Lateral compartmentalization of continuous mitochondria. (A–C) Dual-color FLIP in wild-type cells expressing GFP-tagged mitochondrial proteins and matrix-mCherry. Representative images, pooled quantification data of GFP and mCherry FLIP, and tX (time to reduce to X% of the total fluorescence) of GFP FLIP from three independent experiments are shown. (A) Wild-type cells expressing Hem1-GFP and matrix-mCherry (n = 30 cells, n = 10 cells for each experiment). (B) Wild-type cells expressing Tom20-GFP and matrix-mCherry (n = 32 cells, n ≥ 10 cells for each experiment). (C) Wild-type cells expressing Atp1-GFP and matrix-mCherry (n = 25 cells, n ≥ 8 cells for each experiment). Photobleach was applied in the GFP and mCherry channels as indicated with white circles. Images are a sum projection of five z-stacks taken at 0.5-μm intervals. Scale bar: 3 μm. Data from three independent clones were pooled to obtain the bleaching curves. Shadows represent mean ± SD. Error bar: mean ± SE. Welch’s two-tailed t test was applied to compare the tX in the mother and bud.