Elucidation of the Regulon and <i>cis</i> -Acting Regulatory Element of HrpB, the AraC-Type Regulator of a Plant Pathogen-Like Type III Secretion System in Burkholderia pseudomallei

Abstract

The human pathogen Burkholderia pseudomallei possesses multiple type III secretion system (T3SS) gene clusters. One of these, the B. pseudomallei T3SS2 (T3SS2(bp)) gene cluster, which apparently plays no role in animal virulence, is also found in six additional Burkholderia spp. and is very similar to T3SSs found in phytopathogenic Xanthomonas spp. and Ralstonia solanacearum. The T3SS2(bp) gene cluster also encodes an AraC-type regulatory protein (HrpB(bp)) that is an ortholog of HrpB, the master regulator of the R. solanacearum T3SS (T3SS(rso)) and its secreted effectors. Transcriptome analysis showed that HrpB(bp) activates the expression of T3SS2(bp) genes, as well as their orthologs in R. solanacearum. In addition to activating T3SS2(bp), HrpB(bp) also upregulates the expression of ~30 additional B. pseudomallei genes, including some that may confer production of adhesive pili, a polyketide toxin, several putative T3SS2(bp)-secreted effectors, and components of a regulatory cascade. T3SS2(bp) promoter regions were found to contain a conserved DNA motif (p2(bp) box) identical in sequence and position to the hrp(II) box required for HrpB-dependent T3SS(rso) transcription activation. The p2(bp) box is also present in the promoter regions of the essentially identical T3SS found in the very closely related species Burkholderia thailandensis (T3SS2(bt)). Analysis of p2(bp) box mutants showed that it is essential for HrpB(bp)-mediated transcription activation in both species. Although it has been suggested that T3SS2(bp) and T3SS2(bt) may function in phytopathogenicity, we were unable to demonstrate a phytopathogenic phenotype for B. thailandensis in three different plant hosts.