Eukaryotic translation initiation factor 3 plays distinct roles at the mRNA entry and exit channels of the ribosomal preinitiation complex

  1. Colin Echeverría Aitken
  2. Petra Beznosková
  3. Vladislava Vlčkova
  4. Wen-Ling Chiu
  5. Fujun Zhou
  6. Leoš Shivaya Valášek  Is a corresponding author
  7. Alan G Hinnebusch  Is a corresponding author
  8. Jon R Lorsch  Is a corresponding author
  1. National Institutes of Health, United States
  2. Institute of Microbiology ASCR, Czech Republic
  3. PharmaEssentia Corporation, Taiwan

Abstract

Eukaryotic translation initiation factor 3 (eIF3) is a central player in recruitment of the pre-initiation complex (PIC) to mRNA. We probed the effects on mRNA recruitment of a library of S. cerevisiae eIF3 functional variants spanning its 5 essential subunits using an in vitro-reconstituted system. Mutations throughout eIF3 disrupt its interaction with the PIC and diminish its ability to accelerate recruitment to a native yeast mRNA. Alterations to the eIF3a CTD and eIF3b/i/g significantly slow mRNA recruitment, and mutations within eIF3b/i/g destabilize eIF2•GTP•Met-tRNAi binding to the PIC. Using model mRNAs lacking contacts with the 40S entry or exit channels, we uncover a critical role for eIF3 requiring the eIF3a NTD, in stabilizing mRNA interactions at the exit channel, and an ancillary role at the entry channel requiring residues of the eIF3a CTD. These functions are redundant: defects at each channel can be rescued by filling the other channel with mRNA.

Article and author information

Author details

  1. Colin Echeverría Aitken

    Laboratory on the Mechanism and Regulation of Protein Synthesis, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, United States
    Competing interests
    No competing interests declared.
  2. Petra Beznosková

    Laboratory of Regulation of Gene Expression, Institute of Microbiology ASCR, Prague, Czech Republic
    Competing interests
    No competing interests declared.
  3. Vladislava Vlčkova

    Laboratory of Regulation of Gene Expression, Institute of Microbiology ASCR, Prague, Czech Republic
    Competing interests
    No competing interests declared.
  4. Wen-Ling Chiu

    PharmaEssentia Corporation, Taipei, Taiwan
    Competing interests
    No competing interests declared.
  5. Fujun Zhou

    Laboratory on the Mechanism and Regulation of Protein Synthesis, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, United States
    Competing interests
    No competing interests declared.
  6. Leoš Shivaya Valášek

    Laboratory of Regulation of Gene Expression, Institute of Microbiology ASCR, Prague, Czech Republic
    For correspondence
    valasekl@biomed.cas.cz
    Competing interests
    No competing interests declared.
  7. Alan G Hinnebusch

    Laboratory of Gene Regulation and Development, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, United States
    For correspondence
    ahinnebusch@nih.gov
    Competing interests
    Alan G Hinnebusch, Reviewing editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1627-8395
  8. Jon R Lorsch

    Laboratory on the Mechanism and Regulation of Protein Synthesis, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, United States
    For correspondence
    jon.lorsch@nih.gov
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4521-4999

Funding

National Institutes of Health (Intramural Research Program)

  • Colin Echeverría Aitken
  • Wen-Ling Chiu
  • Fujun Zhou
  • Alan G Hinnebusch
  • Jon R Lorsch

Wellcome (090812/B/09/Z)

  • Colin Echeverría Aitken
  • Leoš Shivaya Valášek

Centrum of Excellence of the Czech Science Foundation (P305/12/G034)

  • Colin Echeverría Aitken
  • Leoš Shivaya Valášek

Leukemia and Lymphoma Society (5199-12)

  • Colin Echeverría Aitken

National Institutes of Health (GM62128)

  • Jon R Lorsch

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Nahum Sonenberg, McGill University, Canada

Publication history

  1. Received: August 25, 2016
  2. Accepted: October 25, 2016
  3. Accepted Manuscript published: October 26, 2016 (version 1)
  4. Accepted Manuscript updated: October 31, 2016 (version 2)
  5. Version of Record published: December 12, 2016 (version 3)

Copyright

This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication.

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  1. Colin Echeverría Aitken
  2. Petra Beznosková
  3. Vladislava Vlčkova
  4. Wen-Ling Chiu
  5. Fujun Zhou
  6. Leoš Shivaya Valášek
  7. Alan G Hinnebusch
  8. Jon R Lorsch
(2016)
Eukaryotic translation initiation factor 3 plays distinct roles at the mRNA entry and exit channels of the ribosomal preinitiation complex
eLife 5:e20934.
https://doi.org/10.7554/eLife.20934

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