title: Mechanism of cargo-directed Atg8 conjugation during selective autophagy.
abstract: Selective autophagy is mediated by cargo receptors that link the cargo to the isolation membrane via interactions with Atg8 proteins. Atg8 proteins are localized to the membrane in an ubiquitin-like conjugation reaction, but how this conjugation is coupled to the presence of the cargo is unclear. Here we show that theS. cerevisiaeAtg19, Atg34 and the human p62, Optineurin and NDP52 cargo receptors interact with the E3-like enzyme Atg12~Atg5-Atg16, which stimulates Atg8 conjugation. The interaction of Atg19 with the Atg12~Atg5-Atg16 complex is mediated by its Atg8-interacting motifs (AIMs). We identify the AIM-binding sites in the Atg5 subunit and mutation of these sites impairs selective autophagy. In a reconstituted system the recruitment of the E3 to the prApe1 cargo is sufficient to drive accumulation of conjugated Atg8 at the cargo. The interaction of the Atg12~Atg5-Atg16 complex and Atg8 with Atg19 is mutually exclusive, which may confer directionality to the system.
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Dorotea
authors.initials: D
authors.lastname: Fracchiolla
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Justyna
authors.initials: J
authors.lastname: Sawa-Makarska
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Bettina
authors.initials: B
authors.lastname: Zens
authors.affiliation: Department of Structural and Computational Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Anita de
authors.initials: A
authors.lastname: Ruiter
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Zaffagnini
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Andrea
authors.initials: A
authors.lastname: Brezovich
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Julia
authors.initials: J
authors.lastname: Romanov
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Runggatscher
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Claudine
authors.initials: C
authors.lastname: Kraft
authors.affiliation: Department of Structural and Computational Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Bojan
authors.initials: B
authors.lastname: Zagrovic
authors.affiliation: Department of Biochemistry and Cell Biology, Max F. Perutz Laboratories (MFPL), University of Vienna, Vienna Biocenter (VBC), Vienna, Austria.
authors.forename: Sascha
authors.initials: S
authors.lastname: Martens
mesh_terms: D000595:Amino Acid Sequence; D001343:Autophagy / Q000235:genetics*; D000071192:Autophagy-Related Protein 12 / Q000235:genetics / Q000378:metabolism; D000071187:Autophagy-Related Protein 5 / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000071190:Autophagy-Related Protein 8 Family / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000071183:Autophagy-Related Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001665:Binding Sites; D001692:Biological Transport; D018797:Cell Cycle Proteins; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005786:Gene Expression Regulation; D006367:HeLa Cells; D006801:Humans; D026901:Membrane Transport Proteins; D062105:Molecular Docking Simulation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D017433:Protein Structure, Secondary; D011956:Receptors, Cell Surface / Q000235:genetics / Q000378:metabolism; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D000071456:Sequestosome-1 Protein / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D035582:Transcription Factor TFIIIA / Q000235:genetics / Q000378:metabolism; D033921:Vesicular Transport Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: C491969:ATG12 protein, S cerevisiae; C498649:ATG16 protein, S cerevisiae; C434480:ATG19 protein, S cerevisiae; C102268:ATG5 protein, S cerevisiae; C113757:ATG8 protein, S cerevisiae; C552963:Atg34 protein, S cerevisiae; D000071192:Autophagy-Related Protein 12; D000071187:Autophagy-Related Protein 5; D000071190:Autophagy-Related Protein 8 Family; D000071183:Autophagy-Related Proteins; C094297:CALCOCO2 protein, human; D018797:Cell Cycle Proteins; D026901:Membrane Transport Proteins; D009687:Nuclear Proteins; C409460:OPTN protein, human; D011956:Receptors, Cell Surface; D018160:Receptors, Cytoplasmic and Nuclear; D011994:Recombinant Proteins; C099718:SQSTM1 protein, human; D029701:Saccharomyces cerevisiae Proteins; D000071456:Sequestosome-1 Protein; D035582:Transcription Factor TFIIIA; D033921:Vesicular Transport Proteins
keywords: S. cerevisiae; biochemical reconstitution; biochemistry; cargo receptor; human; membrane biology; protein conjugation; selective autophagy