Autophagy is one of cellular survival mechanisms under starvation condition. When eukaryotic cells are in poor nutritional state, such as starvation, cells tend to utilize intracellular autophagic substrates (e.g., defective, surplus or damaged cellular macromolecules, or organelles) to generate building blocks such as amino acids for temporal adaptation to adverse condition. The forkhead/winged helix transcription factor FoxO1 promotes hepatic glucose production (HGP) by increasing expression of genes responsible for gluconeogenesis. In this study, we propose a possible mechanism how autophagy regulates gluconeogenesis in hepatocytes under starvation condition. Using an autophagy inhibitor, chloroquine (CQ) and mice primary hepatocytes, we found that HGP was suppressed under autophagy deficiency. Interestingly, the effect of autophagy inhibitor on HGP suppression was significantly reduced in FoxO1-knockout mice primary hepatocytes. Furthermore, autophagy deficiency decreased FoxO1 protein amount and expression of genes encoding gluconeogenesis rate-limiting enzymes Pck1 and G6pc, but barely affected FoxO1 mRNA level and FoxO1-S253 phosphorylation. Immunofluorescent data also suggested that less FoxO1 was translocated into nuclear under CQ-treatment. By analyzing the amino acid profile in hepatocytes in vitro with HPLC, we found that CQ-treatment significantly decreased the concentration of amino acids for leucine, isoleucine, alanine, phenylalanine, tyrosine, glutamine, glycine and valine. Importantly, when complete 20 amino acids mixture was added in CQ-treated hepatocytes under starvation, the reduction of FoxO1 protein levels was largely attenuated. Thus, we conclude that autophagy regulates gluconeogenesis in hepatocytes through supporting intracellular amino acids turnover which helps biosynthesis of FoxO1.
Z. Shen: None. Q. Pan: None. W. Yang: None. Y. Chen: None. X. Li: None. W. Ai: None. S. Guo: None.
American Diabetes Association (1-15-CD-09 to S.G.)