, C. and Li, H. and Rosso, M. and Weisshaar, B. and Schwechheimer, C. and}, title = {{LucTrap Vectors Are Tools to Generate Luciferase Fusions for the Quantification of Transcript and Protein Abundance in Vivo}}, year = {2006}, pages = {3-14}, journal = {Plant Physiol.}, doi = {10.1104/pp.106.078097}, volume = {141}, abstract = {Proper plant growth and development strongly rely on the plant\'s ability to respond dynamically to signals and cues from the intra- and extracellular environment. Whereas many of these responses require specific changes at the level of gene expression, in recent years it has become increasingly clear that many plant responses are at least in part also controlled at the level of protein turnover. It is a challenge for signal transduction research to understand how distinct incoming signals are integrated to generate specific changes at the transcript or protein level. The activity of luciferase (LUC) reporters can be detected in nondestructive qualitative and quantitative assays in vivo. Therefore,z LUC reporters are particularly well suited for the detection of changes at the transcript and protein level. To the best of our knowledge, the number of plant transformation vectors for LUC fusions is very limited. In this article, we describe the LucTrap plant transformation vectors that allow generation of targeted and random transcriptional and translational fusions with the modified firefly LUC reporter LUC\+. We demonstrate that LucTrap-based fusions can be used to monitor rapid changes in gene expression and protein abundance in vivo.} } @Article{IPB-1970, author = {Ludwig, A. A. and Saitoh, H. and Felix, G. and Freymark, G. and Miersch, O. and Wasternack, C. and Boller, T. and Jones, J. D. G. and Romeis, T. and}, title = {{Ethylene-mediated cross-talk between calcium-dependent protein kinase and MAPK signaling controls stress responses in plants}}, year = {2005}, pages = {10736-10741}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.0502954102}, volume = {102}, abstract = {Plants are constantly exposed to environmental changes and need to integrate multiple external stress cues. Calcium-dependent protein kinases (CDPKs) are implicated as major primary Ca2\+ sensors in plants. CDPK activation, like activation of mitogen-activated protein kinases (MAPKs), is triggered by biotic and abiotic stresses, although distinct stimulus-specific stress responses are induced. To investigate whether CDPKs are part of an underlying mechanism to guarantee response specificity, we identified CDPK-controlled signaling pathways. A truncated form of Nicotiana tabacum CDPK2 lacking its regulatory autoinhibitor and calcium-binding domains was ectopically expressed in Nicotiana benthamiana. Infiltrated leaves responded to an abiotic stress stimulus with the activation of biotic stress reactions. These responses included synthesis of reactive oxygen species, defense gene induction, and SGT1-dependent cell death. Furthermore, N-terminal CDPK2 signaling triggered enhanced levels of the phytohormones jasmonic acid, 12-oxo-phytodienoic acid, and ethylene but not salicylic acid. These responses, commonly only observed after challenge with a strong biotic stimulus, were prevented when the CDPK\'s intrinsic autoinhibitory peptide was coexpressed. Remarkably, elevated CDPK signaling compromised stress-induced MAPK activation, and this inhibition required ethylene synthesis and perception. These data indicate that CDPK and MAPK pathways do not function independently and that a concerted activation of both pathways controls response specificity to biotic and abiotic stress.} } @Article{IPB-1965, author = {Levy, M. and Rachmilevitch, S. and Abel, S. and}, title = {{Transient Agrobacterium-mediated gene expression in the Arabidopsis hydroponics root system for subcellular localization studies}}, year = {2005}, pages = {179-184}, journal = {Plant Mol. Biol. Rep.}, doi = {10.1007/BF02772708}, volume = {23}, abstract = {To a great extent, the cellular compartmentalization and molecular interactions are indicative of the function of a protein. The development of simple and efficient tools for testing the subcellular location of proteins is indispensable to elucidate the function of genes in plants. In this report, we assessed the feasibility ofAgrobacterium-mediated transformation of hydroponically grown roots to follow intracellular targeting of proteins fused to green fluorescent protein (GFP). We developed a simple in planta assay for subcellular localization of proteins inArabidopsis roots via transient transformation and tested this method by expressing a GFP fusion of a known nuclear protein, IQD1. Visualization of transiently expressed GFP fusion proteins in roots by means of confocal microscopy is superior to the analysis of green tissues because the roots are virtually transparent and free of chlorophyll autofluorescence.} } @Article{IPB-1964, author = {Levy, M. and Wang, Q. and Kaspi, R. and Parrella, M. P. and Abel, S. and}, title = {{Arabidopsis IQD1, a novel calmodulin-binding nuclear protein, stimulates glucosinolate accumulation and plant defense}}, year = {2005}, pages = {79-96}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2005.02435.x}, volume = {43}, abstract = {Glucosinolates are a class of secondary metabolites with important roles in plant defense and human nutrition. To uncover regulatory mechanisms of glucosinolate production, we screened Arabidopsis thaliana T‐DNA activation‐tagged lines and identified a high‐glucosinolate mutant caused by overexpression of IQD1 (At3g09710). A series of gain‐ and loss‐of‐function IQD1 alleles in different accessions correlates with increased and decreased glucosinolate levels, respectively. IQD1 encodes a novel protein that contains putative nuclear localization signals and several motifs known to mediate calmodulin binding, which are arranged in a plant‐specific segment of 67 amino acids, called the IQ67 domain. We demonstrate that an IQD1‐GFP fusion protein is targeted to the cell nucleus and that recombinant IQD1 binds to calmodulin in a Ca2\+‐dependent fashion. Analysis of steady‐state messenger RNA levels of glucosinolate pathway genes indicates that IQD1 affects expression of multiple genes with roles in glucosinolate metabolism. Histochemical analysis of tissue‐specific IQD1 ::GUS expression reveals IQD1 promoter activity mainly in vascular tissues of all organs, consistent with the expression patterns of several glucosinolate‐related genes. Interestingly, overexpression of IQD1 reduces insect herbivory, which we demonstrated in dual‐choice assays with the generalist phloem‐feeding green peach aphid (Myzus persicae ), and in weight‐gain assays with the cabbage looper (Trichoplusia ni ), a generalist‐chewing lepidopteran. As IQD1 is induced by mechanical stimuli, we propose IQD1 to be novel nuclear factor that integrates intracellular Ca2\+ signals to fine‐tune glucosinolate accumulation in response to biotic challenge.} } @Article{IPB-1959, author = {Isayenkov, S. and Mrosk, C. and Stenzel, I. and Strack, D. and Hause, B. and}, title = {{Suppression of Allene Oxide Cyclase in Hairy Roots of Medicago truncatula Reduces Jasmonate Levels and the Degree of Mycorrhization with Glomus intraradices}}, year = {2005}, pages = {1401-1410}, journal = {Plant Physiol.}, doi = {10.1104/pp.105.069054}, volume = {139}, abstract = {During the symbiotic interaction between Medicago truncatula and the arbuscular mycorrhizal (AM) fungus Glomus intraradices, an endogenous increase in jasmonic acid (JA) occurs. Two full-length cDNAs coding for the JA-biosynthetic enzyme allene oxide cyclase (AOC) from M. truncatula, designated as MtAOC1 and MtAOC2, were cloned and characterized. The AOC protein was localized in plastids and found to occur constitutively in all vascular tissues of M. truncatula. In leaves and roots, MtAOCs are expressed upon JA application. Enhanced expression was also observed during mycorrhization with G. intraradices. A partial suppression of MtAOC expression was achieved in roots following transformation with Agrobacterium rhizogenes harboring the MtAOC1 cDNA in the antisense direction under control of the cauliflower mosaic virus 35S promoter. In comparison to samples transformed with 35S∷uidA, roots with suppressed MtAOC1 expression exhibited lower JA levels and a remarkable delay in the process of colonization with G. intraradices. Both the mycorrhization rate, quantified by fungal rRNA, and the arbuscule formation, analyzed by the expression level of the AM-specific gene MtPT4, were affected. Staining of fungal material in roots with suppressed MtAOC1 revealed a decreased number of arbuscules, but these did not exhibit an altered structure. Our results indicate a crucial role for JA in the establishment of AM symbiosis.} } @Article{IPB-1951, author = {Gerhardt, B. and Fischer, K. and Balkenhohl, T. J. and Pohnert, G. and Kühn, H. and Wasternack, C. and Feussner, I. and}, title = {{Lipoxygenase-mediated metabolism of storage lipids in germinating sunflower cotyledons and β-oxidation of (9Z,11E,13S)-13-hydroxy-octadeca-9,11-dienoic acid by the cotyledonary glyoxysomes}}, year = {2005}, pages = {919-930}, journal = {Planta}, doi = {10.1007/s00425-004-1408-1}, volume = {220}, abstract = {During the early stages of germination, a lipid-body lipoxygenase is expressed in the cotyledons of sunflowers (Helianthus annuus L.). In order to obtain evidence for the in vivo activity of this enzyme during germination, we analyzed the lipoxygenase-dependent metabolism of polyunsaturated fatty acids esterified in the storage lipids. For this purpose, lipid bodies were isolated from etiolated sunflower cotyledons at different stages of germination, and the storage triacylglycerols were analyzed for oxygenated derivatives. During the time course of germination the amount of oxygenated storage lipids was strongly augmented, and we detected triacylglycerols containing one, two or three residues of (9Z,11E,13S)-13-hydro(pero)xy-octadeca-9,11-dienoic acid. Glyoxysomes from etiolated sunflower cotyledons converted (9Z,11E,13S)-13-hydroxy-octadeca-9,11-dienoic acid to (9Z,11E)-13-oxo-octadeca-9,11-dienoic acid via an NADH-dependent dehydrogenase reaction. Both oxygenated fatty acid derivatives were activated to the corresponding CoA esters and subsequently metabolized to compounds of shorter chain length. Cofactor requirement and formation of acetyl-CoA indicate degradation via β-oxidation. However, β-oxidation only proceeded for two consecutive cycles, leading to accumulation of a medium-chain metabolite carrying an oxo group at C-9, equivalent to C-13 of the parent (9Z,11E,13S)-13-hydroxy-octadeca-9,11-dienoic acid. Short-chain β-oxidation intermediates were not detected during incubation. Similar results were obtained when 13-hydroxy octadecanoic acid was used as β-oxidation substrate. On the other hand, the degradation of (9Z,11E)-octadeca-9,11-dienoic acid was accompanied by the appearance of short-chain β-oxidation intermediates in the reaction mixture. The results suggest that the hydroxyl/oxo group at C-13 of lipoxygenase-derived fatty acids forms a barrier to continuous β-oxidation by glyoxysomes.} } @Article{IPB-1950, author = {Gago, S. and De la Peña, M. and Flores, R. and}, title = {{A kissing-loop interaction in a hammerhead viroid RNA critical for its in vitro folding and in vivo viability}}, year = {2005}, pages = {1073-1083}, journal = {RNA}, doi = {10.1261/rna.2230605}, volume = {11}, abstract = {Chrysanthemum chlorotic mottle viroid (CChMVd) RNA (398–401 nucleotides) can form hammerhead ribozymes that play a functional role in its replication through a rolling-circle mechanism. In contrast to most other viroids, which adopt rod-like or quasi-rod-like secondary structures of minimal free energy, the computer-predicted conformations of CChMVd and Peach latent mosaic viroid (PLMVd) RNAs are branched. Moreover, the covariations found in a number of natural CChMVd variants support that the same or a closely related conformation exists in vivo. Here we report that the CChMVd natural variability also supports that the branched conformation is additionally stabilized by a kissing-loop interaction resembling another one proposed in PLMVd from in vitro assays. Moreover, site-directed mutagenesis combined with bioassays and progeny analysis showed that: (1) single CChMVd mutants affecting the kissing loops had low or no infectivity at all, whereas infectivity was recovered in double mutants restoring the interaction; (2) mutations affecting the structure of the regions adjacent to the kissing loops reverted to wild type or led to rearranged stems, also supporting their interaction; and (3) the interchange between 4 nucleotides of each of the two kissing loops generated a viable CChMVd variant with eight mutations. PAGE analysis under denaturing and nondenaturing conditions revealed that the kissing-loop interaction determines proper in vitro folding of CChMVd RNA. Preservation of a similar kissing-loop interaction in two hammerhead viroids with an overall low sequence similarity suggests that it facilitates in vivo the adoption and stabilization of a compact folding critical for viroid viability.} } @Article{IPB-1948, author = {Fortes, A. M. and Miersch, O. and Lange, P. R. and Malhó, R. and Testillano, P. S. and Risueño, M. d. C. and Wasternack, C. and Pais, M. S. and}, title = {{Expression of Allene Oxide Cyclase and Accumulation of Jasmonates during Organogenic Nodule Formation from Hop (Humulus lupulus var. Nugget) Internodes}}, year = {2005}, pages = {1713-1723}, journal = {Plant Cell Physiol.}, doi = {10.1093/pcp/pci187}, volume = {46}, abstract = {A crucial step in the biosynthesis of jasmonic acid (JA) is the formation of its stereoisomeric precursor, cis-(\+)-12-oxophytodienoic acid (OPDA), which is catalyzed by allene oxide cyclase (AOC, EC 5.3.99.6). A cDNA of AOC was isolated from Humulus lupulus var. Nugget. The ORF of 765 bp encodes a 255 amino acid protein, which carries a putative chloroplast targeting sequence. The recombinant protein without its putative chloroplast target sequence showed significant AOC activity. Previously we demonstrated that wounding induces organogenic nodule formation in hop. Here we show that the AOC transcript level increases in response to wounding of internodes, peaking between 2 and 4 h after wounding. In addition, Western blot analysis showed elevated levels of AOC peaking 24 h after internode inoculation. The AOC increase was accompanied by increased JA levels 24 h after wounding, whereas OPDA had already reached its highest level after 12 h. AOC is mostly present in the vascular bundles of inoculated internodes. During prenodule and nodule formation, AOC levels were still high. JA and OPDA levels decreased down to 10 and 118 pmol (g FW)–1, respectively, during nodule formation, but increased during plantlet regeneration. Double immunolocalization analysis of AOC and Rubisco in connection with lugol staining showed that AOC is present in amyloplasts of prenodular cells and in the chloroplasts of vacuolated nodular cells, whereas meristematic cells accumulated little AOC. These data suggest a role of AOC and jasmonates in organogenic nodule formation and plantlet regeneration from these nodules.} } @Article{IPB-1944, author = {Ebeler, S. E. and Dingley, K. H. and Ubick, E. and Abel, S. and Mitchell, A. E. and Burns, S. A. and Steinberg, F. M. and Clifford, A. J. and}, title = {{Animal Models and Analytical Approaches for Understanding the Relationships Between Wine and Cancer}}, year = {2005}, pages = {19-27}, journal = {Drugs Exp. Clin. Res.}, url = {https://pubmed.ncbi.nlm.nih.gov/15921026/}, volume = {31}, abstract = {We used two approaches for studying the relationships between wine consumption, wine composition and cancer In the first approach, a transgenic mouse model of human neurofibromatosis, combined with the use of well-defined, chemically purified diets, showed that red wine contains nonalcoholic components that can delay tumor onset. In additional studies, catechin, the main monomeric polyphenol of red wine, delayed tumor onset in this mouse model in a positive, linear relationship when incorporated into the diet at levels of 0.5-4 mmol/kg diet. In the second approach, low doses of the chemical carcinogen 2-amino-1-methyl-6-phenylimidazo(4, 5-b)pyridine (PhlP) were administered to rats, and formation of DNA adducts was evaluated by accelerator mass spectrometry. Consumption of red wine solids (the residue from red wine remaining after removal of alcohol and water) and the wine polyphenol quercetin did not influence PhlP-DNA adduct levels or induce liver enzymes (glutathione-S-transferase and quinone reductase). However, quercetin did alter distribution of PhlP in the rat tissues compared to control animals and animals fed other potential dietary chemopreventive agents, including phenylethyl isothiocyanate and sulforaphane. These studies demonstrate the feasibility of these approaches for studying the chemopreventive potential of dietary components at physiologic levels in} } @Article{IPB-1943, author = {Durgbanshi, A. and Arbona, V. and Pozo, O. and Miersch, O. and Sancho, J. V. and Gómez-Cadenas, A. and}, title = {{Simultaneous Determination of Multiple Phytohormones in Plant Extracts by Liquid Chromatography−Electrospray Tandem Mass Spectrometry}}, year = {2005}, pages = {8437-8442}, journal = {J. Agr. Food Chem.}, doi = {10.1021/jf050884b}, volume = {53}, abstract = {A rapid multiresidue method to quantify three different classes of plant hormones has been developed. The reduced concentrations of these metabolites in real samples with complex matrixes require sensitive techniques for their quantification in small amounts of plant tissue. The method described combines high-performance liquid chromatography with electrospray ionization tandem mass spectrometry. Deuterium-labeled standards were added prior to sample extraction to achieve an accurate quantification of abscisic acid, indole-3-acetic acid, and jasmonic acid in a single run. A simple method of extraction and purification involving only centrifugation, a partition against diethyl ether, and filtration was developed and the analytical method validated in four different plant tissues, citrus leaves, papaya roots, barley seedlings, and barley immature embryos. This method represents a clear advantage because it extensively reduces sample preparation and total time for routine analysis of phytohormones in real plant samples.} } @Article{IPB-1941, author = {Danon, A. and Miersch, O. and Felix, G. and op den Camp, R. G. L. and Apel, K. and}, title = {{Concurrent activation of cell death-regulating signaling pathways by singlet oxygen in Arabidopsis thaliana}}, year = {2005}, pages = {68-80}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2004.02276.x}, volume = {41}, abstract = {Upon a dark/light shift the conditional flu mutant of Arabidopsis starts to generate singlet oxygen (1O2), a non‐radical reactive oxygen species that is restricted to the plastid compartment. Immediately after the shift, plants stop growing and develop necrotic lesions. We have established a protoplast system, which allows detection and characterization of the death response in flu induced by the release of 1O2. Vitamin B6 that quenches 1O2 in fungi was able to protect flu protoplasts from cell death. Blocking ethylene production was sufficient to partially inhibit the death reaction. Similarly, flu mutant seedlings expressing transgenic NahG were partially protected from the death provoked by the release of 1O2, indicating a requirement for salicylic acid (SA) in this process, whereas in cells depleted of both, ethylene and SA, the extent of cell death was reduced to the wild‐type level. The flu mutant was also crossed with the jasmonic acid (JA)‐depleted mutant opr3 , and with the JA, OPDA and dinor OPDA (dnOPDA)‐depleted dde2‐2 mutant. Analysis of the resulting double mutants revealed that in contrast to the JA‐induced suppression of H2O2/superoxide‐dependent cell death reported earlier, JA promotes singlet oxygen‐mediated cell death in flu , whereas other oxylipins such as OPDA and dnOPDA antagonize this death‐inducing activity of JA.} } @Article{IPB-1940, author = {Cenzano, A. and Vigliocc, A. and Miersch, O. and Abdala, G. and}, title = {{Hydroxylated jasmonate levels during stolon to tuber transition in Solarium tuberosum L}}, year = {2005}, pages = {107}, journal = {Potato Res.}, doi = {10.1007/BF02742370}, volume = {48}, abstract = {Various octadecanoids and derived compounds have been identified in potato leaves. However, information regarding jasmonate hydroxylated forms in stolons or tubers is scarce. We investigated endogenous jasmonates in stolon material ofSolarium tuberosum cv. Spunta. Stolons and incipient tubers were collected from 8 weeks old plants. The material was cut into apical regions and stolons. We identified jasmonic acid (JA), methyl jasmonate, 11-OH-JA, 12-OH-JA, 12-oxo-phytodienoic acid (OPDA) and a conjugate. The content of JA and 12OH-JA decreased in the apical region but remained high in stolons during tuberization. Thus the apical region might be a site of JAs-utilization or metabolization and stolons might supply JAs to that region. The content of 12-OH-JA was higher than that of 11-OH-JA in all stages analyzed, both in apical regions and stolons. However, these compounds showed a different time-course in the apical region: while 11-OH-JA increased, 12-OH-JA decreased. Thus, JA from leaves or roots could be transported as 12-OH-JA to the apical region, stimulating tuber formation.} } @Article{IPB-1939, author = {Calderon-Villalobos, L. I. and Kuhnle, C. and Dohmann, E. M. and Li, H. and Bevan, M. and Schwechheimer, C. and}, title = {{The Evolutionarily Conserved TOUGH Protein Is Required for Proper Development of Arabidopsis thaliana}}, year = {2005}, pages = {2473-2485}, journal = {Plant Cell}, doi = {10.1105/tpc.105.031302}, volume = {17}, abstract = {In this study, we characterize the evolutionarily conserved TOUGH (TGH) protein as a novel regulator required for Arabidopsis thaliana development. We initially identified TGH as a yeast two-hybrid system interactor of the transcription initiation factor TATA-box binding protein 2. TGH has apparent orthologs in all eukaryotic model organisms with the exception of the budding yeast Saccharomyces cerevisiae. TGH contains domains with strong similarity to G-patch and SWAP domains, protein domains that are characteristic of RNA binding and processing proteins. Furthermore, TGH colocalizes with the splicing regulator SRp34 to subnuclear particles. We therefore propose that TGH plays a role in RNA binding or processing. Arabidopsis tgh mutants display developmental defects, including reduced plant height, polycotyly, and reduced vascularization. We found TGH expression to be increased in the amp1-1 mutant, which is similar to tgh mutants with respect to polycotyly and defects in vascular development. Interestingly, we observed a strong genetic interaction between TGH and AMP1 in that tgh-1 amp1-1 double mutants are extremely dwarfed and severely affected in plant development in general and vascular development in particular when compared with the single mutants.} } @Article{IPB-1930, author = {Andrade, A. and Vigliocco, A. and Alemano, S. and Miersch, O. and Botella, M. A. and Abdala, G. and}, title = {{Endogenous jasmonates and octadecanoids in hypersensitive tomato mutants during germination and seedling development in response to abiotic stress}}, year = {2005}, pages = {309-318}, journal = {Seed Sci. Res.}, doi = {10.1079/SSR2005219}, volume = {15}, abstract = {Although jasmonates (JAs) are involved in germination and seedling development, the regulatory mechanism of JAs, and their relation with endogenous level modifications in these processes, is not well understood. We report here the detection of 12-oxo-phytodienoic acid (OPDA), jasmonic acid (JA), 11-hydroxyjasmonate (11-OH-JA), 12-hydroxyjasmonate (12-OH-JA) and methyljasmonate (JAME) in unimbibed seeds and seedlings of tomato Lycopersicon esculentum Mill cv. Moneymaker (wild type) and tss1, tss2, tos1 mutants. The main compounds in wild-type and tss1, tss2, tos1 seeds were the hydroxylate-JAs; 12-OH-JA was the major component in dry seeds of the wild type and in tss2 and tos1. The amounts of these derivatives were higher in seeds than in seedlings. Changes in JAs during wild-type and tss1 imbibition were analysed in seeds and the imbibition water. In wild-type imbibed seeds, 11-OH-JA content was higher than in tss1. 12-OH-JA showed a different tendency with respect to 11-OH-JA, with high levels in the wild type at early imbibition. In tss1, levels of 12-OH-JA rose from 24 to 48 h of imbibition. At 72 h of imbibition, when radicles had emerged, the amounts of both hydroxylates in wild-type and tss1 seeds were minimal. An important release of the hydroxylate forms was observed in the imbibition water. 11-OH-JA decreased in the imbibition water of wild-type seeds at 48 h. On the contrary, a high and sustained liberation of this compound was observed in tss1 after 24 h. 12-OH-JA increased in wild-type as well in tss1 until 24 h. Thereafter, a substantial reduction in the content of this compound was registered. NaCl-treated wild-type seedlings increased their 12-OH-JA, but tss1 seedlings increased their JA in response to salt treatment. In tss2 seedlings, NaCl caused a slight decrease in 11-OH-JA and JAME, whereas tos1 seedlings showed a dramatic OPDA and 12-OH-JA decrease in response to salt treatment. Under salt stress the mutant seedlings showed different patterns of JAs according to their differential hypersensitivity to abiotic stress. The JA-hydroxylate forms found, and the differential accumulation of JAs during germination, imbibition and seedling development, as well as their response to NaCl stress, provide new evidence about the control of many developmental processes by JA.} } @Article{IPB-1929, author = {Abel, S. and Savchenko, T. and Levy, M. and}, title = {{Genome-wide comparative analysis of the IQD gene families in Arabidopsis thaliana and Oryza sativa}}, year = {2005}, pages = {72}, journal = {BMC Evol. Biol.}, doi = {10.1186/1471-2148-5-72}, volume = {5}, abstract = {BackgroundCalcium signaling plays a prominent role in plants for coordinating a wide range of developmental processes and responses to environmental cues. Stimulus-specific generation of intracellular calcium transients, decoding of calcium signatures, and transformation of the signal into cellular responses are integral modules of the transduction process. Several hundred proteins with functions in calcium signaling circuits have been identified, and the number of downstream targets of calcium sensors is expected to increase. We previously identified a novel, calmodulin-binding nuclear protein, IQD1, which stimulates glucosinolate accumulation and plant defense in Arabidopsis thaliana. Here, we present a comparative genome-wide analysis of a new class of putative calmodulin target proteins in Arabidopsis and rice.ResultsWe identified and analyzed 33 and 29 IQD1-like genes in Arabidopsis thaliana and Oryza sativa, respectively. The encoded IQD proteins contain a plant-specific domain of 67 conserved amino acid residues, referred to as the IQ67 domain, which is characterized by a unique and repetitive arrangement of three different calmodulin recruitment motifs, known as the IQ, 1-5-10, and 1-8-14 motifs. We demonstrated calmodulin binding for IQD20, the smallest IQD protein in Arabidopsis, which consists of a C-terminal IQ67 domain and a short N-terminal extension. A striking feature of IQD proteins is the high isoelectric point (~10.3) and frequency of serine residues (~11%). We compared the Arabidopsis and rice IQD gene families in terms of gene structure, chromosome location, predicted protein properties and motifs, phylogenetic relationships, and evolutionary history. The existence of an IQD-like gene in bryophytes suggests that IQD proteins are an ancient family of calmodulin-binding proteins and arose during the early evolution of land plants.ConclusionComparative phylogenetic analyses indicate that the major IQD gene lineages originated before the monocot-eudicot divergence. The extant IQD loci in Arabidopsis primarily resulted from segmental duplication and reflect preferential retention of paralogous genes, which is characteristic for proteins with regulatory functions. Interaction of IQD1 and IQD20 with calmodulin and the presence of predicted calmodulin binding sites in all IQD family members suggest that IQD proteins are a new class of calmodulin targets. The basic isoelectric point of IQD proteins and their frequently predicted nuclear localization suggest that IQD proteins link calcium signaling pathways to the regulation of gene expression. Our comparative genomics analysis of IQD genes and encoded proteins in two model plant species provides the first step towards the functional dissection of this emerging family of putative calmodulin targets.