Glomerular Capillaries-- Normal and Pathologic

肾小球毛细血管——正常和病理

基本信息

批准号：
8060511
负责人：
MARILYN Gist FARQUHAR
金额：
$ 40.56万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN DIEGO
依托单位国家：
美国
项目类别：
财政年份：
1979
资助国家：
美国
起止时间：
1979-05-01 至 2014-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8060511
关键词：
1-Phosphatidylinositol 3-Kinase Abbreviations Actin-Binding Protein Actins Acute Adaptor Signaling Protein Affect Albumins Animal Disease Models Antibodies Apical Architecture Back Binding Biochemical Biological Cell Shape Cell membrane Cells Centrosome Clathrin Co-Immunoprecipitations Coated vesicle Complex Cytoskeleton Data Databases Diabetic Nephropathy Disease Down-Regulation Dynein ATPase Early Endosome Electron Microscopy Endocytosis Endosomes Epithelial Cells Epithelium Equipment and supply inventories Event Filtration Focal glomerulosclerosis Foot Process G Actin G Protein-Coupled Receptor Genes G-Protein-Coupled Receptors Glomerular Capillary Glomerulonephritis Goals Grant Green Fluorescent Proteins Growth Factor Health Immunofluorescence Immunologic In Situ Injury Insulin-Like Growth Factor Receptor LDL-Receptor Related Protein 2 Lead Life Light Link Low Density Lipoprotein Receptor Low-Density Lipoproteins MDCK cell Maps Mediating Membranous Glomerulonephritis Microtubule-Organizing Center Microtubules Mitosis Molecular Motor Multiprotein Complexes Mus Nephrosis Organelles Pathogenesis Pathologic Pathology Pathway interactions Permeability Play Procedures Process Protein Binding Proteins Proteinuria Proteomics Puromycin Aminonucleoside RGS19 gene Rattus Receptor Protein-Tyrosine Kinases Recycling Regulation Renal function Renal glomerular disease Research Role Route Shapes Signal Transduction Signaling Molecule Small Interfering RNA Spectrin System Tail Testing Tight Junctions Tubulin Up-Regulation Vascular Endothelial Growth Factors Work Yolk Sac absorption base cell growth cell motility cellular imaging coated pit expectation ezrin glomerular basement membrane glomerular filtration glomerular function hypercholesterolemia insight interest molecular marker nephrin novel occludin podocalyxin podocyte protein complex receptor receptor mediated endocytosis response slit diaphragm trafficking ubiquitin-protein ligase uptake

项目摘要

DESCRIPTION (provided by applicant): During the last 25 years our research has been directed toward understanding the cellular and molecular mechanisms of glomerular permeability and protein absorption as well as understanding the derangements in these processes that occur in glomerular diseases. Major findings of the previous renewal period were: 1) Demonstration that the adaptor protein ARH facilitates endocytosis and recycling of megalin; 2) Discovery that ARH associates with centrosomal proteins and dynein; 3) Identification of novel components of the nephrin multiprotein complex, including IQGAP, MAGI- 2, CASK, and spectrins; 4) Demonstration that podocalyxin expression activates RhoA and induces actin reorganization; 5) Demonstration that podocalyxin can bind directly to ezrin and is hyperphosphorylated in PAN nephrosis; 6) Discovery of GIPN, a putative E3 ubiquitin ligase that interacts with RGS-GAIP and promotes downregulation of Gai3. The studies proposed in this application represent a direct continuation of our ongoing work on characterization of the slit diaphragm proteins, role of actin and actin binding proteins in regulating foot process organization, and the role of ARH and megalin in protein uptake. Our specific aims are: 1) To isolate and characterize the glomerular junctional complexes from normal and PAN nephrotic rats; 2) To investigate and regulation of the foot process organization through its interaction with actin. We will explore the possibility that GIV expression can serve as a marker for glomerular injury in proteinuria involving changes in the unique foot process organization of the podocyte; and 3) To further define the functions of the adaptor protein ARH in megalin's uptake of filtered proteins in the podocyte and proximal tubule cell as well as to explore its role in connecting endocytosis to centrosomal functions. In this work we will use a combination of morphological (EM, immunofluorescence), biochemical, molecular biological and proteomics approaches on glomeruli and cultured podocytes as well as live cell imaging on mouse podocytes and proximal tubule cells in culture. It is our hope and expectation that these studies will continue to provide new insights into our understanding of the cellular and molecular mechanisms of glomerular filtration, glomerular injury, protein absorption, and their alterations in glomerular diseases associated with proteinuria. PUBLIC HEALTH RELEVANCE: Under this grant we have discovered and characterized a number of new proteins present on the podocyte that are important in normal glomerular permeability and in the pathogenesis of glomerular diseases associated with proteinuria, including minimal change disease, membranous glomerulonephritis and focal glomerulosclerosis. The studies planned can be expected to shed light not only on how normal podocytes function, but also they should provide key insights into the molecular basis of the signaling that directs changes in the actin cytoskeleton and organization of the foot processes of podocytes that occur in diseases associated with proteinuria and in the survival pathways for podocytes and proximal tubule cells in response to glomerular injury that lead to proteinuria.

