IADR Abstract Archives
C-di-AMP Signaling Pathway Regulates the Competitiveness of Streptococcus Mutans
Objectives: The oral microecological balance is closely associated with the development of dental caries. S. mutans inhibits the growth of oral symbiotic bacteria by synthesizing mutacins, and resists hydrogen peroxide (H2O2) stress produced by oral symbiotic bacteria such as Streptococcus sanguinis. c-di-AMP is a key factor regulating the biofilm formation and environmental stress response of S. mutans. This study aimed to elucidate the role of bacterial H2O2 in regulating the survival competitiveness of S. mutans via c-di-AMP signaling pathway.
Methods: Quantitative RT-PCR was used to quantify gene expression. CdaA protein was purified and HPLC was used to determine the concentration of c-di-AMP. Whole-genome microarrays was performed to identify the global transcriptional changes in cdaA mutant. EMSA was performed to investigate the binding of c-di-AMP to MubR protein. Dual-species biofilm was constructed and the spatio-temporal interaction was investigated using fluorescence in situ hybridization (FISH). A rat model of dental caries was constructed to determine the cariogenicity of cdaA mutant.
Results: The expression of cdaA was specifically induced by oxidative stress (0.002% H2O2). The exogenous H2O2 also stimulated the Mn2+ uptake and c-di-AMP synthesis. The biological activity of CdaA protein showed Mn2+ concentration dependence, suggesting that exogenous H2O2 promotes the synthesis of c-di-AMP by regulating the uptake of Mn2+. Microarray data showed down-regulation of a number of genes within the mub gene cluster in cdaA mutant. EMSA assay showed that c-di-AMP could bind to the transcriptional regulator of mub (MubR), suggesting that c-di-AMP regulates the oxidative stress of S. mutans by regulating mub expression. The S. mutans cdaA mutant exhibited increased sensitivity to oxidative stress and compromised competitiveness against S. sanguinis. The number of cdaA mutant colonized in mouse oral cavity decreased, and its cariogenicity also decreased based on the Keyes score.
Conclusions: Bacterial H2O2 promotes the synthesis of c-di-AMP by regulating the uptake of Mn2+. c-di-AMP regulates the oxidative stress and competitiveness of S. mutans by regulating mub expression. Results obtained from this study suggest that c-di-AMP may be a novel target for the ecological prevention and treatment of dental caries.
Methods: Quantitative RT-PCR was used to quantify gene expression. CdaA protein was purified and HPLC was used to determine the concentration of c-di-AMP. Whole-genome microarrays was performed to identify the global transcriptional changes in cdaA mutant. EMSA was performed to investigate the binding of c-di-AMP to MubR protein. Dual-species biofilm was constructed and the spatio-temporal interaction was investigated using fluorescence in situ hybridization (FISH). A rat model of dental caries was constructed to determine the cariogenicity of cdaA mutant.
Results: The expression of cdaA was specifically induced by oxidative stress (0.002% H2O2). The exogenous H2O2 also stimulated the Mn2+ uptake and c-di-AMP synthesis. The biological activity of CdaA protein showed Mn2+ concentration dependence, suggesting that exogenous H2O2 promotes the synthesis of c-di-AMP by regulating the uptake of Mn2+. Microarray data showed down-regulation of a number of genes within the mub gene cluster in cdaA mutant. EMSA assay showed that c-di-AMP could bind to the transcriptional regulator of mub (MubR), suggesting that c-di-AMP regulates the oxidative stress of S. mutans by regulating mub expression. The S. mutans cdaA mutant exhibited increased sensitivity to oxidative stress and compromised competitiveness against S. sanguinis. The number of cdaA mutant colonized in mouse oral cavity decreased, and its cariogenicity also decreased based on the Keyes score.
Conclusions: Bacterial H2O2 promotes the synthesis of c-di-AMP by regulating the uptake of Mn2+. c-di-AMP regulates the oxidative stress and competitiveness of S. mutans by regulating mub expression. Results obtained from this study suggest that c-di-AMP may be a novel target for the ecological prevention and treatment of dental caries.