Costamere Defects in Muscular Dystrophies

描述(由申请人提供)：肋骨是横纹肌细胞中肌膜下的蛋白质集合，在物理上将产生力量的肌节与肌膜相连。由于在人类肌肉营养不良和扩张型心肌病中，几种组成蛋白是主要的缺陷部位，因此，衣壳对于正常的肌肉功能是很重要的。我们先前证明，胞核细胞骨架富含非肌肉“细胞质”细胞肌动蛋白，肌肉特异性的细胞肌动蛋白消融导致一种新形式的进行性肌病。这次更新的主要目的是阐明骨骼肌中细胞肌动蛋白亚型和细胞肌动蛋白亚型的重要但知之甚少的功能。我们将利用我的团队在最初的项目期间开发的新的条件性动物模型和异构体特异性试剂来解决细胞质肌动蛋白在正常骨骼肌功能和人类骨骼肌组织疾病中的几个基本问题。在目标1中，我们将研究细胞肌动蛋白在骨骼肌力学功能中的具体作用，这也可能有助于揭示正常肌肉和dystrophin缺陷肌肉收缩损伤的机制。在目标2中，将通过在骨骼肌中特定敲除细胞肌动蛋白或细胞和细胞肌动蛋白的小鼠品系的特征来研究细胞肌动蛋白和细胞肌动蛋白在发育中和成年骨骼肌中的不同和重叠的作用，特别是在尾部骨骼肌中。在目标3中，我们将在体内和体外测试细胞肌动蛋白异常定位在脊髓性肌萎缩症发病机制中的假设作用。拟议的研究结果将最明确地阐述细胞质肌动蛋白亚型对正常和疾病骨骼肌功能的独特和重要贡献。 公共卫生相关性：肌节肌动蛋白亚型因其在骨骼肌和心肌收缩中的重要作用而广为人知。然而，非肌浆肌动蛋白异构体在特殊肌肉结构的发育/维持中的作用以及它们在骨骼肌疾病中的作用也很重要，但仍然知之甚少。通过对转基因小鼠骨骼肌的严格表征，结合对分离的细胞和蛋白质的补充实验，选择性地消除其中一个或两个非肌肉胞浆肌动蛋白，拟议的研究将最明确地解决非肌肉胞浆肌动蛋白亚型对正常和患病骨骼肌功能的独特和重要贡献。特别是，这个项目对于了解Duchenne肌营养不良症和脊髓性肌萎缩的病理机制具有很高的相关性。

DESCRIPTION (provided by applicant): Costameres are subsarcolemmal protein assemblies in striated muscle cells that physically couple force-generating sarcomeres with the sarcolemma. Costameres are clearly important for normal muscle function because several constituent proteins are the primary sites of defect in human muscular dystrophies and dilated cardiomyopathies. We previously demonstrated that the costameric cytoskeleton is enriched in non-muscle "cytoplasmic" ?cyto-actin and that muscle-specific ablation of ?cyto-actin causes a novel form of progressive myopathy. The major objective of this renewal is to elucidate the important, but poorly understood functions of both ¿cyto- and ?cyto-actin isoforms in skeletal muscle. We will make use of novel conditional animal models and isoform-specific reagents generated by my group during the original project period to address several fundamental questions about cytoplasmic actins in normal skeletal muscle function and in human diseases of skeletal muscle tissue. In aim 1, we will investigate the specific contribution of ?cyto-actin to the mechanical function of skeletal muscle, which may also shed light into the mechanism of contraction-induced injury of normal and dystrophin-deficient muscle. In aim 2, the distinct and overlapping roles of ¿cyto- and ?cyto- actins in developing and adult skeletal muscle, particularly in costameres, will be investigated through the characterization of mouse lines in which either ¿cyto-actin, or ¿cyto- and ?cyto-actins have been knocked out specifically in skeletal muscle. In aim 3, we will test the hypothesized role for aberrant ¿cyto-actin localization in the pathogenesis of spinal muscular atrophy both in vivo and in vitro. The results of the proposed studies will most definitively address the unique and important contributions of cytoplasmic actin isoforms to the function of normal and diseased skeletal muscle. PUBLIC HEALTH RELEVANCE: The sarcomeric actin isoforms are well known for their important role in contraction of skeletal and cardiac muscle. However, the roles of non-muscle cytoplasmic actin isoforms in the development/maintenance of specialized muscle structures and their contribution to diseases of skeletal muscle are also important, but remain poorly understood. Through rigorous characterization of skeletal muscle in genetically-modified lines of mice where one or both non-muscle cytoplasmic actins are selectively eliminated in combination with complementary experiments on isolated cells and proteins, the proposed studies will most definitively address the unique and important contributions of non-muscle cytoplasmic actin isoforms to the function of normal and diseased skeletal muscle. In particular, this project is highly relevant to understanding the pathological mechanism of Duchenne muscular dystrophy and spinal muscular atrophy.