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. Author manuscript; available in PMC: 2021 Aug 20.
Published in final edited form as: J Cell Physiol. 2018 Dec 19;234(8):13220–13232. doi: 10.1002/jcp.27994

FIGURE 1.

FIGURE 1

BRAF upregulates ERK3 expression. (a) Western blot analysis of ERK3 reduction upon the knockdown of BRAF by siRNA in three different melanoma cell lines: A375, WM3211, and uveal BRAFV600E melanoma OCM3. β-Actin serves as a loading control. Western blots are representative of multiple experiments and quantified. (b) Expression plasmids for wild-type (WT) BRAF or constitutively active BRAFV600E were transiently transfected into melanoma cell lines A375 (BRAFV600E) and WM3211 (BRAF WT) or embryonic kidney noncancer cell line 293T (BRAF WT). Western blot analysis analysis reveals an increase in ERK3 protein concomitant with increased BRAF. (c) RT-qPCR was used to measure ERK3 mRNA levels in three melanoma cell lines A375, OCM3, and WM3211. BRAF was knocked down by siRNA for 2 days. Reduced ERK3 mRNA expression corresponds to the reduction in BRAF. Significance (p values) were determined by Student’s t test and expressed as mean ± standard deviation (n = 3). (d) ERK3 protein level was reduced upon BRAF knockdown using a silencer select siRNA (Ambion) in both A375 (left panel) and OCM3 (right panel). ERK3: extracellular signal-regulated kinase 3; mRNA: messenger RNA; RT-qPCR: quantitative reverse transcription PCR; siRNA: small interfering RNA; WT: wild-type