Fig. 3: Genetic knockdowns and pharmacological treatments reversing the direction of the actin cytoskeleton chirality.

a–d Typical examples of actin organisation visualised by LifeAct-GFP in control siRNA (a), profilin 1 (Pfn1) siRNA (b), profilin 1 siRNA plus Pfn1 full-length plasmid (c) and profilin 2 (Pfn2) siRNA (d) transfected cells. The histograms in (a–d) show the distribution of mean RF tilt in the 6−10 μm annulus in cells treated as indicated and imaged for 12–16 h after plating. Histograms and mean±SEM values are based on measurements during the entire period of imaging of 70 control cells, 127 Pfn1 knockdown cells, 35 Pfn1 knockdown cells rescued by co-transfection with full-length Pfn1 plasmid and 182 Pfn2 knockdown cells . e Typical example of actin organisation visualised by phalloidin-staining in CapZβ siRNA-transfected cells fixed at 6 h after cell plating. Histogram under the image shows the distribution of mean RF tilt in the 8−12 μm annulus in cells at 6 h after plating. Histogram and mean±SEM value are based on 208 cells. f Average values of RF tilts (mean±SEM) as a function of the distance of annuli from the cell edge for profilins and CapZβ experiments (the same cell samples as a–e). Control siRNA #1 and #2 represent the 70 and 146 control cells used in experiments with profilins and CapZβ respectively. g Typical example of actin organisation visualised by phalloidin-labelling in cells treated with 20 nM of latrunculin A (LatA). Histogram under image shows the distribution of mean RF tilt in the 6−10 μm annulus of cells fixed at 6 h after cell plating. Histogram and mean±SEM value are based on 243 cells. h Average values of RF tilts (mean±SEM) as a function of the distance of annuli from the cell edge for untreated control cells (n = 274), 20 nM LatA-treated cells (n = 243), and 5 nM swinholide A (SwinA)-treated cells (n = 153) at 6 h after cell plating. Colour coding in histograms (b–e) and (g) correspond to those indicated in graphs (f) and (h) respectively. Scale bars, 10 μm (a–e and g). See also Supplementary Movie 3. For statistical analysis, see Supplementary Table 1, lines 15–29.