Effects of active oxygen species and methyl jasmonate on expression of the gene for a wound-inducible 1-aminocyclopropane-1-carboxylate synthase in winter squash (Cucurbita maxima)

Abstract.

The production of ethylene by mesocarp tissue of winter squash is induced by mechanical wounding. We isolated a cDNA fragment (WSACS2) from the wounded mesocarp tissue of winter-squash (Cucurbita maxima Duch.) fruit by the reverse transcriptase-polymerase chain reaction using primers that corresponded to two conserved amino acid sequences found in 1-aminocyclopropane-1-carboxylate (ACC) synthases. The sequence of WSACS2 cDNA was almost identical to that of the cDNA for a wound-inducible ACC synthase (CM-ACS1) reported by N. Nakajima et␣al. 1990, Plant Cell Physiol 31: 1021–1029, with only one nucleotide difference. The expression of the WSACS2 transcript in tissue disks was examined by Northern blotting analysis. The accumulation of WSACS2 mRNA in the wounded tissue disks was inhibited by treatment with diphenyleneiodonium, which is known as an inhibitor of superoxide-generating NAD(P)H oxidases. The addition of xanthine and xanthine oxidase, which form a superoxide-generating system, strongly induced the accumulation of the WSACS2 transcript within 30 min and the level of the transcript fell thereafter. We measured levels of active oxygen species that were generated in the tissue disks by using chemiluminescence reagents and found that the chemiluminescence signal became apparent within a few seconds after wounding. The synthesis of the WSACS2 transcript was slightly inhibited by inhibitors of lipoxygenase, which is involved in the synthesis of jasmonic acid. Methyl jasmonate stimulated the synthesis of the WSACS2 transcript in the tissue disks.