Regulation of the dnaK operon of Streptomyces coelicolor A3(2) is governed by HspR, an autoregulatory repressor protein

Abstract

The dnaK operon of Streptomyces coelicolor contains four genes (5'-dnaK-grpE-dnaJ-hspR). The fourth gene encodes a novel heat shock protein, HspR, which appears so far to be unique to the high-G+C actinomycete group of bacteria. HspR binds with high specificity to three inverted repeat sequences in the promoter region of the S. coelicolor dnaK operon, strongly suggesting a direct role for HspR in heat shock gene regulation. Here we present genetic and biochemical evidence that HspR is the repressor of the dnaK operon. Disruption of hspR leads to high-level constitutive transcription of the dnaK operon. Parallel transcriptional analyses of groESL1 and groEL2 expression demonstrated that heat shock regulation of the groE genes was essentially unaffected in an hspR null mutant, although the basal (uninduced) level of groEL2 transcription was slightly elevated compared with the wild type. The results of HspR titration experiments, where the dnaK operon promoter region was cloned at ca. 50 copies per chromosome, were consistent with the prediction that HspR functions as a negative autoregulator. His-tagged HspR, overproduced and purified from Escherichia coli, was shown to repress transcription from the dnaK operon promoter in vitro, providing additional evidence for the proposal that HspR directly regulates transcription of the dnaK operon. These studies indicate that there are at least two transcriptional mechanisms for controlling heat shock genes in S. coelicolor--one controlling the dnaK operon and another controlling the groE genes.