Oncogenesis By Jaagsiekte Sheep Retrovirus

描述（由申请人提供）：我们将继续研究绵羊肺腺瘤逆转录病毒（JSRV）的致癌转化，这是一种传染性肺癌（与人肺腺癌非常相似的绵羊肺癌（OPA））的病因。肿瘤发生的靶细胞是II型肺细胞和Clara细胞。JSRV的独特之处在于它的包膜基因也是一种致癌基因;它可以在培养中转化细胞并在动物中诱导肿瘤。与其他病毒癌蛋白一样，Env的多个结构域参与转化。跨膜蛋白（TM）的胞质尾区是必需的，但表面蛋白（SU）也参与其中。此外，JSRV编码一种调节活性rej，这是从未剪接的病毒mRNA表达蛋白质所必需的。Rej编码在env区域内，这增加了它也可能参与转化的可能性。本研究的目的有三：1）Env蛋白TM和rej的结构与功能分析。将通过溶液NMR确定TM的胞质尾区的结构，将研究TM的跨膜和胞外域的潜在作用，并将评价rej在转化中的潜在作用。2)参与转化的细胞蛋白质的鉴定。将测试通过酵母双杂交筛选鉴定的候选蛋白与JSRV转化的相关性，并通过串联亲和纯化（TAP）/蛋白质组学方法寻找额外的细胞结合蛋白。3)JSRV体外和体内转化。为了更接近地反映肺上皮细胞的体内肿瘤发生，还将测试原代大鼠和绵羊II型肺细胞（具有或不具有协同致癌基因）和鼠细支气管肺泡干细胞（BASC）的体外转化。将开发JSRV诱导的肺癌的转基因小鼠模型，并将在腺相关病毒（AAV）载体中测试野生型和突变型JSRV Env的致瘤性。这些实验将大大增加我们对这种独特的人类肺癌模型的理解，并且所揭示的致癌途径可能阐明人类肺癌中的类似途径。 敷设总结：这项提案将研究一种引起传染性肺癌（OPA）的绵羊病毒（JSRV），这种肺癌与人类肺癌的一种形式非常相似。这些发现可能会导致更好地了解人类肺癌，从而改善治疗和预防。

DESCRIPTION (provided by applicant): We will continue studies on oncogenic transformation by jaagsiekte sheep retrovirus (JSRV), the cause of a transmissible lung cancer, ovine pulmonary carcinoma (OPA) that closely resembles human lung adenocarcinoma. The target cells for oncogenesis are type II pneumocytes and Clara cells. JSRV is unique in that its envelope gene also is an oncogene; it can transform cells in culture and induce tumors in animals. Like other viral onco-proteins, multiple domains of Env are involved in transformation. The cytoplasmic tail of the transmembrane protein (TM) is essential, but the surface protein (SU) also participates. In addition, JSRV encodes a regulatory activity rej, necessary for expression of proteins from unspliced viral mRNA. Rej is encoded within the env region, which raises the possibility that it may also be involved in transformation. The proposed research has three aims: 1) Structure-function analysis of the Env protein TM, and of rej. The structure of the cytoplasmic tail of TM will be determined by solution NMR, the potential roles of the membrane-spanning and ectodomains of TM will be investigated, and a potential role for rej in transformation will be evaluated. 2) Identification of cellular proteins involved in transformation. Candidate proteins identified by yeast two-hybrid screening will be tested for relevance to JSRV transformation, and additional cellular binding proteins will be sought by tandem affinity purification (TAP)/proteomic approaches. 3) JSRV transformation in vitro and in vivo. To more closely mirror in vivo oncogenesis of lung epithelial cells, in vitro transformation of primary rat and ovine type II pneumocytes (with or without cooperating oncogenes), and of murine bronchiole-alveolar stem cells (BASCs) will also be tested. A transgenic mouse model for JSRV-induced lung cancer will be developed, and wild-type and mutant JSRV Env's will be tested for tumorigenicity in an adeno-associated virus (AAV) vector. These experiments will substantially increase our understanding of this unique model for human lung cancer, and the oncogenic pathways uncovered may elucidate similar ones in human lung cancer. Lay summary: This proposal will study a virus of sheep (JSRV) that causes a transmissible lung cancer (OPA), very similar to one form of human lung cancer. The discoveries may lead to a better understanding of human lung cancer, which may result in improved therapies and prevention.