Role of surface exposed proteins of M. paratuberculosis in the pathogenesis of Johne's disease in ruminants

鸟分枝杆菌亚种副结核（Map）是约翰氏病（JD）的病原体，这是反刍动物中一种使人衰弱的慢性胃肠炎。由于过早扑杀、胴体价值降低和产奶量下降，JD对奶牛有很大的经济影响。人们也越来越关注Map与克罗恩病的关联。由于无法治愈JD，控制计划受阻，迫切需要通过识别和表征参与发病机制和宿主免疫反应的细菌成分来改进诊断工具。我们在Map中发现了两个蛋白家族，PE和PPE蛋白，它们是这方面有希望的候选蛋白。这些家族是分枝杆菌特有的，并且在该属的致病成员中高度扩展。在结核分枝杆菌（Mtb）中，它们与毒力因子有关，已知在感染期间具有免疫刺激作用。一些Mtb PE/PPE蛋白在细胞表面表达，并被认为提供可能导致免疫逃避的抗原变异。Map中有10个PE基因和37个PPE基因，占基因组的1%，但它们在JD中的潜在作用从未被研究过。提出的研究假设是，Map的一些PPE蛋白通过与细胞壁结合而表面暴露，并与宿主细胞相互作用以促进感染。我们最近在体外实验中发现了4个Map的PPE蛋白表面暴露在细胞壁上。我们将进一步研究PPE蛋白在Map中的亚细胞定位，因为这暗示了它们的功能。通过分析巨噬细胞吞噬Map细胞后PPE蛋白在上皮细胞中的表达，我们将更好地了解它们在感染中的作用。我们将研究PE和PPE蛋白之间可能形成的复合物，并鉴定PPE蛋白与宿主蛋白、细胞和组织之间的其他蛋白-蛋白相互作用。最后，我们将研究PPE蛋白与细胞壁的具体相互作用。我们预计，Map中PE和PPE蛋白的功能和相互作用的发现将对了解JD的发病机制产生重大影响。

Mycobacterium avium subsp. paratuberculosis (Map) is the causative agent of Johne's disease (JD), a debilitating chronic gastroenteritis in ruminants. JD has a great economic impact in dairy cattle due to premature culling, reduced carcass value and decreased milk production. There is also a growing concern about the association of Map with Crohn's disease. With no available cure for JD and hampered control programs, there is an urgent need to improve diagnostic tools by the identification and characterization of the bacterial components involved in the pathogenesis and the host immune response. We have identified two protein families in Map, the PE and PPE proteins, which are promising candidates in this respect. Those families are unique to Mycobacteria and are highly expanded in the pathogenic members of this genus. In M. tuberculosis (Mtb), they are implicated as virulence factors and known to be immunostimulatory during infection. Some of the Mtb PE/PPE proteins are expressed on the cell surface and are suggested to provide the antigenic variation that could lead to immune evasion. There are 10 PE and 37 PPE genes in Map, comprising 1% of the genome, but their potential role in JD has never been studied. The hypothesis of the proposed studies is that some PPE proteins of Map are surface exposed by binding to the cell wall where they interact with host cells to promote infection. We have recently discovered four PPE proteins of Map to be surface exposed on the cell wall in vitro. We will further study the subcellular localization of the PPE proteins in Map since this is suggestive of their function. We will gain a better understanding of their role in infection by analyzing the expression of the PPE proteins after ingestion of Map cells by macrophages and invasion in epithelial cells. We will study the putative complex formation between PE and PPE proteins and identify other protein-protein interactions of the PPE proteins with host proteins, cells and tissues. Finally, we will investigate the specific interactions of the PPE proteins with the cell wall. We anticipate that the discovery of the function(s) and interactions of the PE and PPE proteins of Map will have a great impact on the understanding of the pathogenesis of JD.