DISCOVERY & CHARACTERIZATION OF PKD PROTEIN INTERACTIONS

ADPKD是一种全身性疾病，以累及多个器官的囊肿、脑动脉瘤和主动脉瘤、心脏瓣膜异常和肾衰竭为特征。导致两种最常见疾病的基因已经被确定。已经确定了导致最常见疾病形式的PKD 1基因。PKD 1基因产物可能是一种膜糖蛋白，被认为介导细胞-细胞或细胞-基质相互作用。PKD 2被预测具有6个跨膜结构域，并且与几种类型的离子通道具有适度的同源性。这两种蛋白质的细胞作用尚未阐明。这项建议汇集了来自多个学科的调查人员的集体专业知识，以解决“没有围墙的中心”的问题。一个项目是竞争性更新，旨在识别和表征人PKD 1的结合伴侣。PI已经显示PKD 1的C-末端可以结合PKD 2和使用多种体外和体内方法的不寻常的Db 1家族成员。该建议旨在确定预先观察到的相互作用的生物学相关性。它还将继续寻找与先前观察到的相互作用相关的PKD 1结合伙伴。它还将继续在胚胎鼠组织中寻找PKD 1结合伴侣，该组织是在PKD 1蛋白已知是必需的阶段获得的。另一个项目将使用模型实验生物果蝇（Drosophila melanogaster）来表征PKD 2在体内和体外的功能。拟议的研究采取了多学科的方法，使用遗传学，电生理学，细胞生物学，生物化学和分子方法相结合的果蝇同源PKD 2的表征。在下一个项目中，C.线虫分子遗传学鉴定和定义基因作用途径将应用于PKD功能。PI发现C. elegans同源蛋白PKD 1（LOV 1）是正常交配行为所必需的，C. PKD 2的elegans同源物共定位于相同的细胞类型。他将使用这种行为表型来确定这些基因产物类型的遗传途径。他将利用这种行为表型来确定这些基因产物参与的遗传途径。最后一个项目将确定PKD 1和PKD 2功能重要结构域的3D结构，这些结构域在其天然的非疾病状态下单独表达，并相互复合。试点和可行性研究的PI先前发现，大约25%的脊椎动物PKD 1与海胆精子的卵胶受体具有高度同源性。在目前的提案中，他将识别和描述海胆PKD 2。

ADPKD is a systemic disorder characterized by cysts involving multiple organs, cerebral and aortic aneurysms, cardiac valvular abnormalities and renal failure. The genes responsible for the two most common forms of the disease have been identified. The PKD1 genes responsible for the most common forms of the disease have been identified. The PKD1 gene product is a probably membrane glycoprotein thought to mediate cell-cell or cell- matrix interactions. PKD2 is predicted to have 6 transmembrane domains and has modest homology to several types of ion channels. The cellular role of either protein has not yet been elucidated. This proposal brings together the collective expert5ise of investigators from multiple disciplines to tackle the problem in a "Center without walls". One project is a competitive renewal that aims to identify and characterize binding partners of human PKD1. The PI has shown that the C-terminus of PKD1 can bind to PKD2 and to an unusual Db1 family member of using multiple in vitro and in vivo methods. This proposal seeks to determine the biological relevance of the pr3eviously observed interactions. It also will continue the search for PKD1 binding partners in relevance of the previously observed interactions. It also will continue the search for PKD1 binding partners in fetal murine tissues obtained at a stage when PKD1 protein is known to be essential. Another project will characterize the functions of PKD2 in vivo and in vitro using a model experimental organism, Drosophila melanogaster. The proposed research takes a multi-disciplinary approach to the characterization of the Drosophila homologue of PKD2 using a combination of genetic, electrophysiological, cell biological, biochemical and molecular approaches. In the next project, the power of C. elegans molecular genetics to identify and define pathways of gene action will be applied to PKD function. The PI has found that the C. elegans homologue of PKD1 (LOV1) is essential for normal mating behavior and that the C. elegans homologue of PKD2 co- localizes to the same cell types. He will use this behavioral phenotype to determine the genetic pathways in which these gene products types. He will use this behavioral phenotype to determine the genetic pathways in which these gene products participate. The last project will determine the 3D structures of functionally important domains of PKD1 and PKD2 expressed in their native, non-disease state alone and in complex with each other. The PI of the Pilot and Feasibility has previously found that approximately 25% of vertebrate PKD1 has high homology to the Receptor for Egg Jelly of sea urchin sperm. In the current proposal, he will identify and characterize the sea urchin PKD2.