Endosome fission defect due to reduction of EHD1 recruitment to endosomes in ORP10 KO cells. (A) Confocal microscopic images of control (Cont) or ORP10 KO cells showing localization of GFP-EHD1 and endogenous SNX1. Scale bars, 2.5 µm (full size) and 1 µm (inset). (B) Quantification of EHD1 fluorescence (Fluo.) at SNX1-positive endosomes (mean ± SEM; n ≥ 10 cells). (C) Gallery of representative confocal time-lapse images showing the fission process visualized with GFP-SNX1 in control or ORP10 KO cells. Scale bar, 1 µm. (D) Quantification of times required for completion of tubule fission in control (Cont), ORP10 KO cells, ORP10 KO cells rescued by GFP fused to ORP10 (ORP10^Full), ΔORD (ORP10^ΔORD), or ORP9 (mean ± SEM, the number of fission events for quantification; n = 35 from 11 cells for control, n = 44 from 16 cells for ORP10 KO, n = 33 from 12 cells for ORP10^Full, n = 21 from 10 cells for ORP10^ΔORD, n = 25 from 10 cells for ORP9). ****, P < 0.0001.