Association between Drp1 and Mff on ER. (A) Left: Merged confocal image of a live GFP-Drp1-KI cell expressing mito-BFP (gray), eBFP2-peroxisome (gray), mStrawberry-Mff-S (red), and pLVX-E2-Crimson-ER (blue). Drp1 in green. Right: Time-lapse confocal images of boxed region show example of a Drp1 puncta maturing from an independent Mff puncta (yellow arrows). See also Video 8. (B) Independent Mff punctae in scramble siRNA–treated cells (left) and Drp1 siRNA–treated cells (right). Left: Merged image of live U2OS cells transiently expressing GFP-Mff-S (Mff, green), eBFP2-peroxisome (Pex, blue), and mCherry-mito7 (Mito, red). Right: Insets from boxed regions in whole-cell image. Yellow arrows denote independent Mff punctae. (C) Density of independent Mff punctae in control siRNA– and Drp1 siRNA–treated U2OS cells, quantified from live-cell images of GFP-Mff as in B. Units, number of independent Mff punctae per square micrometer in the ROI. 368 independent punctae from nine control cell ROIs and 106 punctae from nine Drp1 KD cell ROIs. ***, P < 0.0001 by Student’s t test. (D) Density of independent Mff punctae in control siRNA– and Drp1 siRNA–treated U2OS cells, quantified from fixed-cell immunofluorescence of endogenous Mff. Units, number of Mff punctae per square micrometer in ROI. 643 independent punctae from five control cell ROIs and 153 puncta from seven Drp1 KD cell ROIs. ***, P < 0.0005 by Student’s t test. Bars: (whole-cell images) 10 µm; (insets) 2 µm. Time in seconds.