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Isolation of a protein target of the FKBP12-rapamycin complex in mammalian cells.

The immunosuppressive drug, rapamycin, interferes with an undefined signaling pathway required for the progression of G1-phase T-cells into S phase. Genetic analyses in yeast indicate that binding of rapamycin to its intracellular receptor, FKBP12, generates a toxic complex that inhibits cell growth in G1 phase. These analyses implicated two related proteins, TOR1 and TOR2, as targets of the FKBP12-rapamycin complex in yeast. In this study, we have used a glutathione S-transferase (GST)-FKBP12-rapamycin affinity matrix to isolate putative mammalian targets of rapamycin (mTOR) from tissue extracts. In the presence of rapamycin, immobilized GST-FKBP12 specifically precipitates similar high molecular mass proteins from both rat brain and murine T-lymphoma cell extracts. Binding experiments performed with rapamycin-sensitive and -resistant mutant clones derived from the YAC-1 T-lymphoma cell line demonstrate that the GST-FKBP12-rapamycin complex recovers significantly lower amounts of the candidate mTOR from rapamycin-resistant cell lines. The latter results suggest that mTOR is a relevant target of rapamycin in these cells. Finally, we report the isolation of a full-length mTOR cDNA that encodes a direct ligand for the FKBP12-rapamycin complex. The deduced amino acid sequence of mTOR displays 42 and 45% identity to those of yeast TOR1 and TOR2, respectively. These results strongly suggest that the FKBP12-rapamycin complex interacts with homologous ligands in yeast and mammalian cells and that the loss of mTOR function is directly related to the inhibitory effect of rapamycin on G1- to S-phase progression in T-lymphocytes and other sensitive cell types.

Pubmed ID: 7822316


  • Sabers CJ
  • Martin MM
  • Brunn GJ
  • Williams JM
  • Dumont FJ
  • Wiederrecht G
  • Abraham RT


The Journal of biological chemistry

Publication Data

January 13, 1995

Associated Grants

  • Agency: NIGMS NIH HHS, Id: GM47286

Mesh Terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • Carrier Proteins
  • Cell Cycle Proteins
  • DNA Primers
  • DNA, Complementary
  • DNA-Binding Proteins
  • Fungal Proteins
  • Heat-Shock Proteins
  • Humans
  • Lymphoma, T-Cell
  • Molecular Sequence Data
  • Phosphatidylinositol 3-Kinases
  • Phosphotransferases (Alcohol Group Acceptor)
  • Polyenes
  • Protein Kinases
  • Rats
  • Rats, Sprague-Dawley
  • Saccharomyces cerevisiae Proteins
  • Sequence Homology, Amino Acid
  • Sirolimus
  • TOR Serine-Threonine Kinases
  • Tacrolimus Binding Proteins
  • Tumor Cells, Cultured