Bacteria have evolved different cell envelope compositions that provide them specific advantages in their natural environments or as human pathogens. Two major types of didermic bacteria, containing cell envelopes with two lipid membranes, have been discovered that differ by the composition of the surface of their outer membranes (OM). Gram-negative bacteria (e.g. Escherichia coli) contain a surface layer of lipopolysaccharides (LPS) that produces a stringent permeability barrier at the OM and make them resistant to many antibiotics. In addition, LPS biogenesis is essential in most Gram-negative bacteria, although the reason why has remained elusive. Other diderms (e.g. many spirochetes) have glycerophospholipids (GPLs) and lipoproteins at the surface of the OM. Surface lipoproteins have many critical roles in pathogenesis, nutrient acquisition, and signaling. However, how lipoproteins reach the surface of the OM has remained unknown for most bacteria in which they are found. The high-priority Gram-negative pathogen, Acinetobacter baumannii, is remarkably able to survive with an OM containing either LPS or GPLs and lipoproteins at the surface. A. baumannii naturally produces an LPS- containing OM. However, this bacterium is able to survive with complete inactivation of LPS biogenesis. Loss of LPS at the cell surface is accompanied by up-regulation of surface-exposed lipoproteins. The primary objective of this application is to investigate the function and assembly of molecules on the surface of the cell, LPS and lipoproteins, using LPS-containing and LPS-deficient A. baumannii.
In Aim 1, we will take both a directed approach, to investigate the role of surface lipoproteins during LPS-deficiency, and a non-biased approach, to identify additional genetic requirements for LPS-deficient A. baumannii.
In Aim 2, we will characterize how lipoproteins reach the surface of the OM in A. baumannii. We will target genes implicated in lipoprotein transport in other bacteria, and perform an unbiased screen to identify genes involved in surface localization. The remarkable ability of A. baumannii to grow with two different OM compositions, provides a unique opportunity to explore the advantages each provides to bacterial cells. This application is built on a strong scientific premise addressing major gaps in our understanding of the pathogen, A. baumannii, that will have broad implications towards cell envelope biogenesis and possible treatment of both Gram-negative and other didermic bacteria. The application will be completed by the principal investigator in the lab of Dr. M. Stephen Trent (sponsor) at the University of Georgia. Both of which are committed to providing trainees with research (e.g. core facilities in genomics, microscopy, etc.) and career development resources (e.g. training in grantsmanship, teaching, mentoring, etc.). Completion of the fellowship will provide training in new techniques (e.g. lipidomics), new fields of research (e.g. genomics), and critical skills to prepare for a career as an independent researcher in academia.
Molecules on the surface of bacterial cells serve critical roles in interacting with the environment, the ability to cause disease, and resisting the human immune system and antibiotics. Gram-negative bacteria (e.g. Escherichia coli) are particularly adept at causing human infections and resisting drug treatment due to unique lipid molecules on their surface that form a protective barrier. This proposal will explore the function of surface molecules and how they are assembled in the Gram-negative bacterium Acinetobacter baumannii, an extensively drug-resistant bacterium causing devastating disease. Understanding how bacteria assemble surface molecules will allow for the development of novel therapies to fight multi-drug resistant infections.