} } @Article{IPB-1989, author = {Stumpe, M. and Carsjens, J.-G. and Stenzel, I. and Göbel, C. and Lang, I. and Pawlowski, K. and Hause, B. and Feussner, I. and}, title = {{Lipid metabolism in arbuscular mycorrhizal roots of Medicago truncatula}}, year = {2005}, pages = {781-791}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2005.01.020}, volume = {66}, abstract = {The peroxidation of polyunsaturated fatty acids, common to all eukaryotes, is mostly catalyzed by members of the lipoxygenase enzyme family of non-heme iron containing dioxygenases. Lipoxygenase products can be metabolized further in the oxylipin pathway by several groups of CYP74 enzymes. One prominent oxylipin is jasmonic acid (JA), a product of the 13-allene oxide synthase branch of the pathway and known as signaling substance that plays a role in vegetative and propagative plant development as well as in plant responses to wounding and pathogen attack. In barley roots, JA level increases upon colonization by arbuscular mycorrhizal fungi. Apart from this first result regarding JA, no information is available on the relevance of lipidperoxide metabolism in arbuscular mycorrhizal symbiosis. Thus we analyzed fatty acid and lipidperoxide patterns in roots of Medicago truncatula during mycorrhizal colonization. Levels of fungus-specific fatty acids as well as palmitic acid (16:0) and oleic acid (18:1 n − 9) were increased in mycorrhizal roots. Thus the degree of arbuscular mycorrhizal colonization of roots can be estimated via analysis of fungal specific esterified fatty acids. Otherwise, no significant changes were found in the profiles of esterified and free fatty acids. The 9- and 13-LOX products of linoleic and α-linolenic acid were present in all root samples, but did not show significant differences between mycorrhizal and non-mycorrhizal roots, except JA which showed elevated levels in mycorrhizal roots. In both types of roots levels of 13-LOX products were higher than those of 9-LOX products. In addition, three cDNAs encoding CYP74 enzymes, two 9/13-hydroperoxide lyases and a 13-allene oxide synthase, were isolated and characterized. The transcript accumulation of these three genes, however, was not increased in mycorrhizal roots of M. truncatula.} } @Article{IPB-1988, author = {Sharma, V. K. and Monostori, T. and Hause, B. and Maucher, H. and Göbel, C. and Hornung, E. and Hänsch, R. and Bittner, F. and Wasternack, C. and Feussner, I. and Mendel, R. R. and Schulze, J. and}, title = {{Genetic transformation of barley to modify expression of a 13-lipoxygenase}}, year = {2005}, pages = {33-34}, journal = {Acta Biol. Szeged.}, url = {http://abs.bibl.u-szeged.hu/index.php/abs/article/view/2409}, volume = {49}, abstract = {Immature scutella of barley were transformed with cDNA coding for a 13-lipoxygenase of barley (LOX-100) via particle bombardment. Regenerated plants were tested by PAT-assay, Western-analysis and PCR-screening. Immunocytochemical assay of T0 plants showed expression of the LOX cDNA both in the chloroplasts and in the cytosol, depending on the presence of the chloroplast signal peptide sequences in the cDNA. A few transgenic plants containing higher amounts of LOX-derived products have been found. These are the candidates for further analysis concerning pathogen resistance.} } @Article{IPB-1987, author = {Schneider, K. and Kienow, L. and Schmelzer, E. and Colby, T. and Bartsch, M. and Miersch, O. and Wasternack, C. and Kombrink, E. and Stuible, H.-P. and}, title = {{A New Type of Peroxisomal Acyl-Coenzyme A Synthetase from Arabidopsis thaliana Has the Catalytic Capacity to Activate Biosynthetic Precursors of Jasmonic Acid}}, year = {2005}, pages = {13962-13972}, journal = {J. Biol. Chem.}, doi = {10.1074/jbc.M413578200}, volume = {280}, abstract = {Arabidopsis thaliana contains a large number of genes that encode carboxylic acid-activating enzymes, including nine long-chain fatty acyl-CoA synthetases, four 4-coumarate:CoA ligases (4CL), and 25 4CL-like proteins of unknown biochemical function. Because of their high structural and sequence similarity with bona fide 4CLs and their highly hydrophobic putative substrate-binding pockets, the 4CL-like proteins At4g05160 and At5g63380 were selected for detailed analysis. Following heterologous expression, the purified proteins were subjected to a large scale screen to identify their preferred in vitro substrates. This study uncovered a significant activity of At4g05160 with medium-chain fatty acids, medium-chain fatty acids carrying a phenyl substitution, long-chain fatty acids, as well as the jasmonic acid precursors 12-oxo-phytodienoic acid and 3-oxo-2-(2′-pentenyl)-cyclopentane-1-hexanoic acid. The closest homolog of At4g05160, namely At5g63380, showed high activity with long-chain fatty acids and 12-oxo-phytodienoic acid, the latter representing the most efficiently converted substrate. By using fluorescent-tagged variants, we demonstrated that both 4CL-like proteins are targeted to leaf peroxisomes. Collectively, these data demonstrate that At4g05160 and At5g63380 have the capacity to contribute to jasmonic acid biosynthesis by initiating the β-oxidative chain shortening of its precursors.} } @Article{IPB-1984, author = {Rudus, I. and Kepczynska, E. and Kepczynski, J. and Wasternack, C. and Miersch, O. and}, title = {{Changes in jasmonates and 12-oxophytodienoic acid contents of Medicago sativa L. during somatic embryogenesis}}, year = {2005}, pages = {497-504}, journal = {Acta Physiol. Plant.}, doi = {10.1007/s11738-005-0055-x}, volume = {27}, abstract = {Jasmonic acid (JA), its methyl ester (MeJA) and the biosynthetic precursor 12-oxophytodienoic acid (OPDA) were detected quantitatively during somatic embryogenesis of Medicago sativa L. Using GC-MS analysis, these compounds were found in initial explants, in calli and in somatic embryos in the nanogram range per gram of fresh weight. In distinct stages of somatic embryogenesis, JA and 12-OPDA accumulated preferentially in cotyledonary embryos. Initial explants exhibited about five-fold higher JA content than OPDA content, whereas in other stages OPDA accumulated predominantly. These data suggest that also in embryogenic tissues OPDA and JA may have individual signalling properties.} } @Article{IPB-1982, author = {Quint, M. and Ito, H. and Zhang, W. and Gray, W. M. and}, title = {{Characterization of a novel temperature-sensitive allele of the CUL1/AXR6 subunit of SCF ubiquitin-ligases}}, year = {2005}, pages = {371-383}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2005.02449.x}, volume = {43}, abstract = {Selective protein degradation by the ubiquitin‐proteasome pathway has emerged as a key regulatory mechanism in a wide variety of cellular processes. The selective components of this pathway are the E3 ubiquitin‐ligases which act downstream of the ubiquitin‐activating and ‐conjugating enzymes to identify specific substrates for ubiquitinylation. SCF‐type ubiquitin‐ligases are the most abundant class of E3 enzymes in Arabidopsis. In a genetic screen for enhancers of the tir1‐1 auxin response defect, we identified eta1 /axr6‐3 , a recessive and temperature‐sensitive mutation in the CUL1 core component of the SCFTIR1 complex. The axr6‐3 mutation interferes with Skp1 binding, thus preventing SCF complex assembly. axr6‐3 displays a pleiotropic phenotype with defects in numerous SCF‐regulated pathways including auxin signaling, jasmonate signaling, flower development, and photomorphogenesis. We used axr6‐3 as a tool for identifying pathways likely to be regulated by SCF‐mediated proteolysis and propose new roles for SCF regulation of the far‐red light/phyA and sugar signaling pathways. The recessive inheritance and the temperature‐sensitive nature of the pleiotropically acting axr6‐3 mutation opens promising possibilities for the identification and investigation of SCF‐regulated pathways in Arabidopsis.} } @Article{IPB-1971, author = {Meixner, C. and Ludwig-Müller, J. and Miersch, O. and Gresshoff, P. and Staehelin, C. and Vierheilig, H. and}, title = {{Lack of mycorrhizal autoregulation and phytohormonal changes in the supernodulating soybean mutant nts1007}}, year = {2005}, pages = {709-715}, journal = {Planta}, doi = {10.1007/s00425-005-0003-4}, volume = {222}, abstract = {Autoregulatory mechanisms have been reported in the rhizobial and the mycorrhizal symbiosis. Autoregulation means that already existing nodules or an existing root colonization by an arbuscular mycorrhizal fungus systemically suppress subsequent nodule formation/root colonization in other parts of the root system. Mutants of some legumes lost their ability to autoregulate the nodule number and thus display a supernodulating phenotype. On studying the effect of pre-inoculation of one side of a split-root system with an arbuscular mycorrhizal fungus on subsequent mycorrhization in the second side of the split-root system of a wild-type soybean (Glycine max L.) cv. Bragg and its supernodulating mutant nts1007, we observed a clear suppressional effect in the wild-type, whereas further root colonization in the split-root system of the mutant nts1007 was not suppressed. These data strongly indicate that the mechanisms involved in supernodulation also affect mycorrhization and support the hypothesis that the autoregulation in the rhizobial and the mycorrhizal symbiosis is controlled in a similar manner. The accumulation patterns of the plant hormones IAA, ABA and Jasmonic acid (JA) in non-inoculated control plants and split-root systems of inoculated plants with one mycorrhizal side of the split-root system and one non-mycorrhizal side, indicate an involvement of IAA in the autoregulation of mycorrhization. Mycorrhizal colonization of soybeans also resulted in a strong induction of ABA and JA levels, but on the basis of our data the role of these two phytohormones in mycorrhizal autoregulation is questionable.} } @Article{IPB-2063, author = {Wasternack, C. and}, title = {{Introductory Remarks on Biosynthesis and Diversity in Actions}}, year = {2004}, pages = {167-169}, journal = {J. Plant Growth Regul.}, doi = {10.1007/s00344-004-0051-1}, volume = {23}, } @Article{IPB-2057, author = {Ticconi, C. A. and Abel, S. and}, title = {{Short on phosphate: plant surveillance and countermeasures}}, year = {2004}, pages = {548-555}, journal = {Trends Plant Sci.}, doi = {10.1016/j.tplants.2004.09.003}, volume = {9}, abstract = {Metabolism depends on inorganic phosphate (Pi) as reactant, allosteric effector and regulatory moiety in covalent protein modification. To cope with Pi shortage (a common situation in many ecosystems), plants activate a set of adaptive responses to enhance Pi recycling and acquisition by reprogramming metabolism and restructuring root system architecture. The physiology of Pi starvation responses has become well understood, and so current research focuses on the initial molecular events that sense, transmit and integrate information about external and internal Pi status. Recent studies have provided evidence for Pi as a signaling molecule and initial insight into the coordination of Pi deficiency responses at the cellular and molecular level.} } @Article{IPB-2056, author = {Ticconi, C. A. and Delatorre, C. A. and Lahner, B. and Salt, D. E. and Abel, S. and}, title = {{Arabidopsis pdr2 reveals a phosphate-sensitive checkpoint in root development}}, year = {2004}, pages = {801-814}, journal = {Plant J.}, doi = {10.1111/j.1365-313x.2004.02005.x}, volume = {37}, abstract = {Plants have evolved complex strategies to maintain phosphate (Pi) homeostasis and to maximize Pi acquisition when the macronutrient is limiting. Adjustment of root system architecture via changes in meristem initiation and activity is integral to the acclimation process. However, the mechanisms that monitor external Pi status and interpret the nutritional signal remain to be elucidated. Here, we present evidence that the Pi deficiency response , pdr2 , mutation disrupts local Pi sensing. The sensitivity and amplitude of metabolic Pi‐starvation responses, such as Pi‐responsive gene expression or accumulation of anthocyanins and starch, are enhanced in pdr2 seedlings. However, the most conspicuous alteration of pdr2 is a conditional short‐root phenotype that is specific for Pi deficiency and caused by selective inhibition of root cell division followed by cell death below a threshold concentration of about 0.1 mm external Pi. Measurements of general Pi uptake and of total phosphorus (P) in root tips exclude a defect in high‐affinity Pi acquisition. Rescue of root meristem activity in Pi‐starved pdr2 by phosphite (Phi), a non‐metabolizable Pi analog, and divided‐root experiments suggest that pdr2 disrupts sensing of low external Pi availability. Thus, PDR2 is proposed to function at a Pi‐sensitive checkpoint in root development, which monitors environmental Pi status, maintains and fine‐tunes meristematic activity, and finally adjusts root system architecture to maximize Pi acquisition.} } @Article{IPB-2053, author = {Schwechheimer, C. and Villalobos, L. I. A. C. and}, title = {{Cullin-containing E3 ubiquitin ligases in plant development}}, year = {2004}, pages = {677-686}, journal = {Curr. Opin. Plant Biol.}, doi = {10.1016/j.pbi.2004.09.009}, volume = {7}, abstract = {In eukaryotes, the ubiquitin–proteasome system participates in the control of signal transduction events by selectively eliminating regulatory proteins. E3 ubiquitin ligases specifically bind degradation substrates and mediate their poly-ubiquitylation, a prerequisite for their degradation by the 26S proteasome. On the basis of the analysis of the Arabidopsis genome sequence, it is predicted that there are more than 1000 E3 ubiquitin ligases in plants. Several types of E3 ubiquitin ligases have already been characterized in eukaryotes. Recently, some of these E3 enzymes have been implicated in specific plant signaling pathways.} } @Article{IPB-2051, author = {Schüler, G. and Mithöfer, A. and Baldwin, I. T. and BERGER, S. and Ebel, J. and Santos, J. G. and Herrmann, G. and Hölscher, D. and Kramell, R. and Kutchan, T. M. and Maucher, H. and Schneider, B. and Stenzel, I. and Wasternack, C. and Boland, W. and}, title = {{Coronalon: a powerful tool in plant stress physiology}}, year = {2004}, pages = {17-22}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(04)00239-X}, volume = {563}, abstract = {Coronalon, a synthetic 6‐ethyl indanoyl isoleucine conjugate, has been designed as a highly active mimic of octadecanoid phytohormones that are involved in insect and disease resistance. The spectrum of biological activities that is affected by coronalon was investigated in nine different plant systems specifically responding to jasmonates and/or 12‐oxo‐phytodienoic acid. In all bioassays analyzed, coronalon demonstrated a general strong activity at low micromolar concentrations. The results obtained showed the induction of (i) defense‐related secondary metabolite accumulation in both cell cultures and plant tissues, (ii) specific abiotic and biotic stress‐related gene expression, and (iii) root growth retardation. The general activity of coronalon in the induction of plant stress responses together with its simple and efficient synthesis suggests that this compound might serve as a valuable tool in the examination of various aspects in plant stress physiology. Moreover, coronalon might become employed in agriculture to elicit plant resistance against various aggressors.} } @Article{IPB-2037, author = {Miersch, O. and Weichert, H. and Stenzel, I. and Hause, B. and Maucher, H. and Feussner, I. and Wasternack, C. and}, title = {{Constitutive overexpression of allene oxide cyclase in tomato (Lycopersicon esculentum cv. Lukullus) elevates levels of some jasmonates and octadecanoids in flower organs but not in leaves}}, year = {2004}, pages = {847-856}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2004.01.016}, volume = {65}, abstract = {The allene oxide cyclase (AOC), an enzyme in jasmonate biosynthesis, occurs in vascular bundles and ovules of tomato flowers which exhibit a tissue-specific oxylipin signature (Plant J. 24, 113-126, 2000). Constitutive overexpression of the AOC did not led to altered levels of jasmonates in leaves, but these levels increased upon wounding or other stresses suggesting regulation of jasmonate biosynthesis by substrate availability (Plant J. 33, 577-589, 2003). Here, we show dramatic changes in levels of jasmonic acid (JA), of 12-oxo-phytodienoic acid (OPDA), their methyl esters (JAME, OPDAME), and of dinor-OPDA in most flower organs upon constitutive overexpression of AOC. Beside a dominant occurrence of OPDAME and JA in most flower organs, the ratio among the various compounds was altered differentially in the organs of transgenic flowers, e.g. OPDAME increased up to 53-fold in stamen, and JA increased about 51-fold in buds and 7.5-fold in sepals. The increase in jasmonates and octadecanoids was accompanied by decreased levels of free lipid hydro(per)oxy compounds. Except for 16:2, the AOC overexpression led to a significant increase in free but not esterified polyunsaturated fatty acids in all flower organs. The data suggest different regulation of JA biosynthesis in leaves and flowers of tomato.Constitutive overexpression of the AOC increases in all flower organs levels of some jasmonates and octadecanoids, alters the ratios among the compounds, decreases levels of free lipid hydro(per)oxy compounds and increases levels of free but not of esterified polyunsaturated fatty acids.} } @Article{IPB-2035, author = {Maucher, H. and Stenzel, I. and Miersch, O. and Stein, N. and Prasad, M. and Zierold, U. and Schweizer, P. and Dorer, C. and Hause, B. and Wasternack, C. and}, title = {{The allene oxide cyclase of barley (Hordeum vulgare L.)—cloning and organ-specific expression}}, year = {2004}, pages = {801-811}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2004.01.009}, volume = {65}, abstract = {The naturally occurring enantiomer of the various octadecanoids and jasmonates is established in a biosynthetic step catalyzed by the allene oxide cyclase (AOC). The AOC converts an allene oxide formed by an allene oxide synthase (AOS). Here, we show cloning and characterization of cDNAs encoding the AOC and a third AOS, respectively, in addition to the two AOSs previously published (Plant J. 21, 199–213, 2000). The ORF of the AOC-cDNA of 717 bp codes for a protein of 238 amino acid residues carrying a putative chloroplast target sequence. Overexpression without chloroplast target sequence revealed AOC activity. The AOC was found to be a single copy gene which mapped on chromosome 6H. AOC mRNA accumulation appeared in leaf segments upon treatment with various jasmonates, octadecanoids and ABA or during stress such as treatment with sorbitol or glucose solutions. Infection with powdery mildew activated AOC expression in susceptible and resistant lines of barley which correlated with PR1b expression. Among different tissues of barley seedlings, the scutellar node and leaf base accumulated AOC mRNA preferentially which correlated with accumulation of mRNAs for other biosynthetic enzymes (lipoxygenases, AOSs). AOC mRNA accumulation appeared also abundantly in parts of the root containing the tip and correlated with elevated levels of jasmonates. The data suggest a link of AOC expression and JA formation and support role of JA in stress responses and development of barley.Barley plants contain one allene oxide cyclase and three allene oxide synthases which are up-regulated during seedling development accompanied by elevated levels of jasmonate.} } @Article{IPB-2028, author = {Köck, M. and Groß, N. and Stenzel, I. and Hause, G. and}, title = {{Phloem-specific expression of the wound-inducible ribonuclease LE from tomato (Lycopersicon esculentum cv. Lukullus)}}, year = {2004}, pages = {233-242}, journal = {Planta}, doi = {10.1007/s00425-004-1227-4}, volume = {219}, abstract = {Ribonuclease LE (RNaseLE) from tomato (Lycopersicon esculentum Mill. cv. Lukullus) belongs to the widespread RNase T2 family of ribonucleases. With the exception of S-RNases of the solanaceous self-incompatibility system the functions of other members of the RNase T2 family are only barely understood. Using a 2.6-kbp putative promoter sequence of RNaseLE in front of the uidA reporter gene, expression of β-glucuronidase in developing phloem tissue and, especially, in the meristematic and elongation zones at root tips was detected. The tissue-specific expression accords with the range of cis-acting elements detected in the RNaseLE promoter. RNaseLE mRNA was localized in developing phloem cells but not in mature phloem tissue, suggesting association of RNaseLE expression with phloem development. Histochemical staining of β-glucuronidase activity as well as detailed inspection of RNaseLE at mRNA, protein and enzyme activity levels revealed that the wound-induced expression of RNaseLE was also restricted to vascular tissue. RNaseLE transcript accumulation detected by in situ hybridization occurred preferentially in phloem and cambial cells of stem sections upon wounding. The data provide evidence for a role of RNaseLE in a tissue-specific wound response and in wound healing of tomato.} } @Article{IPB-2019, author = {Grubb, C. D. and Zipp, B. J. and Ludwig-Müller, J. and Masuno, M. N. and Molinski, T. F. and Abel, S. and}, title = {{Arabidopsis glucosyltransferase UGT74B1 functions in glucosinolate biosynthesis and auxin homeostasis}}, year = {2004}, pages = {893-908}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.2004.02261.x}, volume = {40}, abstract = {Glucosinolates are a class of secondary metabolites with important roles in plant defense and human nutrition. Here, we characterize a putative UDP‐glucose:thiohydroximate S‐glucosyltransferase, UGT74B1, to determine its role in the Arabidopsis glucosinolate pathway. Biochemical analyses demonstrate that recombinant UGT74B1 specifically glucosylates the thiohydroximate functional group. Low K m values for phenylacetothiohydroximic acid (approximately 6 μ m ) and UDP‐glucose (approximately 50 μm ) strongly suggest that thiohydroximates are in vivo substrates of UGT74B1. Insertional loss‐of‐function ugt74b1 mutants exhibit significantly decreased, but not abolished, glucosinolate accumulation. In addition, ugt74b1 mutants display phenotypes reminiscent of auxin overproduction, such as epinastic cotyledons, elongated hypocotyls in light‐grown plants, excess adventitious rooting and incomplete leaf vascularization. Indeed, during early plant development, mutant ugt74b1 seedlings accumulate nearly threefold more indole‐3‐acetic acid than the wild type. Other phenotypes, however, such as chlorosis along the leaf veins, are likely caused by thiohydroximate toxicity. Analysis of UGT74B1 promoter activity during plant development reveals expression patterns consistent with glucosinolate metabolism and induction by auxin treatment. The results are discussed in the context of known mutations in glucosinolate pathway genes and their effects on auxin homeostasis. Taken together, our work provides complementary in vitro and in vivo evidence for a primary role of UGT74B1 in glucosinolate biosynthesis.} } @Article{IPB-2018, author = {Groß, N. and Wasternack, C. and Köck, M. and}, title = {{Wound-induced RNaseLE expression is jasmonate and systemin independent and occurs only locally in tomato (Lycopersicon esculentum cv. Lukullus)}}, year = {2004}, pages = {1343-1350}, journal = {Phytochemistry}, doi = {10.1016/j.phytochem.2004.04.036}, volume = {65}, abstract = {Tomato RNaseLE is induced by phosphate deficiency and wounding and may play a role in macromolecular recycling as well as wound healing. Here, we analyzed the role of jasmonate and systemin in the wound-induced RNaseLE activation. The rapid expression of RNaseLE upon wounding of leaves leading to maximal RNase activity within 10 h, appeared only locally. Jasmonic acid (JA) or its molecular mimic ethyl indanoyl isoleucine conjugate did not induce RNaseLE expression. Correspondingly, RNaseLE was expressed upon wounding of 35S::allene oxide cyclase antisense plants known to be JA deficient. RNaseLE was not expressed upon systemin treatment, but was locally expressed in the spr1 mutant which is affected in systemin perception. In tomato plants carrying a PromLE::uidA construct, GUS activity could be detected upon wounding, but not following treatment with JA or systemin. The data indicate a locally acting wound-inducible systemin- and JA-independent signaling pathway for RNaseLE expression.RNaseLE expression was analyzed by pharmacological studies of different tomato lines and upon wounding of leaves. The gene is only locally activated via a new type of wound-induced signaling pathway in a jasmonate/systemin-independent manner.} } @Article{IPB-2015, author = {Frisch, M. and Quint, M. and Lübberstedt, T. and Melchinger, A. E. and}, title = {{Duplicate marker loci can result in incorrect locus orders on linkage maps}}, year = {2004}, pages = {305-316}, journal = {Theor. Appl. Genet.}, doi = {10.1007/s00122-003-1578-4}, volume = {109}, abstract = {Genetic linkage maps, constructed from multi-locus recombination data, are the basis for many applications of molecular markers. For the successful employment of a linkage map, it is essential that the linear order of loci on a chromosome is correct. The objectives of this theoretical study were to (1) investigate the occurrence of incorrect locus orders caused by duplicate marker loci, (2) develop a statistical test for the detection of duplicate markers, and (3) discuss the implications for practical applications of linkage maps. We derived conditions, under which incorrect locus orders do or do not occur with duplicate marker loci for the general case of n markers on a chromosome in a BC1 mapping population. We further illustrated these conditions numerically for the special case of four markers. On the basis of the extent of segregation distortion, an exact test for the presence of duplicate marker loci was suggested and its power was investigated numerically. Incorrect locus orders caused by duplicate marker loci can (1) negatively affect the assignment of target genes to chromosome regions in a map-based cloning experiment, (2) hinder indirect selection for a favorable allele at a quantitative trait locus, and (3) decrease the efficiency of reducing the length of the chromosome segment attached to a target gene in marker-assisted backcrossing.} } @Article{IPB-2012, author = {Flores, R. and Delgado, S. and Gas, M.-E. and Carbonell, A. and Molina, D. and Gago, S. and De la Peña, M. and}, title = {{Viroids: the minimal non-coding RNAs with autonomous replication}}, year = {2004}, pages = {42-48}, journal = {FEBS Lett.}, doi = {10.1016/j.febslet.2004.03.118}, volume = {567}, abstract = {Viroids are small (246–401 nucleotides), non‐coding, circular RNAs able to replicate autonomously in certain plants. Viroids are classified into the families Pospiviroidae and Avsunviroidae , whose members replicate in the nucleus and chloroplast, respectively. Replication occurs by an RNA‐based rolling‐circle mechanism in three steps: (1) synthesis of longer‐than‐unit strands catalyzed by host DNA‐dependent RNA polymerases forced to transcribe RNA templates, (2) processing to unit‐length, which in family Avsunviroidae is mediated by hammerhead ribozymes, and (3) circularization either through an RNA ligase or autocatalytically. Disease induction might result from the accumulation of viroid‐specific small interfering RNAs that, via RNA silencing, could interfere with normal developmental pathways.} } @Article{IPB-2010, author = {Bücking, H. and Förster, H. and Stenzel, I. and Miersch, O. and Hause, B. and}, title = {{Applied jasmonates accumulate extracellularly in tomato, but intracellularly in barley}}, year = {2004}, pages = {45-50}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(04)00178-4}, volume = {562}, abstract = {Jasmonic acid (JA) and its derivatives are well‐characterized signaling molecules in plant defense and development, but the site of their localization within plant tissue is entirely unknown. To address the question whether applied JA accumulates extracellularly or intracellularly, leaves of tomato and barley were fed with 14C‐labeled JA and the label was localized in cryofixed and lyophilized leaf tissues by microautoradiography. In tomato the radioactivity was detectable within the apoplast, but no label was found within the mesophyll cells. By contrast, in barley leaf tissues, radioactivity was detected within the mesophyll cells suggesting a cellular uptake of exogenously applied JA. JA, applied to leaves of both plants as in the labeling experiments, led in all leaf cells to the expression of JA‐inducible genes indicating that the perception is completed by JA signal transduction.