Genetic Analysis of EBV's Immortalizing Gene, LMP-1

描述（由申请人提供）：EB病毒（EBV）是一种人类肿瘤 与几种淋巴瘤和癌有因果关系的病毒。它 有效地诱导和维持B淋巴细胞的增殖， 直到免疫反应可以限制这种感染。EBV的 对受感染细胞的增殖作用可能是其致病性的基础。一 在引起这些增殖效应的少数EBV基因中， 膜蛋白-1（LMP-1）。LMP-1存在于质膜上， 羧基末端结构域与几种细胞信号分子相关 其也被TNF和CD 40受体结合。LMP-1的信号结构域具有 在没有其他病毒基因的细胞中进行了广泛的研究。它有 也在重组EBV中进行了研究，由于技术原因， 只有定性的发现。我们发明了一种方法 重组EBV，称为“maxiEBV”，具有高滴度，使我们能够评估 在转化测定中定量测定LMP-1的单个元素。我们 假设LMP-1多种活性有助于其有效的 支持EBV诱导和维持细胞增殖的信号传导 被感染的细胞我们将用Aim中的maxi-EBV基因来测试这一假设 通过识别LMP直接靶向的细胞基因， l在Aim IV中具有独特的信号活动。LMP-1信号在表观 缺乏配体，其六个跨膜结构域支持其 聚集代替由配体结合贡献的聚集。它交通到 质膜和膜筏，在膜筏中发现它作为信号复合物 同样是在明显缺乏配体的情况下。我们假设LMP-1代表了 目前尚未定义的调节信号问题的解决方案。我们将 通过阐明LMP-1过渡到 质膜、聚集所需的部分以及时间 它移动到膜筏并形成信号的顺序和要求 目标二和目标三的复杂性。这些研究将得到确认和延长 将它们与maxi-EBV进行遗传学研究，以将它们置于EBV感染的背景下。 所有拟议的研究将量化已知的贡献， 活性和新的活动LMP-1，它提供了EBV的感染细胞。

DESCRIPTION (provided by applicant): Epstein-Barr Virus (EBV) is a human tumor virus causally associated with several lyrnphomas and carcinomas. It efficiently induces and maintains proliferation of the B-lymphocytes it naturally infects until the immune response can limit that infection. EBV's proliferative effects on infected cells likely underlie its pathogenicity. One of the few EBV genes contributing these proliferative effects is its latent membrane protein-1 (LMP-1). LMP-1 is found at the plasma membrane where its carboxy terminal domain associates with several cellular signaling molecules which are also bound by TNF and CD40 receptors. LMP-1's signaling domain has been studied extensively in cells in the absence of other viral genes. It has also been studied in recombinant EBV's which for technical reasons have allowed only qualitative findings. We have developed a means to isolate pure recombinant EBV's, termed "maxiEBV's," with high titers allowing us to assess quantitatively individual elements of LMP-1 in transformation assays. We hypothesize that multiple activities of LMP-1 contribute to its efficient signaling to support EBV's induction and maintenance of proliferation of infected cells. We shall test this hypothesis genetically with maxi-EBVs in Aim I and by identifying the cellular genes which are the immediate targets of LMP- l's distinct signaling activities in Aim IV. LMP-1 signals in the apparent absence of a ligand with its six membrane spanning domains supporting its aggregation in lieu of that contributed by ligand-binding. It traffics to the plasma membrane and to membrane rafts where it is found as a signaling complex again in the apparent absence of a ligand. We hypothesize that LMP-1 represents a currently undefined solution to the problem of regulating signaling. We shall test this hypothesis by elucidating the path by which LMP-1 transits to the plasma membrane, the moieties it requires for aggregation, and the temporal order and requirements for it to move to membrane rafts and form a signaling complex in Aims II and III. These studies will be confirmed and extended genetically with maxi-EBVs to place them in the context of infection with EBV. All of the proposed research will quantify the contributions of known activities and new activities of LMP-1 that it provides EBV's infected cells.