Figure 2.

ERK activation by NDP-MSH in human melanoma cells expressing variant MC1R. A, Clones of HBL cells stably expressing the MC1R forms indicated on top of each blot were stimulated with NDP-MSH (10⁻⁷ m) for the times shown, PMA (0.1 ng/μl, 15 min), or FSK (10⁻⁵ m, 15 min). Cell extracts were blotted for pERK. The experiment was repeated at least three times with similar results. B, Agonist-induced cAMP levels after stimulation with NDP-MSH in HBL cells expressing WT or variant MC1R. Cells were serum deprived for 3 h before agonist challenge for 30 or 180 min. Results are given as mean ± sem (n ≥ 4). C, Dose dependence of NDP-MSH-induced ERK phosphorylation in HBL cells expressing endogenous WT MC1R. Cells were serum deprived and challenged with the indicated concentrations of NDP-MSH for 5 min before estimation of pERK levels by Western blot. A representative blot of three is shown. D, Comparison of the changes in pERK levels (squares, left axis) and cAMP accumulation (triangles, right axis) in HBL cells treated with the indicated concentrations of NDP-MSH. Results are the mean ± sem (n ≥ 3).