Actin filament elasticity and actin-binding protein function

肌动蛋白丝弹性和肌动蛋白结合蛋白功能

• 批准号: 9029502
• 负责人: ENRIQUE M DE LA CRUZ
• 金额: $ 42.43万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-15 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9029502
• 关键词: Actin-Binding Protein; Actins; Award; Binding; Binding Proteins; Binding Sites; Biochemical; Biological Process; Biology; Biomimetics; Cations; Cell Polarity; Cells; Chemistry; Complex; Computer Simulation; Contractile Proteins; Cryoelectron Microscopy; Cytokinesis; Cytoskeleton; Dissociation; Elasticity; Endocytosis; Eukaryotic Cell; Filament; Growth; In Vitro; Ions; Knowledge; Laboratories; Link; Magnetism; Mechanics; Medical; Methods; Microfilaments; Molecular; Molecular Conformation; Molecular Models; Motor; Mutate; Myosin ATPase; Organism; Phosphorylation; Physics; Physiology; Plastics; Play; Post-Translational Protein Processing; Probability; Property; Proteins; Publishing; Recruitment Activity; Regulation; Research; Research Activity; Resolution; Role; Shapes; Stretching; Structure; System; Testing; Translating; Work; biophysical analysis; biophysical techniques; cell motility; chemical property; cofilin; density; experience; experimental analysis; genetic regulatory protein; in vivo; insight; mathematical model; models and simulation; molecular modeling; novel; pathogenic bacteria; physical property; polymerization; predictive modeling; protein function; public health relevance; sensor

 DESCRIPTION (provided by applicant): Polymerization of the protein actin into helical filaments powers the directed motility of eukaryotic cells and some pathogenic bacteria. Actin assembly also plays critical roles in endocytosis, cytokinesis, and establishment of cell polarity. The essential regulatory protein, cofilin, is one of four actin-binding proteins that precisely choreograph actin assembly and organization in living systems. It acts by severing filaments, which increases the concentration of filament ends available for subunit addition and dissociation, thereby accelerating overall actin network dynamics and reorganization. It is therefore of general medical importance to understand how cofilin fragments actin filaments. Although the effects of cofilin binding to actin filaments have been extensively studied, the molecular mechanism of how cofilin severs filaments, which have stiffness comparable to commercial laboratory plastics, remains a central and unresolved mystery of cellular actin cytoskeleton reorganization. Elucidating the cofilin severing mechanism demands a multi-disciplinary approach integrating biology, chemistry, physics and mathematical modeling. Proposed research efforts focus on identifying how specific cation binding, post-translational modification, competition with other regulatory proteins, and filament shape deformations modulate actin filament structure and severing by vertebrate cofilin. Five general hypotheses will be tested. The first is that vertebrate cofilin severs filaments by dissociating a specific filamen-associated cation that controls filament structure and mechanical properties. The second is that competitive displacement of cofilin by other filament binding proteins can promote cofilactin filament severing by introducing boundaries of bare and cofilin-decorated segments. The third is that phosphorylation enhances cooperative cofilin binding and inhibits severing, not by lowering cofilin occupancy along filaments, but by reducing the density of boundaries where severing can occur. The fourth is that contractile protein- driven deformations in filament shape enhance severing by cofilin. The fifth is that actin filaments can act as tension sensors that recruit or exclude cofilin depending on the magnitude and mode of filament shape deformation. We will integrate biochemical and biophysical approaches, including experimental manipulation of single filaments, with mathematical modeling and simulations to develop predictive molecular models of actin filament elasticity and fragmentation, and directly test hypotheses formulated from biochemical and biophysical analysis of cofilin-actin interactions completed during the prior award period. The proposed research activities will advance knowledge of actin filament physiology by providing multi-scale relationships between filament mechanics, structure, and the biological function (e.g. severing activity) of essential regulatory proteins. New experimental and methods of analysis readily applicable to other filament binding proteins will be developed. Novel insight regarding the relationship between actin filament elasticity, conformation and regulatory protein occupancy will emerge from the work.

 描述(申请人提供)：将蛋白质肌动蛋白聚合成螺旋细丝，为真核细胞和一些致病细菌的定向运动提供动力。肌动蛋白组装也在内吞作用、胞质分裂和细胞极性的建立中起着关键作用。 关键的调节蛋白cofilin是四种肌动蛋白结合蛋白之一，它精确地编排了肌动蛋白在生命系统中的组装和组织。它的作用是切断细丝，从而增加可用于亚基添加和解离的细丝末端的浓度，从而加速整个肌动蛋白网络的动态和重组。因此，了解Cofilin如何使肌动蛋白细丝断裂具有普遍的医学意义。虽然cofilin与肌动蛋白细丝结合的作用已被广泛研究，但cofilin如何切断肌动蛋白细丝的分子机制仍然是细胞肌动蛋白细胞骨架重组的中心和未解之谜。阐明cofilin的切割机制需要综合生物学、化学、物理学和数学模型的多学科方法。建议的研究工作集中在确定特定的阳离子结合，翻译后修饰，与其他调节蛋白的竞争，以及细丝形状变形如何调节肌动蛋白细丝结构，并被脊椎动物的cofilin切断。五个一般性的假设将被检验。第一种是脊椎动物的纤丝蛋白通过解离特定的细丝相关阳离子来切断细丝，这种阳离子控制着细丝的结构和机械性能。第二种是其他微丝结合蛋白竞争性置换粘连蛋白，通过引入裸段和修饰粘连蛋白片段的边界来促进粘连蛋白细丝的断裂。第三种是，磷酸化增强了协同胶丝蛋白结合并抑制了切割，这不是通过降低沿着纤维的胶丝蛋白占有率，而是通过降低发生切割的边界密度来实现的。第四，收缩蛋白驱动的细丝状变形增强了粘连蛋白的切断作用。第五，肌动蛋白微丝可以作为张力传感器，根据微丝形状变形的大小和方式招募或排除粘连蛋白。我们将把生化和生物物理方法，包括对单丝的实验操作，与数学建模和模拟相结合，以开发肌动蛋白细丝弹性和断裂的预测分子模型，并直接测试在之前获奖期间完成的对粘连蛋白-肌动蛋白相互作用的生化和生物物理分析得出的假说。拟议的研究活动将通过提供肌丝机械、结构和基本调节蛋白的生物学功能(例如，切断活性)之间的多尺度关系，来促进对肌动蛋白细丝生理学的了解。新实验 并将开发适用于其他细丝结合蛋白的分析方法。这项工作将对肌动蛋白细丝弹性、构象和调节蛋白占有率之间的关系产生新的见解。