Regulation of transcription of the Bacillus subtilis pyrG gene, encoding cytidine triphosphate synthetase

The B. subtilis pyrG gene, which encodes CTP synthetase, is located far fro m the pyrimidine biosynthetic operon on the chromosome and is independently regulated. The pyrG promoter and 5 ' leader were fused to lacZ and integra ted into the chromosomes of several B. subtilis strains having mutations in genes of pyrimidine biosynthesis and salvage. These mutations allowed the intracellular pools of cytidine and uridine nucleotides to be manipulated b y the composition of the growth medium. These experiments indicated that py rG expression is repressed by cytidine nucleotides but is largely independe nt of uridine nucleotides. The start of pyrG transcription was mapped by pr imer extension to a position 178 nucleotides upstream of the translation in itiation codon. A factor-independent termination hairpin lying between the pyrG promoter and its coding region is essential for regulation of pyrG exp ression. Primer-extended transcripts were equally abundant in repressed and derepressed cells when the primer bound upstream of the terminator, but th ey were much less abundant in repressed cells when the primer bound downstr eam of the terminator. Furthermore, deletion of the terminator from pyrG-la cZ fusions integrated into the chromosome yielded elevated levels of expres sion that was not repressible by cytidine. We suggest that cytidine repress ion of pyrG expression is mediated by an antitermination mechanism in which antitermination by a putative trans-acting protein is reduced by elevated levels of cytidine nucleotides. Conservation of sequences and secondary str uctural elements in the pyrG 5 ' leaders of several other gram-positive bac teria indicates that their pyrG genes are regulated by a similar mechanism.