} } @Article{IPB-2089, author = {Hause, B. and Stenzel, I. and Miersch, O. and Wasternack, C. and}, title = {{Occurrence of the allene oxide cyclase in different organs and tissues of Arabidopsis thaliana}}, year = {2003}, pages = {971-980}, journal = {Phytochemistry}, doi = {10.1016/S0031-9422(03)00447-3}, volume = {64}, abstract = {Occurrence of an essential enzyme in jasmonate (JA) biosynthesis, the allene oxide cyclase, (AOC) was analyzed in different developmental stages and various organs of Arabidopsis thaliana plants by immuno blot analysis and immunocytological approaches. Levels of AOC and of the two preceding enzymes in JA biosynthesis increased during seedling development accompanied by increased levels of JA and 12-oxophytodienoic acid levels after 4 and 8 weeks. Most tissues including all vascular bundles and that of flower buds contain AOC protein. Flowers shortly before opening, however, contain AOC protein preferentially in ovules, stigma cells and vascular bundles, whereas in anthers and pollen AOC could not be detected. The putative roles of AOC and JA in development are discussed.The allene oxide cyclase (AOC) is an important enzyme in jasmonate biosynthesis. Levels and occurrence of AOC in different organs and tissues are altered during development of Arabidopsis thaliana.} } @Article{IPB-2088, author = {Hause, B. and Hause, G. and Kutter, C. and Miersch, O. and Wasternack, C. and}, title = {{Enzymes of Jasmonate Biosynthesis Occur in Tomato Sieve Elements}}, year = {2003}, pages = {643-648}, journal = {Plant Cell Physiol.}, doi = {10.1093/pcp/pcg072}, volume = {44}, abstract = {The allene oxide cyclase (AOC) is a plastid-located enzyme in the biosynthesis of the signaling compound jasmonic acid (JA). In tomato, AOC occurs specifically in ovules and vascular bundles [Hause et al. (2000)PlantJ. 24; 113]. Immunocytological analysis of longitudinal sections of petioles and flower stalks revealed the occurrence of AOC in companion cells (CC) and sieve elements (SE). Electron microscopic analysis led to the conclusion that the AOC-containing structures of SE are plastids. AOC was not detected in SE of 35S::AOCantisense plants. The enzymes preceding AOC in JA biosynthesis, the allene oxide synthase (AOS) and the lipoxygenase, were also detected in SE. In situ hybridization showed that the SE are free of AOC-mRNA suggesting AOC protein traffic from CC to SE via plasmodesmata. A control by in situ hybridization of AOS mRNA coding for a protein with a size above the exclusion limit of plasmodesmata indicated mRNA in CC and SE. The data suggest that SE carry the capacity to form 12-oxo-phytodienoic acid, the unique precursor of JA. Together with preferential generation of JA in vascular bundles [Stenzel et al. (2003)Plant J. 33: 577], the data support a role of JA in systemic wound signaling.} } @Article{IPB-2086, author = {Gidda, S. K. and Miersch, O. and Levitin, A. and Schmidt, J. and Wasternack, C. and Varin, L. and}, title = {{Biochemical and Molecular Characterization of a Hydroxyjasmonate Sulfotransferase from Arabidopsis thaliana}}, year = {2003}, pages = {17895-17900}, journal = {J. Biol. Chem.}, doi = {10.1074/jbc.M211943200}, volume = {278}, abstract = {12-Hydroxyjasmonate, also known as tuberonic acid, was first isolated from Solanum tuberosum and was shown to have tuber-inducing properties. It is derived from the ubiquitously occurring jasmonic acid, an important signaling molecule mediating diverse developmental processes and plant defense responses. We report here that the gene AtST2a from Arabidopsis thaliana encodes a hydroxyjasmonate sulfotransferase. The recombinant AtST2a protein was found to exhibit strict specificity for 11- and 12-hydroxyjasmonate with Km values of 50 and 10 μm, respectively. Furthermore, 12-hydroxyjasmonate and its sulfonated derivative are shown to be naturally occurring inA. thaliana. The exogenous application of methyljasmonate to A. thaliana plants led to increased levels of both metabolites, whereas treatment with 12-hydroxyjasmonate led to increased level of 12-hydroxyjasmonate sulfate without affecting the endogenous level of jasmonic acid. AtST2a expression was found to be induced following treatment with methyljasmonate and 12-hydroxyjasmonate. In contrast, the expression of the methyljasmonate-responsive gene Thi2.1, a marker gene in plant defense responses, is not induced upon treatment with 12-hydroxyjasmonate indicating the existence of independent signaling pathways responding to jasmonic acid and 12-hydroxyjasmonic acid. Taken together, the results suggest that the hydroxylation and sulfonation reactions might be components of a pathway that inactivates excess jasmonic acid in plants. Alternatively, the function of AtST2a might be to control the biological activity of 12-hydroxyjasmonic acid.} } @Article{IPB-2084, author = {Färber, K. and Schumann, B. and Miersch, O. and Roos, W. and}, title = {{Selective desensitization of jasmonate- and pH-dependent signaling in the induction of benzophenanthridine biosynthesis in cells of Eschscholzia californica}}, year = {2003}, pages = {491-500}, journal = {Phytochemistry}, doi = {10.1016/S0031-9422(02)00562-9}, volume = {62}, abstract = {The biosynthesis of benzophenanthridine alkaloids, phytoalexins of Eschscholzia californica, in cultured cells can be induced by a glycoprotein preparation from yeast, methyljasmonate, artificial acidification with permeant acids, or mild osmotic stress. Each of these stimuli strongly attenuated the subsequent response to the same stimulus (homologous desensitization). Elicitor contact and artificial acidification mutually desensitized the cells for either signal. In contrast, elicitor-treated cells maintained their responsiveness to methyljasmonate or hyperosmolarity (sorbitol). Elicitor concentrations that nearly saturated the alkaloid response did not cause a detectable increase of jasmonate content. Transient acidification of the cytoplasm is a necessary step of signaling by low elicitor concentrations but was not detectable after jasmonate treatment. Seen together, the data indicate the existence of a jasmonate-dependent and jasmonate-independent (ΔpH controlled) signal pathway towards the expression of benzophenanthridine biosynthesis. Selective desensitization allows either stimulus to activate a distinct share of the biosynthetic capacity of the cell and limits the accumulation of toxic defense metabolites.Yeast elicitor and jasmonate trigger alkaloid production via different signal pathways that show selective desensitization. Elicitor treatment (bottom cells) but not jasmonate (top cells) evokes intracellular pH shifts.} } @Article{IPB-2082, author = {Dußle, C. and Quint, M. and Melchinger, A. and Xu, M. and Lübberstedt, T. and}, title = {{Saturation of two chromosome regions conferring resistance to SCMV with SSR and AFLP markers by targeted BSA}}, year = {2003}, pages = {485-493}, journal = {Theor. Appl. Genet.}, doi = {10.1007/s00122-002-1107-x}, volume = {106}, abstract = {Quantitative trait loci (QTLs) and bulked segregant analyses (BSA) identified the major genes Scmv1 on chromosome 6 and Scmv2 on chromosome 3, conferring resistance against sugarcane mosaic virus (SCMV) in maize. Both chromosome regions were further enriched for SSR and AFLP markers by targeted bulked segregant analysis (tBSA) in order to identify and map only markers closely linked to either Scmv1 or Scmv2. For identification of markers closely linked to the target genes, symptomless individuals of advanced backcross generations BC5 to BC9 were employed. All AFLP markers, identified by tBSA using 400 EcoRI/MseI primer combinations, mapped within both targeted marker intervals. Fourteen SSR and six AFLP markers mapped to the Scmv1 region. Eleven SSR and 18 AFLP markers were located in the Scmv2 region. Whereas the linear order of SSR markers and the window size for the Scmv2 region fitted well with publicly available genetic maps, map distances and window size differed substantially for the Scmv1 region on chromosome 6. A possible explanation for the observed discrepancies is the presence of two closely linked resistance genes in the Scmv1 region.} } @Article{IPB-2080, author = {Dingley, K. H. and Ubick, E. A. and Chiarappa-Zucca, M. L. and Nowell, S. and Abel, S. and Ebeler, S. E. and Mitchell, A. E. and Burns, S. A. and Steinberg, F. M. and Clifford, A. J. and}, title = {{Effect of Dietary Constituents With Chemopreventive Potential on Adduct Formation of a Low Dose of the Heterocyclic Amines PhIP and IQ and Phase II Hepatic Enzymes}}, year = {2003}, pages = {212-221}, journal = {Nutr. Cancer}, doi = {10.1207/S15327914NC4602_15}, volume = {46}, abstract = {We conducted a study to evaluate dietary chemopreventive strategies to reduce genotoxic effects of the carcinogens 2-amino-1-methyl-6-phenyl-imidazo[4,5-b]pyridine (PhIP) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ). PhIP and IQ are heterocyclic amines (HCAs) that are found in cooked meat and may be risk factors for cancer. Typical chemoprevention studies have used carcinogen doses many thousand-fold higher than usual human daily intake. Therefore, we administered a low dose of [14C] PhIPand [3H] IQand utilized accelerator mass spectrometry to quantify PhIP adducts in the liver, colon, prostate, and blood plasma and IQadducts in the liver and blood plasma with high sensitivity. Diets supplemented with phenethylisothiocyanate (PEITC), genistein, chlorophyllin, or lycopene were evaluated for their ability to decrease adduct formation of [14C] PhIPand [3H] IQin rats. We also examined the effect of treatments on the activity of the phase II detoxification enzymes glutathione S-transferase (GST), UDP-glucuronyltransferase (UGT), phenol sulfotransferase (SULT) and quinone reductase (QR). PEITC and chlorophyllin significantly decreased PhIP-DNA adduct levels in all tissues examined, which was reflected by similar changes in PhIP binding to albumin in the blood. In contrast, genistein and lycopene tended to increase PhIP adduct levels. The treatments did not significantly alter the level of IQ-DNA or -protein adducts in the liver.With the exception of lycopene, the treatments had some effect on the activity of one or more hepatic phase II detoxification enzymes. We conclude that PEITC and chlorophyllin are protective of PhIP-induced genotoxicity after a low exposure dose of carcinogen, possibly through modification of HCA metabolism.} } @Article{IPB-2079, author = {De la Peña, M. and Gago, S. and Flores, R. and}, title = {{Peripheral regions of natural hammerhead ribozymes greatly increase their self-cleavage activity}}, year = {2003}, pages = {5561-5570}, journal = {EMBO J.}, doi = {10.1093/emboj/cdg530}, volume = {22}, abstract = {Natural hammerhead ribozymes are mostly found in some viroid and viroid‐like RNAs and catalyze their cis cleavage during replication. Hammerheads have been manipulated to act in trans and assumed to have a similar catalytic behavior in this artificial context. However, we show here that two natural cis‐acting hammerheads self‐cleave much faster than trans‐acting derivatives and other reported artificial hammerheads. Moreover, modifications of the peripheral loops 1 and 2 of one of these natural hammerheads induced a \>100‐fold reduction of the self‐cleavage constant, whereas engineering a trans‐acting artificial hammerhead into a cis derivative by introducing a loop 1 had no effect. These data show that regions external to the central conserved core of natural hammerheads play a role in catalysis, and suggest the existence of tertiary interactions between these peripheral regions. The interactions, determined by the sequence and size of loops 1 and 2 and most likely of helices I and II, must result from natural selection and should be studied in order to better understand the hammerhead requirements in vivo.} } @Article{IPB-2070, author = {Abdala, G. and Miersch, O. and Kramell, R. and Vigliocco, A. and Agostini, E. and Forchetti, G. and Alemano, S. and}, title = {{Jasmonate and octadecanoid occurrence in tomato hairy roots. Endogenous level changes in response to NaCl}}, year = {2003}, pages = {21-27}, journal = {Plant Growth Regul.}, doi = {10.1023/A:1023016412454}, volume = {40}, abstract = {Jasmonic acid biosynthesis occurs in leaves and there is also evidence of a similar pathway in roots. The expression of lipoxygenase, allene oxide cyclase and low amounts of transcripts of allene oxide synthase in tomato roots indicates that some steps of the jasmonate synthesis may occur in these organs. Thus, the aim of the present work was to study the jasmonate and octadecanoid occurrence in tomato roots using isolated cultures of hairy roots. These were obtained by the transformation of cv. Pera roots with Agrobacterium rhyzogenes. Also we investigated the effect of NaCl stress on the endogenous levels of these compounds. Jasmonic acid, 12-oxophytodienoic acid and their methylated derivatives, as well as a jasmonate-isoleucine conjugate, were present in control hairy roots of 30 d of culture. The 12-oxophytodienoic acid and its methylated derivative showed higher levels than jasmonic acid and its methylated form, although the content of the conjugate was the same as that of jasmonic acid. After salinization of hairy roots for 14, 20 and 30 d, free jasmonates and octadecanoids were measured. Fourteen days after salt treatment, increased levels of these compounds were found, jasmonic acid and 12-oxophytodienoic acid showed the most remarkable rise. 11-OH-jasmonic acid was found at 14 d of culture in control and salt-treated hairy roots; whereas the 12-OH- form of jasmonic acid was only detected in the salt-treated hairy roots. Agrobacterium rhizogenes cultures did not produce jasmonates and/or octadecanoids.} } @Article{IPB-2124, author = {Stenzel, I. and Hause, B. and Miersch, O. and Kurz, T. and Maucher, H. and Weichert, H. and Ziegler, J. and Feussner, I. and Wasternack, C. and}, title = {{Jasmonate biosynthesis and the allene oxide cyclase family of Arabidopsis thaliana}}, year = {2003}, pages = {895-911}, journal = {Plant Mol. Biol.}, doi = {10.1023/A:1023049319723}, volume = {51}, abstract = {In biosynthesis of octadecanoids and jasmonate (JA), the naturally occurring enantiomer is established in a step catalysed by the gene cloned recently from tomato as a single-copy gene (Ziegler et al., 2000). Based on sequence homology, four full-length cDNAs were isolated from Arabidopsis thaliana ecotype Columbia coding for proteins with AOC activity. The expression of AOCgenes was transiently and differentially up-regulated upon wounding both locally and systemically and was induced by JA treatment. In contrast, AOC protein appeared at constitutively high basal levels and was slightly increased by the treatments. Immunohistochemical analyses revealed abundant occurrence of AOC protein as well as of the preceding enzymes in octadecanoid biosynthesis, lipoxygenase (LOX) and allene oxide synthase (AOS), in fully developed tissues, but much less so in 7-day old leaf tissues. Metabolic profiling data of free and esterified polyunsaturated fatty acids and lipid peroxidation products including JA and octadecanoids in wild-type leaves and the jasmonate-deficient mutant OPDA reductase 3 (opr3) revealed preferential activity of the AOS branch within the LOX pathway. 13-LOX products occurred predominantly as esterified derivatives, and all 13-hydroperoxy derivatives were below the detection limits. There was a constitutive high level of free 12-oxo-phytodienoic acid (OPDA) in untreated wild-type and opr3 leaves, but an undetectable expression of AOC. Upon wounding opr3 leaves exhibited only low expression of AOC, wounded wild-type leaves, however, accumulated JA and AOC mRNA. These and further data suggest regulation of JA biosynthesis by OPDA compartmentalization and a positive feedback by JA during leaf development.} } @Article{IPB-2123, author = {Stenzel, I. and Ziethe, K. and Schurath, J. and Hertel, S. C. and Bosse, D. and Köck, M. and}, title = {{Differential expression of the LePS2 phosphatase gene family in response to phosphate availability, pathogen infection and during development}}, year = {2003}, pages = {138-146}, journal = {Physiol. Plant.}, doi = {10.1034/j.1399-3054.2003.00091.x}, volume = {118}, abstract = {In this study, we report the cloning of the three‐member LePS2 gene family of acid phosphatases via subtractive screening of a cDNA library of Pi‐starved cultivated tomato cells (Lycopersicon esculentum Mill. cv. Lukullus). As members of the plant Pi‐starvation response, LePS2 genes were tightly regulated in cultivated cells and tomato seedlings by Pi availability. The LePS2 enzymes which are most likely expressed in the cytoplasma could be involved in processes that are accompanied by degradation of phosphorylated organic substrates. Independently from exogenous phosphate supply LePS2 expression was detected in tomato endosperm during germination. LePS2 genes were differentially induced after infection with the bacterial pathogen Pseudomonas syringae and in the early stages of flower development. Using RT–PCR it was found that the gene LePS2B was the most abundant transcript in phosphate‐depleted cells, but a reduced expression was determined in floral buds and it was not found during pathogen interaction. In this respect, it is interesting that the promoter sequences of the LePS2 genes are also divergent. LePS2 gene products may have functions in developmental processes which are restricted to distinct plant tissues or cell types.} } @Article{IPB-2122, author = {Stenzel, I. and Hause, B. and Maucher, H. and Pitzschke, A. and Miersch, O. and Ziegler, J. and Ryan, C. A. and Wasternack, C. and}, title = {{Allene oxide cyclase dependence of the wound response and vascular bundle-specific generation of jasmonates in tomato - amplification in wound signalling}}, year = {2003}, pages = {577-589}, journal = {Plant J.}, doi = {10.1046/j.1365-313X.2003.01647.x}, volume = {33}, abstract = {The allene oxide cyclase (AOC)‐catalyzed step in jasmonate (JA) biosynthesis is important in the wound response of tomato. As shown by treatments with systemin and its inactive analog, and by analysis of 35S::prosysteminsense and 35S::prosysteminantisense plants, the AOC seems to be activated by systemin (and JA) leading to elevated formation of JA. Data are presented on the local wound response following activation of AOC and generation of JA, both in vascular bundles. The tissue‐specific occurrence of AOC protein and generation of JA is kept upon wounding or other stresses, but is compromised in 35S::AOCsense plants, whereas 35S::AOCantisense plants exhibited residual AOC expression, a less than 10% rise in JA, and no detectable expression of wound response genes. The (i) activation of systemin‐dependent AOC and JA biosynthesis occurring only upon substrate generation, (ii) the tissue‐specific occurrence of AOC in vascular bundles, where the prosystemin gene is expressed, and (iii) the tissue‐specific generation of JA suggest an amplification in the wound response of tomato leaves allowing local and rapid defense responses.} } @Article{IPB-2117, author = {Schilling, S. and Manhart, S. and Hoffmann, T. and Ludwig, H.-H. and Wasternack, C. and Demuth, H.-U. and}, title = {{Substrate Specificity of Glutaminyl Cyclases from Plants and Animals}}, year = {2003}, pages = {1583-1592}, journal = {Biol. Chem.}, doi = {10.1515/BC.2003.175}, volume = {384}, abstract = {Glutaminyl cyclases (QC) catalyze the intramolecular cyclization of N-terminal glutamine residues of peptides and proteins. For a comparison of the substrate specificity of human and papaya QC enzymes, a novel continuous assay was established by adapting an existing discontinuous method. Specificity constants (kcat/Km) of dipeptides and dipeptide surrogates were higher for plant QC, whereas the selectivity for oligopeptides was similar for both enzymes. However, only the specificity constants of mammalian QC were dependent on size and composition of the substrates. Specificity constants of both enzymes were equally pH-dependent in the acidic pH-region, revealing a pKa value identical to the pKa of the substrate, suggesting similarities in the substrate conversion mode. Accordingly, both QCs converted the L-?homoglutaminyl residue in the peptide H-?homoGln-Phe-Lys-Arg-Leu-Ala-NH2 and the glutaminyl residues of the branched peptide H-Gln-Lys(Gln)-Arg-Leu-Ala-NH2 as well as the partially cyclized peptide H-Gln-cyclo( N?-Lys-Arg-Pro-Ala-Gly-Phe). In contrast, only QC from C. papaya was able to cyclize a methylated glutamine residue, while this compound did not even inhibit human QC-catalysis, suggesting distinct substrate recognition pattern. The conversion of the potential physiological substrates gastrin, neurotensin and [GlN1]-fertilization promoting peptide indicates that human QC may play a key role in posttranslational modification of most if not all pGlu-containing hormones.} } @Article{IPB-2116, author = {Schilling, S. and Niestroj, A. J. and Rahfeld, J.-U. and Hoffmann, T. and Wermann, M. and Zunkel, K. and Wasternack, C. and Demuth, H.-U. and}, title = {{Identification of Human Glutaminyl Cyclase as a Metalloenzyme}}, year = {2003}, pages = {49773-49779}, journal = {J. Biol. Chem.}, doi = {10.1074/jbc.M309077200}, volume = {278}, abstract = {Human glutaminyl cyclase (QC) was identified as a metalloenzyme as suggested by the time-dependent inhibition by the heterocyclic chelators 1,10-phenanthroline and dipicolinic acid. The effect of EDTA on QC catalysis was negligible. Inactivated enzyme could be fully restored by the addition of Zn2\+ in the presence of equimolar concentrations of EDTA. Little reactivation was observed with Co2\+ and Mn2\+. Other metal ions such as K\+, Ca2\+, and Ni2\+ were inactive under the same conditions. Additionally, imidazole and imidazole derivatives were identified as competitive inhibitors of QC. An initial structure activity-based inhibitor screening of imidazole-derived compounds revealed potent inhibition of QC by imidazole N-1 derivatives. Subsequent data base screening led to the identification of two highly potent inhibitors, 3-[3-(1H-imidazol-1-yl)propyl]-2-thioxoimidazolidin-4-one and 1,4-bis-(imidazol-1-yl)-methyl-2,5-dimethylbenzene, which exhibited respective Ki values of 818 ± 1 and 295 ± 5 nm. The binding properties of the imidazole derivatives were further analyzed by the pH dependence of QC inhibition. The kinetically obtained pKa values of 6.94 ± 0.02, 6.93 ± 0.03, and 5.60 ± 0.05 for imidazole, methylimidazole, and benzimidazole, respectively, match the values obtained by titrimetric pKa determination, indicating the requirement for an unprotonated nitrogen for binding to QC. Similarly, the pH dependence of the kinetic parameter Km for the QC-catalyzed conversion of H-Gln-7-ami-no-4-methylcoumarin also implies that only N-terminally unprotonated substrate molecules are bound to the active site of the enzyme, whereas turnover is not affected. The results reveal human QC as a metal-dependent transferase, suggesting that the active site-bound metal is a potential site for interaction with novel, highly potent competitive inhibitors.} } @Article{IPB-2111, author = {Quint, M. and Dußle, C. M. and Melchinger, A. E. and Lübberstedt, T. and}, title = {{Identification of genetically linked RGAs by BAC screening in maize and implications for gene cloning, mapping and MAS}}, year = {2003}, pages = {1171-1177}, journal = {Theor. Appl. Genet.}, doi = {10.1007/s00122-002-1105-z}, volume = {106}, abstract = {The resistance gene analogue (RGA) pic19 in maize, a candidate for sugarcane mosaic virus (SCMV) resistance gene (R gene) Scmv1, was used to screen a maize BAC library to identify homologous sequences in the maize genome and to investigate their genomic organisation. Fifteen positive BAC clones were identified and could be classified into five physically independent contigs consisting of overlapping clones. Genetic mapping clustered three contigs into the same genomic region as Scmv1 on chromosome 6S. The two remaining contigs mapped to the same region as a QTL for SCMV resistance on chromosome 1. Thus, RGAs mapping to a target region can be successfully used to identify further-linked candidate sequences. The pic19 homologous sequences of these clones revealed a sequence similarity of 94–98% on the nucleotide level. The high sequence similarity reveals potential problems for the use of RGAs as molecular markers. Their application in marker-assisted selection (MAS) and the construction of high-density genetic maps is complicated by the existence of closely linked homologues resulting in \'ghost\' marker loci analogous to \'ghost\' QTLs. Therefore, implementation of genomic library screening, including genetic mapping of potential homologues, seems necessary for the safe application of RGA markers in MAS and gene isolation.} } @Article{IPB-2104, author = {Naum-Onganı́a, G. and Gago-Zachert, S. and Peña, E. and Grau, O. and Laura Garcia, M. and}, title = {{Citrus psorosis virus RNA 1 is of negative polarity and potentially encodes in its complementary strand a 24K protein of unknown function and 280K putative RNA dependent RNA polymerase}}, year = {2003}, pages = {49-61}, journal = {Virus Res.}, doi = {10.1016/S0168-1702(03)00172-2}, volume = {96}, abstract = {Citrus psorosis virus (CPsV), the type member of genus Ophiovirus, has three genomic RNAs. Complete sequencing of CPsV RNA 1 revealed a size of 8184 nucleotides and Northern blot hybridization with chain specific probes showed that its non-coding strand is preferentially encapsidated. The complementary strand of RNA 1 contains two open reading frames (ORFs) separated by a 109-nt intergenic region, one located near the 5′-end potentially encoding a 24K protein of unknown function, and another of 280K containing the core polymerase motifs characteristic of viral RNA-dependent RNA polymerases (RdRp). Comparison of the core RdRp motifs of negative-stranded RNA viruses, supports grouping CPsV, Ranunculus white mottle virus (RWMV) and Mirafiori lettuce virus (MiLV) within the same genus (Ophiovirus), constituting a monophyletic group separated from all other negative-stranded RNA viruses. Furthermore, RNAs 1 of MiLV, CPsV and RWMV are similar in size and those of MiLV and CPsV also in genomic organization and sequence.} } @Article{IPB-2102, author = {Monostori, T. and Schulze, J. and Sharma, V. K. and Maucher, H. and Wasternack, C. and Hause, B. and}, title = {{Novel plasmid vectors for homologous transformation of barley (Hordeum vulgare L.) with JIP23 cDNA in sense and antisense orientation}}, year = {2003}, pages = {17-24}, journal = {Cereal Res. Commun.}, doi = {10.1007/BF03543245}, volume = {31}, abstract = {The most abundant jasmonate-induced protein (JIP) in barley leaves is a 23 kDa protein (JIP23). Its function, however, is unknown. In order to analyze its function by homologous transformation, new plasmid vectors have been constructed. They carry the cDNA coding for JIP23 in sense or antisense orientation under the control of the Ubi-1-promoter as well as the pat resistance gene under the control of the 35S promoter. Barley mesophyll protoplasts were transiently transformed with the sense constructs. PAT activity and immunological detection of JIP23 could be achieved in transformed protoplasts but not in untransformed protoplasts indicating that the construct was active. Thus, these new vectors are suitable for stable transformation of barley. Carrying a multiple cloning site (MCS), these vectors can be used now in a wide range of transformation of barley.} } @Article{IPB-2106, author = {O'Donnell, P. J. and Schmelz, E. and Block, A. and Miersch, O. and Wasternack, C. and Jones, J. B. and Klee, H. J. and}, title = {{Multiple Hormones Act Sequentially to Mediate a Susceptible Tomato Pathogen Defense Response}}, year = {2003}, pages = {1181-1189}, journal = {Plant Physiol.