For immunoblotting, cells were lysed in Cell Lysis Buffer (Cell Signaling Technology), and 10 μg of protein was subjected to SDS–polyacrylamide gel electrophoresis [using 4 to 12% (w/v) gradient gels (Life Technologies, NuPAGE Novex, Bis-Tris)], transferred to nitrocellulose membranes, and assayed by immunoblotting. The primary antibodies were mouse anti-actin (Thermo Fisher Scientific, β-actin loading control, 1:5000) and mouse anti-myosin heavy chain (R&D Systems, 1:2000). The secondary antibody was horseradish peroxidase (HRP)–conjugated anti-mouse IgG antibody (Cell Signaling Technology, Danvers, MA, USA). An HRP substrate was used for chemiluminescent analysis (Bio-Rad, Hercules, CA, USA).

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