Fig. 5: Directional actin flow is accompanied by a local polymerization–depolymerization gradient.

a–e, Photoconversion of mEos2-actin to measure actin polymerization and depolymerization dynamics. a, Left: actin structure before photoconversion (in green) in a square region of interest (yellow dashed line); the edge of the fibronectin pattern is marked by a white dashed line. Right: actin structure immediately after photoconversion (in magenta). Scale bars, 5 μm. b, Kymograph along the white arrow in a of the green channel shows hotspots of actin polymerization. c, Kymograph along the white arrow in a of the red channel shows actin depolymerization. Horizontal scale bar, 1 μm, vertical scale bar, 10 s. d, Actin polymerization measured from photoconversion experiments (n = 47). Error bars indicate s.e.m. e, Actin depolymerization measured from photoconversion experiments (n = 47). Error bars indicate s.e.m. f, mEmerald-arp-p34 in tranfected cells shows Arp2/3 localization. Scale bar, 2 μm. g, Cofilin staining shows depolymerization activity in the front. Scale bar, 2 μm. For f and g, the white dashed line marks the edge of the fibronectin pattern and the green dashed line marks the edge of the cell. h, Polymerization–depolymerization gradient at negative curvature. n = 8 for Arp2/3 and n = 3 for cofilin. Error bars indicate s.e.m., i, GFP-FAK localizes on RFs. The magenta dashed line marks the edge of the fibronectin pattern and the green dashed line marks the edge of the cell. Scale bar, 2 μm. j, SIM-TIRF imaging of GFP-FAK near the substrate at the edge of the adhesive pattern (magenta dashed line) shows mobilization of particles of GFP-FAK on radial actin fibres. Scale bar, 2 μm. k, Kymograph along the yellow dashed line in j; the magenta dashed line marks the edge of the fibronectin (FN) pattern. GFP-FAK particles engage in continuous flow (filled arrowhead) in the non-adherent region on the left side whereas most particles stay immobile (open arrowhead) in the adherent region on the right side. Horizontal scale bar, 1 μm; vertical scale bar, 1 min. l, Schematic showing the polarized organization of the actin cytoskeleton at negative curvature.