}, doi = {10.1104/pp.103.030379}, volume = {133}, abstract = {Phytohormones regulate plant responses to a wide range of biotic and abiotic stresses. How a limited number of hormones differentially mediate individual stress responses is not understood. We have used one such response, the compatible interaction of tomato (Lycopersicon esculentum) and Xanthomonas campestris pv vesicatoria (Xcv), to examine the interactions of jasmonic acid (JA), ethylene, and salicylic acid (SA). The role of JA was assessed using an antisense allene oxide cyclase transgenic line and the def1 mutant to suppress Xcv-induced biosynthesis of jasmonates. Xcv growth was limited in these lines as was subsequent disease symptom development. No increase in JA was detected before the onset of terminal necrosis. The lack of a detectable increase in JA may indicate that an oxylipin other than JA regulates basal resistance and symptom proliferation. Alternatively, there may be an increase in sensitivity to JA or related compounds following infection. Hormone measurements showed that the oxylipin signal must precede subsequent increases in ethylene and SA accumulation. Tomato thus actively regulates the Xcv-induced disease response via the sequential action of at least three hormones, promoting expansive cell death of its own tissue. This sequential action of jasmonate, ethylene, and SA in disease symptom development is different from the hormone interactions observed in many other plant-pathogen interactions.} } @Article{IPB-2164, author = {Hause, B. and Maier, W. and Miersch, O. and Kramell, R. and Strack, D. and}, title = {{Induction of Jasmonate Biosynthesis in Arbuscular Mycorrhizal Barley Roots}}, year = {2002}, pages = {1213-1220}, journal = {Plant Physiol.}, doi = {10.1104/pp.006007}, volume = {130}, abstract = {Colonization of barley (Hordeum vulgare cv Salome) roots by an arbuscular mycorrhizal fungus, Glomus intraradices Schenck \& Smith, leads to elevated levels of endogenous jasmonic acid (JA) and its amino acid conjugate JA-isoleucine, whereas the level of the JA precursor, oxophytodienoic acid, remains constant. The rise in jasmonates is accompanied by the expression of genes coding for an enzyme of JA biosynthesis (allene oxide synthase) and of a jasmonate-induced protein (JIP23). In situ hybridization and immunocytochemical analysis revealed that expression of these genes occurred cell specifically within arbuscule-containing root cortex cells. The concomitant gene expression indicates that jasmonates are generated and act within arbuscule-containing cells. By use of a near-synchronous mycorrhization, analysis of temporal expression patterns showed the occurrence of transcript accumulation 4 to 6 d after the appearance of the first arbuscules. This suggests that the endogenous rise in jasmonates might be related to the fully established symbiosis rather than to the recognition of interacting partners or to the onset of interaction. Because the plant supplies the fungus with carbohydrates, a model is proposed in which the induction of JA biosynthesis in colonized roots is linked to the stronger sink function of mycorrhizal roots compared with nonmycorrhizal roots.} } @Article{IPB-2162, author = {Grubb, C. D. and Gross, H. B. and Chen, D. L. and Abel, S. and}, title = {{Identification of Arabidopsis mutants with altered glucosinolate profiles based on isothiocyanate bioactivity}}, year = {2002}, pages = {143-152}, journal = {Plant Sci.}, doi = {10.1016/S0168-9452(01)00550-7}, volume = {162}, abstract = {Glucosinolates are a diverse class of nitrogen- and sulfur-containing secondary metabolites. They are rapidly hydrolyzed on tissue disruption to a number of biologically active compounds that are increasingly attracting interest as anticarcinogenic phytochemicals and crop protectants. Several glucosinolate-derived isothiocyanates are potent chemopreventive agents that favorably modulate carcinogen metabolism in mammals. Methylsulfinylalkyl isothiocyanates, in particular the 4-methylsulfinylbutyl derivative, are selective and potent inducers of mammalian detoxification enzymes such as quinone reductase (QR). Cruciferous plants including Arabidopsis thaliana (L.) Heyhn, synthesize methylsulfinylalkyl glucosinolates, which are derived from methionine. Using a colorimetric assay for QR activity in murine hepatoma cells and high performance liquid chromatography (HPLC) analysis of desulfoglucosinolates, we have demonstrated a strong positive correlation between leaf QR inducer potency and leaf content of methionine-derived glucosinolates in various A. thaliana ecotypes and available glucosinolate mutants. In a molecular genetic approach to glucosinolate biosynthesis, we screened 3000 chemically mutagenized M2 plants of the Columbia ecotype for altered leaf QR inducer potency. Subsequent HPLC analysis of progeny of putative mutants identified six lines with significant and heritable changes in leaf glucosinolate content and composition.} } @Article{IPB-2156, author = {Feussner, I. and Wasternack, C. and}, title = {{The lipoxygenase pathway}}, year = {2002}, pages = {275-297}, journal = {Annu. Rev. Plant Biol.}, doi = {10.1146/annurev.arplant.53.100301.135248}, volume = {53}, abstract = {Lipid peroxidation is common to all biological systems, both appearing in developmentally and environmentally regulated processes of plants. The hydroperoxy polyunsaturated fatty acids, synthesized by the action of various highly specialized forms of lipoxygenases, are substrates of at least seven different enzyme families. Signaling compounds such as jasmonates, antimicrobial and antifungal compounds such as leaf aldehydes or divinyl ethers, and a plant-specific blend of volatiles including leaf alcohols are among the numerous products. Cloning of many lipoxygenases and other key enzymes within the lipoxygenase pathway, as well as analyses by reverse genetic and metabolic profiling, revealed new reactions and the first hints of enzyme mechanisms, multiple functions, and regulation. These aspects are reviewed with respect to activation of this pathway as an initial step in the interaction of plants with pathogens, insects, or abiotic stress and at distinct stages of development.} } @Article{IPB-2151, author = {Ellis, C. and Karafyllidis, I. and Wasternack, C. and Turner, J. G. and}, title = {{The Arabidopsis Mutant cev1 Links Cell Wall Signaling to Jasmonate and Ethylene Responses}}, year = {2002}, pages = {1557-1566}, journal = {Plant Cell}, doi = {10.1105/tpc.002022}, volume = {14}, abstract = {Biotic and abiotic stresses stimulate the synthesis of jasmonates and ethylene, which, in turn, induce the expression of genes involved in stress response and enhance defense responses. The cev1 mutant has constitutive expression of stress response genes and has enhanced resistance to fungal pathogens. Here, we show that cev1 plants have increased production of jasmonate and ethylene and that its phenotype is suppressed by mutations that interrupt jasmonate and ethylene signaling. Genetic mapping, complementation analysis, and sequence analysis revealed that CEV1 is the cellulose synthase CeSA3. CEV1 was expressed predominantly in root tissues, and cev1 roots contained less cellulose than wild-type roots. Significantly, the cev1 mutant phenotype could be reproduced by treating wild-type plants with cellulose biosynthesis inhibitors, and the cellulose synthase mutant rsw1 also had constitutive expression of VSP. We propose that the cell wall can signal stress responses in plants.} } @Article{IPB-2150, author = {Dussle, C. and Quint, M. and Xu, M. and Melchinger, A. and Lübberstedt, T. and}, title = {{Conversion of AFLP fragments tightly linked to SCMV resistance genes Scmv1 and Scmv2 into simple PCR-based markers}}, year = {2002}, pages = {1190-1195}, journal = {Theor. Appl. Genet.}, doi = {10.1007/s00122-002-0964-7}, volume = {105}, abstract = {In a previous study, bulked segregant analysis with amplified fragment length polymorphisms (AFLPs) identified several markers closely linked to the sugarcane mosaic virus resistance genes Scmv1 on chromosome 6 and Scmv2 on chromosome 3. Six AFLP markers (E33M61-2, E33M52, E38M51, E82M57, E84M59 and E93M53) were located on chromosome 3 and two markers (E33M61-1 and E35M62-1) on chromosome 6. Our objective in the present study was to sequence the respective AFLP bands in order to convert these dominant markers into more simple and reliable polymerase chain reaction (PCR)-based sequence-tagged site markers. Six AFLP markers resulted either in complete identical sequences between the six inbreds investigated in this study or revealed single nucleotide polymorphisms within the inbred lines and were, therefore, not converted. One dominant AFLP marker (E35M62-1) was converted into an insertion/deletion (indel) marker and a second AFLP marker (E33M61-2) into a cleaved amplified polymorphic sequence marker. Mapping of both converted PCR-based markers confirmed their localization to the same chromosome region (E33M61-2 on chromosome 3; E35M62-1 on chromosome 6) as the original AFLP markers. Thus, these markers will be useful for marker-assisted selection and facilitate map-based cloning of SCMV resistance genes.} } @Article{IPB-2137, author = {Bachmann, A. and Hause, B. and Maucher, H. and Garbe, E. and Vörös, K. and Weichert, H. and Wasternack, C. and Feussner, I. and}, title = {{Jasmonate-Induced Lipid Peroxidation in Barley Leaves Initiated by Distinct 13-LOX Forms of Chloroplasts}}, year = {2002}, pages = {1645-1657}, journal = {Biol. Chem.}, doi = {10.1515/BC.2002.185}, volume = {383}, abstract = {In addition to a previously characterized 13-lipoxygenase of 100 kDa encoded by LOX2:Hv:1 [Vörös et al., Eur. J. Biochem. 251 (1998), 36 44], two fulllength cDNAs (LOX2:Hv:2, LOX2:Hv:3) were isolated from barley leaves (Hordeum vulgare cv. Salome) and characterized. Both of them encode 13-lipoxygenases with putative target sequences for chloroplast import. Immunogold labeling revealed preferential, if not exclusive, localization of lipoxygenase proteins in the stroma. The ultrastructure of the chloroplast was dramatically altered following methyl jasmonate treatment, indicated by a loss of thylakoid membranes, decreased number of stacks and appearance of numerous osmiophilic globuli. The three 13-lipoxygenases are differentially expressed during treatment with jasmonate, salicylate, glucose or sorbitol. Metabolite profiling of free linolenic acid and free linoleic acid, the substrates of lipoxygenases, in water floated or jasmonatetreated leaves revealed preferential accumulation of linolenic acid. Remarkable amounts of free 9- as well as 13-hydroperoxy linolenic acid were found. In addition, metabolites of these hydroperoxides, such as the hydroxy derivatives and the respective aldehydes, appeared following methyl jasmonate treatment. These findings were substantiated by metabolite profiling of isolated chloroplasts, and subfractions including the envelope, the stroma and the thylakoids, indicating a preferential occurrence of lipoxygenasederived products in the stroma and in the envelope. These data revealed jasmonateinduced activation of the hydroperoxide lyase and reductase branch within the lipoxygenase pathway and suggest differential activity of the three 13-lipoxygenases under different stress conditions.} } @Article{IPB-2196, author = {Weichert, H. and Kolbe, A. and Kraus, A. and Wasternack, C. and Feussner, I. and}, title = {{Metabolic profiling of oxylipins in germinating cucumber seedlings - lipoxygenase-dependent degradation of triacylglycerols and biosynthesis of volatile aldehydes}}, year = {2002}, pages = {612-619}, journal = {Planta}, doi = {10.1007/s00425-002-0779-4}, volume = {215}, abstract = {A particular isoform of lipoxygenase (LOX) localized on lipid bodies was shown by earlier investigations to play a role in initiating the mobilization of triacylglycerols during seed germination. Here, further physiological functions of LOXs within whole cotyledons of cucumber (Cucumis sativus L.) were analyzed by measuring the endogenous amounts of LOX-derived products. The lipid-body LOX-derived esterified (13S)-hydroperoxy linoleic acid was the dominant metabolite of the LOX pathway in this tissue. It accumulated to about 14 µmol/g fresh weight, which represented about 6% of the total amount of linoleic acid in cotyledons. This LOX product was not only reduced to its hydroxy derivative, leading to degradation by β-oxidation, but alternatively it was metabolized by fatty acid hydroperoxide lyase leading to formation of hexanal as well. Furthermore, the activities of LOX forms metabolizing linolenic acid were detected by measuring the accumulation of volatile aldehydes and the allene oxide synthase-derived metabolite jasmonic acid. The first evidence is presented for an involvement of a lipid-body LOX form in the production of volatile aldehydes.} } @Article{IPB-2195, author = {Wang, Q. and Grubb, C. D. and Abel, S. and}, title = {{Direct analysis of single leaf disks for chemopreventive glucosinolates}}, year = {2002}, pages = {152-157}, journal = {Phytochem. Anal.}, doi = {10.1002/pca.636}, volume = {13}, abstract = {Natural isothiocyanates, produced during plant tissue damage from methionine‐derived glucosinolates, are potent inducers of mammalian phase 2 detoxification enzymes such as quinone reductase (QR). A greatly simplified bioassay for glucosinolates based on induction and colorimetric detection of QR activity in murine hepatoma cells is described. It is demonstrated that excised leaf disks of Arabidopsis thaliana (ecotype Columbia) can directly and reproducibly substitute for cell‐free leaf extracts as inducers of murine QR, which reduces sample preparation to a minimum and maximizes throughput. A comparison of 1 and 3 mm diameter leaf disks indicated that QR inducer potency was proportional to disk circumference (extent of tissue damage) rather than to area. When compared to the QR inducer potency of the corresponding amount of extract, 1 mm leaf disks were equally effective, whereas 3 mm disks were 70% as potent. The QR inducer potency of leaf disks correlated positively with the content of methionine‐derived glucosinolates, as shown by the analysis of wild‐type plants and mutant lines with lower or higher glucosinolate content. Thus, the microtitre plate‐based assay of single leaf disks provides a robust and inexpensive visual method for rapidly screening large numbers of plants in mapping populations or mutant collections and may be applicable to other glucosinolate‐producing species.} } @Article{IPB-2191, author = {Vigliocco, A. and Bonamico, B. and Alemano, S. and Miersch, O. and Abdala, G. and}, title = {{Stimulation of jasmonic acid production in Zea Mays L. infected by the maize rough dwarf virus - Río Cuarto. Reversion of symptoms by salicylic acid}}, year = {2002}, pages = {369-374}, journal = {Biocell}, url = {https://www.techscience.com/biocell/v26n3/34012}, volume = {26}, abstract = {In the present paper we study the possible biological relevance of endogenous jasmonic acid (JA) and exogenous salicylic acid (SA) in a plant-microbial system maize-virus. The virus disease \"Mal de Río Cuarto\" is caused by the maize rough dwarf virus - Río Cuarto. The characteristic symptoms are the appearance of galls or \"enations\" in leaves, shortening of the stem internodes, poor radical system and general stunting. Changes in JA and protein pattern in maize control and infected plants of a virus-tolerant cultivar were investigated. Healthy and infected-leaf discs were collected for JA measurement at different post-infection times (20, 40, 60 and 68 days). JA was also measured in roots on day 60 after infection. For SDS-PAGE protein analysis, leaf discs were also harvested on day 60 after infection. Infected leaves showed higher levels of JA than healthy leaves, and the rise in endogenous JA coincided with the enation formation. The soluble protein amount did not show differences between infected and healthy leaves; moreover, no difference in the expression of soluble protein was revealed by SDS-PAGE. Our results show that the octadecanoid pathway was stimulated in leaves and roots of the tolerant maize cultivar when infected by this virus. This finding, together with fewer plants with the disease symptoms, suggest that higher foliar and roots JA content may be related to disease tolerance. SA exogenous treatment caused the reversion of the dwarfism symptom.} } @Article{IPB-2184, author = {Schilling, S. and Hoffmann, T. and Rosche, F. and Manhart, S. and Wasternack, C. and Demuth, H.-U. and}, title = {{Heterologous Expression and Characterization of Human Glutaminyl Cyclase: Evidence for a Disulfide Bond with Importance for Catalytic Activity}}, year = {2002}, pages = {10849-10857}, journal = {Biochemistry}, doi = {10.1021/bi0260381}, volume = {41}, abstract = {Glutaminyl cyclase (QC, EC 2.3.2.5) catalyzes the formation of pyroglutamate residues from glutamine at the N-terminus of peptides and proteins. In the current study, human QC was functionally expressed in the secretory pathway of Pichia pastoris, yielding milligram quantities after purification from the supernatant of a 5 L fermentation. Initial characterization studies of the recombinant QC using MALDI-TOF mass spectrometry revealed correct proteolytic processing and N-glycosylation at both potential sites with similar 2 kDa extensions. CD spectral analysis indicated a high α-helical content, which contrasts with plant QC from Carica papaya. The kinetic parameters for conversion of H-Gln-Tyr-Ala-OH by recombinant human QC were almost identical to those previously reported for purified bovine pituitary QC. However, the results obtained for conversion of H-Gln-Gln-OH, H-Gln-NH2, and H-Gln-AMC were found to be contradictory to previous studies on human QC expressed intracellularly in E. coli. Expression of QC in E. coli showed that approximately 50% of the protein did not contain a disulfide bond that is present in the entire QC expressed in P. pastoris. Further, the enzyme was consistently inactivated by treatment with 15 mM DTT, whereas deglycosylation had no effect on enzymatic activity. Analysis of the fluorescence spectra of the native, reduced, and unfolded human QC point to a conformational change of the protein upon treatment with DTT. In terms of the different enzymatic properties, the consequences of QC expression in different environments are discussed.} } @Article{IPB-2183, author = {Schilling, S. and Hoffmann, T. and Wermann, M. and Heiser, U. and Wasternack, C. and Demuth, H.-U. and}, title = {{Continuous Spectrometric Assays for Glutaminyl Cyclase Activity}}, year = {2002}, pages = {49-56}, journal = {Anal. Biochem.}, doi = {10.1006/abio.2001.5560}, volume = {303}, abstract = {The enzymatic conversion of one chromogenic substrate, -glutamine-p-nitroanilide, and two fluorogenic substrates, -glutaminyl-2-naphthylamide and -glutaminyl-4-methylcoumarinylamide, into their respective pyroglutamic acid derivatives by glutaminyl cyclase (QC) was estimated by introducing a new coupled assay using pyroglutamyl aminopeptidase as the auxiliary enzyme. For the purified papaya QC, the kinetic parameters were found to be in the range of those previously reported for other glutaminyl peptides, such as Gln-Gln, Gln-Ala, or Gln-tert-butyl ester. The assay can be performed in the presence of ammonia up to a concentration of 50 mM. Increasing ionic strength, e.g., potassium chloride up to 300 mM, resulted in an increase in enzymatic activity of about 20%. This is the first report of a fast, continuous, and reliable determination of QC activity, even in the presence of ammonium ions, during the course of protein purification and enzymatic analysis.} } @Article{IPB-2181, author = {Quint, M. and Mihaljevic, R. and Dussle, C. and Xu, M. and Melchinger, A. and Lübberstedt, T. and}, title = {{Development of RGA-CAPS markers and genetic mapping of candidate genes for sugarcane mosaic virus resistance in maize}}, year = {2002}, pages = {355-363}, journal = {Theor. Appl. Genet.}, doi = {10.1007/s00122-002-0953-x}, volume = {105}, abstract = {Three previously published resistance gene analogues (RGAs), pic13, pic21 and pic19, were mapped in relation to sugarcane mosaic virus (SCMV) resistance genes (Scmv1, Scmv2) in maize. We cloned these RGAs from six inbreds including three SCMV-resistant lines (D21, D32, FAP1360A) and three SCMV-susceptible lines (D145, D408, F7). Pairwise sequence alignments among the six inbreds revealed a frequency of one single nucleotide polymorphism (SNP) per 33 bp for the three RGAs, indicating a high degree of polymorphism and a high probability of success in converting RGAs into codominant cleaved amplified polymorphic sequence (CAPS) markers compared to other sequences. SNPs were used to develop CAPS markers for mapping of the three RGAs in relation to Scmv1 (chromosome 6) and Scmv2 (chromosome 3), and for pedigree analyses of resistant inbred lines. By genetic mapping pic21 was shown to be different from Scmv2, whereas pic19 and pic13 are still candidates for Scmv1 and Scmv2, respectively, due to genetic mapping and consistent restriction patterns of ancestral lines.} } @Article{IPB-2179, author = {Nibbe, M. and Hilpert, B. and Wasternack, C. and Miersch, O. and Apel, K. and}, title = {{Cell death and salicylate- and jasmonate-dependent stress responses in Arabidopsis are controlled by single cet genes}}, year = {2002}, pages = {120-128}, journal = {Planta}, doi = {10.1007/s00425-002-0907-1}, volume = {216}, abstract = {The jasmonic acid (JA)-dependent regulation of the Thi2.1 gene had previously been exploited for setting up a genetic screen for the isolation of signal transduction mutants of Arabidopsis thaliana (L.) Heynh. that constitutively express the thionin gene. Several cet mutants had been isolated which showed a constitutive expression of the thionin gene. These cet mutants, except for one, also showed spontaneous leaf cell necrosis and were up-regulated in the expression of the PR1 gene, reactions often associated with the systemic acquired resistance (SAR) pathway. Four of these cet mutants, cet1, cet2, cet3 and cet4.1 were crossed with the fad triple and coi1 mutants that are blocked at two steps within the JA-dependent signaling pathway, and with transgenic NahG plants that are deficient in salicylic acid (SA) and are unable to activate SAR. Analysis of the various double-mutant lines revealed that the four cet genes act within a signaling cascade at or prior to branch points from which not only JA-dependent signals but also SA-dependent signaling and cell death pathways diverge.} } @Article{IPB-2174, author = {Laskowski, M. J. and Dreher, K. A. and Gehring, M. A. and Abel, S. and Gensler, A. L. and Sussex, I. M. and}, title = {{FQR1, a Novel Primary Auxin-Response Gene, Encodes a Flavin Mononucleotide-Binding Quinone Reductase}}, year = {2002}, pages = {578-590}, journal = {Plant Physiol.}, doi = {10.1104/pp.010581}, volume = {128}, abstract = {FQR1 is a novel primary auxin-response gene that codes for a flavin mononucleotide-binding flavodoxin-like quinone reductase. Accumulation of FQR1 mRNA begins within 10 min of indole-3-acetic acid application and reaches a maximum of approximately 10-fold induction 30 min after treatment. This increase in FQR1 mRNA abundance is not diminished by the protein synthesis inhibitor cycloheximide, demonstrating thatFQR1 is a primary auxin-response gene. Sequence analysis reveals that FQR1 belongs to a family of flavin mononucleotide-binding quinone reductases. Partially purified His-tagged FQR1 isolated fromEscherichia coli catalyzes the transfer of electrons from NADH and NADPH to several substrates and exhibits in vitro quinone reductase activity. Overexpression of FQR1 in plants leads to increased levels of FQR1 protein and quinone reductase activity, indicating that FQR1 functions as a quinone reductase in vivo. In mammalian systems, glutathione S-transferases and quinone reductases are classified as phase II detoxification enzymes. We hypothesize that the auxin-inducible glutathioneS-transferases and quinone reductases found in plants also act as detoxification enzymes, possibly to protect against auxin-induced oxidative stress.} } @Article{IPB-2134, author = {Abel, S. and Ticconi, C. A. and Delatorre, C. A. and}, title = {{Phosphate sensing in higher plants}}, year = {2002}, pages = {1-8}, journal = {Physiol. Plant.}, doi = {10.1034/j.1399-3054.2002.1150101.x}, volume = {115}, abstract = {Phosphate (Pi) plays a central role as reactant and effector molecule in plant cell metabolism. However, Pi is the least accessible macronutrient in many ecosystems and its low availability often limits plant growth. Plants have evolved an array of molecular and morphological adaptations to cope with Pi limitation, which include dramatic changes in gene expression and root development to facilitate Pi acquisition and recycling. Although physiological responses to Pi starvation have been increasingly studied and understood, the initial molecular events that monitor and transmit information on external and internal Pi status remain to be elucidated in plants. This review summarizes molecular and developmental Pi starvation responses of higher plants and the evidence for coordinated regulation of gene expression, followed by a discussion of the potential involvement of plant hormones in Pi sensing and of molecular genetic approaches to elucidate plant signalling of low Pi availability. Complementary genetic strategies in Arabidopsis thaliana have been developed that are expected to identify components of plant signal transduction pathways involved in Pi sensing. Innovative screening methods utilize reporter gene constructs, conditional growth on organophosphates and the inhibitory properties of the Pi analogue phosphite, which hold the promise for significant advances in our understanding of the complex mechanisms by which plants regulate Pi‐starvation responses.} } @Article{IPB-2133, author = {Abdala, G. and Castro, G. and Miersch, O. and Pearce, D. and}, title = {{Changes in jasmonate and gibberellin levels during development of potato plants (Solanum tuberosum)}}, year = {2002}, pages = {121-126}, journal = {Plant Growth Regul.}, doi = {10.1023/A:1015065011536}, volume = {36}, abstract = {Among the multiple environmental signals and hormonal factors regulatingpotato plant morphogenesis and controlling tuber induction, jasmonates (JAs)andgibberellins (GAs) are important components of the signalling pathways in theseprocesses. In the present study, with Solanum tuberosum L.cv. Spunta, we followed the endogenous changes of JAs and GAs during thedevelopmental stages of soil-grown potato plants. Foliage at initial growthshowed the highest jasmonic acid (JA) concentration, while in roots the highestcontent was observed in the stage of tuber set. In stolons at the developmentalstage of tuber set an important increase of JA was found; however, in tubersthere was no change in this compound during tuber set and subsequent growth.Methyl jasmonate (Me-JA) in foliage did not show the same pattern as JA; Me-JAdecreased during the developmental stages in which it was monitored, meanwhileJA increased during those stages. The highest total amount of JAs expressed asJA \+ Me-JA was found at tuber set. A very important peak ofJA in roots was coincident with that observed in stolons at tuber set. Also, aprogressive increase of this compound in roots was shown during the transitionof stolons to tubers. Of the two GAs monitored, gibberellic acid(GA3) was the most abundant in all the organs. While GA1and GA3 were also found in stolons at the time of tuber set, noothermeasurements of GAs were obtained for stolons at previous stages of plantdevelopment. Our results indicate that high levels of JA and GAs are found indifferent tissues, especially during stolon growth and tuber set.} } @Article{IPB-2242, author = {Weichert, H. and Kohlmann, M. and Wasternack, C. and Feussner, I. and}, title = {{Metabolic profiling of oxylipins upon sorbitol treatment in barley leaves}}, year = {2001}, pages = {861-862}, journal = {Biochem. Soc. Trans.}, doi = {10.1042/bst0280861}, volume = {28}, abstract = {In barley leaves 13-lipoxygenases (LOXs) are induced by salicylate and jasmonate. Here, we analyse by metabolic profiling the accumulation of oxylipins upon sorbitol treatment. Although 13-LOX-derived products are formed and specifically directed into the reductase branch of the LOX pathway, accumulation is much later than in the cases of salicylate and jasmonate treatment. In addition, under these conditions only the accumulation of jasmonates as additional products of the LOX pathway has been found.