Regulation of σ(B) levels and activity in Bacillus subtilis

The sigB operon of Bacillus subtilis encodes σ(B) plus three additional proteins (RsbV, RsbW, and RsbX) that regulate σ(B) activity. Using an anti- σ(B) monoclonal antibody to monitor the levels of σ(B) protein, P(SPAC) to control the expression of the sigB operon, and a ctc-lacZ reporter system to monitor σ(B) activity, we observed that the rsbV and rsbW products control σ(B) activity at the ctc promoter independently of their effects on σ(B) levels. In contrast, RsbX was found to have no effect on expression of ctc when the sigB operon was controlled by P(SPAC). The data are consistent with RsbV and RsbW being regulators of σ(B) activity and RsbX acting primarily as a negative regulator of sigB operon expression. Evidence that stationary- phase induction of the σ(B)-dependent ctc promoter is accomplished by a reduction in RsbW-dependent inhibition of σ(B) activity is also presented. In addition, Western blot (immunoblot) analyses of sigB operon expression demonstrated that σ(B) accumulation is coupled to the synthesis of its primary inhibitor (RsbW). This finding is consistent with RsbW and σ(B) being present within the cell in equivalent amounts, a circumstance that would permit RsbW to directly influence σ(B) activity by a direct protein- protein interaction.