Active Subversion of Innate Immunity by Bacterial LysM Protein

描述(申请人提供)：细胞内病原体导致惊人数量的人类感染和死亡。破坏宿主先天免疫反应的能力是这些病原体感染的关键因素。单核细胞增多性李斯特菌是一种兼性的人和动物细胞内致病菌，经常被用作研究免疫机制和发病机制的模型。我们之前对单核细胞增多性乳杆菌的研究发现了一种细菌蛋白p60，它的表达和细菌分泌是系统感染小鼠模型致病所必需的。在培养的细胞中，Lm感染不需要P60。我们发现p60刺激宿主自然杀伤细胞(NK细胞)的激活，NK细胞是一种免疫细胞群，与宿主对单核细胞增多性乳杆菌和其他几种致病菌的易感性增加有关。我们已经证明，p60通过与树突状细胞结合，并诱导IL-1和IL-18的产生来刺激NK细胞。 P60激活DC和NK细胞的能力映射到含有LysM结构域的蛋白质区域。纯化的p60蛋白，或者仅仅是含有LysM结构域的p60区域就足以刺激NK细胞的激活。在DC、NK细胞和p60的培养中，宿主因子NLRP3也是产生IL-18和激活NK细胞所必需的。在我们的第一个目标中，我们将进一步确定含有LysM结构域的多肽独特地结合和刺激DC产生IL-18和激活NK细胞的机制。将使用分子、成像、免疫学和细胞生物学方法。在我们的第二个目标中，我们将调查NLRP3在系统性细菌感染过程中如何影响宿主耐药性。在我们的第三个目标中，我们将表征NK细胞在单核细胞增多性李氏杆菌感染和p60反应中产生IL-10的特征。我们还测试了NK细胞产生IL-10如何影响宿主免疫反应和对细菌感染的敏感性。我们的工作将共同揭示含有LysM的细菌 毒力因子颠覆性地激活先天免疫的特定方面，以促进系统性感染的建立。

DESCRIPTION (provided by applicant): Intracellular pathogens cause a staggering number of human infections and deaths. The ability to subvert host innate immune responses is a key factor in the establishment of infections by these pathogens. Listeria monocytogenes is a facultative intracellular bacterial pathogen of humans and animals and is frequently used as a model to dissect mechanisms of immunity and pathogenesis. Our prior studies with L. monocytogenes identified a bacterial protein, p60, whose expression and secretion from the bacterium is required for pathogenicity in the mouse model of systemic infection. p60 is not required for Lm infection in cultured cells. We found that p60 stimulates the activation of host natural killer (NK) cells, an immune cell population associated with increased host susceptibility to L. monocytogenes and several other pathogenic bacteria. We have shown that p60 stimulates NK cells by binding to dendritic cells, and eliciting the production of IL-1¿ and IL-18. The ability of p60 to activate DCs and NK cells maps to a region of the protein containing a LysM domain. Purified p60 protein, or just the region of p60 containing the LysM domain is sufficient to stimulate NK cell activation. The host factor NLRP3 is also required for IL-18 production and NK cell activation in cultures with DCs, NK cells, and p60. In our first Aim, we will further define the mechanisms by which the LysM domain containing peptide uniquely binds and stimulates DCs for IL-18 production and NK cell activation. Molecular, imaging, immunological, and cell biological approaches will be used. In our second Aim, we will investigate how NLRP3 impacts host resistance during systemic bacterial infections. In our third Aim, we will characterize NK cells producing IL-10 in response to L. monocytogenes infection and p60. We also test how IL-10 production by NK cells impacts host immune responses and susceptibility to bacterial infection. Together, our work will reveal how LysM-containing bacterial virulence factors subversively activate specific aspects of innate immunity in order to promote the establishment of systemic infections.