Fig. 5: Initiation hubs for selective autophagy are conserved in human cells.

a, U2OS cells transfected with FKBP–GFP–ULK1 and mCherry–Parkin were stained with MitoTracker DeepRed. Mitophagy was induced with antimycin A and oligomycin (AO). Live cells were visualized by fluorescence microscopy. Scale bar, 10 µm; Scale bar inset, 1 µm. Quantification: cells containing GFP–ULK1 foci. Data are mean values (n > 130 cells per condition across replicates, four biological replicates). Circles show mean values of each replicate, bars show the mean. Statistical analysis was conducted by a one-tailed unpaired t-test, ****P < 0.0001. b, U2OS cells transfected with mCherry–LC3B and YFP–Parkin were stained with MitoTracker DeepRed and treated with AO. Images of one out of three biological replicates are shown. Scale bar, 10 µm. Quantification: cells containing mCherry–LC3B foci. Data are mean values (n > 50 cells per condition across replicates, three biological replicates). Circles show mean values of each replicate, bars show mean. Statistical analysis was conducted by a one-tailed unpaired t-test, ****P = 0.0001. c, U2OS cells were transfected with FKBP–GFP or FKBP–GFP–ULK1 and mCherry–p62, and cultured in nutrient-rich medium or starvation medium (Earle’s balanced salt solution; EBSS) for 4 h to induce bulk autophagy. Live cells were visualized by fluorescence microscopy. Images of one out of three biological replicates are shown. Scale bar, 10 µm, scale bar inset, 1 µm. Quantification: co-localization events between GFP and p62. Data are mean values (n > 100 cells per condition across replicates, three biological replicates). Circles show mean values of each replicate, bars show mean. Statistical analysis was conducted by one-way ANOVA followed by Sidak’s multiple comparison test, ****P < 0.0001. Source numerical data are available in Source data.