67. イネ培養細胞におけるファイトアレキシン生合成に関与するジテルペン環化酵素遺伝子 OsDTC2 の cDNA クローニングと機能解析

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In suspension-cultured rice cells, diterpenoid phytoalexins such as momilactones, (-)-phytocassanes, and oryzalexins are produced in responce to exogenously applied elicitors such as chitin oligomers and cerebrosides. Previously we reported that at least two species of mRNA encoding putative diterpene cyclases, OsDTC1 and OsDTC2,were expressed in suspension-cultured rice cells treated with a chitin elicitor, N-acetylchitoheptaose, and that OsDTC1 functioned as ent-cassa-12,15-diene synthase, a diterpene cyclase involved in biosynthesis of (-)-phytocassanes. We report here cloning and functional analysis of OsDTC2 cDNA in rice (Oryza sativa L. cv. Nipponbare). A cDNA encoding the full-length ORF of OsDTC2 was cloned by PCR using primers designed based on information from a rice genome database (RiceBlast.dna.affrc.go.jp) and a cDNA library from elicitor-treated rice cells as a template. The OsDTC2 cDNA was overexpressed in Escherichia coli as a fusion protein with glutathione S-transferase. In the presence of OsCycl that catalyzes the conversion of geranylgeranyl diphosphate (GGDP) into syn-copalyl diphosphate (syn-CDP), the recombinant OsDTC2 converted GGDP into stemar-13-ene, a putative diterpene hydrocarbon precursor of oryzalexin S. It was thus indicated that OsDTC2 functioned as stmar-13-ene synthase. The levels of OsDTC2 mRNA in suspension-cultured rice cells began to increase from 3h after addition of the elicitor, and the level reached the maximum after 8h, and gradually decreased. It was also shown that the endogenous levels of stemar-13-ene increased 8h after addition of the elicitor in suspension-cultured rice cells.
植物化学調節学会の論文
2003-10-10

67. イネ培養細胞におけるファイトアレキシン生合成に関与するジテルペン環化酵素遺伝子 OsDTC2 の cDNA クローニングと機能解析

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