} } @Article{IPB-2239, author = {Ticconi, C. A. and Delatorre, C. A. and Abel, S. and}, title = {{Attenuation of Phosphate Starvation Responses by Phosphite in Arabidopsis}}, year = {2001}, pages = {963-972}, journal = {Plant Physiol.}, doi = {10.1104/pp.010396}, volume = {127}, abstract = {When inorganic phosphate is limiting, Arabidopsis has the facultative ability to metabolize exogenous nucleic acid substrates, which we utilized previously to identify insensitive phosphate starvation response mutants in a conditional genetic screen. In this study, we examined the effect of the phosphate analog, phosphite (Phi), on molecular and morphological responses to phosphate starvation. Phi significantly inhibited plant growth on phosphate-sufficient (2 mm) and nucleic acid-containing (2 mmphosphorus) media at concentrations higher than 2.5 mm. However, with respect to suppressing typical responses to phosphate limitation, Phi effects were very similar to those of phosphate. Phosphate starvation responses, which we examined and found to be almost identically affected by both anions, included changes in: (a) the root-to-shoot ratio; (b) root hair formation; (c) anthocyanin accumulation; (d) the activities of phosphate starvation-inducible nucleolytic enzymes, including ribonuclease, phosphodiesterase, and acid phosphatase; and (e) steady-state mRNA levels of phosphate starvation-inducible genes. It is important that induction of primary auxin response genes by indole-3-acetic acid in the presence of growth-inhibitory Phi concentrations suggests that Phi selectively inhibits phosphate starvation responses. Thus, the use of Phi may allow further dissection of phosphate signaling by genetic selection for constitutive phosphate starvation response mutants on media containing organophosphates as the only source of phosphorus.} } @Article{IPB-2201, author = {BERGER, S. and Weichert, H. and Porzel, A. and Wasternack, C. and Kühn, H. and Feussner, I. and}, title = {{Enzymatic and non-enzymatic lipid peroxidation in leaf development}}, year = {2001}, pages = {266-276}, journal = {BBA-Mol. Cell Biol. Lipids}, doi = {10.1016/S1388-1981(01)00161-5}, volume = {1533}, abstract = {Enzymatic and non-enzymatic lipid peroxidation has been implicated in programmed cell death, which is a major process of leaf senescence. To test this hypothesis we developed a high-performance liquid chromatography (HPLC) method for a simultaneous analysis of the major hydro(pero)xy polyenoic fatty acids. Quantities of lipid peroxidation products in leaves of different stages of development including natural senescence indicated a strong increase in the level of oxygenated polyenoic fatty acids (PUFAs) during the late stages of leaf senescence. Comprehensive structural elucidation of the oxygenation products by means of HPLC, gas chromatography/mass spectrometry and 1H nuclear magnetic resonance suggested a non-enzymatic origin. However, in some cases a small share of specifically oxidized PUFAs was identified suggesting involvement of lipid peroxidizing enzymes. To inspect the possible role of enzymatic lipid peroxidation in leaf senescence, we analyzed the abundance of lipoxygenases (LOXs) in rosette leaves of Arabidopsis. LOXs and their product (9Z,11E,13S,15Z)-13-hydroperoxy-9,11,15-octadecatrienoic acid were exclusively detected in young green leaves. In contrast, in senescing leaves the specific LOX products were overlaid by large amounts of stereo-random lipid peroxidation products originating from non-enzymatic oxidation. These data indicate a limited contribution of LOXs to total lipid peroxidation, and a dominant role of non-enzymatic lipid peroxidation in late stages of leaf development.} } @Article{IPB-2227, author = {Li, G. and Riaz, A. and Goyal, S. and Abel, S. and Quiros, C. F. and}, title = {{Inheritance of Three Major Genes Involved in the Synthesis of Aliphatic Glucosinolates in Brassica oleracea}}, year = {2001}, pages = {427-431}, journal = {J. Am. Soc. Hortic. Sci.}, doi = {10.21273/JASHS.126.4.427}, volume = {126}, abstract = {Inheritance of three major genes involved in synthesis of aliphatic glucosinolates (GSL) was followed in segregating populations of Brassica oleracea L. generated from three crosses: broccoli × cauliflower, collard × broccoli, and collard × cauliflower. Two of these genes, GSL-PRO and GSL-ELONG, regulate sidechain length. The action of the former results in three-carbon GSL, whereas action of the latter produces four-carbon GSL. We determined that these two genes act and segregate independently from each other in B. oleracea. The double recessive genotype produces only trace amounts of aliphatic GSL. The third gene, GSL-ALK controls sidechain desaturation and, as it has been observed in Arabidopsis thaliana (L.) Heynh., we found that this gene cosegregates with a fourth gene, GSL-OH, that is responsible for sidechain hydroxylation. Elucidation of the inheritance of major genes controlling biosynthesis of GSL will allow for manipulation of these genes and facilitate development of lines with specific GSL profiles. This capability will be important for improvement of Brassica breeding lines with high content of desirable GSL, like glucoraphanin, a demonstrated precursor of anticarcinogenic compounds. Additionally, this work is the first step towards cloning the major genes of the aliphatic GSL pathway, and to use these clones in transformation strategies for further crop enhancement.} } @Article{IPB-2220, author = {Hilpert, B. and Bohlmann, H. and Den Camp, R. o. and Przybyla, D. and Miersch, O. and Buchala, A. and Apel, K. and}, title = {{Isolation and characterization of signal transduction mutants of Arabidopsis thaliana that constitutively activate the octadecanoid pathway and form necrotic microlesions}}, year = {2001}, pages = {435-446}, journal = {Plant J.}, doi = {10.1046/j.1365-313X.2001.2641036.x}, volume = {26}, abstract = {Thionins are a group of antimicrobial polypeptides that form part of the plant\'s defense mechanism against pathogens. The Thi 2.1 thionin gene of Arabidopsis thaliana has been shown to be inducible by jasmonic acid (JA), an oxylipin‐like hormone derived from oxygenated linolenic acid and synthesized via the octadecanoid pathway. The JA‐dependent regulation of the Thi 2.1 gene has been exploited for setting up a genetic screen for the isolation of signal transduction mutants that constitutively express the Thi 2.1 gene. Ten cet‐mutants have been isolated which showed a c onstitutive e xpression of the t hionin gene. Allelism tests revealed that they represent at least five different loci. Some mutants are dominant, others recessive, but all cet mutations behaved as monogenic traits when backcrossed with Thi 2.1‐GUS plants. Some of the mutants overproduce JA and its bioactive precursor 12‐oxophytodienoic acid (OPDA) up to 40‐fold while others have the same low levels as the control wildtype plants. Two of the mutants showed a strong induction of both the salicylic acid (SA)‐ and the JA‐dependent signaling pathways, while the majority seems to be affected only in the octadecanoid pathway. The Thi 2.1 thionin gene and the Pdf 1.2 defensin gene are activated independently, though both are regulated by JA. The cet‐mutants, except for one, also show a spontaneous leaf cell necrosis, a reaction often associated with the systemic acquired resistance (SAR) pathway.} } @Article{IPB-2213, author = {Feussner, I. and Kühn, H. and Wasternack, C. and}, title = {{Lipoxygenase-dependent degradation of storage lipids}}, year = {2001}, pages = {268-273}, journal = {Trends Plant Sci.}, doi = {10.1016/S1360-1385(01)01950-1}, volume = {6}, abstract = {Oilseed germination is characterized by the mobilization of storage lipids as a carbon source for the germinating seedling. In spite of the importance of lipid mobilization, its mechanism is only partially understood. Recent data suggest that a novel degradation mechanism is initiated by a 13-lipoxygenase during germination, using esterified fatty acids specifically as substrates. This 13-lipoxygenase reaction leads to a transient accumulation of ester lipid hydroperoxides in the storage lipids, and the corresponding oxygenated fatty acid moieties are preferentially removed by specific lipases. The free hydroperoxy fatty acids are subsequently reduced to their hydroxy derivatives, which might in turn undergo β-oxidation.} } @Article{IPB-2270, author = {Maucher, H. and Hause, B. and Feussner, I. and Ziegler, J. and Wasternack, C. and}, title = {{Allene oxide synthases of barley (Hordeum vulgare cv. Salome): tissue specific regulation in seedling development}}, year = {2000}, pages = {199-213}, journal = {Plant J.}, doi = {10.1046/j.1365-313x.2000.00669.x}, volume = {21}, abstract = {Allene oxide synthase (AOS) is the first enzyme in the lipoxygenase (LOX) pathway which leads to formation of jasmonic acid (JA). Two full‐length cDNAs of AOS designated as AOS1 and AOS2, respectively, were isolated from barley (H. vulgare cv. Salome) leaves, which represent the first AOS clones from a monocotyledonous species. For AOS1, the open reading frame encompasses 1461 bp encoding a polypeptide of 487 amino acids with calculated molecular mass of 53.4 kDa and an isoelectric point of 9.3, whereas the corresponding data of AOS2 are 1443 bp, 480 amino acids, 52.7 kDa and 7.9. Southern blot analysis revealed at least two genes. Despite the lack of a putative chloroplast signal peptide in both sequences, the protein co‐purified with chloroplasts and was localized within chloroplasts by immunocytochemical analysis. The barley AOSs, expressed in bacteria as active enzymes, catalyze the dehydration of LOX‐derived 9‐ as well as 13‐hydroperoxides of polyenoic fatty acids to the unstable allene oxides. In leaves, AOS mRNA accumulated upon treatment with jasmonates, octadecanoids and metabolizable carbohydrates, but not upon floating on abscisic acid, NaCl, Na‐salicylate or infection with powdery mildew. In developing seedlings, AOS mRNA strongly accumulated in the scutellar nodule, but less in the leaf base. Both tissues exhibited elevated JA levels. In situ hybridizations revealed the preferential occurrence of AOS mRNA in parenchymatic cells surrounding the vascular bundles of the scutellar nodule and in the young convoluted leaves as well as within the first internode. The properties of both barley AOSs, their up‐regulation of their mRNAs and their tissue specific expression suggest a role during seedling development and jasmonate biosynthesis.} } @Article{IPB-2265, author = {Kramell, R. and Miersch, O. and Atzorn, R. and Parthier, B. and Wasternack, C. and}, title = {{Octadecanoid-Derived Alteration of Gene Expression and the “Oxylipin Signature” in Stressed Barley Leaves. Implications for Different Signaling Pathways}}, year = {2000}, pages = {177-188}, journal = {Plant Physiol.}, doi = {10.1104/pp.123.1.177}, volume = {123}, abstract = {Stress-induced gene expression in barley (Hordeum vulgare cv Salome) leaves has been correlated with temporally changing levels of octadecanoids and jasmonates, quantified by means of gas chromatography/mass spectrometry-single ion monitoring. Application of sorbitol-induced stress led to a low and transient rise of jasmonic acid (JA), its precursor 12-oxophytodienoic acid (OPDA), and the methyl esters JAME and OPDAME, respectively, followed by a large increase in their levels. JA and JAME peaked between 12 and 16 h, about 4 h before OPDA and OPDAME. However, OPDA accumulated up to a 2.5-fold higher level than the other compounds. Dihomo-JA and 9,13-didehydro-OPDA were identified as minor components. Kinetic analyses revealed that a transient threshold of jasmonates or octadecanoids is necessary and sufficient to initiate JA-responsive gene expression. Although OPDA and OPDAME applied exogenously were metabolized to JA in considerable amounts, both of them can induce gene expression, as evidenced by those genes that did not respond to endogenously formed JA. Also, coronatine induces JA-responsive genes independently from endogenous JA. Application of deuterated JA showed that endogenous synthesis of JA is not induced by JA treatment. The data are discussed in terms of distinct signaling pathways.} } @Article{IPB-2257, author = {Hause, B. and Stenzel, I. and Miersch, O. and Maucher, H. and Kramell, R. and Ziegler, J. and Wasternack, C. and}, title = {{Tissue-specific oxylipin signature of tomato flowers: allene oxide cyclase is highly expressed in distinct flower organs and vascular bundles}}, year = {2000}, pages = {113-126}, journal = {Plant J.}, doi = {10.1046/j.1365-313x.2000.00861.x}, volume = {24}, abstract = {A crucial step in the biosynthesis of jasmonic acid (JA) is the formation of its correct stereoisomeric precursor, cis (\+)12‐oxophytodienoic acid (OPDA). This step is catalysed by allene oxide cyclase (AOC), which has been recently cloned from tomato . In stems, young leaves and young flowers, AOC mRNA accumulates to a low level , contrasting with a high accumulation in flower buds, flower stalks and roots. The high levels of AOC mRNA and AOC protein in distinct flower organs correlate with high AOC activity, and with elevated levels of JA, OPDA and JA isoleucine conjugate. These compounds accumulate in flowers to levels of about 20 nmol g−1 fresh weight, which is two orders of magnitude higher than in leaves. In pistils, the level of OPDA is much higher than that of JA, whereas in flower stalks, the level of JA exceeds that of OPDA. In other flower tissues, the ratios among JA, OPDA and JA isoleucine conjugate differ remarkably, suggesting a tissue‐specific oxylipin signature. Immunocytochemical analysis revealed the specific occurrence of the AOC protein in ovules, the transmission tissue of the style and in vascular bundles of receptacles, flower stalks, stems, petioles and roots. Based on the tissue‐specific AOC expression and formation of JA, OPDA and JA amino acid conjugates, a possible role for these compounds in flower development is discussed in terms of their effect on sink–source relationships and plant defence reactions. Furthermore, the AOC expression in vascular bundles might play a role in the systemin‐mediated wound response of tomato.} } @Article{IPB-2255, author = {Gross, H. B. and Dalebout, T. and Grubb, C. D. and Abel, S. and}, title = {{Functional detection of chemopreventive glucosinolates in Arabidopsis thaliana}}, year = {2000}, pages = {265-272}, journal = {Plant Sci.}, doi = {10.1016/S0168-9452(00)00354-X}, volume = {159}, abstract = {Natural isothiocyanates, derived from glucosinolates by myrosinase-catalyzed hydrolysis, are potent chemopreventive agents that favorably modify carcinogen metabolism in mammals by inhibiting metabolic activation of carcinogens and/or by inducing carcinogen-detoxifying enzymes. Methylsulfinylalkyl isothiocyanates are potent selective inducers of mammalian Phase 2 detoxification enzymes such as quinone reductase [NADP(H):quinone-acceptor oxidoreductase, EC 1.6.99.2]. Members of the Cruciferae family, including the model plant species Arabidopsis thaliana (L.) Heyhn, synthesize methylsulfinylalkyl glucosinolates. We have adapted a colorimetric bioassay for quinone reductase activity in Hepa 1c1c7 murine hepatoma cells as a versatile tool to rapidly monitor methylsulfinylalkyl glucosinolate content in A. thaliana leaf extracts. Using wild type plants and mutant plants defective in the synthesis of 4-methylsulfinylbutyl glucosinolate (glucoraphanin), we have demonstrated that A. thaliana (ecotype Columbia) is a rich source of Phase 2 enzyme inducers and that methylsulfinylalkyl glucosinolates, predominantly glucoraphanin, account for about 80% of the quinone reductase inducer potency of Columbia leaf extracts. We have optimized leaf extraction conditions and the quinone reductase bioassay to allow for screening of large numbers of plant extracts in a molecular genetic approach to dissecting glucosinolate biosynthesis in A. thaliana.} } @Article{IPB-2253, author = {Colón-Carmona, A. and Chen, D. L. and Yeh, K.-C. and Abel, S. and}, title = {{Aux/IAA Proteins Are Phosphorylated by Phytochrome in Vitro}}, year = {2000}, pages = {1728-1738}, journal = {Plant Physiol.}, doi = {10.1104/pp.124.4.1728}, volume = {124}, abstract = {Auxin/indole-3-acetic acid (Aux/IAA) genes encode short-lived transcription factors that are induced as a primary response to the plant growth hormone IAA or auxin. Gain-of-function mutations in Arabidopsis genes,SHY2/IAA3, AXR3/IAA17, andAXR2/IAA7 cause pleiotropic phenotypes consistent with enhanced auxin responses, possibly by increasing Aux/IAA protein stability. Semidominant mutations shy2-1D,shy2-2, axr3-1, and axr2-1induce ectopic light responses in dark-grown seedlings. Because genetic studies suggest that the shy2-1D andshy2-2 mutations bypass phytochrome requirement for certain aspects of photomorphogenesis, we tested whether SHY2/IAA3 and related Aux/IAA proteins interact directly with phytochrome and whether they are substrates for its protein kinase activity. Here we show that recombinant Aux/IAA proteins from Arabidopsis and pea (Pisum sativum) interact in vitro with recombinant phytochrome A from oat (Avena sativa). We further show that recombinant SHY2/IAA3, AXR3/IAA17, IAA1, IAA9, and Ps-IAA4 are phosphorylated by recombinant oat phytochrome A in vitro. Deletion analysis of Ps-IAA4 indicates that phytochrome A phosphorylation occurs on the N-terminal half of the protein. Metabolic labeling and immunoprecipitation studies with affinity-purified antibodies to IAA3 demonstrate increased in vivo steady-state levels of mutant IAA3 in shy2-2 plants and phosphorylation of the SHY2-2 protein in vivo. Phytochrome-dependent phosphorylation of Aux/IAA proteins is proposed to provide one molecular mechanism for integrating auxin and light signaling in plant development.} } @Article{IPB-2252, author = {Chen, D. L. and Delatorre, C. A. and Bakker, A. and Abel, S. and}, title = {{Conditional identification of phosphate-starvation-response mutants in Arabidopsis thaliana}}, year = {2000}, pages = {13-22}, journal = {Planta}, doi = {10.1007/s004250000271}, volume = {211}, abstract = {Plants have evolved elaborate metabolic and developmental adaptations to low phosphorus availability. Biochemical responses to phosphate limitation include increased production and secretion of phosphate-acquisition proteins such as nucleases, acid phosphatases, and high-affinity phosphate transporters. However, the signal transduction pathways that sense phosphate availability and integrate the phosphate-starvation response in plants are unknown. We have devised a screen for conditional mutants in Arabidopsis thaliana (L.) Heynh. to dissect signaling of phosphate limitation. Our genetic screen is based on the facultative ability of wild-type Arabidopsis plants to metabolize exogenous DNA when inorganic phosphate is limiting. After screening 50,000 M2 seedlings, we isolated 22 confirmed mutant lines that showed severely impaired growth on medium containing DNA as the only source of phosphorus, but which recovered on medium containing soluble inorganic phosphate. Characterization of nine such mutant lines demonstrated an inability to utilize either DNA or RNA. One mutant line, psr1 (phosphate starvation response), had significantly reduced activities of phosphate-starvation-inducible isoforms of ribonuclease and acid phosphatase under phosphate-limiting conditions. The data suggest that a subset of the selected mutations impairs the expression of more than one phosphate-starvation-inducible enzyme required for utilization of exogenous nucleic acids, and may thus affect regulatory components of a Pi starvation response pathway in higher plants.} } @Article{IPB-2244, author = {Abel, S. and Nürnberger, T. and Ahnert, V. and Krauss, G.-J. and Glund, K. and}, title = {{Induction of an Extracellular Cyclic Nucleotide Phosphodiesterase as an Accessory Ribonucleolytic Activity during Phosphate Starvation of Cultured Tomato Cells}}, year = {2000}, pages = {543-552}, journal = {Plant Physiol.}, doi = {10.1104/pp.122.2.543}, volume = {122}, abstract = {During growth under conditions of phosphate limitation, suspension-cultured cells of tomato (Lycopersicon esculentum Mill.) secrete phosphodiesterase activity in a similar fashion to phosphate starvation-inducible ribonuclease (RNase LE), a cyclizing endoribonuclease that generates 2′:3′-cyclic nucleoside monophosphates (NMP) as its major monomeric products (T. Nürnberger, S. Abel, W. Jost, K. Glund [1990] Plant Physiol 92: 970–976). Tomato extracellular phosphodiesterase was purified to homogeneity from the spent culture medium of phosphate-starved cells and was characterized as a cyclic nucleotide phosphodiesterase. The purified enzyme has a molecular mass of 70 kD, a pH optimum of 6.2, and an isoelectric point of 8.1. The phosphodiesterase preparation is free of any detectable deoxyribonuclease, ribonuclease, and nucleotidase activity. Tomato extracellular phosphodiesterase is insensitive to EDTA and hydrolyzes with no apparent base specificity 2′:3′-cyclic NMP to 3′-NMP and the 3′:5′-cyclic isomers to a mixture of 3′-NMP and 5′-NMP. Specific activities of the enzyme are 2-fold higher for 2′:3′-cyclic NMP than for 3′:5′-cyclic isomers. Analysis of monomeric products of sequential RNA hydrolysis with purified RNase LE, purified extracellular phosphodiesterase, and cleared −Pi culture medium as a source of 3′-nucleotidase activity indicates that cyclic nucleotide phosphodiesterase functions as an accessory ribonucleolytic activity that effectively hydrolyzes primary products of RNase LE to substrates for phosphate-starvation-inducible phosphomonoesterases. Biosynthetical labeling of cyclic nucleotide phopshodiesterase upon phosphate starvation suggests de novo synthesis and secretion of a set of nucleolytic enzymes for scavenging phosphate from extracellular RNA substrates.} } @Article{IPB-2292, author = {Ziegler, J. and Stenzel, I. and Hause, B. and Maucher, H. and Hamberg, M. and Grimm, R. and Ganal, M. and Wasternack, C. and}, title = {{Molecular Cloning of Allene Oxide Cyclase}}, year = {2000}, pages = {19132-19138}, journal = {J. Biol. Chem.}, doi = {10.1074/jbc.M002133200}, volume = {275}, abstract = {Allene oxide cyclase (EC 5.3.99.6) catalyzes the stereospecific cyclization of an unstable allene oxide to (9S,13S)-12-oxo-(10,15Z)-phytodienoic acid, the ultimate precursor of jasmonic acid. This dimeric enzyme has previously been purified, and two almost identical N-terminal peptides were found, suggesting allene oxide cyclase to be a homodimeric protein. Furthermore, the native protein was N-terminally processed. Using degenerate primers, a polymerase chain reaction fragment could be generated from tomato, which was further used to isolate a full-length cDNA clone of 1 kilobase pair coding for a protein of 245 amino acids with a molecular mass of 26 kDa. Whereas expression of the whole coding region failed to detect allene oxide cyclase activity, a 5′-truncated protein showed high activity, suggesting that additional amino acids impair the enzymatic function. Steric analysis of the 12-oxophytodienoic acid formed by the recombinant enzyme revealed exclusive (\>99%) formation of the 9S,13Senantiomer. Exclusive formation of this enantiomer was also found in wounded tomato leaves. Southern analysis and genetic mapping revealed the existence of a single gene for allene oxide cyclase located on chromosome 2 of tomato. Inspection of the N terminus revealed the presence of a chloroplastic transit peptide, and the location of allene oxide cyclase protein in that compartment could be shown by immunohistochemical methods. Concomitant with the jasmonate levels, the accumulation of allene oxide cyclase mRNA was transiently induced after wounding of tomato leaves.} } @Article{IPB-2289, author = {Weichert, H. and Kolbe, A. and Wasternack, C. and Feussner, I. and}, title = {{Formation of 4-hydroxy-2-alkenals in barley leaves}}, year = {2000}, pages = {850-851}, journal = {Biochem. Soc. Trans.}, doi = {10.1042/bst0280850}, volume = {28}, abstract = {In barley leaves 13-lipoxygenases are induced by jasmonates. This leads to induction of lipid peroxidation. Here we show by in vitro studies that these processes may further lead to autoxidative formation of (2E)-4-hydroxy-2-hexenal from (3Z)-hexenal.} } @Article{IPB-2288, author = {Wasternack, C. and Hause, B. and}, title = {{Stressabwehr und Entwicklung: Jasmonate — chemische Signale in Pflanzen}}, year = {2000}, pages = {312-320}, journal = {Biologie in unserer Zeit}, doi = {10.1002/1521-415X(200011)30:6<312::AID-BIUZ312>3.0.CO;2-8}, volume = {30}, abstract = {Chemische Signale wurden bereits im 19.Jahrhundert als Regulatoren von Wachstum und Entwicklung der Pflanzen postuliert.In den letzten 70 Jahren wurde die Wirkungsweise der klassischen Pflanzenhormone wie der Auxine, Gibberelline, Cytokinine, Ethylen und Abscisinsäure aufgeklärt. Doch erst im letzten Jahrzehnt entdeckte man die Bedeutung der Brassinosteroide, der Peptidhormone und der Jasmonate.} } @Article{IPB-2278, author = {Quint, M. and Melchinger, A. E. and Dußle, C. M. and Lübberstedt, T. and}, title = {{Breeding for virus resistance in maize}}, year = {2000}, pages = {529-545}, journal = {Genetika}, url = {http://www.dgsgenetika.org.rs/abstrakti/vol32_2000_no3_en.htm#Rad26}, volume = {32}, abstract = {Sugarcane mosaic virus (SCMV) is an important disease in maize, which is emerging in Germany since 1983. Using this pest as a model for the inheritance of oligogenic traits, we clarified the genetic ba­sis for resistance in early maturing European maize germplasm. Screening of 122 adapted European inbred lines identified three completely resistant lines, which were used for further analyses. The genetics of SCMV resis­tance was investigated by allelism tests in field experiments combined with QTL and bulked segregant analyses (BSA) on the marker level. QTL analyses revealed the presence of two major genes Scm1 and Scm2 plus three minor QTL. Involvement of Scm1 and Scm2 in the inheritance of SCMV resistance could be confirmed by BSA in a second cross. Breeders can make use of tightly linked STS markers for marker-assisted selection (MAS) as well as our SCMV resistant flint lines to improve their elite germplasm. Currently, recurrent backcrossing with phenotypic selection is the most appropriate and cost effective breeding method. With de­creasing costs of DNA chip technology, MAS can be competitive with phenotypic selection in the near future. Further objectives of our research are the isolation and cloning of Scm1 and Scm2. To achieve this goal we follow two different approaches. (1) Positional cloning based on more than 500 AFLP primer combinations resulted in Scm1/Scm2 specific markers with a resolution of approximately 0.2 cM in the respective re­gions. (2) Resistance gene analogues (RGAs), cosegregating with the tar­get genes are used to identify further candidate genes for transformation experiments.} } @Article{IPB-2271, author = {Miersch, O. and Wasternack, C. and}, title = {{Octadecanoid and Jasmonate Signaling in Tomato (Lycopersicon esculentum Mill.) Leaves: Endogenous Jasmonates Do Not Induce Jasmonate Biosynthesis}}, year = {2000}, pages = {715-722}, journal = {Biol. Chem.}, doi = {10.1515/BC.2000.092}, volume = {381}, abstract = {Jasmonates and their precursors, the octadecanoids, are signals in stress-induced alteration of gene expression. Several mRNAs coding for enzymes of jasmonic acid (JA) biosynthesis are up-regulated upon JA treatment or endogenous increase of the JA level. Here we investigated the positive feedback of endogenous JA on JA formation, as well as its β-oxidation steps. JA-responsive gene expression was recorded in terms of proteinase inhibitor2 (pin2) mRNA accumulation. JA formed upon treatment of tomato (Lycopersicon esculentum cv. Moneymaker) leaves with JA derivatives carrying different lengths of the carboxylic acid side chain was quantified by gas chromatography-mass spectrometry (GC-MS). The data revealed that β-oxidation of the side chain occurs up to a butyric acid moiety. The amount of JA formed from side-chain modified JA derivatives correlated with pin2-mRNA accumulation. JA derivatives with a carboxylic side chain of 3, 5 or 7 carbon atoms were unable to form JA and to express on pin2, whereas evennumbered derivatives were active.After treatment of tomato leaves with (10-2H)-(–)-12-oxophytoenoic acid, (4-2H)-(–)-JA and its methyl ester were formed and could be quantified separately from the endogenously nonlabeled JA pool by GC-MS analysis via isotopic discrimination. The level of 8 nmol per g fresh weight JA and its methyl ester originated exclusively from labeled 12-oxophytoenic acid. This and further data indicate that endogenous synthesis of the JA precursor 12-oxophytodienoic acid, as well as of JA and its methyl ester, are not induced in tomato leaves, suggesting that positive feedback in JA biosynthesis does not function in vivo.