CodY Regulates Expression of the Bacillus subtilis Extracellular Proteases Vpr and Mpr
ABSTRACTCodY is a global transcriptional regulator in low-G+C Gram-positive bacteria that is responsive to GTP and branched-chain amino acids. By interacting with its two cofactors, it is able to sense the nutritional and energetic status of the cell and respond by regulating expression of adaptive genetic programs. InBacillus subtilis, more than 200 genes, including those for peptide transporters, intracellular proteolytic enzymes, and amino acid degradative pathways, are controlled by CodY. In this study, we demonstrated that expression of two extracellular proteases, Vpr and Mpr, is negatively controlled by CodY. By gel mobility shift and DNase I footprinting assays, we showed that CodY binds to the regulatory regions of both genes, in the vicinity of their transcription start points. Themprgene is also characterized by the presence of a second, higher-affinity CodY-binding site located at the beginning of its coding sequence. Using strains carryingvpr- ormpr-lacZtranscriptional fusions in which CodY-binding sites were mutated, we demonstrated that repression of both protease genes is due to the direct effect by CodY and that themprinternal site is required for regulation. Thevprpromoter is a rare example of a sigma H-dependent promoter that is regulated by CodY. In acodYnull mutant, Vpr became one of the more abundant proteins of theB. subtilisexoproteome.IMPORTANCECodY is a global transcriptional regulator of metabolism and virulence in low-G+C Gram-positive bacteria. InB. subtilis, more than 200 genes, including those for peptide transporters, intracellular proteolytic enzymes, and amino acid degradative pathways, are controlled by CodY. However, no role forB. subtilisCodY in regulating expression of extracellular proteases has been established to date. In this work, we demonstrate that by binding to the regulatory regions of the corresponding genes,B. subtilisCodY negatively controls expression of Vpr and Mpr, two extracellular proteases. Thus, inB. subtilis, CodY can now be seen to regulate the entire protein utilization pathway.