Figure 6. The Arp2/3 complex is dispensable for chemotaxis to CSF and CX3CL1, but is required for fibronectin haptotaxis.

A, C, E) Wind rose plots generated from WT or Arpc2−/− macrophages migrating in a CSF gradient (A), CX3CL1 gradient (C) or FN gradient (E). Each plot is from one representative experiment of mixed WT and Arpc2−/− cells. 0°on the rose plot is the direction of the gradient. B, D, F) Forward Migration Index (FMI) of WT and Arpc2−/− macrophages plotted as mean ± 95% CI in CSF (B), CX3CL1 (D), or FN gradients (F). Data tables relate mean ± SEM of cell speed (V), FMI, d/T and total cells tracked (N). Quantitative measurements were pooled from multiple experiments using identical chamber conditions. G) WT and Arpc2−/− macrophages plated on 1 µg/mL collagen stained for αM integrin, vinculin and p-Tyr (4G10), β1 integrin, WAVE2 and F-actin at bound 6 micron Cy5-fibronectin (Cy5-FN) beads after 15 minute incubation. Arrowheads denote bound beads on Arpc2−/− macrophages. Scale = 10 µm. H) Localization of β1 integrin, p-Tyr and F-actin to Cy5-FN beads 5, 15 or 30 minutes after bead binding by WT or Arpc2−/− macrophages plotted as mean integrated pixel density (A.U.) ± SEM. **p = 0.0011, ***p < 0.0001. I) Schematic depiction of integrin-Arp2/3 complex coordination. Upon ligation integrin initiates an outside-in signal (black arrows) that induces the Arp2/3 complex to nucleate a branched actin network (an inside-out response). The branched actin network then reinforces the ‘upstream’ pathway (red arrows), perhaps by coordinating or concentrating these factors at new sites of integrin engagement. Without the Arp2/3 complex, macrophages still produce an outside-in signal but lack the ability to reinforce the initial signal, leading to loss of ECM sensing and inhibition of CR3 phagocytosis. See also Figure S6, Movies S4 and S5.