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CaV1.2 beta-subunit coordinates CaMKII-triggered cardiomyocyte death and afterdepolarizations.

ABSTRACT: Excessive activation of calmodulin kinase II (CaMKII) causes arrhythmias and heart failure, but the cellular mechanisms for CaMKII-targeted proteins causing disordered cell membrane excitability and myocardial dysfunction remain uncertain. Failing human cardiomyocytes exhibit increased CaMKII and voltage-gated Ca(2+) channel (Ca(V)1.2) activity, and enhanced expression of a specific Ca(V)1.2 beta-subunit protein isoform (beta(2a)). We recently identified Ca(V)1.2 beta(2a) residues critical for CaMKII phosphorylation (Thr 498) and binding (Leu 493), suggesting the hypothesis that these amino acids are crucial for cardiomyopathic consequences of CaMKII signaling. Here we show WT beta(2a) expression causes cellular Ca(2+) overload, arrhythmia-triggering cell membrane potential oscillations called early afterdepolarizations (EADs), and premature death in paced adult rabbit ventricular myocytes. Prevention of intracellular Ca(2+) release by ryanodine or global cellular CaMKII inhibition reduced EADs and improved cell survival to control levels in WT beta(2a)-expressing ventricular myocytes. In contrast, expression of beta(2a) T498A or L493A mutants mimicked the protective effects of ryanodine or global cellular CaMKII inhibition by reducing Ca(2+) entry through Ca(V)1.2 and inhibiting EADs. Furthermore, Ca(V)1.2 currents recorded from cells overexpressing CaMKII phosphorylation- or binding-incompetent beta(2a) subunits were incapable of entering a CaMKII-dependent high-activity gating mode (mode 2), indicating that beta(2a) Thr 498 and Leu 493 are required for Ca(V)1.2 activation by CaMKII in native cells. These data show that CaMKII binding and phosphorylation sites on beta(2a) are concise but pivotal components of a molecular and biophysical and mechanism for EADs and impaired survival in adult cardiomyocytes.

SUBMITTER: Koval OM

PROVIDER: S-EPMC2841943 | biostudies-literature | 2010 Mar

REPOSITORIES: biostudies-literature

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CaV1.2 beta-subunit coordinates CaMKII-triggered cardiomyocyte death and afterdepolarizations.

Koval Olha M OM Guan Xiaoquan X Wu Yuejin Y Joiner Mei-Ling ML Gao Zhan Z Chen Biyi B Grumbach Isabella M IM Luczak Elizabeth D ED Colbran Roger J RJ Song Long-Sheng LS Hund Thomas J TJ Mohler Peter J PJ Anderson Mark E ME

Proceedings of the National Academy of Sciences of the United States of America 20100301 11

Excessive activation of calmodulin kinase II (CaMKII) causes arrhythmias and heart failure, but the cellular mechanisms for CaMKII-targeted proteins causing disordered cell membrane excitability and myocardial dysfunction remain uncertain. Failing human cardiomyocytes exhibit increased CaMKII and voltage-gated Ca(2+) channel (Ca(V)1.2) activity, and enhanced expression of a specific Ca(V)1.2 beta-subunit protein isoform (beta(2a)). We recently identified Ca(V)1.2 beta(2a) residues critical for C ...[more]

PMID: 20194790

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Project description:ObjectivePathologically increased activity of Ca2+/calmodulin-dependent protein kinase II (CaMKII) and the associated Ca2+-leak from the sarcoplasmic reticulum are recognized to be important novel pharmacotherapeutic targets in heart failure and cardiac arrhythmias. However, CaMKII-inhibitory compounds for therapeutic use are still lacking. We now report on the cellular and molecular effects of a novel pyrimidine-based CaMKII inhibitor developed towards clinical use.Methods and resultsOur findings demonstrate that AS105 is a high-affinity ATP-competitive CaMKII-inhibitor that by its mode of action is also effective against autophosphorylated CaMKII (in contrast to the commonly used allosteric CaMKII-inhibitor KN-93). In isolated atrial cardiomyocytes from human donors and ventricular myocytes from CaMKIIδC-overexpressing mice with heart failure, AS105 effectively reduced diastolic SR Ca2+ leak by 38% to 65% as measured by Ca2+-sparks or tetracaine-sensitive shift in [Ca2+]i. Consistent with this, we found that AS105 suppressed arrhythmogenic spontaneous cardiomyocyte Ca2+-release (by 53%). Also, the ability of the SR to accumulate Ca2+ was enhanced by AS105, as indicated by improved post-rest potentiation of Ca2+-transient amplitudes and increased SR Ca2+-content in the murine cells. Accordingly, these cells had improved systolic Ca2+-transient amplitudes and contractility during basal stimulation. Importantly, CaMKII inhibition did not compromise systolic fractional Ca2+-release, diastolic SR Ca2+-reuptake via SERCA2a or Ca2+-extrusion via NCX.ConclusionAS105 is a novel, highly potent ATP-competitive CaMKII inhibitor. In vitro, it effectively reduced SR Ca2+-leak, thus improving SR Ca2+-accumulation and reducing cellular arrhythmogenic correlates, without negatively influencing excitation-contraction coupling. These findings further validate CaMKII as a key target in cardiovascular disease, implicated by genetic, allosteric inhibitors, and pseudo-substrate inhibitors.

Dataset Information

CaV1.2 beta-subunit coordinates CaMKII-triggered cardiomyocyte death and afterdepolarizations.

Publications

CaV1.2 beta-subunit coordinates CaMKII-triggered cardiomyocyte death and afterdepolarizations.

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