Imaging Two-Component Systems in E. Coli

University Of Pennsylvania, Philadelphia PA

Investigators

Abstract

Two-component signaling systems are one of the major modes of signal transduction in bacteria. These regulatory circuits, which in their simplest forms are characterized by two proteins, a sensor kinase and a response regulator, play a central role in controlling basic aspects of microbial physiology and mediate responses to diverse environmental signals. Genetics and biochemical studies have made substantial progress in establishing the mechanisms that govern signal transduction in two-component systems. However, there has been relatively little work on the spatial and temporal organization of this important class of circuits within cells. This research will focus on the EnvZ/OmpR system in Escherichia coli, which is one of the simplest and best characterized examples of two-component signaling. In preliminary work, functional fusions of green fluorescent protein (GFP) to the histidine kinase EnvZ and to the response regulator OmpR were constructed. It was found that both proteins exhibit a heterogeneous distribution within cells. The present research will study the spatial localization of these regulatory proteins, using a combination of molecular biology, fluorescence microscopy, genetics, and quantitative analysis. The work will focus on the molecular determinants and dynamics of EnvZ and OmpR localization and on the correlation with signaling activity. This research will provide the first steps towards formulating a detailed picture of the spatial organization of this simple regulatory circuit and towards understanding its physiological significance. The results will also have important implications for the cell biology of other two-component systems and more generally provide new insights into the spatial organization of the bacterial cell. The broader impacts of this project will provide valuable interdisciplinary training for graduate students, undergraduates, and high-school students in aspects of molecular biology and microbiology as well as biophysics and mathematical analysis. The research will thus train scientists who are able to effectively bridge the gaps between the biological sciences and traditionally more quantitative disciplines such as mathematics, physics, and computer science.