Abstract Body: Introduction:
Hypertrophic cardiomyopathy (HCM)–associated mutations in MYH7 alter sarcomere mechanics, yet how these mutations influence mechanobiological signaling in cardiomyocytes (CMs) remains poorly understood. The early-onset H251N β-cardiac myosin mutation increases actin motility, intrinsic force, and ATPase activity, consistent with a hypercontractile molecular phenotype. Here, we investigate the transmission of these forces from the cell to the matrix.
Hypothesis:
We hypothesized that H251N-associated increases in molecular motor activity would enhance focal adhesion (FA) organization and mechanobiological responses.
Methods:
Human induced pluripotent stem cell-derived CMs (hiPSC-CMs) carrying the H251N mutation and isogenic controls were seeded onto deformable substrates to mimic phyhsiological and pathological stiffness. A Förster resonance energy transfer (FRET)- based molecular tension sensor was employed to assess intracellular FA organization and adhesion-associated tension.
Results:
Two-way ANOVA revealed a significant interaction between genotype and substrate stiffness for FA area density (p < 0.0001), FRET index (p = 0.0121), and donor fluorescence intensity (p < 0.0001), indicating a stiffness-dependent effect of H251N (Figure 1). FA area density was lower in H251N relative to control on soft substrates but slightly higher on stiff substrates. Consistent with this pattern, FRET index was lower in H251N on soft substrates and higher on stiff substrates, while donor fluorescence intensity exhibited a similar but more pronounced stiffness-dependent shift.
Conclusion:
Our results show that H251N-associated increases in molecular motor activity are linked to stiffness-dependent alterations in FA remodeling and mechanotransductive coupling.
This finding supports our hypothesis that the H251N myosin mutation promotes mechanosensitive remodelling through cell substrate adhesion complexes.