Figure 5. FAK regulates Rac1-dependent cortactin recruitment and lamellipodial stability.

LacZ- or Cre-infected fak^flox/flox SMC were treated with 20 ng/ml PDGF-BB and immediately imaged at 5 second intervals for 60 min. (A) Representative images of cells 20 min following PDGF treatment. Cell polarization as shown by formation of leading edge (arrowhead) and trailing edge (arrow) was observed in control but not in FAK-deficient SMC. (B) Representative kymographs of LacZ and Cre-infected cells 0-20 min following PDGF-BB treatment. Ascending slopes represent protrusion (arrows) and descending slopes represent retraction (arrowheads). (C-E) Kymographic analysis of lamellipodial dynamics following PDGF treatment. Calculated protrusion rate (C), lamellipodial persistence (D) and leading edge displacement (E) from 8-10 cells from 4 independent experiments (mean±SEM; *p < .05.) (F) Average 2-D cell speed within 4 hr following PDGF treatment. Results are mean±SEM of 30-60 cells from 4 independent experiments. *p < .05. (G-I) LacZ or Cre infected fak^flox/flox cells stained for Rac1 showing dorsal ruffle localization in both control (G, top left) and FAK-deficient (G, top right) cells at 2.5 min following PDGF treatment. At 15 min, leading edge localization of Rac1 (arrows) was observed in control cells, whereas FAK-deficient cells showed un-polarized Rac1 localization (arrowheads). (H, I) Quantification of cells exhibiting Rac1-labeled dorsal ruffles 2.5 min post PDGF treatment (H) and Rac1-labeled dominant leading edge lamellipodia 15 min post PDGF treatment (I). Results are mean±SEM of 150-200 cells from 3 independent experiments. ***p< .001. (J-L) SMC treated as above were stained with anti-cortactin antibodies and processed as described in G-I. Similar dorsal ruffle but not leading edge recruitment of cortactin was observed in FAK-deficient cells. Results are mean±SEM of 200-250 cells from 4 independent experiments. **p< .01. Scale bar = 10μm.