Fig. 9

hFis1 is required for MIEF-induced mitochondrial fusion. a Generation of hFis1 KO 293T cell lines by CRISPR/Cas9-based genome editing was confirmed by Western blot analysis obtained from the single cell colonies derived hFis1 KO 293T cells. b, c WT 293T cells co-transfected with mito-PAGFP (0.5 μg) and mito-DsRed (0.2 μg), or hFis1 KO 293T cells co-transfected with mito-PAGFP (0.5 μg), mito-DsRed (0.2 μg), and either empty vector (0.5 μg), MIEF1 (0.5 μg), or MIEF2 (0.5 μg) were photoactivated in the ROI (white circle, 3 μm diameter) in preactivation images of mitochondria (red). After photoactivation, mito-PAGFP fluorescence (green) intensity and mitochondrial marker (mito-DsRed) were collected at indicated time points (b). Mitochondrial fusion was quantified by analyzing changes in fluorescence intensity of photoactivated mito-PAGFP in ROIs at 40 s and 10, 20, and 30 min. The dilution rates (percentage) of the GFP fluorescence intensity at different time points were normalized by the fluorescence intensity at 40 s after photoactivation (c) (n represents the number of cells analyzed)