Bacillus subtilis RNA Polymerase Recruits the Transcription Factor Spo0A∼P to Stabilize a Closed Complex during Transcription Initiation

Abstract

The Bacillus subtilis response regulator Spo0A∼P activates transcription from the spoIIG promoter by stimulating a rate-limiting transition between the initial interaction of RNA polymerase with the promoter and initiation of RNA synthesis. Previous work showed that Spo0A exerts its effect on RNA polymerase prior to the formation of an open complex in which the DNA strands at the initiation site have been separated. To isolate the effect of Spo0A∼P on events prior to DNA strand separation at spoIIG we studied RNA polymerase binding to DNA fragments that were truncated to contain only promoter sequences 5′ to the − 10 element by electrophoretic mobility shift assays. RNA polymerase bound to these fragments readily though highly reversibly, and polymerase–promoter complexes recruited Spo0A∼P. Sequence-independent interactions between the RNA polymerase and the DNA upstream of the core promoter were important for RNA polymerase binding and essential for Spo0A∼P recruitment, while sequence-specific Spo0A∼P–DNA interactions positioned and stabilized RNA polymerase binding to the DNA. Spo0A∼P decreased the dissociation rate of the complexes formed with truncated promoter templates which could contribute to the means by which Spo0A∼P stimulates spoIIG expression.