Identification of stress-responsive genes in Streptococcus mutans by differential display reverse transcription-PCR

Streptococcus mutans, which causes dental caries in the human oral cavity a nd occasionally causes infective endocarditis in the heart, withstands adve rse environmental stress through diverse alterations in protein synthesis. Differential gene expression in response to environmental stress was analyz ed by RNA fingerprinting using arbitrarily primed PCR with a panel of 11mer primers designed for differential display in Enterobacteriaceae. Dot and N orthern blot hybridization confirmed that the transcription of several gene s was up- or down-regulated following exposure to acid shock from pH 7.5 to 5.5. RNA of a gene designated AP-185 (acid stress protein) was induced spe cifically by acid treatment, while RNA of GSP-781 (general-stress protein) was up-regulated significantly when bacteria were exposed to high osmolarit y and temperature, as well as low pH. The deduced amino acid sequence of AP -185 shares homology (78% identity) with branched-chain amino acid aminotra nsferase. Cloning and sequence analysis of GSP-781 revealed a potential sec reted protein of a molecular mass of about 43 kDa and with a pi predicted t o be 5.5. Transcriptional levels of another gene, designated AR-186 (acid-r epressed protein), which encodes putative aconitase, were repressed by acid treatment but were enhanced by plasma or serum components. Analogous resul ts were identified in icd and citZ genes, and repression of these genes, al ong with AR-186, was also observed when they were exposed to high osmolarit y and temperature. These results indicate that differential regulation of s pecific genes at the transcriptional level is triggered by different stress and that genes responsible for glutamate biosynthesis in the citrate pathw ay are coordinately regulated during the stress response of S. mutans.