Characterization of the SarA virulence gene regulator of Staphylococcus aureus

Staphylococcus aureus is a potent human pathogen that expresses a large number of virulence factors in a temporally regulated fashion. Two pleiotropically acting regulatory loci were identified in previous mutational studies. The agr locus comprises two operons that express a quorum-sensing system from the P2 promoter and a regulatory RNA molecule from the P3 promoter. The sar locus encodes a DNA-binding protein that activates the expression of both agr operons. We have cloned the sarA gene, expressed SarA in Escherichia coli and purified the recombinant protein to apparent homogeneity. The purified protein was found to be dimeric in the presence and absence of DNA and to consist mostly of alpha-helices. DNase I footprinting of SarA on the putative regulatory region cis to the agr promoters revealed three high-affinity binding sites composed of two half-sites each. Quantitative electrophoretic mobility shift assays (EMSAs) were used to derive equilibrium binding constants (KD) for the interaction of SarA with these binding sites. An unusual ladder banding pattern was observed in EMSA with a large DNA fragment including all three binding sites. Our data indicate that SarA regulation of the agr operons involves binding to multiple half-sites and may involve other sites located downstream of the promoters.     

Evidence that AphB, essential for the virulence of Vibrio vulnificus, is a global regulator

The Vibrio vulnificus aphB mutant was significantly less virulent than the wild type and was impaired in motility and adherence to host cells. Microarray analysis revealed that AphB of V. vulnificus (AphB(Vv)) influences the expression of over 10% of the V. vulnificus genome. The combined results indicated that AphB(Vv) is a global regulator contributing to the pathogenesis of V. vulnificus.     

Regulation of the intercellular adhesin locus regulator (icaR) by SarA, sigmaB, and IcaR in Staphylococcus aureus

The staphylococcal accessory regulator SarA and the alternative sigma factor σ^B have been previously identified as positive regulators, and IcaR as a negative regulator, of icaADBC expression. Here, we show that in Staphylococcus aureus SarA and σ^B are also required for icaR expression and that IcaR does not have a significant effect on its own expression.     

Autogenous transcriptional regulation of the regA gene, encoding an AraC-Like, essential virulence regulator in Citrobacter rodentium

We identified several promoters responsible for the expression of regA, which encodes a global virulence regulator in Citrobacter rodentium. Expression of some of the promoters was strongly autoactivated by RegA in conjunction with bicarbonate. Biochemical and mutational analyses were used to determine the consensus sequence of the RegA-binding sites.