title: Atg8 family LC3/GABARAP proteins are crucial for autophagosome-lysosome fusion but not autophagosome formation during PINK1/Parkin mitophagy and starvation.
abstract: Members of the Atg8 family of proteins are conjugated to autophagosomal membranes, where they have been proposed to drive autophagosome formation and selective sequestration of cargo. In mammals, the Atg8 family consists of six members divided into the LC3 and GABARAP subfamilies. To define Atg8 function, we used genome editing to generate knockouts of the LC3 and GABARAP subfamilies as well as all six Atg8 family members in HeLa cells. We show that Atg8s are dispensable for autophagosome formation and selective engulfment of mitochondria, but essential for autophagosome-lysosome fusion. We find that the GABARAP subfamily promotes PLEKHM1 recruitment and governs autophagosome-lysosome fusion, whereas the LC3 subfamily plays a less prominent role in these processes. Although neither GABARAPs nor LC3s are required for autophagosome biogenesis, loss of all Atg8s yields smaller autophagosomes and a slowed initial rate of autophagosome formation. Our results clarify the essential function of the Atg8 family and identify GABARAP subfamily members as primary contributors to PINK1/Parkin mitophagy and starvation autophagy.
authors.affiliation: Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Melbourne, Victoria 3800, Australia.
authors.forename: Thanh Ngoc
authors.identifier: http://orcid.org/0000-0001-9698-0020
authors.initials: TN
authors.lastname: Nguyen
authors.affiliation: Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Melbourne, Victoria 3800, Australia.
authors.forename: Benjamin Scott
authors.identifier: http://orcid.org/0000-0002-5710-6100
authors.initials: BS
authors.lastname: Padman
authors.affiliation: Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Melbourne, Victoria 3800, Australia.
authors.forename: Joanne
authors.identifier: http://orcid.org/0000-0002-4926-802X
authors.initials: J
authors.lastname: Usher
authors.affiliation: Monash Ramaciotti Centre for Cryo Electron Microscopy, Monash University, Melbourne, Victoria 3800, Australia.
authors.forename: Viola
authors.identifier: http://orcid.org/0000-0002-8122-8271
authors.initials: V
authors.lastname: Oorschot
authors.affiliation: Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Melbourne, Victoria 3800, Australia.|Monash Ramaciotti Centre for Cryo Electron Microscopy, Monash University, Melbourne, Victoria 3800, Australia.
authors.forename: Georg
authors.identifier: http://orcid.org/0000-0003-3596-2288
authors.initials: G
authors.lastname: Ramm
authors.affiliation: Department of Biochemistry and Molecular Biology, Biomedicine Discovery Institute, Monash University, Melbourne, Victoria 3800, Australia michael.lazarou@monash.edu.
authors.forename: Michael
authors.identifier: http://orcid.org/0000-0003-2150-5545
authors.initials: M
authors.lastname: Lazarou
mesh_terms: D000143:Acids / Q000378:metabolism; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D051017:Apoptosis Regulatory Proteins; D000071182:Autophagosomes / Q000378:metabolism* / Q000648:ultrastructure; D000071190:Autophagy-Related Protein 8 Family / Q000378:metabolism*; D000071183:Autophagy-Related Proteins; D055786:Gene Knockout Techniques; D006367:HeLa Cells; D006801:Humans; D008247:Lysosomes / Q000378:metabolism* / Q000648:ultrastructure; D008561:Membrane Fusion; D008562:Membrane Glycoproteins / Q000378:metabolism; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D063306:Mitophagy; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*
chemical_list: D000143:Acids; D048868:Adaptor Proteins, Signal Transducing; D051017:Apoptosis Regulatory Proteins; D000071190:Autophagy-Related Protein 8 Family; D000071183:Autophagy-Related Proteins; C116741:GABARAP protein, human; C477012:MAP1LC3A protein, human; D008562:Membrane Glycoproteins; D008869:Microtubule-Associated Proteins; C476215:PLEKHM1 protein, human; D044767:Ubiquitin-Protein Ligases; C111567:parkin protein