Figure 5
Tip60 binds to and acetylates cortactin through Fe65 to inhibit cell motility.
(A), Inducible Fe65 stable knockdown clones of 293T cells were transfected with 1.5 μg Flag-cortactin and 1.5 μg of HA-Tip60 and treated with DMSO or 1 μg/mL doxycycline as shown. Cellular extracts were subjected to co-immunoprecipitation analyses with antibodies as indicated. (B), Cells were transfected as in panel A and treated with DMSO, 300 ng/mL TSA and/or 1 μg/mL doxycycline. Cellular extracts were subjected to co-immunoprecipitation analyses with antibodies as indicated. Acetyl-lysine (Ac-K) antibody is an antibody that specifically recognizes lysine-acetylated proteins. (C) and (D), 293T cells were transfected with HA-Tip60 together with control or Fe65 siRNA. 36 h post transfections, 5 × 105 transfected cells were plated into the inserts for trans-well assays. Western blots were performed to show Fe65 knockdown and β-actin blots were included as loading controls (panel C). Cells were fixed and photographed 24 h after plating (panel D). (E) and (F), Quantitative data were presented as bar graphs and data statistics performed as described in Fig. 1.
