Quantitative proteomics reveals the selectivity of ubiquitin-binding autophagy receptors in the turnover of damaged lysosomes by lysophagy

  1. Vinay V Eapen
  2. Sharan Swarup
  3. Melissa J Hoyer
  4. Joao A Paulo
  5. Wade Harper  Is a corresponding author
  1. Harvard Medical School, United States

Abstract

Removal of damaged organelles via the process of selective autophagy constitutes a major form of cellular quality control. Damaged organelles are recognized by a dedicated surveillance machinery, leading to the assembly of an autophagosome around the damaged organelle, prior to fusion with the degradative lysosomal compartment. Lysosomes themselves are also prone to damage and are degraded through the process of lysophagy. While early steps involve recognition of ruptured lysosomal membranes by glycan-binding Galectins and ubiquitylation of transmembrane lysosomal proteins, many steps in the process, and their inter-relationships, remain poorly understood, including the role and identity of cargo receptors required for completion of lysophagy. Here, we employ quantitative organelle capture and proximity biotinylation proteomics of autophagy adaptors, cargo receptors, and Galectins in response to acute lysosomal damage, thereby revealing the landscape of lysosome-associated proteome remodeling during lysophagy. Among proteins dynamically recruited to damaged lysosomes were ubiquitin-binding autophagic cargo receptors. Using newly developed lysophagic flux reporters including Lyso-Keima, we demonstrate that TAX1BP1, together with its associated kinase TBK1, are both necessary and sufficient to promote lysophagic flux in both HeLa cells and induced neurons (iNeurons). While the related receptor OPTN can drive damage-dependent lysophagy when overexpressed, cells lacking either OPTN or CALCOCO2 still maintain significant lysophagic flux in HeLa cells. Mechanistically, TAX1BP1-driven lysophagy requires its N-terminal SKICH domain, which binds both TBK1 and the autophagy regulatory factor RB1CC1, and requires upstream ubiquitylation events for efficient recruitment and lysophagic flux. These results identify TAX1BP1 as a central component in the lysophagy pathway and provide a proteomic resource for future studies of the lysophagy process.

Data availability

All proteomic .RAW files have been deposited in the PRIDE component of Proteome xchange with the identifier PXDO27476, and will be released upon publication

The following data sets were generated

Article and author information

Author details

  1. Vinay V Eapen

    Harvard Medical School, Boston, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-8023-387X
  2. Sharan Swarup

    Harvard Medical School, Boston, United States
    Competing interests
    No competing interests declared.
  3. Melissa J Hoyer

    Harvard Medical School, Boston, United States
    Competing interests
    No competing interests declared.
  4. Joao A Paulo

    Harvard Medical School, Boston, United States
    Competing interests
    No competing interests declared.
  5. Wade Harper

    Harvard Medical School, Boston, United States
    For correspondence
    wade_harper@hms.harvard.edu
    Competing interests
    Wade Harper, J.W.H. is a consultant and founder of Caraway Therapeutics and a founding scientific advisory board member of Interline Therapeutics.Reviewing editor, eLife.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6944-7236

Funding

aligning science across parkinsons (ASAP-000282)

  • Wade Harper

National Institutes of Health (NS083524)

  • Wade Harper

National Institutes of Health (NS110395)

  • Wade Harper

National Institutes of Health (DK098285)

  • Joao A Paulo

Jane Coffin Childs Memorial Fund for Medical Research

  • Melissa J Hoyer

Jane Coffin Childs Memorial Fund for Medical Research

  • Vinay V Eapen

Canadian Institutes of Health Research

  • Sharan Swarup

Ned Goodnow Fund

  • Wade Harper

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

Reviewing Editor

  1. Ivan Dikic, Institute of Biochemistry II, Germany

Version history

  1. Preprint posted: July 19, 2021 (view preprint)
  2. Received: July 20, 2021
  3. Accepted: September 25, 2021
  4. Accepted Manuscript published: September 29, 2021 (version 1)
  5. Version of Record published: October 18, 2021 (version 2)

Copyright

© 2021, Eapen 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. Vinay V Eapen
  2. Sharan Swarup
  3. Melissa J Hoyer
  4. Joao A Paulo
  5. Wade Harper
(2021)
Quantitative proteomics reveals the selectivity of ubiquitin-binding autophagy receptors in the turnover of damaged lysosomes by lysophagy
eLife 10:e72328.
https://doi.org/10.7554/eLife.72328

Share this article

https://doi.org/10.7554/eLife.72328

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