TY - JOUR ID - 2124 TI - Jasmonate biosynthesis and the allene oxide cyclase family of Arabidopsis thaliana JO - Plant Mol. Biol. PY - 2003 SP - 895-911 AU - Stenzel, I. AU - Hause, B. AU - Miersch, O. AU - Kurz, T. AU - Maucher, H. AU - Weichert, H. AU - Ziegler, J. AU - Feussner, I. AU - Wasternack, C. AU - VL - 51 UR - DO - 10.1023/A:1023049319723 AB - 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. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2089 TI - Occurrence of the allene oxide cyclase in different organs and tissues of Arabidopsis thaliana JO - Phytochemistry PY - 2003 SP - 971-980 AU - Hause, B. AU - Stenzel, I. AU - Miersch, O. AU - Wasternack, C. AU - VL - 64 UR - DO - 10.1016/S0031-9422(03)00447-3 AB - 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. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2088 TI - Enzymes of Jasmonate Biosynthesis Occur in Tomato Sieve Elements JO - Plant Cell Physiol. PY - 2003 SP - 643-648 AU - Hause, B. AU - Hause, G. AU - Kutter, C. AU - Miersch, O. AU - Wasternack, C. AU - VL - 44 UR - DO - 10.1093/pcp/pcg072 AB - 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. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2086 TI - Biochemical and Molecular Characterization of a Hydroxyjasmonate Sulfotransferase from Arabidopsis thaliana JO - J. Biol. Chem. PY - 2003 SP - 17895-17900 AU - Gidda, S. K. AU - Miersch, O. AU - Levitin, A. AU - Schmidt, J. AU - Wasternack, C. AU - Varin, L. AU - VL - 278 UR - DO - 10.1074/jbc.M211943200 AB - 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. A2 - C1 - Bioorganic Chemistry; Molecular Signal Processing ER - TY - JOUR ID - 2084 TI - Selective desensitization of jasmonate- and pH-dependent signaling in the induction of benzophenanthridine biosynthesis in cells of Eschscholzia californica JO - Phytochemistry PY - 2003 SP - 491-500 AU - Färber, K. AU - Schumann, B. AU - Miersch, O. AU - Roos, W. AU - VL - 62 UR - DO - 10.1016/S0031-9422(02)00562-9 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2082 TI - Saturation of two chromosome regions conferring resistance to SCMV with SSR and AFLP markers by targeted BSA JO - Theor. Appl. Genet. PY - 2003 SP - 485-493 AU - Dußle, C. AU - Quint, M. AU - Melchinger, A. AU - Xu, M. AU - Lübberstedt, T. AU - VL - 106 UR - DO - 10.1007/s00122-002-1107-x AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2080 TI - 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 JO - Nutr. Cancer PY - 2003 SP - 212-221 AU - Dingley, K. H. AU - Ubick, E. A. AU - Chiarappa-Zucca, M. L. AU - Nowell, S. AU - Abel, S. AU - Ebeler, S. E. AU - Mitchell, A. E. AU - Burns, S. A. AU - Steinberg, F. M. AU - Clifford, A. J. AU - VL - 46 UR - DO - 10.1207/S15327914NC4602_15 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2079 TI - Peripheral regions of natural hammerhead ribozymes greatly increase their self-cleavage activity JO - EMBO J. PY - 2003 SP - 5561-5570 AU - De la Peña, M. AU - Gago, S. AU - Flores, R. AU - VL - 22 UR - DO - 10.1093/emboj/cdg530 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2070 TI - Jasmonate and octadecanoid occurrence in tomato hairy roots. Endogenous level changes in response to NaCl JO - Plant Growth Regul. PY - 2003 SP - 21-27 AU - Abdala, G. AU - Miersch, O. AU - Kramell, R. AU - Vigliocco, A. AU - Agostini, E. AU - Forchetti, G. AU - Alemano, S. AU - VL - 40 UR - DO - 10.1023/A:1023016412454 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2117 TI - Substrate Specificity of Glutaminyl Cyclases from Plants and Animals JO - Biol. Chem. PY - 2003 SP - 1583-1592 AU - Schilling, S. AU - Manhart, S. AU - Hoffmann, T. AU - Ludwig, H.-H. AU - Wasternack, C. AU - Demuth, H.