title: Mammalian target of rapamycin is a direct target for protein kinase B: identification of a convergence point for opposing effects of insulin and amino-acid deficiency on protein translation.
abstract: Growth factor induced activation of phosphoinositide 3-kinase and protein kinase B (PKB) leads to increased activity of the mammalian target of rapamycin (mTOR). This subsequently leads to increased phosphorylation of eIF4E binding protein-1 (4EBP1) and activation of p70 ribosomal S6 protein kinase (p70(S6K)), both of which are important steps in the stimulation of protein translation. The stimulation of translation is attenuated in cells deprived of amino acids and this is associated with the attenuation of 4EBP1 phosphorylation and p70(S6K) activation. It has been suggested that PKB regulates mTOR function by phosphorylation although direct phosphorylation of mTOR by PKB has not been demonstrated previously. In the present work, we have found that PKB directly phosphorylates mTOR and, using phosphospecific antibodies, we have shown this phosphorylation occurs at Ser(2448). Insulin also induces phosphorylation on Ser(2448) and this effect is blocked by wortmannin but not rapamycin, consistent with the effect being mediated by PKB. Amino-acid starvation rapidly attenuated the reactivity of the Ser(2448) phosphospecific antibody with mTOR and this could not be restored by either insulin stimulation of cells or incubation with PKB in vitro. Our findings demonstrate that mTOR is a direct target for PKB and support the conclusion that regulation of phosphorylation of Ser(2448) is a point of convergence for the counteracting regulatory effects of growth factors and amino acid levels.
authors.affiliation: Department of Biochemistry, University College London, Gower St, London WC1E 6BT, U.K.
authors.forename: B T
authors.initials: BT
authors.lastname: Navé
authors.forename: M
authors.initials: M
authors.lastname: Ouwens
authors.forename: D J
authors.initials: DJ
authors.lastname: Withers
authors.forename: D R
authors.initials: DR
authors.lastname: Alessi
authors.forename: P R
authors.initials: PR
authors.lastname: Shepherd
mesh_terms: D000596:Amino Acids / Q000172:deficiency*; D000730:Androstadienes / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D014176:Protein Biosynthesis; D011494:Protein Kinases; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt; D015398:Signal Transduction; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D000077191:Wortmannin
chemical_list: D000596:Amino Acids; D000730:Androstadienes; D007328:Insulin; D000081082:Phosphoinositide-3 Kinase Inhibitors; D011518:Proto-Oncogene Proteins; D011494:Protein Kinases; D017853:Phosphotransferases (Alcohol Group Acceptor); D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus; D000077191:Wortmannin