Combined action of two transcription factors regulates genes encoding spore coat proteins of Bacillus subtilis

During sporulation of Bacillus subtilis, spore coat proteins encoded by cot genes are expressed in the mother cell and deposited on the forespore, Transcription of the cotB, cotC, and cotX genes by sigma(K) RNA polymerase is activated by a small, DNA-binding protein called GerE, The promoter region of each of these genes has two GerE binding sites. 5' deletions that eliminated the more upstream GerE site decreased expression of lacZ fused to cotB and cotX by approximately 80% and 60%, respectively but had no effect on cotC-lacZ expression. The cotC-lacZ fusion was expressed later during sporulation than the other two fusions, Primer extension analysis confirmed that cotB mRNA increases first during sporulation, followed by cotX and cotC mRNAs over a 2-h period. In vitro transcription experiments suggest that the differential pattern of cot gene expression results from the combined action of GerE and another transcription factor, SpoIIID. A low concentration of GerE activated cotB transcription by sigma(K) RNA polymerase, whereas a higher concentration was needed to activate transcription of cotX or cotC, SpoIIID at low concentration repressed cotC transcription, whereas a higher concentration only partially repressed cotX transcription and had little effect on cotB transcription. DNase I foot-printing showed that SpoIIID binds strongly to two sites in the cotC promoter region, binds weakly to one site in the cotX promoter, and does not bind specifically to cotB, We propose that late in sporulation the rising level of GerE and the falling level of SpoIIID, together with the position and affinity of binding sites for these transcription factors in cot gene promoters, dictates the timing and level of spore coat protein synthesis, ensuring optimal assembly of the protein shell on the forespore surface.