Effects of MYBPC3 loss-of-function mutations preceding hypertrophic cardiomyopathy
Effects of MYBPC3 loss-of-function mutations preceding hypertrophic cardiomyopathy
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DOI：
10.1172/jci.insight.133782
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发表时间：
2020-01-30
期刊：
影响因子：
8
通讯作者：
Day, Sharlene M.
中科院分区：
文献类型：
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作者：
Helms, Adam S.;Tang, Vi T.;Day, Sharlene M.
心肌肌球蛋白结合蛋白C（MyBP-C，由MYBPC 3编码）的突变是肥厚型心肌病（HCM）的最常见原因。大多数MYBPC 3突变导致过早终止密码子（PTC），导致HCM患者心脏中RNA降解和MyBP-C减少。然而，在MYBPC 3突变体诱导的多能干细胞心肌细胞（iPSCMs）中并未一致观察到MyBP-C的减少。为了确定早期MYBPC 3突变的影响，我们使用了患者和基因组工程iPSCM。将具有移码突变的IPSCM与具有MYBPC 3启动子和翻译起始位点缺失的IPSCM进行比较，揭示等位基因功能丧失是导致PTC的突变的主要激发后果。尽管在所有杂合子iPSCM中野生型mRNA减少，但没有观察到MyBP-C蛋白减少，表明蛋白质水平的补偿通过我们认为是以前未表征的机制。虽然纯合突变iPSCM表现出收缩失调，但杂合突变iPSCM在补偿MyBP-C水平的背景下具有正常的收缩功能。不可知的RNA-Seq分析揭示了参与蛋白质折叠的基因中的差异表达作为唯一的失调基因集。为了确定MYBPC 3突变iPSCM如何实现补偿的MyBP-C水平，用稳定同位素标记测量肌节蛋白合成和降解。杂合突变iPSCM显示MyBP-C合成速率降低，但MyBP-C降解速率较慢。这些发现表明，尽管由于截短突变导致MYBPC 3等位基因功能丧失，但心肌细胞具有获得正常MyBP-C化学计量的先天能力。调节MyBP-C降解以维持MyBP-C蛋白水平可能是HCM收缩功能障碍上游的新治疗方法。
Mutations in cardiac myosin binding protein C (MyBP-C, encoded by MYBPC3) are the most common cause of hypertrophic cardiomyopathy (HCM). Most MYBPC3 mutations result in premature termination codons (PTCs) that cause RNA degradation and a reduction of MyBP-C in HCM patient hearts. However, a reduction in MyBP-C has not been consistently observed in MYBPC3-mutant induced pluripotent stem cell cardiomyocytes (iPSCMs). To determine early MYBPC3 mutation effects, we used patient and genome-engineered iPSCMs. IPSCMs with frameshift mutations were compared with iPSCMs with MYBPC3 promoter and translational start site deletions, revealing that allelic loss of function is the primary inciting consequence of mutations causing PTCs. Despite a reduction in wild-type mRNA in all heterozygous iPSCMs, no reduction in MyBP-C protein was observed, indicating protein-level compensation through what we believe is a previously uncharacterized mechanism. Although homozygous mutant iPSCMs exhibited contractile dysregulation, heterozygous mutant iPSCMs had normal contractile function in the context of compensated MyBP-C levels. Agnostic RNA-Seq analysis revealed differential expression in genes involved in protein folding as the only dysregulated gene set. To determine how MYBPC3-mutant iPSCMs achieve compensated MyBP-C levels, sarcomeric protein synthesis and degradation were measured with stable isotope labeling. Heterozygous mutant iPSCMs showed reduced MyBP-C synthesis rates but a slower rate of MyBP-C degradation. These findings indicate that cardiomyocytes have an innate capacity to attain normal MyBP-C stoichiometry despite MYBPC3 allelic loss of function due to truncating mutations. Modulating MyBP-C degradation to maintain MyBP-C protein levels may be a novel treatment approach upstream of contractile dysfunction for HCM.