MYOSIN GENE DIVERSITY AND FUNCTION

肌肉收缩需要有序的组装和功能 许多蛋白质的相互作用，其中许多是由 多种异构体。我们计划的重点是马达分子， 肌球蛋白，在哺乳动物中由至少10个基因编码。的程度 肌球蛋白基因家族仍不清楚，也不清楚其影响 肌球蛋白的组成对肌肉的发育和功能的影响。我们 建议使用酵母人工染色体来定义人类骨骼 17号染色体上的肌球蛋白重链（MHC）基因座。 我们将确定 我们发现的10个MHC基因的顺序、方向和同一性。 肌球蛋白的标志性功能之一是它能自我组装成粗丝。 负责肌球蛋白自我组装的序列开始被 但很明显，重组的肌球蛋白丝在 长度和组成。我们将调查 肌球蛋白粗丝组装的决定因素和粗丝的作用 在这个过程中，使用细胞培养物， 转染系统此外，我们还将描述 肌球蛋白突变对粗肌丝形成和功能的影响 分离的细胞和完整的肌肉。一个重要的问题是 肌球蛋白基因多样性是否提供功能特化，或 它是否代表系统内的冗余。为了解决这个 问题，我们将使用胚胎干细胞中的同源重组， 小鼠围产期骨骼MHC基因的克隆。这将使我们能够 确定围产期MHC基因是否是正常肌肉所必需的 发展和功能。肌球蛋白突变的鉴定 常染色体显性遗传性心脏病，家族性肥厚型心肌病 (FHC)使我们使用转基因方法来询问以下2个问题 问题：1.骨骼肌肌球蛋白的突变， 会导致骨骼肌疾病吗2.意志突变 心脏MHC分子中已知的功能域导致了 表型与FHC相似？这些方法应该使我们能够 了解肌肉发育，结构， 和功能

Muscle contraction requires the orderly assembly and functional interaction of numerous proteins, many of which are represented by multiple isoforms. The focus of our proposal is the motor molecule, myosin, which is encoded by at least 10 genes in mammals. The extent of the myosin gene family is still not known; nor is it clear what impact myosin composition has on the development and function of muscle. We propose to use yeast artificial chromosomes to define the human skeletal muscle myosin heavy chain (MHC) locus on chromosome 17. We will determine the order, orientation and identity of the 10 MHC genes we have found. One of myosin's hallmark functions is its self-assembly into thick filaments. The sequences responsible for myosin self assembly are beginning to be understood, but it is clear that reconstituted myosin filaments differ in length and composition from native thick filaments. We will investigate the determinants of myosin thick filament assembly and the roles of thick filament-associated proteins in this process, using a cell culture transfection system. in addition, we will characterize the impact of myosin mutations on thick filament formation and function in the context of both isolated cells and the intact muscle. An important question is whether myosin gene diversity provides functional specialization, or whether it represents redundancy within the system. To address this question, we will use homologous recombination in embryonic stem cells to inactivate the perinatal skeletal MHC gene in mice. This will allow us to determine whether the perinatal MHC gene is essential for normal muscle development and function. The identification of myosin mutations in the autosomal dominant heart disease, familial hypertrophic cardiomyopathy (FHC) has led us to use transgenic approaches to ask the following 2 questions: 1. Will mutations in skeletal myosin that are analogous to those found in FHC result in skeletal muscle disease? 2. Will mutation of known functional domains in the cardiac MHC molecule result in a dominant phenotype similar to that seen in FHC? These approaches should allow us to understand a number of important aspects of muscle development, structure and function.