Rebalancing Protein Phosphorylation in Heart Failure to Prevent Arrhythmias

European Journal of Heart FailureVolume 20, Issue 12 p. 1686-1689 Editorial CommentFree Access Rebalancing protein phosphorylation in heart failure to prevent arrhythmias Vasco Sequeira, Vasco Sequeira Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Würzburg, GermanySearch for more papers by this authorChristoph Maack, Corresponding Author Christoph Maack maack_c@ukw.de Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Würzburg, GermanyCorresponding author. Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Am Schwarzenberg 15, Haus A15, 97078 Würzburg, Germany. Tel: +49 931 201-46502, Email: maack_c@ukw.deSearch for more papers by this author Vasco Sequeira, Vasco Sequeira Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Würzburg, GermanySearch for more papers by this authorChristoph Maack, Corresponding Author Christoph Maack maack_c@ukw.de Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Würzburg, GermanyCorresponding author. Comprehensive Heart Failure Center (CHFC), University Clinic Würzburg, Am Schwarzenberg 15, Haus A15, 97078 Würzburg, Germany. Tel: +49 931 201-46502, Email: maack_c@ukw.deSearch for more papers by this author First published: 17 October 2018 https://doi.org/10.1002/ejhf.1315Citations: 3 The opinions expressed in this article are not necessarily those of the Editors of the European Journal of Heart Failure or of the European Society of Cardiology. doi: 10.1002/ejhf.1297 AboutSectionsPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat This article refers to ‘Activation of protein phosphatase 1 by a selective phosphatase disrupting peptide reduces sarcoplasmic reticulum Ca2+ leak in human heart failure’ by T.H. Fischer et al., published in this issue on pages xxx. In Europe, more than 15 million people are diagnosed with heart failure (HF), and this number continues to increase, representing a major cause of hospitalization and death. About half of the HF patient population presents with reduced ejection fraction (HFrEF).1 The most common forms of death or hospitalization of HFrEF patients are pump failure and arrhythmias,2, 3 and for both conditions, alterations of cardiac myocyte calcium (Ca2+) handling play an important role. Cardiac myocytes tightly control their Ca2+ homeostasis in a process coined excitation–contraction (EC) coupling. Upon membrane depolarization during the cardiac action potential, Ca2+ enters myocytes via L-type Ca2+ channels. This Ca2+ influx triggers a much larger intracellular Ca2+ release from the Ca2+ store, the sarcoplasmic reticulum (SR)4 (Figure 1). This Ca2+ activates the myofilament machinery to contract during systole. During diastole, Ca2+ diffuses from the myofilaments and is taken back up into the SR (∼70%) by the SR Ca2+ ATPase (SERCA), or exported by the Na+/Ca2+ exchanger, and the ensuing decrease of intracellular Ca2+ initiates relaxation (Figure 1). The energy for these processes comes from mitochondria, which turn over the cellular ATP pool in less than a minute. Figure 1Open in figure viewerPowerPoint Mechanisms of excitation–contraction coupling in cardiac myocytes and its control by Ca2+/calmodulin-dependent protein kinase II (CaMKII) and protein kinase A (PKA). For details see text. ADP, adenosine diphosphate; AR, adrenergic receptor; ATP, adenosine triphosphate; Ca, calcium; cAMP, cyclic adenosine monophosphate; Epac, exchange protein directly activated by cyclic AMP; INa, Na current; Na, sodium; NCX, Na+/Ca2+ exchanger; ROS, reactive oxygen species; RyR, ryanodine receptor; SERCA, sarcoplasmic reticulum Ca2+ ATPase; T-tubule, transversal tubule. The spatiotemporal control of Ca2+ homeostasis is governed by protein phosphorylation through kinases, increasing and accelerating contractility (inotropy), relaxation (lusitropy), and heart rate (chronotropy). Noradrenaline-induced activation of β-adrenoceptors (β-AR) stimulates cyclic adenosine monophosphate (cAMP) production, followed by protein kinase A (PKA) activation (Figure 1). Phosphorylation by PKA activates Ca2+ channels to increase Ca2+ influx, promotes Ca2+ release from the SR via ryanodine receptors, accelerates Ca2+ uptake by SERCA and reduces myofilament Ca2+ affinity (Figure 1, right).4 Activation of Epac (exchange protein directly activated by cAMP) additionally activates the Ca2+/calmodulin-dependent protein kinase II (CaMKII), which has many functions that are redundant to PKA (such as activation of Ca2+ channels, ryanodine receptors and SERCA), but in addition, activates Na+ channels to increase Na+ influx via the cell membrane (Figure 1, right).5, 6 In failing myocardium, contractile dysfunction is directly linked to disturbed Ca2+ handling, and a central defect is decreased Ca2+ load of — and release from the Ca2+ store — the SR.4 Two main mechanisms underlie this defect: (i) increased diastolic Ca2+ leakage from the SR via ryanodine receptors, and (ii) decreased expression and activity of SERCA.4 As a consequence, less Ca2+ is released during each heartbeat to activate the myofilaments, reducing systolic force generation, and more Ca2+ accumulates in the cytosol during diastole, increasing diastolic tension (Figure 1). Moreover, intracellular Na+ levels are elevated in failing myocytes due to abnormal activities of Na+ channels and exchangers.7 While Na+ elevation partly compensates for reduced systolic Ca2+ release from the SR via facilitating cytosolic Ca2+ influx via the reverse mode of the Na+/Ca2+ exchanger, this slows relaxation (Figure 1, left)4 and impairs mitochondrial Ca2+ uptake required for energy supply and demand matching.8 Furthermore, elevated diastolic Ca2+ prolongs action potential duration and thereby increases the risk for early afterdepolarizations, which can trigger ventricular arrhythmias.4 In HF, sustained β-adrenergic stimulation causes β-AR desensitization and downregulation through β-AR phosphorylation by PKA and/or G-protein-coupled receptor kinase, the latter facilitating β-arrestin-mediated receptor internalization upon longer-term stimulation.9 In contrast to PKA, expression and activity of CaMKII is increased in HF patients10, 11 and maintains its effects on EC coupling beyond desensitization of the β-AR/PKA system.12 This increased activity is likely the result of elevated diastolic Ca2+ levels, β1-AR stimulation with subsequent β-arrestin activation, oxidative and/or nitrosative stress, indicating that CaMKII is a central signalling hub that integrates various forms of stress toward cellular Ca2+ handling (Figure 1, left), but also pro-hypertrophic and death pathway signalling.13 In fact, increased CaMKII activity is considered a central defect underlying aberrant Ca2+ handling and, thereby, contractile dysfunction and the risk of arrhythmias in HF.5, 6 While PKA and CaMKII are the central protein kinases that phosphorylate key channels and transporters involved in EC coupling, their natural counter-players are protein phosphatases (PP) that dephosphorylate these targets. In fact, about one third of all protein phosphorylation can be reversed by PP,14 and in the heart, this is achieved by PP1, PP2A and PP2B.15 In particular, PP1 counterbalances β-AR-induced (and PKA- and/or CaMKII-mediated) activation of L-type Ca channels, ryanodine receptors and SERCA activity, thereby opposing the positive inotropic, lusitropic and chronotropic effects of these kinases (Figure 2, right).16, 17 In animal models18, 19 and human HF,20, 21 expression and activity of PP1 is increased, while the one of its endogenous inhibitor I-1 is reduced, presumably in an attempt to counterbalance hyper-phosphorylation imposed by increased CaMKII activity. Figure 2Open in figure viewerPowerPoint Impact of the novel cell-permeable peptide drug PDP3 on cytosolic Ca2+ handling and arrhythmias. PDP3 activates protein phosphatase 1 (PP1), which in turn dephosphorylates ryanodine receptors (RyR). This reduces the leak of Ca2+ from the sarcoplasmic reticulum, which in turn reduces the frequency of spontaneous Ca2+ release events (Ca2+ sparks) and, thereby, arrhythmias. ADP, adenosine diphosphate; ATP, adenosine triphosphate; Ca, calcium; CaMKII, Ca2+/calmodulin-dependent protein kinase II; INa, Na current; Na, sodium; NCX, Na+/Ca2+ exchanger; SERCA, sarcoplasmic reticulum Ca2+ ATPase; T-tubule, transversal tubule. Despite a wealth of knowledge on the mechanisms underlying arrhythmias in HF, it is a therapeutic challenge to tackle these without interfering with inotropy.2, 3 In this issue of the Journal, Fischer and colleagues22 provide evidence that instead of inhibiting CaMKII, which may have undesired implications for several important physiological functions not only in the heart,23 it may be a more promising approach to tackle the problem from the other side: by activating specifically PP1. In an elegant and technically demanding study performed on isolated cardiac myocytes from patients with either compensated hypertrophy (obtained from patients with aortic stenosis upon surgical valve replacement) or HF (obtained from explanted hearts during heart transplantation), they show that activation of PP1 with the novel cell-penetrating peptide compound PDP324, 25 ameliorates cellular arrhythmias without reducing steady-state cytosolic Ca2+ transients, which govern contractility (Figure 2). In a careful experimental approach, they first analyse the human cardiac tissue biochemically to show that