Fig. 8.

MC1R polymorphisms can decrease MAPK activity induced by αMSH and/or BMS-470539 relative to the wild-type control. Forty-eight hours after transfection of MC1R encoding cDNA, HEK293 cells were stimulated with the indicated concentrations of either αMSH or BMS-470539 for 5 or 20 min, or cells were not treated with ligand (0 min). After stimulation, cells were lysed, and levels of phospho- and total ERK were measured by Western blot analysis as described under Materials and Methods. A, a representative Western blot illustrates the concentration- and time-dependent effects of αMSH on phospho-ERK levels in cells expressing wild-type versus variant receptors. B, quantitation of Western blot signals at the 5-min point. Data are expressed as a percentage of the αMSH-induced effect at the corresponding ligand concentration in cells expressing the wild-type MC1R. Bars represent the mean ± S.E.M from at least three independent experiments. Comparison of ligand effects at a given concentration in cells expressing wild-type versus variant receptors: *, p < 0.05; **, p < 0.01; ***, p < 0.001. C, a representative Western blot illustrates the concentration- and time-dependent effects of BMS-470539 on phospho-ERK levels in cells expressing wild-type versus variant receptors. D, quantitation of Western blot data at the 5-min point. Data are expressed as a percentage of the BMS-470539 induced effect at the corresponding ligand concentration in cells expressing the wild-type MC1R. Bars represent the mean ± S.E.M from at least three independent experiments. Comparison of ligand effects at a given concentration in cells expressing wild-type versus variant receptors: *, p < 0.05; **, p < 0.01.