Regulation of Actin Dynamics

肌动蛋白动力学的调节

• 批准号: 7462650
• 负责人: DAVID C AMBERG
• 金额: $ 27.65万
• 依托单位: UPSTATE MEDICAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-05-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7462650
• 关键词: Actin-Binding Protein; Actins; Adopted; Affect; Age; Automobile Driving; Binding; Biological Models; Cell Cycle Progression; Cell Death; Cell physiology; Cells; Cellular Morphology; Cellular Stress; Charge; Complement; Complex; Crystallography; Cysteine; Cytoskeleton; DNA Sequence Rearrangement; Data; Disulfides; Elements; Eukaryotic Cell; F-Actin; Filament; Genes; Genetic Screening; Grant; Homologous Gene; Human; In Vitro; Investigation; Kidney; Lead; MAP Kinase Kinase Kinase; MEKKs; Mammalian Cell; Mass Spectrum Analysis; Membrane; Microfilaments; Modeling; Molecular; Mutate; Mutation; NADPH Dehydrogenase; NADPH Oxidoreductases NADH; Osmolar Concentration; Oxidation-Reduction; Oxidative Stress; Pathway interactions; Patients; Phosphotransferases; Proteins; Reactive Oxygen Species; Recovery; Regulation; Reporting; Resolution; Roentgen Rays; Role; Saccharomyces cerevisiae; Set protein; Sickle Cell; Signal Transduction; Stress; Structure; Structure-Activity Relationship; Testing; Work; Yeasts; biological adaptation to stress; cell motility; cofactor; cofilin; disulfide bond; gain of function; human MAP3K1 protein; in vivo; inhibitor/antagonist; insight; loss of function; mutant; novel; oxidation; response

Continual dynamic remodeling of the actin cytoskeleton, in response to intrinsic and extrinsic signals, is critical for the execution of many eukaryotic cell functions including cell cycle progression, cell motility, secretion and recovery from cellular/environmental stress. These dynamic rearrangements are regulated by a large, and as yet incompletely defined or understood, battery of actin binding proteins. This proposal seeks to continue investigations on the functions of three novel regulators of actin dynamics: the NADPH oxidoreductase Old Yellow Enzyme (Oye2p; Aim #1), the MAPKKK Ssk2p (Aim #2), and the cofilin activator Aip1p (Aim #3). Our previous work on Oye2p suggests that it controls the redox state of a C285-C374 disulfide bond in actin that can in turn affect F-actin stability, sensitivity to oxidative stress and cell death/aging. We propose to extend those studies to investigate how actin oxidation alters actin dynamics and how the cell regulates the organization of its F-actin structures during the response to, and recovery from, oxidative stress. In particular we seek to identify the components of F-actin containing oxidized actrin bodies that form upon a severe oxidative stress. The Ssk2p kinase facilitates re-polarization of the actin cytoskeleton following osmotic stress. Our studies on this conserved protein, and the adaptation to osmotic stress, will be extended by identifying the relevant substrates of the kinase that drive re-polarization of the actin cytoskeleton employing a candidate protein approach and by identifying associated proteins by mass-spectrometry. Aip1p is a conserved cofactor of the small actin binding protein cofilin. These two proteins act in concert to destabilize actin filaments in vitro and drive actin dynamics in vivo in diverse actin networks. Structure/function analysis of the Aip1p-cofilin complex has led to a model for the complex; we seek to continue these studies in order to further refine this model to gain further insight into the mechanism of F-actin de-stabilization by cofilin. This approach will take advantage of our recently identified gain of function mutants in cofilin for which we have sub-two angstrom crystallography data.

肌动蛋白细胞骨架的持续动态重构，对内在和外在的反应 信号对包括细胞周期在内的许多真核细胞功能的执行至关重要 进展、细胞运动、分泌和从细胞/环境应激中恢复。这些 动态重排由一个尚未完全定义或理解的大的， 肌动蛋白结合蛋白电池。该提案寻求继续对这些职能进行调查 三种新的肌动蛋白动力学调节因子：NADPH氧化还原酶--老黄酶 (Oye2p；Aim#1)、MAPKKK Ssk2p(Aim#2)和Cofilin激活剂Aip1p(Aim#3)。我们的 之前对Oye2p的研究表明，它控制着C285-C374二硫键的氧化还原状态 在肌动蛋白中，它可以反过来影响F-肌动蛋白的稳定性、对氧化应激的敏感性和细胞死亡/衰老。 我们建议扩展这些研究，以调查肌动蛋白氧化如何改变肌动蛋白动力学和 细胞如何在对和/或 从氧化应激中恢复过来。特别是，我们试图确定F-肌动蛋白的成分 含有氧化的Actrin小体，在严重的氧化应激下形成。Ssk2p蛋白激酶 促进渗透胁迫后肌动蛋白细胞骨架的再极化。我们对此的研究 保守的蛋白质，以及对渗透胁迫的适应，将通过识别 驱动肌动蛋白细胞骨架再极化的激酶的相关底物 候选蛋白质的方法和通过质谱学鉴定相关蛋白质。Aip1p是 小肌动蛋白结合蛋白cofilin的保守辅因子。这两种蛋白质协同作用。 在体外破坏肌动蛋白细丝的稳定性，并在体内驱动不同肌动蛋白网络中的肌动蛋白动力学。 对Aip1p-cofilin复合体的结构/功能分析导致了该复合体的模型；我们 寻求继续这些研究，以便进一步完善这一模型，以进一步深入了解 Cofilin对F-肌动蛋白去稳定作用的机制。这种方法将利用我们的 最近确定的功能突变体在粘连蛋白中的增益，我们对其具有亚2埃 结晶数据。