Thomas C Vary1, Gina Deiter, Rachel Lantry. 1. Department of Cellular and Molecular Physiology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania 17033, USA. tvary@psu.edu
Abstract
BACKGROUND: Chronic alcohol administration impairs protein synthesis ultimately causing a loss of proteins in cardiac muscle. Inhibition of protein synthesis resides in the process of mRNA translation. The present set of experiments were designed to examine the potential regulatory effect of chronic alcohol consumption on mammalian target of rapamycin (mTOR), a serine/threonine kinase important in controlling signaling cascades in the mRNA translation initiation pathway in rat hearts. METHODS: Rats were fed a diet containing ethanol for 20 to 26 weeks. Pair-fed rats served as controls. Rates of protein synthesis were measured following intravenous infusion of [(3)H]-L-phenylalanine (150 mM, 30 microCi/ml; 1 ml/100 g body weight). The phosphorylation state of mTOR, eukaryotic initiation factor 4G (eIF4G), protein kinase B (PKB) and S6K1 in heart were measured using immunoblot techniques with phospho-specific antibodies. RESULTS: Protein synthesis was reduced by 35% in animals consuming a diet containing ethanol. The fall in protein synthesis was accompanied by diminished S6K1(Thr(389)) and eIF4G (Ser(1108)) phosphorylation, both downstream effectors of mTOR signaling. These changes in phosphorylation of S6K1 and eIF4G were not associated with differences in the distribution of mTOR between TORC1 and TORC2. Instead, phosphorylation of mTOR on Ser(2448) but not on Ser(2481) was significantly reduced following feeding rats an ethanol containing diet. Decreased phosphorylation of mTOR(Ser(2448)) was not associated with a corresponding lessening of tumor suppressor complex 2 phosphorylation or expression of regulated in development and DNA damage 1, both upstream regulators of mTOR. Likewise, phosphorylation of PKB on either Ser(473) or Thr(308) was unaffected by long-term alcohol consumption. CONCLUSIONS: Chronic ethanol consumption does not alter the distribution of mTOR between TORC1 and TORC2, but instead diminishes mTOR phosphorylation on Ser(2448) independent of changes in tumor suppressor complex 2 and PKB phosphorylation. Furthermore, the data suggest that protein synthesis in rats fed a diet containing ethanol is limited by mTOR-dependent reduction in phosphorylation of S6K1(Thr(389)) and eIF4G(Ser(1108)) secondary to reduced phosphorylation of mTOR(Ser(2448)).
BACKGROUND: Chronic alcohol administration impairs protein synthesis ultimately causing a loss of proteins in cardiac muscle. Inhibition of protein synthesis resides in the process of mRNA translation. The present set of experiments were designed to examine the potential regulatory effect of chronic alcohol consumption on mammalian target of rapamycin (mTOR), a serine/threonine kinase important in controlling signaling cascades in the mRNA translation initiation pathway in rat hearts. METHODS:Rats were fed a diet containing ethanol for 20 to 26 weeks. Pair-fed rats served as controls. Rates of protein synthesis were measured following intravenous infusion of [(3)H]-L-phenylalanine (150 mM, 30 microCi/ml; 1 ml/100 g body weight). The phosphorylation state of mTOR, eukaryotic initiation factor 4G (eIF4G), protein kinase B (PKB) and S6K1 in heart were measured using immunoblot techniques with phospho-specific antibodies. RESULTS: Protein synthesis was reduced by 35% in animals consuming a diet containing ethanol. The fall in protein synthesis was accompanied by diminished S6K1(Thr(389)) and eIF4G (Ser(1108)) phosphorylation, both downstream effectors of mTOR signaling. These changes in phosphorylation of S6K1 and eIF4G were not associated with differences in the distribution of mTOR between TORC1 and TORC2. Instead, phosphorylation of mTOR on Ser(2448) but not on Ser(2481) was significantly reduced following feeding rats an ethanol containing diet. Decreased phosphorylation of mTOR(Ser(2448)) was not associated with a corresponding lessening of tumor suppressor complex 2 phosphorylation or expression of regulated in development and DNA damage 1, both upstream regulators of mTOR. Likewise, phosphorylation of PKB on either Ser(473) or Thr(308) was unaffected by long-term alcohol consumption. CONCLUSIONS: Chronic ethanol consumption does not alter the distribution of mTOR between TORC1 and TORC2, but instead diminishes mTOR phosphorylation on Ser(2448) independent of changes in tumor suppressor complex 2 and PKB phosphorylation. Furthermore, the data suggest that protein synthesis in rats fed a diet containing ethanol is limited by mTOR-dependent reduction in phosphorylation of S6K1(Thr(389)) and eIF4G(Ser(1108)) secondary to reduced phosphorylation of mTOR(Ser(2448)).
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