-U. AU - VL - 384 UR - DO - 10.1515/BC.2003.175 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2116 TI - Identification of Human Glutaminyl Cyclase as a Metalloenzyme JO - J. Biol. Chem. PY - 2003 SP - 49773-49779 AU - Schilling, S. AU - Niestroj, A. J. AU - Rahfeld, J.-U. AU - Hoffmann, T. AU - Wermann, M. AU - Zunkel, K. AU - Wasternack, C. AU - Demuth, H.-U. AU - VL - 278 UR - DO - 10.1074/jbc.M309077200 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2111 TI - Identification of genetically linked RGAs by BAC screening in maize and implications for gene cloning, mapping and MAS JO - Theor. Appl. Genet. PY - 2003 SP - 1171-1177 AU - Quint, M. AU - Dußle, C. M. AU - Melchinger, A. E. AU - Lübberstedt, T. AU - VL - 106 UR - DO - 10.1007/s00122-002-1105-z AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2106 TI - Multiple Hormones Act Sequentially to Mediate a Susceptible Tomato Pathogen Defense Response JO - Plant Physiol. PY - 2003 SP - 1181-1189 AU - O'Donnell, P. J. AU - Schmelz, E. AU - Block, A. AU - Miersch, O. AU - Wasternack, C. AU - Jones, J. B. AU - Klee, H. J. AU - VL - 133 UR - DO - 10.1104/pp.103.030379 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2104 TI - 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 JO - Virus Res. PY - 2003 SP - 49-61 AU - Naum-Onganı́a, G. AU - Gago-Zachert, S. AU - Peña, E. AU - Grau, O. AU - Laura Garcia, M. AU - VL - 96 UR - DO - 10.1016/S0168-1702(03)00172-2 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2102 TI - Novel plasmid vectors for homologous transformation of barley (Hordeum vulgare L.) with JIP23 cDNA in sense and antisense orientation JO - Cereal Res. Commun. PY - 2003 SP - 17-24 AU - Monostori, T. AU - Schulze, J. AU - Sharma, V. K. AU - Maucher, H. AU - Wasternack, C. AU - Hause, B. AU - VL - 31 UR - DO - 10.1007/BF03543245 AB - 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. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2123 TI - Differential expression of the LePS2 phosphatase gene family in response to phosphate availability, pathogen infection and during development JO - Physiol. Plant. PY - 2003 SP - 138-146 AU - Stenzel, I. AU - Ziethe, K. AU - Schurath, J. AU - Hertel, S. C. AU - Bosse, D. AU - Köck, M. AU - VL - 118 UR - DO - 10.1034/j.1399-3054.2003.00091.x AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2122 TI - Allene oxide cyclase dependence of the wound response and vascular bundle-specific generation of jasmonates in tomato - amplification in wound signalling JO - Plant J. PY - 2003 SP - 577-589 AU - Stenzel, I. AU - Hause, B. AU - Maucher, H. AU - Pitzschke, A. AU - Miersch, O. AU - Ziegler, J. AU - Ryan, C. A. AU - Wasternack, C. AU - VL - 33 UR - DO - 10.1046/j.1365-313X.2003.01647.x AB - 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. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - CHAP ID - 149 TI - Shift in Fatty Acid and Oxylipin Pattern of Tomato Leaves Following Overexpression of the Allene Oxide Cyclase T2 - Advanced Research on Plant Lipids PB - PY - 2003 SP - 275-278 AU - Weichert, H. AU - Maucher, H. AU - Hornung, E. AU - Wasternack, C. AU - Feussner, I. AU - VL - UR - DO - 10.1007/978-94-017-0159-4_64 AB - Polyunsaturated fatty acids (PUFAs) are a source of numerous oxidation products, the oxylipins. In leaves, α-linolenic acid (α-LeA) is the preferential substrate for lipid peroxidation reactions. This reaction may be catalyzed either by a 9-lipoxygenase (9-LOX) or by a 13-LOX and oxygen is inserted regioselectively as well as stereospecifically leading to formation of 13S- or 9S-hydroperoxy octadecatrienoic acid (13-/9-HPOT; Brash, 1999). At least, seven different enzyme families or reaction branches within the LOX pathway can use these HPOTs or other hydroperoxy PUFAs leading to (i) keto-PUFAs (LOX); (ii) epoxy hydroxy-PUFAs (epoxy alcohol synthase, EAS); (iii) octadecanoids and jasmonates (allene oxide synthase, AOS); (iv) leaf aldehydes and leaf