Figure 4.

Upregulated RNF44 expression is related to hyperactivation of ERK and AKT in BR cells. (A) Immunoblotting demonstrated that BR and BMR cells possessed downregulated AMPK‐α1 and upregulated RNF44. (B) RNA levels of RNF44 in parental and BR cell lines were determined by qRT‐PCR and normalized by GAPDH. (C) Higher protein levels of p‐AKT, p‐ERK, and RNF44 appeared in BR cells even in the presence of BRAFi (vemurafenib). (D) BR cell lines were treated with MEKi (trametinib, 5 nm), ERKi (SCH772984, 3 μm), or AKTi (MK‐2206, 2 μm) for 24 h, and their RNF44 levels were determined by qRT‐PCR. Data shown in a bar graph were represented as mean ± SEM (n = 3, *P < 0.05, **P < 0.01, and ***P < 0.005). (E) Protein levels of ERK, AKT, AMPK‐α1, and RNF44 in BR and BMR cells were determined by immunoblotting following treatment with AKTi or ERKi.