The Role of RelA of Streptococcus mutans in Global Control of Gene Expression

The production of (p)ppGpp by Streptococcus mutans UA159 is catalyzed by three gene products, RelA, RelP and RelQ. Here, we investigate the role of the RelA (Rel) homologue of S. mutans in the stringent response and in global control of gene expression. RelA of S. mutans was shown to synthesize pppGpp in vitro from GTP and ATP in the absence of added ribosomes, as well as in vivo in an E. coli relA-spoT mutant. Mupirocin (MUP) was shown to induce high levels of (p)ppGpp production in S. mutans in a relA-dependent manner, with a concommitant reduction in GTP pools. Keywords: (p)ppGpp, nutrient starvation, biofilm, virulence, stress For microarray analysis, cells were grown in the chemically-defined medium FMC to an optical density at 600 nm (OD600) of 0.3 and the cultures were divided into two aliquots. To one aliquot, 500 ng ml-1 of MUP was added and the cells were incubated at 37oC for 20 minutes (MUP-treated cells), while the other aliquot was collected by centrifugation and immediately frozen (MUP-control cells). Since MUP treatment resulted in rapid growth arrest, the final OD600 of experimental and control samples were essentially identical. Four individual Cy3-labeled cDNA samples originating from four different cultures of UA159 and JLrelA strains grown under control (FMC) and experimental conditions (FMC containing MUP) were hybridized to the arrays along with Cy5-labeled reference cDNA, generating a total of 16 slides. The ratios of the signals of the test RNAs with that of the uniform reference RNA were analyzed within and between slides.