EPSTEIN-BARR VIRUS INDUCED GENOMIC INSTABILITY

这项研究的长期目标是了解分子 爱泼斯坦-巴尔病毒(EBV)致癌机制EBV是 与鼻咽癌、地方性Burkitt淋巴瘤相关 (Bl)和霍奇金淋巴瘤。我们认为与艾滋病相关的淋巴瘤 为EBV肿瘤发生提供新的见解，因为免疫抑制 揭开病毒固有的属性，这些属性可能是其核心 病理生物学。11a缺陷EB病毒在艾滋病患者中的鉴定 破坏病毒潜伏期的基因组，2/a独特的EBV复制损伤 (口腔毛状白斑)和31种溶解抗原在艾滋病相关淋巴瘤中的表达 所有这些都证实了这一概念的正确性。鉴于6种核抗原和 3膜蛋白在淋巴母细胞潜伏感染中的表达 被认为有助于EBV诱导的淋巴增殖，这是一个作用 对于复制的循环蛋白，还没有被认识到。我们现在 假设，作为病毒复制过程的一部分，这 嗜淋巴疱疹病毒激活相关部位特异性重组酶 免疫球蛋白和T细胞受体基因的多样化。生物学 后果包括产生有缺陷的EBV基因组和新的 致病潜力，病毒整合到宿主基因组，以及 基因组不稳定性的介绍。具体目标是测试这一点 假设是：II确定诱导表达的EBV基因产物 重组酶激活基因L和2(RAG1&2)；2)疗效评价 V(D)J重组酶对EBV基因组组织的影响；3)阐明致病性 EB病毒相关RAG表达的后果。 EB病毒诱导病毒DNA所需基因表达RAG 复制(EBNA1和BZLF1)将在 RNA、蛋白质和功能水平(目标1)。EB病毒与细胞DNA 克隆的BL细胞整合位点的断裂点将被测序 以确定V(D)J识别信号序列(RS)的存在。EB病毒 将对分子间和基因内重排进行测序 V(D)J重组酶连接的特征：存在七聚体/纳米RSS， 核苷酸的连接损失和加成(目标2)。最后，利用艾滋病-- 与淋巴瘤相关的临床相关性将通过平行进行 单位长度和缺陷EBV DNA的整合分析(目标3)。 病毒DNA复制过程中V(D)J重组酶的异常表达 提供慢性病毒中EBV重新激活的第一个迹象 携带者在致癌过程中直接发挥作用，并将提供 在严重免疫抑制期间进行抗病毒干预的理由 爱滋病患者。

The long term goal of this research is to understand the molecular mechanisms by which the Epstein-Barr virus (EBV) causes cancer. EBV is associated with nasopharyngeal carcinoma, endemic Burkitt's lymphoma (BL), and Hodgkin's lymphoma. We contend that AIDS-related lymphomas offer novel insights into EBV oncogenesis because immunosuppression unmasks properties intrinsic to the virus that may be central to its pathobiology. Identification in AIDS patients of 11 a defective EBV genome that disrupts viral latency, 2/ a unique EBV replicative lesion (oral hairy leukoplakia), and 31 lytic antigens in AIDS-related lymphomas all confirm the validity of this notion. Whereas 6 nuclear antigens and 3 membrane proteins expressed in latent infection of lymphoblastoid cells are recognized as contributing to EBV-induced lymphoproliferation, a role for replicative cycle proteins has not been appreciated. We now hypothesize that, as part of the viral replicative process, this lymphotropic herpesvirus activates site-specific recombinases involved in diversification of immunoglobulin and T cell receptor genes. Biologic consequences include generation of defective EBV genomes with novel pathogenic potential, virus integration into the host genome, and the introduction of genomic instability. Specific aims to test this hypothesis are: II determine the EBV gene product that induces expression of recombinase activating genes l and 2 (RAG1&2); 2) evaluate the effect of V(D)J recombinase on EBV genomic organization; 3) elucidate pathogenic consequences of EBV-associated RAG expression. EBV induction of RAG expression by genes required for viral DNA replication (EBNA1 and BZLF1) will be examined in transfection assays at the RNA, protein and functional levels (aim 1). EBV and cellular DNA breakpoints from cloned integration sites in BL cells will be sequenced to determine presence of V(D)J recognition signal sequences (RSS). EBV intermolecular and intragenic rearrangements will be sequenced for hallmarks of V(D)J recombinase joining: presence of heptamer/nanomer RSS, junctional loss and addition of nucleotides (aim 2). Finally, using AIDS- related lymphomas, clinical correlations will be made by parallel analyses for integration of unit length and defective EBV DNA (aim 3). Aberrant V(D)J recombinase expression during viral DNA replication would provide the first indication that EBV reactivation in chronic virus carriers has a direct role in the oncogenic process and would provide a rationale for antiviral interventions during the severe immunosuppression of AIDS.