FIG. 6.

DU145 cells exhibit TGF-β-dependent activation of Smad signaling. (A) DU145 cells were treated for 24 h with normal growth medium (C), 10 ng of TGF-β1/ml (T), 100 nM rapamycin (R), or 10 ng of TGF-β1/ml plus 100 nM rapamycin (T + R), and immunoblotting experiments on whole-cell extracts were performed using antibodies specific for the indicated proteins. Total extracts from HEK 293 (293) cells were used as a positive control in Rb, p107, and p130 immunoblotting experiments. (B) DU145 cells were treated as for panel A, and (CAGA)₁₂-luciferase assays were performed as described in Materials and Methods. Assays were performed in triplicate, and results are presented as relative luciferase units (RLU) normalized to micrograms of protein assayed. (C) Luciferase assays were performed using an E2F-luciferase reporter construct, and the results are presented as for panel B.