1. Biochemistry and Chemical Biology
  2. Structural Biology and Molecular Biophysics
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Structural and mutational analysis of the ribosome-arresting human XBP1u

  1. Vivekanandan Shanmuganathan
  2. Nina Schiller
  3. Anastasia Magoulopoulou
  4. Jingdong Cheng
  5. Katharina Braunger
  6. Florian Cymer
  7. Otto Berninghausen
  8. Birgitta Beatrix
  9. Kenji Kohno
  10. Gunnar von Heijne  Is a corresponding author
  11. Roland Beckmann  Is a corresponding author
  1. Ludwig Maximilian University of Munich, Germany
  2. Stockholm University, Sweden
  3. Nara Institute of Science and Technology, Japan
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Cite this article as: eLife 2019;8:e46267 doi: 10.7554/eLife.46267

Abstract

XBP1u, a central component of the unfolded protein response (UPR), is a mammalian protein containing a functionally critical translational arrest peptide (AP). Here, we present a 3 Å cryo-EM structure of the stalled human XBP1u AP. It forms a unique turn in the ribosomal exit tunnel proximal to the peptidyl transferase center where it causes a subtle distortion, thereby explaining the temporary translational arrest induced by XBP1u. During ribosomal pausing the hydrophobic region 2 (HR2) of XBP1u is recognized by SRP, but fails to efficiently gate the Sec61 translocon. An exhaustive mutagenesis scan of the XBP1u AP revealed that only 8 out of 20 mutagenized positions are optimal; in the remaining 12 positions, we identify 55 different mutations increase the level of translational arrest. Thus, the wildtype XBP1u AP induces only an intermediate level of translational arrest, allowing efficient targeting by SRP without activating the Sec61 channel.

Article and author information

Author details

  1. Vivekanandan Shanmuganathan

    Gene Center, Department of Biochemistry, Ludwig Maximilian University of Munich, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  2. Nina Schiller

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  3. Anastasia Magoulopoulou

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  4. Jingdong Cheng

    Gene Center, Department of Biochemistry, Ludwig Maximilian University of Munich, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  5. Katharina Braunger

    Gene Center, Department of Biochemistry, Ludwig Maximilian University of Munich, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-9067-2155
  6. Florian Cymer

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    Competing interests
    The authors declare that no competing interests exist.
  7. Otto Berninghausen

    Gene Center, Department of Biochemistry, Ludwig Maximilian University of Munich, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  8. Birgitta Beatrix

    Gene Center, Department of Biochemistry, Ludwig Maximilian University of Munich, Munich, Germany
    Competing interests
    The authors declare that no competing interests exist.
  9. Kenji Kohno

    Institute for Research Initiatives, Nara Institute of Science and Technology, Ikoma, Japan
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3503-6551
  10. Gunnar von Heijne

    Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden
    For correspondence
    gunnar.von.heijne@dbb.su.se
    Competing interests
    The authors declare that no competing interests exist.
  11. Roland Beckmann

    Gene Center, Department of Biochemistry, Ludwig Maximilian University of Munich, Munich, Germany
    For correspondence
    beckmann@genzentrum.lmu.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-4291-3898

Funding

Deutsche Forschungsgemeinschaft (QBM (Quantitative Biosciences Munich) Graduate School Fellowship)

  • Vivekanandan Shanmuganathan

Knut och Alice Wallenbergs Stiftelse (2012.0282)

  • Gunnar von Heijne

Vetenskapsrådet (621-2014-3713)

  • Gunnar von Heijne

Cancerfonden (15 0888)

  • Gunnar von Heijne

Japan Society for the Promotion of Science (JP26116006)

  • Kenji Kohno

Deutsche Forschungsgemeinschaft (GRK1721)

  • Roland Beckmann

Boehringer Ingelheim Fonds (Graduate Fellowship)

  • Katharina Braunger

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

Reviewing Editor

  1. Ramanujan S Hegde, MRC Laboratory of Molecular Biology, United Kingdom

Publication history

  1. Received: February 20, 2019
  2. Accepted: June 20, 2019
  3. Accepted Manuscript published: June 27, 2019 (version 1)
  4. Version of Record published: July 11, 2019 (version 2)

Copyright

© 2019, Shanmuganathan 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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