Changes in myosin biomechanics influence growth and maturation of iPSC-cardiomyocytes.

Abstract

Hypertrophic cardiomyopathy (HCM) is characterized by hypercontractility, cardiomyocyte hypertrophy, myofibril disarray, and altered energetics. Despite the identification of >1000 mutations in sarcomeric proteins, one-third of which are in β-cardiac myosin (MYH7), the mechanism by which altered force at the level of the sarcomere is transduced into cellular hypertrophy and other phenotypes is still incompletely understood. Our biochemical studies reveal surprising heterogeneity in the impact of different MYH7 mutations on myosin force generation and ATPase activity with some mutations reducing activity at the molecular level. Using CRISPR-edited hiPSC-cardiomyocytes (hiPSC-CMs), we studied the impact of myosin HCM mutations on the generation of contractile forces, hypertrophy, myofibril organization, and mitochondrial function, and compared cellular phenotypes with the effects of the mutations on myosin biomechanics. Our studies reveal a critical role for alterations in the myosin super-relaxed state (SRX) as a mechanism for cellular hypercontractility. To validate the fidelity of our findings to human disease, we compared hiPSC-CM results to those in 26 septal myectomy samples from HCM patients and 13 donor heart controls. In both hiPSC-CMs and myectomy samples we observed fiber disarray, altered mitochondrial metabolism, and widening of the Z-disc. A genome-wide key driver analysis revealed altered expression of 52 Z-disc-related genes in patient samples; 24 Z-disc genes in hiPSC-CMs; with 14 Z-disc genes shared between the two platforms. Activation of calcineurin pro-hypertrophic signaling provides a potential link between Z-disc mechano-sensation of sarcomeric forces and downstream hypertrophic signaling, which we explore using a vinculin FRET-tension sensor to directly measure intracellular force at the Z-disc. Our multi-scale approach, combined with both patient samples and CRISPR-edited hiPSC-CMs, confirms the fidelity of our in vitro model in recapitulating human disease and increases the potential translation of our findings to develop new precision-based HCM therapies.