alcohols (hydroperoxide lyase, HPL); (v) hydroxy PUFAs (reductase); (vi) divinyl ether PUFAs (divinyl ether synthase, DES); and (vii) epoxy- or dihydrodiol-PUFAs (peroxygenase, PDX; Fig. 1; Feussner and Wasternack, 2002). A2 - C1 - Molecular Signal Processing ER - TY - CHAP ID - 147 TI - The Lipoxygenase Pathway in Mycorrhizal Roots of Medicago Truncatula T2 - Advanced Research on Plant Lipids PB - PY - 2003 SP - 287-290 AU - Stumpe, M. AU - Stenzel, I. AU - Weichert, H. AU - Hause, B. AU - Feussner, I. AU - VL - UR - DO - 10.1007/978-94-017-0159-4_67 AB - Mycorrhizas are by far the most frequent occurring beneficial symbiotic interactions between plants and fungi. Species in >80% of extant plant families are capable of establishing an arbuscular mycorrhiza (AM). In relation to the development of the symbiosis the first molecular modifications are those associated with plant defense responses, which seem to be locally suppressed to levels compatible with symbiotic interaction (Gianinazzi-Pearson, 1996). AM symbiosis can, however, reduce root disease caused by several soil-borne pathogens. The mechanisms underlying this protective effect are still not well understood. In plants, products of the enzyme lipoxygenase (LOX) and the corresponding downstream enzymes, collectively named LOX pathway (Fig. 1B), are involved in wound healing, pest resistance, and signaling, or they have antimicrobial and antifungal activity (Feussner and Wasternack, 2002). The central reaction in this pathway is catalyzed by LOXs leading to formation of either 9- or 13-hydroperoxy octadeca(di/trien)oic acids (9/13-HPO(D/T); Brash, 1999). Thus LOXs may be divided into 9- and 13-LOXs (Fig. 1A). Seven different reaction branches within this pathway can use these hydroperoxy polyenoic fatty acids (PUFAs) leading to (i) keto PUFAs by a LOX; (ii) epoxy hydroxy-fatty acids by an epoxy alcohol synthase (EAS); (iii) octadecanoids and jasmonates via allene oxide synthase (AOS); (iv) leaf aldehydes and leaf alcohols via fatty acid hydroperoxide lyase (HPL); (v) hydroxy PUFAs (reductase); (vi) divinyl ether PUFAs via divinyl ether synthase (DES); and (vii) epoxy- or dihydrodiolPUFAs via peroxygenase (PDX; Feussner and Wasternack, 2002). AOS, HPL and DES belong to one subfamily of P450-containing enzymes, the CYP74 family (Feussner and Wasternack, 2002). Here, the involvement of this CYP74 enzyme family in mycorrhizal roots of M. truncatula during early stages of AM symbiosis formation was analyzed. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - CHAP ID - 145 TI - Transcriptional Activation of Jasmonate Biosynthesis Enzymes is not Reflected at Protein Level T2 - Advanced Research on Plant Lipids PB - PY - 2003 SP - 267-270 AU - Stenzel, I. AU - Hause, B. AU - Feussner, I. AU - Wasternack, C. AU - VL - UR - DO - 10.1007/978-94-017-0159-4_62 AB - Jasmonic acid (JA) and its precursor 12-oxo phytodienoic acid (OPDA) are lipid-derived signals in plant stress responses and development (Wasternack and Hause, 2002). Within the wound-response pathway of tomato, a local response of expression of defense genes such as the proteinase inhibitor 2 gene (PIN2) is preceded by a rise in JA (Herde et al., 1996; Howe et al., 1996) and ethylene (O’Donnell et al., 1996). Mutants affected in JA biosynthesis such as defl (Howe et al., 1996) or spr-2 (Li et al., 2002) clearly indicated that JA biosynthesis is an ultimate part of wound signaling. It is less understood, however, how the rise in JA is regulated. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2184 TI - Heterologous Expression and Characterization of Human Glutaminyl Cyclase: Evidence for a Disulfide Bond with Importance for Catalytic Activity JO - Biochemistry PY - 2002 SP - 10849-10857 AU - Schilling, S. AU - Hoffmann, T. AU - Rosche, F. AU - Manhart, S. AU - Wasternack, C. AU - Demuth, H.