Role of dynamin for podocytes structure and function

动力对足细胞结构和功能的作用

基本信息

批准号：
8730142
负责人：
Sanja Sever
金额：
$ 36.93万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-15 至 2016-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8730142
关键词：
Actins Adaptor Signaling Protein Affect Affinity Antibodies Applications Grants Arginine Binding C-terminal Clathrin Clathrin-Coated Vesicles Complex Cytoskeleton Data Down-Regulation Dynamin Endocytosis Epithelial Cells Fluorescence Microscopy Foot Process Functional disorder Gelsolin Glomerular Capillary Goals Guanosine Triphosphate Phosphohydrolases Higher Order Chromatin Structure In Vitro Integral Membrane Protein Intercellular Junctions Kidney Diseases Kidney Glomerulus Laboratories Link Lipids Mediating Membrane Microfilaments Molecular NCK adaptor protein 1 NPHS2 protein Phenotype Phosphorylation Play Proline Proteins Recruitment Activity Regulation Role SH3 Domains Signal Transduction Staging Structure Testing United States Work base blood filtration coated pit in vitro Assay mutant nephrin neuronal cell body novel podocyte polymerization slit diaphragm small molecule

项目摘要

DESCRIPTION (provided by applicant): The goal of this proposal is to elucidate the molecular mechanism(s) that underlie pathophysiology of proteinuric kidney diseases. Work from this laboratory recently demonstrated direct regulation of actin assembly in vitro and in podocytes by dynamin, and suggested an intricate interplay between dynamin's oligomerization cycle and actin cytoskeleton dynamics. Our findings motivate the hypothesis that signaling at the slit diaphragm alters dynamin oligomerization cycle through direct dynamin-Nck interactions, which in turn initiates actin polymerization in the membrane vicinity. In Specific Aim 1 we test whether clustering of Nck on the membrane promotes dynamin oligomerization into rings, which in turn drives actin polymerization in cultured podocytes. Using small molecules that specifically promote or inhibit dynamin oligomerization, as well as our novel actin binding domain mutants of dynamin, we test whether dynamin oligomerization cycle and actin-binding interactions are essential and/or sufficient for Nck-dependent polymerization of cortical actin cytoskeleton. In Specific Aim 2 we test whether direct interactions between dynamin and Nck alter the dynamin oligomerization cycle and dynamin-dependent actin polymerization using in-vitro assays. In Specific Aim 3, using TIR-FM we test whether direct dynamin-actin interactions are essential for clathrin mediated endocytosis.

描述（由申请人提供）：该提案的目的是阐明蛋白尿肾脏疾病病理生理的分子机制。该实验室的工作最近证明了肌动蛋白对肌动蛋白组装的直接调节，并提出了Dynamin的低聚循环和肌动蛋白细胞骨架动力学之间的复杂相互作用。我们的发现激发了这样的假设，即缝隙隔膜处的信号通过直接Dynamin-NCK相互作用改变了动力学的寡聚循环，这反过来又启动了膜附近的肌动蛋白聚合。在特定的目标1中，我们测试NCK在膜上的聚类是否会促进动力蛋白的寡聚为环，这反过来驱动培养的足细胞中的肌动蛋白聚合。使用特异性促进或抑制动力蛋白低聚的小分子，以及我们的新型肌动蛋白结合结构蛋白的突变体，我们测试元素蛋白寡聚循环和肌动蛋白结合相互作用是否必不可少，并且/或足以对NCK依赖性依赖性依赖性的皮质肌动蛋白蛋白细胞骨架。在特定的目标2中，我们使用视野测定法测试了动力蛋白和NCK之间的直接相互作用是否改变了动力蛋白的寡聚循环和依赖性肌动蛋白聚合的肌动蛋白聚合。在特定的目标3中，使用TIR-FM我们测试直接动力蛋白 - 肌动蛋白相互作用是否对于网格蛋白介导的内吞作用至关重要。