Competence for Genetic Transformation in Streptococcus pneumoniae: Mutations in σ A Bypass the ComW Requirement for Late Gene Expression

Abstract

ABSTRACT Streptococcus pneumoniae is able to integrate exogenous DNA into its genome by natural genetic transformation. Transient accumulation of high levels of the only S. pneumoniae alternative σ factor is insufficient for development of full competence without expression of a second competence-specific protein, ComW. The Δ comW mutant is 10 4 -fold deficient in the yield of recombinants, 10-fold deficient in the amount of σ X activity, and 10-fold deficient in the amount of σ X protein. The critical role of ComW during transformation can be partially obviated by σ A mutations clustered on surfaces controlling affinity for core RNA polymerase (RNAP). While strains harboring σ A mutations in the comW mutant background were transforming at higher rates, the mechanism of transformation restoration was not clear. To investigate the mechanism of transformation restoration, we measured late gene expression in σ A * suppressor strains. Restoration of late gene expression was observed in Δ comW σ A * mutants, indicating that a consequence of the σ A * mutations is, at least, to restore σ X activity. Competence kinetics were normal in Δ comW σ A * strains, indicating that strains with restored competence exhibit the same pattern of transience as wild-type (WT) strains. We also identified a direct interaction between ComW and σ X using the yeast two-hybrid (Y2H) assay. Taken together, these data are consistent with the idea that ComW increases σ X access to core RNAP, pointing to a direct role of ComW in σ factor exchange during genetic transformation. However, the lack of late gene shutoff in Δ comW mutants also points to a potential new role for ComW in competence shutoff. IMPORTANCE The sole alternative sigma factor of the streptococci, SigX, regulates development of competence for genetic transformation, a widespread mechanism of adaptation by horizontal gene transfer in this genus. The transient appearance of this sigma factor is strictly controlled at the levels of transcription and stability. This report shows that it is also controlled at the point of its substitution for SigA by a second transient competence-specific protein, ComW.