The CovR response regulator of group A streptococcus (GAS) acts directly to repress its own promoter.
The CovR/S (CsrR/S) two component system is a global regulator of virulence gene expression in the group A streptococcus (GAS, Streptococcus pyogenes). The response regulator, CovR, regulates about 15% of the genes of GAS, including its own operon. Using in vitro DNA binding assays with purified CovR protein, we found that CovR binds a DNA fragment including the covR promoter (Pcov). DNaseI footprint analyses showed that phosphorylation of CovR enhanced and extended the protected regions. The proposed CovR consensus binding sequence (ATTARA) was present at most, but not all protected regions. The effect of replacing the two thymine residues in the consensus binding sequence (CB) with guanine residues was evaluated both in vitro and in vivo. Most, but not all, CB mutations reduced binding of CovR in vitro. Using a transcriptional reporter introduced in single copy into the GAS chromosome, we found that mutations at each CB completely or partially relieved CovR-mediated repression in vivo. This suggests that CovR regulation of Pcov is direct. Further support for this conclusion comes from use of an in vitro GAS transcription system in which CovR was sufficient to mediate repression of Pcov. This repression was enhanced by phosphorylation of the protein. In addition, we found that the CovR binding region overlapping the promoter was essential for wild type repression of Pcov both in vitro and in vivo, suggesting that promoter occlusion is a primary mechanism of Pcov repression by CovR.
Gusa
Asiya A
AA
Department of Microbiology and Immunology, Emory University School of Medicine, Atlanta, GA 30322, USA.
Scott
June R
JR
D019677:Artificial Gene Fusion; D001426:Bacterial Proteins / Q000235:genetics* / Q000502:physiology*; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D016384:Consensus Sequence; D004247:DNA / Q000378:metabolism; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D005966:Glucuronidase / Q000032:analysis / Q000235:genetics; D008969:Molecular Sequence Data; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D011485:Protein Binding; D012097:Repressor Proteins / Q000235:genetics* / Q000502:physiology*; D013297:Streptococcus pyogenes / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; C432143:CsrR protein, Streptococcus pyogenes; D004268:DNA-Binding Proteins; D012097:Repressor Proteins; D004247:DNA; D005966:Glucuronidase