Molecular Mechanisms of Actin Cytoskeleton Dynamics

肌动蛋白细胞骨架动力学的分子机制

• 批准号: 9187980
• 负责人: JIANPENG MA
• 金额: $ 34.08万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9187980
• 关键词: ANGPTL2 gene; ATP Hydrolysis; Actin-Binding Protein; Actins; Adenylyl Imidodiphosphate; Adopted; Binding; Binding Sites; Biological Process; Cell physiology; Cells; Communicable Diseases; Complex; Computer Simulation; Crystallization; Cytokinesis; Cytoskeleton; Development; Dimerization; Disease; Elongation Factor; F-actin-binding proteins; Family; Filament; Goals; Hereditary Disease; Homo; Infection; Investigation; Mediating; Molecular; Molecular Conformation; Morphogenesis; Morphology; Muscle Development; Muscular Dystrophies; Mutation; Myomatous neoplasm; Neoplasm Metastasis; Neurodegenerative Disorders; Pathogenicity; Process; Protein Fragment; Proteins; Refractory; Research; Role; Structure; Surface; Testing; Tropomyosin; base; cell motility; design; dimer; drug development; experimental study; human disease; innovation; member; monomer; mutant; neurogenesis; novel; prevent; profilin; public health relevance; success; tumor initiation; tumorigenesis

 DESCRIPTION (provided by applicant): The inability to obtain atomic structures on actin cytoskeleton has severely hindered our understanding of this most abundant eukaryotic protein and its dynamic turnovers in performing a myriad of cellular functions such as cell motility, cytokinesis and morphogenesis through interactions with hundreds of actin-binding proteins. This application describes an innovative double-mutant strategy to overcome this longstanding barrier in the actin cytoskeleton field. Indeed, this novel double-mutant strategy has enabled the PIs' successful determination of the very first atomic crystal structure of actin-nucleator complex (actin-Cobl complex). In addition, a similar approach has been used to solve the structure of actin with a bacterial effector, VopL. The observed non-filament-like conformation in actin-Cobl structure and filament-like conformation in actin-VopL structure together suggest that both types of conformation are fully accessible to an actin complex obtained via the double-mutant strategy, thus its true conformation is most likely preserved. The experiments proposed in this application will apply the double-mutant strategy to three most divergent members of the latest class actin nucleators characterized by the presence of tandem actin-binding sites. The goal is to decipher their molecular mechanisms of actin nucleation, the roles of ATP hydrolysis in their functional cycle, and how they collaborate with specific cellular components to fulfill their functions. This goal will be achieved in three Specific Aims using combined structural and functional approaches: Aim 1: Mechanisms of Cobl-mediated actin nucleation; Aim 2: Mechanistic study of actin nucleation by Lmod; and Aim 3: Mechanistic study of APC-mediated actin nucleation. The extensive preliminary studies presented in this application suggest a high promise of success for the proposed research. Results from this study will be significant not only in elucidating the molecular mechanisms of their respective roles in neurogenesis, muscle development and tumor initiation, but also in unraveling some long-sought-after general underpinnings for de novo actin nucleation, a process underlying every stage of mammalian development as well as many types of pathogenic infection. By providing a detailed atomic gallery of how de novo actin nucleation is accomplished and regulated, this study will stimulate deeper mechanistic investigations on these nucleators as well as discovery and characterization of new actin nucleators. More importantly, this application will validate the double-mutant strategy on proteins/protein fragments with and without actin nucleation activities, thus providing sufficient proof-of-principle for extending this approach to many other actin-involved biological processes beyond actin nucleation. Ultimately, the research along this line is expected to directly benefit the treatment of many forms of human diseases due to actin cytoskeleton malfunctions including neurodegenerative disorders, muscular dystrophy, tumorigenesis and metastasis.

 描述（应用程序提供）：无法获得肌动蛋白细胞骨架上的原子结构严重阻碍了我们对这种最丰富的真核蛋白的理解及其在执行无数细胞功能（例如细胞运动，细胞因子和形成性）中的动态失误，这是通过与数百种肌动蛋白结合蛋白相互作用的相互作用。该应用程序描述了一种创新的双重突变策略，以克服肌动蛋白细胞骨架领域的这一长期障碍。的确，这种新颖的双重突变策略使PIS成功地确定了肌动蛋白 - 核复合物的第一个原子晶体结构 （肌动蛋白-COBL复合体）。另外，使用细菌效应子VOPL来求解肌动蛋白的结构。在肌动蛋白-COBL结构中观察到的非丝样组成和肌动蛋白 - vopl结构中的细丝形成在一起表明，通过双重抗衡策略获得的肌动蛋白复合物可以完全访问两种类型的考虑，因此其真正的构象很可能保留得很可能。本应用程序中提出的实验将对以串联肌动蛋白结合位点存在为特征的最新类肌动蛋白成核器中的三个最不同的策略应用。目的是破译其肌动蛋白成核的分子机制，ATP水解在其功能周期中的作用以及它们如何与特定的细胞成分合作以实现其功能。使用结构和功能方法的三个特定目标将实现此目标：目标1：COBL介导的肌动蛋白成核的机理；目标2：LMOD肌动蛋白成核的机理研究；和目标3：APC介导的肌动蛋白成核的机理研究。本应用程序中介绍的广泛初步研究表明，拟议研究对成功的良好承诺。这项研究的结果不仅在阐明其在神经发生，肌肉发育和肿瘤倡议中的分子机制方面也将是重要的，而且还阐明了一些从头开始的肌动蛋白成核的一般基础，这是一个基于哺乳动物发育的每个阶段的过程。通过提供一个详细的原子画廊，了解如何完成和调节从头肌动蛋白成核，这项研究将刺激对这些成核器的更深层机械投资，以及新肌动蛋白成核器的发现和表征。更重要的是，该应用将验证有或没有肌动蛋白成核活性的蛋白质/蛋白质片段的双重突变策略，从而提供 改进的原理证明将这种方法扩展到肌动蛋白成核之外的许多其他参与肌动蛋白的生物学过程。最终，由于肌动蛋白细胞骨架功能不当，包括神经退行性疾病，肌肉营养不良，肿瘤发生和转移，预计沿该线的研究将直接受益于许多形式的人类疾病的治疗。