EXPRESSION AND LOCALIZATION OF HRPA1 - A PROTEIN OF XANTHOMONAS-CAMPESTRIS PV VESICATORIA ESSENTIAL FOR PATHOGENICITY AND INDUCTION OF THE HYPERSENSITIVE REACTION

The hrp cluster of the pepper and tomato pathogen Xanthomonas campestr is pv, vesicatoria is required for both pathogenicity on susceptible h ost plants and induction of the hypersensitive reaction on resistant p lants. The hrpA locus is located at the left end of the 25-kb hrp regi on and encodes a single 64-kDa Hrp protein, HrpA1, which belongs to th e PuID superfamily of proteins involved in type II and type III protei n secretion. In this study, we developed a defined medium without any plant-derived molecules that induces expression of hrpA in vitro. The hrpA transcription start site was mapped in the coding region of the h rpB8 gene, which is the last gene of the hrpB operon. The inducible hr pA promoter shows no homology to known promoter elements or other hip loci of X. campestris pv. vesicatoria. hrpA expression was shown to be independent of the hrp regulatory gene hrpX. The amino acid sequence of the HrpA1 protein is predicted to contain an N-terminal signal sequ ence and no further transmembrane domains and to be rich in beta-sheet stretches. Expression of HrpA1 in Escherichia coli cells causes induc tion of the psp operon like some of its counterparts, suggesting some commonality of function and that HrpA1 forms multimers. The protein pr oduct of hrpA1 was identified by using a specific polyclonal antibody. Cell fractionation studies demonstrated that the HrpA1 protein is loc alized in the outer membrane of X, campestris pv. vesicatoria. BrpA1 i s the first component of the Hrp secretion system whose localization h as been determined in the original organism.