The hypertrophic cardiomyopathy mutations R403Q and R663H increase the number of myosin heads available to interact with actin

Sarkar, Saswata S.; Trivedi, Darshan V.; Morck, Makenna M.; Adhikari, Arjun S.; Pasha, Shaik Naseer; Ruppel, Kathleen M.; Spudich, James A.

doi:10.1126/sciadv.aax0069

Cited by 76 publications

(118 citation statements)

References 53 publications

(88 reference statements)

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“…4). This is consistent with the behavior observed in purified myosin and muscle fibers containing the myosin R403Q mutation (13,41). Interestingly, unlike WT, the SRX population in STFs made of R403Q myosin was insensitive to both the ionic strength and ADP chase.…”

Section: Discussionsupporting

confidence: 87%

Synthetic thick filaments: A new avenue for better understanding the myosin super-relaxed state in healthy, diseased, and mavacamten-treated cardiac systems

Gollapudi¹,

Yu²,

Gan³

et al. 2021

Journal of Biological Chemistry

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A hallmark feature of myosin-II is that it can spontaneously self-assemble into bipolar synthetic thick filaments (STFs) in low ionic strength buffers, thereby serving as a reconstituted in-vitro model for muscle thick filament. While these STFs have been extensively used for structural characterization, their functional evaluation has been limited. In this report, we show that myosins in STFs mirror the more electrostatic and cooperative interactions that underlie the energy-sparing super-relaxed (SRX) state, which are not seen using shorter myosin sub-fragments, heavy meromyosin (HMM) and myosin subfragment-1 (S1). Using these STFs, we show several pathophysiological insults in hypertrophic cardiomyopathy, including the R403Q myosin mutation, phosphorylation of myosin light chains, and increased ADP:ATP ratio destabilize the SRX population. Furthermore, wild-type myosin containing STFs, but not S1, HMM, or STFs-containing R403Q myosin, recapitulated the ADP-induced destabilization of the SRX state. Studies involving a clinical-stage small molecule inhibitor, mavacamten, showed that it is not only more effective in increasing myosin SRX population in STFs than in S1 or HMM , but it also increases myosin SRX population equally well in STFs made of healthy and disease-causing R403Q myosin. Importantly, we also found that pathophysiological perturbations such as elevated ADP concentration weakens the mavacamten’s ability to increase the myosin SRX population, suggesting that mavacamten-bound myosin heads are not permanently protected in the SRX state but can be recruited into action. These findings collectively emphasize that STFs serve as a valuable tool to provide novel insights into the myosin SRX state in healthy, disease, and therapeutic conditions.

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Section: Discussionsupporting

confidence: 87%

Synthetic thick filaments: A new avenue for better understanding the myosin super-relaxed state in healthy, diseased, and mavacamten-treated cardiac systems

Gollapudi¹,

Yu²,

Gan³

et al. 2021

Journal of Biological Chemistry

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“…This increases the number of heads available for binding to actin, which we hypothesized would increase force production. We have measured similar changes in SRX across a range of HCM-causing mutations [8,14], and others have confirmed these effects in cellular and animal models [13,47], suggesting that SRX regulation may be a major cause of hypercontractility in HCM. This is further corroborated by the finding that mavacamten, a small-molecule inhibitor of myosin that is known to increase the SRX state, is effective in reducing symptoms in patients with HCM [13,48].…”

Section: Discussionsupporting

confidence: 67%

Hypertrophic cardiomyopathy ß-cardiac myosin mutation (P710R) leads to hypercontractility by disrupting super-relaxed state

Roest

Liu

Morck

et al. 2020

Preprint

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Hypertrophic cardiomyopathy (HCM) is the most common inherited form of heart disease, associated with over 1000 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 decreases in vitro motility 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 larger, two-headed myosin constructs, freeing more heads to generate force. Micropatterned 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 integrates measured molecular changes to demonstrate that closely predict the measured traction forces. These results confirm a key role for regulation of the super-relaxed state in driving hypercontractility in HCM and demonstrate the value of a multiscale approach in revealing key mechanisms of disease.Significance StatementHeart disease is the leading cause of death world wide, and hypertrophic cardiomyopathy (HCM) is the most common inherited form of heart disease, affecting over 1 in 200 people. Mutations in myosin, the motor protein responsible for contraction of the heart, are a common cause of HCM but have diverse effects on the biomechanics of the myosin protein that can make it difficult to predict the combined effects of each mutation. We demonstrate that complex biomechanical effects of mutations associated with heart disease can be effectively studied and understood using a multi-scale experimental and computational modeling approach. This work can be extended to aid in the development of new targeted therapies for patients with different mutations.

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“…It was first reported in a study with skinned cardiac muscle fibers that homozygous but not heterozygous MyBPC knockout mice have a significant decrease of the myosin population in the SRX state as compared to WT ( McNamara et al, 2016 ). Consistent with this top-down observation, a bottom-up approach with purified human β-cardiac myosin recently demonstrated that the population of myosin in the SRX state increased in the presence of the C0-C7 MyBPC fragment ( Sarkar et al, 2020 ). Both these studies suggest that MyBPC increases the population of the myosin in the SRX state.…”

Section: Muscle Physiology and The Myosin Srx Statesupporting

confidence: 55%

“…A follow-up study by Adhikari et al, 2019 demonstrated the depopulation of the SRX state in recombinant human β-cardiac myosin constructs harboring mutations in the myosin mesa (R249Q, H251N) and converter (D382Y, R719W) hotspots of HCM mutations in myosin. Sarkar et al, 2020 most recently reported depopulated SRX state in recombinant human β-cardiac myosin harboring the R403Q and R663H mutations, thus extending the Anderson et al, 2018 fiber work.…”

Section: Diseased State and Therapeutic Modulators Of Myosin Srx Statmentioning

confidence: 97%

To lie or not to lie: Super-relaxing with myosins

Nag¹,

Trivedi

2021

eLife

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Since the discovery of muscle in the 19th century, myosins as molecular motors have been extensively studied. However, in the last decade, a new functional super-relaxed (SRX) state of myosin has been discovered, which has a 10-fold slower ATP turnover rate than the already-known non-actin-bound, disordered relaxed (DRX) state. These two states are in dynamic equilibrium under resting muscle conditions and are thought to be significant contributors to adaptive thermogenesis in skeletal muscle and can act as a reserve pool that may be recruited when there is a sustained demand for increased cardiac muscle power. This report provides an evolutionary perspective of how striated muscle contraction is regulated by modulating this myosin DRX↔SRX state equilibrium. We further discuss this equilibrium with respect to different physiological and pathophysiological perturbations, including insults causing hypertrophic cardiomyopathy, and small-molecule effectors that modulate muscle contractility in diseased pathology.

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The hypertrophic cardiomyopathy mutations R403Q and R663H increase the number of myosin heads available to interact with actin

Cited by 76 publications

References 53 publications

Synthetic thick filaments: A new avenue for better understanding the myosin super-relaxed state in healthy, diseased, and mavacamten-treated cardiac systems

Synthetic thick filaments: A new avenue for better understanding the myosin super-relaxed state in healthy, diseased, and mavacamten-treated cardiac systems

Hypertrophic cardiomyopathy ß-cardiac myosin mutation (P710R) leads to hypercontractility by disrupting super-relaxed state

To lie or not to lie: Super-relaxing with myosins

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