Acute Regulation of the Renal Na/H Exchanger NHE-3

描述（改编自申请人的摘要）：肾脏呈现一种 在维持酸碱平衡、细胞外液 通过调节NaHCO 3和NaCl实现容量和血压稳态 排泄在所有这些过程中起核心作用的一种蛋白质是Na/H 交换异构体3（NHE 3）。NHE 3介导绝大多数NaHCO 3和NaCl 吸收，并通过血液动力学和神经体液因素快速调节， 满足有机体的需求。几个实验室最近的工作 显示NHE 3可以通过许多机制进行剧烈调节。我们有 表明激素如甲状旁腺激素（PTH）和多巴胺（DA）急性 抑制NHE 3转运活性，而不改变其在细胞表面的蛋白丰度。 质膜这种抑制与NHE 3的复杂变化有关 NHE 3蛋白的磷酸化和成簇。我们假设， NHE 3磷酸化导致NHE 3聚簇，这导致NHE 3磷酸化抑制。 活动在这个提议中。我们将重点关注：一，绘制出氨基酸 NHE 3中的残基被调节。2.确定NHE 3是否发生变化 磷酸化本身改变其活性。3.确定NHE 3是否 聚集是NHE 3磷酸化变化的结果， 聚集改变其活性。4.确定两个NHE 3结合的作用 蛋白质介导NHE 3聚集和磷酸化的变化。我们将 在运输生理学、生物化学、重组DNA中应用协议 技术、免疫组织学和生物物理荧光光谱学来研究 整个动物、培养细胞和纯化的蛋白质。这些研究将揭示 一种重要的上皮细胞Na+的高度基本的调节机制 转运蛋白和进一步我们的理解01酸碱平衡紊乱， 钠潴留和高血压。

DESCRIPTION (Adapted from the Applicant's Abstract): The kidney assumes a critical role in the maintenance of acid-base balance, extracellular fluid volume and blood pressure homeostasis via regulation of NaHCO3 and NaCI excretion. One protein that is central to all these processes is the Na/H exchanger isoform 3 (NHE3). NHE3 mediates a great majority of NaHCO3 and NaCI absorption and is regulated rapidly by hemodynamic and neurohumoral factors to match the demands of the organism. Recent work from several laboratories have shown that NHE3 can be acutely regulated via a number O mechanisms. We have shown that hormones such as parathyroid hormone (PTH) and dopamine (DA) acutely inhibits NHE3 transport activity without changing its protein abundance on the plasma membrane. This inhibition is associated with complex changes in NHE3 phosphorylation and clustering of NHE3 proteins. We hypothesize that changes in NHE3 phosphorylation leads to NHE3 clustering which results in inhibition of activity. In this proposal. we will focus on: 1. Mapping out the amino acid residues n NHE3 that is regulated. 2. Determine whether changes in NHE3 phosphorylation per se alters its activity. 3. Determine whether NHE3 clustering is a consequence of changes in NHE3 phosphorylation and whether NHE3 clustering alters its activity. 4. Determine the role of two NHE3 binding proteins in mediating NHE3 clustering and changes in phosphorylation. We will apply protocols in transport physiology, biochemistry, recombinant DNA technology, immunohistology, and biophysical fluorescence spectroscopy to study whole animals, culture cells, and purified proteins. These studies will uncover highly fundamental mechanisms of regulation of an important epithelial Na transporter and further our understanding 01 disturbance in acid-base balance, Na retention and hypertension.