No abstract

AMP-activated protein kinase (AMPK) is viewed as an energy sensor that acts to modulate glucose uptake and fatty acid oxidation in skeletal muscle. Given that protein synthesis is a high energy-consuming process, it may be transiently depressed during cellular energy stress. Thus, the intent of this investigation was to examine whether AMPK activation modulates the translational control of protein synthesis in skeletal muscle. Injections of 5-aminoimidazole-4-carboxamide 1-␤-D-ribonucleoside (AICAR) were used to activate AMPK in male rats. The activity of ␣ 1 AMPK remained unchanged in gastrocnemius muscle from AICAR-treated animals compared with controls, whereas ␣ 2 AMPK activity was significantly increased (51%). AICAR treatment resulted in a reduction in protein synthesis to 45% of the control value. This depression was associated with decreased activation of protein kinases in the mammalian target of rapamycin (mTOR) signal transduction pathway as evidenced by reduced phosphorylation of protein kinase B on Ser 473 , mTOR on Ser 2448 , ribosomal protein S6 kinase on Thr 389 , and eukaryotic initiation factor eIF4E-binding protein on Thr 37 . A reduction in eIF4E associated with eIF4G to 10% of the control value was also noted. In contrast, eIF2B activity remained unchanged in response to AICAR treatment and therefore would not appear to contribute to the depression in protein synthesis. This is the first investigation to demonstrate changes in translation initiation and skeletal muscle protein synthesis in response to AMPK activation.Considerable attention has focused on understanding the role of AMP-activated protein kinase (AMPK) 1 in monitoring the energy status of the cell and mediating subsequent metabolic events. AMPK has been referred to as an energy-sensing/ signaling protein within the cell that responds to changes in the ratio of ATP/AMP as well as phosphocreatine/creatine (1, 2). Changes in the cellular energy state activate AMPK through various mechanisms involving allosteric regulation of AMPK, activation by an upstream AMPK kinase, and diminished activity of phosphatases (3). AMPK activation increases glucose uptake and fatty acid oxidation in muscle (4) as well as up-regulates expression of various metabolic genes (e.g. the glucose transporter, GLUT4, uncoupling protein-3, and cytochrome c) (5-7). Consequently AMPK serves as a sensor/modulator of intermediary metabolism by directing cellular events to increase energy availability and sustain high energy phosphate levels. Research using in vitro systems has shown that AMPK can be activated under artificial conditions such as treatment with high fructose or 2-deoxyglucose, heat shock, and inhibitors of oxidative phosphorylation (3). Pharmacological use of 5-aminoimidazole-4-carboxamide 1-␤-D-ribonucleoside (AICAR) has been commonly utilized to directly activate AMPK without altering cellular concentrations of ATP, ADP, and AMP (8). Additionally, starvation and endurance exercise result in increased activity of AMPK in skeletal muscle (9 -11). Ex...

The objectives of the present study were twofold: 1) to determine whether leucine is unique among the branched-chain amino acids (BCAA) in its ability to stimulate protein synthesis in skeletal muscle of food-deprived rats; and 2) to investigate whether changes in muscle protein synthesis after leucine administration involve a signaling pathway that includes the protein kinase mammalian target of rapamycin (mTOR). In the first set of experiments, food-deprived (18 h) male rats (200 g) were orally administered saline or 270 mg valine, isoleucine or leucine. In the second set of experiments, food-deprived rats were injected intravenously with rapamycin (0.75 mg/kg), a specific inhibitor of mTOR, before leucine administration. Only leucine stimulated protein synthesis in skeletal muscle above saline-treated controls (P: < 0.05). Furthermore, leucine was most effective among the BCAA at enhancing phosphorylation of eukaryotic initiation factor (eIF), 4E binding protein 1 (4E-BP1) and the 70-kDa ribosomal protein S6 kinase (S6K1). Leucine-dependent hyperphosphorylation of 4E-BP1 increased the availability of eIF4E to form the active eIF4G.eIF4E complex. To a lesser extent, isoleucine also enhanced phosphorylation of 4E-BP1 and S6K1. Rapamycin inhibited protein synthesis in both leucine-treated and food-deprived rats. Additionally, rapamycin prevented the stimulatory effects of leucine on eIF4E availability for binding eIF4G and inhibited leucine-dependent phosphorylation of S6K1. The data demonstrate that leucine is unique among the BCAA in its ability to stimulate protein synthesis in muscle of food-deprived rats. We show for the first time that leucine-dependent stimulation of translation initiation in vivo occurs via a rapamycin-sensitive pathway.