Thin Filaments and Muscle Regulation

细丝和肌肉调节

• 批准号: 6819255
• 负责人: WILLIAM J LEHMAN
• 金额: $ 32.3万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-09-30 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6819255
• 关键词: actins; animal tissue; calcium; caldesmon; cytoskeleton; digital imaging; dystrophin; electron microscopy; image processing; microfilaments; mitogen activated protein kinase; muscle function; muscle proteins; myocardium; phosphorylation; protein binding; protein protein interaction; protein reconstitution; protein structure function; smooth muscle; striated muscles; tropomyosin; troponin

EXCEED THE SPACE PROVIDED. Thin filament-associated actin-binding proteins control both actomyosin-based contractilityand cytoskeletal formation in muscle and non-muscle cells. To elucidate these mechanisms, it is crucial to determine the structural interactionsof the proteins involved. We address several interrelated questions fundamental to understanding muscle thin filament function: (1) What is the architecture of the thin filament in skeletal, cardiac and smooth muscles? (2) What are the changing structural interactionsof thin filament-linked proteinsthat regulate muscle activity? (3) How do these proteins interact with actin to form the muscle cytoskeleton? We use state-of-the-art electron microscopy, image analysis and 3-D reconstruction to establishthe macromolecular structureof components bound on thin filament actin. Using these techniques: (A) We aim to determine the structural basis of troponin-tropomyosinregulation of skeletal and cardiac muscle activity by analyzing interactions of tropomyosin and troponin on thin filaments and the effects of Ca2¿and myosin-crossbridgebinding. (B) To help understandthe organization of the cortical actin cytoskeletonof muscle cells, we aim to determine the structural interactions of dystrophin and utrophin with filamentous F-actin by examining the binding of their distinctcalponin homology (CH)-domains. (C) We aim to assess the structural role of thin filament- associated caldesmon as a possible modulator of actomyosin-based motility and cytoskeletal assembly in smooth muscle. (D) Studies of the structureof nebulin bound to actinwill be part of our continuing investigationof the functional design of thin filaments. In each study, reconstructionswill be fitted to the atomic resolution maps of F-actin to define molecular contacts of binding proteins with actin. Further, such "hybrid crystallography" will be used to fit newly solved crystal structures of troponin, tropomyosin, dystrophin and utrophin domains within EM density maps to attain near atomic resolution. Our ongoing studies on troponin-trepomyosin regulated filaments will lead to an elucidation of the molecular mechanism of relaxation and activation in skeletal and cardiac muscle. Our successfully initiated structural studies on utrophin and dystrophinwill establish their unique features as cytoskeletal elements, informationapplicable to designing genetic therapies for muscular dystrophy.Studies on smooth muscle filaments will contributeto understanding the fine-tuning of the smooth muscle contractile response. Such modulation affects vascular tone and pulmonary airway resistance, determinants in, e.g., hypertension and asthma. The wider significance of our goals is underscored by the role of actin and associated proteins indiverse and vital cellular mechanisms that can become aberrant in malignancy. PERFORMANCE SITE ========================================Section End===========================================

超出所提供的空间。薄的肌动蛋白相关肌动蛋白结合蛋白控制肌动球蛋白为基础的收缩性和细胞骨架的形成在肌肉和非肌肉细胞。为了阐明这些机制，确定相关蛋白质的结构相互作用至关重要。我们解决了几个相互关联的问题，了解肌肉细丝功能的基础：（1）骨骼肌，心肌和平滑肌中细丝的结构是什么？(2)调节肌肉活动的细丝连接蛋白的结构相互作用有哪些变化？(3)这些蛋白质如何与肌动蛋白相互作用形成肌肉细胞骨架？我们使用最先进的电子显微镜，图像分析和3-D重建来建立细丝肌动蛋白上的大分子结构。使用这些技术：（A）我们的目的是通过分析原肌球蛋白和肌钙蛋白在细肌丝上的相互作用以及Ca 2+和肌球蛋白的作用来确定肌钙蛋白-原肌球蛋白调节骨骼肌和心肌活动的结构基础。(B)为了帮助理解肌细胞的皮质肌动蛋白细胞骨架的组织，我们的目标是通过检测肌营养不良蛋白和肌营养不良蛋白与丝状F-肌动蛋白不同的钙调蛋白同源（CH）结构域的结合来确定它们的结构相互作用。(C)我们的目的是评估细丝相关钙调蛋白作为一种可能的平滑肌肌动蛋白为基础的运动和细胞骨架组装的调节剂的结构作用。(D)对与肌动蛋白结合的星云蛋白结构的研究将是我们继续研究细丝功能设计的一部分。在每项研究中，重构将适合于F-肌动蛋白的原子分辨率图，以定义结合蛋白与肌动蛋白的分子接触。此外，这种“混合晶体学”将用于在EM密度图内拟合肌钙蛋白、原肌球蛋白、肌营养不良蛋白和肌营养不良蛋白结构域的新解析的晶体结构，以获得近原子分辨率。我们正在进行的研究肌钙蛋白-环肌球蛋白调节的细丝将导致骨骼肌和心肌的松弛和激活的分子机制的阐明。我们对utrophin和dystrophin的结构研究将确立它们作为细胞骨架元素的独特特征，这些信息可用于设计肌营养不良症的基因疗法。对平滑肌细丝的研究将有助于理解平滑肌收缩反应的微调。这种调节影响血管张力和肺气道阻力，这些是例如，高血压和哮喘。肌动蛋白和相关蛋白质在恶性肿瘤中可能变得异常的多样化和重要的细胞机制中的作用强调了我们目标的更广泛的意义。性能现场=