Multiscale investigation of cardiomyopathy-associated mutations in metavinculin

美维库林心肌病相关突变的多尺度研究

基本信息

批准号：
10403271
负责人：
Samantha Kirstin Barrick
金额：
$ 0.25万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-02-01 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10403271
关键词：
Actins Address Affect Amino Acids Area Attention Binding Biological Assay Biological Process Cardiac Cardiac Myocytes Cardiomyopathies Cell-Matrix Junction Cells Coupling Dependence Development Dilated Cardiomyopathy Disease Educational process of instructing Extracellular Matrix Fellowship Generations Heart Heart Diseases Human Hydrogels Impairment Individual Induced Mutation Intercalated disc Intercellular Junctions Investigation Lead Ligands Measurement Measures Mediating Microfilaments Mission Molecular Muscle Muscle Cells Mutation Organizational Change Pathogenicity Pathologic Patients Physiological Play Process Protein Isoforms Proteins RNA Splicing Research Role Sarcomeres Site Stress Structure Tail Testing Traction Traction Force Microscopy Training United States National Institutes of Health Universities VCL gene Ventricular Remodeling Vinculin Washington alpha helix biophysical techniques disease phenotype disease-causing mutation education research experimental study induced pluripotent stem cell inherited cardiomyopathy live cell imaging live cell microscopy mechanical force mechanotransduction medical schools mouse model mutant novel therapeutics optical traps programs research and development single molecule skills stem cells transmission process

项目摘要

Project Summary/Abstract Familial cardiomyopathies are genetic heart diseases that involve ventricular remodeling and altered cardiac contractility. These diseases are often caused by mutations in proteins within the sarcomere, the fundamental contractile unit of cardiomyocytes. Mutations in non-sarcomeric proteins involved in mechanotransduction, the process by which cells sense and respond to mechanical force, have also been implicated in cardiomyopathy but have received considerably less attention. For instance, studies of human patients have identified cardiomyopathy-associated mutations in metavinculin, the muscle-specific isoform of the ubiquitous mechanotransducer vinculin, but how these mutations lead to the disease phenotype is not well-understood. Structural studies have shown that the 68-amino acid insert that differentiates metavinculin from vinculin replaces the first alpha-helix in the actin-binding vinculin tail domain. Although this alpha-helix does not directly bind actin, the metavinculin insert results in drastic changes of the organization of actin filaments by metavinculin compared to vinculin, suggesting an allosteric effect on actin binding. The proposed research will test the hypothesis that the pathogenic mechanism of cardiomyopathy caused by mutations in metavinculin involves disruption of cardiac mechanotransduction through impairment of the force-dependent binding of metavinculin to actin. Single- molecule force measurements of metavinculin binding to actin will directly address whether force stabilizes binding of metavinculin to actin, as has been previously demonstrated for vinculin, as well as the effect of pathogenic mutations on this force-dependent binding. The cellular consequences of altered force dependence of metavinculin-actin binding will be investigated in stem cell-derived cardiomyocytes that carry the disease- causing mutations using live-cell imaging of sarcomerogenesis (the establishment of new sarcomeres) and traction force microscopy. These experiments will also be carried out on metavinculin-null cardiomyocytes to elucidate the role of metavinculin in sarcomerogenesis and cellular contractility. The training provided under this fellowship will take place at the Washington University School of Medicine, a world leader in biomedical education and research. The proposed research aligns with the strategic objectives of the NIH by addressing the normal biological function of metavinculin and the pathobiological mechanism underlying the onset and progression of cardiomyopathy caused by mutations in metavinculin. In addition, the proposed training plan will contribute to the strategic objective of developing a scientific workforce capable of accomplishing the NIH’s mission by supporting the development of research, teaching, and professional skills required for the PI to establish a successful independent research program in the field of cardiac mechanobiology.

项目概要/摘要家族性心肌病是遗传性心脏病，涉及心室重塑和心脏改变这些疾病通常是由肌节内的蛋白质突变引起的，肌节是基本的蛋白质。参与机械转导的非肌节蛋白的突变。细胞感知和响应机械力的过程也与心肌病有关但受到的关注却少得多，例如，对人类患者的研究发现。心肌病相关的美维库林突变，这是普遍存在的肌肉特异性亚型机械转导纽蛋白，但这些突变如何导致疾病表型尚不清楚。结构研究表明，区分metavinculin 和vinculin 的68 个氨基酸插入物取代了肌动蛋白结合纽蛋白尾部结构域中的第一个 α 螺旋虽然该 α 螺旋不直接结合肌动蛋白，与metavinculin相比，metavinculin插入导致肌动蛋白丝组织发生巨大变化纽蛋白，表明对肌动蛋白结合的变构作用。拟议的研究将检验以下假设：美维库林突变引起的心肌病的致病机制涉及心脏功能的破坏通过破坏metavinculin 与肌动蛋白的力依赖性结合来进行机械转导。 Metavinculin 与肌动蛋白结合的分子力测量将直接解决力是否稳定的问题 Metavinculin 与肌动蛋白的结合（如之前针对纽蛋白所证明的那样），以及这种力依赖性结合的致病突变。力依赖性的细胞后果。将在携带疾病的干细胞衍生的心肌细胞中研究metavinculin-actin结合的情况- 使用肌节发生的活细胞成像（新肌节的建立）引起的突变和这些实验还将在无美维库林的心肌细胞上进行。阐明美维库林在肌瘤生成和细胞收缩性中的作用。奖学金将在生物医学教育领域世界领先的华盛顿大学医学院进行拟议的研究通过解决正常问题与 NIH 的战略目标保持一致。美维库林的生物学功能以及疾病发生和进展的病理生物学机制此外，所提出的训练计划将有助于由metavinculin突变引起的心肌病。培养一支能够完成 NIH 使命的科学队伍的战略目标支持 PI 建立所需的研究、教学和专业技能的发展心脏机械生物学领域成功的独立研究项目。