Abstract 438: Familial Cardiomyopathy Mutations Affect Mechanosensing at the Molecular and Cellular Levels

Familial cardiomyopathies are the leading cause of sudden cardiac death in young people, affecting more than 1 in 500 people. Two common forms, hypertrophic (HCM) and dilated (DCM) cardiomyopathies, are characterized by remodeling of the heart tissue, frequently accompanied by fibrosis and myocyte disarray. These diseases are primarily caused by mutations in sarcomeric proteins that regulate the contraction of the heart; however, it is not clear how these point mutations at the molecular level lead to the disease phenotype seen in patients. To better understand this process, we have examined the molecular and cellular consequences of two mutations in troponin T, R92Q and ΔK210, that cause HCM and DCM, respectively. Using recombinant proteins and a battery of in vitro biochemical and biophysical techniques, we find that R92Q increases activation of the thin filament by stabilizing myosin force generating states while ΔK210 reduces the ability of myosin to access these states at submaximal calcium levels. To study these mutations at the single cell level, we used CRISPR/Cas9 to generate human stem cell derived cardiomyocytes bearing these mutations. We examined the structural and contractile properties of single cells on engineered hydrogels that mimic either healthy or diseased hearts. Consistent with our molecular studies, we find that R92Q causes increased force production and power output. Interestingly, the ΔK210 mutation does not affect cellular force or power output on substrates that mimic the stiffness of the healthy heart, but it causes disordering of the sarcomeres on stiff substrates. This suggests a mechanism for ΔK210 where fibrosis leads to progressive loss of contractile function via mechanosensitive structural remodeling. Taken together, these results (1) demonstrate that alterations in force production and mechanosensing contribute to the disease pathogenesis in familial cardiomyopathies and (2) illuminate the differential mechanisms that lead to either HCM or DCM.