miR-1 Overexpression Enhances Ca2 Release and Promotes Cardiac Arrhythmogenesis by Targeting PP2A Regulatory Subunit B56 and Causing CaMKII-Dependent

Abstract—MicroRNAs are small endogenous noncoding RNAs that regulate protein expression by hybridization to imprecise complementary sequences of target mRNAs. Changes in abundance of muscle-specific microRNA, miR-1, have been implicated in cardiac disease, including arrhythmia and heart failure. However, the specific molecular targets and cellular mechanisms involved in the action of miR-1 in the heart are only beginning to emerge. In this study we investigated the effects of increased expression of miR-1 on excitation–contraction coupling and Ca2 cycling in rat ventricular myocytes using methods of electrophysiology, Ca2 imaging and quantitative immunoblotting. Adenoviral-mediated overexpression of miR-1 in myocytes resulted in a marked increase in the amplitude of the inward Ca2 current, flattening of Ca2 transients voltage dependency, and enhanced frequency of spontaneous Ca2 sparks while reducing the sarcoplasmic reticulum Ca2 content as compared with control. In the presence of isoproterenol, rhythmically paced, miR-1–overexpressing myocytes exhibited spontaneous arrhythmogenic oscillations of intracellular Ca2, events that occurred rarely in control myocytes under the same conditions. The effects of miR-1 were completely reversed by the CaMKII inhibitor KN93. Although phosphorylation of phospholamban was not altered, miR-1 overexpression increased phosphorylation of the ryanodine receptor (RyR2) at S2814 (CaMKII) but not at S2808 (protein kinase A). Overexpression of miR-1 was accompanied by a selective decrease in expression of the protein phosphatase PP2A regulatory subunit B56 involved in PP2A targeting to specialized subcellular domains. W