Abstract
Calcium (Ca2+) is the key intracellular signalling molecule in the heart. It is stored within the sarcoplasmic reticulum (SR) and triggers myocardial contraction upon release into the cytosol. The release of Ca2+ from the SR is gated by the ryanodine receptor II (RyR2). Thus, timely opening and closing of RyR2 dictate cardiac contraction and relaxation periods. To effectively mediate Ca2+ release, neighbouring RyR2 proteins form clusters. However, in disease, excessive RyR2 phosphorylation via Protein kinase A (PKA) and Ca2+/calmodulin dependent kinase II (CaMKII) disrupts typical cluster organisation, leading to abnormal spontaneous Ca2+ release (SCR) and arrhythmia. The Jones lab has identified novel protein kinase CK2 phosphorylation sites of RyR2 that reduce SCR.
To investigate whether CK2 phosphorylation reduces SCR due to changes in RyR2 cluster organisation, phosphomimetic RyR2 mutants that mimic permanent CK2 phosphorylation (CK2+/+) and permanent dephosphorylation (CK2-/-) were compared to wild-type (WT) controls (N=8 for all genotypes). Three-month-old mice were euthanised, hearts were extracted, and ventricular samples were prepared for microscopy. Cluster area, the number of RyR2 channels within a cluster (cluster size), and the number of clusters within a cardiomyocyte (cluster density) were determined. All comparisons were made using one-way ANOVA.
Surprisingly, cluster area, cluster size, and cluster density between genotypes were not different in CK2+/+ or CK2-/- mice compared to WT controls (P > 0.05).
The lack of difference indicates that CK2 phosphorylation does not affect RyR2 cluster organisation in cardiomyocytes. This suggests that the underlying mechanism by which CK2 phosphorylation suppresses SCR is independent of cluster organisation, unlike other phosphorylation events that disrupt RyR2 cluster organisation in disease. Future studies on how CK2 phosphorylation of RyR2 mediates its anti-arrhythmic effect may provide insights that reveal a possible novel therapeutic target against arrhythmia.