Products of the Transforming Genes of Polyomavirus

小鼠多瘤病毒为肿瘤转化提供了有价值的模型。这种病毒会在动物身上引起广泛的肿瘤。这意味着，在实验室开发的铅可以很容易地在宿主中进行测试。对多发性瘤的研究已经反复为细胞生长调节的基本机制提供了洞察力。酪氨酸激酶信号和磷脂酰肌醇3-激酶(PI3K)信号是多发性肿瘤研究中发现的两个基本机制。中T抗原(MT)是多瘤病毒的主要转化蛋白。对于多发性瘤的细胞转化来说，它总是必要的，通常也是足够的。这个子项目将根据前一个项目期间开发的线索，扩展我们对MT及其信令的知识。有4个具体目标。1)我们已经证明MT能激活小的GTP酶ral。我们将研究MT是如何激活ral的。我们将测试ral在MT转化中的作用。由于Ral在包括分泌在内的贩运过程中发挥作用，我们将在那里测试MT的功能。2)MT诱导骨桥蛋白，这是一种被认为与转化和转移有关的分泌性蛋白。我们将确定MT如何调节骨桥蛋白。我们将确定骨桥蛋白的作用 在MT转化中，通过检查肿瘤的轮廓。利用乳腺癌和血管瘤模型，我们将测试骨桥蛋白在转移和细胞募集中的作用。我们将使用核磁共振来启动MT的结构分析，以确定MT与下游靶标如Shc和PI3K相互作用的结构。然后我们将绘制MT和PI3K或Shc的P85之间的相互作用面。最后，有强有力的遗传证据表明，存在未发现的MT结合蛋白。我们将使用串联亲和纯化和质谱法对它们进行鉴定。我们将确定它们相互作用所需的MT序列。我们将在MT检测中测试它们的功能以进行转化。

Murine polyomavirus provides a valuable model for neoplastic transformation. The virus causes a broad spectrum of tumors in animals. This means that leads developed in the laboratory can be readily tested in the host. Studies on polyoma have repeatedly provided insight into basic mechanisms of cellular growth regulation. Tyrosine kinase signaling and phosphatidylinositol 3-kinase (PI3K) signaling are two fundamental mechanisms uncovered from studies on polyoma. Middle T antigen (MT) is the major transforming protein of polyomavirus. It is always necessary, and often sufficient, for cellular transformation by polyoma. This subproject will extend our knowledge of MT and its signaling based on leads developed in the previous project period. There are 4 specific aims. 1) We have shown that MT activates the small GTPase Ral. We will examine how MT activates Ral. We will test the role of Ral in MT transformation. Since Ral functions in trafficking processes including secretion, we will test MT function there. 2) MT induces osteopontin, a secreted protein thought to be linked to transformation and metastasis. We will determine how MT regulates osteopontin. We will determine the role of osteopontin in MT transformation by examining tumor profiles. Using breast cancer and hemangioma models, we will test the role of osteopontin in metastasis and in cell recruitment. We will initiate structural analysis of MT using NMR to determine the structure of the portion of MT that interacts with downstream targets such as Shc and PI3K. We will then map the interaction surface between MT and p85 of PI3K or Shc. Finally, there is strong genetic evidence suggesting that there are undiscovered MT binding proteins. We will use tandem affinity purification and mass spectrometry to identify them. We will determine the MT sequences required for their interaction. We will test their function in MT assays for transformation.