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Mechanisms of Kv1.2 regulation by tyrosine kinase

酪氨酸激酶调节Kv1.2的机制

基本信息

批准号：
7446702
负责人：
ANTHONY D MORIELLI
金额：
$ 33.22万
依托单位：
UNIVERSITY OF VERMONT & ST AGRIC COLLEGE
依托单位国家：
美国
项目类别：
财政年份：
2004
资助国家：
美国
起止时间：
2004-09-15 至 2010-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7446702
关键词：
Actin-Binding Protein Actins Acute Address Affect Attention Binding Biochemical Biochemistry Biological Blood Vessels Cardiovascular system Cell physiology Cells Complex Coupled Cytoskeleton Data Development Dissociation Dynamin Electrophysiology (science)Endocytosis GTP-Binding Proteins Gated Ion Channel Goals Human Pathology Hypertension Immunofluorescence Microscopy Individual Ion Channel Link Mediating Membrane Methods Microfilaments Microscopy Modeling Molecular Molecular Biology Monomeric GTP-Binding Proteins Mutation Neurons Numbers Pathway interactions Phosphorylation Phosphotransferases Physiology Principal Investigator Process Property Protein Kinase Protein Tyrosine Kinase Proteins Publishing Range Receptor Protein-Tyrosine Kinases Regulation Reporting Research Role Seizures Serine/Threonine Phosphorylation Signal Transduction Stroke System Testing Thinking Tyrosine Tyrosine Phosphorylation Ubiquitin Vascular Smooth Muscle Voltage-Gated Potassium Channel Work actin kinase cell type human EMS1 protein innovation insight interest intracellular protein transport multidisciplinary neuronal excitability novel programs protein transport research study rhoA GTP-Binding Protein scaffold voltage voltage gated channel

项目摘要

DESCRIPTION (provided by applicant): The long range objective of our research is to understand the cellular mechanisms governing neuronal excitability. This proposal focuses on the tyrosine kinase dependent suppression of Kv1.2, a voltage gated potassium channel. Kv1.2 is widely expressed throughout the nervous and cardiovascular systems and its suppression is hypothesized to have a key role in human pathologies ranging from neuronal hyperexcitability associated with seizure and stroke to increased vascular tone associated with hypertension. Despite its importance, the mechanism for Kv1.2 suppression remains almost completely unknown. The actin cytoskeleton appears to be central for Kv1.2 regulation since the actin-regulating proteins RhoA and cortactin bind to Kv1.2 and participate in its suppression by tyrosine kinases. Kv1.2 also undergoes tyrosine phosphorylation and actin cytoskeleton dependent endocytosis. This suggests a model in which the physical mechanism of channel suppression involves tyrosine phosphorylation dependent alteration of Kv1.2 interaction with the actin cytoskeleton leading to channel endocytosis and consequent loss of channel function. The goal of this proposal is to understand the molecular mechanisms by which tyrosine kinases, dynamic actin and the endocytotic machinery converge at Kv1.2 to regulate cellular excitability. To do so, a range of biochemical, molecular biological, immunofluorescence microscopy and electrophysiology methods will be used to address the following specific aims: Specific Aim 1: Determine the role of the actin binding protein cortactin in the regulation of Kv1.2 by testing the hypothesis that cortactin acts as a physical conduit between Kv1.2, the actin cytoskeleton, tyrosine kinases and proteins involved in endocytosis. Specific Aim 2: Determine the mechanisms of Kv1.2 endocytosis by testing the hypothesis that individual tyrosines within the channel have specific and divergent roles in ubiquitin dependent channel endocytosis. Specific Aim 3: Elucidate the role of the small G-protein RhoA in Kv1.2 suppression by testing the hypotheses that RhoA bound to Kv1.2 evokes channel suppression by activating effector proteins known to participate in actin filament reorganization. Collectively these aims explore the novel idea that tyrosine kinase signaling, cytoskeletal physiology and protein trafficking act as coordinated players in the regulation of Kv1.2. Thus, the experiments proposed here will provide fundamentally new perspectives into the mechanisms of ion channel regulation and the processes governing neuronal excitability.

描述（由申请人提供）：我们研究的长期目标是了解控制神经元兴奋性的细胞机制。该建议的重点是酪氨酸激酶依赖性抑制Kv1.2，电压门控钾通道。Kv1.2在整个神经和心血管系统中广泛表达，并且其抑制被假设在人类病理学中具有关键作用，所述人类病理学从与癫痫发作和中风相关的神经元过度兴奋到与高血压相关的血管张力增加。尽管其重要性，Kv1.2抑制的机制仍然几乎完全未知。肌动蛋白细胞骨架似乎是Kv1.2调节的中心，因为肌动蛋白调节蛋白RhoA和corneum结合Kv1.2并参与酪氨酸激酶的抑制。Kv1.2还经历酪氨酸磷酸化和肌动蛋白细胞骨架依赖性内吞作用。这表明了一种模型，其中通道抑制的物理机制涉及Kv1.2与肌动蛋白细胞骨架相互作用的酪氨酸磷酸化依赖性改变，导致通道内吞作用和随后的通道功能丧失。这个建议的目标是了解酪氨酸激酶，动态肌动蛋白和内吞机制收敛在Kv1.2调节细胞兴奋性的分子机制。为此，一系列的生物化学，分子生物学，免疫荧光显微镜和电生理学方法将被用来解决以下具体目标：具体目标1：确定肌动蛋白结合蛋白corneumn在调节Kv1.2通过测试的假设，corneumn作为一个物理管道之间的Kv1.2，肌动蛋白细胞骨架，酪氨酸激酶和蛋白质参与内吞作用的作用。具体目标二：通过检验通道内的单个酪氨酸在泛素依赖性通道内吞作用中具有特定和不同作用的假设，确定Kv1.2内吞作用的机制。具体目标3：通过测试RhoA与Kv1.2结合通过激活已知参与肌动蛋白丝重组的效应蛋白引起通道抑制的假设，阐明小G蛋白RhoA在Kv1.2抑制中的作用。总的来说，这些目标探索了酪氨酸激酶信号传导，细胞骨架生理学和蛋白质运输作为协调的球员在Kv1.2的调节新的想法。因此，本文提出的实验将为离子通道调节机制和神经元兴奋性的调控过程提供全新的视角。