The Role of Carbohydrate Availability, pH, and Carbon Catabolite Control Protein A (CcpA) in the Regulation of Exopolysaccharide Production by Streptococcus mutans

Streptococcus mutans, the leading etiological agent of dental caries, is capable of producing a variety of exopolysaccharides (EPS) from sucrose, including glucans via glucosyltransferases (Gtf B, C, and D) and fructans via fructosyltransferase (Ftf). Expression of both gtfBC and ftf has been shown to be constitutive, and increased by sucrose. In Bacillus subtilis and other Gram positive bacteria, CcpA is key in controlling carbohydrate metabolism. Objectives: The purpose of this study was to examine the role carbohydrate availability, pH, and CcpA in the regulation of EPS synthesis by S. mutans. Methods: Two recombinant strains that carry fusions of a cat gene to the ftf promoter or to the gtfBC promoter were grown in a tryptone-vitamins base medium at a dilution rate of 0.1 h-1 and CAT activity was monitored in cells grown to steady state at pH 7.0, 6.0, and 5.0 under glucose-limiting or -excess conditions. The gene fusions were then introduced into a ccpA- strain of S. mutans UA159, and grown under the same conditions. Results: The expression of ftf was optimal at pH 7.0 and 6.0 under carbohydrate excess conditions. There was strong repression of ftf at pH 5.0 and a 7-10 fold reduction in cultures growing under carbohydrate limitation, compared to glucose-excess conditions. Expression of gtf was influenced modestly by pH and carbohydrate availability. Inactivation of ccpA resulted in a decrease of 5-7 fold in ftf expression, depending on the pH, and in a 10-fold decrease in gtf expression. Conclusions: These novel findings highlight the importance of pH and carbohydrate availability in expression of the genes for EPS production and reveal that CcpA is critical for the control of gtf and ftf expression by S. mutans, perhaps through the regulation of carbon flux, or by alteration of the phosphorylation state of the phosphotransferase system. This work was supported by NIDCR Award # DE12236.