Protomer alignment modulates specificity of RNA substrate recognition by Ire1

  1. Weihan Li
  2. Kelly Crotty
  3. Diego Garrido Ruiz
  4. Mark Voorhies
  5. Carlos Rivera
  6. Anita Sil
  7. R Dyche Mullins
  8. Matthew P Jacobson
  9. Jirka Peschek  Is a corresponding author
  10. Peter Walter  Is a corresponding author
  1. University of California, San Francisco, United States
  2. Yale School of Medicine, United States
  3. Howard Hughes Medical Institute, University of California, San Francisco, United States

Abstract

The unfolded protein response (UPR) maintains protein folding homeostasis in the endoplasmic reticulum (ER). In metazoan cells, the Ire1 branch of the UPR initiates two functional outputs—non-conventional mRNA splicing and selective mRNA decay (RIDD). By contrast, Ire1 orthologs from Saccharomyces cerevisiae and Schizosaccharomyces pombe are specialized for only splicing or RIDD, respectively. Previously, we showed that the functional specialization lies in Ire1's RNase activity, which is either stringently splice-site specific or promiscuous (W. Li et al., 2018). Here, we developed an assay that reports on Ire1's RNase promiscuity. We found that conversion of two amino acids within the RNase domain of S. cerevisiae Ire1 to their S. pombe counterparts rendered it promiscuous. Using biochemical assays and computational modeling, we show that the mutations rewired a pair of salt bridges at Ire1 RNase domain's dimer interface, changing its protomer alignment. Thus, Ire1 protomer alignment affects its substrates specificity.

Data availability

All data generated or analysed during this study are included in the manuscript and supporting files.

Article and author information

Author details

  1. Weihan Li

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4718-1884
  2. Kelly Crotty

    Department of Biochemistry and Biophysics, Howard Hughes Medical Institute, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  3. Diego Garrido Ruiz

    Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2441-385X
  4. Mark Voorhies

    Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8815-7384
  5. Carlos Rivera

    Department of Molecular Biophysics and Biochemistry, Yale School of Medicine, New Haven, United States
    Competing interests
    The authors declare that no competing interests exist.
  6. Anita Sil

    Microbiology & Immunology, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  7. R Dyche Mullins

    Microbiology & Immunology, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  8. Matthew P Jacobson

    Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, United States
    Competing interests
    The authors declare that no competing interests exist.
  9. Jirka Peschek

    Biochemistry and Biophysics, University of California, San Francisco, San Francisco, United States
    For correspondence
    jirka@walterlab.ucsf.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8158-9301
  10. 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
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6849-708X

Funding

Howard Hughes Medical Institute

  • R Dyche Mullins
  • Peter Walter

Deutsche Forschungsgemeinschaft (Emmy Noether fellow)

  • Jirka Peschek

UCSF-Zaffaroni Fellowship

  • Weihan Li

Human Frontier Science Program

  • Jirka Peschek

National Institute of Allergy and Infectious Diseases (2R37 AI066224)

  • Anita Sil

National Institute of General Medical Sciences (R01 GM032384)

  • Peter Walter

National Institute of General Medical Sciences (1R35 GM118119-01)

  • R Dyche Mullins

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

Version history

  1. Received: February 12, 2021
  2. Accepted: April 27, 2021
  3. Accepted Manuscript published: April 27, 2021 (version 1)
  4. Accepted Manuscript updated: April 29, 2021 (version 2)
  5. Version of Record published: May 7, 2021 (version 3)

Copyright

© 2021, Li 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.

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  1. Weihan Li
  2. Kelly Crotty
  3. Diego Garrido Ruiz
  4. Mark Voorhies
  5. Carlos Rivera
  6. Anita Sil
  7. R Dyche Mullins
  8. Matthew P Jacobson
  9. Jirka Peschek
  10. Peter Walter
(2021)
Protomer alignment modulates specificity of RNA substrate recognition by Ire1
eLife 10:e67425.
https://doi.org/10.7554/eLife.67425

Share this article

https://doi.org/10.7554/eLife.67425

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