Biogenesis of bacterial membranes: from fundamental mechanisms to novel antimicrobial targets

Abstract

The last decade marked a paradigm shift in the way we understand and study the subcellular organization of bacteria. The idea of "bags of enzymes" was replaced by the realization that essential processes such as DNA replication, cell division and cell wall expansion are performed by higher-order protein complexes, whose subcellular localization and activity are precisely regulated in time and space. Despite these advances, we still do not understand how the plasma membrane of bacteria is constructed. The enzymes and chemical reactions that produce phospholipids have been worked out, but how the insertion of new building blocks into an expanding membrane is organized in space and time, and how membrane synthesis is coordinated with cell growth remain as fundamental questions of bacterial cell biology. The main goal of this project is to provide a detailed description of the molecular mechanisms of bacterial membrane biogenesis, with focus on two prominent questions: 1) Where is new membrane being synthesized? and 2) How bacteria knows how much membrane needs to be synthesized? To answer (1), we will apply modern fluorescence microscopy methods to determine the subcellular localization and dynamics of phospholipid-synthesizing enzymes and whether they function alone or in a multiprotein complex. To answer (2), we will apply a complementary array of methods (molecular genetics, biochemistry, proteomics and metabolomics) to investigate the interplay between lipid and membrane synthesis and the second messenger ppGpp, a key regulator of bacterial growth. Since proper control of lipid metabolism is essential for survival, an additional goal of this project will be to identify inhibitors of key targets in membrane biogenesis. We will leverage the basic knowledge gathered along the project to set up "target-based whole-cell assays" and screen different collections of compounds (small molecules, fragments, natural products). Because the targets selected are novel, the inhibitors identified may represent leads to completely new classes of antibiotics. (AU)