The Molecular Mechanisms of Polycystin-1 Proteolytic Cleavage in Kidney Health and Polycystic Kidney Disease

多囊蛋白-1 蛋白水解切割在肾脏健康和多囊肾病中的分子机制

基本信息

批准号：
9348875
负责人：
Feng Qian
金额：
$ 11.56万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-21 至 2017-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9348875
关键词：
Adhesions Affect Atomic Force Microscopy Autosomal Dominant Polycystic Kidney Back Biochemical Biogenesis Biophysical Process Birth CRISPR/Cas technology Cells Cilia Cleaved cell Computer Simulation Cyst Defect Development Disease Distal Embryo Figs - dietary Goals Health Homeostasis Human Image In Vitro Individual Kidney Kidney Failure Knock-in Mouse Length Link Liquid substance Measures Mediating Mendelian disorder Missense Mutation Molecular Mus Mutant Strains Mice Mutation N-terminal Nephrons PKD1 gene Pathogenesis Pathway interactions Patients Pattern Phenotype Polycystic Kidney Diseases Process Property Proteins Recruitment Activity Renal function Renal tubule structure Role Series Signal Transduction Staging Structure Testing Therapeutic Transmembrane Domain Tubular formation Uncertainty Work abstracting base biophysical techniques ciliopathy falls flexibility functional restoration in vivo insight molecular dynamics mouse model mutant nephrogenesis novel novel strategies polycystic kidney disease 1 protein postnatal success trafficking transmission process

项目摘要

Project Summary/Abstract Our long-term goal is to understand specific functions of the heterodimeric (PC1cFL) and uncleaved (PC1U) states of Polycystin-1 (PC1) in kidney development, homeostasis, and PKD pathogenesis. This project builds on our previous key findings involving cleavage of PC1 at the GPS motif and investigates molecular and biophysical mechanisms by which cleavage regulates the trafficking and function of PC1. The central hypothesis is that the GPS motif and the adjacent linker form a bipartite force-transduction module that mediates critical functions of PC1 in both the heterodimeric (PC1cFL) and uncleaved (PC1U) states. In PC1cFL, GPS cleavage generates a conformational change of the last β-strand (β-1) within the GPS motif, which enables ciliary trafficking and function via signal transmittance from the rigid linker to the N-terminal transmembrane domain. In PC1U, the linker transduces cleavage-independent forces. This proposal will use a multi-disciplinary approach to test the functional role of the GPS-linker module. 1) We will test the hypothesis that a tight association of the β1-strand within the GPS is required for PC1 ciliary trafficking and function, and is disrupted by PKD1-associated mutations. We predict that tight association of this β-strand within the GPS motif is required to enable PC1 to recruit Rabep1/GGA1/Arl3, traffic to cilia, and induce in vitro tubulogenesis. 2) We will test the hypothesis that high rigidity and short length in the linker is required for PC1 ciliary trafficking and function, and is disrupted by PKD1-associated mutations. We predict that the rigidity of the linker is required to enable PC1 to recruit Rabep1/GGA1/Arl3, traffic to cilia, and induce in vitro tubulogenesis. 3) We will determine the in vivo role and mechanism of the GPS-linker module by generating two Pkd1 knockin mouse models, one with impaired GPS association and the other with a flexible linker. We will examine their kidney phenotypes at development and after birth, and analyze the biogenesis and trafficking of mutant PC1 in proximal and distal nephron cells. The proposed studies will provide novel mechanistic insights into the GPS-linker module and dissect their functional role for the two PC1 forms at kidney development and in different nephron-segments in postnatal periods. These insights should result in therapeutic strategies that restore normal PC1 function in a significant proportion of ADPKD patients by manipulating the GPS-linker module force-transduction process.

项目概要/摘要我们的长期目标是了解异二聚体 (PC1cFL) 和未切割 (PC1U) 的特定功能该项目建立了多囊蛋白-1 (PC1) 在肾脏发育、稳态和 PKD 发病机制中的状态。基于我们之前涉及 GPS 基序处 PC1 裂解的关键发现，并研究了分子和裂解调节 PC1 运输和功能的生物物理机制。假设 GPS 基序和相邻的连接子形成一个二分力传导模块，在异二聚体 (PC1cFL) 和未切割 (PC1U) 状态下介导 PC1 的关键功能。 GPS 裂解产生 GPS 基序内最后一个 β 链 (β-1) 的构象变化，通过从刚性接头到 N 端的信号传输实现纤毛运输和功能在 PC1U 中，连接子转换独立于切割的力。多学科方法来测试 GPS 链接器模块的功能作用 1) 我们将测试假设。 PC1 纤毛运输和功能需要 GPS 内 β1 链的紧密结合，并且我们预测 GPS 基序内该 β 链的紧密关联。需要使 PC1 能够募集 Rabep1/GGA1/Arl3、运输至纤毛并诱导体外肾小管形成 2) 我们。将检验 PC1 纤毛运输需要接头具有高刚性和短长度的假设，并且我们预测连接子的刚性是必需的，并且会被 PKD1 相关突变破坏。使 PC1 能够招募 Rabep1/GGA1/Arl3，运输至纤毛，并诱导体外肾小管形成 3) 我们将确定。通过生成两个 Pkd1 敲入小鼠模型来研究 GPS-linker 模块的体内作用和机制，其中一个 GPS 关联受损，另一个具有灵活的连接器，我们将在以下位置检查它们的肾脏表型。发育和出生后，并分析突变PC1在近端和远端的生物发生和运输拟议的研究将为 GPS 链接器模块和肾单位细胞提供新的机制见解。剖析两种 PC1 形式在肾脏发育和不同肾单位段中的功能作用这些见解应该会产生恢复正常 PC1 功能的治疗策略。通过操纵 GPS 连接器模块力传导过程来治疗很大一部分 ADPKD 患者。