Auto-regulation of Rab5 GEF activity in Rabex5 by allosteric structural changes, catalytic core dynamics and ubiquitin binding

  1. Janelle Lauer
  2. Sandra Segeletz
  3. Alice Cezanne
  4. Giambattista Guaitoli
  5. Francesco Raimondi
  6. Marc Gentzel
  7. Vikram Alva
  8. Michael Habeck
  9. Yannis Kalaidzidis
  10. Marius Ueffing
  11. Andrei N Lupas
  12. Christian Johannes Gloeckner
  13. Marino Zerial  Is a corresponding author
  1. Max Planck Institute of Molecular Cell Biology and Genetics, Germany
  2. German Center for Neurodegenerative Diseases, Germany
  3. Heidelberg University, Germany
  4. Technical University Dresden, Germany
  5. Max Planck Institute for Developmental Biology, Germany
  6. Max Planck Institute for Biophysical Chemistry, Germany
  7. University of Tübingen, Germany

Abstract

Intracellular trafficking depends on the function of Rab GTPases, whose activation is regulated by guanine exchange factors (GEFs). The Rab5 GEF, Rabex5, was previously proposed to be auto-inhibited by its C-terminus. Here, we studied full-length Rabex5 and Rabaptin5 proteins as well as domain deletion Rabex5 mutants using hydrogen deuterium exchange mass spectrometry. We generated a structural model of Rabex5, using chemical cross-linking mass spectrometry and integrative modeling techniques. By correlating structural changes with nucleotide exchange activity for each construct, we uncovered new auto-regulatory roles for the Ubiquitin binding domains and the Linker connecting those domains to the catalytic core of Rabex5. We further provide evidence that enhanced dynamics in the catalytic core are linked to catalysis. Our results suggest a more complex auto-regulation mechanism than previously thought and imply that Ubiquitin binding serves not only to position Rabex5 but to also control its Rab5 GEF activity through allosteric structural alterations.

Data availability

Data generated for figures 1a and 3 are included in the supporting files.

Article and author information

Author details

  1. Janelle Lauer

    Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1412-6766
  2. Sandra Segeletz

    Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany
    Competing interests
    The authors declare that no competing interests exist.
  3. Alice Cezanne

    Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany
    Competing interests
    The authors declare that no competing interests exist.
  4. Giambattista Guaitoli

    Translational Biomarker in Neurodegenerative Diseases, German Center for Neurodegenerative Diseases, Tübingen, Germany
    Competing interests
    The authors declare that no competing interests exist.
  5. Francesco Raimondi

    Bioquant, Heidelberg University, Heidelberg, Germany
    Competing interests
    The authors declare that no competing interests exist.
  6. Marc Gentzel

    Molecular Analysis Mass Spectrometry Center for Molecular and Cellular Bioimaging, Technical University Dresden, Dresden, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4482-6010
  7. Vikram Alva

    Department of Protein Evolution, Max Planck Institute for Developmental Biology, Tübingen, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1188-473X
  8. Michael Habeck

    Statistical Inverse Problems in Biophysics, Max Planck Institute for Biophysical Chemistry, Göttingen, Germany
    Competing interests
    The authors declare that no competing interests exist.
  9. Yannis Kalaidzidis

    Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany
    Competing interests
    The authors declare that no competing interests exist.
  10. Marius Ueffing

    Institute for Ophthalmic Research, Center for Ophthalmology, University of Tübingen, Tübingen, Germany
    Competing interests
    The authors declare that no competing interests exist.
  11. Andrei N Lupas

    Department of Protein Evolution, Max Planck Institute for Developmental Biology, Tübingen, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-1959-4836
  12. Christian Johannes Gloeckner

    Translational Biomarker in Neurodegenerative Diseases, German Center for Neurodegenerative Diseases, Tübingen, Germany
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6494-6944
  13. Marino Zerial

    Max Planck Institute of Molecular Cell Biology and Genetics, Dresden, Germany
    For correspondence
    zerial@mpi-cbg.de
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-7490-4235

Funding

Max-Planck-Gesellschaft (Open-access funding)

  • Marino Zerial

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

Reviewing Editor

  1. Suzanne R Pfeffer, Stanford University School of Medicine, United States

Publication history

  1. Received: February 21, 2019
  2. Accepted: October 22, 2019
  3. Accepted Manuscript published: November 13, 2019 (version 1)
  4. Version of Record published: November 14, 2019 (version 2)
  5. Version of Record updated: August 18, 2022 (version 3)

Copyright

© 2019, Lauer 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. Janelle Lauer
  2. Sandra Segeletz
  3. Alice Cezanne
  4. Giambattista Guaitoli
  5. Francesco Raimondi
  6. Marc Gentzel
  7. Vikram Alva
  8. Michael Habeck
  9. Yannis Kalaidzidis
  10. Marius Ueffing
  11. Andrei N Lupas
  12. Christian Johannes Gloeckner
  13. Marino Zerial
(2019)
Auto-regulation of Rab5 GEF activity in Rabex5 by allosteric structural changes, catalytic core dynamics and ubiquitin binding
eLife 8:e46302.
https://doi.org/10.7554/eLife.46302
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