Acetyl phosphate-dependent activation of a mutant PhoP response regulator that functions independently of its cognate sensor kinase.
The two-component system is a signal communication network generally consisting of a sensor kinase that receives inputs from the environment and modifies the phosphorylated state of a response regulator that executes an adaptive behavior. PhoP is a response regulator that controls virulence gene expression in Salmonella enterica. Transcription of PhoP-regulated genes is modulated by the Mg(2+) levels detected by the sensor PhoQ. Here, we describe a PhoP mutant protein, PhoP*, that functions in the absence of its cognate sensor, thereby allowing transcription of PhoP-activated genes independently of the Mg(2+ )concentration in the environment. The PhoP* protein harbors a S93N substitution in the response regulator receiver domain. PhoP*-mediated transcription is abolished by either mutation of the aspartate residue that is conserved among response regulators as the site of phosphorylation or inactivation of the pta-encoded phosphotransacetylase. This enzyme mediates the production of acetyl phosphate, which has been shown to serve as a low molecular mass phosphate donor for certain response regulators. The purified PhoP* protein autophosphorylated from acetyl phosphate more efficiently than the wild-type PhoP protein in vitro. The PhoP* protein retained the capacity to interact with the PhoQ protein, which promoted phosphorylation of the PhoP* protein in vitro and abolished PhoP*-mediated transcription under high Mg(2+ )concentrations in vivo. Cumulatively, our results uncover a role of PhoQ in transcriptional repression during growth in millimolar Mg(2+ )and define a mutant response regulator form with an increased capacity to be phosphorylated by acetyl phosphate.
Chamnongpol
S
S
Howard Hughes Medical Institute, Department of Molecular Microbiology, Washington University School of Medicine, 660 S. Euclid Ave., St. Louis, MO 63110, USA.
Groisman
E A
EA
D000483:Alleles; D019943:Amino Acid Substitution / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004777:Environment; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D005798:Genes, Bacterial / Q000235:genetics; D005809:Genes, Regulator / Q000235:genetics; D008274:Magnesium / Q000494:pharmacology; D008565:Membrane Proteins / Q000737:chemistry; D000072236:Methyl-Accepting Chemotaxis Proteins; D008958:Models, Molecular; D009154:Mutation / Q000235:genetics*; D009876:Operon / Q000235:genetics; D010755:Organophosphates / Q000494:pharmacology*; D010766:Phosphorylation; D011485:Protein Binding; D014176:Protein Biosynthesis / Q000187:drug effects; D017434:Protein Structure, Tertiary; D011993:Recombinant Fusion Proteins; D019779:Salmonella enterica / Q000187:drug effects / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D001426:Bacterial Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; D010755:Organophosphates; C079792:PhoQ protein, Bacteria; D011993:Recombinant Fusion Proteins; D015534:Trans-Activators; C061064:PhoP protein, Bacteria; C011632:acetyl phosphate; D008274:Magnesium