Benno Parthier

Accumulation of jasmonate, abscisic acid, specific transcripts and proteins in osmotically stressed barley leaf segments

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.

Occurrence and identification of jasmonic acid and its amino acid conjugates induced by osmotic stress in barley leaf tissue

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.

Novel natural substances acting in plant growth regulation

This review includes novel natural products of microbial and plant origin reported to exert plant growth-regulating activities. The well-known “classic” plant hormones are excluded. The substances described with their biological activities are restricted to those that have been isolated from microbial or plant sources later than 1990 or that have been intensively investigated during the last few years, such as jasmonic acid and brassinosteroids.

Structure–activity relations of substituted, deleted or stereospecifically altered jasmonic acid in gene expression of barley leaves

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.

Amino acid conjugates of jasmonic acid induce jasmonate-responsive gene expression in barley (Hordeum vulgare L.) leaves

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.

Purification and Characterization of Allene Oxide Cyclase from Dry Corn Seeds

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.

Jasmonate signalling can be uncoupled from abscisic acid signalling in barley: identification of jasmonate-regulated transcripts which are not induced by abscisic acid

Jasmonate and abscisic acid induce several identical mRNAs and proteins in barley. In order to study whether both hormones act through the same signalling pathway, we identified four transcripts induced by jasmonic acid methylester (JM) in leaf segments of barley (Hordeum vulgare L. cv. Salome). These newly identified transcripts were not induced by abscisic acid within the tested times of 2–72 h. This finding supports the conclusion that jasmonate signalling in barley is independent of abscisic acid, in contrast to the wound-induction signal cascade of proteinase-inhibitor II in tomato and potato. Of the four isolated cDNAs, the putative translation frame of one was homologous to caffeic acid methyltransferase, another was homologous to chalcone synthase, and the C-terminus of the third showed homology to two proteins from rice (a salt-induced protein and a root-specific protein); the last cDNA was not homologous to any sequences in the databases. The new cDNAs will be valuable tools for studying jasmonate signal transduction in barley.

Diversity in octadecanoid-induced gene expression of tomato

Summary 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 s-oxidized to give a JA response, (ii) Octadecanoids which can not be s-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 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.

Methyljasmonate-induced accumulation of coumaroyl conjugates in barley leaf segments.

The effect of methyljasmonate on the induction of phenolic components in barley leaf segments was investigated. RP-HPLC of methanol extracts showed that three compounds accumulate to high concentrations in response to methyljasmonate treatment. Two of them were identified as N-(E)-4-coumaroylputrescine and N-(E)-4-coumaroylagmatine by UV-spectroscopy and mass spectrometry.

Jasmonic acid: biosynthesis, signal transduction, gene expression

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 stress-induced 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 he presented and discussed in terms of JA as a master switch among various lipid-derived signals.