PINK1 is needed to target tubule components for ER-phagy at three-way junctions. (A and B) YFP-RTN3L (A) and LNPK-GFP (B) were quantitated in LAMP1-mCherry marked structures. YFP-RTN3L samples were treated with Baf for 4 h, while LNPK-GFP samples were treated for 6 h. (C) PINK1 and FIP200 are needed for the lysosomal delivery of the ER-phagy reporter ssRFP-GFP-KDEL. The number of red dots per cell are reported for the indicated samples. (D) Stills (3.6, 8.1 and 9.6 s) from Video 1 of cells transfected with mCherry-KDEL and YFP-PINK1. (E–G) Quantitation and representative images used to calculate the percent cells with large Akita puncta (≥0.5 µm²) in Ctrl and siRNA-treated cells. In E, the siPINK1-treated cells were transfected with mCherry-RNAi-resistant PINK1 or mCherry-RNAi-resistant PINK1 kinase dead (KD). Arrows mark large Akita puncta. Error bars represent SEM, n = 3 independent experiments. The results were quantified from 58 to 69 cells in A, 47–59 cells in B, 69–85 cells in C, 99–111 cells in E, 79–105 cells in F, and 79–101 cells in G. NS: not significant (P ≥ 0.05), * (P < 0.05), **(P < 0.01), *** (P < 0.001), Student’s unpaired t test.