Factors Mediating Host Resistance to Chlamydia trachomatis

描述（由申请方提供）：沙眼衣原体感染是世界范围内发病率较高的原因。慢性生殖道感染C.沙眼可导致女性盆腔炎、不孕不育等并发症。我们的总体目标是确定C。沙眼衣原体在感染期间与哺乳动物宿主相互作用并操纵哺乳动物宿主。在前一个项目期间，我们集中在小鼠中进行大规模的正向遗传筛选，以绘制影响对C.沙眼我们鉴定了一个免疫相关的GTP酶（IRGs）家族，该家族负责小鼠对C.沙眼然而，在人类中，IRG不负责C。沙眼抗性;相反，人类通过吲哚-2，3-双加氧酶（IDO）的表达抵抗感染。IRG和IDO的鉴定是小鼠和人类之间关于IFN-介导的抗性的关键差异，这使得我们现在提出开发模拟人类感染C.沙眼为了实现这一目标，我们将改造人源化小鼠品系，其中小鼠特异性免疫应答驱动的IRG被依赖于IDO表达的人应答替代。目前，不存在小动物模型来概括慢性疾病，因为它在人类中表现出来，特别是导致人类生殖道病理和不育的长期或复发性感染。 我们还证明了一些C。沙眼蛋白效应物在感染过程中易位到宿主细胞胞质溶胶中，在那里它们切割或改变宿主蛋白质。由于改变宿主细胞蛋白质的稳定性似乎是C.为了研究沙眼衣原体感染后的蛋白质变化，我们采用了两种新的筛选方法，即“细胞培养中氨基酸稳定同位素标记法”（SILAC）和“整体蛋白质稳定性法”（GPS），在全球范围内分析哪些宿主蛋白质在沙眼衣原体感染过程中受到了干扰。沙眼一旦我们确定了在C.沙眼衣原体感染后，我们将测试是否减少或增加这些蛋白在细胞中的流行损害C。沙眼发育尽管病原体诱导的宿主改变是发病机制的关键，但以前不可能以这些方法现在可能的规模同时评估感染对单个宿主蛋白质的影响。本申请中探索的方法首次允许了解细菌感染如何在转录水平之外全面调节宿主细胞蛋白质和途径。 理解C.沙眼衣原体与其人类宿主的关系需要一种大规模的方法来编目C.沙眼衣原体在感染过程中诱导宿主蛋白。一旦了解了这些蛋白质及其作用途径，就只能使用准确反映人类衣原体发病机制的小动物模型来理解破坏这些衣原体诱导的操纵的影响。沙眼感染

DESCRIPTION (provided by applicant): Infection with Chlamydia trachomatis is responsible for significant morbidity throughout the world. Chronic genital infections with C. trachomatis can lead to pelvic inflammatory disease, infertility, and other complications in women. Our overall goal in this grant is to define how C. trachomatis interacts with and manipulates the mammalian host during infection. In the previous project period we focused on a large-scale forward genetic screen in mice to map variant alleles that affect resistance to C. trachomatis. We identified a family of Immunity-Related-GTPases (IRGs) as responsible for mouse resistance to C. trachomatis. However in humans, IRGs are not responsible for C. trachomatis resistance; instead humans resist infection through the expression of indole-2,3-dioxygenase (IDO). The identification of IRGs and IDO as the key differences between mice and humans with regard to IFNg-mediated resistance leads us now to propose the development of a mouse model that mimics human infection with C. trachomatis. To achieve this goal we will engineer strains of humanized mice in which the mouse-specific immune response driven IRGs is replaced with the human response that depends on IDO expression. Currently, no small animal model exists that recapitulates the chronic disease as it manifests itself in humans, particularly the prolonged or recurring infections that cause pathologies of the genital tract and infertility in humans. We have also shown that a number of C. trachomatis protein effectors are translocated into the host cell cytosol during infection where they cleave or otherwise alter host proteins. Because alteration of host cell protein stability appears to be a general strategy used by C. trachomatis, we are applying two novel screening methods called "stable isotope labeling with amino acids in cells culture" (SILAC) and "Global Protein Stability" (GPS) to analyze, on a global scale, which host proteins are perturbed during infection with C. trachomatis. Once we have identified host proteins stabilized or destabilized during C. trachomatis infection, we will test whether reducing or increasing the prevalence of these proteins in cells impairs C. trachomatis development. Although pathogen-induced alterations of the host are a key to pathogenesis, it has not previously been possible to simultaneously assess the impact of infection on individual host proteins at the scale now possible with these approaches. The methods explored in this application allow, for the first time, an appreciation of how bacterial infection globally regulates host cell proteins and pathways beyond the transcriptional level. Understanding the interaction of C. trachomatis with its human host requires a large-scale approach to catalog the changes C. trachomatis induces in host proteins during infection. Once these proteins and the pathways in which they act are understood, the impact of disrupting these Chlamydia-induced manipulations can only be appreciated using small animal models that accurately reflect the pathogenesis of human C. trachomatis infection.