Spo0A activates transcription in Bacillus subtilis from promoters that are used by two types of RNA polymerase, RNA polymerase containing the primary sigma factor, ^A, and RNA polymerase containing a secondary sigma factor, known as ^H. The region of ^A near positions 356 to 359 is required for Spo0A-dependent promoter activation, possibly because Spo0A interacts with this region of ^A at these promoters. To determine if the amino acids in the corresponding region of ^H are also important in Spo0A-dependent promoter activation, we examined the effects of single alanine substitutions at 10 positions in ^H (201 to 210). Two alanine substitutions in ^H, at glutamine 201 (Q201A) and at arginine 205 (R205A), significantly decreased activity from the Spo0A-dependent, ^H-dependent promoter spoIIA but did not affect expression from the ^H-dependent, Spo0A-independent promoters citGp2 and spoVG. Therefore, promoter activation by Spo0A requires homologous regions in ^A and ^H. A mutant form of Spo0A, S231F, that suppresses the sporulation defect caused by several amino acid substitutions in ^A did not suppress the sporulation defects caused by the Q201A and R205A substitutions in ^H. This result and others indicate that different surfaces of Spo0A probably interact with ^A and ^H RNA polymerases.