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bioRxiv · 10.1101/229732

A new CRISPR screening approach for identifying novel autophagy-related factors and cytoplasm-to-lysosome trafficking routes

Shoemaker, C. J.·Huang, T. Q.·Weir, N. R.·Polyakov, N.·Denic, V.
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Abstract

Selective autophagy comprises cytoplasm-to-lysosome trafficking routes that transport cargos using double-membrane vesicles (autophagosomes). Cargos are detected by receptor proteins, which typically also bind to lipid-conjugated LC3 proteins on autophagosome membranes. We dissected lysosomal delivery of four SQSTM1-like receptors by genome-wide CRISPR screening looking for novel autophagy-related (ATG) factors and trafficking routes. We uncovered new mammalian ATG factors including TMEM41B, an endoplasmic reticulum membrane protein required for autophagosome membrane expansion and/or closure. Furthermore, we found that certain receptors remain robustly targeted to the lysosome even in the absence of ATG7 or other LC3 conjugation factors. Lastly, we identified a unique genetic fingerprint behind receptor flux in ATG7KO cells, which includes factors implicated in nucleating autophagosome formation and vesicle trafficking factors. Our work uncovers new ATG factors, reveals a malleable network of autophagy receptor genetic interactions, and provides a valuable resource (http://crispr.deniclab.com) for further mining of novel autophagy mechanisms.

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Shoemaker, C. J., Huang, T. Q., Weir, N. R., Polyakov, N., Denic, V.. 2017-12-06. A new CRISPR screening approach for identifying novel autophagy-related factors and cytoplasm-to-lysosome trafficking routes. https://doi.org/10.1101/229732

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