Original magnification, ×1,000 (upper and lower panels); ×2,000 (middle panels). (A–F) Cardiomyocytes were transfected with control siRNA (A–C) or si-cMK (D–F). Middle panels show higher magnification of boxed regions in top panels. In serum-containing conditions, si-cMK–transfected cardiomyocytes showed reduced levels of MLC2v phosphorylation (D) compared with control siRNA–transfected cardiomyocytes (A), although both exhibited regularly organized sarcomere structures. Actin staining in cardiomyocytes cultured in serum-free conditions revealed a punctuated pattern in the sarcomeres (B and E); moreover, the degree of MLC2v phosphorylation was reduced in the si-cMK–transfected cardiomyocytes compared with the control siRNA–transfected cardiomyocytes. Stimulation with 2 μM epinephrine provoked upregulation of MLC2v phosphorylation and sarcomere reassembly in control siRNA–transfected cardiomyocytes (C), but not in si-cMK–transfected cardiomyocytes (F). (G) We confirmed the levels of MLC2v phosphorylation shown in A–F using immunoblot analysis. (H) Quantitation of the levels of phosphorylated MLC2v shown in G. Values are mean ± SEM. (I) Percentage of the cells with organized sarcomeres. There was no significant difference between the populations of cardiomyocytes transfected with control siRNA and si-cMK under either serum-containing or serum-free conditions. The percentage of the cells with organized sarcomeres was significantly higher for the control siRNA–transfected cardiomyocytes than for the si-cMK–transfected cardiomyocytes. Values are mean ± SEM. p-MLC2v, phosphorylated MLC2v. **P < 0.001.