Translational initiation factor eIF5 replaces eIF1 on the 40S ribosomal subunit to promote start-codon recognition
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
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Structure of a partial yeast 48S preinitiation complex with eIF5 N-terminal domain (Map A)Electron Microscopy Data Bank, 4330.
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Structure of a partial yeast 48S preinitiation complex with eIF5 N-terminal domain (Sample Map 1)Electron Microscopy Data Bank, 4328.
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Structure of a partial yeast 48S preinitiation complex with eIF5 N-terminal domain (Map B)Electron Microscopy Data Bank, 4331.
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Structure of a partial yeast 48S preinitiation complex with eIF5 N-terminal domain (Map C1)Electron Microscopy Data Bank, 4327.
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Structure of a partial yeast 48S preinitiation complex with eIF5 N-terminal domain (Map C2)Electron Microscopy Data Bank, 4329.
Article and author information
Author details
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
- Rachel Green, Johns Hopkins School of Medicine, United States
Publication history
- Received: June 26, 2018
- Accepted: November 21, 2018
- Accepted Manuscript published: November 26, 2018 (version 1)
- Accepted Manuscript updated: November 29, 2018 (version 2)
- Accepted Manuscript updated: November 30, 2018 (version 3)
- Version of Record published: December 18, 2018 (version 4)
- 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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