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、心脏瓣膜异常和肾功能衰竭为特征。导致这种疾病最常见的两种形式的基因已经确定。与最常见的疾病形式有关的PKD1基因已经被确定。PKD1基因产物可能是一种膜糖蛋白，被认为介导细胞与细胞或细胞与基质的相互作用。据预测，PKD2有6个跨膜结构域，与几种类型的离子通道有一定的同源性。这两种蛋白质在细胞中的作用还没有被阐明。这项建议汇集了来自多个学科的研究人员的集体专家，在一个“没有围墙的中心”解决这个问题。一个项目是竞争性更新，旨在确定和表征人PKD1的结合伙伴。等电点研究表明，通过多种体外和体内方法，PKD1的C末端可以与PKD2和一个不常见的DB1家族成员结合。这一提议试图确定之前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PKD1结合伙伴。它还将继续在胚胎小鼠组织中寻找PKD1结合伙伴，该组织是在已知PKD1蛋白必不可少的阶段获得的。另一个项目将利用一种模型实验生物--黑腹果蝇，在体内和体外表征PKD2的功能。这项拟议的研究采用多学科方法，结合遗传学、电生理学、细胞生物学、生化和分子方法来表征PKD2的果蝇同源物。在下一个项目中，线虫分子遗传学识别和定义基因作用途径的能力将应用于PKD功能。PI已经发现线虫的PKD1同源物(LOV1)对于正常的交配行为是必不可少的，并且线虫的PKD2的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PKD1和PKD2的功能重要结构域的3D结构，它们在各自的天然、非疾病状态下单独表达，以及在彼此复杂的状态下表达。Pilot的等电点和可行性先前已经发现大约25%的脊椎动物PKD1与海胆精子的卵冻受体有很高的同源性。在目前的提案中，他将识别和描述海胆PKD2。

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.