Uncovering specialized roles for membrane phospholipids in protein localization and function in Escherichia coli.

摘要

Bacteria execute remarkably complex processes by coordinating the spatial and temporal localization of macromolecules. The physical, chemical and geometric cues that bacteria utilize to perform such precise and coordinated localization is a subject that has been the focus of much study in recent years. In rod-shaped bacteria such as Escherichia coli, the curvature at the hemispherical poles is much larger than that at the sidewall of the cell. The E. coli inner membrane contains three major phospholipids-phosphatidylethanolamine (PE), phosphatidylglycerol (PG) and cardiolipin (CL). Of these, cardiolipin has a high intrinsic curvature and clusters of phase separated CL that form within the inner membrane localize to the poles and are stabilized by the high curvature at this region. The heterogeneity in membrane composition observed in this region of the cell has been hypothesized to serve as landmarks for the recruitment of proteins to the poles. This dissertation describes how RecA-the primary enzyme required for recombination repair in E. coli, is stabilized at the poles by its interaction with PG and CL. This interaction is not only necessary to anchor these 'storage' forms of RecA but also facilitates the formation of RecA filament bundles after DNA is damaged and helps promote the SOS response. As an additional layer of regulation, the SOS response also regulates the synthesis of anionic phospholipids through the presence of gene regulatory elements upstream of PG synthase. Studies of the SOS response and other stress responses have been advanced by the discovery and characterization of novel small molecule inhibitors that target these processes. In this dissertation, I also describe the characterization of a DNA gyrase inhibitor-the Gyramides-that inhibit cell division via the SOS response but does not inflict breaks or cuts in the DNA. Together these studies provide interesting insights into the role of the membrane in DNA repair and the interplay between protein localization and gene regulation.