Genome-wide Analyses to Identify Acid-Inducible Genes in Streptococcus mutans

Streptococcus mutans is considered as a leading cariogenic pathogen worldwide. A significant virulence attribute of S. mutans in dental caries is its acidogenicity and aciduricity. Objective: To understand its acid tolerance mechanism, we used two genome-wide approaches to analyze and identify acid-inducible genes in S. mutans. Methods: We first used microarray to perform a global transcriptional analysis of acid-inducible gene expression of S. mutans. Total RNAs were isolated from S. mutans UA159 grown under a condition of either pH5.0 or pH7.5. The purified RNAs were used for cDNA synthesis, Cy-dye coupling and array hybridization using an indirect labeling protocol. The S. mutans microarray slides obtained from the PFGRC were scanned with Perkin ScanArray 5000XL Reader. The data were analyzed from TIFF images using TM4 suite software. To validate the array data, qRT-PCR was performed using a SYBR green PCR kit in a Cepheid Smart Cycler. In the second approach, we random screened 5,000 insertion mutants of a transposon library to identify mutants defective in acid resistance. A single primer PCR strategy was used to identify and sequence the genes flanking each transposon insertion. Results: Microarray analysis showed that 247 genes were up regulated and 240 genes down regulated after the cells were exposed to pH 5.0 for 2 hours. The upregulated genes could be classified into seven functional groups, including cell envelope components, energy metabolism, cellular processes, signaling transduction, transcriptional regulators, transports and hypothetical proteins. The down-regulated genes largely included those encoding proteins of unknown function, amino acid and protein biosynthesis. By screening the transposon library, we identified 25 mutants defective in acid resistance, although the phenotypic characterization is under investigation. Conclusion: We used two genome-wide approaches to identify a number of acid-inducible genes in S. mutans. The data provide new insights into the acid tolerance mechanisms of S. mutans.