Hierarchical phosphorylation of the translation inhibitor 4E-BP1.

By A. Gingras, B. Raught, Steven P. Gygi, A. Niedźwiecka, M. Miron, Stephen K. Burley, R. Polakiewicz, A. Wysłouch-Cieszyńska, R. Aebersold, Nahum Sonenberg. Genes & Development, 2001

946 citations recorded by the source corpus.

In most instances, translation is regulated at the initiation phase, when a ribosome is recruited to the 5' end of an mRNA. The eIF4E-binding proteins (4E-BPs) interdict translation initiation by binding to the translation factor eIF4E, and preventing recruitment of the translation machinery to mRNA. The 4E-BPs inhibit translation in a reversible manner. Hypophosphorylated 4E-BPs interact avidly with eIF4E, whereas 4E-BP hyperphosphorylation, elicited by stimulation of cells with hormones, cytokines, or growth factors, results in an abrogation of eIF4E-binding activity. We reported previously that phosphorylation of 4E-BP1 on Thr 37 and Thr 46 is relatively insensitive to serum deprivation and rapamycin treatment, and that phosphorylation of these residues is required for the subsequent phosphorylation of a set of unidentified serum-responsive sites. Here, using mass spectrometry, we identify the serum-responsive, rapamycin-sensitive sites as Ser 65 and Thr 70. Utilizing a novel combination of two-dimensional isoelectric focusing/SDS-PAGE and Western blotting with phosphospecific antibodies, we also establish the order of 4E-BP1 phosphorylation in vivo; phosphorylation of Thr 37/Thr 46 is followed by Thr 70 phosphorylation, and Ser 65 is phosphorylated last. Finally, we show that phosphorylation of Ser 65 and Thr 70 alone is insufficient to block binding to eIF4E, indicating that a combination of phosphorylation events is necessary to dissociate 4E-BP1 from eIF4E.

Subfields: Molecular Biology, Radiology, Nuclear Medicine and Imaging

View Paper: https://www.semanticscholar.org/paper/8e09d8c64bd5909a3d25bbd1b3db9308816deabf

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Citations in the corpus

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Targeting mTOR for cancer therapy (2019, background): “mTORC1 phosphorylates S6K1 at T389 and 4E-BP1 at multiple residues [64].”
Active-Site Inhibitors of mTOR Target Rapamycin-Resistant Outputs of mTORC1 and mTORC2 (2009, background): “Additional Analysis of 4EBP1 Phosphorylation (A) 4EBP1 phosphorylation at T70 is not inhibited by either PP242 or rapamycin.”
Upstream and downstream of mTOR. (2004, background): “We shall not discuss the literature here, as several recent reviews deal critically with this issue (Gingras et al. 2001b; Harris and Lawrence 2003).”
PRAS40 is an insulin-regulated inhibitor of the mTORC1 protein kinase. (2007, background): “The best-characterized downstream effectors of mTORC1 are S6 Kinase 1 (S6K1) and 4E-BP1, two translational regulators that mTORC1 directly phosphorylates (Brunn et al., 1997; Burnett et al., 1998; Gingras et al., 2001).”
Regulation of the mTOR complex 1 pathway by nutrients, growth factors, and stress (2010, background): “Interestingly, for unknown reasons, rapamycin-FKBP12 inhibits mTORC1 activity only toward a subset of mTORC1 substrates (Gingras et al., 2001; Jiang et al., 2001; Kudchodkar et al., 2004; Thoreen et al., 2009).”
DEPTOR is an mTOR Inhibitor Whose Frequent Overexpression in Multiple Myeloma Cells Promotes their Survival (2009, background): “Interestingly, many of the phosphorylation sites fall within ‘‘proline-directed’’ motifs like those in 4E-BP1 (Gingras et al., 2001), which are known to be phosphorylated by mTOR (Brunn et al., 1997; Burnett et al., 1998; Gingras et al., 1999).”
The TSC1-TSC2 complex: a molecular switchboard controlling cell growth. (2008, background): “Four separate proline-directed sites (/Thr-Pro) on 4E-BP1 (Thr37, Thr47, Ser65 and Thr70) can be phosphorylated by mTORC1 and are sensitive to rapamycin treatment [39,40].”
mTORC1 controls mitochondrial activity and biogenesis through 4E-BP-dependent translational regulation. (2013, background): “Phosphorylation of 4E-BPs by mTORC1 leads to their dissociation from eIF4E, thereby allowing association of eIF4E with eIF4G and the assembly of the eIF4F translation initiation complex at the mRNA 50 end (Gingras et al., 1999, 2001; Pause et al., 1994).”
A Novel Function of eIF2α Kinases as Inducers of the Phosphoinositide-3 Kinase Signaling Pathway (2007, background): “PKR to induce the phosphorylation of 4E-BP1 mediated through PI3K and the downstream effector FRAP/mTOR.”
Raptor, a binding partner of target of rapamycin (TOR), mediates TOR action. (2002, background): “The similarity of the phenotypes observed for RNAi against the(Gingras et al., 2001; Mothe-Satney et al., 2000).”