Titin in Skeletal Muscle Health and Disease

肌联蛋白在引起神经肌肉疾病中的重要性已被越来越多的人所认识 （肌联蛋白病），其中两个，我们集中在这项工作中，中枢性肌病（CNM）和遗传性肌病 早期呼吸衰竭（HMERF）Titinopathies经常表现为早发性肌无力， 呼吸困难，但了解其潜在的机制是阻碍了我们仍然有限的 了解肌联蛋白在骨骼肌中的功能作用。肌联蛋白构成肌肉的第三条肌丝 并沿肌节沿着从Z盘到M带。Titin的I带区域起着分子弹簧的作用 最近的研究表明，被动刚度影响主动力， 最大激活水平。然而，目前还不知道肌联蛋白的硬度对整个骨骼肌有多重要 健康和如果改变僵硬可能会导致肌病。重要性可能很高是由我们的 初步研究揭示了患有CNM的肌联蛋白病患者的肌联蛋白刚度紊乱和主动张力降低。 目的1和2解决如何改变肌联蛋白为基础的刚度影响被动和主动肌肉属性，如果 它可能是致病的。我们将对肌钙蛋白病患者的活检进行机械研究（重点是 CNM）以及研究两种对比小鼠模型，其中肌联蛋白弹簧区域的刚度是 增加（PEVK截短）或减少（Rbm20失活）。目前的假设是改变肌联蛋白的 弹簧区改变被动肌肉僵硬和主动张力，这导致肌病。目的2 还研究了一种新的小鼠模型，该模型模拟了肌联蛋白弹簧区剪接位点突变的CNM， 将检查在RNA，蛋白质，结构和功能水平的功能后果。我们还研究 肌联蛋白的A带片段，特别是C带。该区域尚未被研究，它在临床上很重要， 这是发现大量致病突变的地方。我们在目标3中研究了 C区外显子343引起HMERF，这是一种呼吸肌受累的肌病， 致命的使用一种新的HMERF小鼠，我们将研究该疾病的机制基础。 通过切除突变的肌联蛋白外显子343，外显子跳跃治疗HMERF的治疗潜力 会得到考验凭借其基础科学和转化目标及其深入和综合的方法， 这项申请旨在继续我们的尖端titin研究的跟踪记录。强大的技术和 新的小鼠模型已经到位，试点数据支持指导假设，我们的研究团队正在 经验丰富该提案将极大地提高对肌联蛋白生物学，肌联蛋白在肌肉疾病中的作用， 和肌联蛋白作为治疗靶点的潜力。

There is a rapidly increasing awareness of the importance of titin in causing neuromuscular disorders (titinopathies), two of which we focus on in this work, centronuclear myopathy (CNM) and hereditary myopathy with early respiratory failure (HMERF). Titinopathies frequently present early-onset muscle weakness and respiratory difficulty but understanding their underlying mechanisms is held back by our still limited understanding of the functional roles of titin in skeletal muscle. Titin comprises the third myofilament of muscle and spans along the sarcomere, from Z-disk to M-band. Titin’s I-band region functions as a molecular spring that generates passive stiffness with recent studies indicating that passive stiffness affects active force at sub- maximal activation levels. However, it is not known how important titin’s stiffness is to overall skeletal muscle health and if altered stiffness can cause a myopathy. That the importance might be high is suggested by our pilot studies that reveal deranged titin stiffness and reduced active tensions in titinopathy patients with CNM. Aims 1 and 2 address how altering titin-based stiffness affects both passive and active muscle properties and if it can be disease-causing. We will perform mechanical studies on biopsies from titinopathy patients (focus on CNM) as well as study two contrasting mouse models in which the stiffness of titin’s spring region is either increased (PEVK truncation) or reduced (inactivation of Rbm20). The working hypothesis is that altering titin’s spring region alters both passive muscle stiffness and active tension and that this causes myopathy. Aim 2 also studies a novel mouse model that mimics CNM with splice site mutations in titin’s spring region and we will examine the functional consequences at the RNA, protein, structural and functional levels. We also study the A-band segment of titin, specifically the C-zone. This zone has not been studied, it is clinically important as this is where a large number of disease-causing mutations are found. We investigate in Aim 3 a mutation in the C-zone exon 343 which causes HMERF, a myopathy with respiratory muscle involvement that can be fatal. Using a novel HMERF mouse that we made the mechanistic basis of the disease will be studied. Through excision of the mutated titin exon 343, the therapeutic potential of exon skipping for treating HMERF will be tested. With its basic science and translational goals and its in-depth and integrative approach, this application seeks to continue our track record of cutting-edge titin research. Powerful techniques and novel mouse models are in place, pilot data support the guiding hypotheses, and our research team is highly experienced. The proposal will greatly enhance insights in titin biology, titin’s role in muscle disease, and titin’s potential as a therapeutic target.