Mechanisms of Receptor Regulated Na+-H+ Exchange

受体调节 Na -H 交换的机制

• 批准号: 7889280
• 负责人: DIANE L BARBER
• 金额: $ 7.86万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-13 至 2010-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7889280
• 关键词: Actin-Binding Protein; Actins; Affinity; Affinity Chromatography; Binding; C-terminal; Cell Proliferation; Cell Shape; Cell Volumes; Cells; Complex; Cytoplasmic Tail; Distal; Fibroblasts; Focal Adhesions; Funding; Guanosine Triphosphate Phosphohydrolases; Homeostasis; Integrins; Ions; Link; Mass Spectrum Analysis; Mediating; Membrane; Microfilaments; Multiprotein Complexes; Mutation; Myosin Light Chains; NHE1; Neoplastic Cell Transformation; Normal Cell; Null Lymphocytes; Pathway interactions; Phosphorylation; Phosphotransferases; Protein Binding; Proteins; Proteomics; Regulation; Rho-associated kinase; Role; Signal Transduction; Signaling Molecule; Site; Sum; Testing; extracellular; ezrin; growth factor-activatable Na-H exchanger NHE-1; moesin; mutant; pH Homeostasis; radixin protein; receptor; scaffold; tumor progression

DESCRIPTION (provided by applicant): The ubiquitously expressed Na-H exchanger NHE1 has an established function in intracellular pH and cell volume homeostasis by catalyzing electroneutral exchange of extracellular Na + for intracellular H+. During the past funding period an ancillary function of NHEI as an anchor for actin filaments was identified. Actin anchoring by NHE1 is mediated by direct binding of ERM (ezrin, radixin, moesin) actin-binding proteins, and in fibroblasts is necessary to retain the predominant localization of NHEI at the distal margin of lamellipodia, normal cell shape, and the assembly of cortical actin filaments. The objective of the current proposal is to establish an additional ancillary function of NHEI as a scaffolding platform for assembling signaling complexes and promoting signal relay. The C-terminal cytoplasmic domain of NHE1 binds directly at least 10 functionally distinct signaling molecules that coordinately regulate NHE1 activity, restrict the localization of NHE1 in specialized membrane domains, and as now proposed, may transmit signals to diverse effector pathways. Focusing on the role of NHE1 in fibroblasts, this proposal investigates the hypothesis that NHE1 acts as a spatially-restricted scaffold to assemble signaling complexes and to promote signal relay. Studies in Aim 1 will determine how NHE1 scaffolding and localization are integrated by asking how scaffolding by NHE1 determines its localization and in turn how the localization of NHE1 in lamellipodia determines its binding to identified interacting proteins. Studies in Aim 2 will test the role of an NHE1 scaffold in the assembly of signaling complexes and in signal relay. Distinct signaling units, each having a component that binds directly to NHE1, will be investigated by asking whether binding to NHE1, the localization of NHE1 in lamellipodia, or ion translocation by NHE1 is necessary for the assembly and signal relay of the unit. Representative signaling units include phosphorylation of ezrin and Arp2/3 by the Ste20-1ike kinase NIK, ezrin binding to the GTPases Rac and Cdc42, and phosphorylation of myosin light chain by the Rho kinase ROCK. Studies in Aim 3 will define the complexity of an NHE1 scaffold by using a proteomics approach to determine the dynamic composition of an NHE1 scaffold ensemble and then will ask whether the profile of associated proteins is different when NHE 1 localization or ion translocation is impaired. Established roles for NHEI in intracellular pH homeostasis, cell proliferation, neoplastic transformation, and tumor progression underscore the importance of understanding its structural and functional regulation of diverse signaling networks.

描述（由申请人提供）：无处不在表达的Na-H交换器NHE1在细胞内pH和细胞体积稳态中具有确定的功能，通过催化细胞外Na +的电荷交换以用于细胞内H +。在过去的资金期间，确定了NHEI作为肌动蛋白丝的锚固功能。 NHE1锚定的肌动蛋白是由ERM（Ezrin，radixin，Moesin）肌动蛋白结合蛋白的直接结合而介导的，而成纤维细胞则是保留NHEI在Lamellipodia远端的主要定位，正常细胞形状，正常的细胞形状，正常的Cortical Catoments的组装。当前建议的目的是建立NHEI作为组装信号复合物和促进信号继电器的脚手架平台的附加辅助功能。 NHE1的C末端细胞质结构域直接结合至少10个在功能上不同的信号分子，这些信号分子协同调节NHE1活性，限制NHE1在专用膜结构域中的定位，并且如今，正如现在所提出的，可能会传递信号到不同的效应器途径。该提案的重点是NHE1在成纤维细胞中的作用，研究了NHE1充当空间限制的支架，以组装信号复合物并促进信号继电器。 AIM 1中的研究将通过询问NHE1的脚手架来确定其定位以及NHE1在Lamellipodia中的定位如何确定其与已识别的相互作用蛋白的结合，从而确定NHE1支架和定位是如何整合的。 AIM 2中的研究将测试NHE1支架在信号复合物组装和信号继电器中的作用。通过询问与NHE1的结合，NHE1在Lamellipodia中的定位或NHE1的离子易位是对单元的组装和信号继电器的必要条件，将研究与NHE1，NHE1的定位，NHE1的定位，NHE1的定位是否与NHE1结合，对NHE1的结合，NHE1的定位是否与NHE1结合来研究。代表性的信号传导单元包括Ste20-1ike激酶Nik对Ezrin和ARP2/3的磷酸化，Ezrin与GTPases RAC和CDC42的结合以及Rho激酶岩石对肌球蛋白轻链的磷酸化。 AIM 3中的研究将通过使用蛋白质组学方法来确定NHE1支架的复杂性来确定NHE1支架集合的动态组成，然后询问相关蛋白的曲线在NHE 1定位或离子易位时是否有所不同。 NHEI在细胞内pH稳态，细胞增殖，肿瘤转化和肿瘤进展中确立了作用，强调了了解其对各种信号网络的结构和功能调节的重要性。