描述（由申请人提供）：在过去 25 年中，我们的研究致力于了解肾小球通透性和蛋白质吸收的细胞和分子机制，以及了解肾小球疾病中发生的这些过程的紊乱。上一个更新期的主要发现是：1）证明接头蛋白ARH促进巨蛋白的内吞作用和再循环； 2）发现ARH与中心体蛋白和动力蛋白相关； 3) 鉴定去氧肾上腺素多蛋白复合物的新成分，包括IQGAP、MAGI-2、CASK和血影蛋白； 4) 证明足萼蛋白表达激活RhoA并诱导肌动蛋白重组； 5) 证明足萼蛋白可以直接与埃兹蛋白结合，并在 PAN 肾病中过度磷酸化； 6) GIPN 的发现，一种推定的 E3 泛素连接酶，与 RGS-GAIP 相互作用并促进 Gai3 的下调。本申请中提出的研究代表了我们正在进行的研究工作的直接延续，这些研究涉及裂隙隔膜蛋白的表征、肌动蛋白和肌动蛋白结合蛋白在调节足突组织中的作用，以及ARH和巨蛋白在蛋白质摄取中的作用。我们的具体目标是： 1) 分离和表征正常和 PAN 肾病大鼠的肾小球连接复合物； 2）通过足突组织与肌动蛋白的相互作用来研究和调节足突组织。我们将探讨 GIV 表达作为蛋白尿肾小球损伤标志物的可能性，涉及足细胞独特的足突组织的变化； 3) 进一步明确接头蛋白ARH在足细胞和近曲小管细胞中巨蛋白摄取过滤蛋白中的功能，并探讨其在连接内吞作用与中心体功能中的作用。在这项工作中，我们将结合形态学（EM、免疫荧光）、生物化学、分子生物学和蛋白质组学方法对肾小球和培养的足细胞进行分析，并对培养的小鼠足细胞和近端小管细胞进行活细胞成像。我们希望和期望这些研究将继续为我们理解肾小球滤过、肾小球损伤、蛋白质吸收的细胞和分子机制及其在与蛋白尿相关的肾小球疾病中的改变提供新的见解。公共健康相关性：在这笔资助下，我们发现并鉴定了足细胞上存在的许多新蛋白质，这些蛋白质对于正常肾小球通透性和与蛋白尿相关的肾小球疾病的发病机制很重要，包括微小病变、膜性肾小球肾炎和局灶性肾小球硬化症。预计计划中的研究不仅可以揭示正常足细胞的功能，而且还可以为指导疾病中发生的肌动蛋白细胞骨架和足细胞足突组织变化的信号传导的分子基础提供重要见解。与蛋白尿相关，并且参与足细胞和近端小管细胞响应导致蛋白尿的肾小球损伤的生存途径。