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Effects of MYBPC3 loss-of-function mutations preceding hypertrophic cardiomyopathy
Adam S. Helms, … , Michael J. Previs, Sharlene M. Day
Adam S. Helms, … , Michael J. Previs, Sharlene M. Day
Published December 26, 2019
Citation Information: JCI Insight. 2020;5(2):e133782. https://doi.org/10.1172/jci.insight.133782.
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Research Article Cardiology

Effects of MYBPC3 loss-of-function mutations preceding hypertrophic cardiomyopathy

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Abstract

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.

Authors

Adam S. Helms, Vi T. Tang, Thomas S. O’Leary, Sabrina Friedline, Mick Wauchope, Akul Arora, Aaron H. Wasserman, Eric D. Smith, Lap Man Lee, Xiaoquan W. Wen, Jordan A. Shavit, Allen P. Liu, Michael J. Previs, Sharlene M. Day

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Figure 3

Intercellular alignment of iPSCMs in micropatterned 2D tissues is not disrupted by MYBPC3 mutations.

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Intercellular alignment of iPSCMs in micropatterned 2D tissues is not di...
Two-dimensional anisotropic cardiac tissues were generated by seeding iPSCMs onto row-micropatterned polydimethylsiloxane (PDMS) substrates. Because the row micropatterning induces cellular and myofibrillar alignment, the technique enables analysis of mutant impact on in vitro tissue formation. All the isogenic MYBPC3-mutant lines, including the homozygous line, formed well-aligned tissues, indicating that a tissue-level phenotype of disorganization does not occur in the isogenic MYBPC3 mutation iPSCM models. The intensity profiles depict alignment of adjacent iPSCMs in each 2D-tissue (cell-cell boundaries indicated by N-cadherin) as evidenced by registration of Z-bands (α-actinin) in adjacent cells. The solid and dotted lines in the graphs indicate the intensity profiles from the solid and dotted line arrows in the images. Scale bar: 10 μm; inset: ×4 magnification.

Copyright © 2023 American Society for Clinical Investigation
ISSN 2379-3708

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