Biosynthesis of Bacterially Induced Sesquiterpenoid Phytoalexins in Cotton: The project goal is elucidation of the biosynthetic pathway from farnesyl pyrophosphate to the phytoalexin 2,7-dihydroxycadalene in cotton. Project objectives are: 1) identification of intermediate metabolites of the biosynthetic pathway, and 2) isolation and purification of the sesquiterpene cyclase involved. A cotton line lacking the potentially interfering pigment glands will be used. Direct MS/MS analysis of powdered, inoculated tissue, as well as GC/MS/MS of tissue extracts will be used to search for resistance-associated sesquiterpenoids. Putative intermediates will be purified for structure elucidation. An assay will be developed for the sesquiterpene cyclase, based on conversion of isotopically labeled substrate to a cyclic intermediate. Purification of the enzyme will be undertaken. 2,7-Dihydroxycadalene and three structurally related compounds are produced during expression of resistance to the bacterial pathogen Xanthomonas campestris pv. malvacearum and accumulate to inhibitory concentrations in fluorescent, necrotic leaf cells adjacent to the bacterial colonies. They thus appear to play an important role in race-specific resistance of cotton to bacterial blight. There is genetic evidence that this resistance response results from a recognition event determined by a host resistance gene and a corresponding bacterial avirulence gene. Identification of the enzymes that are regulated will facilitate the study of how the recognition event regulates phytoalexin production. The information gained should identify traits that have to be bred or engineered into cotton to make it more resistant to infection.
