Sensitivity of global translation to mTOR inhibition in REN cells depends on the equilibrium between eIF4E and 4E-BP1

Our plain-English summary of Sensitivity of global translation to mTOR inhibition in REN cells depends on the equilibrium between eIF4E and 4E-BP1, by S. Grosso, E. Pesce, D. Brina, et al (PloS One, 2011).

The study

• Authors: S. Grosso, E. Pesce, D. Brina, A. Beugnet, F. Loreni, S. Biffo
• Journal: PloS One
• Published: 2011;6(12):e29136
• Cited by: 21 articles, as counted by NIH iCite
• Read it: PubMed · Publisher · Free full text

Abstract

Initiation is the rate-limiting phase of protein synthesis, controlled by signaling pathways regulating the phosphorylation of translation factors. Initiation has three steps, 43S, 48S and 80S formation. 43S formation is repressed by eIF2α phosphorylation. The subsequent steps, 48S and 80S formation are enabled by growth factors. 48S relies on eIF4E-mediated assembly of eIF4F complex; 4E-BPs competitively displace eIF4E from eIF4F. Two pathways control eIF4F: 1) mTORc1 phosphorylates and inactivates 4E-BPs, leading to eIF4F formation; 2) the Ras-Mnk cascade phosphorylates eIF4E. We show that REN and NCI-H28 mesothelioma cells have constitutive activation of both pathways and maximal translation rate, in the absence of exogenous growth factors. Translation is rapidly abrogated by phosphorylation of eIF2α. Surprisingly, pharmacological inhibition of mTORc1 leads to the complete dephosphorylation of downstream targets, without changes in methionine incorporation. In addition, the combined administration of mTORc1 and MAPK/Mnk inhibitors has no additive effect. The inhibition of both mTORc1 and mTORc2 does not affect the metabolic rate. In spite of this, mTORc1 inhibition reduces eIF4F complex formation, and depresses translocation of TOP mRNAs on polysomes. Downregulation of eIF4E and overexpression of 4E-BP1 induce rapamycin sensitivity, suggesting that disruption of eIF4F complex, due to eIF4E modulation, competes with its recycling to ribosomes. These data suggest the existence of a dynamic equilibrium in which eIF4F is not essential for all mRNAs and is not displaced from translated mRNAs, before recycling to the next.

© 2011 Grosso et al.

In plain English

This laboratory study of human cancer cells did not examine asbestos exposure or patients. The authors examined how translation (protein production) in malignant mesothelioma cell lines, mainly REN and NCI-H28, responded to drugs blocking growth signals, including rapamycin and PP242, which inhibit mTOR. In REN cells, insulin did not significantly change protein production. Arsenite, dithiothreitol and thapsigargin blocked it; brief brefeldin A did not. Rapamycin and other signal blockers switched off their targets but only minimally reduced translation. In these short-term REN experiments, combining drugs had no additive effect, and PP242 left translation unaffected. In NCI-H28 cells, PP242 did not significantly reduce translation, though one figure legend reported a reduction at one concentration. Rapamycin reduced assembly of eIF4F, a complex that helps start most, but not all, translation. Rapamycin and PP242 slightly reduced translation of TOP mRNAs. Adding 4E-BP1 or lowering eIF4E made REN cells' translation sensitive to rapamycin. Rapamycin reduced translation in mesMM98 and ME-180 cells, but not REN or NCI-H28. The authors concluded complex inhibition did not match overall translation; they noted short-term treatments may explain the small effects.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: laboratory (in vitro) experiments on cultured human cancer cell lines
• Who was studied: Human malignant mesothelioma cell lines: REN (inflammatory epithelial subtype), NCI-H28 (epithelial subtype) and mesMM98 (from pleural effusion of a sarcomatous mesothelioma), plus ME-180 cancer cells for comparison. No people or animals were studied.
• Exposure: No asbestos exposure was studied. Cells were treated briefly with signaling-inhibitor drugs: rapamycin (mTORc1 inhibitor), PP242 (mTORc1 and mTORc2 inhibitor), U0126 (MEK inhibitor) and an Mnk1 inhibitor. They were also treated with insulin and with stress-inducing chemicals (thapsigargin, dithiothreitol, arsenite, brefeldin A).
• Main results:
  • In serum-starved REN cells, insulin stimulation did not significantly affect methionine incorporation (a measure of protein production).
  • In REN cells, the stress-inducing chemicals arsenite, dithiothreitol and thapsigargin blocked translation.
  • In REN cells, short brefeldin A treatment caused only minor eIF2a phosphorylation, which was not sufficient to block the translation rate.
  • In REN cells, blocking mTOR, ERK1/2 or Mnk1 only minimally reduced translation. Combining drugs had no additive effect.
  • In REN cells, methionine incorporation was unaffected by PP242 treatment.
  • In REN cells, rapamycin increased 4E-BP1 binding to eIF4E and displaced eIF4G, meaning less cap complex was formed. The article notes that most translation is cap-dependent, but some mRNAs do not require eIF4F.
  • In REN cells, rapamycin and PP242 slightly reduced the association of TOP mRNAs with polysomes (the ribosome clusters that translate messages), although TOP mRNA translation remained fairly high.
  • Overexpressing wild-type 4E-BP1 in REN cells made their translation sensitive to rapamycin.
  • Lowering eIF4E in REN cells made their translation sensitive to rapamycin.
  • Rapamycin reduced translation in mesMM98 and ME-180 cells but was ineffective on REN and NCI-H28 cells.
  • In NCI-H28 cells, the main text says PP242 did not reduce translation to a statistically significant level.
  • The Figure 8 legend for NCI-H28 cells reports that mTOR kinase inactivation reduced translation at only one tested concentration (the unit is likely garbled in the text).
• The authors' conclusion: The authors concluded that inhibition of the cap complex does not match global translation, and that global translation is not necessarily reduced by mTOR inhibition in all cell types.
• Limitations the authors state:
  • The authors stated that the small effect of pathway inhibition may reflect their use of short-term treatments.
  • The authors stated that the reduction in TOP mRNA translation may be below the sensitivity of their assay.
  • The authors could not establish a relationship between p53 and rapamycin sensitivity.

Topics

Medical terms in this study, explained in our Lexicon: Mesothelioma, Translocation, Polysome.

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