Characterization of Aquaporin 2 Interacting Proteins

描述（由申请人提供）：加压素（VP）是主要的抗利尿激素，参与调节哺乳动物肾脏的水重吸收。大量研究表明，VP通过将AQP2水通道从细胞质囊泡募集到集管主细胞的质膜来发挥作用。VP-AQP2信号通路的损伤导致尿崩症和一些心力衰竭患者的液体潴留。AQP2通过复杂的贩运途径受到调节。我们的假设是，AQP2的调节运输需要在细胞内易位、胞吐和内吞过程中直接和间接的蛋白质-蛋白质相互作用。拟议的研究分为三个具体目标，分别是鉴定和表征新的AQP2相互作用蛋白，以及研究这些新的蛋白相互作用对体外和体内AQP2运输的功能意义。该研究将利用多学科方法（分子生物学、细胞生物学、生物化学和免疫细胞化学）和强大的技术（酵母双杂交筛选、结构功能分析、基因转移）来鉴定和表征新的蛋白质-蛋白质相互作用及其生物功能。这些研究将为尚不清楚的AQP2转运的分子机制提供重要信息，并可能最终为尿浓度缺陷患者提供更好的治疗方法。

DESCRIPTION (provided by applicant): Vasopressin (VP) is the major antidiuretic hormone involved in the regulation of water reabsorption by mammalian kidney. Intensive studies have established that VP functions by recruiting the AQP2 water channel from cytoplasmic vesicles to the plasma membrane of collecting duct principal cells. The impairment of VP-AQP2 signaling pathways results in diabetes insipidus as well as fluid retention seen in some heart failure patients. AQP2 is regulated through complex trafficking pathways. Our hypothesis is that regulated trafficking of AQP2 requires direct and indirect protein-protein interactions during intracellular translocation, exocytosis as well as endocytosis. The proposed studies are divided into three specific aims addressing identification and characterization of novel AQP2 interacting proteins as well as investigation of the functional significance of these novel protein interactions on AQP2 trafficking in vitro and in vivo. The proposed study will utilize multidisciplinary approaches (molecular biology, cell biology, biochemistry and immunocytochemistry) and powerful technology (yeast two hybrid screen, structural functional analysis, gene transfer) to identify and characterize novel protein-protein interactions and their biologic functions. These studies will provide important information of the poorly understood molecular mechanisms underlying AQP2 trafficking, and may ultimately lead to better therapies for patients with defective urinary concentration.