-U. AU - VL - 41 UR - DO - 10.1021/bi0260381 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2183 TI - Continuous Spectrometric Assays for Glutaminyl Cyclase Activity JO - Anal. Biochem. PY - 2002 SP - 49-56 AU - Schilling, S. AU - Hoffmann, T. AU - Wermann, M. AU - Heiser, U. AU - Wasternack, C. AU - Demuth, H.-U. AU - VL - 303 UR - DO - 10.1006/abio.2001.5560 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2181 TI - Development of RGA-CAPS markers and genetic mapping of candidate genes for sugarcane mosaic virus resistance in maize JO - Theor. Appl. Genet. PY - 2002 SP - 355-363 AU - Quint, M. AU - Mihaljevic, R. AU - Dussle, C. AU - Xu, M. AU - Melchinger, A. AU - Lübberstedt, T. AU - VL - 105 UR - DO - 10.1007/s00122-002-0953-x AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2179 TI - Cell death and salicylate- and jasmonate-dependent stress responses in Arabidopsis are controlled by single cet genes JO - Planta PY - 2002 SP - 120-128 AU - Nibbe, M. AU - Hilpert, B. AU - Wasternack, C. AU - Miersch, O. AU - Apel, K. AU - VL - 216 UR - DO - 10.1007/s00425-002-0907-1 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2174 TI - FQR1, a Novel Primary Auxin-Response Gene, Encodes a Flavin Mononucleotide-Binding Quinone Reductase JO - Plant Physiol. PY - 2002 SP - 578-590 AU - Laskowski, M. J. AU - Dreher, K. A. AU - Gehring, M. A. AU - Abel, S. AU - Gensler, A. L. AU - Sussex, I. M. AU - VL - 128 UR - DO - 10.1104/pp.010581 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2164 TI - Induction of Jasmonate Biosynthesis in Arbuscular Mycorrhizal Barley Roots JO - Plant Physiol. PY - 2002 SP - 1213-1220 AU - Hause, B. AU - Maier, W. AU - Miersch, O. AU - Kramell, R. AU - Strack, D. AU - VL - 130 UR - DO - 10.1104/pp.006007 AB - 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. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2162 TI - Identification of Arabidopsis mutants with altered glucosinolate profiles based on isothiocyanate bioactivity JO - Plant Sci. PY - 2002 SP - 143-152 AU - Grubb, C. D. AU - Gross, H. B. AU - Chen, D. L. AU - Abel, S. AU - VL - 162 UR - DO - 10.1016/S0168-9452(01)00550-7 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2156 TI - The lipoxygenase pathway JO - Annu. Rev. Plant Biol. PY - 2002 SP - 275-297 AU - Feussner, I. AU - Wasternack, C. AU - VL - 53 UR - DO - 10.1146/annurev.arplant.53.100301.135248 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2151 TI - The Arabidopsis Mutant cev1 Links Cell Wall Signaling to Jasmonate and Ethylene Responses JO - Plant Cell PY - 2002 SP - 1557-1566 AU - Ellis, C. AU - Karafyllidis, I. AU - Wasternack, C. AU - Turner, J. G. AU - VL - 14 UR - DO - 10.1105/tpc.002022 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2150 TI - Conversion of AFLP fragments tightly linked to SCMV resistance genes Scmv1 and Scmv2 into simple PCR-based markers JO - Theor. Appl. Genet. PY - 2002 SP - 1190-1195 AU - Dussle, C. AU - Quint, M. AU - Xu, M. AU - Melchinger, A. AU - Lübberstedt, T. AU - VL - 105 UR - DO - 10.1007/s00122-002-0964-7 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2196 TI - Metabolic profiling of oxylipins in germinating cucumber seedlings - lipoxygenase-dependent degradation of triacylglycerols and biosynthesis of volatile aldehydes JO - Planta PY - 2002 SP - 612-619 AU - Weichert, H. AU - Kolbe, A. AU - Kraus, A. AU - Wasternack, C. AU - Feussner, I. AU - VL - 215 UR - DO - 10.1007/s00425-002-0779-4 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2195 TI - Direct analysis of single leaf disks for chemopreventive glucosinolates JO - Phytochem. Anal. PY - 2002 SP - 152-157 AU - Wang, Q. AU - Grubb, C. D. AU - Abel, S. AU - VL - 13 UR - DO - 10.1002/pca.636 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2191 TI - Stimulation of