M-Line Proteins and A-Band Assembly in Skeletal Muscle

骨骼肌中的 M 线蛋白和 A 带组装

• 批准号: 7576798
• 负责人: Aikaterini Kontrogianni-Konstantopoulos
• 金额: $ 27.98万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7576798
• 关键词: Adhesions; Adult; Ankyrins; Binding; Binding Sites; Biological Assay; Cells; Co-Immunoprecipitations; Complex; Cytoskeleton; DNA Library; Dominant-Negative Mutation; Electron Microscopy; Family; Gene Transfer; Head; Human; Immunoglobulin Domain; In Vitro; Knock-in Mouse; Knowledge; Laboratories; Maintenance; Mediating; Membrane; Molecular; Molecular Profiling; Muscle Cells; Muscle Contraction; Muscle Fibers; Mutagenesis; Myofibrillogenesis; Myofibrils; Myopathy; Myosin ATPase; N-terminal; Peptides; Physiological; Play; Positioning Attribute; Process; Protein Isoforms; Proteins; RNA; Research; Research Personnel; Role; Sarcomeres; Sarcoplasmic Reticulum; Signal Transduction; Signaling Protein; Skeletal Muscle; Structure; Surface Plasmon Resonance; Technology; Terminator Codon; Testing; Thick Filament; To specify; Two-Hybrid System Techniques; Work; Yeasts; connectin; in vivo; insight; knock-down; member; molecular array; myosin-binding protein C; nebulin; novel; obscurin; overexpression; postnatal; programs; scaffold; skeletal; tool; yeast two hybrid system

Myofibrillogenesis is a highly complex process that depends on the coordinated assembly and integration of a number of contractile, cytoskeletal and signaling proteins into regular arrays, the sarcomeres. Myosin Binding Protein C (MyBP-C), a protein associated with thick filaments, is believed to have both a structural and a regulatory role within the muscle cell, by contributing to the normal assembly and stabilization of thick filaments and modulating the number of myosin heads available for involvement in the contractile cycle. Obscurin, a giant myofibrillar protein, that closely surrounds Z-disks and M-lines, interacts specifically and directly with a novel isoform of MyBP-C slow, MyBP-C(+) slow, that appears to selectively concentrate at the M-line. Consequently, we hypothesize that the binding of obscurin with MyBP-C(+) slow contributes to the assembly, stabilization and maintenance of sarcomeric myosin into regular A-bands, through formation of primitive M-lines that may serve as scaffolding structures. We propose to test this hypothesis through three specific aims: (I) to characterize the novel MyBP-C(+) slow isoform in differentiating and adult skeletal muscle fibers, using a wide array of molecular, cellular and immunological approaches; (II) to study the interaction between obscurin and MyBP-C(+) slow qualitatively, by in vitro and in vivo binding assays, and quantitatively, by surface plasmon resonance technology and (III) to investigate the physiological significance of the binding of obscurin to MyBP-C(+) slow in the assembly and organization of myosin thick filaments into periodic A-bands, using adenovirally-mediated gene transfer and small inhibitory RNA technology. Knowledge gained from the proposed studies will provide new insights into the molecular mechanisms that integrate thick filaments into sarcomeres of normal skeletal fibers and how they are compromised in human muscle disease.

肌原纤维形成是一个高度复杂的过程，依赖于 一些收缩、细胞骨架和信号蛋白排列成规则的肌节。肌球蛋白 结合蛋白C(MyBP-C)是一种与粗丝相关的蛋白质，被认为既具有结构上的 以及在肌肉细胞内的调节作用，有助于正常组装和稳定厚壁 细丝和调节可用于参与收缩周期的肌球蛋白头的数量。 Obscurin是一种巨大的肌原纤维蛋白，紧密围绕着Z盘和M线，与 直接与一种新的MyBP-C慢速亚型MyBP-C(+)慢速结合，它似乎选择性地集中在 M线。因此，我们假设黑点蛋白与MyBP-C(+)的结合缓慢有助于 通过形成肌节肌球蛋白将肌球蛋白组装、稳定和维持为规则的A带 可用作脚手架结构的原始M线。我们建议通过三个方面来检验这一假设 具体目标：(I)鉴定新的MyBP-C(+)慢异构体在分化和成人骨骼中的作用 肌肉纤维，使用广泛的分子、细胞和免疫学方法；(Ii)研究 通过体外和体内结合试验，Oblcurin和MyBP-C(+)之间的相互作用定性地缓慢，并且 定量地，通过表面等离子激元共振技术和(III)研究生理 肌球蛋白厚度组装和组织过程中肌球蛋白与MyBP-C(+)结合缓慢的意义 使用腺病毒介导的基因转移和小抑制性RNA将细丝转化为周期性的A-带 技术从拟议的研究中获得的知识将提供对分子的新见解 将粗丝整合到正常骨骼纤维的肌节中的机制及其如何 在人类肌肉疾病方面受到了损害。