Mechanisms of MHCII expression and CD4+ T cell activation during Chlamydia trachomatis infection

项目摘要 哺乳动物宿主需要CD4+T细胞才能在感染了许多不同的细胞内病原体的情况下存活下来。 CD4+T细胞丢失导致对沙眼衣原体感染的易感性 肺结核和其他疾病。抗原提呈细胞(APC)位于入侵病原体和 适应性免疫，感知受感染的环境，并直接向CD4+T细胞发出信号。的表达方式 APC上的主要组织相容性复合体II(MHCII)是CD4+T细胞检测同源细胞的关键 并激活适当的宿主反应。许多病原体操纵MHCII的表达以 逃避保护豁免权。在完全丧失MHCII阻止CD4+T细胞功能的同时，如何调节 感染期间MHCII水平影响病原体特异性CD4+T细胞反应的有效性 未知。我们假设MHCII的表达水平是激活和效应的一个重要因素 抗原特异性的CD4+T细胞的功能。通过了解MHCII如何在全球范围内受到监管，我们将 定位于开发调节MHCII表达的干预措施，以克服病原体的毒力 旨在削弱保护性宿主反应的策略。我们进行了全基因组CRISPR-Cas9筛查 确定激活的APC中MHCII表面表达的调节因子。我们发现超过200个基因显著地 改变表面MHCII的表达，但在MHCII调节中的功能未知。这里，使用CRISPR-Cas9 中介功能增益法和功能损失法我们将系统地确定优先候选者， 增强或抑制MHCII的表面表达，并开始剖析其调控MHCII的机制。我们会 接下来，确定MHCII表达的调节如何直接改变病原体特异性的CD4+T细胞反应 在生殖器感染鼠衣原体期间。我们正在调查衣原体，因为它是一种重要的 病原体在临床上被列为美国头号细菌性传播感染。衣原体 导致严重的病理，导致不孕、宫外孕和盆腔炎。我们会 使用大多数情况下不存在的免疫学工具跟踪衣原体特异性的CD4+T细胞反应 病原体。与我们的CRISPR方法相结合，我们处于独特的地位，可以直接检查 MHCII的表达影响衣原体特异性的体内和体外的CD4+T细胞反应。我们将决定 APC表面MHCII表达增强或降低如何改变一系列CD4+T细胞表型，包括 生殖道病理的激活、增殖、效应器功能、保护和发展。我们的 关于MHCII表达在沙眼衣原体感染过程中作用的重点研究将广泛确定调控因素 MHCII的作用，然后可以在一系列需要CD4+T细胞激活的疾病状态下进行探索 MHCII被操纵，包括其他细胞内病原体或癌症。

Project Summary The mammalian host requires CD4+ T cells to survive infections with many distinct intracellular pathogens. Loss of CD4+ T cells leads to susceptibility to infections caused by Chlamydia trachomatis, Mycobacterium tuberculosis and others. Antigen presenting cells (APCs) are at the interface of invading pathogens and adaptive immunity, sensing the infected environment and signaling directly to CD4+ T cells. The expression of Major Histocompatibility Complex II (MHCII) on APCs is critical for CD4+ T cells to detect their cognate antigens and activate an appropriate host response. Many pathogens manipulate the expression of MHCII to evade protective immunity. While complete loss of MHCII prevents CD4+ T cell function, how modulation of MHCII levels during infection impacts the effectiveness of pathogen-specific CD4+ T cell responses remains unknown. We hypothesize that MHCII expression levels are an important factor in the activation and effector function of antigen-specific CD4+ T cells. By understanding how MHCII is regulated at a global scale we will be positioned to develop interventions that modulate MHCII expression in order to overcome pathogen virulence strategies aimed at blunting protective host responses. We performed a genome-wide CRISPR-Cas9 screen to identify regulators of MHCII surface expression in activated APCs. We found over 200 genes that significantly alter surface MHCII expression with no described function in MHCII regulation. Here, using CRISPR-Cas9 mediated gain-of-function and loss-of-function approaches we will systemically identify priority candidates that enhance or inhibit MHCII surface expression and begin to dissect their mechanisms of MHCII control. We will next determine how modulation of MHCII expression directly alters the pathogen-specific CD4+ T cell response during genital infections with Chlamydia muridarum. We are investigating Chlamydia because it is an important pathogen clinically as the number one bacterial sexually transmitted infection in the United States. Chlamydia causes severe pathologies that lead to infertility, ectopic pregnancy and pelvic inflammatory disease. We will track the Chlamydia-specific CD4+ T cell response using immunological tools that do not exist for most pathogens. In combination with our CRISPR approaches we are uniquely positioned to directly examine how MHCII expression impacts the Chlamydia-specific CD4+ T cell response ex vivo and in vivo. We will determine how enhanced or reduced MHCII expression on APCs alters a range of CD4+ T cell phenotypes including activation, proliferation, effector function, protection and the development of genital tract pathologies. Our focused studies on the role of MHCII expression during C. trachomatis infection will broadly identify regulators of MHCII that can then be explored in a range of disease states where CD4+ T cell activation is required and MHCII is manipulated including other intracellular pathogens or cancers.