Abstract:Hypertrophic cardiomyopathies (HCMs) are the leading cause of acute cardiac failure in young individuals. Over 300 mutations throughout β-cardiac myosin, including in the motor domain, are associated with HCM. A β-cardiac myosin motor mutation (R712L) leads to a severe form of HCM. Actin-gliding motility of R712L-myosin is inhibited, despite near-normal ATPase kinetics. By optical trapping, the working stroke of R712L-myosin was decreased 4-fold, but actin-attachment durations were normal. A prevalent hypothes… Show more
“…As the effects of OM and P710R share some similarities, it is important to note that residue P710 is part of the OM binding pocket in the converter domain (65). A recent preprint reported a reduction in myosin's step size due to a different HCM mutation in the same region (P712L) (66). Taken together, these results emphasize the converter domain's critical facilitation of myosin's power stroke and load sensitivity and reveal various consequences of perturbing this domain.…”
Hypertrophic cardiomyopathy (HCM) is the most common inherited form of heart disease, associated with over 1,000 mutations, many in β-cardiac myosin (MYH7). Molecular studies of myosin with different HCM mutations have revealed a diversity of effects on ATPase and load-sensitive rate of detachment from actin. It has been difficult to predict how such diverse molecular effects combine to influence forces at the cellular level and further influence cellular phenotypes. This study focused on the P710R mutation that dramatically decreased in vitro motility velocity and actin-activated ATPase, in contrast to other MYH7 mutations. Optical trap measurements of single myosin molecules revealed that this mutation reduced the step size of the myosin motor and the load sensitivity of the actin detachment rate. Conversely, this mutation destabilized the super relaxed state in longer, two-headed myosin constructs, freeing more heads to generate force. Micropatterned human induced pluripotent derived stem cell (hiPSC)–cardiomyocytes CRISPR-edited with the P710R mutation produced significantly increased force (measured by traction force microscopy) compared with isogenic control cells. The P710R mutation also caused cardiomyocyte hypertrophy and cytoskeletal remodeling as measured by immunostaining and electron microscopy. Cellular hypertrophy was prevented in the P710R cells by inhibition of ERK or Akt. Finally, we used a computational model that integrated the measured molecular changes to predict the measured traction forces. These results confirm a key role for regulation of the super relaxed state in driving hypercontractility in HCM with the P710R mutation and demonstrate the value of a multiscale approach in revealing key mechanisms of disease.
“…As the effects of OM and P710R share some similarities, it is important to note that residue P710 is part of the OM binding pocket in the converter domain (65). A recent preprint reported a reduction in myosin's step size due to a different HCM mutation in the same region (P712L) (66). Taken together, these results emphasize the converter domain's critical facilitation of myosin's power stroke and load sensitivity and reveal various consequences of perturbing this domain.…”
Hypertrophic cardiomyopathy (HCM) is the most common inherited form of heart disease, associated with over 1,000 mutations, many in β-cardiac myosin (MYH7). Molecular studies of myosin with different HCM mutations have revealed a diversity of effects on ATPase and load-sensitive rate of detachment from actin. It has been difficult to predict how such diverse molecular effects combine to influence forces at the cellular level and further influence cellular phenotypes. This study focused on the P710R mutation that dramatically decreased in vitro motility velocity and actin-activated ATPase, in contrast to other MYH7 mutations. Optical trap measurements of single myosin molecules revealed that this mutation reduced the step size of the myosin motor and the load sensitivity of the actin detachment rate. Conversely, this mutation destabilized the super relaxed state in longer, two-headed myosin constructs, freeing more heads to generate force. Micropatterned human induced pluripotent derived stem cell (hiPSC)–cardiomyocytes CRISPR-edited with the P710R mutation produced significantly increased force (measured by traction force microscopy) compared with isogenic control cells. The P710R mutation also caused cardiomyocyte hypertrophy and cytoskeletal remodeling as measured by immunostaining and electron microscopy. Cellular hypertrophy was prevented in the P710R cells by inhibition of ERK or Akt. Finally, we used a computational model that integrated the measured molecular changes to predict the measured traction forces. These results confirm a key role for regulation of the super relaxed state in driving hypercontractility in HCM with the P710R mutation and demonstrate the value of a multiscale approach in revealing key mechanisms of disease.
“…During the power stroke, the light chain-binding domain rotates by ∼70°, generating a ∼4 nm displacement. ADP is then released and the light chain-binding domain rotates further to the post-power stroke state, generating an additional ∼2 nm displacement ( 20 , 21 ) for a total displacement of ∼6 nm ( 22 , 23 , 24 , 25 , 26 ). Figure 3 The myosin mechanochemical cycle and its regulation.…”
Section: The Actomyosin Biochemical Cyclementioning
confidence: 99%
“…Therefore, the rate of ADP release sets the rate of unloaded shortening, since this rate limits crossbridge detachment at physiologically relevant ATP concentrations ( 34 ). The speed is also proportional to the size of the myosin working stroke ( 32 ), which can be altered by disease-causing mutations ( 21 ). It should be noted that other models of muscle contraction have been proposed, including a recent model where the speed depends on both attachment and detachment kinetics ( 2 ).…”
Section: The Actomyosin Biochemical Cyclementioning
confidence: 99%
“…The effects of OM could be specific to mutations. An elegant example of this was the demonstration that despite suppressing the working stroke of wild-type myosin, OM rescues the reduced working stroke of myosin carrying the R712L mutation ( 21 ). This surprising finding was suggested to result from OM binding to the post-power stroke state of R712L myosin rather than the pre-power stroke state, highlighting the complexity of the therapeutic mechanism of OM and of myosin force generation more broadly.…”
Section: Small Molecules That Target Myosin Contractilitymentioning
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“…This strategy is particularly important because there is evidence that different HCM mutations can lead to hypocontractility (i.e., reduced Ca 2+ -sensitivity of force or ATPase activity) or other alterations of force generation and relaxation (e.g., Kirschner et al, 2005 ; Moore et al, 2012 ; van Dijk et al, 2012 ; Kraft et al, 2013 ). For example, an HCM-linked myosin mutation (R712L) decreased the working stroke, which was rescued by a myosin activator (omecamtiv mecarbil) instead of an inhibitor ( Snoberger et al, 2021 ). Some HCM-related mutations in cardiac troponin T (cTnT) caused an increase in Ca 2+ -sensitivity of force, which was, however, dependent on the mutation location and dose level of the mutant protein ( Schuldt et al, 2021 ).…”
Section: Why Make a Strong Muscle Weaker?mentioning
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