The ctc gene of Bacillus subtilis is transcribed in vitro by the minor RNA polymerase holoenzyme forms, E sigma 37 and E sigma 32. To study the expression and regulation of ctc in vivo, we constructed operon and translational fusions of the ctc promoter region to the lacZ gene of Escherichia coli. Our results indicate that ctc is regulated at the transcriptional level, and that this RNA synthesis is maximally induced at the end of the exponential phase of growth under nutritional conditions which inhibit the activity of the tricarboxylic acid cycle. Analysis of in vitro-constructed deletion mutations extending into the ctc promoter region demonstrated that the region required for this regulation is no greater than 53 base-pairs in length. We also compared the expression of ctc to that of another B. subtilis gene, which is transcribed by E sigma 37 and E sigma 32 in vitro, the sporulation gene spoVG. Although the ctc and spoVG promoter regions are recognized by the same forms of RNA polymerase in vitro, our results show that they differ strikingly in the nutritional and genetic requirements for their expression in vivo.