BPS2026 – The dilated cardiomyopathy-associated MYH7 mutation R369Q exhibits varied contractile phenotypes in a multiscale study of stem cell-derived cardiomyocytes

Mutations in the sarcomeric protein β-myosin heavy chain (MHC) are commonly implicated in familial cardiomyopathies. Such mutations can lead to progressive remodeling of cardiac muscle, significantly impairing the function of the heart. Our lab aims to understand the link between molecular mechanisms of disease and cardiomyopathy phenotype. One mutation of interest, R369Q, is associated with dilated cardiomyopathy. Recent work by our collaborators and us, using targeted molecular dynamics, has suggested that the R369 residue is involved in repelling tropomyosin during the C-state to M-state transition, further exposing binding spots on actin for strong binding to myosin. To examine the effects of the R369Q mutation on contractile function experimentally, we leveraged a CRISPR-Cas9 gene-edited homozygous MYH7 R369Q/R369Q human induced pluripotent stem cell-derived cardiomyocyte (hiPSC-CM) model. In engineered heart tissue constructs, mutant tissues generated 51% as much twitch force as wild type tissues yet had unchanged activation and relaxation kinetics. By contrast, single-cell imaging experiments yielded no appreciable difference in average sarcomere shortening when comparing wild type and R369Q cells. At the organelle scale, mutant myofibrils generated 44% the specific force of wild type myofibrils and exhibited a slow phase of relaxation with a rate approximately 3.5 times faster than the wild type, suggesting that the thin filament deactivates more quickly in the R369Q mutant. More rapid relaxation kinetics imply faster crossbridge cycling, suggesting that ADP or phosphate release could be accelerated in R369Q myofibrils. To further examine how the R369Q mutation contributes to pathological contractile dysfunction, we are modeling the post-powerstroke state using molecular dynamics and utilizing stopped flow kinetics to understand changes in the rates of specific steps in the crossbridge cycle.