The octadecanoic pathway: signal molecules for the regulation of secondary pathways.

Plant defense against microbial pathogens and herbivores relies heavily on the induction of defense proteins and low molecular weight antibiotics. The signals between perception of the aggression, gene activation, and the subsequent biosynthesis of secondary compounds are assumed to be pentacylic oxylipin derivatives. The rapid, but transient, synthesis of cis-jasmonic acid was demonstrated after insect attack on a food plant and by microbial elicitor addition to plant suspension cultures. This effect is highly specific and not caused by a number of environmental stresses such as light, heavy metals, or cold or heat shock. Elicitation of Eschscholtzia cell cultures also led to a rapid alkalinization of the growth medium prior to jasmonate formation. Inhibition of this alkalinization process by the protein kinase inhibitor staurosporine also inhibited jasmonate formation. The induction of specific enzymes in the benzo[c]phenanthridine alkaloid pathway leading to the antimicrobial sanguinarine was induced to a qualitatively and quantitatively similar extent by fungal elicitor, methyl jasmonate, and its linolenic acid-derived precursor 12-oxophytodienoic acid. It is herein proposed that a second oxylipid cascade may exist in plants starting from linoleic acid via 15,16-dihydro-12-oxophytodienoic acid to 9,10-dihydrojasmonate. Experiments with synthetic trihomojasmonate demonstrated that beta-oxidation is not a prerequisite for biological activity and that 12-oxophytodienoic acid and derivatives are most likely fully active as signal transducers. Octadecanoic acid-derived compounds are essential elements in modulating the synthesis of antibiotic compounds and are thus integral to plant defense.

Quantification of jasmonic acid by capillary gas chromatography-negative chemical ionization-mass spectrometry.

Jasmonic acid can be assayed by the highly sensitive and reproducible gas chromatography-negative ion chemical ionization-mass spectrometry (GC-NICI-MS) technique. We describe an optimized sample preparation method for routine analysis of jasmonic acids which allows the analysis of more than 10 samples/day. The basis of the method resides in a one-step extraction and phase partition, selective adsorption/elution of acids with an aminopropyl column, conversion of acids to pentafluorobenzyl esters, final purification using a silica column, and GC-NICI-MS. Extracted standard curves were linear over an assay range of 1 to 1000 ng; correlation coefficients were typically greater than 0.9999. The limit of detection was 500 fg (2.4 fmol) jasmonic acid/injection at a signal to noise ratio of 10:1. Furthermore, the method was modified for steric analysis of jasmonic acid stereoisomers at the nanogram level. (3R,7S)-(+)-7-Iso-jasmonic acid was found to be biosynthesized from plant tissue cell cultures upon elicitation. This stereoisomer epimerizes rapidly under alkaline and acidic conditions and in the presence of albumin. Epimerization rate constants were determined at different pH values.

The Pseudomonas phytotoxin coronatine mimics octadecanoid signalling molecules of higher plants.

The phytotoxic principle, coronatine, which is present in several pathovars of the plant pathogen, Pseudomonas syringae was shown to be highly active in completely different, jasmonate-selective bioassays. At nanomolar to micromolar concentrations, coronatine induced the accumulation of defense-related secondary metabolites in several plant cell cultures, induced transcript accumulation of the elicitor-responsive gene encoding the berberine bridge enzyme of Eschscholtzia californica, as well as the coiling response of Bryonia dioica tendrils. Biological activity critically depended upon the structure of coronatine, and slight modifications, such as methylation of the carboxyl moiety or reduction of the carbonyl group, rendered the molecules almost inactive. Coronafacic acid, obtained by hydrolysis of coronatine, was also nearly inactive. Coronatine did not elicit the accumulation of endogenous jasmonic acid in the systems analyzed. While coronafacic acid is similar in structure to jasmonic acid, we found coronatine to be a close structural analogue of the cyclic C18-precursor of jasmonic acid, 12-oxo-phytodienoic acid. The phytotoxic symptoms produced by coronatine can now be understood on the basis of the toxin's action as a mimic of the octadecanoid signalling molecules of higher plants.

Signaling in the elicitation process is mediated through the octadecanoid pathway leading to jasmonic acid.

Fungal cell walls and fragments thereof (elicitors) induce the formation of low and high molecular weight defense compounds in plant cell suspension cultures. This induced synthesis requires a signal molecule transmitting the message between the elicitor plant cell wall receptor and gene activation. We demonstrate in this study that cis-jasmonic acid is rapidly synthesized in plant cell cultures of diverse taxonomic origin (gymnosperms and mono- and dicotyledonous plants) after challenge with a fungal elicitor preparation. The rapid decline of cis-jasmonic acid in some of these tissues is attributed to rapid metabolism of this pentacyclic acid. The induction of alkaloids by several different molecules provoking the elicitation process is strictly correlated with the synthesis of jasmonates. Elicitation leads to a rapid release of alpha-linolenic acid from the lipid pool of the plant cell. alpha-Linolenic acid and 12-oxophytodienoic acid, the formation of which is also induced, are known to be distant precursors of jasmonic acid. We assume cis-jasmonic acid and its precursors to be the signaling molecules in the elicitation process.