doi: 10.1161/01.RES.0000225873.63162.c4
© 2006 American Heart Association, Inc.
Editorials |
Myosin Binding Protein C in the Heart
From the Program in Cardiovascular Sciences, University of Illinois at Chicago.
Correspondence to Pieter P. de Tombe, PhD, Department of Physiology & Biophysics, 835 South Wolcott Ave MC 901, University of Illinois at Chicago, Chicago, IL 60612. E-mail pdetombe{at}uic.edu
See related article, pages 1290–1298
Key Words: myosin accessory proteins • sarcomere • cross-bridge cycling • regulation
Myocardial force generation is initiated on the release of calcium ions from the sarcoplasmic reticulum and subsequent activation of the contractile apparatus.1 A schematic of the cardiac sarcomere and some of its component proteins is shown in the Figure.2 Contractile activation involves binding calcium to Tn-C, redistribution of the affinity of Tn-I for Tn-C away from Tn-T, and movement of tropomyosin on the thin filament, thereby exposing binding sites on actin. Binding of the myosin S1 domain to actin leads to cross-bridge force development. However, this simple two state model of muscle contraction is not sufficient to explain many features of muscle contraction.2 For example, the rate of force development critically depends on the activation state of the thin filament.2 Also, binding of a modified myosin (NEM-S1 which cannot hydrolyze ATP) has been shown to abolish this phenomenon.3 This, and other structural evidence, has led to the hypothesis that the thin filament must have at least three states: a blocked state (top left), where tropomyosin sterically blocks S1 binding; a closed state, induced by calcium binding to Tn-C, where tropomyosin has shifted away somewhat allowing weak interaction with the S1 domain; and an open state, where tropomyosin has moved away fully allowing for a strong interaction with S1 and force generation (bottom).2 One interesting aspect of this theory is that the binding of myosin to actin by itself induces further activation of the thin filament, thus facilitating S1 domain binding along the filament; an elegant, yet simple molecular model of cooperative activation. Cardiac muscle activation is highly cooperative, even though calcium binding to isolated cardiac troponin-C itself is not cooperative.2,4 However, it is less clear how modulation of cross-bridge cycling rate could be accommodated by such a theory.2,4 To further complicate matters, phosphorylation of contractile proteins, in particular troponin-I and troponin-T, leads to marked alterations in cross-bridge cycling rate apart from its impact on cooperative activation.5 Thus, modulation of thin filament activation, either directly by varying the level of thin filament activation2,4 or indirectly by posttranslational modification of regulatory proteins,5 appears to modulate the kinetics of the actin–myosin interaction. Data presented by Herron et al in this issue suggest that myosin binding protein C (MyBP-C) now needs to be added to this list.6
MyBP-C, or C-protein, was first identified by Starr and Offer in myosin extracts from muscle.7 The protein was named on the basis of mobility on an SDS-gel (the third protein, ie, C-protein after the first and second proteins on the gel). MyBP-C is a large protein, consisting of 10 domains: 7 immunoglobulin motifs and 3 fibronectin motifs.8,9 MyBP-C exists in a 
1:8 molar ratio with myosin, and there are several muscle type specific isoforms.8,9 Notably, as is the case for troponin-I, the cardiac isoform of MyBP-C is larger and contains several phosphorylation sites. The protein has been located in the sarcomere by immunohistochemistry to 8 10-nm-wide transverse stripes, at 43 nm intervals along the thick filament in two zones located on either side of the M-line. It has been speculated that 3 MyBP-C proteins join to form a "collar structure" surrounding the thick filament.8,9 It is important to note that because of the stoichiometry and this "collar," only 1 of 7 to 8 myosin molecules in the sarcomere are actually exposed to MyBP-C. The precise function of MyBP-C has been somewhat of a mystery. It has been suggested that MyBP-C is important as a molecular "ruler" (as has the giant protein titin10,11) to regulate the length of the thick filament during assembly of the sarcomere.8,9 Recent data support this notion, albeit in a slightly modified way; the role of MyBP-C here is proposed to be a binding partner to a LIM scaffold protein involved in the regulation of myosin filament formation and sarcomere assembly.12 However, murine knockout models of MyBP-C do not show a lack of sarcomere formation,13–15 a result that either implies a minor role for MyBP-C in sarcomere assembly or that this important function has been taken over by some other as yet unidentified mechanism. Second, MyBP-C may play a role in the regulation of muscle contraction.8,9 Indeed, early studies in which MyBP-C was chemically extracted from skinned myocardium showed an increase in calcium sensitivity and velocity,16 consistent with recent results obtained in the MyBP-C knockout models.13–15 Cardiac MyBP-C is a substrate for PKA; stimulation of beta-adrenergic receptors in the heart leads to marked phosphorylation of both MyBP-C and Tn-I with a similar time course.5 Earlier work by the Winegrad group suggested that PKA-mediated phosphorylation allowed myosin heads to move away from the backbone of the thick filament.8 Overall, these results paint a picture for a molecular model where MyBP-C plays a critical role in the activation process of the cardiac sarcomere by restraining or limiting the reach of myosin heads toward the thin filament. An interesting twist to this theory was recently provided by Cazorla et al,17 who showed that PKA mediated phosphorylation of Tn-I in a MyBP-C knockout model failed to affect myofilament calcium sensitivity, similar to earlier work using a slow skeletal muscle Tn-I replacement transgenic animal model.5 The implication of these two studies is that MyBP-C needs to be phosphorylated, or at least be present, for the PKA signal to be effective in modulating contractile function in the heart.