} } @Article{IPB-2322, author = {Ortel, B. and Atzorn, R. and Hause, B. and Feussner, I. and Miersch, O. and Wasternack, C. and}, title = {{Jasmonate-induced gene expression of barley (Hordeum vulgare) leaves - the link between jasmonate and abscisic acid}}, year = {1999}, pages = {113-122}, journal = {Plant Growth Regul.}, doi = {10.1023/A:1006212017458}, volume = {29}, abstract = {In barley leaves a group of genes is expressed in response to treatment with jasmonates and abscisic acid (ABA) [21]. One of these genes coding for a jasmonate-induced protein of 23 kDa (JIP-23) was analyzed to find out the link between ABA and jasmonates by recording its expression upon modulating independently, the endogenous level of both of them. By use of inhibitors of JA synthesis and ABA degradation, and the ABA-deficient mutant Az34, as well as of cultivar-specific differences, it was shown that endogenous jasmonate increases are necessary and sufficient for expression of this gene. The endogenous rise of ABA did not induce synthesis of JIP-23, whereas exogenous ABA did not act via jasmonates. Different signalling pathways are suggested and discussed.} } @Article{IPB-2320, author = {Morgan, K. E. and Zarembinski, T. I. and Theologis, A. and Abel, S. and}, title = {{Biochemical characterization of recombinant polypeptides corresponding to the predicted βαα fold in Aux/IAA proteins}}, year = {1999}, pages = {283-287}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(99)00819-4}, volume = {454}, abstract = {The plant hormone indoleacetic acid (IAA or auxin) transcriptionally activates a select set of early genes. The Auxl IAA class of early auxin-responsive genes encodes a large family of short-lived, nuclear proteins. Aux/IAA polypeptides homo-and heterodimerize, and interact with auxin-response transcription factors (ARFs) via C-terminal regions conserved in both protein families. This shared region contains a predicted βαα motif similar to the prokaryotic β-Ribbon DNA binding domain, which mediates both protein dimerization and DNA recognition. Here, we show by circular dichroism spectroscopy and by chemical cross-linking experiments that recombinant peptides corresponding to the predicted βαα region of three Aux/IAA proteins from Arabidopsis thaliana contain substantial α-helical secondary structure and undergo homo- and heterotypic interactions in vitro. Our results indicate a similar biochemical function of the plant βαα domain and suggest that the βαα fold plays an important role in mediating combinatorial interactions of Aux/IAA and ARF proteins to specifically regulate secondary gene expression in response to auxin.} } @Article{IPB-2319, author = {Miersch, O. and Kramell, R. and Parthier, B. and Wasternack, C. and}, title = {{Structure–activity relations of substituted, deleted or stereospecifically altered jasmonic acid in gene expression of barley leaves}}, year = {1999}, pages = {353-361}, journal = {Phytochemistry}, doi = {10.1016/S0031-9422(98)00597-4}, volume = {50}, abstract = {Jasmonic acid and 66 structurally related compounds were tested to find the structural requirements which induce the expression of jasmonate-responsive genes in barley. An intact cyclopentanone ring as well as a pentenyl side chain exhibiting only minor alterations are necessary for this activity. The (−)-enantiomeric and the (\+)-7-iso-enantiomeric structure increase activity of jasmonoyl compounds.} } @Article{IPB-2318, author = {Miersch, O. and Porzel, A. and Wasternack, C. and}, title = {{Microbial conversion of jasmonates - hydroxylations by Aspergillus niger}}, year = {1999}, pages = {1147-1152}, journal = {Phytochemistry}, doi = {10.1016/S0031-9422(98)00698-0}, volume = {50}, abstract = {Aspergillus niger is able to hydroxylate the pentenyl side chain of (−)-jasmonic acid (JA) leading to (11S)- (−)-hydroxy-JA/ (11R)- (−)-hydroxy-JA (2:1) and (−)-11,12-didehydro-JA. Methyl (−)-jasmonate (JA-Me) is converted upon hydrolysis. During prolonged cultivation or at non-optimized isolation procedures, the 11-hydroxy- (9Z)-pentenyl side chain may isomerize to (10E)-9-hydroxy- and (9E)-11-hydroxy-compounds by allylic rearrangement. The fungus hydroxylates (±)-9,10-dihydro-JA at position C-11 into 11j-hydroxy-9,10-dihydro-JA. As JA-Me, the methyl dihydro-JA is hydroxylated only upon hydrolysis into the free acid.} } @Article{IPB-2317, author = {Miersch, O. and Bohlmann, H. and Wasternack, C. and}, title = {{Jasmonates and related compounds from Fusarium oxysporum}}, year = {1999}, pages = {517-523}, journal = {Phytochemistry}, doi = {10.1016/S0031-9422(98)00596-2}, volume = {50}, abstract = {The culture filtrate of Fusarium oxysporum f sp matthiolae was inspected on the occurrence of jasmonates and related compounds. Among compounds described for the first time of biological origin are 7-iso-cucurbic acid, (1S,2S)- and (1S,2R)-3-oxo-2-pentylcyclopentane-1-butyric acid, (1S,2S)- and (1S,2R)-3-oxo-2-(2Z-pentenyl)cyclopentane-1-hexanoic acid, (1S,2S)- and (1S,2R)-3-oxo-2-pentylcyclopentane-1-hexanoic acid, (1S,2S)-3-oxo-2-(2Z-pentenyl)cyclopentane-1-octanoic acid, (1S,2S)-3-oxo-2-pentylcyclopentane-1-octanoic acid and N-[9,10-dihydro-7-iso-jasmonoyl]-(S)-isoleucine. The following metabolites were identified for the first time for this fungus: (−)-Jasmonic acid, 9,10-dihydrojasmonic acid and N-[(−)-jasmonoyl-(S)]-isoleucine were major constituents of the culture filtrate, whereas as minor metabolites occurred N-[9,10-dihydrojasmonoyl]-(S)-isoleucine, cucurbic acid and 3-oxo-2-(2Z-pentenyl)cyclopentane-1-butyric acid, 3-oxo-2-(2Z-pentenyl)cyclopentane-1-octanoic acid and 3-oxo-2-pentylcyclopentane-1-octanoic acid. All cyclopentanones found carried a cis- or trans-attached side chain. Didehydro-jasmonates, hydroxylated jasmonates or 12-oxophytodienoic acid could not be detected in the culture filtrate.} } @Article{IPB-2313, author = {Kramell, R. and Miersch, O. and Schneider, G. and Wasternack, C. and}, title = {{Liquid chromatography of jasmonic acid amine conjugates}}, year = {1999}, pages = {42-46}, journal = {Chromatographia}, doi = {10.1007/BF02467185}, volume = {49}, abstract = {Racemic jasmonic acid (3R,7R/3S,7S)-(±)-JA) was chemically conjugated with different biogenic amines originating from aliphatic and aromatic α-amino acids by decarboxylation. The resulting isomeric compounds were subjected to reversed-phase high-performance liquid chromatography (HPLC) and to HPLC on the chiral stationary phases Chiralpak AS and Nucleodex β-PM. Under reversed-phase conditions, all the homologous amine derivatives tested could be separated from each other except the JA-conjugates containing 2-phenyl-ethylamine and 3-methylbutylamine. On both chiral supports the (3R,7R)-(−)-JA conjugates eluted earlier than those of the enantiomeric counterpart (3S,7S)-(\+)-JA. On Chiralpak AS all the isomers studied could be separated to baseline with a mobile phase containingn-hexane and 2-propanol. The calculated resolution factors were between 1.80 and 4.17. The pairs of isomers were also chromatographed on the cyclodextrin stationary phase Nucleodex β-PM with methanol-triethylammonium acetate buffer as mobile phase. Under these conditions resolution factors were between 0.74 and 1.29. The individual isomers were chiroptically characterized by measurement of their circular dichroism.} } @Article{IPB-2312, author = {Kramell, R. and Porzel, A. and Miersch, O. and Schneider, G. and Wasternack, C. and}, title = {{Chromatographic resolution of peptide-like conjugates of jasmonic acid and of cucurbic acid isomers}}, year = {1999}, pages = {103-107}, journal = {J. Chromatogr. A}, doi = {10.1016/S0021-9673(99)00335-0}, volume = {847}, abstract = {The chiral separation of peptide-like conjugates of jasmonic acid and of cucurbic acid isomers was investigated by liquid chromatography on Chiralpak AS and Nucleodex β-PM. The retention sequences reflect distinct chromatographic properties with respect to the chirality of the jasmonic acid part or of the cucurbic acid isomers. The chromatographic behaviour of the amide conjugates on a reversed-phase C18 column provides evidence for the resolution of diastereomeric conjugates depending on the chirality of both constituents of the conjugate molecule. The chromatographic procedures are suitable for the analytical and preparative separation of such conjugates.} } @Article{IPB-2310, author = {Kenton, P. and Mur, L. A. J. and Atzorn, R. and Wasternack, C. and Draper, J. and}, title = {{(—)-Jasmonic Acid Accumulation in Tobacco Hypersensitive Response Lesions}}, year = {1999}, pages = {74-78}, journal = {Mol. Plant Microbe Interact.}, doi = {10.1094/MPMI.1999.12.1.74}, volume = {12}, abstract = {Tobacco infected with Pseudomonas syringae pv. phaseolicola undergoes a hypersensitive response (HR). Jasmonic acid (JA) accumulated within the developing lesion 3 to 9 h after infection and this accumulation preceded protein loss, cell death, and malondialdehyde accumulation. Accumulating JA consisted largely of the (—)-JA stereoisomer and was essentially restricted to the HR lesion.} } @Article{IPB-2309, author = {Herde, O. and Peña Cortés, H. and Wasternack, C. and Willmitzer, L. and Fisahn, J. and}, title = {{Electric Signaling and Pin2 Gene Expression on Different Abiotic Stimuli Depend on a Distinct Threshold Level of Endogenous Abscisic Acid in Several Abscisic Acid-Deficient Tomato Mutants}}, year = {1999}, pages = {213-218}, journal = {Plant Physiol.}, doi = {10.1104/pp.119.1.213}, volume = {119}, abstract = {Experiments were performed on three abscisic acid (ABA)-deficient tomato (Lycopersicon esculentum Mill.) mutants, notabilis,flacca, and sitiens, to investigate the role of ABA and jasmonic acid (JA) in the generation of electrical signals and Pin2 (proteinaseinhibitor II) gene expression. We selected these mutants because they contain different levels of endogenous ABA. ABA levels in the mutant sitiens were reduced to 8% of the wild type, in notabilis they were reduced to 47%, and in flacca they were reduced to 21%. In wild-type and notabilis tomato plants the induction ofPin2 gene expression could be elicited by heat treatment, current application, or mechanical wounding. Inflacca and sitiens only heat stimulation induced Pin2 gene expression. JA levels inflacca and sitiens plants also accumulated strongly upon heat stimulation but not upon mechanical wounding or current application. Characteristic electrical signals evolved in the wild type and in the notabilis andflacca mutants consisting of a fast action potential and a slow variation potential. However, in sitiens only heat evoked electrical signals; mechanical wounding and current application did not change the membrane potential. In addition, exogenous application of ABA to wild-type tomato plants induced transient changes in membrane potentials, indicating the involvement of ABA in the generation of electrical signals. Our data strongly suggest the presence of a minimum threshold value of ABA within the plant that is essential for the early events in electrical signaling and mediation of Pin2 gene expression upon wounding. In contrast, heat-induced Pin2 gene expression and membrane potential changes were not dependent on the ABA level but, rather, on the accumulation of JA.} } @Article{IPB-2308, author = {Hause, B. and Hertel, S. C. and Klaus, D. and Wasternack, C. and}, title = {{Cultivar-Specific Expression of the Jasmonate-Induced Protein of 23 kDa (JIP-23) Occurs in Hordeum vulgare L. by Jasmonates but not During Seed Germination}}, year = {1999}, pages = {83-89}, journal = {Plant Biol.}, doi = {10.1111/j.1438-8677.1999.tb00712.x}, volume = {1}, abstract = {Treatment of barley leaf segments with jasmonic acid methyl ester (JM) leads to the accumulation of a set of newly formed abundant proteins. Among them, the most abun dant protein exhibits a molecular mass of 23 kDa (JIP‐23). Here, data are presented on the occurrence and expression of the lIP‐23 genes in different cultivars of Hordeum vulgare . Southern blot analysis of 80 cultivars revealed the occurrence of 2 to 4 genes coding for JIP‐23 in all cultivars. By means of Northern blot and immunoblot analysis it is shown that some cultivars lack the ex pression of jip‐23 upon treatment of primary leaves with JM as well as upon stress performed by incubation with 1 M sorbitol solution. During germination, however, all tested cultivars ex hibited developmental expression of jip‐23 . The results are dis cussed in terms of possible functions of JIP‐23 in barley.} } @Article{IPB-2307, author = {Hause, B. and Vörös, K. and Kogel, K.-H. and Besser, K. and Wasternack, C. and}, title = {{A Jasmonate-responsive Lipoxygenase of Barley Leaves is Induced by Plant Activators but not by Pathogens}}, year = {1999}, pages = {459-462}, journal = {J. Plant Physiol.}, doi = {10.1016/S0176-1617(99)80283-1}, volume = {154}, abstract = {Using the recently isolated eDNA clone LOX2 : Hv : 1 which codes for the most abundant jasmonateinducible lipoxygenase (LOX) in barley leaves (Vörös et al., 1998), we analysed the capability of different activators of systemic activated resistance (SAR) to induce the expression of that LOX. Upon treatment of barley leaves with salicylate, 2,6-dichloroisonicotinic acid and benzo-(1,2,3)-thiadiazole-7-carbothioic acid S-methyl ester, all these compounds were able to induce the expression of the LOX2 : Hv : 1 gene, whereas upon infection with the powdery mildew fungus (Blumeria graminis f. sp. hordei) mRNA accumulation was not detectable in compatible or in incompatible interactions. The induction of the LOX2 : Hv : 1 protein by SAR activators and the expression of different sets of genes induced by jasmonate and salicylate, respectively, are discussed in relation to defense responses against pathogenic fungi.} } @Article{IPB-2305, author = {Gago-Zachert, S. and Costa, N. and Semorile, L. and Grau, O. and}, title = {{Sequence variability in p27 gene of Citrus Tristeza Virus (CTV) revealed by SSCP analysis}}, year = {1999}, pages = {41-50}, journal = {Electron. J. Biotechnol.}, doi = {10.2225/vol2-issue1-fulltext-3}, volume = {2}, abstract = {Citrus tristeza closterovirus (CTV), is a phloem-limited virus transmitted by aphids in a semipersistent manner. The genome of CTV is composed of a ssRNA with two capsid proteins: CP, covering about 95% of the particle length, and a diverged coat protein (dCP), present only in one end of the particle, forming a rattlesnake structure. dCP is the product of p27 gene for which it is also postulated a function in the transmissibility by aphid vectors. Hybridization analysis showed a p27 gene region, which exhibits different patterns with two probes derived from two biological distinct CTV isolates. In an attempt to screen whether that gene region differs in mild and severe strains, six CTV isolates belonging to different biogroups were compared for variations in their p27 gene by analysis of single-strand conformation polymorphism (SSCP). The p27 gene was reverse transcribed and amplified by PCR and thirty clones of each isolate were obtained. From each clone, two fragments of the gene were amplified by PCR: fragment (a), 459 bp long, and fragment (b), 281 bp long. Sequence variations in both gene fragments were studied by SSCP analysis. A variety of SSCP patterns was obtained from each isolate, being isolates belonging to the groups II-IV and III those with the higher and lower number of them. Moreover, SSCP analysis provided a rapid procedure to screen the genetic heterogeneity of the viral isolates reducing considerably the amount of nucleic acid sequenciation necessary to gain that knowledge.} } @Article{IPB-2364, author = {Ratajczak, R. and Feussner, I. and Hause, B. and Böhm, A. and Parthier, B. and Wasternack, C. and}, title = {{Alteration of V-type H\+-ATPase during methyljasmonate-induced senescence in barley (Hordeum vulgare L. cv. Salome)}}, year = {1998}, pages = {199-206}, journal = {J. Plant Physiol.}, doi = {10.1016/S0176-1617(98)80133-8}, volume = {152}, abstract = {In barley leaves, the application of (−)-jasmonic acid or its methyl ester (JAME) induces a senescencelike phenotype. This is accompanied by the synthesis of abundant proteins, so-called jasmonate-induced proteins (JlPs). Here, we show that modifications of vacuolar H\+-ATPase (V-ATPase) subunits are jasmo-nate inducible. Using immunofluorescence analysis, we demonstrate that V-ATPase of barley leaves is exclusively located at the tonoplast also upon JAME treatment. Total ATP-hydrolysis activity of microsomal fractions increased by a factor of 10 during 72 h of JAME-treatment, while Bafilomycin Ai-sensitive ATP-hydrolysis activity, which is usually referred to V-ATPase activity, increased by a factor of about 2 in tono-plast-enriched membrane fractions. Moreover, due to JAME treatment there was a pronounced increase in ATP-hydrolysis activity at pH 6.2. This activity was not affected by inhibitors of P-, F-, or V-ATPases. However, biochemical analysis of partially purified V-ATPase suggests, that this activity might be due at least in part to the V-ATPase. JAME-treatment seems to change biochemical properties of the V-ATPase, i.e. a shift of the pH optimum of activity to a more acidic pH and a decrease in Bafilomycin A1 sensitivity. This is accompanied by the appearance of several additional forms of V-ATPase subunits which might represent either different isoforms or post-translationally modified proteins. We suggest that these changes in properties of the V-ATPase, which is involved in house-keeping and stress responses, may be due to JAME-induced senescence to overcome concomitant changes of the vacuolar membrane.} } @Article{IPB-2357, author = {Miersch, O. and Knöfel, H.-D. and Schmidt, J. and Kramell, R. and Parthier, B. and}, title = {{A jasmonic acid conjugate, N-[(—)-jasmonoyl]-tyramine, from Petunia pollen}}, year = {1998}, pages = {327-329}, journal = {Phytochemistry}, doi = {10.1016/S0031-9422(97)00617-1}, volume = {47}, abstract = {A new jasmonate, N-[(—)-jasmonoyl]-tyramine, was identified from petunia pollen in which (—)-jasmonic acid was detected and quantified.} } @Article{IPB-2351, author = {Feussner, I. and Wasternack, C. and}, title = {{Lipoxygenase catalyzed oxygenation of lipids}}, year = {1998}, pages = {146-152}, journal = {Fett/Lipid}, doi = {10.1002/(SICI)1521-4133(19985)100:4/5<146::AID-LIPI146>3.0.CO;2-D}, volume = {100}, abstract = {Lipoxygenases (LOXs) and other LOX pathway enzymes are potentially able to form a large set of compounds being of commercial interest. Among them are conjugated dienic acids, jasmonates, and volatile aldehydes. Additionally, fatty acid hydroperoxides, formed by LOX, can serve as precursors for further transformation by either enzymes of the so‐called LOX pathway or by chemical reactions. In the case of linoleic acid more than one hundred products generated from its LOX‐derived fatty acid hydroperoxides have been described. Many of these products exhibit biological activity, suggesting a significant biological function of LOXs. This will be described for two different 13‐LOXs. (I) In various oilseeds we found that specific 13‐LOXs are localized at the lipid body membrane. They are capable of oxygenating esterified polyenoic fatty acids, such as triacylglycerols and phospho‐lipids. In addition, they form with arachidonic acid as substrate preferentially either 8‐ or 11‐hydroperoxy eicosatetraenoic acid, which is a very unusual positional specificity for plant LOXs. (II) From barley leaves we isolated another linoleate 13‐LOX form, which is localized within chloroplasts and is induced by jasmonic acid methyl ester. It is suggested, that this LOX form is capable of oxygenating linolenic acid residues of galactolipids. Examples will be presented for barley leaves of oxygenated derivatives of linolenic acid and compounds resulting from the hydroperoxide lyase‐branch of the LOX pathway.} } @Article{IPB-2346, author = {Churin, J. and Hause, B. and Feussner, I. and Maucher, H. P. and Feussner, K. and Börner, T. and Wasternack, C. and}, title = {{Cloning and expression of a new cDNA from monocotyledonous plants coding for a diadenosine 5′,5′′′-P1,P4-tetraphosphate hydrolase from barley (Hordeum vulgare)}}, year = {1998}, pages = {481-485}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(98)00819-9}, volume = {431}, abstract = {From a cDNA library generated from mRNA of white leaf tissues of the ribosome‐deficient mutant ‘albostrians\' of barley (Hordeum vulgare cv. Haisa) a cDNA was isolated carrying 54.2% identity to a recently published cDNA which codes for the diadenosine‐5′,5′′′‐P1,P4‐tetraphosphate (Ap4A) hydrolase of Lupinus angustifolius (Maksel et al. (1998) Biochem. J. 329, 313–319), and 69% identity to four partial peptide sequences of Ap4A hydrolase of tomato. Overexpression in Escherichia coli revealed a protein of about 19 kDa, which exhibited Ap4A hydrolase activity and cross‐reactivity with an antibody raised against a purified tomato Ap4A hydrolase (Feussner et al. (1996) Z. Naturforsch. 51c, 477–486). Expression studies showed an mRNA accumulation in all organs of a barley seedling. Possible functions of Ap4A hydrolase in plants will be discussed.} } @Article{IPB-2343, author = {Bohlmann, H. and Vignutelli, A. and Hilpert, B. and Miersch, O. and Wasternack, C. and Apel, K. and}, title = {{Wounding and chemicals induce expression of the Arabidopsis thaliana gene Thi2.1, encoding a fungal defense thionin, via the octadecanoid pathway}}, year = {1998}, pages = {281-286}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(98)01251-4}, volume = {437}, abstract = {In seedlings of Arabidopsis thaliana the thionin gene Thi2.1 is inducible by methyl jasmonate, wounding, silver nitrate, coronatine, and sorbitol. We have used a biochemical and genetic approach to test the signal transduction of these different inducers. Both exogenously applied jasmonates and jasmonates produced endogenously upon stress induction, lead to GUS expression in a Thi2.1 promoter-uidA transgenic line. No GUS expression was observed in a coi1 mutant background which lacks jasmonate perception whereas methyl jasmonate and coronatine but not the other inducers were able to overcome the block in jasmonic acid production in a fad3-2 fad7-2 fad8 mutant background. Our results show conclusively that all these inducers regulate Thi2-1 gene expression via the octadecanoid pathway.} } @Article{IPB-2342, author = {Binarová, P. and Hause, B. and Doležel, J. and Dráber, P. and}, title = {{Association of γ-tubulin with kinetochore/centromeric region of plant chromosomes}}, year = {1998}, pages = {751-757}, journal = {Plant J.}, doi = {10.1046/j.1365-313x.1998.00166.x}, volume = {14}, abstract = {Monoclonal antibodies raised against a phylogenetically conserved peptide from the C‐terminal domain of γ‐tubulin molecule were used for immunofluorescence detection of γ‐tubulin in acentriolar mitotic spindles of plant cells. The antibodies stained kinetochore fibres along their whole length, including the close vicinity of kinetochores. After microtubule disassembly by the antimicrotubular drugs amiprophos‐methyl, oryzalin and colchicine, γ‐tubulin was found on remnants of kinetochore fibres attached to chromosomes. In cells recovering from the amiprophos‐methyl treatment, γ‐tubulin was localized with the re‐growing kinetochore microtubule fibres nucleated or captured by kinetochore/centromeric regions. On isolated chromosomes, γ‐tubulin co‐localized with α‐tubulin in the kinetochore/centromeric region. The data presented suggest that in acentriolar higher plant cells γ‐tubulin might be directly or indirectly involved in modulation and/or stabilization of kinetochore–microtubule interactions.} } @Article{IPB-2376, author = {Wasternack, C. and Miersch, O. and Kramell, R. and Hause, B. and Ward, J. and Beale, M. and Boland, W. and Parthier, B. and Feussner, I. and}, title = {{Jasmonic acid: biosynthesis, signal transduction, gene expression}}, year = {1998}, pages = {139-146}, journal = {Fett/Lipid}, doi = {10.1002/(SICI)1521-4133(19985)100:4/5<139::AID-LIPI139>3.0.CO;2-5}, volume = {100}, abstract = {Jasmonic acid (JA) is an ubiquitously occurring plant growth regulator which functions as a signal of developmentally or environmentally regulated expression of various genes thereby contributing to the defense status of plants [1–5]. The formation of jasmonates in a lipid‐based signalling pathway via octadecanoids seems to be a common principle for many plant species to express wound‐ and stressinduced genes [4, 5].There are various octadecanoid‐derived signals [3]. Among them, jasmonic acid and its amino acid conjugates are most active in barley, supporting arguments that β‐oxidation is an essential step in lipid‐based JA mediated responses. Furthermore, among derivatives of 12‐oxophytodienoic acid (PDA) carrying varying length of the carboxylic acid side‐chain, only those with a straight number of carbon atoms are able to induce JA responsive genes in barley leaves after treatment with these compounds. Barley leaves stressed by treatment with sorbitol solutions exhibit mainly an endogenous rise of JA and JA amino acid conjugates suggesting that both of them are stress signals. Data on organ‐ and tissue‐specific JA‐responsive gene expression will be presented and discussed in terms of “JA as a master switch” among various lipid‐derived signals.} } @Article{IPB-2375, author = {Wasternack, C. and Ortel, B. and Miersch, O. and Kramell, R. and Beale, M. and Greulich, F. and Feussner, I. and Hause, B. and Krumm, T. and Boland, W. and Parthier, B. and}, title = {{Diversity in octadecanoid-induced gene expression of tomato}}, year = {1998}, pages = {345-352}, journal = {J. Plant Physiol.}, doi = {10.1016/S0176-1617(98)80149-1}, volume = {152}, abstract = {In tomato plants wounding leads to up-regulation of various plant defense genes via jasmonates (Ryan, 1992; Bergey et al., 1996). Using this model system of jasmonic acid (JA) signalling, we analyzed activity of octadecanoids to express JA-responsive genes. Leaf treatments were performed with naturally occurring octadecanoids and their molecular mimics such as coronatine or indanone conjugates. JA responses were recorded in terms of up- or down-regulation of various genes by analyzing transcript accumulation, and at least partially in vitro translation products and polypeptide pattern of leaf extracts. The data suggest: (i) 12-Oxo-phytodienoic acid and other intermediates of the octadecanoid pathway has to be ß-oxidized to give a JA response, (ii) Octadecanoids which can not be ß-oxidized are inactive, (iii) JA, its methyl ester (JM), and its amino acid conjugates are most active signals in tomato leaves leading to up regulation of mainly wound-inducible genes and down-regulation of mainly \<house-keeping\> genes, (iv) Some compounds carrying a JA/JM- or JA amino acid conjugate-like structure induce/repress only a subset of genes suggesting diversity of JA signalling.} } @Article{IPB-2374, author = {Vörös, K. and Feussner, I. and Kühn, H. and Lee, J. and Graner, A. and Löbler, M. and Parthier, B. and Wasternack, C. and}, title = {{Characterization of a methyljasmonate-inducible lipoxygenase from barley (Hordeum vulgare cv. Salome) leaves}}, year = {1998}, pages = {36-44}, journal = {Eur. J. Biochem.}, doi = {10.1046/j.1432-1327.1998.2510036.x}, volume = {251}, abstract = {We found three methyl jasmonate−induced lipoxygenases with molecular masses of 92 kDa, 98 kDa, and 100 kDa (LOX‐92, ‐98 and ‐100) [Feussner, I., Hause, B., Vörös, K., Parthier, B. \& Wasternack, C. (1995) Plant J. 7 , 949−957]. At least two of them (LOX‐92 and LOX‐100), were shown to be localized within chloroplasts of barley leaves. Here, we describe the isolation of a cDNA (3073 bp) coding for LOX‐100, a protein of 936 amino acid residues and a molecular mass of 106 kDa. By sequence comparison this lipoxygenase could be identified as LOX2‐type lipoxygenase and was therefore designated LOX2 : Hv : 1 . The recombinant lipoxygenase was expressed in Escherichia coli and characterized as linoleate 13‐LOX and arachidonate 15‐LOX, respectively. The enzyme exhibited a pH optimum around pH 7.0 and a moderate substrate preference for linoleic acid. The gene was transiently expressed after exogenous application of jasmonic acid methyl ester with a maximum between 12 h and 18 h. Its expression was not affected by exogenous application of abscisic acid. Also a rise of endogenous jasmonic acid resulting from sorbitol stress did not induce LOX2 : Hv : 1 , suggesting a separate signalling pathway compared with other jasmonate‐induced proteins of barley. The properties of LOX2 : Hv : 1 are discussed in relation to its possible involvement in jasmonic acid biosynthesis and other LOX forms of barley identified so far.