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Journal of Bacteriology, January 2000, p. 14-22, Vol. 182, No. 1
0021-9193/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Role of HrcA and CIRCE in the Heat Shock Regulatory Network of Bradyrhizobium japonicum

Alexander C. Minder, Hans-Martin Fischer, Hauke Hennecke, and Franz Narberhaus*

Institut für Mikrobiologie, Eidgenössische Technische Hochschule, CH-8092 Zürich, Switzerland

Received 11 August 1999/Accepted 6 October 1999

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 using a complementation approach that screened for GrpE function. In vivo and in vitro transcript mapping demonstrated that both genes are transcribed separately from RpoH (sigma 32)-dependent promoters. To investigate the supposed negative regulatory function of HrcA, we compared the expression of putative target genes in the wild type with that in an hrcA mutant. Transcription of the CIRCE-associated chaperonin operons groESL4 and groESL5, as well as the beta -galactosidase activity derived from corresponding groE-lacZ fusions, was strongly elevated in the hrcA mutant even at physiological temperatures. Expression of other heat shock regulons (RpoH or ROSE dependent) was not affected. To study the activity of HrcA in vitro, we purified a histidine-tagged version of the protein under nondenaturing conditions. Specific binding to the CIRCE element was obtained with a soluble fraction of HrcA in gel retardation experiments.

* Corresponding author. Mailing address: Institut für Mikrobiologie, ETH-Zentrum, Schmelzbergstrasse 7, CH-8092 Zürich, Switzerland. Phone: 41-1-632-2586. Fax: 41-1-632-1148. E-mail: fnarber{at}micro.biol.ethz.ch.

Journal of Bacteriology, January 2000, p. 14-22, Vol. 182, No. 1
0021-9193/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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