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item Keith, Lisa

Submitted to: Plasmid Journal
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 9/28/2002
Publication Date: 9/28/2003
Citation: Plasmid 49, 2003, pgs. 106 - 117.

Interpretive Summary: Pseudomonas syringae pv. glycinea, a plant pathogenic bacterium, causes blight of soybeans. One of the symptoms is yellowing, or chlorosis, of the leaves which is caused by the toxin coronatine. The bacterial genes involved in the production of the toxin are located on a 90-kb plasmid named p4180A. The genes involved in the global regulation for coronatine production are unknown. In our study we evaluated the role of rpoN, an alternate sigma factor, in the abiliby of P. syringae pv. glycinea PG 4180 to cause disease. A mutant of PG 4180 which no longer contained rpoN could not grow in minimal media unless the media was supplemented with glutamate, glutamine or aspartate. The mutant bacteria could not grow or cause disease on soybeans and did not produce the toxin coronatine. Additional genes were affected by the rpoN mutation, including hrpL which is involved in secretion. Upon the addition of a copy of the rpoN gene, the PG4180 mutant could grow on minimal medium and produce the toxin, indicating that rpoN is required for growth and production of coronatine.

Technical Abstract: The plant pathogen Pseudomonas syringae pv. glycinea PG4180 produces coronatine (COR), a phytotoxin which functions as a virulence factor in bacterial blight of soybeans. The COR biosynthetic gene cluster in PG4180 is borne on a 90-kb plasmid named p4180A. Although pathway-specific regulatory genes for COR have been identified, global regulatory genes for COR production in PG4180 remain undefined. In the present study, we evaluated the role of rpoN, which encodes rpoN, in the virulence of strain PG4180. A rpoN mutant of PG4180, designated PG4180.K2, was unable to grow in M9 minimal medium; however, the addition of exogenous glutamate, glutamine or aspartate to M9 medium enabled PG4180.K2 to grow in vitro. PG4180.K2 could not induce disease symptoms or multiply in soybean plants and was defective in COR production and cor gene expression. Furthermore, PG4180.K2 was impaired in transcription of hrpL, an alternate sigma factor that mediates expression of genes in the type III secretion system of P. syringae. PG4180.K2 transconjugants with a wild-type copy of rpoN were complemented for hrpL and cor gene expression, COR biosynthesis, and growth in vitro. Our results indicate that rpoN is required for growth and the expression of both chromosomal and plasmid-encoded virulence factors in P. syringae pv. glycinea PG4180.