EMBO J. 2023 Jun 05. e112845
Ruheena Javed,
Ashish Jain,
Thabata Duque,
Emily Hendrix,
Masroor Ahmad Paddar,
Sajjad Khan,
Aurore Claude-Taupin,
Jingyue Jia,
Lee Allers,
Fulong Wang,
Michal Mudd,
Graham Timmins,
Keith Lidke,
Tor Erik Rusten,
Prithvi Reddy Akepati,
Yi He,
Fulvio Reggiori,
Eeva-Liisa Eskelinen,
Vojo Deretic.
The canonical autophagy pathway in mammalian cells sequesters diverse cytoplasmic cargo within the double membrane autophagosomes that eventually convert into degradative compartments via fusion with endolysosomal intermediates. Here, we report that autophagosomal membranes show permeability in cells lacking principal ATG8 proteins (mATG8s) and are unable to mature into autolysosomes. Using a combination of methods including a novel in vitro assay to measure membrane sealing, we uncovered a previously unappreciated function of mATG8s to maintain autophagosomal membranes in a sealed state. The mATG8 proteins GABARAP and LC3A bind to key ESCRT-I components contributing, along with other ESCRTs, to the integrity and imperviousness of autophagic membranes. Autophagic organelles in cells lacking mATG8s are permeant, are arrested as amphisomes, and do not progress to functional autolysosomes. Thus, autophagosomal organelles need to be maintained in a sealed state in order to become lytic autolysosomes.
Keywords: ATG8; ESCRT; LC3; amphisome; autophagy