both CaMKII phosphorylation and PP1 expression are increased in failing compared to nonfailing or hypertrophied myocardium, confirming previous results.10, 11, 20, 21 Isolated cardiac myocytes from failing hearts displayed a stronger Ca2+ leak from the SR than myocytes from patients with compensated hypertrophy, which (despite decreased SR Ca2+ load) predisposes to spontaneous Ca2+ release events (so-called ‘Ca2+ sparks’) as triggers of delayed afterdepolarizations that can induce arrhythmias in whole hearts. Dismantling the strong control of Ca2+ handling exerted by PPs, the non-specific PP inhibitor okadaic acid substantially increased SR Ca2+ leak and arrhythmias, indicating that PPs act as natural endogenous ‘breaks’ on arrhythmias. Conversely, activating specifically PP1 with PDP3 ‘sealed’ the Ca2+ leak of the SR and prevented spontaneous SR Ca2+ release events and, thereby, cellular arrhythmias in hypertrophied cardiac myocytes (Figure 2). Despite its favourable effect on arrhythmias, steady-state cytosolic Ca2+ transients and SR Ca2+ load were not reduced, what might have been expected on simplistic grounds by PP1-mediated indirect antagonization of CaMKII-induced phosphorylation. This implies that PP2 activation by PDP3 may exert its anti-arrhythmic effects without compromising systolic function in patients with HF. The authors have to be congratulated for addressing a clinically relevant issue with a sophisticated and yet highly translational approach by going the long and rocky way of using cells and myocardium from patients with HF and cardiac hypertrophy. This effort, however, is well justified since (i) EC coupling is fundamentally different in mice compared to humans, and (ii) animal models of HF rarely reflect the real complexity of human disease. However, while the presented results are a promising first step toward developing novel anti-arrhythmic drugs, several open issues need to be resolved and some additional hurdles to be overcome. First, since dephosphorylation of the myofilaments by PPs could increase their affinity for Ca2+, PDP3 may have a relevant impact on the metabolic consequences of contraction and relaxation. While it is currently still controversial whether in patients with HFrEF, myofilament Ca2+ sensitivity is increased or decreased,26, 27 it is important to note that in patients with hypertrophic cardiomyopathy, mutations of genes encoding sarcomeric proteins commonly increase myofilament Ca2+ sensitivity, and this induces energetic imbalance and paves the way toward arrhythmias and maladaptive cardiac remodelling.28 Therefore, phospho-proteomic and functional studies should clarify how PDP3 impacts on sarcomeric function and energetic aspects of EC coupling (Figure 2). Second, the compound PDP3 is still a long way from being applicable in humans. In fact, the development of compounds targeting PP1 has been challenging due to the vast abundance of PP1-interacting proteins (up to 200 in vertebrates), which govern PP1 localization and function.25, 29 In fact, PDP3 specifically targets PP1 by disrupting its interaction with a subset of its interacting proteins.24 However, to achieve sufficient concentrations within cells, quite high concentrations of PDP3 are required, and the accessibility of whole tissue (such as myocardium) is not fully resolved yet. Furthermore, prolonged treatment duration with PDP3 was toxic for non-cardiac cells,24 and its systemic effects in vivo are currently unknown. Nevertheless, the current study is an important proof of concept that may stimulate further research and drug development toward fine-tuning the fragile balance between protein phosphorylation and dephosphorylation in cardiac myocytes, thereby optimizing EC coupling and preventing arrhythmias in patients with HF. Funding Christoph Maack is funded by the Deutsche Forschungsgemeinschaft (DFG; Ma 2528/7-1, SFB-894, TRR-219) and the German Ministry of Education and Science (BMBF). Conflict of interest: C.M. is an advisor to Servier and received speaker honoraria from Bayer, Bristol-Myers Squibb, Boehringer Ingelheim, Berlin Chemie, Daiichi-Sankyo, Novartis, and Pfizer. V.S. has no conflict of interest to disclose. References 1Borlaug BA, Paulus WJ. Heart failure with preserved ejection fraction: pathophysiology, diagnosis, and treatment. Eur Heart J 2011; 32: 670– 679. 2Lip GY, Heinzel FR, Gaita F, Juanatey JR, Le Heuzey JY, Potpara T, Svendsen JH, Vos MA, Anker SD, Coats AJ, Haverkamp W, Manolis AS, Chung MK, Sanders P, Pieske B, Gorenek B, Lane D, Boriani G, Linde C, Hindricks G, Tsutsui H, Homma S, Brownstein S, Nielsen JC, Lainscak M, Crespo-Leiro M, Piepoli M, Seferovic P, Savelieva I. 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