Products of the Transforming Genes of Polyomavirus

描述（由申请方提供）：鼠多瘤病毒为肿瘤转化提供了重要模型。这种病毒能引起多种肿瘤。 实验室得出的结论可以很容易地在动物身上进行测试。 对多瘤的研究反复提供了对细胞生长调节的基本机制的见解。 酪氨酸激酶和磷脂酰肌醇3-激酶（PI 3 K）信号传导是从多瘤研究中发现的两个基本机制。 转化由三种病毒基因产物的作用引起：大T（LT）、中T（MT）和小T（ST）抗原。 有四个具体的目标，将提供新的见解，他们是如何工作的：1）第一个具体的目标涉及大T。 将确定由不同的大T通路引起的宿主细胞RNA表达模式。 LT调控含CREB/ATF位点基因的机制将得以建立。 由于乙酰化与转录调控有关，因此将建立LT上的乙酰化位点，并通过定点突变测试其功能。 最近的研究结果表明，LT导致G2/M期阻滞。 将进行实验以确定LT引起这种停滞的机制。 我们已经确定了两个新的LT结合伙伴，Pinl和Bubl，这是已知的G2/M的调节剂。 它们在LT功能中的作用将被探讨。 2)第二个具体目标涉及小T。 在多瘤和SV 40小T之间已经检测到功能上的重要差异。 表达谱分析将用于确定它们之间差异的程度和基础。 新的多瘤小T合作伙伴将寻求串联亲和纯化和质谱。蛋白磷酸酶2A是一个关键的小T目标。将进行实验以区分小T靶向PP 2A的不同机制。 3)人乳腺上皮细胞提供了一种有用的人类癌症模型。 第三个具体目标是利用该模型来探讨中小企业的作用。 将确定转化所需的MT信号传导途径。 将在这些细胞中测试结合新伴侣的小T突变体。 4)我们的第四个具体目标涉及使用NMR的结构研究。 LT的N末端结构域（NT）的结构将被确定。 该结构域足以促进细胞生长，引起细胞凋亡和调节细胞RNA转录。 我们还将开展多瘤小T的结构测定研究。

DESCRIPTION (provided by applicant): Murine polyomavirus provides an important model for neoplastic transformation. The virus causes a broad spectrum of tumors. Conclusions reached in the laboratory can be readily tested in animals. Studies on polyoma have repeatedly provided insight into basic mechanisms of cellular growth regulation. Tyrosine kinase and phosphatidylinositol 3-kinase (PI3K) signaling are two fundamental mechanisms uncovered from studies on polyoma. Transformation results from the action of three viral gene products: large T (LT), middle T (MT) and small T (ST) antigens. There are four specific aims that will provide new insight into how they work: 1) the first specific aim concerns large T. Patterns of host cellular RNA expression that result from different large T pathways will be determined. The mechanism will be established by which LT regulates genes containing CREB/ATF sites. Because acetylation has been connected to transcriptional regulation, the sites of acetylation on LT will be established and their function tested by site-directed mutagenesis. Recent results show that LT causes G2/M arrest. Experiments will be carried out to determine the mechanism by which LT causes this arrest. We have identified two new LT-binding partners, Pinl and Bubl, which are known regulators of G2/M. Their role in LT function will be probed. 2) The second specific aim concerns small T. Important differences in function have been detected between polyoma and SV40 small T. Expression profiling will be used to identify the extent of, and the basis for, the differences between them. New polyoma small T partners will be sought by tandem affinity purification and mass spec. Protein phosphatase 2A is a critical small T target. Experiments will be performed to distinguish between different mechanisms by which small T targets PP2A. 3) Human mammary epithelial cells provide a useful model of human cancer. The third specific aim will use the model to probe the function of middle T and small T. The MT signaling pathways required for transformation will be determined. Small T mutant in binding novel partners will be tested in these cells. 4) Our fourth specific aim involves structural studies using NMR. The structure of the N-terminal domain (NT) of LT will be determined. This domain is sufficient to promote cell growth, to cause apoptosis and to regulate cellular RNA transcription. We will also initiate studies to carry out structure determination of polyoma small T.