Abstract

Leucine-rich repeat kinase 2 (LRRK2) variants associated with Parkinson's disease (PD) and Crohn's disease lead to increased phosphorylation of its Rab substrates. While it has been recently shown that perturbations in cellular homeostasis including lysosomal damage can increase LRRK2 activity and localization to lysosomes, the molecular mechanisms by which LRRK2 activity is regulated have remained poorly defined. We performed a targeted siRNA screen to identify regulators of LRRK2 activity and identified Rab12 as a novel modulator of LRRK2-dependent phosphorylation of one of its substrates, Rab10. Using a combination of imaging and immunopurification methods to isolate lysosomes, we demonstrated that Rab12 is actively recruited to damaged lysosomes and leads to a local and LRRK2-dependent increase in Rab10 phosphorylation. PD-linked variants, including LRRK2 R1441G and VPS35 D620N, lead to increased recruitment of LRRK2 to the lysosome and a local elevation in lysosomal levels of pT73 Rab10. Together, these data suggest a conserved mechanism by which Rab12, in response to damage or expression of PD-associated variants, facilitates the recruitment of LRRK2 and phosphorylation of its Rab substrate(s) at the lysosome.

Data availability

All data generated or analyzed during this study are included in the manuscript and supporting files; source data files for western blots have been provided for all figures.

Article and author information

Author details

  1. Xiang Wang

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Xiang Wang, is an employee of Denali Therapeutics..
  2. Vitaliy V Bondar

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Vitaliy V Bondar, was an employees of Denali Therapeutics when these studies were conducted and is currently an employee of REGENEXBIO Inc..
  3. Oliver B Davis

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Oliver B Davis, is an employee of Denali Therapeutics..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-3622-8651
  4. Michael T Maloney

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Michael T Maloney, is an employee of Denali Therapeutics..
  5. Maayan Agam

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Maayan Agam, is an employee of Denali Therapeutics..
  6. Marcus Y Chin

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Marcus Y Chin, is an employee of Denali Therapeutics..
  7. Audrey Cheuk-Nga Ho

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Audrey Cheuk-Nga Ho, was an employee of Denali Therapeutics when these studies were conducted and is currently an employee of Cellares..
  8. Rajarshi Ghosh

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Rajarshi Ghosh, is an employee of Denali Therapeutics..
  9. Dara E Leto

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Dara E Leto, is an employee of Denali Therapeutics..
  10. David Joy

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    David Joy, is an employee of Denali Therapeutics..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-9941-9538
  11. Meredith EK Calvert

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Meredith EK Calvert, is an employee of Denali Therapeutics..
  12. Joseph W Lewcock

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Joseph W Lewcock, is an employee of Denali Therapeutics..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-3012-7881
  13. Gilbert Di Paolo

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Gilbert Di Paolo, is an employee of Denali Therapeutics..
  14. Robert G Thorne

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Robert G Thorne, is an employee of Denali Therapeutics..
  15. Zachary K Sweeney

    Denali Therapeutics, South San Francisco, United States
    Competing interests
    Zachary K Sweeney, was an employee of Denali Therapeutics when these studies were conducted and is currently an employee of Interline Therapeutics Inc..
  16. Anastasia G Henry

    Denali Therapeutics, South San Francisco, United States
    For correspondence
    henry@dnli.com
    Competing interests
    Anastasia G Henry, is an employee of Denali Therapeutics..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8124-5477

Funding

No external funding was received for this work.

Reviewing Editor

  1. Shaeri Mukherjee, University of California, San Francisco, United States

Version history

  1. Preprint posted: February 22, 2023 (view preprint)
  2. Received: February 24, 2023
  3. Accepted: October 23, 2023
  4. Accepted Manuscript published: October 24, 2023 (version 1)

Copyright

© 2023, Wang 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. Xiang Wang
  2. Vitaliy V Bondar
  3. Oliver B Davis
  4. Michael T Maloney
  5. Maayan Agam
  6. Marcus Y Chin
  7. Audrey Cheuk-Nga Ho
  8. Rajarshi Ghosh
  9. Dara E Leto
  10. David Joy
  11. Meredith EK Calvert
  12. Joseph W Lewcock
  13. Gilbert Di Paolo
  14. Robert G Thorne
  15. Zachary K Sweeney
  16. Anastasia G Henry
(2023)
Rab12 is a regulator of LRRK2 and its activation by damaged lysosomes
eLife 12:e87255.
https://doi.org/10.7554/eLife.87255

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