Skeletal myosin binding protein-C isoforms regulate thin filament activity in a Ca2+-dependent manner

Mutations in genes encoding myosin, the molecular motor that powers cardiac muscle contraction, and its accessory protein, cardiac myosin binding protein C (cMyBP-C), are the two most common causes of hypertrophic cardiomyopathy (HCM). Recent studies established that the N-terminal domains (NTDs) of cMyBP-C (e.g., C0, C1, M, and C2) can bind to and activate or inhibit the thin filament (TF). However, the molecular mechanism(s) by which NTDs modulate interaction of myosin with the TF remains unknown and the contribution of each individual NTD to TF activation/inhibition is unclear. Here we used an integrated structure–function approach using cryoelectron microscopy, biochemical kinetics, and force measurements to reveal how the first two Ig-like domains of cMyPB-C (C0 and C1) interact with the TF. Results demonstrate that despite being structural homologs, C0 and C1 exhibit different patterns of binding on the surface of F-actin. Importantly, C1 but not C0 binds in a position to activate the TF by shifting tropomyosin (Tm) to the “open” structural state. We further show that C1 directly interacts with Tm and traps Tm in the open position on the surface of F-actin. Both C0 and C1 compete with myosin subfragment 1 for binding to F-actin and effectively inhibit actomyosin interactions when present at high ratios of NTDs to F-actin. Finally, we show that in contracting sarcomeres, the activating effect of C1 is apparent only once low levels of Ca2+ have been achieved. We suggest that Ca2+ modulates the interaction of cMyBP-C with the TF in the sarcomere.

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Skeletal Myosin Binding Protein-C Isoforms Modulate Actomyosin Contractility and are Regulated by Phosphorylation

Biophysical Journal ◽

10.1016/j.bpj.2014.11.2304 ◽

2015 ◽

Vol 108 (2) ◽

pp. 421a

Author(s):

Amy Li ◽

Samantha Beck Previs ◽

Michael Previs ◽

Brian Lin ◽

Cristobal dos Remedios ◽

...

Keyword(s):

Protein C ◽

Binding Protein ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Actomyosin Contractility ◽

Skeletal Myosin ◽

Myosin Binding

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Fast skeletal myosin-binding protein-C regulates fast skeletal muscle contraction

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2003596118 ◽

2021 ◽

Vol 118 (17) ◽

pp. e2003596118

Author(s):

Taejeong Song ◽

James W. McNamara ◽

Weikang Ma ◽

Maicon Landim-Vieira ◽

Kyoung Hwan Lee ◽

...

Keyword(s):

Skeletal Muscle ◽

Protein C ◽

Binding Protein ◽

Calcium Sensitivity ◽

Fast Skeletal Muscle ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Skeletal Myosin ◽

Myosin Binding ◽

Edl Muscle

Fast skeletal myosin-binding protein-C (fMyBP-C) is one of three MyBP-C paralogs and is predominantly expressed in fast skeletal muscle. Mutations in the gene that encodes fMyBP-C, MYBPC2, are associated with distal arthrogryposis, while loss of fMyBP-C protein is associated with diseased muscle. However, the functional and structural roles of fMyBP-C in skeletal muscle remain unclear. To address this gap, we generated a homozygous fMyBP-C knockout mouse (C2−/−) and characterized it both in vivo and in vitro compared to wild-type mice. Ablation of fMyBP-C was benign in terms of muscle weight, fiber type, cross-sectional area, and sarcomere ultrastructure. However, grip strength and plantar flexor muscle strength were significantly decreased in C2−/− mice. Peak isometric tetanic force and isotonic speed of contraction were significantly reduced in isolated extensor digitorum longus (EDL) from C2−/− mice. Small-angle X-ray diffraction of C2−/− EDL muscle showed significantly increased equatorial intensity ratio during contraction, indicating a greater shift of myosin heads toward actin, while MLL4 layer line intensity was decreased at rest, indicating less ordered myosin heads. Interfilament lattice spacing increased significantly in C2−/− EDL muscle. Consistent with these findings, we observed a significant reduction of steady-state isometric force during Ca2+-activation, decreased myofilament calcium sensitivity, and sinusoidal stiffness in skinned EDL muscle fibers from C2−/− mice. Finally, C2−/− muscles displayed disruption of inflammatory and regenerative pathways, along with increased muscle damage upon mechanical overload. Together, our data suggest that fMyBP-C is essential for maximal speed and force of contraction, sarcomere integrity, and calcium sensitivity in fast-twitch muscle.

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