The spoIIE operon is a developmentally regulated transcription unit activated in the second hour of sporulation in Bacillus subtilis. Its promoter has an unusual structure, containing sequences which conform perfectly to the consensus for vegetative promoters recognized by sigma A-associated RNA polymerase (E sigma A), but with a spacing of 21 bp between the apparent -10 and -35 elements instead of the 17- or 18-bp spacing typical of promoters utilized by E sigma A. Mutations introduced into the apparent -10 element affected transcription in a manner consistent with its functioning as a polymerase recognition sequence. The deleterious effect of one -10 mutation was also suppressed in an allele-specific manner by a mutation in sigA known to suppress analogous -10 mutations in conventional vegetative promoters recognized by E sigma A. Similar suppression experiments failed to provide evidence for a direct interaction between E sigma A and the "-35-like" element, however, and DNase I protection experiments suggested instead that the Spo0A protein binds to a site overlapping this -35-like hexamer. Moreover, the effects of mutations within the -35-like hexamer on the binding of Spo0A in vitro paralleled their effects on transcription in vivo. We suggest that spoIIE belongs to a class of early-intermediate sporulation genes whose transcription by E sigma A is activated by the Spo0A protein.