This website uses cookies so that we can provide you with the best user experience possible. Cookie information is stored in your browser and performs functions such as recognising you when you return to our website and helping our team to understand which sections of the website you find most interesting and useful.
Proximity biotinylation of ATG8 proteins and selective autophagy receptors
Output Details
Description
Autophagy involves the formation of an autophagosome around a cellular cargo, and once encapsulated in this double membrane structure, the autophagosome fuses with the lysosome to facilitate degradation of the cargo. Membrane-bounded organelles such as mitochondria are frequent targets. This protocol concerns the use of proximity biotinylation with the engineered peroxidase APEX2 (doi.org/10.1038/nprot.2016.018) fused with either ATG8 proteins or autophagic cargo receptors in mammalian cells. There are six ATG8 proteins in mammals (MAP1LC3A, B, and C & GABARAP, L1, and L2). These proteins bind LC3 interacting motifs (LIR motifs) on cargo receptors. Therefore, proximity biotinylation can be used to identify proteins that are nearby ATG8 proteins in response to activation of selective autophagy in cells. This protocol uses nutrient stress as the autophagy activator. The protocol builds on a previously published method (Hung et al., 2016)
Identifier (DOI)
10.17504/protocols.io.dm6gp3jb1vzp/v1