eEF3 promotes late stages of tRNA translocation on the ribosome

Namit Ranjan; Agnieszka A. Pochopien; Colin Chih Chien Wu; Bertrand Beckert; Sandra Blanchet; Rachel Green; Marina V. Rodnina; Daniel N. Wilson

doi:10.1101/2020.07.01.182105

eEF3 promotes late stages of tRNA translocation on the ribosome

Namit Ranjan, Agnieszka A. Pochopien, Colin Chih Chien Wu, Bertrand Beckert, Sandra Blanchet, Rachel Green, Marina V. Rodnina, Daniel N. Wilson

School of Medicine

Research output: Contribution to journal › Article › peer-review

Abstract

In addition to the conserved translation elongation factors eEF1A and eEF2, fungi require a third essential elongation factor, eEF3. While eEF3 has been implicated in tRNA binding and release at the A and E sites, its exact mechanism of action is unclear. Here we show that eEF3 acts at the mRNA–tRNA translocation step by promoting the dissociation of the tRNA from the E site, but independent of aminoacyl-tRNA recruitment to the A site. Depletion of eEF3 in vivo leads to a general slow-down in translation elongation due to accumulation of ribosomes with an occupied A site. Cryo-EM analysis of ex vivo eEF3-ribosome complexes shows that eEF3 facilitates late steps of translocation by favoring nonrotated ribosomal states as well as by opening the L1 stalk to release the E-site tRNA. Additionally, our analysis provides structural insights into novel translation elongation states, enabling presentation of a revised yeast translation elongation cycle.

Original language	English (US)
Journal	Unknown Journal
DOIs	https://doi.org/10.1101/2020.07.01.182105
State	Published - Jul 1 2020

Keywords

ABC ATPase
Cryo-EM
E-site tRNA
EEF3
L1 stalk
Translation elongation
Translocation
Yeast

ASJC Scopus subject areas

Biochemistry, Genetics and Molecular Biology(all)
Agricultural and Biological Sciences(all)
Immunology and Microbiology(all)
Neuroscience(all)
Pharmacology, Toxicology and Pharmaceutics(all)

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title = "eEF3 promotes late stages of tRNA translocation on the ribosome",

abstract = "In addition to the conserved translation elongation factors eEF1A and eEF2, fungi require a third essential elongation factor, eEF3. While eEF3 has been implicated in tRNA binding and release at the A and E sites, its exact mechanism of action is unclear. Here we show that eEF3 acts at the mRNA–tRNA translocation step by promoting the dissociation of the tRNA from the E site, but independent of aminoacyl-tRNA recruitment to the A site. Depletion of eEF3 in vivo leads to a general slow-down in translation elongation due to accumulation of ribosomes with an occupied A site. Cryo-EM analysis of ex vivo eEF3-ribosome complexes shows that eEF3 facilitates late steps of translocation by favoring nonrotated ribosomal states as well as by opening the L1 stalk to release the E-site tRNA. Additionally, our analysis provides structural insights into novel translation elongation states, enabling presentation of a revised yeast translation elongation cycle.",

keywords = "ABC ATPase, Cryo-EM, E-site tRNA, EEF3, L1 stalk, Translation elongation, Translocation, Yeast",

author = "Namit Ranjan and Pochopien, {Agnieszka A.} and Wu, {Colin Chih Chien} and Bertrand Beckert and Sandra Blanchet and Rachel Green and Rodnina, {Marina V.} and Wilson, {Daniel N.}",

note = "Publisher Copyright: The copyright holder for this preprint (which was not certified by peer review) is the author/funder. It is made available under a CC-BY-NC-ND 4.0 International license. Copyright: Copyright 2020 Elsevier B.V., All rights reserved.",

year = "2020",

month = jul,

day = "1",

doi = "10.1101/2020.07.01.182105",

language = "English (US)",

journal = "Advances in Water Resources",

issn = "0309-1708",

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AU - Ranjan, Namit

AU - Pochopien, Agnieszka A.

AU - Wu, Colin Chih Chien

AU - Beckert, Bertrand

AU - Blanchet, Sandra

AU - Green, Rachel

AU - Rodnina, Marina V.

AU - Wilson, Daniel N.

N1 - Publisher Copyright: The copyright holder for this preprint (which was not certified by peer review) is the author/funder. It is made available under a CC-BY-NC-ND 4.0 International license. Copyright: Copyright 2020 Elsevier B.V., All rights reserved.

PY - 2020/7/1

Y1 - 2020/7/1

N2 - In addition to the conserved translation elongation factors eEF1A and eEF2, fungi require a third essential elongation factor, eEF3. While eEF3 has been implicated in tRNA binding and release at the A and E sites, its exact mechanism of action is unclear. Here we show that eEF3 acts at the mRNA–tRNA translocation step by promoting the dissociation of the tRNA from the E site, but independent of aminoacyl-tRNA recruitment to the A site. Depletion of eEF3 in vivo leads to a general slow-down in translation elongation due to accumulation of ribosomes with an occupied A site. Cryo-EM analysis of ex vivo eEF3-ribosome complexes shows that eEF3 facilitates late steps of translocation by favoring nonrotated ribosomal states as well as by opening the L1 stalk to release the E-site tRNA. Additionally, our analysis provides structural insights into novel translation elongation states, enabling presentation of a revised yeast translation elongation cycle.

AB - In addition to the conserved translation elongation factors eEF1A and eEF2, fungi require a third essential elongation factor, eEF3. While eEF3 has been implicated in tRNA binding and release at the A and E sites, its exact mechanism of action is unclear. Here we show that eEF3 acts at the mRNA–tRNA translocation step by promoting the dissociation of the tRNA from the E site, but independent of aminoacyl-tRNA recruitment to the A site. Depletion of eEF3 in vivo leads to a general slow-down in translation elongation due to accumulation of ribosomes with an occupied A site. Cryo-EM analysis of ex vivo eEF3-ribosome complexes shows that eEF3 facilitates late steps of translocation by favoring nonrotated ribosomal states as well as by opening the L1 stalk to release the E-site tRNA. Additionally, our analysis provides structural insights into novel translation elongation states, enabling presentation of a revised yeast translation elongation cycle.

KW - ABC ATPase

KW - Cryo-EM

KW - E-site tRNA

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KW - L1 stalk

KW - Translation elongation

KW - Translocation

KW - Yeast

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