Regulation of Actin Dynamics

肌动蛋白动力学的调节

基本信息

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
项目类别：
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

项目摘要

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塑，响应内在和外在的信号对于执行许多真核细胞功能至关重要从细胞/环境压力中恢复的进展，细胞运动，分泌和恢复。这些动态重排受一个大的，但尚未完全定义或理解的调节肌动蛋白结合蛋白的电池。该提案旨在继续对功能进行调查肌动蛋白动力学的三个新型调节剂：NADPH氧化还原酶旧黄色酶（OYE2P; AIM＃1），MAPKKK SSK2P（AIM＃2）和Cofilin Activator AIP1P（AIM＃3）。我们的 OYE2P的先前工作表明，它控制了C285-C374二硫键的氧化还原状态在肌动蛋白中可能影响F-肌动蛋白稳定性，对氧化应激和细胞死亡/衰老的敏感性。我们建议将这些研究扩展到研究肌动蛋白氧化如何改变肌动蛋白动力学和细胞如何调节其在响应和从氧化应激中恢复。特别是我们寻求识别F-肌动蛋白的组成部分含有在严重氧化应激上形成的氧化的actrin体。 SSK2P激酶渗透应激后促进肌动蛋白细胞骨架的重新叠成。我们对此的研究保守的蛋白质以及对渗透应激的适应激酶的相关底物，该激酶驱动肌动蛋白细胞骨架的重新彻底化候选蛋白方法和通过质谱法鉴定相关蛋白质。 AIP1P是小肌动蛋白结合蛋白cofilin的保守辅助因子。这两种蛋白质演唱会起作用在体外破坏肌动蛋白丝的稳定，并在各种肌动蛋白网络中驱动肌动蛋白动力学。 AIP1P-COFILIN复合物的结构/功能分析已导致该复合物的模型。我们寻求继续这些研究，以进一步完善该模型，以进一步了解 F-肌动蛋白去稳定的机理通过Cofilin进行。这种方法将利用我们最近确定了Cofilin中的功能突变体的增益，我们具有亚两个晶体学数据。