The Role of TamAB in Salmonella Pathogenesis

摘要 鼠伤寒沙门氏菌是一种主要的食源性病原体，在巨噬细胞内繁殖， 导致危及生命的疾病。我们研究的长期目标是了解 沙门氏菌绕过宿主免疫系统引起疾病的分子机制。 细菌的外膜是与免疫系统的界面。外膜β- 桶蛋白通常由BAM复合物组装。巴姆的同系物，TamAB 易位模块在γ-变形菌中高度保守。最初是作为一个系统提出的 为了组装β-桶蛋白的一个子集，即所谓的自转运蛋白， Tamab的功能未知。我们发现tamAB是一个以前未被识别的成员， 当沙门氏菌在巨噬细胞中复制时， 该系统对于毒力是重要的。事实上，我们的初步数据有力地支持了我们的主要观点。 假设TamAB被诱导，以协助BAM功能的压力条件下，在 吞噬体我们的次要假设是TamAB是正确组装 在吞噬体的条件下，外膜中的特定蛋白质， 这些蛋白质的减少或损失减弱沙门氏菌的毒力。在目标1中，我们将确定 外膜组成，并监测包膜应力的表达 反应系统，以确定由TamAB损失引起的生化和调节作用， 从而鉴定直接或间接需要TamAB进行正确组装的蛋白质。我们还将 抑制体外外膜通透性缺陷的分离突变赋予或 因失去TamAB而加剧。在目标2中，我们将描述已鉴定的基因座及其在以下方面的作用： 巨噬细胞存活和毒力。巨噬细胞存活中已识别因子的总体作用 将通过与tamAB的基因相互作用来揭示。了解TamAB的作用将使我们了解 关于吞噬体中导致外膜组装应力的条件和 沙门氏菌用来对抗它们的机制。识别受蛋白质丢失影响的特定蛋白质 TamAB还鉴定了对沙门氏菌致病至关重要的其他毒力因子。

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.