Functional Characterization of WTX in Renal Development

描述(由申请人提供)： 肾母细胞瘤是一种起源于胚胎肾脏前体的多潜能肿瘤，是儿童最常见的肾脏肿瘤，也是儿童第四常见的恶性肿瘤。Wilms肿瘤中第一个失活的基因是肿瘤抑制基因WT1，其功能特征强调了这些肿瘤与肾脏发育之间的联系。WT1对小鼠的肾脏发育是必不可少的，它也是WAGR、Denys-Drash和Frasier综合征中生殖道缺陷的原因。然而，WT1在仅5-10%的散发性肾母细胞瘤中失活。已经描述了β-连环蛋白、胰岛素样生长因子2(IGF2)和P53的变化，但没有已知的特定基因缺陷可以解释大多数病例。KO8奖的候选人Miguel Rivera博士已经发现并克隆了一种新的肿瘤抑制基因WTX，该基因在30%的散发性肾母细胞瘤病例中失活。值得注意的是，WTX在肾脏发育过程中以高度调控的方式表达，部分与WT1重叠。因此，通过与WT1的类比，WTX有望成为肾母细胞瘤发生和肾脏发育的关键基因。候选人现在建议在这些初步结果的基础上，描述WTX在肾脏发育中的功能特性。目的1：研究WTX的生化性质。慢病毒载体将被用来在接近生理水平表达WTX，并确定与其他蛋白质的关联。目的2：明确WTX在肾脏发育组织培养模型中的作用。WTX上调和下调的功能后果将在早期肾脏发育的细胞培养模型(RSTEM细胞-多能肾细胞系)和完全分化的肾上皮细胞(小鼠足细胞系)中进行评估。目的3：研究条件失活WTX对小鼠肾脏的影响。与Nabeel Bardeesy博士的实验室合作，正在培育一种有条件的WTX基因敲除小鼠。作为这项合作的一部分，候选人将分析这只小鼠在肾脏发育中的表型。预计通过加深我们对早期肾脏发育的了解，该项目将在儿童和成人肾脏疾病、肾组织修复和肾癌方面有公共卫生应用。

DESCRIPTION (provided by applicant): Wilms tumor is a pluripotent tumor which arises from embryonic renal precursors and is the most common pediatric kidney tumor and the fourth most common childhood malignancy. The first gene shown to be inactivated in Wilms tumor was the tumor suppressor WT1 and its functional characterization has emphasized the connection between these tumors and renal development. WT1 is essential for kidney development in mice and it is also responsible for genitourinary defects in the WAGR, Denys-Drash and Frasier syndromes. WT1, however, is inactivated in only 5-10% of sporadic Wilms' tumors. Alterations in beta-catenin, insulin-like growth factor 2 (IGF2) and p53 have been described but there are no known specific genetic defects that account for the majority of cases. The candidate for this KO8 award, Dr. Miguel Rivera, has identified and cloned a novel tumor suppressor gene, WTX, which is inactivated in 30% of sporadic Wilms tumor cases. Remarkably, WTX is expressed in a highly regulated pattern that partially overlaps WT1 during kidney development. Thus, by analogy to WT1, WTX is expected be a critical gene in both Wilms tumorigenesis and kidney development. The candidate now proposes to build on these initial results and characterize the functional properties of WTX in renal development. Aim 1: To characterize the biochemical properties of WTX. Lentiviral vectors will be used to express WTX at near physiologic levels and to determine associations with other proteins. Aim 2: To define the role of WTX in tissue culture models of renal development. The functional consequences of WTX upregulation and downregulation will be assessed in cell culture models of early renal development (RSTEM cells - a pluripotent renal cell line ) and fully differentiated kidney epithelia (a mouse podocyte cell line). Aim 3: To characterize the consequences of conditional WTX inactivation in the mouse kidney. A conditional WTX knockout mouse is being generated in collaboration with the laboratory of Dr. Nabeel Bardeesy. As part of this collaboration, the candidate will analyze the phenotype of this mouse in kidney development. It is anticipated that, by furthering our knowledge of early kidney development, this project will have public health applications in pediatric and adult kidney disease, kidney tissue repair and kidney cancer.