Acetyl phosphate-dependent activation of a mutant PhoP response regulator that functions independently of its cognate sensor kinase

Abstract

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 Mg2+ 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 Mg2+ 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 Mg2+ concentrations in vivo. Cumulatively, our results uncover a role of PhoQ in transcriptional repression during growth in millimolar Mg2+ and define a mutant response regulator form with an increased capacity to be phosphorylated by acetyl phosphate.