} } @Article{IPB-2373, author = {Vignutelli, A. and Wasternack, C. and Apel, K. and Bohlmann, H. and}, title = {{Systemic and local induction of an Arabidopsis thionin gene by wounding and pathogens}}, year = {1998}, pages = {285-295}, journal = {Plant J.}, doi = {10.1046/j.1365-313X.1998.00117.x}, volume = {14}, abstract = {The Arabidopsis Thi2.1 thionin gene was cloned and sequenced. The promoter was fused to the uidA gene and stably transformed into Arabidopsis to study its regulation. GUS expression levels correlated with the steady‐state levels of Thi2.1 mRNA, thus demonstrating that the promoter is sufficient for the regulation of the Thi2.1 gene. The sensitivity of the Thi2.1 gene to methyl jasmonate was found to be developmentally determined. Systemic and local expression could be induced by wounding and inoculation with Fusarium oxysporum f sp. matthiolae . A deletion analysis of the promoter identified a fragment of 325 bp upstream of the start codon, which appears to contain all the elements necessary for the regulation of the Thi2.1 gene. These results support the view that thionins are defence proteins, and indicate the possibility that resistance of Arabidopsis plants to necrotrophic fungal pathogens is mediated through the octadecanoid pathway.} } @Article{IPB-2425, author = {Ziegler, J. and Hamberg, M. and Miersch, O. and Parthier, B. and}, title = {{Purification and Characterization of Allene Oxide Cyclase from Dry Corn Seeds}}, year = {1997}, pages = {565-573}, journal = {Plant Physiol.}, doi = {10.1104/pp.114.2.565}, volume = {114}, abstract = {Allene oxide cyclase (AOC; EC 5.3.99.6) catalyzes the cyclization of 12,13(S)-epoxy-9(Z),11,15(Z)-octadecatrienoic acid to 12-oxo- 10,15(Z)-phytodienoic acid, the precursor of jasmonic acid (JA). This soluble enzyme was purified 2000-fold from dry corn (Zea mays L.) kernels to apparent homogeneity. The dimeric protein has a molecular mass of 47 kD. Allene oxide cyclase activity was not affected by divalent ions and was not feedback-regulated by its product, 12-oxo-l0,15(Z)-phytodienoic acid, or by JA. ([plus or minus])-cis- 12,13-Epoxy-9(Z)-octadecenoic acid, a substrate analog, strongly inhibited the enzyme, with 50% inhibition at 20 [mu]M. Modification of the inhibitor, such as methylation of the carboxyl group or a shift in the position of the epoxy group, abolished the inhibitory effect, indicating that both structural elements and their position are essential for binding to AOC. Nonsteroidal anti-inflammatory drugs, which are often used to interfere with JA biosynthesis, did not influence AOC activity. The purified enzyme catalyzed the cyclization of 12,13(S)-epoxy-9(Z),11,15(Z)-octadecatrienoic acid derived from linolenic acid, but not that of 12,13(S)-epoxy-9(Z),11- octadecadienoic acid derived from linoleic acid.} } @Article{IPB-2424, author = {Ziegler, J. and Vogt, T. and Miersch, O. and Strack, D. and}, title = {{Concentration of Dilute Protein Solutions Prior to Sodium Dodecyl Sulfate–Polyacrylamide Gel Electrophoresis}}, year = {1997}, pages = {257-260}, journal = {Anal. Biochem.}, doi = {10.1006/abio.1997.2248}, volume = {250}, } @Article{IPB-2415, author = {Wasternack, C. and Parthier, B. and}, title = {{Jasmonate-signalled plant gene expression}}, year = {1997}, pages = {302-307}, journal = {Trends Plant Sci.}, doi = {10.1016/S1360-1385(97)89952-9}, volume = {2}, abstract = {Jasmonic acid is distributed throughout higher plants, synthesized from linolenic acid via the octadecanoic pathway. An important and probably essential role seems to be its operation as a ‘master switch’, responsible for the activation of signal transduction pathways in response to predation and pathogen attack. Proteins encoded by jasmonate-induced genes include enzymes of alkaloid and phytoalexin synthesis, storage proteins, cell wall constituents and stress protectants. The wound-induced formation of proteinase inhibitors is a well-studied example, in which jasmonic acid combines with abscisic acid and ethylene to protect the plant from predation.} } @Article{IPB-2414, author = {Ward, J. L. and Gaskin, P. and Beale, M. H. and Sessions, R. and Koda, Y. and Wasternack, C. and}, title = {{Molecular modelling, synthesis and biological activity of methyl 3-methyljasmonate and related derivatives}}, year = {1997}, pages = {8181-8194}, journal = {Tetrahedron}, doi = {10.1016/S0040-4020(97)00485-7}, volume = {53}, abstract = {Methyl 3-methyljasmonate was synthesised from methyl jasmonate via methyl 3,7-dehydrojasmonate. Molecular modelling predicted an increase in the proportion of cis-orientated side-chains for equilibrated 3-methyl-substituted jasmonate. The synthetic 3-methyljasmonate was shown by gc-ms analysis to equilibrate to a 2:1 ratio of isomers, which appeared from the NMR spectra to comprise mainly the cis-isomer. Surprisingly, both 3,7-dehydro- and 3-methyl-derivatives were inactive in four well established jasmonate bioassays. Methyl-2-methyljasmonate was synthesised and also found to be inactive. Methyl 4,5-dehydrojasmonate was prepared, via the 5-diazo derivative. Both of these compounds have low activity. Our results are discussed with reference to previous knowledge of jasmonate structure-activity relationships and indicate that there are stringent steric demands in jasmonate-receptor interactions.} } @Article{IPB-2402, author = {Kramell, R. and Schneider, G. and Miersch, O. and}, title = {{Chiral separation of amide conjugates of jasmonic acid by liquid chromatography}}, year = {1997}, pages = {104-108}, journal = {Chromatographia}, doi = {10.1007/BF02505545}, volume = {45}, abstract = {Synthetic amide conjugates of (−)-jasmonic acid and its (\+)-enantiomer were resolved by means of chiral liquid chromatography. The diastereomeric pairs prepared by chemical reaction of (±)-jasmonic acid with a series of (S)- or (R)-amino acids and with some (S)-amino acid alcohols were completely separated on Chiralpak AS using a mixture of n-hexane/2-propanal as mobile phase. The retention data indicate that the (−)-jasmonic acid conjugates eluted faster than those of the (\+)-enantiomer, independent on the configuration of the bound amino acid. Likewise, enantiomeric derivatives of (±)-jasmonic acid and non-chiral amino acids were completely separated on the chiral stationary phase and showed the same elution sequence. The resolution factors,Rs, were found to range between 1.13 and 6.64. The separated compounds were chiropatically analyzed by measurement of the circular dichroism.} } @Article{IPB-2401, author = {Kramell, R. and Miersch, O. and Hause, B. and Ortel, B. and Parthier, B. and Wasternack, C. and}, title = {{Amino acid conjugates of jasmonic acid induce jasmonate-responsive gene expression in barley (Hordeum vulgare L.) leaves}}, year = {1997}, pages = {197-202}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(97)01005-3}, volume = {414}, abstract = {Leaves of barley (Hordeum vulgare L. cv. Salome ) treated with jasmonic acid (JA), its methyl ester (JM), or its amino acid conjugates exhibit up‐regulation of specific genes and down‐regulation of house‐keeping genes. This transcriptional regulation exhibits several specificities. (i) The (−)‐enantiomers are more active, and conjugates are mainly active if they carry an l ‐amino acid moiety. (ii) The various JA‐responsive genes respond differentially to enantiomeric and chiralic forms. (iii) Both JA and its amino acid conjugates exhibiting no or negligible interconversion induce/repress genes.} } @Article{IPB-2397, author = {Hertel, S. C. and Knöfel, H.-D. and Kramell, R. and Miersch, O. and}, title = {{Partial purification and characterization of a jasmonic acid conjugate cleaving amidohydrolase from the fungus Botryodiplodia theobromae}}, year = {1997}, pages = {105-110}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(97)00307-4}, volume = {407}, abstract = {A protein preparation from the mycelium of the tropical pathogenic fungus Botryodiplodia theobromae revealed a novel peptidase activity. This enzyme was capable of cleaving conjugates of jasmonic acid with α-amino acids. The protein was enriched 108-fold by gel filtration, ion exchange and hydrophobic interaction chromatography. The enzyme was found to be a glycoprotein with a molecular mass of about 107 kDa. The amidohydrolase seems to be very specific with regard to (−)-jasmonic acid and α-amino acids with (S)-configuration.} } @Article{IPB-2396, author = {Hause, B. and Feussner, K. and Wasternack, C. and}, title = {{Nuclear Location of a Diadenosine 5′,5′”-P1,P4Tetraphosphate (Ap4A) Hydrolase in Tomato Cells Grown in Suspension Cultures}}, year = {1997}, pages = {452-457}, journal = {Bot. Acta}, doi = {10.1111/j.1438-8677.1997.tb00662.x}, volume = {110}, abstract = {Diadenosine 5′,5′”‐P1,P4‐tetraphosphate (Ap4A) cleaving enzymes are assumed to regulate intracellular levels of Ap4A, a compound known to affect cell proliferation and stress responses. From plants an Ap4A hydrolase was recently purified using tomato cells grown in suspension. It was partially sequenced and a peptide antibody was prepared (Feussner et al., 1996). Using this polyclonal monospecific antibody, an abundant nuclear location of Ap4A hydrolase in 4‐day‐old cells of atomato cell suspension culture is demonstrated here by means of immunocytochemical techniques using FITC (fluorescein‐5‐isothiocyanate) labeled secondary antibodies. The microscopic analysis of the occurrence of Ap4A hydrolase performed for different stages of the cell cycle visualized by parallel DAPI (4,6‐diamidino‐2‐phenylindole) staining revealed that the protein accumulates within nuclei of cells in the interphase, but is absent in the nucleus as well as cytoplasm during all stages of mitosis. This first intracellular localization of an Ap4A degrading enzyme within the nucleus and its pattern of appearance during the cell cycle is discussed in relation to the suggested role of Ap4A in triggering DNA synthesis and cell proliferation.} } @Article{IPB-2395, author = {Hause, B. and Kogel, K.-H. and Parthier, B. and Wasternack, C. and}, title = {{In barley leaf cells, jasmonates do not act as a signal during compatible or incompatible interactions with the powdery mildew fungus (Erysiphe graminis f. sp. hordei)}}, year = {1997}, pages = {127-132}, journal = {J. Plant Physiol.}, doi = {10.1016/S0176-1617(97)80191-5}, volume = {150}, abstract = {We have studied a possible function of jasmonates as mediators in the host-pathogen interaction of barley (Hordeum vulgare L.) with the powdery mildew fungus Egh (Erysiphe graminis f. sp. hordei). Previous findings from whole-leaf extracts demonstrated that (i) extracts from infected barley leaves did not contain enhanced levels of jasmonates, (ii) transcripts of jasmonate-inducible genes were not expressed upon infection, and (iii) exogenous application of jasmonates did not induce resistance to Egh (Kogel et al., 1995). Nevertheless, the question arises whether or not jasmonates are involved in the interaction of barley with the powdery mildew fungus at the local site of infection. Using an immunocytological approach the analysis of leaf cross-sections from a susceptible barley cultivar and its near-isogenic mlo5-resistant line revealed no accumulation of JIP-23, the most abundant jasmonate inducible protein, neither in epidermal cells attacked by the pathogen nor in adjacent mesophyll cells. As a positive control, cross-sections from methyl jasmonate-treated leaf segments showed a strong signal for JIP-23 accumulation. Because the presence of the jasmonate-inducible protein is highly indicative for an already low threshold level of endogenous jasmonate (Lehmann et al., 1995), the lack of JIP-23 accumulation at the sites of attempted fungal infection clearly demonstrates the absence of enhanced levels of jasmonates. This excludes even a local rise of jasmonate confined to those single cells penetrated (Mlo genotype) or attacked (mlo5 genotype) by the fungus.} } @Article{IPB-2394, author = {Görschen, E. and Dunaeva, M. and Reeh, I. and Wasternack, C. and}, title = {{Overexpression of the jasmonate-inducible 23 kDa protein (JIP 23) from barley in transgenic tobacco leads to the repression of leaf proteins}}, year = {1997}, pages = {58-62}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(97)01433-6}, volume = {419}, abstract = {We investigated transgenic tobacco lines which express different amounts of the barley JIP 23. In these plants the amount of several proteins decreased proportionally to increasing amounts of JIP 23 whereas the transcript levels were constant as determined for the small and the large subunit of RuBPCase. However, the translation initiation of the rbcS transcript was found to be less efficient than in the wild type. In contrast, the jip 23 transcript was efficiently initiated, indicating that no unspecific impairment of initiation occurred. The data suggest that the barley JIP 23 leads to discrimination among certain tobacco transcripts during translation initiation.} } @Article{IPB-2393, author = {Görschen, E. and Dunaeva, M. and Hause, B. and Reeh, I. and Wasternack, C. and Parthier, B. and}, title = {{Expression of the ribosome-inactivating protein JIP60 from barley in transgenic tobacco leads to an abnormal phenotype and alterations on the level of translation}}, year = {1997}, pages = {470-478}, journal = {Planta}, doi = {10.1007/s004250050151}, volume = {202}, abstract = {In this paper we report the in-planta activity of the ribosome-inactivating protein JIP60, a 60-kDa jasmonate-induced protein from barley (Hordeum vulgare L.), in transgenic tobacco (Nicotiana tabacum L.) plants. All plants expressing the complete JIP60 cDNA under the control of the cauliflower mosaic virus (CaMV) 35S promoter exhibited conspicuous and similar phenotypic alterations, such as slower growth, shorter internodes, lanceolate leaves, reduced root development, and premature senescence of leaves. Microscopic inspection of developing leaves showed a loss of residual meristems and higher degree of vacuolation of mesophyll cells as compared to the wild type. When probed with an antiserum which was immunoreactive against both the N- and the C-terminal half of JIP60, a polypeptide with a molecular mass of about 30 kDa, most probably a processed JIP60 product, could be detected. Phenotypic alterations could be correlated with the differences in the detectable amount of the JIP60 mRNA and processed JIP60 protein. The protein biosynthesis of the transformants was characterized by an increased polysome/monosome ratio but a decreased in-vivo translation activity. These findings suggest that JIP60 perturbs the translation machinery in planta. An immunohistological analysis using the JIP60 antiserum indicated that the immunoreactive polypeptide(s) are located mainly in the nucleus of transgenic tobacco leaf cells and to a minor extent in the cytoplasm.} } @Article{IPB-2389, author = {Feussner, I. and Balkenhohl, T. J. and Porzel, A. and Kühn, H. and Wasternack, C. and}, title = {{Structural Elucidation of Oxygenated Storage Lipids in Cucumber Cotyledons}}, year = {1997}, pages = {21635-21641}, journal = {J. Biol. Chem.}, doi = {10.1074/jbc.272.34.21635}, volume = {272}, abstract = {At early stages of germination, a special lipoxygenase is expressed in cotyledons of cucumber and several other plants. This enzyme is localized at the lipid storage organelles and oxygenates their storage triacylglycerols. We have isolated this lipid body lipoxygenase from cucumber seedlings and found that it is capable of oxygenating in vitro di- and trilinolein to the corresponding mono-, di-, and trihydroperoxy derivatives. To investigate the in vivo activity of this enzyme during germination, lipid bodies were isolated from cucumber seedlings at different stages of germination, and the triacylglycerols were analyzed for oxygenated derivatives by a combination of high pressure liquid chromatography, gas chromatography/mass spectrometry, and nuclear magnetic resonance spectroscopy. We identified as major oxygenation products triacylglycerols that contained one, two, or three 13S-hydroperoxy-9(Z),11(E)-octadecadienoic acid residues. During germination, the amount of oxygenated lipids increased strongly, reaching a maximum after 72 h and declining afterward. The highly specific pattern of hydroperoxy lipids formed suggested the involvement of the lipid body lipoxygenase in their biosynthesis.These data suggest that this lipoxygenase may play an important role during the germination process of cucumber and other plants and support our previous hypothesis that the specific oxygenation of the storage lipids may initiate their mobilization as a carbon and energy source for the growing seedling.} } @Article{IPB-2388, author = {Feussner, I. and Porzel, A. and Wasternack, C. and Kühn, H. and}, title = {{Quantitative Analyse von Lipoxygenase-Metaboliten in Lipiden durch NMR-Spektroskopie}}, year = {1997}, pages = {54-58}, journal = {BIOspektrum}, volume = {3}, } @Article{IPB-2387, author = {Feussner, K. and Feussner, I. and Leopold, I. and Wasternack, C. and}, title = {{Isolation of a cDNA coding for an ubiquitin-conjugating enzyme UBC1 of tomato - the first stress-induced UBC of higher plants}}, year = {1997}, pages = {211-215}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(97)00509-7}, volume = {409}, abstract = {A clone of an ubiquitin‐conjugating enzyme (UBC) was isolated from a λ‐ZAP‐cDNA library, generated from mRNA of tomato (Lycopersicon esculentum) cells grown in suspension for 3 days. The open reading frame called Le UBC1, encodes for a polypeptide with a predicted molecular mass of 21.37 kDa, which was confirmed by bacterial overexpression and SDS‐PAGE. Database searches with Le UBC1 showed highest sequence similarities to UBC1 of bovine and yeast. By Southern blot analysis Le UBC1 was identified as a member of a small E2 subfamily of tomato, presumably consisting of at least two members. As revealed by Northern blot analysis Le UBC1 is constitutively expressed in an exponentially growing tomato cell culture. In response to heat shock an increase in Le UBC1‐mRNA was detectable. A strong accumulation of the Le UBC1‐transcript was observed by exposure to heavy metal stress which was performed by treatment with cadmium chloride (CdCl2). The cellular uptake of cadmium was controlled via ICP‐MS measurements. The data suggest that like in yeast, in plants a certain subfamily of UBC is specifically involved in the proteolytic degradation of abnormal proteins as result of stress.} } @Article{IPB-2386, author = {Feussner, I. and Fritz, I. G. and Hause, B. and Ullrich, W. R. and Wasternack, C. and}, title = {{Induction of a new Lipoxygenase Form in Cucumber Leaves by Salicylic Acid or 2,6-Dichloroisonicotinic Acid}}, year = {1997}, pages = {101-108}, journal = {Bot. Acta}, doi = {10.1111/j.1438-8677.1997.tb00616.x}, volume = {110}, abstract = {Changes in lipoxygenase (LOX) protein pattern and/or activity were investigated in relation to acquired resistance of cucumber (Cucumis sativus L.) leaves against two powdery mildews, Sphaerotheca fuliginea (Schlecht) Salmon and Erysiphe cichoracearum DC et Merat. Acquired resistance was established by spraying leaves with salicylic acid (SA) or 2,6‐dichloroisonicotinic acid (INA) and estimated in whole plants by infested leaf area compared to control plants. SA was more effective than INA. According to Western blots, untreated cucumber leaves contained a 97 kDa LOX form, which remained unchanged for up to 48 h after pathogen inoculation. Upon treatment with SA alone for 24 h or with INA plus pathogen, an additional 95 kDa LOX form appeared which had an isoelectric point in the alkaline range. For the induction of this form, a threshold concentration of 1 mM SA was required, higher SA concentrations did not change LOX‐95 expression which remained similar between 24 h and 96 h but further increased upon mildew inoculation. Phloem exudates contained only the LOX‐97 form, in intercellular washing fluid no LOX was detected. dichloroisonicotinic localization revealed LOX protein in the cytosol of the mesophyll cells without differences between the forms.} } @Article{IPB-2385, author = {Feussner, I. and Kühn, H. and Wasternack, C. and}, title = {{Do specific linoleate 13-lipoxygenases initiate β-oxidation?}}, year = {1997}, pages = {1-5}, journal = {FEBS Lett.}, doi = {10.1016/S0014-5793(97)00218-4}, volume = {406}, abstract = {The germination process of oilseed plants is characterized by a mobilization of the storage lipids which constitute the major carbon source for the growing seedling. Despite the physiological importance of the lipid mobilization, the mechanism of this process is not well understood. Recently, it was found that a specific linoleate 13-lipoxygenase is induced during the stage of lipid mobilization in various oilseed plants and that this enzyme is translocated to the membranes of the lipid storage organelles, the so called lipid bodies. Lipoxygenase expression was paralleled by the occurrence of enantiospecific hydro(pero)xy polyenoic fatty acid derivatives in the storage lipids suggesting the in vivo action of the enzyme. Furthermore, it was reported that oxygenated polyenoic fatty acids, in particular as 13(S)-hydro(pero)xy-9(Z),11(E)-octadecanoic acid [13(S)-H(P)ODE], are cleaved preferentially from the storage lipids when compared with their non-oxygenated linoleate residues. These findings may suggest that 13(S)-H(P)ODE may constitute the endogenous substrate for β-oxidation during lipid mobilization of oilseed plants.} } @Article{IPB-2450, author = {Wong, L. M. and Abel, S. and Shen, N. and de la Foata, M. and Mall, Y. and Theologis, A. and}, title = {{Differential activation of the primary auxin response genes, PS-IAA4/5 and PS-IAA6, during early plant development}}, year = {1996}, pages = {587-599}, journal = {Plant J.}, doi = {10.1046/j.1365-313X.1996.9050587.x}, volume = {9}, abstract = {The plant growth hormone auxin typified by indoleacetic acid (IAA) transcriptionally activates early genes in pea, PS‐IAA4/5 and PS‐IAA6 , that are members of a multigene family encoding short‐lived nuclear proteins. To gain first insight into the biological role of PS‐IAA4/5 and PSIAA6 , promoter‐β‐glucuronidase (GUS) gene fusions were constructed and their expression during early development of transgenic tobacco seedlings was examined. The comparative analysis reveals spatial and temporal expression patterns of both genes that correlate with cells, tissues, and developmental processes known to be affected by auxin. GUS activity in seedlings of both transgenic lines is located in the root meristem, sites of lateral root initiation and in hypocotyls undergoing rapid elongation. In addition, mutually exclusive cell‐specific expression is evident. For instance, PS‐IAA4/5—GUS but not PS‐IAA6—GUS is expressed in root vascular tissue and in guard cells, whereas only PS‐IAA6—GUS activity is detectable in glandular trichomes and redistributes to the elongating side of the hypocotyl upon gravitropic stimulation. Expression of PS‐IAA4/5 and PS‐IAA6 in elongating, dividing, and differentiating cell types indicates multiple functions during development. The common and yet distinct activity patterns of both genes suggest a combinatorial code of spatio‐temporal co‐expression of the various PS‐IAA4/ 5‐like gene family members in plant development that may mediate cell‐specific responses to auxin.} } @Article{IPB-2446, author = {Wasternack, C. and Atzorn, R. and Peña-Cortés, H. and Parthier, B. and}, title = {{Alteration of Gene Expression by Jasmonate and ABA in Tobacco and Tomato}}, year = {1996}, pages = {503-510}, journal = {J. Plant Physiol.}, doi = {10.1016/S0176-1617(96)80038-1}, volume = {147}, abstract = {The synthesis of jasmonate-induced proteins in leaves of tobacco (Nicotiana plumbaginifolia) and tomato (Lycopersicon esculentum) was studied in order to find a possible functional link in the actions of abscisic acid (ABA) and jasmonates. ABA-deficient mutants of tobacco (CKR1) and of tomato (sitiens, flacca), and their corresponding wild-types, were compared with respect to endogenous contents of jasmonates and ABA, and polypeptide and transcript patterns in water- or jasmonate-floated leaves, leaves stressed by floating on sorbitol, or by weak desiccation. Our results indicate that in tobacco the synthesis of proteins induced by jasmonate differed from those induced by ABA, whereas in tomato some jasmonate-induced proteins were also induced by ABA. The results provide further evidence that different signalling pathways exist for jasmonate/ABA-responsive gene expression in various plant species.} } @Article{IPB-2441, author = {Peña-Cortés, H. and Prat, S. and Atzorn, R. and Wasternack, C. and Willmitzer, L. and}, title = {{Abscisic acid-deficient plants do not accumulate proteinase inhibitor II following systemin treatment}}, year = {1996}, pages = {447-451}, journal = {Planta}, doi = {10.1007/BF00620062}, volume = {198}, abstract = {The role of systemin in Pin2 gene expression was analyzed in wild-type plants of potato (Solanum tuberosum L.) and tomato (Lycopersicon esculentum Mill.), as well as in abscisic acid (ABA)-deficient tomato (sitiens) and potato (droopy) plants. The results showed that systemin initiates Pin2 mRNA accumulation only in wildtype tomato and potato plants. As in the situation after mechanical wounding,Pin2 gene expression in ABA-deficient plants was not activated by systemin. Increased endogenous levels of jasmonic acid (JA) and accumulation of Pin2 mRNA were observed following treatment with α-linolenic acid, the precursor of JA biosynthesis, suggesting that these ABA mutants still have the capability to synthesize de novo JA. Measurement of endogenous levels of ABA and JA showed that systemin leads to an increase of both phytohormones (ABA and JA) only in wild-type but not in ABA-deficient plants.} } @Article{IPB-2440, author = {O'Donnell, P. J. and Calvert, C. and Atzorn, R. and Wasternack, C. and Leyser, H. M. O. and Bowles, D. J. and}, title = {{Ethylene as a Signal Mediating the Wound Response of Tomato Plants}}, year = {1996}, pages = {1914-1917}, journal = {Science}, doi = {10.1126/science.274.5294.1914}, volume = {274}, abstract = {Plants respond to physical injury, such as that caused by foraging insects, by synthesizing proteins that function in general defense and tissue repair. In tomato plants, one class of wound-responsive genes encodes proteinase inhibitor (pin) proteins shown to block insect feeding. Application of many different factors will induce or inhibit pin gene expression. Ethylene is required in the transduction pathway leading from injury, and ethylene and jasmonates act together to regulate pin gene expression during the wound response.} } @Article{IPB-2438, author = {Leopold, J. and Hause, B. and Lehmann, J. and Graner, A. and Parthier, B. and Wasternack, C. and}, title = {{Isolation, characterization and expression of a cDNA coding for a jasmonate-inducible protein of 37 kDa in barley leaves}}, year = {1996}, pages = {675-684}, journal = {Plant Cell Environ.}, doi = {10.1111/j.1365-3040.1996.tb00402.x}, volume = {19}, abstract = {In barley leaves, there is a dramatic alteration of gene expression upon treatment with jasmonates leading to the accumulation of newly formed proteins, designated as jasmonate‐inducible proteins (JIPs). In the present study, a new jasmonate‐inducible cDNA, designated pHvJS37, has been isolated by differential screening of a γgt10 cDNA library constructed from mRNA of jasmonate‐treated barley leaf segments. The open reading frame (ORF) encodes a 39‐9 kDa polypeptide which cross‐reacts with antibodies raised against the in vivo JIP‐37. The hydropathic plot suggests that the protein is mainly hydrophilic, containing two hydrophilic domains near the C‐terminus. Database searches did not show any sequence homology of pHv.JS37 to known sequences. Southern analysis revealed at least two genes coding for JIP‐37 which map to the distal portion of the long arm of chromosome 3 and are closely related to genes coding for JIP‐23. The expression pattern of the JIP‐37 genes over time shows differential responses to jasmonate, abscisic acid (ABA), osmotic stress (such as sorbitol treatment) and desiccation stress. No expression was found under salt stress. From experiments using an inhibitor and intermediates of jasmonate synthesis such as α‐linolenic acid and 12‐oxophytodienoic acid, we hypothesize that there is a stress‐induced lipid‐based signalling pathway in which an endogenous rise of jasmonate switches on JIP‐37 gene expression. Using immunocytochemical techniques, JIP‐37 was found to be simultaneously located in the nucleus, the cytoplasm and the vacuoles.