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This Article Full Text Full Text (PDF) Supplemental Figures All Versions of this Article: 297/2/E304    most recent 91007.2008v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Google Scholar Articles by Williamson, D. L. Articles by Alway, S. E. PubMed PubMed Citation Articles by Williamson, D. L. Articles by Alway, S. E.

AMPK inhibits myoblast differentiation through a PGC-1-dependent mechanism

Division of Exercise Physiology, West Virginia University School of Medicine, Morgantown, West Virginia

Submitted 16 December 2008 ; accepted in final form 26 May 2009

Elevated phosphorylation of AMP-activated protein kinase (AMPK) has been shown to inhibit skeletal muscle growth in both culture and animal models, but its role in differentiation of muscle cells is less clear. p21 is known to have an important role in differentiation, but AMPK's role regulating p21 in differentiation in muscle cultures is unknown. Therefore, the purpose of this study was to determine the role of p21 in differentiation of skeletal muscle cells under conditions of elevated AMPK phosphorylation. Treating C₂C₁₂ myoblast cultures with 1 mM 5-aminoimidazole-4-carboxamide 1-β-D-ribonucleoside (AICAR) for up to 24 h induced AMPK phosphorylation. Activation of AMPK reduced p21 protein and mRNA expression, which was associated with reduced G₁/S cell cycle transition and p21 promoter activity. AICAR-treated myoblasts undergoing differentiation also had reduced p21 protein expression, reduced myotube formation, and myosin accumulation. When myotube cultures were treated with AICAR for 24 h, p21, myosin protein expression, and MyoD were significantly reduced. Myotube atrophy was also apparent compared with control conditions. Addition of compound C, an AMPK inhibitor, attenuated AICAR's negative effects on the myotube cultures. The nuclear expression of p21 protein appeared to be more affected by AICAR-treated myotubes than the cytosolic portion of p21 protein, which was attenuated with compound C treatment. Further analysis revealed that AICAR treatment increased PGC-1 and decreased FOXO3A protein expression, which was reversed with compound C cotreatment. Knockdown of PGC-1 with shRNA corroborated the compound C data, preserving nuclear FOXO3A and p21 protein expression. These data demonstrate that AICAR-induced AMPK phosphorylation inhibits cell cycle transition, reducing differentiation of myoblasts into myotubes, through PGC-1-FOXO3A-p21.

adenosine 5'-monophosphate-activated protein kinase; myotube; peroxisome proliferator-activated receptor- coactivator-1; 5-aminoimidazole-4-carboxamide-1-β-D-ribonucleoside; cell cycle; myogenesis