Fig. 3: Airyscan live-cell imaging reveals asymmetric actin organization at CME sites.

a A representative single time frame image of an Airyscan movie (Supplementary Movie 3) of DNM2-tagGFP2 (red) and JF635 ligand-conjugated ARPC3-HaloTag (cyan) on the ventral plasma membrane surface of ADA cells. The highlighted region is boxed by a dashed line. Scale bar: 1 µm. b Histogram of distance between the center of mass of DNM2 and ARPC3 at frame corresponding to scission. Mean and standard deviation are shown on the graph. Data were analyzed using Prism 9. Source data are provided in the Source Data file. c Montage of average intensity projection of representative ARPC3 positive CME sites. Frame corresponding to scission was determined by maximum DNM2 intensity. 33 × 33 pixels square region centered at DNM2 maximum intensity pixel was cropped for six continuous frames ending in scission. Cropped frame series were rotated by 90 or 180 degrees as needed to align the ARPC3 signal to the left of the center of the image at the frame corresponding to scission. *: Averaged image of the frame corresponding to scission (maximum DNM2 intensity) is marked to show the displacement between the CME neck (DNM2) and branched actin (ARPC3). Scale bar: 1 µm. Intervals: 0.2 s. N = 72. d The line scan function in ImageJ software was used to measure the fluorescence intensity along a one pixel wide, 33 pixel long horizontal line drawn across the center of the averaged intensity image of the frame corresponding to scission* in c. The signals were normalized to the minimum signal along the line, and intensity was calculated as a ratio to the maximum signal along the line. Source data are provided in the Source Data file.