Induction kinetics of the PhoQ-PhoP two-component system in Escherichia coli

Abstract

Cells rely on signal transduction systems to sense and respond to changes in their enviroments. When a stimulus is present, the corresponding signal transduction system will activate and enact the appropriate biological response, often by modulating target gene expression. In many cases, the temporal dynamics of the activation of target gene expression in the presence of constant stimulus is complex, and often exhibits one or several pulses. How these complex temporal dynamics are regulated at the molecular level is unknown for many signal transduction systems. In this thesis, I discuss the molecular regulation of the temporal dynamics of PhoQ-PhoP induction in Escherichia coli. The PhoQ-PhoP pathway is a canonical two-component system that responds to low extracellular Mg'+, certain antimicrobial peptides, and potentially other unknown factors. Upon activation, the bifunctional histidine kinase PhoQ autophosphorylates and subsequently phosphotransfers to the response regulator PhoP, thereby activating it to increase transcription of PhoP target genes. Because PhoQ is bifunctional, PhoQ acts as a phosphatase on phosphorylated PhoP in the absence of stimulus, thereby keeping the system inactivated. When the PhoQ-PhoP system is strongly induced, PhoP target genes exhibit impulse kinetics, meaning gene expression increases to a maximal level and subsequently decreases to an eventual steady state. We discovered that this impulse response is caused by a negative feedback loop in which active PhoP transcribes mgrB, a gene encoding a small membrane protein that interacts directly with PhoQ to repress the output of the system. MgrB selectively inhibits the ability of PhoQ to phosphorylate PhoP, and permits PhoQ to act as a phosphatase on phosphorylated PhoP. This change in PhoQ activity causes a decrease in the level of active PhoP and the level of PhoP target genes. This thesis reveals how negative feedback loops and histidine kinase bifunctionality can drive the kinetics of two-component system induction in bacteria, and more generally explores how cells regulate changes in gene expression over time.