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Salicylic acid (SA) and acetylsalicylic acid (ASA), previously shown to inhibit proteinase inhibitor synthesis induced by wound- Willmitzer [19931 Planta 191: 123-128), are shown here t o be potent inhibitors of systemin-induced and jasmonic acid (JA)-induced synthesis of proteinase inhibitor mRNAs and proteins. The inhibition by SA and ASA of proteinase inhibitor synthesis induced by systemin and JA, as well as by wounding and oligosaccharide elicitors, provides further evidence that both oligosaccharide and polypeptide inducer molecules utilize the octadecanoid pathway to signal the activation of proteinase inhibitor genes. Tomato (Lycopersicon esculentum) leaves were pulse labeled with [35S]methionine, followed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and the inhibitory effects of SA are shown to be specific for the synthesis of a small number of JA-inducible proteins that includes the proteinase inhibitors. Previous results have shown that SA inhibits the conversion of 1 3s-hydroperoxy linolenic acid t o 12-0x0-phytodienoic acid, thereby inhibiting the signaling pathway by blocking synthesis of JA. Here we report that the inhibition of synthesis of proteinase inhibitor proteins and mRNAs by SA in both light and darkness also occurs at a step i n the signal transduction pathway, after JA synthesis but preceding transcription of the inhibitor genes.

An intracellular signaling pathway for activating plant defense genes against attacking herbivores and pathogens is mediated by a lipid-based signal transduction cascade. I n this pathway, linolenic acid (18:3) is proposed to be liberated from cell membranes and is converted to cyclopentanones that are involved in transcriptional regulation of defense genes, analogously to prostaglandin synthesis and function in animals. Levels of 18:3 and linoleic acid in tomato (Lycopersicon esculentum) leaves increased within 1 h when the leaves were wounded with a hemostat across the main vein t o simulate herbivore attacks. The increase correlated with the time course of accumulation of jasmonic acid, a cyciopentanone product of 18:3, that had previously been shown t o increase i n leaves in response both to wounding and to elicitors of plant defense genes. One hour after wounding, at least a 15-fold excess of 18:3 was found over that required to account for the levels of newly synthesized jasmonic acid. The free fatty acids in both control and wounded leaves accounted for less than 0.25% of the total fatty acids. However, the total lipid contents of the leaves remained relatively unchanged up to 8 h after wounding, indicating that extensive loss of lipids did not occur, although a gradual decrease in polar lipids was observed, mainly in monogalactosyl diacylglycerol of chloroplast lipids. The data support a role for lipid release as a key step in the signaling events that activate defense genes in tomato leaves in response t o wounding by attacking herbivores. In leaves of tomato (Lycopersicon esculentum) plants, severa1defense genes are activated by herbivore attacks or other mechanical wounds that crush the tissues (Ryan, 1992; Schaller and Ryan, 1996). Among the activated genes are those encoding Ser, aspartyl, and Cys proteinase i h b i t o r s and the enzyme polyphenol oxidase. These genes are activated in leaves both nearby and distant from the wound sites (Ryan, 1992; Schaller and Ryan, 1996). The signals released to local cells have been attributed to (a) oligosaccharide fragments of the plant and/or pathogen cell walls (Bishop et al., 1984;Darvill and Albersheim, 1984), (b) 18:3 (Farmer and Ryan, 1992), a major component of plant membranes, and (c)

Many plant genes that respond to environmental and developmental changes are regulated by jasmonic acid, which is derived from linolenic acid via the octadecanoid pathway. Linolenic acid is an important fatty-acid constituent of membranes in most plant species and its intracellular levels increase in response to certain signals. Here we report that irradiation of tomato leaves with ultraviolet light induces the expression of several plant defensive genes that are normally activated through the octadecanoid pathway after wounding. The response to ultraviolet light is blocked by an inhibitor of the octadecanoid pathway and it does not occur in a tomato mutant defective in this pathway. The ultraviolet irradiation maximally induces the defence genes at levels where cyclobutane pyrimidine dimer formation, an indicator of DNA damage, is less than 0.2 dimers per gene. Our evidence indicates that this plant defence response to certain wavelengths of ultraviolet radiation requires the activation of the octadecanoid defence signalling pathway.

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