HPV AND ANTIGEN PRESENTING CELLS

项目摘要 人乳头瘤病毒（HPV）是常见的性传播病毒，其感染是一个重要的问题。 给人民带来健康负担。高危HPV基因型的持续感染导致 多种人类癌症，最突出的是宫颈癌、肛门癌和头颈部癌症的亚型。HPV 通过逃避免疫系统在宿主体内建立持续感染。HPV感染的一种方式 完成这一点是通过操纵朗格汉斯细胞（LC），这是抗原呈递细胞的网站 负责启动针对上皮入侵病毒的免疫反应。的 HPV通过膜联蛋白A2异四聚体（A2 t）与LC的相互作用和随后的抑制性信号传导 事件充当停止LC激活的制动机制，导致LC成熟不完全， 预防有效的抗病毒T细胞免疫应答。我们的长期目标是了解HPV如何 操纵LC激活和抗原处理以逃避免疫系统，并测试新的治疗方法。 HPV相关疾病。AIM 1将专注于使用“刹车关闭，油门踏板下降”的效果 用于激活暴露于HPV的LC的组合方法。这将通过细胞表面标志物进行研究 以及细胞因子分析和功能性免疫测定。AIM 2将研究信号通路， 与A2 t介导的LC免疫功能抑制相关的A2 t相互作用伴侣， HPV使用生物化学和细胞生物学方法。这将通过广泛的询问来分析， 磷蛋白活化和细胞信号网络，抗原呈递细胞特异性分析基因阵列， 以及新开发的单分子下拉（SimPull）测定。AIM 3将评估A2 t的贡献 LC Birbeck颗粒的结构、功能和在HPV抗原加工中的作用。这将通过以下方式进行调查： LC中Birbeck颗粒组分免疫电镜分析，A2 t蛋白评估 敲低对Birbeck颗粒形成的影响，以及阐明蛋白敲低对Birbeck颗粒形成的影响。 LC正确加工HPV衍生肽并将其呈递给HLA匹配的T细胞克隆的能力。此外，本发明还 这些目标将利用创新的皮肤外植体和三维培养系统来验证所提出的HPV 16-A2 t 相互作用对于体内LC生物学是重要的。总的来说，拟议的研究将深入了解 基础LC生物学和功能。此外，我们将研究一种新的双管齐下的方法的影响， 防止LC中的负信号事件，同时提供正刺激以激活LC 作为治疗HPV感染的新策略，从而预防HPV相关癌症。

Project Summary Human papillomaviruses (HPV) are common sexually transmitted viruses and their infections are a significant health burden for the population. Persistent infection with high-risk HPV genotypes result in the development of a variety of human cancers, most prominently cervical, anal, and subtypes of head and neck cancers. HPV establishes persistent infection within the host through evasion of the immune system. One of the ways HPV accomplishes this is by manipulating Langerhans cells (LC), which are the antigen presenting cells at the site of infection that are responsible for initiating immune responses against epithelium invading viruses. The interaction of HPV with LC through the annexin A2 heterotetramer (A2t) and subsequent suppressive signaling events act as a braking mechanism that stops LC activation, resulting in incomplete LC maturation and prevention of an effective antiviral T cell immune response. Our long-term goals are to understand how HPV manipulates LC activation and antigen processing to evade the immune system and test new ways to treat HPV-associated diseases. AIM 1 will focus on the effects of utilizing a “brakes off, gas pedal down” combination approach for activating LC exposed to HPV. This will be investigated through cell surface marker and cytokine analysis and functional immunological assays. AIM 2 will investigate the signaling pathways and A2t interaction partners associated with A2t-mediated suppression of LC immune function that are hijacked by HPV using biochemical and cell biological approaches. This will be analyzed through broad interrogation of phosphoprotein activation and cell signaling networks, antigen presenting cell specific profiling gene arrays, and a newly developed Single Molecule Pulldown (SiMPull) assay. AIM 3 will evaluate the contribution of A2t to LC Birbeck granule structure, function, and role in HPV antigen processing. This will be investigated through immuno-electron microscopic analysis of Birbeck granule components in LC, assessment of A2t protein knockdown effects on Birbeck granule formation, and elucidation of the effects of protein knockdown on the ability of LC to properly process and present HPV-derived peptides to HLA-matched T cell clones. Additionally, these aims will utilize innovative skin explant and 3-D culture systems to validate that the proposed HPV16-A2t interactions are important for LC biology in vivo. Collectively the proposed studies will provide deep insight into basic LC biology and function. In addition we will examine the effects of a novel two-pronged approach focused on preventing negative signaling events in LC while simultaneously providing a positive stimulus to activate LC as a novel strategy to treat HPV infections, thereby preventing HPV-associated cancers.