A mutation in GerE that affects cotC promoter activation in Bacillus subtilis

The DNA-binding protein GerE acts as both a repressor and an activator of transcription of genes transcribed by sigma(K)-RNA polymerase (RNA-P) during the later stages of endospore formation in Bacillus subtilis. GerE represses transcription from the sigK promoter, and activates transcription from other promoters, including cotC and cotX. Two different regions of GerE (AR1 and AR2) are required for activation of cotC and cotX, respectively. We used a genetic screen to seek mutations that would define additional regions of GerE required for promoter activation. We found that a substitution of proline for leucine at position 12 of GerE (L12P) decreased cotC promoter activity but did not interfere with GerE-dependent repression of the sigK promoter or with activation of the cotX promoter in vivo. We also found that the L12P substitution had no effect on binding to cotC in vitro. However, the L12P-substituted GerE failed to stimulate cotC transcription in vitro, whereas it stimulated transcription from PcotX. The crystal structure of GerE suggests that L12 is not exposed on the surface of the molecule. Therefore, we propose that the L12P substitution reduces the flexibility of the N-terminal arm, preventing an interaction of AR1 with RNA-P that is essential for activation of the cotC promoter.