Role of HrcA and CIRCE in the heat shock regulatory network of Bradyrhizobium japonicum

A large number of bacteria regulate chaperone gene expression by the CIRCE- HrcA system in which a DNA element called CIRCE serves as binding site for the repressor protein HrcA under non-heat-shock conditions. We have cloned the two consecutive genes hrcA and grpE of Bradyrhizobium japonicum by usin g a complementation approach that screened for GrpE function. In vivo and i n vitro transcript mapping demonstrated that both genes are transcribed sep arately from RpoH (sigma(32))-dependent promoters. To investigate the suppo sed negative regulatory function of HrcA, we compared the expression of put ative target genes in the wild type with that in an hrcA mutant. Transcript ion of the CIRCE-associated chaperonin operons groESL(4) and groESL(5), as well as the beta-galactosidase activity derived from corresponding groE-lac Z fusions, was strongly elevated in the hrcA mutant even at physiological t emperatures. Expression of other heat shock regulons (RpoH or ROSE dependen t) was not affected. To study the activity of HrcA in vitro, we purified a histidine-tagged version of the protein under nondenaturing conditions. Spe cific binding to the CIRCE element was obtained with a soluble fraction of HrcA in gel retardation experiments.