Figure 2.

Overexpression of cell-surface NEP inhibits FAK phosphorylation and cell migration. (a) TSU-Pr1–derived cell lines containing wild-type NEP (WT-5), control empty vector (TN-12), or mutated, enzymatically inactive NEP (M-22) were cultured with (+) and without (–) 1 μg tetracycline (Tet). Total cell lysates (20 μg) were analyzed for NEP protein by Western blot as described in Methods using the anti-NEP antibody 5B5. (b) WT-5, TN-12, and M-22 cells were cultured with (+) and without (–) 1 μg tetracycline (with the addition of 100 nM CGS24592 in lanes 2 and 4). Cells were lysed and analyzed as described in Figure 1b. (c) Cell migration assays were performed using the identical conditions described in b. Bars represent SD. Experiments were repeated twice with similar results.