How MyBP-C functions in the heart, and which are the domains within MyBP-C that are responsible for its action, are still actively being investigated. The current study6 reveals some new secrets of MyBP-C. Application of various amounts of recombinant MyBP-C to skinned myocardium induced force production in the absence of calcium. The effect was localized to the C0-C1 N amino terminus domain of cardiac MyBP-C (Figure). These results imply that the normal role of MyBP-C (left panels) is to limit binding of S1 to inactivated thin filaments; competitive binding of exogenously applied MyBP-C Co-C1 (right panels) relieves this restriction. In addition, Herron et al6 clearly demonstrated that MyBP-C-C0-C1 induced cross-bridges behave quite like regular activated cross-bridges in terms of the kinetics of force development. A problem with the model presented in the Figure is that the total absence of MyBP-C, as is the case in the various knockout models, should lead to substantial contractile activation in diastole incompatible with life. Yet this is not what was observed.13–15 A likely explanation for this conundrum may lie in the fact that MyBP-C normally exists in a 1:8 stoichiometry to myosin.8,9 Hence, application of a saturating concentration of MyBP-C fragment will likely lead to binding of the protein to all remaining sites on myosin along the filament. Thus, the results may simply be attributable to the induction of an unphysiological state where every myosin molecule in the sarcomere is no longer "limited" by being tethered to MyBP-C. This is a limitation, as acknowledged by Herron et al,6 of any study where MyBP-C fragments are applied to skinned muscle. To address this potential pitfall, Herron and colleagues also studied the impact of a much lower concentration of MyBP-C fragment, that is, a concentration where there was no observable impact on inactivated myofilament function. Surprisingly, this low concentration of MyBP-C-C0-C1 resulted in marked increases in cross-bridge cycling kinetics at sub-maximal activation levels and at saturating levels of calcium activation. In addition, the impact of sarcomere length on myofilament calcium sensitivity, a property regularly referred to as myofilament length dependent activation,18 was completely abolished in the presence of a low concentration of the protein fragment. Although the molecular mechanisms that underlie myofilament length dependent activation are not known at present, these data strongly suggest that MyBP-C plays a major role in this important physiological regulatory process. MyBP-C may be well positioned within the contractile apparatus to perform such a role (Figure). It is possible that a change in sarcomere length indirectly affects the interaction between myosin and MyBP-C via changes in inter-filament spacing.18 Alternatively, sarcomere length changes may be transmitted directly to MyBP-C via the giant molecule titin.10,11,18 Regardless of the underlying mechanisms, however, the data by Herron et al6 demonstrate a prominent role for MyBP-C in the regulation of myofilament force development, calcium sensitivity, cross-bridge cycling rate, and length dependent activation.
|
It is now recognized that mutations in the MyBP-C gene underlie a large proportion of familial hypertrophic cardiomyopathy (FHC) cases.9 Many of these mutations involve the loss of critical binding sites for myosin from MyBP-C, thus often causing reduced levels of MyBP-C in the sarcomeres of the individuals affected by the mutation. As in the murine knockout models, loss of MyBP-C in humans does not lead to complete impairment of sarcomere assembly nor its function. Nevertheless, lack of functional MyBP-C clearly sets the stage for severe cardiac dysfunction, consistent with the notion that MyBP-C is an important regulatory protein in the cardiac sarcomere. Unraveling the precise molecular mechanisms will require future studies, using all of the tools that modern biology can offer, such as transgenic animals (including knock-out and knock-in models), isolated human myocardium studies, viral transfection studies, and studies using recombinant contractile proteins applied to skinned myocardium.19
 |
Acknowledgments |
|---|
This work was supported in part by National Institutes of Health grants HL-62426, HL75494, and HL77195.
|
Footnotes |
|---|
The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.
|
References |
|---|
 Top References |
|---|
1. Bers DM. Excitation-Contraction Coupling and Cardiac Contractile Force, 2nd ed. Dordrecht, The Netherlands: Kluwer Academic Press; 2001.
2. Gordon AM, Homsher E, Regnier M. Regulation of contraction in striated muscle. Physiol Rev. 2000; 80: 853–924.
3. Fitzsimons DP, Patel JR, Moss RL. Cross-bridge interaction kinetics in rat myocardium are accelerated by strong binding of myosin to the thin filament. J Physiol. 2001; 530 (Pt 2): 263–272.
