Rationale: Failing cardiomyocytes exhibit decreased efficiency of excitation-contraction (E-C) coupling. The downregulation of junctophilin-2 (JP2), a protein anchoring the sarcoplasmic reticulum to T-tubules, has been identified as a major mechanism underlying the defective E-C coupling. However, the regulatory mechanism of JP2 remains unknown.

Objective: To determine whether microRNAs regulate JP2 expression.

Methods and Results: Bioinformatic analysis predicted 2 potential binding sites of miR-24 in the 3′-untranslated regions of JP2 mRNA. Luciferase assays confirmed that miR-24 suppressed JP2 expression by binding to either of these sites. In the aortic stenosis model, miR-24 was upregulated in failing cardiomyocytes. Adenovirus-directed overexpression of miR-24 in cardiomyocytes decreased JP2 expression and reduced Ca²⁺ transient amplitude and E-C coupling gain.

Conclusions: MiR-24–mediated suppression of JP2 expression provides a novel molecular mechanism for E-C coupling regulation in heart cells and suggests a new target against heart failure.