Supplementary Figure 4: Enrichment and turnover of cortical actin during mitosis is not dependent on myosin II.

(a) Images showing a cross sectional region of an STC-arrested mitotic β-actin-GFP expressing HeLa cell used in a photobleaching experiment. The region inside the red circle (diameter, 2 μm) was photobleached for three seconds before time zero. The actin cortex/cytoplasm ratio of the unbleached cell cortex is the ratio of mean pixel intensity in the yellow square (0.32 μm²), outside the bleach zone, and the magenta rectangle (0.32 × 1.18 μm²), which is 0.32 μm away from the unbleached region of the cortex in the cytosol. The actin cortex/cytoplasm ratio of the bleached cell cortex is the ratio of mean pixel intensity in the green square (0.32 μm²), within the bleach zone, and the magenta rectangle. Scale bar, 1.25 μm. (b) The actin cortex/cytoplasm ratio for unbleached (black) and bleached (magenta) cell cortex. The blue trace is the exponential fit of the data, post-bleaching. The half-time and recovery for the cell are indicated within the plot. (c) Plot of the time taken after photobleaching to reach half the final actin cortex/cytoplasm ratio (Half-time) and the percentage recovery of actin cortex/cytoplasm ratio compared to pre-bleached cortex. Cells were transfected at least 48 hours prior to the experiment with control (n = 9 cells) or MYH9 (n = 9 cells) siRNA. The diamond-box contains 25–75% percentiles of the data and the bar denotes the median. n indicates the number of single-cell experiments. Mann–Whitney significance tests of cells with respect to data plotted in the first column are indicated: (NS) U > 0.05, (^∗) 0.05 ≥ U > 0.005, (^∗∗) U ≤ 0.005.