Virulence gene expression in covR/vicK mutants of Streptococcus mutans

Background: Expression of genes encoding glucosyltransferases  B/C (gtfB,gtfC) and glucan-binding protein B (gbpB) by S. mutans (SM) strains is associated with their distinctive capacity to accumulate into biofilms. Genes encoding these virulence proteins appear to be controlled by global two-component-transduction-systems (TCS), Vic and Cov. Little is known, however, whether these systems work similarly among different strain backgrounds.

Objective: The aim of this study was to compare the effect of inactivation of the vicK and covR genes in the expression of gtfB/C and gbpB among different SM strains.

Methods: Nonpolar deletion mutations of vicK or covR were performed by PCR ligation mutagenesis in three SM strains (UA159, 20A3, 5ST1). covR and vicK mutants (MT) and the respective wild type (WT) strains were grown in BHI (37°C;10%CO₂;5h) to absorbances (A_550nm) of 0.3; cells and culture fluids were then collected. RNAs extracted from cell samples were analyzed in semi-quantitative-RT-PCR assays. Amounts of cell-associated and secreted GbpB were also analyzed in western blot assays probed with specific GbpB anti-serum.

Results: Inactivation of covR resulted in a 20 to 30% increase in levels of gtfB and gtfC transcripts (p<0.05,ANOVA) in all the strains analyzed, while the vicK deletion resulted in 10 to 20% decrease in the transcription of gtfB, gtfC and gbpB (p<0.05, ANOVA). Forty to 60% reduction in levels of GbpB were observed among the three vicK- mutants in comparison with the respective WT strains (p<0.05,ANOVA). covR mutants also produced 60 to 250% more GbpB than WT strains (p<0.01, ANOVA).

Conclusions: Data indicate that Vic/Cov systems play roles in the expression of virulence genes among the strains analyzed, in that gtfB, gtfC and gbpB are under the negative control of covR, while the same genes are positively regulated by the vicK TCS. Supported by FAPESP 02/07156-1,06/55933-8 and NIH-Fogarty TW-06324.