Fig. 4: In vitro recruitment of lipid membrane by p62 bodies.
From: Local membrane source gathering by p62 body drives autophagosome formation
a Schematic diagram of the sedimentation assay to separate the condensed liquid droplets and the supernatant. Phase separation assay of mCherry-p62 with linear Ub8 in vitro. Atg2ab DKO cytosol was added after the phase separation. mCherry-p62, 6 µM; His-Ub8: 2 µM; △PB1, 6 µM. b The sedimentation and supernatant from (a) were separated by centrifugation and analyzed by western blot using antibodies against FIP200, ATG9A, ATG16L1, LC3 and GAPDH. S: supernatant, P: pellet. c p62 body and different protein-interacting vesicles from (a) were co-stained with antibodies against p62, FIP200, ATG9A, ATG16L1 and LC3. Scale bar, 2 µm. d Schematic diagram of the sedimentation assay to separate the condensed liquid droplets and the supernatant. Phase separation assay of mCherry-p62 with linear His-Ub8 in vitro. Liposome was added after the phase separation. mCherry-p62, 50 µM; His-Ub8: 15 µM; Liposome, 1 µM. e Liposomes (PE-NDB) around the p62 bodies were observed from the (d). Scale bar, 1 µm. f Liposomes (PE-NDB) around the p62 bodies were observed by TEM from the (d). Scale bar, 500 nm.