jasmonic acid production in Zea Mays L. infected by the maize rough dwarf virus - Río Cuarto. Reversion of symptoms by salicylic acid JO - Biocell PY - 2002 SP - 369-374 AU - Vigliocco, A. AU - Bonamico, B. AU - Alemano, S. AU - Miersch, O. AU - Abdala, G. AU - VL - 26 UR - https://www.techscience.com/biocell/v26n3/34012 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2137 TI - Jasmonate-Induced Lipid Peroxidation in Barley Leaves Initiated by Distinct 13-LOX Forms of Chloroplasts JO - Biol. Chem. PY - 2002 SP - 1645-1657 AU - Bachmann, A. AU - Hause, B. AU - Maucher, H. AU - Garbe, E. AU - Vörös, K. AU - Weichert, H. AU - Wasternack, C. AU - Feussner, I. AU - VL - 383 UR - DO - 10.1515/BC.2002.185 AB - 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. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER - TY - JOUR ID - 2134 TI - Phosphate sensing in higher plants JO - Physiol. Plant. PY - 2002 SP - 1-8 AU - Abel, S. AU - Ticconi, C. A. AU - Delatorre, C. A. AU - VL - 115 UR - DO - 10.1034/j.1399-3054.2002.1150101.x AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - JOUR ID - 2133 TI - Changes in jasmonate and gibberellin levels during development of potato plants (Solanum tuberosum) JO - Plant Growth Regul. PY - 2002 SP - 121-126 AU - Abdala, G. AU - Castro, G. AU - Miersch, O. AU - Pearce, D. AU - VL - 36 UR - DO - 10.1023/A:1015065011536 AB - 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. A2 - C1 - Molecular Signal Processing ER - TY - CHAP ID - 159 TI - Jasmonates and octadecanoids: Signals in plant stress responses and development T2 - PB - Prog. Nucleic Acid Res. Mol. Biol. PY - 2002 SP - 165-221 AU - Wasternack, C. AU - Hause, B. AU - VL - 72 UR - DO - 10.1016/S0079-6603(02)72070-9 AB - Plants are sessile organisms. Consequently they have to adapt constantly to fluctuations in the environment. Some of these changes involve essential factors such as nutrients, light, and water. Plants have evolved independent systems to sense nutrients such as phosphate and nitrogen. However, many of the environmental factors may reach levels which represent stress for the plant. The fluctuations can range between moderate and unfavorable, and the factors can be of biotic or abiotic origin. Among the biotic factors influencing plant life are pathogens and herbivores. In case of bacteria and fungi, symbiotic interactions such as nitrogen-fixating nodules and mycorrhiza, respectively, may be established. In case of insects, a tritrophic interaction of herbivores, carnivores, and plants may occur mutualistically or parasitically. Among the numerous abiotic factors are low temperature, frost, heat, high light conditions, ultraviolet light, darkness, oxidation stress, hypoxia, wind, touch, nutrient imbalance, salt stress, osmotic adjustment, water deficit, and desiccation.In the last decade jasmonates were recognized as being signals in plant responses to most of these biotic and abiotic factors. Signaling via jasmonates was found to occur intracellularly, and systemically as well as interorganismically. Jasmonates are a group of ubiquitously occurring plant growth regulators originally found as the major constituents in the etheric oil of jasmine, and were first suggested to play a role in senescence due to a strong senescence-promoting effect. Subsequently, numerous developmental processes were described in which jasmonates exhibited hormone-like properties. Recent knowledge is reviewed here on jasmonates and their precursors, the octadecanoids. After discussing occurrence and biosynthesis, emphasis is placed upon the signal transduction pathways in plant stress responses in which jasmonates act a signal. Finally, examples are described on the role of jasmonates in developmental processes. A2 - C1 - Molecular Signal Processing; Cell and Metabolic Biology ER -