Effects Of Nitric Oxide On Secondary Metabolite Biosynthesis Of Plant Cells And Its Molecular Basis Of Signaling Transduction

Production of secondary metabolites with clinical importance in plants by cell cultures has been one of the most extensively explored areas in recent years owing to the enormous commercial value of those compounds and the scarcity of the plants in the world. Application of plant cell culture for the production of usful secondary metabolites, however, is still limited due to the low yield of the desired compounds. The biosynthesis of secondary metabolites in plants is tightly regulated by internal and external factors. However, the molecular basis for the regulation of the secondary metabolite biosynthesis is largely unknown. Investigation of the signaling transduction chains (pathways) related to secondary metabolite biosynthesis is helpful for elucidating the mechanism of secondary metabolite biosynthesis in plant cells. Nitric oxide (NO) is a small, water and lipid soluble gas that in recent years has emerged as a major signalling molecule of ancient origin and ubiquitous importance. NO burst is one of the common reactions of plant cells to fungal elicitors. In this paper, the role of NO and its signaling transduction in elicitor-induced secondary metabolite biosynthesis of plant cells have been extensively investigated. The followings are the main results of the experiments.1) NO is essential for elilicitor-inducecd Taxol biosynthesis in Taxus chinensis suspension cell cultures. Elicitor prepared from the cell walls of Penicillium citrinum induces multiple responses of Taxus chinensis cells, including nitric oxide (NO) generation, sequentially followed by phenylalanine ammonia-lyase (PAL) activation and Taxol production. NO scavenger 2-4-carboxyphenyl-4,4,5,5-tetramethylimidazoline-l-oxyl-3-oxide (cPITO) and nitric oxide synthase (NOS) inhibitor S,S' -1,3-phenylene-bis-(1,2- ethanediyl)-bis-isothiourea (PBITU) suppress not only the elicitor-induced NO burst, but also the elicitor-induced PAL activation and Taxol production, which suggests that NO is necessary for elicitor-indced PAL activation and Taxol biosynthesis in Taxus chinensis suspension cells. External application of NO via its donor sodium nitroprusside (SNP) triggers PAL activation and Taxol biosyntesis of the cells in the absence of fungal elicitor, showing that NO is sufficient for inducing PAL activation and Taxol production. The results also show that elicitor-induced NO release of Taxus chinensis suspension cells is strongly inhibited by PBITU. Thus, our results demonstrate a causal relationship between NO generation.and PAL acitivation and Taxol production of Taxus chinensis cells, and indicate that NO, produced via NOS in Taxus chinensis cells treated with fungal elicitor, is an essential signaling molecule for elicitor-induced PAL activation and Taxol...