AMP-activated protein kinase stabilizes FOXO3 in primary myotubes

AMP-activated protein kinase (AMPK) is a critical enzyme in conditions of cellular energy deficit such as exercise, hypoxia or nutritional stress. AMPK is well known to regulate protein degradation pathways notably through FOXO-related axis. In this study, we investigated the implication of AMPK activation in FOXO3 expression and stability in skeletal muscle primary myotubes. First, time course and dose response studies revealed optimal AICAR treatment duration and dose in skeletal muscle cells. Then, experiments with cycloheximide treatment of primary myotubes highlighted that AICAR infusion extends FOXO3 protein half-life. Our results also showed that AICAR treatment or nutrient depletion increases FOXO3 expression in primary myotubes and the expression of the mitochondrial E3 ligase Mul1 involved in mitochondrial turnover (mitophagy). In AMPK KO cells, nutrient depletion failed to alter the level of some FOXO3-dependent atrophic genes, including LC3B, BNIP3, and the mitochondrial E3 ligase Mul1, but not the expression of other genes (i.e. FOXO1, Gabarapl1, MAFbx, MuRF1). In summary, our data highlight that AMPK stabilizes FOXO3 and suggest a role in the first initiation step of mitochondrial segregation in muscle cells.