sirolimus and 7-methylguanosine-triphosphate

In response to nutrients, energy sufficiency, hormones, and mitogenic agents, S6K1 phosphorylates several targets linked to translation. However, the molecular mechanisms whereby S6K1 is activated, encounters substrate, and contributes to translation initiation are poorly understood. We show that mTOR and S6K1 maneuver on and off the eukaryotic initiation factor 3 (eIF3) translation initiation complex in a signal-dependent, choreographed fashion. When inactive, S6K1 associates with the eIF3 complex, while the S6K1 activator mTOR/raptor does not. Cell stimulation promotes mTOR/raptor binding to the eIF3 complex and phosphorylation of S6K1 at its hydrophobic motif. Phosphorylation results in S6K1 dissociation, activation, and subsequent phosphorylation of its translational targets, including eIF4B, which is then recruited into the complex in a phosphorylation-dependent manner. Thus, the eIF3 preinitiation complex acts as a scaffold to coordinate a dynamic sequence of events in response to stimuli that promote efficient protein synthesis.

Topics: Adaptor Proteins, Signal Transducing; Amino Acids; Cell Line; Cell Line, Tumor; Eukaryotic Initiation Factor-3; Eukaryotic Initiation Factors; Gene Expression Regulation; HeLa Cells; Hormones; Humans; Insulin; Kinetics; Mitogens; Models, Biological; Multiprotein Complexes; Mutation; Peptide Chain Initiation, Translational; Phorbol Esters; Phosphorylation; Protein Binding; Protein Kinases; Protein Subunits; Proteins; Regulatory-Associated Protein of mTOR; Ribosomal Protein S6 Kinases; Ribosomal Protein S6 Kinases, 90-kDa; RNA Cap Analogs; Sirolimus; TOR Serine-Threonine Kinases

The cap-binding protein eIF4E-binding protein 3 (4E-BP3) was identified some years ago, but its properties have not been investigated in detail. In this report, we investigated the regulation and localisation of 4E-BP3. We show that 4E-BP3 is present in the nucleus as well as in the cytoplasm in primary T cells, HEK293 cells and HeLa cells. 4E-BP3 was associated with eIF4E in both cell compartments. Furthermore, 4E-BP3/eIF4E association in the cytoplasm was regulated by serum or interleukin-2 starvation in the different cell types. Rapamycin did not affect the association of eIF4E with 4E-BP3 in the cytoplasm or in the nucleus.

Topics: Blotting, Western; Carrier Proteins; Cell Line; Cell Nucleus; Culture Media, Serum-Free; Cytoplasm; Electrophoresis, Polyacrylamide Gel; Eukaryotic Initiation Factor-4E; Gene Expression Regulation; HeLa Cells; Humans; Immunosuppressive Agents; Interleukin-2; Peptides; Protein Binding; RNA Cap Analogs; Sirolimus; T-Lymphocytes; Transfection