Genetic and Biochemical Studies of KSHV LANA

KSHV LANA 的遗传和生化研究

• 批准号: 10599894
• 负责人: Kenneth M Kaye
• 金额: $ 65.55万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10599894
• 关键词: Acquired Immunodeficiency Syndrome; Africa South of the Sahara; Animal Model; Binding; Biochemical; Biological; Biological Assay; C-terminal; Cell Nucleus; Cell Proliferation; Cell Survival; Cells; DNA; DNA Binding Domain; DNA biosynthesis; Daughter; Development; Disease; Elements; Epidemic; Episome; Etiology; Genes; Genetic; Genome; Glycogen Synthase Kinase 3; Herpesviridae Infections; Histone H2A; Human; Human Herpesvirus 8; Immunocompromised Host; Investigation; Kaposi Sarcoma; Link; Lysine; Maintenance; Malignant Neoplasms; Mediating; Mitotic Chromosome; Molecular; Multicentric Angiofollicular Lymphoid Hyperplasia; Mus; N-terminal; Nucleosomes; Oncoproteins; Oral cavity; Organ; Phosphorylation Site; Plasmids; Proliferating; Proteins; Reader; Repetitive Sequence; Role; Sequence Deletion; Sequence Homologs; Specificity; Surface; TP53 gene; Terminal Repeat Sequences; Viral; Viral Genome; Virus; Virus Replication; Visceral; Work; antigen binding; daughter cell; experimental study; gamma-2 herpesvirus; gammaherpesvirus; genetic approach; in vivo; insight; interest; latency-associated nuclear antigen; neoplastic cell; novel; novel strategies; prevent; primary effusion lymphoma; segregation; tumor; ubiquitin-protein ligase; viral DNA

Kaposi's sarcoma (KS)-associated herpesvirus (KSHV or HHV8) is the causative agent of KS, primary effusion lymphoma (PEL) and multicentric Castleman's disease. These tumors occur in AIDS and other immunocompromised states. There are no specific therapies for these diseases. KS is the leading AIDS malignancy, and is epidemic in sub Saharan Africa. KS often involves the oral cavity and can disseminate to visceral organs. KSHV latently infects tumor cells and viral genomes persist as circular, multiple copy, extrachromosomal, episomes (plasmids). In order to persist in proliferating cells, episomes must first replicate and then efficiently segregate to daughter nuclei. Tumor cell viability is dependent on latent KSHV infection. The latency-associated nuclear antigen (LANA) mediates KSHV episome persistence and this function is essential for virus survival. Therefore, LANA function is of fundamental importance. LANA represents the Achilles heel of KSHV as its disruption is lethal to the virus. LANA is necessary and sufficient for KSHV episome maintenance in the absence of other virus genes. Episome persistence is comprised of two components: 1) replication of viral DNA and 2) segregation of the replicated episomes to progeny cell nuclei. LANA mediates both these functions. C-terminal LANA binds specific sequence in KSHV terminal repeat (TR) DNA to mediate DNA replication. LANA tethers TR DNA to mitotic chromosomes to segregate genomes to daughter nuclei. To form a molecular tether, the C-LANA DNA binding domain (DBD) binds TR DNA and N-LANA simultaneously binds histones H2A/H2B at the folded portion of the nucleosome. In addition to N- and C-LANA, internal LANA sequence is also critical for episome persistence. We have discovered two distinct internal LANA regions, one comprised of unique sequence, and one within repetitive sequence, that are critical for LANA mediated episome persistence. The unique sequence contains three distinct functional motifs, whereas the repetitive region is homologous to a host cell “reader” sequence. Both of these regions will be investigated in this work. This work will use a detailed, in depth approach to investigate each of the recently identified LANA functional sequences. Experiments will use a rigorously developed panel of assays to identify the functional motif and determine the mechanism through which the unique sequence mediates episome persistence. The repetitive LANA effector sequence homologous to host “reader” sequence will be investigated to determine the mechanism through which it exerts its function. This work will provide important insight into LANA’s fundamental function of episome maintenance, and may lead to novel strategies to prevent or treat KSHV associated malignancies.

Kaposi的肉瘤（KS）相关疱疹病毒（KSHV或HHV8）是KS的病因，主要 出现淋巴瘤（PEL）和多中心Castleman病。这些肿瘤发生在艾滋病和其他 免疫功能低下的状态。这些疾病没有具体的疗法。 KS是领先的艾滋病 恶性肿瘤，在撒哈拉以南非洲是流行病。 KS通常涉及口腔，并且可以传播到 内脏器官。 KSHV潜在感染的肿瘤细胞和病毒基因组持续为圆形，多拷贝， 外染色体，个性（质粒）。为了持续存在增殖的细胞，必须首先复制插花 然后有效地分离为核里。肿瘤细胞活力取决于潜在的KSHV感染。 潜伏相关的核抗原（LANA）介导了KSHV发作的持久性和此功能 对于病毒生存至关重要。因此，LANA功能至关重要。拉娜代表 KSHV的阿喀琉斯高跟鞋对病毒的破坏是致命的。拉娜（Lana）是必要的，足以满足KSHV 在没有其他病毒基因的情况下，发作维持。 沿着持久性由两个组成部分组成：1）病毒DNA的复制和2）隔离 复制的偶发到后代细胞核。 Lana介导了这两个功能。 C末端LANA结合 KSHV末端重复（TR）DNA中的特定序列介导DNA复制。 Lana Tethers tr dna至 有丝分裂染色体将基因组隔离到女儿核。为了形成分子系绳，C-LANA DNA 结合结构域（DBD）结合TR DNA和N-LANA仅在折叠处结合组蛋白H2A/H2B 核小体的一部分。除N-和C-LANA外，内部LANA序列对于发作也很重要 持久性。 我们发现了两个不同的内部拉娜地区，一个是独特的序列，一个 在重复序列中，这对于LANA中介发作持久性至关重要。独特的序列 包含三个不同的功能基序，而重复区域与宿主细胞“读取器”同源 顺序。这两个区域将在这项工作中进行调查。 这项工作将采用详细的深入方法来调查最近确定的LANA 功能序列。实验将使用严格开发的测定面板来识别功能 主题并确定独特序列介导情节持久性的机制。 重复的LANA效应子序列将研究与主机“读取器”序列同源，以确定 通过其导出功能的机制。这项工作将为Lana提供重要的见解 情节维护的基本功能，并可能导致预防或治疗KSHV的新型策略 相关的恶性肿瘤。