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: 7215657
• 负责人: Aikaterini Kontrogianni-Konstantopoulos
• 金额: $ 28.55万
• 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/7215657
• 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; Localized; Maintenance; Mediating; Membrane; Molecular; Molecular Profiling; Muscle Cells; Muscle Contraction; Muscle Fibers; Mutagenesis; Myofibrillogenesis; Myofibrils; Myopathy; Myosin ATPase; N-terminal; Numbers; Peptides; Physiological; Play; Positioning Attribute; Process; Protein Isoforms; Protein Overexpression; Proteins; RNA; Research; Research Personnel; Role; Sarcomeres; Sarcoplasmic Reticulum; Signal Transduction; Signaling Protein; Skeletal Muscle; Skeletal system; 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; postnatal; programs; scaffold; tool; yeast two hybrid system

DESCRIPTION (provided by applicant): 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.

描述（由申请人提供）：肌原纤维发生是一个高度复杂的过程，取决于许多收缩蛋白、细胞骨架蛋白和信号蛋白协调组装和整合成规则阵列（肌节）。肌球蛋白结合蛋白C（MyBP-C）是一种与粗肌丝相关的蛋白质，被认为通过促进粗肌丝的正常组装和稳定以及调节可用于参与收缩周期的肌球蛋白头的数量，在肌细胞内具有结构和调节作用。Obscurin是一种巨大的肌原纤维蛋白，紧密围绕Z盘和M线，与MyBP-C慢的新亚型MyBP-C（+）慢特异性直接相互作用，该亚型似乎选择性地集中在M线。因此，我们假设obscurin与MyBP-C（+）slow的结合有助于肌节肌球蛋白通过形成可作为支架结构的原始M线组装、稳定和维持成规则的A带。我们提出通过三个具体目标来验证这一假设：（I）使用广泛的分子、细胞和免疫学方法来表征分化和成年骨骼肌纤维中新的MyBP-C（+）慢亚型;（II）通过体外和体内结合试验定性和定量研究obscurin和MyBP-C（+）slow之间的相互作用，通过表面等离子体共振技术和（III）研究obscurin与MyBP-C（+）slow结合在肌球蛋白粗丝组装和组织成周期性A带中的生理意义，使用腺病毒介导的基因转移和小抑制RNA技术。从拟议的研究中获得的知识将为将粗丝整合到正常骨骼纤维的肌节中的分子机制以及它们如何在人类肌肉疾病中受到损害提供新的见解。