A point mutation in the nucleotide exchange factor eIF2B constitutively activates the integrated stress response by allosteric modulation

  1. Morgane Boone
  2. Lan Wang
  3. Rosalie Lawrence
  4. Adam Frost
  5. Peter Walter  Is a corresponding author
  6. Michael Schoof  Is a corresponding author
  1. Howard Hughes Medical Institute, University of California, San Francisco, United States
  2. University of California, San Francisco (Adjunct), United States

Abstract

In eukaryotic cells, stressors reprogram the cellular proteome by activating the integrated stress response (ISR). In its canonical form, stress-sensing kinases phosphorylate the eukaryotic translation initiation factor eIF2 (eIF2-P), which ultimately leads to reduced levels of ternary complex required for initiation of mRNA translation. Previously we showed that translational control is primarily exerted through a conformational switch in eIF2's nucleotide exchange factor, eIF2B, which shifts from its active A-State conformation to its inhibited I-State conformation upon eIF2-P binding, resulting in reduced nucleotide exchange on eIF2 (Schoof et al. 2021). Here, we show functionally and structurally how a single histidine to aspartate point mutation in eIF2B's β subunit (H160D) mimics the effects of eIF2-P binding by promoting an I-State like conformation, resulting in eIF2-P independent activation of the ISR. These findings corroborate our previously proposed A/I-State model of allosteric ISR regulation.

Data availability

All data generated or anaysed during this study are included in the manuscript and source data files. The final structural model has been deposited in PDB under the accession code 7TRJ. Amplicon sequencing data for the CRISPR clones has been deposited in NCBI's Sequence Read Archive (SRA) under accession number PRJNA821864.

The following data sets were generated

Article and author information

Author details

  1. Morgane Boone

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7807-5542
  2. Lan Wang

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8931-7201
  3. Rosalie Lawrence

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    Competing interests
    No competing interests declared.
  4. Adam Frost

    Department of Biochemistry and Biophysics, University of California, San Francisco (Adjunct), San Francisco, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2231-2577
  5. Peter Walter

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    For correspondence
    peter@walterlab.ucsf.edu
    Competing interests
    Peter Walter, is an inventor on U.S. Patent 9708247 held by the Regents of the University of California that describes ISRIB and its analogs. Rights to the invention have been licensed by UCSF to Calico. For the remaining authors, no competing financial interests exist..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6849-708X
  6. Michael Schoof

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    For correspondence
    michael@walterlab.ucsf.edu
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3531-5232

Funding

Calico Life Sciences LLC

  • Peter Walter

The George and Judy Marcus Family Foundation

  • Peter Walter

Belgian-American Educational Foundation

  • Morgane Boone

Damon-Runyon Cancer Research Foundation

  • Lan Wang

Jan Coffin Child Foundation

  • Rosalie Lawrence

Chan Zuckerberg Biohub Investigator Award

  • Adam Frost

HHMI Faculty Scholar grant

  • Adam Frost

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: December 8, 2021
  2. Accepted: April 11, 2022
  3. Accepted Manuscript published: April 13, 2022 (version 1)

Copyright

© 2022, Boone et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 574
    Page views
  • 123
    Downloads
  • 0
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Morgane Boone
  2. Lan Wang
  3. Rosalie Lawrence
  4. Adam Frost
  5. Peter Walter
  6. Michael Schoof
(2022)
A point mutation in the nucleotide exchange factor eIF2B constitutively activates the integrated stress response by allosteric modulation
eLife 11:e76171.
https://doi.org/10.7554/eLife.76171

Further reading

    1. Biochemistry and Chemical Biology
    Tiantian Wei et al.
    Research Article Updated

    The dual-specificity tyrosine phosphorylation-regulated kinase DYRK2 has emerged as a critical regulator of cellular processes. We took a chemical biology approach to gain further insights into its function. We developed C17, a potent small-molecule DYRK2 inhibitor, through multiple rounds of structure-based optimization guided by several co-crystallized structures. C17 displayed an effect on DYRK2 at a single-digit nanomolar IC50 and showed outstanding selectivity for the human kinome containing 467 other human kinases. Using C17 as a chemical probe, we further performed quantitative phosphoproteomic assays and identified several novel DYRK2 targets, including eukaryotic translation initiation factor 4E-binding protein 1 (4E-BP1) and stromal interaction molecule 1 (STIM1). DYRK2 phosphorylated 4E-BP1 at multiple sites, and the combined treatment of C17 with AKT and MEK inhibitors showed synergistic 4E-BP1 phosphorylation suppression. The phosphorylation of STIM1 by DYRK2 substantially increased the interaction of STIM1 with the ORAI1 channel, and C17 impeded the store-operated calcium entry process. These studies collectively further expand our understanding of DYRK2 and provide a valuable tool to pinpoint its biological function.

    1. Biochemistry and Chemical Biology
    2. Structural Biology and Molecular Biophysics
    Lukas P Feilen et al.
    Research Article

    Cleavage of membrane proteins in the lipid bilayer by intramembrane proteases is crucial for health and disease. Although different lipid environments can potently modulate their activity, how this is linked to their structural dynamics is unclear. Here we show that the carboxy-peptidase-like activity of the archaeal intramembrane protease PSH, a homolog of the Alzheimer's disease-associated presenilin/γ-secretase is impaired in micelles and promoted in a lipid bilayer. Comparative molecular dynamics simulations revealed that important elements for substrate binding such as transmembrane domain 6a of PSH are more labile in micelles and stabilized in the lipid bilayer. Moreover, consistent with an enhanced interaction of PSH with a transition-state analog inhibitor, the bilayer promoted the formation of the enzyme´s catalytic active site geometry. Our data indicate that the lipid environment of an intramembrane protease plays a critical role in structural stabilization and active site arrangement of the enzyme-substrate complex thereby promoting intramembrane proteolysis.