Fig. 1: Actin marks the site of mitochondrial fusion.

a Quantification of fission and fusion events in cells in which mitochondria were stained with Mitotracker (MTR) and cells transfected with mCherry-mito and AC-mito (AC). Each point represents an individual cell, with at 12 cells (MTR) and 8 cells (AC) quantified in 4 independent experiments. Bars show the average ±SD. b, c AC-positive fission and fusion events. Cells were transfected with mCherry-mito (mitochondria, magenta) and AC-mito (actin, green). Quantification of fission (Fis.) and fusion (Fus.) events positive for AC-mito. Each point represents an individual cell, with 10 cells quantified in 4 independent experiments (same cells as in (a) AC). Bars show the average ± SD. Two-sided t-test. Representative fusion event showing the enrichment of AC-mito at the fusion site (arrowhead) is shown in (c). Scale bar 2 µm. d Number of AC-mito-positive fusion events compared to the expected number AC-mito-positive fusion events predicted from the mitochondrial area covered AC-mito). Fisher’s exact test. e, f AC-mito signal enrichment at fusion (e) and fission (f) sites in cells transfected as in B. Signal intensity at the event site relative to an adjacent site on the mitochondrial network was quantified for AC-mito (AC) and mCherry-mito (mCh). Each point represents an individual cell, with 8 cells quantified in 4 independent experiments. Bars show the average ±SD. Two-sided t-test. g, h SIM imaging of U2OS transfected with AC-mito and stained with mitotracker. g Representative image showing AC-mito (Green) and mitochondria (Mitotracker, Magenta). Scale bar 1 µm. The arrowheads point to the fusion site. h Quantification of the number of fusion events where actin bridges the two fusing mitochondria as in the bottom image in (g). The total number of fusion events from 5 cells is shown. The same data was also separated into tip-to-tip and tip-to-side fusion events (see Fig. 2).