Catabolite repression-resistant mutations of the Bacillus subtilis alpha-amylase promoter affect transcription levels and are in an operator-like sequence

Abstract

The amyR1 locus controls the regulated transcription of amyE, the structural gene encoding alpha-amylase in Bacillus subtilis. Transcription of amyE is activated in early stationary phase cells, and can be repressed by rapidly metabolized carbon sources such as glucose. Transcription of amyE initiates in vitro from a promoter recognized by the major vegetative form of RNA polymerase, E sigma 43. S1 nuclease mapping of in-vivo amylase transcripts suggests that this promoter is also used in vivo. Two independently isolated cis-acting mutations, gra-5 and gra-10, which abolish glucose-mediated repression of amylase synthesis without altering temporal activation, were determined by DNA sequencing to result from a G.C to A.T transition at a position located five base-pairs downstream from the start site of transcription. While this is the first example of a site involved in catabolite repression of gene expression in a Gram-positive micro-organism, the region surrounding the gra mutations shows considerable homology to certain cis-acting regulatory loci in Escherichia coli, suggesting that such sequences have been evolutionarily conserved.