Figure 2

RA inhibits ATF-2 phosphorylation in B16 mouse melanoma cells. B16 cells were treated with either vehicle (DMSO) (C-control) or RA. At the end of the indicated incubation times, cells were harvested and protein (40 μg) from each group was analyzed by western blotting using polyclonal anti-phospho-ATF-2 or total ATF-2 antibodies as described in Materials and Methods. The relative amount of phospho- or total ATF-2 protein in each sample was determined by densitometry using GAPDH as an internal control. The data shown are from a representative experiment, which was replicated three additional times with similar results. A. Cells were treated with the indicated concentrations of RA and harvested after a 48-hour incubation. B. Cells were treated for the indicated time periods with 10 μM RA. C. The effect of different treatment times with 10 μM RA on total ATF-2 protein level.