Control of Enzyme II(scr) and Sucrose-6-Phosphate Hydrolase Activities in Streptococcus mutans by Transcriptional Repressor ScrR Binding to the cis-Active Determinants of the scr Regulon

In Streptococcus mutans, enzyme II(scr) and sucrose-6-phosphate hydrolase are two important enzymes in the transport and metabolism of dietary sucrose. The scr regulon of S. mutans is composed of three genes, scrA and scrB, which code for enzyme II(scr) and sucrose-6-phosphate hydrolase, respectively, and scrR, which codes for a GalR-LacI-type transcription regulator. It was previously shown that expression of both scrA and scrB is similarly induced by sucrose. Mutation in the scrR gene resulted in increased expression of scrB relative to that in the wild-type strain. In this study, we employed DNA mobility shift and DNase I protection assays with a purified ScrR-histidine tag fusion protein to examine the DNA binding properties of ScrR to the promoter regions of the scrA and scrB genes. The results showed that ScrR bound specifically to the promoter regions of both scrA and scrB. Two regions with high affinity for ScrR in the promoter sequences of the scrA and scrB genes were identified by DNase I protection assays. One, O(C), which includes a 20-bp imperfect inverted-repeat sequence, is located between the two promoters, and the other, O(B), is located within the scrB promoter region containing a 37-bp imperfect direct-repeat sequence. Mutations of O(B) and O(C) resulted in constitutive transcription and expression of both the scrA and scrB genes. Our results indicated that S. mutans coordinates the activities of enzyme II(scr) and sucrose-6-phosphate hydrolase by transcriptional repressor ScrR binding to the promoter regions of the scr regulon

Genetic Analysis of a Unique Bacteriocin, Smb, Produced by Streptococcus mutans GS5

A dipeptide lantibiotic, named Smb, in Streptococcus mutans GS5 was characterized by molecular genetic approaches. The Smb biosynthesis gene locus is encoded by a 9.5-kb region of chromosomal DNA and consists of seven genes in the order smbM1, -T, -F, -M2, -G, -A, -B. This operon is not present in some other strains of S. mutans, including strain UA159. The genes encoding Smb were identified as smbA and smbB. Inactivation of smbM1, smbA, or smbB attenuated the inhibition of the growth of the indicator strain RP66, confirming an essential role for these genes in Smb expression. Mature Smb likely consists of the 30-amino-acid SmbA together with the 32-amino-acid SmbB. SmbA exhibited similarity with the mature lantibiotic lacticinA2 from Lactococcus lactis, while SmbB was similar to the mersacidin-like peptides from Bacillus halodurans and L. lactis. We also demonstrated that Smb expression is induced by the competence-stimulating peptide (CSP) and that a com box-like sequence is located in the smb promoter region. These results suggest that Smb belongs to the class I bacteriocin family, and its expression is dependent on CSP-induced quorum sensing

Multiple Streptococcus mutans Genes Are Involved in Biofilm Formation

Streptococcus mutans has been strongly implicated as the principal etiological agent in dental caries. One of the important virulence properties of these organisms is their ability to form biofilms known as dental plaque on tooth surfaces. Since the roles of sucrose and glucosyltransferases in S. mutans biofilm formation have been well documented, we focused our attention on sucrose-independent factors. We have initially identified several mutants that appear to be defective in biofilm formation on abiotic surfaces by an insertional inactivation mutagenesis strategy applied to S. mutans. A total of 27 biofilm-defective mutants were isolated and analyzed in this study. From these mutants, three genes were identified. One of the mutants was defective in the Bacillus subtilis lytR homologue. Another of the biofilm-defective mutants isolated was a yulF homologue, which encodes a hypothetical protein of B. subtilis whose function in biofilm formation is unknown. The vast majority of the mutants were defective in the comB gene required for competence. We therefore have constructed and examined comACDE null mutants. These mutants were also found to be attenuated in biofilm formation. Biofilm formation by several other regulatory gene mutants were also characterized using an in vitro biofilm-forming assay. These results suggest that competence genes as well as the sgp and dgk genes may play important roles in S. mutans biofilm formation