PDF(Pubmed)
Abstract：
Determining the pathogenicity of hypertrophic cardiomyopathy-associated mutations in the β-myosin heavy chain (MYH7) can be challenging due to its variable penetrance and clinical severity. This study investigates the early pathogenic effects of the incomplete-penetrant MYH7 G256E mutation on myosin function that may trigger pathogenic adaptations and hypertrophy. We hypothesized that the G256E mutation would alter myosin biomechanical function, leading to changes in cellular functions. We developed a collaborative pipeline to characterize myosin function across protein, myofibril, cell, and tissue levels to determine the multiscale effects on structure-function of the contractile apparatus and its implications for gene regulation and metabolic state. The G256E mutation disrupts the transducer region of the S1 head and reduces the fraction of myosin in the folded-back state by 33%, resulting in more myosin heads available for contraction. Myofibrils from gene-edited MYH7WT/G256E human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) exhibited greater and faster tension development. This hypercontractile phenotype persisted in single-cell hiPSC-CMs and engineered heart tissues. We demonstrated consistent hypercontractile myosin function as a primary consequence of the MYH7 G256E mutation across scales, highlighting the pathogenicity of this gene variant. Single-cell transcriptomic and metabolic profiling demonstrated upregulated mitochondrial genes and increased mitochondrial respiration, indicating early bioenergetic alterations. This work highlights the benefit of our multiscale platform to systematically evaluate the pathogenicity of gene variants at the protein and contractile organelle level and their early consequences on cellular and tissue function. We believe this platform can help elucidate the genotype-phenotype relationships underlying other genetic cardiovascular diseases.
摘要：
确定β-肌球蛋白重链（MYH7）中肥厚型心肌病相关突变的致病性可能具有挑战性，因为其外显率和临床严重程度不同。本研究调查了不完全渗透MYH7G256E突变对肌球蛋白功能的早期致病作用，该作用可能引发致病性适应和肥大。我们假设G256E突变会改变肌球蛋白的生物力学功能，导致细胞功能的变化。我们开发了一个协作管道来表征肌球蛋白跨蛋白质的功能，肌原纤维,cell,和组织水平，以确定对收缩器结构功能的多尺度影响及其对基因调节和代谢状态的影响。G256E突变破坏了S1头部的换能器区域，并将折回状态下的肌球蛋白分数降低了33％，导致更多的肌球蛋白头可用于收缩。来自基因编辑的MYH7WT/G256E人诱导的多能干细胞衍生的心肌细胞（hiPSC-CM）的肌原纤维表现出更大和更快的张力发展。这种过度收缩表型在单细胞hiPSC-CM和工程化心脏组织中持续存在。我们证明了一致的过度收缩肌球蛋白功能是MYH7G256E突变的主要结果，突出该基因变异的致病性。单细胞转录组学和代谢谱分析表明线粒体基因上调和线粒体呼吸增加，表明早期生物能量改变。这项工作强调了我们的多尺度平台的好处，以系统地评估蛋白质和收缩细胞器水平的基因变体的致病性及其对细胞和组织功能的早期后果。我们相信这个平台可以帮助阐明其他遗传性心血管疾病的基因型-表型关系。
