Autoinduction of Bacillus subtilis phoPR operon transcription results from enhanced transcription from EsigmaA- and EsigmaE-responsive promoters by phosphorylated PhoP.
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 EsigmaA, EsigmaB, and EsigmaE holoenzymes identified PA4 and PA3 as EsigmaA promoters and PE2 as an EsigmaE promoter. Phosphorylated PhoP (PhoP approximately P) enhanced transcription from each of these promoters. EsigmaB 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 PhoP approximately P in activation of promoters that also have activity in the absence of Pho regulon induction and an activation role for PhoP approximately P at an EsigmaE promoter. Information concerning PB1 and P5 creates a basis for further exploration of the regulatory coordination or overlap of the PhoPR and SigB regulons during phosphate starvation.
Paul
Salbi
S
Laboratory for Molecular Biology, Department of Biological Sciences, University of Illinois at Chicago, Chicago, Illinois 60607, USA.
Birkey
Stephanie
S
Liu
Wei
W
Hulett
F Marion
FM
D001412:Bacillus subtilis / Q000235:genetics*; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism / Q000502:physiology; D001483:Base Sequence; D001665:Binding Sites; D012321:DNA-Directed RNA Polymerases / Q000502:physiology*; D015964:Gene Expression Regulation, Bacterial; D008969:Molecular Sequence Data; D009876:Operon; D010766:Phosphorylation; D011401:Promoter Regions, Genetic; D012808:Sigma Factor / Q000502:physiology*; D014157:Transcription Factors / Q000502:physiology*; D014158:Transcription, Genetic
D001426:Bacterial Proteins; C079011:SigB protein, Bacteria; D012808:Sigma Factor; D014157:Transcription Factors; C063202:sporulation-specific sigma factors; C061064:PhoP protein, Bacteria; C063505:RNA polymerase sigma A; D012321:DNA-Directed RNA Polymerases