The Role of TamAB in Salmonella Pathogenesis

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ABSTRACT Salmonella Typhimurium is a major food-borne pathogen that propagates within macrophages, leading to life-threatening disease. The long-term objectives of our research are to understand the molecular mechanisms by which Salmonella circumvents the host immune system to cause disease. The outer membrane of the bacterium is the interface with the immune system. Outer membrane beta- barrel proteins are normally assembled by the Bam complex. A homolog of Bam, the TamAB translocation module is highly conserved in the Gammaproteobacteria. Originally proposed as a system for assembly of a subset of beta-barrel proteins, the so-called autotransporter proteins, the actual function of TamAB is unknown. We have discovered that tamAB is a previously unrecognized member of the PhoPQ regulon and is induced when Salmonella is replicating in macrophages, suggesting that this system is important for virulence. Indeed, our preliminary data strongly support our primary hypothesis that TamAB is induced to assist Bam function under the stressful conditions in the phagosome. Our secondary hypothesis is that TamAB is required for proper assembly of particular proteins in the outer membrane under the conditions in the phagosome, and that decreases or loss of these proteins attenuate Salmonella virulence. In Aim 1, we will determine outer membrane composition using mass spectrometry, and monitor expression of envelope stress response systems, to determine the biochemical and regulatory effects caused by loss of TamAB, thereby identifying proteins that directly or indirectly require TamAB for proper assembly. We will also isolate mutations that suppress in vitro defects in outer membrane permeability conferred or exacerbated by loss of TamAB. In Aim 2, we will characterize identified loci and their role in macrophage survival and virulence. The overall role of the identified factors in macrophage survival will be revealed by genetic interactions with tamAB. Understanding the roles of TamAB will inform us about the conditions in the phagosome leading to outer membrane assembly stress and the mechanisms used by Salmonella to combat them. Identifying the specific proteins affected by loss of TamAB also identifies additional virulence factors critical for Salmonella pathogenesis.