Abstract P338: Myopeptide Improves Contractile Mechanics In A Mouse Model Of Heart Failure Expressing Phosphoablated Cardiac Myosin Binding Protein-C

Rationale: Normal heart function depends on cardiac myosin binding protein-C (cMyBP-C) phosphorylation. Its decrease is associated with heart failure (HF) by inhibiting actomyosin interactions. In absence of cMyBP-C phosphorylation, the protein is bound to myosin S2, but released when phosphorylated, allowing myosin to form cross-bridges with actin. Challenging cMyBP-C/myosin S2 interaction by myopeptide (the first 126 amino acids of myosin S2) could promote actomyosin interaction in vitro , but its ability to improve contractility in HF remains untested. Objective: To test contractile function in skinned papillary fibers of a cMyBP-C dephosphorylated mouse model using myopeptide. Methods and Results: To mimic constitutive phosphoablation, a knock-in mouse model was established to express cMyBP-C in which serines 273, 282 and 302 were mutated to alanine (cMyBP-C AAA ). Western blotting revealed 50% and 100% of cMyBP-C AAA in het and homo mouse hearts, respectively. Echocardiography showed a decreased percentage of ejection fraction (28%, p<0.01) and fractional shortening (30%, p< 0.05) in both het and homo cMyBP-C AAA mice at 3 months of age, compared to knock-in negative controls. These mice also developed diastolic dysfunction with elevated ratio of E/A and E/e’ waves. Next, pCa-force measurements using skinned papillary fibers determined that maximal force (F max ) and rate of cross-bridge formation ( k tr ) were decreased in the cMyBP-C AAA groups, compared to the control. However, administration of dose-dependent myopeptide increased F max and k tr in wild-type and cMyBP-C AAA permeabilized skinned papillary fibers without affecting myofilament Ca 2+ sensitivity. Conclusions: Myopeptide can increase contractile force and rate of cross-bridge formation by releasing cMyBP-C/myosin S2 and promoting actomyosin formation of cross-bridges, thus validating its therapeutic potential.

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Abstract 125: Cardiac Myosin Binding Protein-C Phosphorylation Is Critical for Normal Diastolic Function

Circulation Research ◽

10.1161/res.111.suppl_1.a125 ◽

2012 ◽

Vol 111 (suppl_1) ◽

Author(s):

Carl W Tong ◽

Mohamed I Abdalla ◽

Yang Liu ◽

Himakarnika Alluri ◽

David E Dostal ◽

...

Keyword(s):

Heart Failure ◽

Ejection Fraction ◽

Diastolic Function ◽

Protein C ◽

Cardiac Myosin ◽

Cross Bridge ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Voluntary Running ◽

Myosin Binding

Heart failure with preserved ejection fraction (HFpEF) accounts for approximately 50% of all cases of heart failure, but there is no effective treatment of this disease. Cardiac myosin binding protein-C (MyBPC3) is a thick filament protein that is thought to slow cross-bridge cycling by inhibiting acto-myosin interaction. When phosphorylated, MyBPC3 releases its inhibition thereby accelerating cross-bridge cycling. Thus, we hypothesize that MyBPC3 phosphorylation enhances heart’s ability to relax (lusitropy). To test this idea, we expressed wild type MyBPC3(tWT), non-phosphorylatable MyBPC3(t3SA), and constitutively phosphorylated mimetic MyBPC3(t3SD) onto MyBPC3(-/-) mouse background. We used echocardiography, voluntary running, and measurements of brain natriuretic peptide (BNP) for comparison. MyBPC3(t3SA) and MyBPC3(t3SD) hearts had similar ejection fraction as MyBPC3(tWT) hearts. MyBPC3(t3SA) hearts show depressed diastolic function revealed by slower myocardial tissue Doppler relaxation velocity at the mitral valve annulus (Ea) and an increased mitral blood flow/myocardium relaxation ratio (E/Ea). In contrast, MyBPC3(t3SD) hearts exhibited enhanced lusitropy with increased Ea and smaller E/Ea. Moreover, the findings of shorter 7-day average voluntary running distances, increased lung/body weight ratios, and increased BNP levels indicated heart failure in MyBPC3(t3SA) mice. Conversely, MyBPC3(t3SD) mice did not show signs of heart failure. Since cardiac function in MyBPC3(t3SA) mice resembles HFpEF and MyBPC3(t3SD) mice exhibit enhanced lusitropy, we conclude that phosphorylation of MyBPC3 is crucial for normal diastolic function.

