Identification of a Helix-Turn-Helix Motif of
            <i>Bacillus thermoglucosidasius</i>
            HrcA Essential for Binding to the CIRCE Element and Thermostability of the HrcA-CIRCE Complex, Indicating a Role as a Thermosensor

Hitomi, Masafumi; Nishimura, Hiroshi; Tsujimoto, Yasuhiro; Matsui, Hiroshi; Watanabe, Kunihiko

doi:10.1128/jb.185.1.381-385.2003

Cited by 24 publications

(25 citation statements)

References 33 publications

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“…Thus, an increased temperature alone is not sufficient for the chlamydial heat shock response, suggesting that additional factors that are not present in our in vitro system play a role. Studies of the thermostability of HrcA-CIRCE complexes in other bacteria have shown that HrcA dissociates from CIRCE at temperatures above physiologic temperatures (24,55), although this effect varies with the bacterial species (14) and the presence of additional factors (24,38). C. trachomatis is an obligate intracellular parasite and, as such, is exposed to a narrower range of growth temperatures than the free-living, gram-positive bacteria in which much of HrcA activity has been characterized previously.…”

Section: Discussionmentioning

confidence: 99%

Stress Response Gene Regulation in Chlamydia Is Dependent on HrcA-CIRCE Interactions

Wilson

Tan

2004

J Bacteriol

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HrcA is a transcriptional repressor that regulates stress response genes in many bacteria by binding to the CIRCE operator. We have previously shown that HrcA regulates the promoter for the dnaK heat shock operon in Chlamydia. Here we demonstrate that HrcA represses a second heat shock promoter that controls the expression of groES and groEL, two other major chlamydial heat shock genes. The CIRCE element of C. trachomatis groEL is the most divergent of known bacterial CIRCE elements, and HrcA had a decreased ability to bind to this nonconsensus operator and repress transcription. We demonstrate that the CIRCE element is necessary and sufficient for transcriptional regulation by chlamydial HrcA and that the inverted repeats of CIRCE are the binding sites for HrcA. Addition of a CIRCE element upstream of a non-heat-shock promoter allowed this promoter to be repressed by HrcA, showing in principle that a chlamydial promoter can be genetically modified to be inducible. These results demonstrate that HrcA is the regulator of the major chlamydial heat shock operons, and we infer that the mechanism of the heat shock response in Chlamydia is derepression. However, derepression is likely to involve more than a direct effect of increased temperature as we found that HrcA binding to CIRCE and HrcA-mediated repression were not altered at temperatures that induce the heat shock response.

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Section: Discussionmentioning

confidence: 99%

Stress Response Gene Regulation in Chlamydia Is Dependent on HrcA-CIRCE Interactions

Wilson

Tan

2004

J Bacteriol

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“…Lund and I. Barber, unpublished data). HrcA instability at elevated temperatures leads to dissociation of HrcA from the CIRCE element which causes increased expression of heat-shock genes (Watanabe et al 2001;Hitomi et al, 2003). A feedback loop involving GroES and GroEL acts to fold HrcA to its active conformation under non-heat-shock conditions (Mogk et al, 1997;Reischl et al, 2002;Roncarati et al, 2007).…”

Section: Introductionmentioning

confidence: 99%

The hrcA and hspR regulons of Campylobacter jejuni

2010

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The human pathogen Campylobacter jejuni has a classic heat shock response, showing induction of chaperones and proteases plus several unidentified proteins in response to a small increase in growth temperature. The genome contains two homologues to known heat shock response regulators, HrcA and HspR. Previous work has shown that HspR controls several heat-shock genes, but the hrcA regulon has not been defined. We have constructed single and double deletions of C. jejuni hrcA and hspR and analysed gene expression using microarrays. Only a small number of genes are controlled by these two regulators, and the two regulons overlap. Strains mutated in hspR, but not those mutated in hrcA, showed enhanced thermotolerance. Some genes previously identified as being downregulated in a strain lacking hspR showed no change in expression in our experiments.

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“…4A) constitutes the recognition helix of the HrcA proteins. The same region was recently proposed as the recognition helix of HrcA for Bacillus thermoglucosidasius and B. subtilis (19,36). Figure 4A also shows the putative stabilization helix of the HTH motif (amino acids 42 to 48 in C. crescentus) in the various HrcA proteins.…”

Section: Resultsmentioning

confidence: 93%

Functional and Structural Analysis of HrcA Repressor Protein from Caulobacter crescentus

et al. 2004

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A large number of bacteria regulate chaperone gene expression during heat shock by the HrcA-CIRCE system, in which the DNA element called CIRCE serves as binding site for the repressor protein HrcA under nonstress conditions. In Caulobacter crescentus, the groESL operon presents a dual type of control. Heat shock induction is controlled by a 32 -dependent promoter and the HrcA-CIRCE system plays a role in regulation of groESL expression under physiological temperatures. To study the activity of HrcA in vitro, we purified a histidine-tagged version of the protein, and specific binding to the CIRCE element was obtained by gel shift assays. The amount of retarded DNA increased significantly in the presence of GroES/GroEL, suggesting that the GroE chaperonin machine modulates HrcA activity. Further evidence of this modulation was obtained using lacZ transcription fusions with the groESL regulatory region in C. crescentus cells, producing different amounts of GroES/GroEL. In addition, we identified the putative DNA-binding domain of HrcA through extensive protein sequence comparison and constructed various HrcA mutant proteins containing single amino acid substitutions in or near this region. In vitro and in vivo experiments with these mutated proteins indicated several amino acids important for repressor activity.Caulobacter crescentus, a member of the ␣-subdivision of proteobacteria, responds to heat shock by elevating the levels of synthesis of over 20 polypeptides (16). Several heat shockinducible genes in C. crescentus have been characterized, including dnaKJ (15), groESL (1), lon (37), hrcA/grpE (30), and ftsH (11), and they all are positively regulated by the alternative sigma factor of RNA polymerase 32 . The rpoH gene encoding 32 in C. crescentus has also been characterized, and one of its promoters was shown to be 32 dependent, indicating an autogenous control of rpoH transcription (28, 38). Furthermore, the levels of 32 were shown to increase transiently during heat shock, and the increased transcription of rpoH seems to account for the induction of 32 levels (28, 38). Recent studies have shown that, as described for Escherichia coli (for a review, see reference 39), the heat shock protein (HSP) DnaK is a negative modulator of the heat shock response in C. crescentus, which acts by inhibiting 32 activity and stimulating its degradation (7). However, despite the strong effect of DnaK levels on the induction phase of the response, downregulation of HSP synthesis is not affected by changes in the amount of this chaperone. Competition between 32 and 73 , the major sigma factor in C. crescentus which was shown to be heat shock inducible, has been proposed as the most important factor controlling the shutoff of HSP synthesis during the recovery phase (7). Another important negative modulator of the heat shock response, the repressor protein HrcA in C. crescentus, has also been described (30). This protein, which is absent in E. coli, has been found to occur in a growing number of diverse bacterial species and pr...

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Identification of a Helix-Turn-Helix Motif of Bacillus thermoglucosidasius HrcA Essential for Binding to the CIRCE Element and Thermostability of the HrcA-CIRCE Complex, Indicating a Role as a Thermosensor

Cited by 24 publications

References 33 publications

Stress Response Gene Regulation in Chlamydia Is Dependent on HrcA-CIRCE Interactions

Stress Response Gene Regulation in Chlamydia Is Dependent on HrcA-CIRCE Interactions

The hrcA and hspR regulons of Campylobacter jejuni

Functional and Structural Analysis of HrcA Repressor Protein from Caulobacter crescentus

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