Role of ubiquitylation in renal cancer

描述（由申请人提供）：肾癌占美国成人恶性肿瘤的3%，是癌症死亡的第六大原因。在这个应用中，我们建议研究导致肾癌最常见形式肾透明细胞癌（RCC）的致癌过程的分子基础。在RCC的发生和von Hippel-Lindau肿瘤抑制蛋白（pVHL）的功能丧失之间有一个很好的联系。众所周知，这种致癌作用可以通过破坏HIFa泛素化来介导。然而，在这个应用中，我们将扩展我们之前的工作，在RCC的发展中发现一种新的pVHL依赖的信号通路：RNA聚合酶II （RNAPII）的大亚基Rpb1的羟基化，以及pVHL肿瘤抑制复合物随后的泛素化。我们的工作假设是P1465羟基化和pvhl依赖的Rpb1泛素化提供了RNAPII复合体转录活性的微调，从而导致基因表达模式促进对环境变化（如氧化应激）的适当反应。我们认为，这种微调的缺失会导致RNAPII功能失调，从而导致肿瘤的发生。这项工作的总体目标是了解pvhl依赖性Rpb1羟基化和泛素化的机制，并评估这一过程的破坏在肿瘤发生中的作用。为了实现这一目标，我们将1)确定泛素和Rpb1中以p1465依赖方式参与pvhl依赖性Rpb1多泛素化的赖氨酸，以响应氧化应激；2)确定参与Rpb1羟基化的脯氨酸羟化酶；3)确定不发生pvhl依赖性羟化和泛素化的Rpb1突变体的致癌特性；4)评估Rpb1羟基化和泛素化在人肾细胞癌肿瘤中的地位，并与正常肾组织进行比较。早期诊断的肾细胞癌患者预后要好得多；然而，早期诊断在肾小细胞癌中并不常见，因为患者通常直到疾病晚期才出现症状。由于本研究探索了一种新的疾病发展的生化机制，它可以从两个方面改善RCC患者的预后：1)通过促进治疗RCC的新药的开发，2)通过发现新的遗传标记，可用于识别发生RCC的高危个体。

DESCRIPTION (provided by applicant): Renal cancer accounts for 3% of adult malignancies in the US, and is the 6th leading cause of cancer mortality. In this application, we propose to examine the molecular basis of the oncogenic process leading to the most common form of kidney cancer, renal clear cell carcinoma (RCC). There is a well established link between RCC oncogenesis and the loss of function of the von Hippel-Lindau tumor suppressor protein (pVHL). It is known that this oncogenic effect can be mediated through disruption of HIFa ubiquitylation. However, in this application, we will expand on our previous work implicating a novel pVHL-dependent signaling pathway in the development of RCC: the hydroxylation of the large subunit of RNA Polymerase II (RNAPII), Rpb1, and its subsequent ubiquitylation by the pVHL tumor suppressor complex. Our working hypothesis is that P1465 hydroxylation and pVHL-dependent ubiquitylation of Rpb1 provide fine tuning of the RNAPII complex's transcriptional activity, which results in patterns of gene expression that facilitate appropriate responses to environmental changes (such as oxidative stress). We propose that loss of that fine-tuning results in dysfunctional activity of RNAPII leading to oncogenesis. The overall goal of this work is to understand the mechanism of pVHL-dependent Rpb1 hydroxylation and ubiquitylation, and to assess the role that disruption of this process plays in oncogenesis. To achieve this, we will 1) identify the lysines within ubiquitin and Rpb1 that participate in pVHL-dependent polyubiquitylation of Rpb1 in a P1465-dependent manner in response to oxidative stress; 2) identify the prolyl hydroxylases involved in hydroxylation of Rpb1; 3) determine the oncogenic properties of Rpb1 mutants that do not undergo pVHL-dependent hydroxylation and ubiquitylation; and 4) assess the status of Rpb1 hydroxylation and ubiquitylation in human RCC tumors as compared to normal kidney tissue. The prognosis for patients with RCC is much better if their cancer is diagnosed in early stages; however, early diagnosis is uncommon in RCC because patients often do not experience symptoms until advanced stages of the disease. Because this research explores a novel biochemical mechanism for disease development, it could improve the prognosis for RCC patients in two ways: 1) by contributing to the development of new drugs to treat RCC, and 2) by identifying new genetic markers that could be used to identify individuals at risk for developing RCC.