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Publikation

Wasternack, C.; Xie, D.; The genuine ligand of a jasmonic acid receptor: Improved analysis of jasmonates is now required Plant Signal Behav. 5, 337-340, (2010) DOI: 10.4161/psb.5.4.11574

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.
Publikation

Wasternack, C.; Kombrink, E.; Jasmonates: Structural Requirements for Lipid-Derived Signals Active in Plant Stress Responses and Development ACS Chem. Biol. 5, 63-77, (2010) DOI: 10.1021/cb900269u

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.
Publikation

Stumpe, M.; Göbel, C.; Faltin, B.; Beike, A. K.; Hause, B.; Himmelsbach, K.; Bode, J.; Kramell, R.; Wasternack, C.; Frank, W.; Reski, R.; Feussner, I.; The moss Physcomitrella patens contains cyclopentenones but no jasmonates: mutations in allene oxide cyclase lead to reduced fertility and altered sporophyte morphology New Phytol. 188, 740-749, (2010) DOI: 10.1111/j.1469-8137.2010.03406.x

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.
Publikation

Sreenivasulu, N.; Radchuk, V.; Alawady, A.; Borisjuk, L.; Weier, D.; Staroske, N.; Fuchs, J.; Miersch, O.; Strickert, M.; Usadel, B.; Wobus, U.; Grimm, B.; Weber, H.; Weschke, W.; De-regulation of abscisic acid contents causes abnormal endosperm development in the barley mutant seg8 Plant J. 64, 589-603, (2010) DOI: 10.1111/j.1365-313X.2010.04350.x

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.
Publikation

Robson, F.; Okamoto, H.; Patrick, E.; Harris, S.-R.; Wasternack, C.; Brearley, C.; Turner, J. G.; Jasmonate and Phytochrome A Signaling in Arabidopsis Wound and Shade Responses Are Integrated through JAZ1 Stability Plant Cell 22, 1143-1160, (2010) DOI: 10.1105/tpc.109.067728

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.
Publikation

Leon-Reyes, A.; Van der Does, D.; De Lange, E. S.; Delker, C.; Wasternack, C.; Van Wees, S. C. M.; Ritsema, T.; Pieterse, C. M. J.; Salicylate-mediated suppression of jasmonate-responsive gene expression in Arabidopsis is targeted downstream of the jasmonate biosynthesis pathway Planta 232, 1423-1432, (2010) DOI: 10.1007/s00425-010-1265-z

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.
Bücher und Buchkapitel

Wasternack, C.; Jasmonates in Stress, Growth, and Development 91-118, (2010) ISBN: 9783527628964 DOI: 10.1002/9783527628964.ch5

This chapter contains sections titled:IntroductionJA BiosynthesisJA MetabolismBound OPDA – ArabidopsidesMutants of JA Biosynthesis and SignalingCOI1–JAZ–JA‐Ile‐Mediated JA SignalingTranscription Factors Involved in JA SignalingJasmonates and Oxylipins in DevelopmentConclusionsAcknowledgmentsReferences
Publikation

Gerhardt, B.; Fischer, K.; Balkenhohl, T. J.; Pohnert, G.; Kühn, H.; Wasternack, C.; Feussner, I.; 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 Planta 220, 919-930, (2005) DOI: 10.1007/s00425-004-1408-1

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.
Publikation

Fortes, A. M.; Miersch, O.; Lange, P. R.; Malhó, R.; Testillano, P. S.; Risueño, M. d. C.; Wasternack, C.; Pais, M. S.; Expression of Allene Oxide Cyclase and Accumulation of Jasmonates during Organogenic Nodule Formation from Hop (Humulus lupulus var. Nugget) Internodes Plant Cell Physiol. 46, 1713-1723, (2005) DOI: 10.1093/pcp/pci187

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.
Publikation

Durgbanshi, A.; Arbona, V.; Pozo, O.; Miersch, O.; Sancho, J. V.; Gómez-Cadenas, A.; Simultaneous Determination of Multiple Phytohormones in Plant Extracts by Liquid Chromatography−Electrospray Tandem Mass Spectrometry J. Agr. Food Chem. 53, 8437-8442, (2005) DOI: 10.1021/jf050884b

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.
  • Die Leibniz-Gemeinschaft
  • Digitalisierung in der Landwirtschaft
  • Martin-Luther Universität Halle
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