title: Target of rapamycin (TOR)-signaling and RAIP motifs play distinct roles in the mammalian TOR-dependent phosphorylation of initiation factor 4E-binding protein 1.
abstract: The translational repressor protein eIF4E-binding protein 1 (4E-BP1, also termed PHAS-I) is regulated by phosphorylation through the rapamycin-sensitive mTOR (mammalian target of rapamycin) pathway. Recent studies have identified two regulatory motifs in 4E-BP1, an mTOR-signaling (TOS) motif in the C terminus of 4E-BP1 and an RAIP motif (named after its sequence) in the N terminus. Other recent work has shown that the protein raptor binds to mTOR and 4E-BP1. We show that raptor binds to full-length 4E-BP1 or a C-terminal fragment containing the TOS motif but not to an N-terminal fragment containing the RAIP motif. Mutation of several residues within the TOS motif abrogates binding to raptor, indicating that the TOS motif is required for this interaction. 4E-BP1 undergoes phosphorylation at multiple sites in intact cells. The effects of removal or mutation of the RAIP and TOS motifs differ. The RAIP motif is absolutely required for phosphorylation of sites in the N and C termini of 4E-BP1, whereas the TOS motif primarily affects phosphorylation of Ser-64/65, Thr-69/70, and also the rapamycin-insensitive site Ser-101. Phosphorylation of N-terminal sites that are dependent upon the RAIP motif is sensitive to rapamycin. The RAIP motif thus promotes the mTOR-dependent phosphorylation of multiple sites in 4E-BP1 independently of the 4E-BP1/raptor interaction.
authors.affiliation: Division of Molecular Physiology, Faculty of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
authors.forename: Anne
authors.initials: A
authors.lastname: Beugnet
authors.forename: Xuemin
authors.initials: X
authors.lastname: Wang
authors.forename: Christopher G
authors.initials: CG
authors.lastname: Proud
mesh_terms: D048868:Adaptor Proteins, Signal Transducing; D020816:Amino Acid Motifs; D000818:Animals; D001665:Binding Sites; D015153:Blotting, Western; D002352:Carrier Proteins / Q000378:metabolism*; D018797:Cell Cycle Proteins; D002460:Cell Line; D018076:DNA, Complementary / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D009154:Mutation; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011485:Protein Binding; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D051381:Rats; D012694:Serine / Q000737:chemistry; D015398:Signal Transduction; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases; D014162:Transfection
chemical_list: D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D018076:DNA, Complementary; C087000:EIF4EBP1 protein, human; C499526:Eif4ebp1 protein, rat; D047908:Intracellular Signaling Peptides and Proteins; D010750:Phosphoproteins; D012694:Serine; D011494:Protein Kinases; C546842:MTOR protein, human; C546845:mTOR protein, rat; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus