Insulin-dependent stimulation of protein synthesis by phosphorylation of a regulator of 5'-cap function

Pause, Arnim; Belsham, Graham J.; Gingras, Anne‐Claude; Donzé, Olivier; Lin, Teng Nan; Lawrence, John C.; Sonenberg, Nahum

doi:10.1038/371762a0

Cited by 1,168 publications

(1,147 citation statements)

References 29 publications

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“…Interestingly, DDX3 stimulates IRES-mediated translation in vivo (Figures 2 and 6). Such in vivo stimulation of IRESmediated translation has been observed with 4EBP3 (Poulin et al, 1998) but not with 4EBP1 or 4EBP2 (Pause et al, 1994;Ohlmann et al, 1996). Furthermore, the eIF4E-binding defective DDX3 mutants, but not the helicase-or ATPase-defective mutants, lost their capacity to stimulate HCV IRES-mediated translation ( Figures 2c and 6b); this suggests that the DDX3-eIF4E interaction is a priming event for the stimulation of IRES-mediated translation.…”

Section: Ddx3 Interacts With Eif4e and Regulates Translation J-w Shihmentioning

confidence: 87%

Candidate tumor suppressor DDX3 RNA helicase specifically represses cap-dependent translation by acting as an eIF4E inhibitory protein

Shih¹,

Tsai²,

Chao³

et al. 2007

Oncogene

161

199

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DDX3 is a human RNA helicase with plethoric functions. Our previous studies have indicated that DDX3 is a transcriptional regulator and functions as a tumor suppressor. In this study, we use a bicistronic reporter to demonstrate that DDX3 specifically represses cap-dependent translation but enhances hepatitis C virus internal ribosome entry site-mediated translation in vivo in a helicase activity-independent manner. To elucidate how DDX3 modulates translation, we identified translation initiation factor eukaryotic initiation factor 4E (eIF4E) as a DDX3-binding partner. Interestingly, DDX3 utilizes a consensus eIF4E-binding sequence YIPPHLR to interact with the functionally important dorsal surface of eIF4E in a similar manner to other eIF4E-binding proteins. Furthermore, cap affinity chromatography analysis suggests that DDX3 traps eIF4E in a translationally inactive complex by blocking interaction with eIF4G. Point mutations within the consensus eIF4E-binding motif in DDX3 impair its ability to bind eIF4E and result in a loss of DDX3's regulatory effects on translation. All these features together indicate that DDX3 is a new member of the eIF4E inhibitory proteins involved in translation initiation regulation. Most importantly, this DDX3-mediated translation regulation also confers the tumor suppressor function on DDX3. Altogether, this study demonstrates regulatory roles and action mechanisms for DDX3 in translation, cell growth and likely viral replication.

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Section: Ddx3 Interacts With Eif4e and Regulates Translation J-w Shihmentioning

confidence: 87%

Candidate tumor suppressor DDX3 RNA helicase specifically represses cap-dependent translation by acting as an eIF4E inhibitory protein

Shih¹,

Tsai²,

Chao³

et al. 2007

Oncogene

161

199

View full text Add to dashboard Cite

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“…To exclude possible involvement of TORC1 in Akt phosphorylation in starfish oocytes, we used a TORC1 inhibitor, rapamycin. As a positive control, rapamycin inhibited the mobility shift of eIF4E-binding protein (4E-BP) (Supplementary Figure S3), a well-known target of TORC1 Pause et al, 1994;Brunn et al, 1997;Ma and Blenis, 2009). However, Akt phosphorylation upon 1-MeAde stimulus was not affected (Supplementary Figure S3), supporting the involvement of TORC2, but not TORC1, in Akt phosphorylation in starfish oocytes.…”

Section: Detection Of Akt Phosphorylation In Starfish Oocytesmentioning

confidence: 99%

Turn motif phosphorylation negatively regulates activation loop phosphorylation in Akt

2011

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Akt, also known as protein kinase B, has a central role in various signaling pathways that regulate cellular processes such as metabolism, proliferation and survival. On stimulation, phosphorylation of the activation loop (Aloop) and hydrophobic motif (HM) of Akt by the kinase phosphoinositide-dependent kinase 1 (PDK1) and the mammalian target of rapamycin complex 2 (mTORC2), respectively, results in Akt activation. A well-conserved threonine in the turn motif (TM) is also constitutively phosphorylated by mTORC2 and contributes to the stability of Akt. The role of TM phosphorylation in HM and A-loop phosphorylation has not been sufficiently evaluated. Using starfish oocytes as a model system, this study provides the first evidence that TM phosphorylation has a negative role in A-loop phosphorylation. In this system, the maturation-inducing hormone, 1-methyladenine, stimulates Akt to reinitiate meiosis through activation of cyclin B-Cdc2. The phosphorylation status of Akt was monitored via introduction of exogenous human Akt (hAkt) in starfish oocytes. TM and HM phosphorylation was inhibited by microinjection of an anti-starfish TOR antibody, but not by rapamycin treatment, suggesting that both phosphorylation events depend on TORC2, as reported in mammalian cells. A single or double alanine substitution at each of three phosphorylation residues revealed that TM phosphorylation renders Akt susceptible to dephosphorylation on the A-loop. When A-loop phosphatase was inhibited by okadaic acid (OA), TM phosphorylation still reduced Aloop phosphorylation, suggesting that the effect is caused at least partially through reduction of sensitivity to PDK1. Negative regulation by TM phosphorylation was also observed in constitutively active Akt and was functionally reflected in meiosis resumption. By contrast, HM phosphorylation enhanced A-loop phosphorylation and achieved full activation of Akt via a mechanism at least partially independent of TM phosphorylation. These observations provide new insight into the mechanism controlling Akt phosphorylation in the cell.

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“…However, under conditions of cellular stress when cap-dependent translation is inhibited, such as nutrient deprivation and hypoxia, certain mRNAs can still be translated. Such conditions favour cap-independent translation of mRNAs containing an internal ribosome entry site (IRES) element within their 5 untranslated region [30]. p27 is a fundamental regulator of proliferation in most cell types that acts by blocking G1/S transition through inhibition of cyclin-dependent kinase 2 (CDK2) [31].…”

Section: Ins-1 Cells Inducibly Expressing Dn-hnf1a But Not Wt-mentioning

confidence: 99%

Early loss of mammalian target of rapamycin complex 1 (mTORC1) signalling and reduction in cell size during dominant-negative suppression of hepatic nuclear factor 1-α (HNF1A) function in INS-1 insulinoma cells

et al. 2008

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Aims/hypothesis Mutations in the HNF1A (previously known as TCF1) gene encoding hepatocyte nuclear factor-1α (HNF1A) lead to the development of maturity-onset diabetes of the young, type 3 (HNF1A-MODY), characterised by impaired insulin secretion and a reduction in beta cell mass. HNF1A plays an important role in pancreatic beta cell differentiation and survival. The mammalian target of rapamycin (mTOR) is a central growth factor-and nutrientactivated protein kinase controlling cell metabolism, growth and survival. We investigated the role of mTOR inactivation in the decline in beta cell mass in a cellular model of HNF1A-MODY. Methods Previously we showed that suppression of HNF1A function via expression of a dominant-negative mutant (DN-HNF1A) decreases insulin gene transcription in insulinoma (INS-1) cells. We investigated the signalling of two distinct mTOR protein complexes, mTORC1 and mTORC2, in response to DN-HNF1A induction.Results We observed delayed inactivation of mTORC2 48 h after DN-HNF1A induction, evidenced by a reduction in serine 473 phosphorylation of thymoma viral protooncogene 1 (AKT1). We also observed an early inactivation of mTORC1 24 h after DN-HNF1A induction, which was detected by decreases in threonine 389 phosphorylation of p70 ribosomal protein S6 kinase (S6K1) and serine 65 phosphorylation of translational inhibitor eukaryotic translation initiation factor 4E binding protein 1 (4E-BP1). Flow cytometry and gene expression analysis demonstrated a preapoptotic decrease in INS-1 cell size in response to DN-HNF1A induction, and an increase in the level of the mTORC1-regulated cell-cycle inhibitor, cyclin-dependent kinase inhibitor 1B p27. Conclusions/interpretation Our data suggest that mTOR kinase and signalling through mTORC1 are highly sensitive to suppression of HNF1A function, and may contribute to disturbance of cell-size regulation and cell-cycle progression in HNF1A-MODY.

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Insulin-dependent stimulation of protein synthesis by phosphorylation of a regulator of 5'-cap function

Cited by 1,168 publications

References 29 publications

Candidate tumor suppressor DDX3 RNA helicase specifically represses cap-dependent translation by acting as an eIF4E inhibitory protein

Candidate tumor suppressor DDX3 RNA helicase specifically represses cap-dependent translation by acting as an eIF4E inhibitory protein

Turn motif phosphorylation negatively regulates activation loop phosphorylation in Akt

Early loss of mammalian target of rapamycin complex 1 (mTORC1) signalling and reduction in cell size during dominant-negative suppression of hepatic nuclear factor 1-α (HNF1A) function in INS-1 insulinoma cells

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