4. de Tombe PP. Cardiac myofilaments: mechanics and regulation. J Biomech. 2003; 36: 721–730.[CrossRef][Medline] [Order article via Infotrieve]
5. Sumandea M, Burkart EM, Kobayashi T, De Tombe PP, Solaro RJ. Molecular and integrated biology of thin filament protein phosphorylation in heart muscle. Ann N Y Acad Sci. 2004; 1015: 39–52.[CrossRef][Medline] [Order article via Infotrieve]
6. Herron TJ, Rostkova E, Kunst G, Chaturvedi R, Gautel M, Kentish JC. Activation of myocardial contraction by the N-terminal domains of myosin binding protein C. Circ Res. 2006; 98: 1290–1298.
7. Starr R, Offer G. The interaction of C-protein with heavy meromyosin and subfragment-2. Biochem J. 1978; 171: 813–816.[Medline] [Order article via Infotrieve]
8. Winegrad S. Cardiac myosin binding protein C. Circ Res. 1999; 84: 1117–1126.
9. Flashman E, Redwood C, Moolman-Smook J, Watkins H. Cardiac myosin binding protein C: its role in physiology and disease. Circ Res. 2004; 94: 1279–1289.
10. Labeit S, Kolmerer B, Linke WA. The giant protein titin: emerging roles in physiology and pathophysiology. Circ Res. 1997; 80: 290–294.
11. Granzier HL, Labeit S. The giant protein titin: a major player in myocardial mechanics, signaling, and disease. Circ Res. 2004; 94: 284–295.
12. McGrath M, Cottle D, Nguyen M, Dyson JM, Coghill ID, Robinson PA, Holdsworth M, Cowling BS, Hardeman EC, Mitchell CA, Brown S. Four and a half LIM protein 1 bins myosin-binding protein C and regulates myosin filament formation and sarcomere assembly. J Biol Chem. 2006; 281: 7666.
13. Harris SP, Bartley CR, Hacker TA, McDonald KS, Douglas PS, Greaser ML, Powers PA, Moss RL. Hypertrophic cardiomyopathy in cardiac myosin binding protein-C knockout mice. Circ Res. 2002; 90: 594–601.
14. McConnell BK, Jones KA, Fatkin D, Arroyo LH, Lee RT, Aristizabal O, Turnbull DH, Georgakopoulos D, Kass D, Bond M, Niimura H, Schoen FJ, Conner D, Fischman DA, Seidman CE, Seidman JG. Dilated cardiomyopathy in homozygous myosin-binding protein-C mutant mice. J Clin Invest. 1999; 104: 1235–1244.[Medline] [Order article via Infotrieve]
15. Carrier L, Knoll R, Vignier N, Keller DI, Bausero P, Prudhon B, Isnard R, Ambroisine ML, Fiszman M, Ross J Jr, Schwartz K, Chien KR. Asymmetric septal hypertrophy in heterozygous cMyBP-C null mice. Cardiovasc Res. 2004; 63: 293–304.
16. Hofmann PA, Greaser ML, Moss RL. C-protein limits shortening velocity of rabbit skeletal muscle fibres at low levels of Ca2+ activation. J Physiol (Lond). 1991; 439: 701–715.
17. Cazorla O, Szilagyi S, Vignier N, Salazar G, Kramer E, Vassort G, Carrier L, Lacampagne A. Length and protein kinase A modulations of myocytes in cardiac myosin binding protein C-deficient mice. Cardiovasc Res. 2006; 69: 370–380.
18. Konhilas JP, Irving TC, De Tombe PP. Frank-Starling law of the heart and the cellular mechanisms of length-dependent activation. Pflugers Arch. 2002; 445: 305–310.[CrossRef][Medline] [Order article via Infotrieve]
19. Solaro RJ, Moss RL. Molecular Control Mechanisms in Striated Muscle Contraction.Vol 1. Dordrecht: Kluwer Academic Publisher; 2002.
Related Article:
Circ. Res. 2006 98: 1290-1298.
This article has been cited by other articles:
 |
|
 |
|
  Y. Ge, I. N. Rybakova, Q. Xu, and R. L. Moss Top-down high-resolution mass spectrometry of cardiac myosin binding protein C revealed that truncation alters protein phosphorylation state PNAS, August 4, 2009; 106(31): 12658 - 12663. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. A. Colson, T. Bekyarova, M. R. Locher, D. P. Fitzsimons, T. C. Irving, and R. L. Moss Protein Kinase A-Mediated Phosphorylation of cMyBP-C Increases Proximity of Myosin Heads to Actin in Resting Myocardium Circ. Res., August 1, 2008; 103(3): 244 - 251. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. L. Granzier and K. B. Campbell New Insights in the Role of Cardiac Myosin Binding Protein C As a Regulator of Cardiac Contractility Circ. Res., October 13, 2006; 99(8): 795 - 797. [Full Text] [PDF]
|
|
|
|
|
|
J. E. Stelzer, J. R. Patel, and R. L. Moss Protein Kinase A-Mediated Acceleration of the Stretch Activation Response in Murine Skinned Myocardium Is Eliminated by Ablation of cMyBP-C Circ. Res., October 13, 2006; 99(8): 884 - 890. [Abstract] [Full Text] [PDF]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||