Sepsis and AMPK Activation by AICAR Differentially Regulate FoxO-1, -3 and -4 mRNA in Striated Muscle

Although much is known regarding the posttranslational regulation of the FoxO transcription factors, there is little appreciation of how stressors which regulate cellular energy status effect the various FoxO family members at the mRNA level. The hypothesis of the present study was that exposure of differentiated muscle cells to agonists of AMP-activated protein kinase (AMPK) would increase the mRNA content of various FoxO mRNA transcripts. Stimulation of AMPK in vivo by the injection of AICAR into mice increased FoxO1 and FoxO3 (but not FoxO4) mRNA in skeletal muscle. A comparable increase in these FoxO mRNAs was seen in skeletal muscle in response to sepsis which also increased AMPK phosphorylation. In contrast to the in vivo data, FoxO1, 3 and 4 mRNA content was decreased dose-dependently, with the decrement in FoxO1 being the largest, in C(2)C1(2) myotubes incubated with the AMPK agonists AICAR or metformin. Treatment of myotubes with 2-deoxyglucose or reducing the media glucose concentration also decreased mRNA content for FoxO1 and FoxO4. All stressors increased AMPK phosphorylation under in vitro conditions. Incubation of myotubes with AICAR decreased the rate of protein synthesis and increased protein degradation. Finally, treatment with the AMPK inhibitor compound C prevented both the AICAR-induced changes in FoxO mRNA and changes in protein metabolism. Our data indicate FoxO mRNA expression is down-regulated by AMPK activation and energy depletion in cultured myotubes, but that a contrasting increase in FoxO1 and FoxO3 mRNA is observed in vivo with the agent (and in response to sepsis) suggesting the expression of these FoxOs may be controlled by other hormonal or energy sensing cues under in vivo conditions.