2024/10/12 by Robin Ketteler, Koshiro Kiso, Lucas von Chamier +1 · 1 voice
Biochemistry, Genetics and Molecular Biology · Medicine · #Autophagy in Disease and Therapy #CRISPR and Genetic Engineering #Endoplasmic Reticulum Stress and Disease
paper · pdf · doi:10.1080/27694127.2024.2392450
openalex publication_date 2024/10/12 · openalex created_date 2025/10/10 · openalex updated_date 2026/06/19
Protein ATG8ylation refers to a post-translational modification involving covalent attachment of ubiquitin-like autophagy-related protein ATG8 (LC3/GABARAP) to other cellular proteins, with reversal mediated by ATG4 proteases. While lipid ATG8ylation is important for autophagosome formation and mechanistically well-characterized, little is known about the mechanism of protein ATG8ylation. Here, we investigated the conjugation machinery of protein ATG8ylation in CRISPR/Cas9-engineered knockout human cell lines, utilizing a deconjugation-resistant (Q116P G120) form of MAP1LC3B. We report that protein ATG8ylation requires the E1-like activating enzyme ATG7 and E2-like conjugating enzyme ATG3, in common with ATG8 lipidation. However, in contrast, the E3-like ATG12-ATG5-ATG16L1 complex involved in lipidation is dispensable for protein ATG8ylation, since ATG5 knockout cells can form ATG8ylated protein conjugates. Further, we uncover that ATG7 itself is a target of ATG8ylation. Overall, our work provides crucial insight into the mechanism of protein ATG8ylation, distinguishing it from ATG8 lipidation, which will aid investigating its functional role.