Identification of the promoter for a spore coat protein gene in Bacillus subtilis and studies on the regulation of its induction at a late stage of sporulation.

Abstract

The cotA (pig) gene of Bacillus subtilis encodes a 65,000 Mr protein that is a component of the spore coat and is responsible for the brown pigment characteristic of colonies in which cells are undergoing sporulation. To study developmental regulation of the cotA gene, we identified its promoter and studied its transcription in a large number of mutants blocked at various stages of sporulation and germination. Deletion analysis showed that induction and efficient transcription of cotA required DNA sequences extending no more than 55 base pairs (bp) upstream (and no more than 130 bp downstream) from the 5' terminus of cotA mRNA. Transcription from the cotA promoter was found to be switched on at approximately the time (4 to 5 h after the onset of sporulation) of spore coat synthesis and deposition. Strikingly, this transcription was substantially inhibited in almost all asporogenous mutants blocked prior to the developmental stage (V) of spore coat formation. cotA transcription was also impaired in several stage V mutants but not in other stage V mutants or in mutants blocked in germination. The germination mutant gerE caused a several-fold overexpression of cotA. The dependence of cotA expression on so many genes required at early to intermediate stages of sporulation suggests that transcription of this spore coat gene is somehow coupled (directly or through several intervening steps) to a morphological or physiological feature(s) of the developing sporangium.