AMP-activated kinase reciprocally regulates triacylglycerol synthesis and fatty acid oxidation in liver and muscle: evidence that sn-glycerol-3-phosphate acyltransferase is a novel target

AMP-activated kinase (AMPK) is activated in response to metabolic stresses that deplete cellular ATP, and in both liver and skeletal muscle, activated AMPK stimulates fatty acid oxidation. To determine whether AMPK might reci procally regulate glycerolipid synthesis, we studied liver and skeletal-mus cle lipid metabolism in the presence of 5-amino-4-imidazolecarboxamide (AIC A) riboside, a cell-permeable compound whose phosphorylated metabolite acti vates AMPK. Adding AICA riboside to cultured rat hepatocytes for 3 h decrea sed [C-14]oleate and [H-3]glycerol incorporation into triacylglycerol (TAG) by 50% and 38% respectively, and decreased oleate labelling of diacylglyce rol by 60%. In isolated mouse soleus, a highly oxidative muscle, incubation with AICA riboside for 90 min decreased [C-14]oleate incorporation into TA G by 37% and increased (CO2)-C-14 production by 48%. When insulin was prese nt, [C-14]oleate oxidation was 49% lower and [C-14]oleate incorporation int o TAG was 62% higher than under basal conditions. AICA riboside blocked ins ulin's antioxidative and lipogenic effects, increasing fatty acid oxidation by 78% and decreasing labelled TAG 43%. Similar results on fatty acid oxid ation and acylglycerol synthesis were observed in C2C12 myoblasts, and in d ifferentiated C2C12 myotubes, AICA riboside also inhibited the hydrolysis o f intracellular TAG. These data suggest that AICA riboside might inhibit sn -glycerol-3-phosphate acyltransferase (GPAT), which catalyses the committed step in the pathway of glycerolipid biosynthesis. Incubating rat hepatocyt es with AICA riboside for both 15 and 30 min decreased mitochondrial GPAT a ctivity 22-34% without affecting microsomal GPAT, diacylglycerol acyltransf erase or acyl-CoA synthetase activities. Finally, purified recombinant AMPK alpha 1 and AMPK alpha 2 inhibited hepatic mitochondrial GPAT in a time- a nd ATP-dependent manner. These data show that AMPK reciprocally regulates a cyl-CoA channelling towards beta-oxidation and away from glycerolipid biosy nthesis, and provide strong evidence that AMPK phosphorylates and inhibits mitochondrial GPAT.