NHERF-1 and PTH regulation of the renal transport of phosphate.

NHERF-1 和 PTH 调节磷酸盐的肾脏转运。

基本信息

批准号：
7523305
负责人：
EDWARD J WEINMAN
金额：
$ 31.88万
依托单位：
UNIVERSITY OF MARYLAND BALTIMORE
依托单位国家：
美国
项目类别：
财政年份：
1999
资助国家：
美国
起止时间：
1999-05-01 至 2013-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7523305
关键词：
8-Bromo Cyclic Adenosine Monophosphate Accounting Adaptor Signaling Protein Adenoviruses Affinity Animals Binding Binding Proteins Biochemical Biological Assay C-terminal Canis familiaris Cell Line Cell membrane Cells Co-Immunoprecipitations Complex Confocal Microscopy Cyclic AMP-Dependent Protein Kinases Cytoskeleton Density Gradient Centrifugation Diet Disease Dissociation Elements Estrogen receptor positive Excretory function Family Functional disorder Gel Genes Goals Hormones In Vitro Ingestion Inorganic Phosphate Transporter Ion Channel Kidney Localized Maps Mediating Membrane Mitogen-Activated Protein Kinases Modeling Mus Mutation NHERF-1 protein Nephrolithiasis Neurofibromin 2 Parathyroid Hormone Receptor Parathyroid Hormones Pathway interactions Phosphoproteins Phosphorylation Phosphorylation Site Physiological Process Protein Binding Protein Dephosphorylation Protein Family Protein Hormone Receptor Protein Kinase Protein Kinase C Proteins Proximal Kidney Tubules Psoriasis Public Health Regulation Reporting Resistance Role Scaffolding Protein Schizophrenia Second Messenger Systems Serine Signal Transduction Signaling Protein Site Site-Directed Mutagenesis Slice Sodium Stimulus Sucrose Surface Plasmon Resonance Threonine Tubular formation apical membrane brush border membrane citrate carrier ezrin hormone regulation human PTH protein in vivo inorganic phosphate insight interest malignant breast neoplasm member moesin radixin protein response second messenger sodium-hydrogen exchanger regulatory factor trafficking urinary

项目摘要

DESCRIPTION (provided by applicant): The Sodium-Hydrogen Exchanger Regulatory Factor-1 (NHERF-1) is an adaptor protein containing two protein-interactive PDZ domains and a C-terminal ERM binding domain that localizes to the brush border membrane of renal proximal convoluted tubule cells and binds to Npt2a, the major sodium-dependent phosphate transporter. Our recent studies have indicated that sodium-dependent phosphate transport in proximal tubule cells from NHERF-1-/- kidneys are resistant to the inhibitory effect of Parathyroid Hormone (PTH). In this current application, we explore the hypothesis that PTH mediates the phosphorylation of specific residues in PDZ I of NHERF-1 thereby regulating Npt2a/NHERF-1 complexes, the abundance of Npt2a in the apical membrane of renal proximal tubule cells, and as a consequence, the tubular reabsorption of phosphate. Elucidation of the factors that regulate the binding of target proteins to PDZ I of NHERF-1 may also provide broader insights into how regulation of PDZ domains of adaptor proteins impact on biologic responses to hormones and on the pathophysiology of NHERF-1 related diseases. In intact animals and cultured proximal tubule cells, we will use physiologic, biochemical, and cell biologic assays to determine how PTH-mediated phosphorylation of PDZ I of NHERF-1 regulateS the binding affinity of target proteins such as Npt2a and the proximal tubule reabsorption of phosphate. We propose three specific aims. First, we will map the serine and/or threonine residues in PDZ I of NHERF-1 that are phosphorylated in response to PTH and downstream protein kinases. Second, we propose to study the association and dissociation of Npt2a/NHERF-1 complexes in response to PTH-mediated phosphorylation of NHERF-1 using in-vitro and in-vivo assays. Third, we will determine the physiologic role of PTH-mediated NHERF-1 phosphorylation on the regulation of phosphate transport in the proximal tubule of the kidney. PUBLIC HEALTH RELEVANCE NHERF-1 is an adaptor protein that binds multiple transporters in the kidney including Npt2a, the major renal proximal tubule phosphate transporter. We will study the hypothesis that Parathyroid Hormone-mediated regulation of renal phosphate transport involves regulation of the binding of Npt2a to NHERF-1 by site-specific phosphorylation of the PDZ I domain of NHERF-1. These observations may provide mechanistic insights into the processes that regulate the binding of target proteins to adaptors such as NHERF-1 and provide broader insights into the pathophysiology of NHERF-1 related diseases.

DESCRIPTION (provided by applicant): The Sodium-Hydrogen Exchanger Regulatory Factor-1 (NHERF-1) is an adaptor protein containing two protein-interactive PDZ domains and a C-terminal ERM binding domain that localizes to the brush border membrane of renal proximal convoluted tubule cells and binds to Npt2a, the major sodium-dependent phosphate transporter.我们最近的研究表明，来自NHERF-1 - / - 肾脏近端小管细胞中钠依赖性磷酸转运具有抗甲状旁腺激素（PTH）的抑制作用。在当前的应用中，我们探讨了PTH介导NHERF-1中PDZ I特定残基的磷酸化的假设，从而调节了NPT2A/NHERF-1复合物，即NPT2A的丰度，在肾近端肾小管细胞的顶膜中，NPT2A的丰度，因此，导致了小管的phosepormatign phosphate。阐明调节靶蛋白与NHERF-1的PDZ I结合的因素也可能提供更广泛的见解，以了解适配蛋白的PDZ域如何影响对激素的生物学反应以及对NHERF-1相关疾病的病理生理学的影响。在完整的动物和培养的近端小管细胞中，我们将使用生理学，生化和细胞生物学测定法来确定NHERF-1的PDZ I的PDZ I磷酸化如何调节NHERF-1的PDZ I，从而调节NPT2A（如NPT2A）和近端管的近代型肾小管蛋白的结合亲和力。我们提出了三个具体目标。首先，我们将在NHERF-1的PDZ I中绘制丝氨酸和/或苏氨酸残基，这些丝氨酸和/或苏氨酸残基是响应于PTH和下游蛋白激酶的磷酸化的。其次，我们建议使用维特罗（Vitro）和体内测定法研究NHERF-1介导的NHERF-1磷酸化，研究NPT2A/NHERF-1复合物的关联和解离。第三，我们将确定PTH介导的NHERF-1磷酸化对肾脏近端小管中磷酸转运的调节的生理作用。公共卫生相关性 NHERF-1是一种衔接蛋白，它结合肾脏中的多个转运蛋白，包括NPT2A，这是主要的肾脏近端小管磷酸转运蛋白。我们将研究以下假设：甲状旁腺激素介导的肾磷酸转运调节涉及通过NHERF-1的PDZ I域的位点特异性磷酸化对NPT2A与NHERF-1的结合。这些观察结果可能会提供对调节靶蛋白与适配器（例如NHERF-1）结合的过程的机械见解，并为NHERF-1相关疾病的病理生理提供了更广泛的见解。