The expression of 21 novel genes located in the region from dnaA to abrB of the Bacillus subtilis chromosome was analyzed. One of the genes, yaaH, had a predicted promoter sequence conserved among SigE-dependent genes. Northern blot analysis revealed that yaaH mRNA was first detected from 2 h after the cessation of logarithmic growth (T(2)) of sporulation in wild-type cells and in spoIIIG (SigG(-)) and spoIVCB (SigK(-)) mutants but not in spoIIAC (SigF(-)) and spoIIGAB (SigE(-)) mutants. The transcription start point was determined by primer extension analysis; the -10 and -35 regions are very similar to the consensus sequences recognized by SigE-containing RNA polymerase. A YaaH-His tag fusion encoded by a plasmid with a predicted promoter for the yaaH gene was produced from T(2) of sporulation in a B. subtilis transformant and extracted from mature spores, indicating that the yaaH gene product is a spore protein. Inactivation of the yaaH gene by insertion of an erythromycin resistance gene did not affect vegetative growth or spore resistance to heat, chloroform, and lysozyme. The germination of yaaH mutant spores in a mixture of L-asparagine, D-glucose, D-fructose, and potassium chloride was almost the same as that of wild-type spores, but the mutant spores were defective in L-alanine-stimulated germination. These results suggest that yaaH is a novel gene encoding a spore protein produced in the mother cell compartment from T(2) of sporulation and that it is required for the L-alanine-stimulated germination pathway.