Autoinduction of Bacillus subtilis phoPR Operon Transcription Results from Enhanced Transcription from EA- and EE-Responsive Promoters by

The phoPR operon encodes a response regulator, PhoP, and a histidine kinase, PhoR, which activate or repress genes of the Bacillus subtilis Pho regulon in response to an extracellular phosphate deficiency. Induction of phoPR upon phosphate starvation required activity of both PhoP and PhoR, suggesting autoregulation of the operon, a suggestion that is supported here by PhoP footprinting on the phoPR promoter. Primer extension analyses, using RNA from JH642 or isogenic sigE or sigB mutants isolated at different stages of growth and/or under different growth conditions, suggested that expression of the phoPR operon represents the sum of five promoters, each responding to a specific growth phase and environmental controls. The temporal expression of the phoPR promoters was investigated using in vitro transcription assays with RNA polymerase holoenzyme isolated at different stages of Pho induction, from JH642 or isogenic sigE or sigB mutants. In vitro transcription studies using reconstituted EA, EB, and EE holoenzymes identified PA4 and PA3 as E A promoters and PE2 as an EE promoter. Phosphorylated PhoP (PhoPP) enhanced transcription from each of these promoters. EB was sufficient for in vitro transcription of the PB1 promoter. P5 was active only in a sigB mutant strain. These studies are the first to report a role for PhoPP in activation of promoters that also have activity in the absence of Pho regulon induction and an activation role for PhoPP at an EE promoter. Informatio