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. 2021 Mar 23;34(12):108862. doi: 10.1016/j.celrep.2021.108862

Figure 3.

Figure 3

MC4R-derived MAPK activation is dependent on receptor internalization and modulated by β-arrestins

(A and B) HEK293 cells transiently expressing MC4R WT were used to assess the effect of dynasore on maximal efficacy of NDP-αMSH in cAMP production (A) and ERK1/2 phosphorylation (B).

(C and D) NDP-αMSH-dependent ERK1/2 phosphorylation in HEK293 cells treated with the β-arrestin/AP2 inhibitor barbadin (C) and siControl, siARRB1, or siARRB2 (D).

(E) Effect of siControl, siARRB1, or siARRB2 on maximal efficacy of NDP-αMSH-induced cAMP production. Representative immunoblots shown were probed for total ERK1/2, phosphorylated ERK1/2 (p-ERK1/2), Gαs, β-arrestin-1/2, and vinculin as an additional loading control. Data are expressed as percentage of maximal control response (% WT) and plotted as mean ± standard error from 3–14 independent experiments. Statistical significance was determined by two-way ANOVA and Dunnet’s multiple comparison test (*p < 0.05, ∗∗∗p < 0.001, compared to NDP-αMSH-stimulated MC4R WT [control]). See also Figure S4.