Shuffled ATG8 interacting motifs form an ancestral bridge between UFMylation and autophagy

Picchianti, Lorenzo and Sánchez de Medina Hernández, Víctor and Zhan, Ni and Irwin, Nicholas AT and Groh, Roan and Stephani, Madlen and Hornegger, Harald and Beveridge, Rebecca and Sawa‐Makarska, Justyna and Lendl, Thomas and Grujic, Nenad and Naumann, Christin and Martens, Sascha and Richards, Thomas A and Clausen, Tim and Ramundo, Silvia and Karagöz, G Elif and Dagdas, Yasin (2023) Shuffled ATG8 interacting motifs form an ancestral bridge between UFMylation and autophagy. The EMBO journal, 42 (10). e112053. ISSN 1460-2075 (

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UFMylation involves the covalent modification of substrate proteins with UFM1 (Ubiquitin‐fold modifier 1) and is important for maintaining ER homeostasis. Stalled translation triggers the UFMylation of ER‐bound ribosomes and activates C53‐mediated autophagy to clear toxic polypeptides. C53 contains noncanonical shuffled ATG8‐interacting motifs (sAIMs) that are essential for ATG8 interaction and autophagy initiation. However, the mechanistic basis of sAIM‐mediated ATG8 interaction remains unknown. Here, we show that C53 and sAIMs are conserved across eukaryotes but secondarily lost in fungi and various algal lineages. Biochemical assays showed that the unicellular alga Chlamydomonas reinhardtii has a functional UFMylation pathway, refuting the assumption that UFMylation is linked to multicellularity. Comparative structural analyses revealed that both UFM1 and ATG8 bind sAIMs in C53, but in a distinct way. Conversion of sAIMs into canonical AIMs impaired binding of C53 to UFM1, while strengthening ATG8 binding. Increased ATG8 binding led to the autoactivation of the C53 pathway and sensitization of Arabidopsis thaliana to ER stress. Altogether, our findings reveal an ancestral role of sAIMs in UFMylation‐dependent fine‐tuning of C53‐mediated autophagy activation.