Identification of S6 kinase 1 as a novel mammalian target of rapamycin (mTOR)-phosphorylating kinase.
Here we demonstrate that mammalian target of rapamycin (mTOR) is phosphorylated in a rapamycin-sensitive manner. We show that S6 kinase 1 (S6K1), but not Akt, directly phosphorylates mTOR in cell-free in vitro system and in cells. Expression of a constitutively active, rapamycin- and wortmannin-resistant S6K1 leads to constitutive phosphorylation of mTOR, whereas knock-down of S6K1 using small inhibitory RNA greatly reduces mTOR phosphorylation despite elevated Akt activity. Importantly, phosphorylation of mTOR by S6K1 occurs at threonine 2446/serine 2448. This region has been shown previously to be part of a regulatory repressor domain. These sites are also constitutively phosphorylated in the breast cancer cell line MCF7 carrying an amplification of the S6K1 gene, but not in a less tumorigenic cell line, MCF10a. Many models for Akt signaling to mTOR have been presented, suggesting direct phosphorylation by Akt. These models must be reconsidered in light of the present findings.
Holz
Marina K
MK
Department of Cell Biology, Harvard Medical School, Boston, Massachusetts 02115, USA.
Blenis
John
J
D000483:Alleles; D000730:Androstadienes / Q000494:pharmacology; D002460:Cell Line; D045744:Cell Line, Tumor; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005822:Genetic Vectors; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D007328:Insulin / Q000378:metabolism; D008954:Models, Biological; D010703:Phorbol Esters / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D038744:Ribosomal Protein S6 Kinases, 90-kDa / Q000378:metabolism / Q000502:physiology*; D012694:Serine / Q000737:chemistry; D015398:Signal Transduction; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D013912:Threonine / Q000737:chemistry; D013997:Time Factors; D000077191:Wortmannin
D000730:Androstadienes; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D007328:Insulin; D010703:Phorbol Esters; D019869:Phosphatidylinositol 3-Kinases; D011494:Protein Kinases; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D038744:Ribosomal Protein S6 Kinases, 90-kDa; D012694:Serine; D020123:Sirolimus; D058570:TOR Serine-Threonine Kinases; D013912:Threonine; D000077191:Wortmannin; C546842:MTOR protein, human; C069329:RPS6KA1 protein, human