Immune Evasion Mechanisms of KSHV Complement and Fc-Receptor Proteins

项目摘要 卡波西肉瘤相关疱疹病毒是卡波西肉瘤的病原体，卡波西肉瘤是一种出现在 皮肤或粘膜表面的肿瘤和两种类型的B细胞淋巴瘤，多中心性Castleman病 和原发性渗出性淋巴瘤。因为没有疫苗或治疗方法可以成功地预防或 根除KSHV，部分原因是它能够逃避宿主的免疫防御，它仍然是一个重要的公众 健康负担。虽然B细胞淋巴瘤很罕见，但超过44,000例新的卡波西肉瘤病例是 每年在全球范围内报告。因此，迫切需要阐明KSHV逃逸的机制。 人类的免疫系统。疱疹病毒，包括KSHV，在数百万年的时间里与它们的宿主共同进化， 劫持主持人的监管因素是为了伪装自己，规避东道主广泛的防御措施。 值得注意的是，像其他疱疹病毒一样，KSHV也获得了一种补体控制蛋白(KCP)，发现它可以调节 宿主补体介导的免疫防御。不幸的是，在生物相关背景下对KCP的研究 由于缺乏产生KCP-Null KSHV和测试KCP-Null KSHV效果的系统而受到阻碍 病毒感染细胞上的补体相互作用。为了解决这些不足，我们产生了重组KSHV 利用新建立的细菌人工染色体缺失KCP(rKSHVΔKCP) 技术，以及新的抗KCP抗体来表征突变病毒。在被指导期间(K99) 在此阶段，我将使用这些资源来描述KCP在逃避补体控制和感染方面的作用。 除了KCP，KSHV还编码一种糖蛋白K1，它具有免疫球蛋白结构域，具有 片断结晶性受体(FCR)发挥功能。其他疱疹病毒中类似的结构域已被证明起作用 作为宿主抗病毒抗体的干扰受体，通常结合存在于免疫细胞上的FCR；然而， K1在Fc介导的KSHV免疫逃避中的作用尚未被研究。因此，在接近尾声时 K99阶段，我建议产生和表征抗K1抗体，并表征我们最近的 获得了稳定的rKSHVΔK1产生细胞系。这些资源将在独立(R00)阶段使用 以表征K1的FC型相互作用和结合能力。我还将研究通过什么机制 K1使用一种新的报告实验干扰宿主FCR的激活。最后，我将评估抗K1抗体的疗效 抗体在阻断Fc介导的免疫逃避中的作用。因此，本文提出的研究将提供更好的 了解KCP和K1在宿主免疫逃避中的作用。这项工作将极大地促进我国经济的发展。 预防和控制KSHV感染和致病的有效策略。此外，这项研究和 为K99阶段提出的培训将提供病毒基因组学/蛋白质组学方面的经验和知识， 免疫生物学和病毒肿瘤学，以及大量可公布的数据和初步数据 许多未来的学习，这将提供一个坚实的基础，以建立我的独立研究事业。

PROJECT ABSTRACT Kaposi sarcoma-associated herpesvirus (KSHV) is the causal agent of Kaposi sarcoma, a cancer that appears as tumors on the skin or mucosal surfaces, and two types of B-cell lymphomas, multicentric Castleman disease and primary effusion lymphoma. As there are no vaccines or therapeutic treatments to successfully prevent or eradicate KSHV, in part due to its ability to evade its host’s immune defenses, it remains a significant public health burden. Although B-cell lymphomas are rare, more than 44,000 new cases of Kaposi sarcoma are reported globally each year. Thus, there is a critical need to elucidate the mechanisms by which KSHV escapes the human immune system. Herpesviruses, including KSHV, co-evolved with their hosts over millions of years, hijacking host regulatory factors to camouflage themselves and evade a wide range of host defense measures. Notably, like other herpesviruses, KSHV has acquired a complement control protein (KCP) found to modulate the host complement-mediated immune defense. Unfortunately, studies of KCP in a biologically relevant setting have been hindered by the absence of a system to produce KCP-null KSHV and test the effects of KCP– complement interactions on virus-infected cells. To address these deficiencies, we generated recombinant KSHV (rKSHV) lacking KCP (rKSHVΔKCP) using newly established KSHV bacterial artificial chromosome (BAC) technology, as well as novel anti-KCP antibodies to characterize the mutant virus. During the mentored (K99) phase, I will use these resources to characterize the role of KCP in evading complement control and infection. In addition to KCP, KSHV encodes a glycoprotein, K1, that has acquired immunoglobulin structure domains with fragment crystallizable receptor (FcR) functions. Similar domains in other herpesviruses have been shown to act as distractor receptors for host antiviral antibodies that typically bind FcRs present on immune cells; however, the role of K1 in Fc-mediated immune evasion by KSHV has not been examined. Therefore, toward the end of the K99 phase, I propose to generate and characterize anti-K1 antibodies and characterize our recently generated stable rKSHVΔK1 producer cell lines. These resources will be utilized in the independent (R00) phase to characterize the Fc-type interactions and binding capabilities of K1. I will also study the mechanism by which K1 interferes with host FcR activation using a novel reporter assay. Finally, I will evaluate the efficacy of anti-K1 antibodies in blocking Fc-mediated immune evasion. Thus, the studies proposed herein will provide a better understanding of the role of KCP and K1 in host immune evasion. This work will greatly enhance the development of effective strategies to prevent and control KSHV infection and pathogenesis. Furthermore, the research and training proposed for the K99 phase will provide experience and knowledge in viral genomics/proteomics, immunobiology, and viral oncology, as well as a significant body of publishable data and preliminary data for many future studies, which will provide a solid foundation upon which to build my independent research career.