We report the cloning and characterization of a cell division gene, herein designated divIC, from the gram-positive, spore-forming bacterium Bacillus subtilis. This gene was previously identified on the basis of a temperature-sensitive mutation, div-355, that blocks septum formation at restrictive temperatures. We show that the divIC gene is a 125-codon open reading frame that is capable of encoding a protein of 14.7 kDa and that div-355 is a 5-bp duplication near the 3' end of the open reading frame. We also show that divIC is an essential gene by use of an in vitro-constructed null mutation. In confirmation and extension of earlier results, we show that divIC is necessary for both vegetative and sporulation septum formation, and we demonstrate that it is required for the activation of genes expressed under the control of the sporulation transcription factors sigma F and sigma E. The divIC gene is located 1.3 kb upstream of the coding sequence for the sporulation gene spoIIE. Between divIC and spoIIE is a 128-codon open reading frame whose predicted product contains a region of similarity to the RNA-binding domains of polynucleotide phosphorylase and ribosomal protein S1 from Escherichia coli and two putative tRNA genes for methionyl-tRNA and glutamyl-tRNA, the gene order being divIC orf128 tRNA(Met) tRNA(Glu) spoIIE.