Pseudomonas syringae pathovar tomato DC3000 produces the phytotoxin coronatine, a major determinant of the leaf chlorosis associated with DC3000 pathogenesis. The DC3000 PSPTO4723 (cmaL) gene is located in a genomic region encoding type III effectors, however it promotes chlorosis in the model plant Nicotiana benthamiana in a manner independent of type III secretion. Coronatine is produced by ligation of two moieties, coronafacic acid (CFA) and coronamic acid (CMA), which are produced by biosynthetic pathways encoded in separate operons. Cross-feeding experiments, performed in N. benthamiana with cfa, cma, and cmaL mutants, implicate CmaL in CMA production. Furthermore, analysis of bacterial supernatants under coronatine-inducing conditions revealed that mutants lacking either the cma operon or cmaL accumulate CFA rather than coronatine, supporting a role for CmaL in the regulation or biosynthesis of CMA. CmaL does not appear to regulate CMA production, as cmaL mutants are unaltered in the expression of proteins with known roles in CMA production. Rather, CmaL is needed for the first step in CMA synthesis, as wild-type levels of coronatine production are restored to a ΔcmaL mutant when supplemented with 50 μg/ml L-allo-isoleucine, the starting unit for CMA production. cmaL is found in all other sequenced P. syringae strains with coronatine biosynthesis genes. This characterization of CmaL identifies a critical missing factor in coronatine production and provides a foundation for further investigation of a member of the widespread DUF1330 protein family.
Jay N. Worley, Alistair B. Russell, Aaron G. Wexler, Philip A. Bronstein, Brian H. Kvitko, Stuart B. Krasnoff, Kathy R. Munkvold, Bryan Swingle, Donna M. Gibson, and Alan Collmer