PKA phosphorylation of cardiac ryanodine receptor modulates SR luminal Ca2+ sensitivity

• Published in: Journal of molecular and cellular cardiology 2012-07, Vol.53 (1), p.33-42
• Main Authors: Ullrich, Nina D, Valdivia, Héctor H, Niggli, Ernst
• Format: Article
• Language: English
• Subjects: Adrenergic beta-Agonists - pharmacology; Animals; Ca2+ imaging; Calcium - metabolism; Cardiovascular; Cyclic AMP-Dependent Protein Kinases - metabolism; Diastole - drug effects; Excitation Contraction Coupling - drug effects; Excitation–contraction coupling; Lipid Bilayers - metabolism; Mice; Mice, Transgenic; Myocardial Contraction - drug effects; Myocardial Contraction - physiology; Myocytes, Cardiac - drug effects; Myocytes, Cardiac - metabolism; Phosphorylation; PKA; Ryanodine Receptor Calcium Release Channel - genetics; Ryanodine Receptor Calcium Release Channel - metabolism; Ryanodine receptor phosphorylation; Sarcoplasmic reticulum; Sarcoplasmic Reticulum - metabolism
• ISSN: 0022-2828; 1095-8584
• DOI: 10.1016/j.yjmcc.2012.03.015

Abstract During physical exercise and stress, the sympathetic system stimulates cardiac contractility via β-adrenergic receptor activation, resulting in protein kinase A (PKA)-mediated phosphorylation of the cardiac ryanodine receptor, RyR2, at Ser2808. Hyperphosphorylation of RyR2-S2808 has been proposed as a mechanism contributing to arrhythmogenesis and heart failure. However, the role of RyR2 phosphorylation during β-adrenergic stimulation remains controversial. We examined the contribution of RyR2-S2808 phosphorylation to altered excitation–contraction coupling and Ca2+ signaling using an experimental approach at the interface of molecular and cellular levels and a transgenic mouse with ablation of the RyR2-S2808 phosphorylation site (RyR2-S2808A). Experimentally challenging the communication between L-type Ca2+ channels and RyR2 led to a spatiotemporal de-synchronization of RyR2 openings, as visualized using confocal Ca2+ imaging. β-Adrenergic stimulation re-synchronized RyR2s, but less efficiently in RyR2-S2808A than in control cardiomyocytes, as indicated by comprehensive analysis of RyR2 activation. In addition, spontaneous Ca2+ waves in RyR2-S2808A myocytes showed significantly slowed propagation and complete absence of acceleration during β-adrenergic stress, unlike wild type cells. Single channel recordings revealed an attenuation of luminal Ca2+ sensitivity in RyR2-S2808A channels upon addition of PKA. This suggests that phosphorylation of RyR2-S2808 may be involved in RyR2 modulation by luminal (intra-SR) Ca2+ ([Ca2+ ]SR ). We show here by three independent experimental approaches that PKA-dependent RyR2-S2808 phosphorylation plays significant functional roles at the subcellular level, namely, Ca2+ release synchronization, Ca2+ wave propagation and functional adaptation of RyR2 to variable [Ca2+ ]SR . These results indicate a direct mechanistic link between RyR2 phosphorylation and SR luminal Ca2+ sensing.