Fig. 7. AF6i3 deletion mutants in wound healing assay: a crucial role of the F-actin-binding site. (A) Schematic representation of AF6i1, AF6i3 and AF6i3 deletion mutants, AF6i3RA1 and AF6i3PDZ (top). Expression levels of shRNA-resistant AF6 constructs stably expressed in AF6i3-knockdown cells (bottom). (B) Graphic representation of wound closure 13 hours after wounding of AF6i3-knockdown cell monolayers reconstituted with mutant constructs as indicated. Values for ±s.d. were derived from three independent experiments. (C) Light microscopy pictures of control and AF6i3-knockdown cells without or with reconstitution by AF6i1 and AF6i3 immediately after wounding (0 h) and 13 hours later. Bar, 0.5 mm. (D) Immunofluorescence pictures of confluent cell monolayers of cells shown in C. Bar, 20 µm. (E) Western blot analysis of detergent-soluble supernatant (S) and the insoluble residue (R) of AF6i3-knockdown cell monolayers reconstituted with AF6i1 or AF6i3.