Modulation ofcovRExpression inStreptococcus mutansUA159

Abstract

The biofilm-forming Streptococcus mutans is a gram-positive bacterium that resides in the human oral cavity and is considered to be the primary etiological agent in the formation of dental caries. The global response regulator CovR, which lacks a cognate sensor kinase, is essential for the pathogenesis and biofilm formation of this bacterium, but it is not clear how covR expression is regulated in S. mutans. In this communication, we present the results of our studies examining various factors that regulate the expression of covR in S. mutans UA159. The results of Southern hybridization and PCR analysis indicated that CovR is an orphan response regulator in various isolates of S. mutans. The transcriptional start site for covR was found to be 221 base pairs upstream of the ATG start codon, and site-directed mutagenesis of the upstream TATAAT box confirmed our findings. The expression of covR is growth phase dependent, with maximal expression observed during exponential-growth phase. While changes to the growth temperature did not significantly affect the expression of covR, increasing the pH or the concentration of Mg 2؉ in the growth medium leads to an increase in covR expression. The results of semiquantitative reverse transcriptase PCR analysis and in vivo transcriptionalfusion reporter assays indicated that CovR autoregulates its own expression; this was verified by the results of electrophoretic mobility shift assays and DNase I protection assays, which demonstrated direct binding of CovR to the promoter region. Apparently, regulation by Mg 2؉ and the autoregulation of covR are not linked. A detailed analysis of the regulation of CovR may lead to a better understanding of the pathogenesis of S. mutans, as well as providing further insight into the prevention of dental caries.

CsrRS Regulates Group BStreptococcusVirulence Gene Expression in Response to Environmental pH: a New Perspective on Vaccine Development

“…We observed no change in the transcript level of CsrRS at an acidic pH, suggesting that pH-dependent regulation is not a function of changes in the abundance of the CsrRS proteins. However, it has been reported recently that, at least for S. mutans, the increase in pH in the growth medium led to an increase in csrR expression (9). An intriguing possibility suggested by our results is that the increase in proton concentration at an acidic pH signals through the CsrS sensor protein to stimulate CsrS autokinase activity, with subsequent downstream phosphorylation of CsrR, thereby increasing the affinity of CsrR for regulated promoters (28).…”

CovR Alleviates Transcriptional Silencing by a Nucleoid-Associated Histone-Like Protein in Streptococcus mutans

“…In total, by DNase I protection assays we have characterized the binding of CovR to promoters of five genes: gtfB, gtfC, gbpC, covR, and SMU.1882 (5,9,19,20). Furthermore, we have also shown CovR binding to 15 additional promoters by EMSA (25).…”

“…S. mutans encodes at least 14 TCSs that play important roles in bacterial adaptation, bacteriocin production, and biofilm formation (1,6). Of these, CovRS is one of the most important and widely studied TCSs, in S. mutans (9,20), as well as in other streptococci (21, 31, 33-35, 42, 45). The CovRS system was first characterized in group A streptococcus (GAS) (30,39,46), where it controls about 15% of the genes, either directly or indirectly (33,34).…”

“…CovR represses half of these genes, including gtfB (glucosyltransferase B), gtfC (glucosyltransferase C), and gbpC (glucan-binding protein C). CovR directly regulates these genes by binding to the promoter region (5,9,20). Among the genes that are activated by CovR, most of them are either uncharacterized or very poorly characterized.…”

Core-Gene-Encoded Peptide Regulating Virulence-Associated Traits in Streptococcus mutans

“…Bacteria in oral biofilms must cope with these changes, and adaptations to fluctuations in environmental conditions are often accomplished by two-component systems (TCSs), which consist of a histidine kinase (HK) and response regulator (RR) that regulate gene expression as a function of a wide variety of inputs (10). In S. mutans UA159, there are 14 putative TCS pairs plus GcrR (also annotated as CovR or TarC), a so-called orphan response regulator that is not encoded in an operon with a cognate histidine kinase (11)(12)(13)(14)(15), and many of these TCSs have been demonstrated to affect the expression of virulence-related traits of this organism. For example, the ComDE TCS consists of the membrane-bound HK ComD and the RR ComE and has a profound impact on the development of competence, biofilm formation, and bacteriocin production (16)(17)(18)(19).…”