Figure 2

Another High SKI expression in melanoma cells does not prevent efficient transcriptional responses to TGF-β. A. 1205Lu melanoma cells were incubated for 60 min. with increasing concentrations of TGF-β. Cell extracts were then prepared and subjected to Western analysis for SKI and P-SMAD3 content. B. 1205Lu melanoma cells were transfected in triplicate dishes with (CAGA)₉-MLP-luc and pRL-TK. Four hours later, TGF-β was added at the indicated concentrations and reporter activity was measured 16 h later. Results are mean+/- SD from one representative experiment. C. Three distinct human melanoma cell lines (501mel, 1205Lu and WM852) were incubated with TGF-β (10 ng/ml) for various time periods. Cell extracts were prepared and subjected to Western analysis to determine SKI content. Actin was used as an internal control. D. 1205Lu cells were incubated for 1 h in the absence (DMSO) or presence (SB) of the TβRI inhibitor SB431542 (5 μM) prior to addition of TGF-β (10 ng/ml). Protein extracts were prepared 60 min later and subjected to Western analysis for SKI content.