Translational initiation factor eIF5 replaces eIF1 on the 40S ribosomal subunit to promote start-codon recognition

  1. José Luis Llácer
  2. Tanweer Hussain
  3. Adesh K Saini
  4. Jagpreet Singh Nanda
  5. Sukhvir Kaur
  6. Yuliya Gordiyenko
  7. Rakesh Kumar
  8. Alan G Hinnebusch  Is a corresponding author
  9. Jon R Lorsch  Is a corresponding author
  10. V Ramakrishnan  Is a corresponding author
  1. MRC Laboratory of Molecular Biology, United Kingdom
  2. Indian Institute of Science, India
  3. Shoolini University of Biotechnology and Management Sciences, India
  4. National Institutes of Health, United States

Abstract

In eukaryotic translation initiation AUG recognition of the mRNA requires accommodation of Met-tRNAi in a 'PIN' state, which is antagonized by the factor eIF1. eIF5 is a GTPase activating protein (GAP) of eIF2 that additionally promotes stringent AUG selection, but the molecular basis of its dual function was unknown. We present a cryo-electron microscopy (cryo-EM) reconstruction of a yeast 48S pre-initiation complex (PIC), at an overall resolution of 3.0 Å, featuring the N-terminal domain (NTD) of eIF5 bound to the 40S subunit at the location vacated by eIF1. eIF5 interacts with and allows a more accommodated orientation of Met-tRNAi. Substitutions of eIF5 residues involved in the eIF5-NTD/tRNAi interaction influenced initiation at near-cognate UUG codons in vivo, and the closed/open PIC conformation in vitro, consistent with direct stabilization of the codon:anticodon duplex by the wild-type eIF5-NTD. The present structure reveals the basis for a key role of eIF5 in start-codon selection.

Data availability

Five maps have been deposited in the EMDB with accession codes EMDB: 4328, EMDB: 4330, EMDB: 4331, EMDB: 4327, EMDB: 4329, for the sample 1 map, Map A, Map B, Map C1 and Map C2, respectively. Two atomic coordinate models have been deposited in the PDB with accession codes PDB: 6FYX, PDB: 6FYY, for models showing TC in conformation 1 and conformation 2, respectively.All data generated or analysed during this study are included in the manuscript and supporting files. Source data files have been provided for Tables 4 and 5 and Figures 4 and 5

The following data sets were generated

Article and author information

Author details

  1. José Luis Llácer

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5304-1795
  2. Tanweer Hussain

    Molecular Reproduction, Development and Genetics (MRDG), Indian Institute of Science, Bangalore, India
    Competing interests
    No competing interests declared.
  3. Adesh K Saini

    Shoolini University of Biotechnology and Management Sciences, Solan, India
    Competing interests
    No competing interests declared.
  4. Jagpreet Singh Nanda

    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.
  5. Sukhvir Kaur

    Shoolini University of Biotechnology and Management Sciences, Solan, India
    Competing interests
    No competing interests declared.
  6. Yuliya Gordiyenko

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    Competing interests
    No competing interests declared.
  7. Rakesh Kumar

    Shoolini University of Biotechnology and Management Sciences, Solan, India
    Competing interests
    No competing interests declared.
  8. Alan G Hinnebusch

    Laboratory of Gene Regulation and Development, Eunice K. Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, United States
    For correspondence
    alanh@mail.nih.gov
    Competing interests
    Alan G Hinnebusch, Reviewing editor, eLife.
  9. 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
  10. V Ramakrishnan

    MRC Laboratory of Molecular Biology, Cambridge, United Kingdom
    For correspondence
    ramak@mrc-lmb.cam.ac.uk
    Competing interests
    No competing interests declared.

Funding

Medical Research Council (MC_U105184332)

  • V Ramakrishnan

Wellcome (WT096570)

  • V Ramakrishnan

Agouron Institute

  • V Ramakrishnan

Department of Science and Technology, Ministry of Science and Technology (Int/NZ/P-2/13)

  • Adesh K Saini

National Institutes of Health (GM62128)

  • Jon R Lorsch

Human Frontier Science Program (RGP-0028/2009)

  • Alan G Hinnebusch

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

Reviewing Editor

  1. Rachel Green, Johns Hopkins School of Medicine, United States

Version history

  1. Received: June 26, 2018
  2. Accepted: November 21, 2018
  3. Accepted Manuscript published: November 26, 2018 (version 1)
  4. Accepted Manuscript updated: November 29, 2018 (version 2)
  5. Accepted Manuscript updated: November 30, 2018 (version 3)
  6. Version of Record published: December 18, 2018 (version 4)
  7. Version of Record updated: January 10, 2019 (version 5)

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. José Luis Llácer
  2. Tanweer Hussain
  3. Adesh K Saini
  4. Jagpreet Singh Nanda
  5. Sukhvir Kaur
  6. Yuliya Gordiyenko
  7. Rakesh Kumar
  8. Alan G Hinnebusch
  9. Jon R Lorsch
  10. V Ramakrishnan
(2018)
Translational initiation factor eIF5 replaces eIF1 on the 40S ribosomal subunit to promote start-codon recognition
eLife 7:e39273.
https://doi.org/10.7554/eLife.39273

Share this article

https://doi.org/10.7554/eLife.39273

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