Regulation of gbpC Expression in Streptococcus mutans

Abstract

Streptococcus mutans, the principal causative agent of dental caries, produces four glucan-binding proteins (Gbp) that play major roles in bacterial adherence and pathogenesis. One of these proteins, GbpC, is an important cell surface protein involved in biofilm formation. GbpC is also important for cariogenesis, bacteremia, and infective endocarditis. In this study, we examined the regulation of gbpC expression in S. mutans strain UA159. We found that gbpC expression attains the maximum level at mid-exponential growth phase, and the half-life of the transcript is less than 2 min. Expression from PgbpC was measured using a PgbpC-gusA transcriptional fusion reporter and was analyzed under various stress conditions, including thermal, osmotic, and acid stresses. Expression of gbpC is induced under conditions of thermal stress but is repressed during growth at low pH, whereas osmotic stress had no effect on expression from PgbpC. The results from the expression analyses were further confirmed using semiquantitative reverse transcription-PCR analysis. Our results also reveal that CovR, a global response regulator in many Streptococcus spp., represses gbpC expression at the transcriptional level. We demonstrated that purified CovR protein binds directly to the promoter region of PgbpC to repress gbpC expression. Using a DNase I protection assay, we showed that CovR binds to DNA sequences surrounding PgbpC from bases ؊68 to 28 (where base 1 is the start of transcription). In summary, our results indicate that various stress conditions modulate the expression of gbpC and that CovR negatively regulates the expression of the gbpC gene by directly binding to the promoter region.

Two-component signal transduction systems in oral bacteria

“…It was proposed that CovR Sm -mediated regulation is dependent on the amounts of CovR Sm present in cells regardless of the phosphorylation state; i.e. CovR Sm would be always active and/or likely phosphorylated by small phosphodonors such as AcP [141,172]. The absence of CovS protein in S. mutans and amino acid differences between CovR Spy and CovR Smu [142] (Figure 4) account for this proposal.…”

Modulation ofcovRExpression inStreptococcus mutansUA159

“…CovR is essential for the biofilm formation and pathogenesis of S. mutans, as the transcription of genes encoding various virulence factors, such as glucosyltransferase and glucan-binding protein C, is essential for the adherence of this pathogen to the tooth surface, leading to the colonization of the oral cavity as well as the onset of cariogenicity and tooth decay (3,6,28). Although CovR of S. mutans shares significant homology with CovR of GAS (84% similarity and 75% identity [1]), CovR in S. mutans functions primarily as a transcriptional activator (Biswas, unpublished), whereas CovR in GAS is predominantly a transcriptional repressor (20).…”

“…The cultures were harvested via centrifugation, resuspended and incubated in RNA protect reagent (Qiagen) at room temperature, and then recentrifuged to form a pellet which was either stored overnight at Ϫ20°C or processed immediately for RNA extraction. RNA extraction was performed as described by Biswas et al (3).…”

“…The regulation of covR expression in S. mutans has not been fully elucidated to date (3,28). In this communication, we report the effects of growth phase and various environmental stimulants on the regulation of the expression of covR.…”

“…mutans lacks covS but does express CovR, which also plays key roles related to biofilm formation and virulence by regulating the transcription of genes such as glucosyltransferase genes (gtfB/C) and glucan-binding protein C genes (gbpC); the expression of these genes is essential for adherence to the tooth surface and for cariogenicity (3,6,31,39).…”