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Cardiac myosin‐binding protein C as a candidate biomarker in heart failure: rational but not revolutionary

European Journal of Heart Failure ◽

10.1002/ejhf.2126 ◽

2021 ◽

Author(s):

A. Mark Richards

Keyword(s):

Heart Failure ◽

Protein C ◽

Binding Protein ◽

Cardiac Myosin ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Myosin Binding

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Phosphorylation of cardiac myosin binding protein C releases myosin heads from the surface of cardiac thick filaments

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1614020114 ◽

2017 ◽

Vol 114 (8) ◽

pp. E1355-E1364 ◽

Cited By ~ 35

Author(s):

Robert W. Kensler ◽

Roger Craig ◽

Richard L. Moss

Keyword(s):

Protein C ◽

Structural Changes ◽

Binding Protein ◽

Cardiac Contraction ◽

Cardiac Myosin ◽

Cross Bridge ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Thick Filaments ◽

Myosin Binding

Cardiac myosin binding protein C (cMyBP-C) has a key regulatory role in cardiac contraction, but the mechanism by which changes in phosphorylation of cMyBP-C accelerate cross-bridge kinetics remains unknown. In this study, we isolated thick filaments from the hearts of mice in which the three serine residues (Ser273, Ser282, and Ser302) that are phosphorylated by protein kinase A in the m-domain of cMyBP-C were replaced by either alanine or aspartic acid, mimicking the fully nonphosphorylated and the fully phosphorylated state of cMyBP-C, respectively. We found that thick filaments from the cMyBP-C phospho-deficient hearts had highly ordered cross-bridge arrays, whereas the filaments from the cMyBP-C phospho-mimetic hearts showed a strong tendency toward disorder. Our results support the hypothesis that dephosphorylation of cMyBP-C promotes or stabilizes the relaxed/superrelaxed quasi-helical ordering of the myosin heads on the filament surface, whereas phosphorylation weakens this stabilization and binding of the heads to the backbone. Such structural changes would modulate the probability of myosin binding to actin and could help explain the acceleration of cross-bridge interactions with actin when cMyBP-C is phosphorylated because of, for example, activation of β1-adrenergic receptors in myocardium.

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S ‐glutathiolation impairs phosphoregulation and function of cardiac myosin‐binding protein C in human heart failure

The FASEB Journal ◽

10.1096/fj.201500048 ◽

2016 ◽

Vol 30 (5) ◽

pp. 1849-1864 ◽

Cited By ~ 13

Author(s):

Konstantina Stathopoulou ◽

Ilka Wittig ◽

Juliana Heidler ◽

Angelika Piasecki ◽

Florian Richter ◽

...

Keyword(s):

Heart Failure ◽

Protein C ◽

Human Heart ◽

Binding Protein ◽

Cardiac Myosin ◽

Human Heart Failure ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Myosin Binding ◽

And Function

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Diagnostic and Prognostic Value of Plasma Levels of Cardiac Myosin Binding Protein-C as a Novel Biomarker in Heart Failure

Pediatric Cardiology ◽

10.1007/s00246-016-1532-2 ◽

2016 ◽

Vol 38 (2) ◽

pp. 418-424 ◽

Cited By ~ 3

Author(s):

Doaa El Amrousy ◽

Hossam Hodeib ◽

Ghada Suliman ◽

Nahed Hablas ◽

Eman Ramadan Salama ◽

...

Keyword(s):

Heart Failure ◽

Plasma Levels ◽

Protein C ◽

Prognostic Value ◽

Binding Protein ◽

Cardiac Myosin ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Myosin Binding ◽

Diagnostic And Prognostic Value

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Decreased phosphorylation levels of cardiac myosin-binding protein-C in human and experimental heart failure

Journal of Molecular and Cellular Cardiology ◽

10.1016/j.yjmcc.2007.05.003 ◽

2007 ◽

Vol 43 (2) ◽

pp. 223-229 ◽

Cited By ~ 111

Author(s):

Ali El-Armouche ◽

Lutz Pohlmann ◽

Saskia Schlossarek ◽

Jutta Starbatty ◽

Yung-Hsin Yeh ◽

...

