Through mainly in vivo studies, the initiation of sporulation in Bacillus subtilis has been shown to depend on the phosphorylation of the Spo0A transcription factor mediated by the multicomponent phosphorelay via KinAB (C), Spo0F, Spo0B, and Spo0A in this order. RNA polymerase containing sigmaA (EsigmaA) or sigmaH (EsigmaH) transcribes the genes of the phosphorelay components. Phosphorylated Spo0A is also involved in their expression and is required for the induction of sigmaH by repressing its repressor gene abrB. We have examined the effects of phosphorylated Spo0A (Spo0A-P) and AbrB on in vitro transcription of the genes involved in the Spo0A phosphorylation and initiation of sporulation. Spo0A-P repressed EsigmaA-dependent transcription of the kinC and EsigmaH-dependent transcription of spo0A and kinA. EsigmaH-dependent transcription of spo0F was stimulated by Spo0A-P at low concentrations but was repressed by higher amounts of Spo0A-P. On the other hand, AbrB repressed EsigmaA-dependent transcription of spo0H (sigmaH gene), kinC, and abrB, although its effect was not strong. With the present results providing in vitro evidence for the roles of Spo0A-P and AbrB as transcriptional regulators, and other results described in the literature, the positive and negative feedback loops controlling the temporal expression of early sporulation genes are discussed.