Molecular Cell
Volume 81, Issue 6, 18 March 2021, Pages 1337-1354.e8
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Systematically defining selective autophagy receptor-specific cargo using autophagosome content profiling

https://doi.org/10.1016/j.molcel.2021.01.009Get rights and content
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Highlights

  • Protease protection coupled APEX2 proximity proteomics of autophagosomes

  • Identification of selective autophagy receptor-dependent cargo

  • Ubl conjugation and protein aggregation only modestly drive cargo engulfment

  • The aggrephagy receptor TOLLIP is implicated in endosomal microautophagy

Summary

Autophagy deficiency in fed conditions leads to the formation of protein inclusions highlighting the contribution of this lysosomal delivery route to cellular proteostasis. Selective autophagy pathways exist that clear accumulated and aggregated ubiquitinated proteins. Receptors for this type of autophagy (aggrephagy) include p62, NBR1, TOLLIP, and OPTN, which possess LC3-interacting regions and ubiquitin-binding domains (UBDs), thus working as a bridge between LC3/GABARAP proteins and ubiquitinated substrates. However, the identity of aggrephagy substrates and the redundancy of aggrephagy and related UBD-containing receptors remains elusive. Here, we combined proximity labeling and organelle enrichment with quantitative proteomics to systematically map the autophagic degradome targeted by UBD-containing receptors under basal and proteostasis-challenging conditions in human cell lines. We identified various autophagy substrates, some of which were differentially engulfed by autophagosomal and endosomal membranes via p62 and TOLLIP, respectively. Overall, this resource will allow dissection of the proteostasis contribution of autophagy to numerous individual proteins.

Keywords

autophagy
aggrephagy
selective autophagy receptors
SQSTM1/p62
TOLLIP
autophagosomes
APEX2
proximity labeling
endosomal microautophagy
proteostasis imbalance

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