Keyword(s):

Heart Failure ◽

Protein C ◽

Binding Protein ◽

Cardiac Myosin ◽

Experimental Heart Failure ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Myosin Binding

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Hypercontractile Properties of Cardiac Muscle Fibers in a Knock-in Mouse Model of Cardiac Myosin-binding Protein-C

Journal of Biological Chemistry ◽

10.1074/jbc.m008691200 ◽

2000 ◽

Vol 276 (7) ◽

pp. 5353-5359 ◽

Cited By ~ 50

Author(s):

Christian C. Witt ◽

Brenda Gerull ◽

Michael J. Davies ◽

Thomas Centner ◽

Wolfgang A. Linke ◽

...

Keyword(s):

Mouse Model ◽

Cardiac Muscle ◽

Protein C ◽

Binding Protein ◽

Muscle Fibers ◽

Cardiac Myosin ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Myosin Binding ◽

Cardiac Muscle Fibers

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Protein kinase A–induced myofilament desensitization to Ca2+ as a result of phosphorylation of cardiac myosin–binding protein C

The Journal of General Physiology ◽

10.1085/jgp.201010448 ◽

2010 ◽

Vol 136 (6) ◽

pp. 615-627 ◽

Cited By ~ 50

Author(s):

Peter P. Chen ◽

Jitandrakumar R. Patel ◽

Inna N. Rybakova ◽

Jeffery W. Walker ◽

Richard L. Moss

Keyword(s):

Protein Kinase ◽

Protein Kinase A ◽

Protein C ◽

Binding Protein ◽

Cardiac Myosin ◽

Cross Bridge ◽

Myosin Binding Protein ◽

Myosin Binding Protein C ◽

Myosin Binding ◽

Kinase A

In skinned myocardium, cyclic AMP–dependent protein kinase A (PKA)-catalyzed phosphorylation of cardiac myosin–binding protein C (cMyBP-C) and cardiac troponin I (cTnI) is associated with a reduction in the Ca2+ responsiveness of myofilaments and an acceleration in the kinetics of cross-bridge cycling, although the respective contribution of these two proteins remains controversial. To further examine the relative roles that cTnI and cMyBP-C phosphorylation play in altering myocardial function, we determined the Ca2+ sensitivity of force (pCa50) and the activation dependence of the rate of force redevelopment (ktr) in control and PKA-treated mouse myocardium (isolated in the presence of 2,3-butanedione monoxime) expressing: (a) phosphorylatable cTnI and cMyBP-C (wild type [WT]), (b) phosphorylatable cTnI on a cMyBP-C–null background (cMyBP-C−/−), (c) nonphosphorylatable cTnI with serines23/24/43/45 and threonine144 mutated to alanines (cTnIAla5), and (d) nonphosphorylatable cTnI on a cMyBP-C–null background (cTnIAla5/cMyBP-C−/−). Here, PKA treatment decreased pCa50 in WT, cTnIAla5, and cMyBP-C−/− myocardium by 0.13, 0.08, and 0.09 pCa units, respectively, but had no effect in cTnIAla5/cMyBP-C−/− myocardium. In WT and cTnIAla5 myocardium, PKA treatment also increased ktr at submaximal levels of activation; however, PKA treatment did not have an effect on ktr in cMyBP-C−/− or cTnIAla5/cMyBP-C−/− myocardium. In addition, reconstitution of cTnIAla5/cMyBP-C−/− myocardium with recombinant cMyBP-C restored the effects of PKA treatment on pCa50 and ktr reported in cTnIAla5 myocardium. Collectively, these results indicate that the attenuation in myofilament force response to PKA occurs as a result of both cTnI and cMyBP-C phosphorylation, and that the reduction in pCa50 mediated by cMyBP-C phosphorylation most likely arises from an accelerated cross-bridge cycling kinetics partly as a result of an increased rate constant of cross-bridge detachment.

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