Quantitative proteomics reveals the selectivity of ubiquitin-binding autophagy receptors in the turnover of damaged lysosomes by lysophagy
- 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
Article and author information
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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.
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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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
