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Cardiovascular Research 2006 69(2):304-306; doi:10.1016/j.cardiores.2005.12.008
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Copyright © 2005, European Society of Cardiology

The role of cardiac myosin binding protein-C as a regulator of myofilament Ca2+ sensitivity

Harald Kögler*

Herzzentrum Göttingen, Kardiologie und Pneumologie, Georg-August-Universität, Göttingen, Germany

* Heart Center Göttingen, Department of Cardiology, Robert-Koch-Str. 40, 37075 Göttingen, Germany. Tel.: +49 551 396380; fax: +49 551 392953. Email address: hkogler{at}med.uni-goettingen.de

Received 29 November 2005; accepted 8 December 2005

Stimulation of β1-adrenergic receptors in the heart by endogenous or exogenous agonists results in enhanced formation of the second messenger cyclic 3',5'-adenosine monophosphate (cAMP), which in turn activates cAMP-dependent protein kinase (PKA). PKA-catalysed phosphorylation ultimately triggers a variety of target proteins to undergo orchestrated functional changes that together comprise the positive inotropic and lusitropic effects caused by β1-adrenoceptor activation. Both these effects serve to enhance cardiac output: the positive inotropic effect boosts myocyte contractility by optimizing Ca2+ homeostasis, mainly via phosphorylation of phospholamban, sarcoplasmic reticulum (SR) Ca2+-ATPase, and L-type Ca2+ channels. The positive lusitropic effect, on the other hand, accelerates myocyte relaxation, thereby facilitating an increase in cardiac output via elevation of heart rate. While the stimulation of Ca2+ reuptake into the sarcoplasmic reticulum to some extent speeds up relaxation, the efficiency of this mechanism is limited by the rate at which Ca2+ ions dissociate (Ca2+ off-rate) from the thin filament regulatory protein troponin-C (TnC), which is a prerequisite for relaxation to occur at the actomyosin cross-bridge level. Hence, mechanisms that decrease the Ca2+ affinity of TnC will favour rapid relaxation.

Treatment of detergent-permeabilised heart preparations with the catalytic subunit of PKA results in a decrease in Ca2+ sensitivity, an increased Ca2+ off-rate, and faster muscle relaxation [1]. Among the target proteins for PKA-dependent phosphorylation, the myofilament proteins cardiac troponin-I (cTnI) [2,3], cardiac myosin binding protein-C (cMyBP-C) [4], and titin [5] have been identified. Due to the prominent role cTnI plays as a molecular switch in the process of Ca2+-dependent activation of muscle contraction, its small size (Mr=24 kDa for human cTnI, which facilitates the production of recombinant protein in transformed bacterial cultures), and the early availability of ultrastructural data, attention initially was focused on the functional consequences of cTnI phosphorylation.

During the last decade, however, interest in the structure and function of cMyBP-C has grown substantially, ignited by the discovery that a large fraction of patients suffering from familial hypertrophic cardiomyopathy (FHC) do so due to mutations in this gene, specifically those affected by chromosome 11-associated FHC [6]. MyBP-C was originally discovered as co-purifying contaminant in myosin preparations [7]. The cardiac-specific isoform cMyBP-C is an 1173-residue polypeptide with a predicted Mr of 137 kDa. Within its N-terminus, the protein contains three PKA phosphorylation sites and an additional site phosphorylated by Ca2+/calmodulin-dependent kinase [6]. cMyBP-C possesses binding sites for myosin, actin, and titin. Modulation of any of these protein–protein interactions could potentially exert pronounced effects on contractility. Early biochemical experiments indicated that cMyBP-C in its dephosphorylated form stimulated actin-activated myosin ATPase activity by stabilising the interaction between actin and myosin, and this stabilising effect was somewhat weakened following phosphorylation of cMyBP-C [8].

New light on the role of cMyBP-C with regard to regulating myofilament Ca2+ sensitivity is now shed by a report published in this issue of Cardiovascular Research [9]: Cazorla et al. examined single permeabilized ventricular myocytes from a knockout (KO) mouse model with a targeted deletion of exons 1 and 2 of the cMyBP-C gene. Since this fragment contains the gene's transcription initiation site, homozygous KO mice do not express intact or truncated cMyBP-C. As a consequence, they develop pronounced eccentric left ventricular hypertrophy and in in vivo hemodynamic measurements exhibit impaired ventricular relaxation under baseline conditions that is accentuated upon treatment with catecholamines [10]. These findings stimulated the authors to hypothesise that cMyBP-C deficiency could sensitise the myofilaments to Ca2+ and that the failure of β1-adrenoceptor stimulation to trigger an appropriate positive lusitropic effect in these KO mice could be due to an attenuated Ca2+-desensitising effect of PKA phosphorylation. As an additional intervention known to exert pronounced effects on myofilament Ca2+ sensitivity, the effect of an increase in sarcomere length (length-dependent activation) was assessed. Indeed, it was demonstrated that myocytes from KO mice exhibited an increased myofilament Ca2+ sensitivity at resting sarcomere length (1.9 µm). However, the Ca2+-sensitisation elicited by stretch of the sarcomeres to 2.3 µm was reduced in KO compared to wild type (WT) mice, such that at this sarcomere length there was no significant difference in the EC50. Moreover, the expected Ca2+-desensitising response to treatment with catalytic subunit of PKA that was present in WT myocytes was largely suppressed in KO myocytes. Care was taken to exclude the possibility that differences in cTnI phosphorylation or re-expression of the non-phosphorylatable slow-skeletal isoform of TnI (ssTnI) interfered with the functional measurements.

The authors concluded that myofilament Ca2+ desensitisation induced by PKA requires the presence of cMyBP-C. As they discuss, however, their findings are not easily reconciled with previous reports and they are, therefore, careful not to overstate their evidence: replacement of cTnI with ssTnI has been shown to likewise suppress the effect of PKA treatment on myofilament Ca2+ sensitivity [11,12]. A conservative interpretation of these studies taken together has to state that the presence of both cTnI and cMyBP-C is necessary for PKA to cause myofilament Ca2+ desensitisation, but that apparently the presence of either cTnI or cMyBP-C alone is insufficient to mediate this effect. It is, however, entirely unclear at present how these two proteins either directly or via additional binding partners might interact in order to modulate Ca2+ sensitivity in a PKA-dependent way. Another issue is the Ca2+ sensitisation afforded in the current animal model by the cMyBP-C deficiency itself. Studies applying various strategies to partially or totally eliminate cMyBP-C caused Ca2+ sensitivity to be either increased, unchanged, or decreased [13–15], and a unifying theory resolving these discrepancies has not been presented thus far. Finally, it was beyond the scope of this study to test for isoform expression changes with regard to titin. It has been shown that under pathological conditions associated with ventricular dilatation, titin isoforms switch to longer, more compliant isoforms [16]. Furthermore, these longer isoforms were demonstrated to undergo a smaller change in passive stiffness upon PKA-dependent phosphorylation, compared to shorter isoforms [17]. In the current model, KO hearts undergo eccentric hypertrophy with ventricular dilatation [10]. It is therefore imaginable that a titin isoform expression switch may have interfered with the experiments investigating the effect of sarcomere length change on Ca2+ sensitivity.

In conclusion, although some unresolved issues remain to be addressed in the future, it has been well established that cMyBP-C, beyond its role as a structural protein important for thick filament assembly, actively takes part in modulating myofilament Ca2+ sensitivity.


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