Roles of DNA sequence and sigma A factor in transcription of the vraSR operon.
Cell wall damage in Staphylococcus aureus induces a rapid genome-wide response, referred to as the cell wall stress stimulon. This response is mediated by a two-component system, the vancomycin resistance-associated sensor/regulator (VraSR). The response regulator protein VraR is a transcription factor. Here, we demonstrate that two VraR binding sites in the vraSR operon control region are involved in the regulation of the vraSR operon. The sites are centered at the -60 and -35 nucleotide positions and are referred to as R1 and R2, respectively. DNase I footprinting and lux operon reporter vector studies showed that both of these sites communicate intimately with each other to fine-tune the activity of the vraSR operon. Mutagenesis of the VraR binding sites showed that dimerization of unphosphorylated VraR at R1 is driven by a hierarchy in VraR binding and by the proximity of the two tandem VraR binding sequences at this site. On the other hand, these studies show that the lack of sequence conservation and the distance between the VraR binding sequences in R2 ensure that VraR is recruited to this site only when phosphorylated (hence, under stress conditions). Furthermore, we demonstrate that sigma A (SigA) factor is involved in the regulation of the vraSR operon. Our study shows that sigma A factor does not bind to the vraSR operon control region in the absence of VraR, suggesting that VraR may interact directly with this factor.
Department of Biology, York University, Toronto, Ontario, Canada.
Antoaneta
A
Belcheva
Vidhu
V
Verma
Artyom
A
Korenevsky
Michael
M
Fridman
Krishan
K
Kumar
Dasantila
D
Golemi-Kotra
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D004269:DNA, Bacterial / Q000235:genetics* / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D003850:Deoxyribonuclease I; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D016680:Genome, Bacterial; D008163:Luminescent Measurements; D008969:Molecular Sequence Data; D009876:Operon / Q000235:genetics; D011485:Protein Binding; D023302:Protein Footprinting; D012808:Sigma Factor / Q000235:genetics / Q000378:metabolism*; D013211:Staphylococcus aureus / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D013312:Stress, Physiological; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D012808:Sigma Factor; C478170:VraR protein, Staphylococcus aureus; C478169:VraS protein, Staphylococcus aureus; D003850:Deoxyribonuclease I