} } @Article{IPB-2433, author = {Herde, O. and Atzorn, R. and Fisahn, J. and Wasternack, C. and Willmitzer, L. and Pena-Cortes, H. and}, title = {{Localized Wounding by Heat Initiates the Accumulation of Proteinase Inhibitor II in Abscisic Acid-Deficient Plants by Triggering Jasmonic Acid Biosynthesis}}, year = {1996}, pages = {853-860}, journal = {Plant Physiol.}, doi = {10.1104/pp.112.2.853}, volume = {112}, abstract = {To test whether the response to electrical current and heat treatment is due to the same signaling pathway that mediates mechanical wounding, we analyzed the effect of electric-current application and localized burning on proteinase inhibitor II (Pin2) gene expression in both wild-type and abscisic acid (ABA)-deficient tomato (Lycopersicon esculentum Mill.) and potato (Solanum phureja) plants. Electric-current application and localized burning led to the accumulation of Pin2 mRNA in potato and tomato wild-type plants. Among the treatments tested, only localized burning of the leaves led to an accumulation of Pin2 mRNA in the ABA-deficient plants. Electric-current application, like mechanical injury, was able to initiate ABA and jasmonic acid (JA) accumulation in wild-type but not in ABA-deficient plants. In contrast, heat treatment led to an accumulation of JA in both wild-type and ABA-deficient plants. Inhibition of JA biosynthesis by aspirin blocked the heat-induced Pin2 gene expression in tomato wild-type leaves. These results suggest that electric current, similar to mechanical wounding, requires the presence of ABA to induce Pin2 gene expression. Conversely, burning of the leaves activates Pin2 gene expression by directly triggering the biosynthesis of JA by an alternative pathway that is independent of endogenous ABA levels.} } @Article{IPB-2432, author = {Hause, B. and Demus, U. and Teichmann, C. and Parthier, B. and Wasternack, C. and}, title = {{Developmental and Tissue-Specific Expression of JIP-23, a Jasmonate-Inducible Protein of Barley}}, year = {1996}, pages = {641-649}, journal = {Plant Cell Physiol.}, doi = {10.1093/oxfordjournals.pcp.a028993}, volume = {37}, abstract = {Developmental expression of a 23 kDa jasmonate-induced protein (JIP-23) of barley leaves (Hordeum vulgare cv. Salome) was studied by measuring the time-dependent accumulation of transcript and protein during germination. Tissue-specific expression of JIP-23 was analyzed immunocytochemically and by in situ hybridizations, respectively. During seed germination JIP-23 mRNA was found to accumulate transiently with a maximum at 32 h, whereas the protein was steadily detectable after the onset of expression. The occurrence of new isoforms of JIP-23 during germination in comparison to jasmonate-treated leaves suggests, that the JIP-23 gene family of barley is able to express different subsets of isoforms dependent on the developmental stage.JIP-23 and its transcript were found mainly in the scutellum, the scutellar nodule and in lower parts of the primary leaf of 6 days old seedlings. All these tissues exhibited high levels of endogenous jasmonates. In situ hybridization revealed specific accumulation of JIP-23 mRNA in companion cells of the phloem in the nodule plate of the scutellum. In accordance with that, JIP-23 was detected immunocytochemically in phloem cells of the root as well as of the scutellar nodule and in parenchymatic cells of the scutellum. The cell type-specific occurrence of JIP-23 was restricted to cells, which are known to be highly stressed osmotically by active solute transport. This observation suggests, that the expression of this protein might be a response to osmotic stress during development.} } @Article{IPB-2431, author = {Feussner, I. and Hause, B. and Nellen, A. and Wasternack, C. and Kindl, H. and}, title = {{Lipid-body lipoxygenase is expressed in cotyledons during germination prior to other lipoxygenase forms}}, year = {1996}, pages = {288-293}, journal = {Planta}, doi = {10.1007/BF00206255}, volume = {198}, abstract = {Lipid bodies are degraded during germination. Whereas some proteins, e.g. oleosins, are synthesized during the formation of lipid bodies of maturating seeds, a new set of proteins, including a specific form of lipoxygenase (LOX; EC 1.13.11.12), is detectable in lipid bodies during the stage of fat degradation in seed germination. In cotyledons of cucumber (Cucumis sativus L.) seedlings at day 4 of germination, the most conspicuous staining with anti-LOX antibodies was observed in the cytosol. At very early stages of germination, however, the LOX form present in large amounts and synthesized preferentially was the lipid-body LOX. This was demonstrated by immunocytochemical staining of cotyledons from 1-h and 24-h-old seedlings: the immunodecoration of sections of 24-h-old seedlings with anti-LOX antiserum showed label exclusively correlated with lipid bodies of around 3 μm in diameter. In accordance, the profile of LOX protein isolated from lipid bodies during various stages of germination showed a maximum at day 1. By measuring biosynthesis of the protein in vivo we demonstrated that the highest rates of synthesis of lipid-body LOX occurred at day 1 of germination. The early and selective appearance of a LOX form associated with lipid bodies at this stage of development is discussed.} } @Article{IPB-2430, author = {Feussner, K. and Guranowski, A. and Kostka, S. and Wasternack, C. and}, title = {{Diadenosine 5′,5‴- P1,P4-tetraphosphate (Ap4A) Hydrolase from Tomato (Lycopersicon esculentum cv. Lukullus) -Purification, Biochemical Properties and Behaviour during Stress}}, year = {1996}, pages = {477-486}, journal = {Z. Naturforsch. C}, doi = {10.1515/znc-1996-7-805}, volume = {51}, abstract = {Dinucleoside 5′,5‴-P1,P4-tetraphosphate hydrolase (EC 3.6.1.17) has been purified to homogeneity from tomato (Lycopersicon esculentum) cells grown in suspension. The purification procedure comprised ammonium sulphate fractionation following five standard chroma­ tography steps and a final chromatography on Ap4A-Sepharose.} } @Article{IPB-2429, author = {Abel, S. and Theologis, A. and}, title = {{Early Genes and Auxin Action}}, year = {1996}, pages = {9-17}, journal = {Plant Physiol.}, doi = {10.1104/pp.111.1.9}, volume = {111}, } @Article{IPB-2428, author = {Abel, S. and Ballas, N. and Wong, L.-M. and Theologis, A. and}, title = {{DNA elements responsive to auxin}}, year = {1996}, pages = {647-654}, journal = {BioEssays}, doi = {10.1002/bies.950180808}, volume = {18}, abstract = {Genes induced by the plant hormone auxin are probably involved in the execution of vital cellular functions and developmental processes. Experimental approaches designed to elucidate the molecular mechanisms of auxin action have focused on auxin perception, genetic dissection of the signaling apparatus and specific gene activation. Auxin‐responsive promoter elements of early genes provide molecular tools for probing auxin signaling in reverse. Functional analysis of several auxin‐specific promoters of unrelated early genes suggests combinatorial utilization of both conserved and variable elements. These elements are arranged into autonomous domains and the combination of such modules generates uniquely composed promoters. Modular promoters allow for auxin‐mediated transcriptional responses to be revealed in a tissue‐ and development‐specific manner.} } @Article{IPB-2427, author = {Abdala, G. and Castro, G. and Guiñazú, M. M. and Tizio, R. and Miersch, O. and}, title = {{Occurrence of jasmonic acid in organs of Solanum tuberosum L. and its effect on tuberization}}, year = {1996}, pages = {139-143}, journal = {Plant Growth Regul.}, doi = {10.1007/BF00024580}, volume = {19}, abstract = {The aims of this study were to demonstrate the endogenous presence of jasmonic acid (JA) in roots, stolons and periderm of new formed tubers, by means of bioassays, ELISA and GC-MS, and to test a microdrop bioassay using the leaflets of potato cuttings cultured in vitro. Our results confirm the existence of JA by bioassays and GC-MS in foliage, stolons, roots and tuber periderm.} } @Article{IPB-2466, author = {Wasternack, C. and Atzorn, R. and Leopold, J. and Feussner, I. and Rademacher, W. and Parthier, B. and}, title = {{Synthesis of jasmonate-induced proteins in barley (Hordeum vulgare) is inhibited by the growth retardant tetcyclacis}}, year = {1995}, pages = {335-341}, journal = {Physiol. Plant.}, doi = {10.1111/j.1399-3054.1995.tb05320.x}, volume = {94}, abstract = {BarJey leaf segments treated with jasmonate respond with the synthesis of specific proseins, referred to as jasmonate‐induced proteins (JIPs). Application of abscisic acid (ABAl also induced JIP synthesis (Weidhase et al. 1987). In this study the effects of inhibitors on sorbitol‐induced increases of endogenous jasmonates and ABA were investigated. The promotion of jasmonates by sorbitol was inhibited by the growth retardant tetcyclacis at concentrations as low as 1 ftM. In parallel with the decrease of jasmonates, JIP gene expression was reduced as reflected by a decline in the level of a 23‐kDa protein UIP‐23) and mRNAs of JIP‐6 and JIP‐23. 12‐Oxo‐phytodienoic acid, an inlermediale in the lipoxygenase (LOX) pathway leading to jasmonic acid was able to overcome the inhibition by tetcyclacis and increases both the endogenous jasmonate content and transcript accumulation. This suggests that tetcyclacis acts upstream of 12‐oxo‐phytodienoic acid and in keeping with this proposal, an increase in relative LOX activity was detected after tetcyclacis treatment. Although tetcyclacis was shown to inhibit the degradation of ABA to phaseic acid, its effect on jasmonate synthesis is much more pronounced.} } @Article{IPB-2460, author = {Lehmann, J. and Atzorn, R. and Brückner, C. and Reinbothe, S. and Leopold, J. and Wasternack, C. and Parthier, B. and}, title = {{Accumulation of jasmonate, abscisic acid, specific transcripts and proteins in osmotically stressed barley leaf segments}}, year = {1995}, pages = {156-192}, journal = {Planta}, doi = {10.1007/BF00239952}, volume = {197}, abstract = {The accumulation of abundant proteins and their respective transcripts, induced by 10−4 M cisabscisic acid or 10−5 M jasmonic acid methyl ester, was studied in barley (Hordeum vulgare L.) leaf segments and compared to that resulting from osmotic stress caused by floating the segments on solutions of sorbitol, glucose, polyethyleneglycol (PEG)-6000 or NaCl. Osmotic stress or treatment with abscisic acid led to the synthesis of novel proteins which were identical to jasmonateinduced proteins (JIPs) with respect to immunological properties and molecular masses. The most prominent polypeptides were characterized by molecular masses of 66, 37 and 23 kDa and were newly synthesized. Whereas sorbitol, mannitol, sucrose, glucose and PEG provoked the synthesis of JIPs, 2deoxyglucose and NaCl did not. We provide evidence that the synthesis of JIPs induced by osmotic stress is directly correlated with a preceding rise in endogenous jasmonates. These jasmonates, quantified by an enzyme immunoassay specific for (−)jasmonic acid and its aminoacid conjugates, increased remarkably in leaf segments treated with sorbitol, glucose or other sugars. In contrast, no increase in jasmonates could be observed in tissues exposed to salts (NaCl). The results strengthen the hypothesis that the accumulation of jasmonates, probably by de-novo synthesis, is an intermediate and essential step in a signalling pathway between (osmotic) stress and activation of genes coding for polypeptides of high abundance.} } @Article{IPB-2459, author = {Kramell, R. and Atzorn, R. and Schneider, G. and Miersch, O. and Brückner, C. and Schmidt, J. and Sembdner, G. and Parthier, B. and}, title = {{Occurrence and identification of jasmonic acid and its amino acid conjugates induced by osmotic stress in barley leaf tissue}}, year = {1995}, pages = {29-36}, journal = {J. Plant Growth Regul.}, doi = {10.1007/BF00212643}, volume = {14}, abstract = {The effect of osmotically active substances on the alteration of endogenous jasmonates was studied in barley (Hordeum vulgare L. cv. Salome) leaf tissue. Leaf segments were subjected to solutions of d-sorbitol, d-mannitol, polyethylene glycol 6000, sodium chloride, or water as a control. Alterations of endogenous jasmonates were monitored qualitatively and quantitatively using immunoassays. The structures of jasmonates isolated were determined on the basis of authentic substances by capillary gas chromatography-mass spectrometry. The stereochemistry of the conjugates was confirmed by high performance liquid chromatography with diastereoisomeric references. In barley leaves, jasmonic acid and its amino acid conjugates, for example, with valine, leucine, and isoleucine, are naturally occurring jasmonates. In untreated leaf segments, only low levels of these native jasmonates were found. After treatment of the leaf tissues with sorbitol, mannitol, as well as with polyethylene glycol, an increase of both jasmonic acid and its conjugates could be observed, depending on the stress conditions used. In contrast, salt stress was without any stimulating effect on the levels of endogenous jasmonates. From barley leaf segments exposed to sorbitol (1m) for 24 h, jasmonic acid was identified as the major accumulating compound. Jasmonic acid-amino acid conjugates increased likewise upon stress treatment.} } @Article{IPB-2458, author = {Kogel, K.-H. and Ortel, B. and Jarosch, B. and Atzorn, R. and Schiffer, R. and Wasternack, C. and}, title = {{Resistance in barley against the powdery mildew fungus (Erysiphe graminis f.sp.hordei) is not associated with enhanced levels of endogenous jasmonates}}, year = {1995}, pages = {319-332}, journal = {Eur. J. Plant Pathol.}, doi = {10.1007/BF01874788}, volume = {101}, abstract = {Onset of acquired resistance of barley (Hordeum vulgare) chemically induced by 2,6-dichloroisonicotinic acid (DCINA) correlated with the accumulation of mRNA homologous to cDNA pHvJ256 which codes for a soluble leaf-thionin with a Mr. of 6 kDa [Wasternacket al., 1994a]. In the present work, we extend this finding by showing that the thionin transcript also accumulated following treatment of barley with the resistance-inducing compounds 3,5-dichlorosalicylic acid (DCSA), salicylic acid (SA), and an extract fromBacillus subtilis. The polypeptide showed antifungal activity against the biotrophic cereal pathogensErysiphe graminis f.sp.hordei andPuccinia graminis f.sp.tritici which may indicate a possible role in the mechanism of acquired resistance in barley. A thionin transcript hybridizing to pHvJ256 accumulated also in response to application of jasmonates, or treatments that elevated endogenous amounts of the plant growth substance, pointing to the possibility that signaling mediating defense responses in barley involves jasmonates. However, a topical spray application of jasmonic acid (JA) or jasmonate methyl ester (JM) did not protect barley leaves against infection byE. graminis. Performing a kinetic analysis by an enzyme immunoassay specific for (−)-JA, (−)-JM, and its amino acid conjugates, accumulation of jasmonates was detected in osmotically stressed barley but not at the onset of chemically induced or genetically based resistance governed by the powdery mildew resistance genesMlg, Mla 12, ormlo 5. Furthermore, the jasmonate-inducible proteins JIP-23 and JIP-60 were strongly induced following JM- but not DCINA-treatment or inoculation withE. graminis. Hence, in barley, no indications were found in favour for the previously proposed model of a lipid-based signaling pathway via jasmonates mediating expression of resistance in plants against pathogens.} } @Article{IPB-2455, author = {Harms, K. and Atzorn, R. and Brash, A. and Kühn, H. and Wasternack, C. and Willmitzer, L. and Pena-Cortes, H. and}, title = {{Expression of a Flax Allene Oxide Synthase cDNA Leads to Increased Endogenous Jasmonic Acid (JA) Levels in Transgenic Potato Plants but Not to a Corresponding Activation of JA-Responding Genes}}, year = {1995}, pages = {1645-1654}, journal = {Plant Cell}, doi = {10.1105/tpc.7.10.1645}, volume = {7}, abstract = {Both jasmonic acid (JA) and its methyl ester, methyl jasmonate (MeJA), are thought to be significant components of the signaling pathway regulating the expression of plant defense genes in response to various stresses. JA and MeJA are plant lipid derivatives synthesized from [alpha]-linolenic acid by a lipoxygenase-mediated oxygenation leading to 13-hydroperoxylinolenic acid, which is subsequently transformed by the action of allene oxide synthase (AOS) and additional modification steps. AOS converts lipoxygenase-derived fatty acid hydroperoxide to allene epoxide, which is the precursor for JA formation. Overexpression of flax AOS cDNA under the regulation of the cauliflower mosaic virus 35S promoter in transgenic potato plants led to an increase in the endogenous level of JA. Transgenic plants had six- to 12-fold higher levels of JA than the nontransformed plants. Increased levels of JA have been observed when potato and tomato plants are mechanically wounded. Under these conditions, the proteinase inhibitor II (pin2) genes are expressed in the leaves. Despite the fact that the transgenic plants had levels of JA similar to those found in nontransgenic wounded plants, pin2 genes were not constitutively expressed in the leaves of these plants. Transgenic plants with increased levels of JA did not show changes in water state or in the expression of water stress-responsive genes. Furthermore, the transgenic plants overexpressing the flax AOS gene, and containing elevated levels of JA, responded to wounding or water stress by a further increase in JA and by activating the expression of either wound- or water stress-inducible genes. Protein gel blot analysis demonstrated that the flax-derived AOS protein accumulated in the chloroplasts of the transgenic plants.} } @Article{IPB-2454, author = {Feussner, I. and Hause, B. and Vörös, K. and Parthier, B. and Wasternack, C. and}, title = {{Jasmonate-induced lipoxygenase forms are localized in chloroplasts of barley leaves (Hordeum vulgare cv. Salome)}}, year = {1995}, pages = {949-957}, journal = {Plant J.}, doi = {10.1046/j.1365-313X.1995.07060949.x}, volume = {7}, abstract = {Barley leaves respond to application of (−)‐jasmonic acid (JA), or its methylester (JM) with the synthesis of abundant proteins, so‐called jasmonate induced proteins (JIPs). Here Western blot analysis is used to show a remarkable increase upon JM treatment of a 100 kDa lipoxygenase (LOX), and the appearance of two new LOX forms of 98 and 92 kDa. The temporal increase of LOX‐100 protein upon JM treatment was clearly distinguishable from the additionally detectable LOX forms. JM‐induced LOX forms in barley leaves were compared with those of Arabidopsis and soybean leaves. Both dicot species showed a similar increase of one LOX upon JM induction, whereas, leaves from soybean responded with additional synthesis of a newly formed LOX of 94 kDa.Using immunofluorescence analysis and isolation of intact chloroplasts, it is demonstrated that JM‐induced LOX forms of barley leaves are exclusively located in the chloroplasts of all chloroplast‐containing cells. Analogous experiments carried out with Arabidopsis and soybean revealed a similar plastidic location of JM‐induced LOX forms in Arabidopsis but a different situation for soybean. In untreated soybean leaves the LOX protein was mainly restricted to vacuoles of paraveinal mesophyll cells. Additionally, LOX forms could be detected in cytoplasm and nuclei of bundle sheath cells. Upon JM treatment cytosolic LOX was detectable in spongy mesophyll cells, too. The intracellular location of JM‐induced LOX is discussed in terms of stress‐related phenomena mediated by JM.} } @Article{IPB-2452, author = {Abel, S. and Nguyen, M. D. and Theologis, A. and}, title = {{The PS-IAA4/5-like Family of Early Auxin-inducible mRNAs in Arabidopsis thaliana}}, year = {1995}, pages = {533-549}, journal = {J. Mol. Biol.}, doi = {10.1006/jmbi.1995.0454}, volume = {251}, abstract = {The plant hormone auxin transcriptionally activates early genes. We have isolated a 14-member family of DNA sequences complementary to indoleacetic acid (IAA)-inducible transcripts inArabidopsis thaliana. The corresponding genes, IAA1 and IAA14, are homologs of PS-1AA4/5 and PS-IAA6 from pea, AUX22 and AUX28 from soybean, ARG3 and ARG4from mungbean, and AtAux2-11 and AtAux2-27 from Arabidopsis. The members of the family are differentially expressed in mature Arabidopsis plants. Characterization of IAA gene expression in etiolated seedlings demonstrates specificity for auxin inducibility. The response of most family members to IAA is rapid (within 4 to 30 minutes) and insensitive to cyclohexamide. Cyclohexamide alone induces all the early genes. Auxin-induction of two late genes, IAA7 and IAA8, is inhibited by cyclohexamide, indicating requirement of protein synthesis for their activation. All IAA genes display a biphasic dose response that is optimal at 10 μM IAA. However, individual genes respond differentially between 10 nM and 5μM IAA. Expression of all genes is defective in the Arabidopsis auxin-resistant mutant lines axr1, axr2, and aux1.The encoded polypeptides share four conserved domains, and seven invariant residues in the intervening regions. The spaces vary considerably in length, rendering the calculated molecular mass of IAA proteins to range from 19 kDa to 36 kDa. Overall sequence identity between members of the family is highly variable (36 to 87%). Their most significant structural features are functional nuclear transport signals, and a putative βαα-fold whose modeled three dimensional structure appears to be compatible with the prokaryotic β-ribbon DNA recognition motif. The data suggest that auxin induces in a differential and hierarchical fashion a large family of early genes that encode a structurally diverse class of nuclear proteins. These proteins are proposed to mediate tissue-specific and cell-type restricted responses to the hormone during plant growth and development.} } @Article{IPB-2451, author = {Abel, S. and Theologis, A. and}, title = {{A polymorphic bipartite motif signals nuclear targeting of early auxin-inducible proteins related to PS-IAA4 from pea (Pisum sativum)}}, year = {1995}, pages = {87-96}, journal = {Plant J.}, doi = {10.1046/j.1365-313X.1995.08010087.x}, volume = {8}, abstract = {The plant hormone, indoleacetic acid (IAA), transcriptionally activates two early genes in pea, PS‐IAA4/5 and PS‐IAA6 , that encode short‐lived nuclear proteins. The identification of the nuclear localization signals (NLS) in PS‐IAA4 and PS‐IAA6 using progressive deletion analysis and site‐directed mutagenesis is reported. A C‐terminal SV40‐type NLS is sufficient to direct the β‐glucuronidase reporter to the nucleus of transiently transformed tobacco protoplasts, but is dispensible for nuclear localization of both proteins. The dominant and essential NLS in PS‐IAA4 and PS‐IAA6 overlap with a bipartite basic motif which is polymorphic and conserved in related proteins from other plant species, having the consensus sequence (KKNEK)KR‐X(24–71)‐(RSXRK)/(RK/RK). Both basic elements of this motif in PS‐IAA4, (KR‐X41‐RSYRK), function interdependently as a bipartite NLS. However, in PS‐IAA6 (KKNEKKR‐X36‐RKK) the upstream element of the corresponding motif contains additional basic residues which allow its autonomous function as an SV40‐type monopartite NLS. The spacer‐length polymorphism, X(24–70), in respective bipartite NLS peptides of several PS‐IAA4‐like proteins from Arabidopsis thaliana does not affect nuclear targeting function. The structural and functional variation of the bipartite basic motif in PS‐IAA4‐like proteins supports the proposed integrated consensus of NLS.} } @Article{IPB-2473, author = {Hause, B. and zur Nieden, U. and Lehmann, J. and Wasternack, C. and Parthier, B. and}, title = {{Intracellular Localization of Jasmonate-Induced Proteins in Barley Leaves}}, year = {1994}, pages = {333-341}, journal = {Bot. Acta}, doi = {10.1111/j.1438-8677.1994.tb00804.x}, volume = {107}, abstract = {The plant growth substance jasmonic acid and its methyl ester (JA‐Me) induce a set of proteins (jasmonate‐induced proteins, JIPs) when applied to leaf segments of barley (Hordeum vulgare L. cv. Salome). Most of these JIPs could be localized within different cell compartments by using a combination of biochemical and histochemical methods. Isolation and purification of various cell organelles of barley mesophyll cells, the separation of their proteins by one‐dimensional polyacrylamide gel electrophoresis and the identification of the major abundant JIPs by Western blot analysis, as well as the immuno‐gold labelling of JIPs in ultrathin sections were performed to localize JIPs intracellularly. JIP‐23 was found to be in vacuoles, peroxisomes, and in the granular parts of the nucleus as well as within the cytoplasm; JIP‐37 was detected in vacuoles and in the nucleoplasm; JIP‐66 is a cytosolic protein. Some less abundant JIPs were also localized within different cell compartments: JIP‐100 was found within the stromal fraction of chloroplasts; JIP‐70 is present in the peroxisome and the nucleus; JIP‐50 and JIP‐6 accumulate in vacuoles. The location of JIP‐66 and JIP‐6 confirms their possible physiological role deduced from molecular analysis of their cDNA.} } @Article{IPB-2469, author = {Abel, S. and Theologis, A. and}, title = {{Transient transformation of Arabidopsis leaf protoplasts: a versatile experimental system to study gene expression}}, year = {1994}, pages = {421-427}, journal = {Plant J.}, doi = {10.1111/j.1365-313X.1994.00421.x}, volume = {5}, abstract = {An improved protocol is reported to isolate and transiently transform mesophyll protoplasts of Arabidopsis thaliana. Transfected leaf protoplasts support high levels of expression of the bacterial reporter gene coding for β‐glucuronidase (GUS), under the control of the cauliflower mosaic virus (CaMV) 35S promoter. Transient expression of GUS activity was monitored spectrophotometrically and reached a maximum between 18 and 48 h after polyethylene glycol (PEG)‐mediated DNA uptake. Histochemical staining for GUS activity revealed reproducible transformation frequencies between 40 and 60%, based on the number of protoplasts survived. To demonstrate the applicability of the transient expression system, the subcellular localization of GUS proteins tagged with different nuclear polypeptides was studied in transfected mesophyll protoplasts, revealing nuclear compartmentalization of the chimeric GUS enzymes. Furthermore, Arabidopsis mesophyll protoplasts support auxin‐mediated induction of chloramphenicol acetyl‐transferase (CAT) activity when transfected with a transcriptional fusion between the CAT reporter gene and the early auxin‐inducible PS‐IAA4/5 promoter. Hence, the method allows in vivo analysis of promoter activity and subcellular localization of fusion proteins in a homologous transformation system.} } @Article{IPB-2468, author = {Abel, S. and Oeller, P. W. and Theologis, A. and}, title = {{Early auxin-induced genes encode short-lived nuclear proteins.}}, year = {1994}, pages = {326-330}, journal = {Proc. Natl. Acad. Sci. U.S.A.}, doi = {10.1073/pnas.91.1.326}, volume = {91}, abstract = {The plant growth hormone indoleacetic acid (IAA) transcriptionally activates gene expression in plants. Some of the genes whose expression is induced by IAA encode a family of proteins in pea (PS-IAA4 and PS-IAA6) and Arabidopsis (IAA1 and IAA2) that contain putative nuclear localization signals that direct a beta-glucuronidase reporter protein into the nucleus. Pulse-chase and immunoprecipitation experiments have defined the t1/2 of the PS-IAA4 and PS-IAA6 proteins to be 8 and 6 min, respectively. Their most prominent feature is the presence of a beta alpha alpha motif similar to the beta-sheet DNA-binding domain found in prokaryotic repressors of the Arc family. Based on these data, we suggest that plant tissues express short-lived nuclear proteins as a primary response to IAA. We propose that these proteins act as activators or repressors of genes responsible for mediating the various auxin responses.} }