The C terminus of initiation factor 4E-binding protein 1 contains multiple regulatory features that influence its function and phosphorylation.
Eukaryotic initiation factor 4E (eIF4E) binds the mRNA cap structure and forms eIF4F complexes that recruit 40S subunits to the mRNA. Formation of eIF4F is blocked by eIF4E-binding proteins such as 4E-BP1, which interacts with eIF4E via a motif in the center of its 118-residue sequence. 4E-BP1 plays key roles in cell proliferation, growth, and survival. Binding of 4E-BP1 to eIF4E is regulated by hierarchical multisite phosphorylation. Here we demonstrate that three different features in the C terminus of 4E-BP1 play distinct roles in regulating its phosphorylation and function. Firstly, we identify a new phosphorylation site in its C terminus (S101). A serine or glutamate at this position is required for efficient phosphorylation at Ser65. A second C-terminal site, S112, directly affects binding of 4E-BP1 to eIF4E without influencing phosphorylation of other sites. Thirdly, a conserved C-terminal motif influences phosphorylation of multiple residues, including rapamycin-insensitive sites. These relatively long-range effects are surprising given the reportedly unstructured nature of 4E-BP1 and may imply that phosphorylation of 4E-BP1 and/or binding to eIF4E induces a more-ordered structure. 4E-BP2 and -3 lack phosphorylatable residues corresponding to both S101 and S112. However, in 4E-BP3, replacement of the alanine at the position corresponding to S112 by serine or glutamate did not confer the ability to be released from eIF4E in response to insulin.
Wang
Xuemin
X
Division of Molecular Physiology, Faculty of Life Sciences, University of Dundee, Dundee DD1 5EH, United Kingdom.
Li
Wei
W
Parra
Josep-Lluis
JL
Beugnet
Anne
A
Proud
Christopher G
CG
D048868:Adaptor Proteins, Signal Transducing; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001665:Binding Sites; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism; D018797:Cell Cycle Proteins; D002455:Cell Division; D002460:Cell Line; D002470:Cell Survival; D004791:Enzyme Inhibitors / Q000494:pharmacology; D039561:Eukaryotic Initiation Factor-4E / Q000378:metabolism; D005822:Genetic Vectors; D018698:Glutamic Acid / Q000737:chemistry; D005973:Glutamine / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D007328:Insulin / Q000494:pharmacology; D008957:Models, Genetic; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D010750:Phosphoproteins / Q000737:chemistry* / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding; D020033:Protein Isoforms; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012694:Serine / Q000737:chemistry / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018797:Cell Cycle Proteins; C087000:EIF4EBP1 protein, human; D004791:Enzyme Inhibitors; D039561:Eukaryotic Initiation Factor-4E; D007328:Insulin; D010455:Peptides; D010750:Phosphoproteins; D020033:Protein Isoforms; D005973:Glutamine; D018698:Glutamic Acid; D012694:Serine