Direct inhibition of the signaling functions of the mammalian target of rapamycin by the phosphoinositide 3-kinase inhibitors, wortmannin and LY294002.
The immunosuppressant, rapamycin, inhibits cell growth by interfering with the function of a novel kinase, termed mammalian target of rapamycin (mTOR). The putative catalytic domain of mTOR is similar to those of mammalian and yeast phosphatidylinositol (PI) 3-kinases. This study demonstrates that mTOR is a component of a cytokine-triggered protein kinase cascade leading to the phosphorylation of the eukaryotic initiation factor-4E (eIF-4E) binding protein, PHAS-1, in activated T lymphocytes. This event promotes G1 phase progression by stimulating eIF-4E-dependent translation initiation. A mutant YAC-1 T lymphoma cell line, which was selected for resistance to the growth-inhibitory action of rapamycin, was correspondingly resistant to the suppressive effect of this drug on PHAS-1 phosphorylation. In contrast, the PI 3-kinase inhibitor, wortmannin, reduced the phosphorylation of PHAS-1 in both rapamycin-sensitive and -resistant T cells. At similar drug concentrations (0.1-1 microM), wortmannin irreversibly inhibited the serine-specific autokinase activity of mTOR. The autokinase activity of mTOR was also sensitive to the structurally distinct PI 3-kinase inhibitor, LY294002, at concentrations (1-30 microM) nearly identical to those required for inhibition of the lipid kinase activity of the mammalian p85-p110 heterodimer. These studies indicate that the signaling functions of mTOR, and potentially those of other high molecular weight PI 3-kinase homologs, are directly affected by cellular treatment with wortmannin or LY294002.
Department of Pharmacology, Mayo Clinic, Rochester, MN 55905, USA.
G J
GJ
Brunn
J
J
Williams
C
C
Sabers
G
G
Wiederrecht
J C
JC
Lawrence
R T
RT
Abraham
D048868:Adaptor Proteins, Signal Transducing; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D000730:Androstadienes / Q000494:pharmacology*; D000818:Animals; D001923:Brain Chemistry; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology*; D004229:Dithiothreitol / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D039642:Eukaryotic Initiation Factors; D007166:Immunosuppressive Agents / Q000494:pharmacology; D007376:Interleukin-2 / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D008213:Lymphocyte Activation; D016399:Lymphoma, T-Cell; D051379:Mice; D009025:Morpholines / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000302:isolation & purification; D011090:Polyenes / Q000494:pharmacology; D011494:Protein Kinases; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus; D013601:T-Lymphocytes / Q000276:immunology; D013602:T-Lymphocytes, Cytotoxic; D058570:TOR Serine-Threonine Kinases; D014407:Tumor Cells, Cultured; D000077191:Wortmannin
D048868:Adaptor Proteins, Signal Transducing; D000255:Adenosine Triphosphate; D000730:Androstadienes; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D002867:Chromones; D004229:Dithiothreitol; D004791:Enzyme Inhibitors; D039642:Eukaryotic Initiation Factors; D007166:Immunosuppressive Agents; D007376:Interleukin-2; D047908:Intracellular Signaling Peptides and Proteins; D009025:Morpholines; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D017853:Phosphotransferases (Alcohol Group Acceptor); D011090:Polyenes; D011494:Protein Kinases; D011993:Recombinant Fusion Proteins; D020123:Sirolimus; D058570:TOR Serine-Threonine Kinases; D000077191:Wortmannin; C499527:Eif4ebp1 protein, mouse; C499526:Eif4ebp1 protein, rat; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; C022571:adenosine 5'-O-(3-thiotriphosphate); C546843:mTOR protein, mouse; C546845:mTOR protein, rat