KSHV latent infection replication

 描述(由申请人提供)：Kaposi肉瘤(KS)相关疱疹病毒(KSHV或HHV8)是KS、原发渗出性淋巴瘤(PEL)和多中心Castleman病的病原体。KSHV通过唾液在人际间传播。KSHV肿瘤主要发生在艾滋病和其他免疫受损状态。目前还没有针对这些肿瘤的特效治疗方法。KS是主要的艾滋病恶性肿瘤，在撒哈拉以南非洲流行，那里与艾滋病毒混合感染很常见。KS常累及口腔和内脏器官。KSHV潜伏感染肿瘤细胞。在潜伏感染期间，病毒基因组以多拷贝、染色体外、环状上体(质粒)的形式存在。为了维持细胞的增殖，上染色体必须在每个细胞周期中进行复制，并有效地分离到子核。潜伏期相关核抗原(LANA)介导了KSHV EB病毒的持续存在，是病毒潜伏期和KSHV在增殖细胞中存活所必需的。附着体的维持由两个部分组成：复制KSHV DNA，以及将复制的附着体分离到子细胞核。拉娜负责这两项职能。为了复制KSHV DNA，LANA与病毒末端重复序列(TrDNA)结合。由于KSHV DNA复制蛋白在潜伏感染期间不表达，因此LANA负责招募宿主细胞复制机制。虽然LANA与多种细胞复制因子相互作用，但对LANA介导复制的潜在机制知之甚少。我们最近发现，LANA招募DNA聚合酶钳位加载器来介导有效的复制和病毒的持久性。这种招募的丧失极大地抑制了LANA介导DNA复制的能力，并导致了病毒感染的丧失。这些发现表明，钳制加载是DNA复制中的一个限速步骤，与病毒的生存不相容。LANA对钳夹负载的增强使KSHV能够复制、持久和存活。这一应用的首要假设是，LANA对钳制加载器的招募是KSHV复制生物学的基本组成部分，是潜在治疗抑制的理想靶点。实验将验证这一假设。这项工作将评估LANA增强钳夹负荷的机制。实验将调查LANA-夹具装载机复合体的结构。此外，实验将针对LANA-钳位加载器复合体进行抑制。这项工作的预期结果是详细了解LANA作用于钳夹加载器的机制，并确认这种相互作用是治疗KSHV恶性肿瘤的潜在靶点。

 DESCRIPTION (provided by applicant): Kaposi's sarcoma (KS)-associated herpesvirus (KSHV or HHV8) is the causative agent of KS, primary effusion lymphoma (PEL) and multicentric Castleman's disease. KSHV is spread person to person through saliva. KSHV tumors occur primarily in AIDS and other immune compromised states. There are no available specific therapies for these tumors. KS is the leading AIDS malignancy and is epidemic in sub Saharan Africa where coinfection with HIV is common. KS often involves the oral cavity and visceral organs. KSHV latently infects tumor cells. During latent infection viral genomes persist as multiple copy, extrachromosomal, circular episomes (plasmids). To persist in proliferating cells, episomes must replicate with each cell cycle and efficiently segregate to daughter nuclei. The latency-associated nuclear antigen (LANA) mediates KSHV episome persistence and is strictly required for viral latency and KSHV survival in proliferating cells. Episome maintanence is comprised of two components: replication of KSHV DNA, and segregation of replicated episomes to daughter cell nuclei. LANA is responsible for both these functions. To replicate KSHV DNA, LANA binds viral terminal repeat (TR) DNA. Since no KSHV DNA replication proteins are expressed during latent infection, LANA is responsible for recruiting host cell replication machinery. Although LANA interacts with several cell replication factors, little is known regarding the underlying mechanisms through which LANA mediates replication. We recently found that LANA recruits the DNA polymerase clamp loader to mediate efficient replication and viral persistence. Loss of this recruitment greatly inhibited LANA's ability to mediate DNA replication and resulted in loss of virus infection. These findings suggested that clamp loading is a rate-limiting step in DNA replication that is incompatible with virus survival. LANA enhancement of clamp loading enables KSHV replication, persistence, and survival. The overarching hypothesis of this application is that LANA's recruitment of the clamp loader is a fundamental component of KSHV replication biology that is an ideal target for potential therapeutic inhibition. Experiments will test this hypothesis. Work will assess the mechanism by which LANA enhances clamp loading. Experiments will investigate the architecture of the LANA-clamp loader complex. In addition, experiments will target the LANA-clamp loader complex for inhibition. The expected outcome of this work is a detailed understanding of the mechanism through which LANA acts on the clamp loader and validation of this interaction as a potential therapeutic target for KSHV malignancy.