Fig 2: PMCA4b is upregulated in both BRAF wild-type and BRAF-mutant melanoma cells after histone deacetylase inhibitor (HDACi) treatment. (A) One BRAF wild-type (MEWO) and two BRAF-mutant (A375, A2058) cell lines were treated with HDACi SAHA or valproate in increasing concentrations for 48 h. DMSO (D) treatment was included as vehicle control. The protein level of PMCA4b and PMCA1 proteins were analyzed by Western blotting of total cell lysates (30 µg per sample) with PMCA4b-specific and pan-PMCA antibodies. Western blots were analyzed by densitometry. (B) Change in PMCA protein expression level was investigated after treatment with SAHA (2 µM) or valproate (2 mM) for 10, 24, 48, and 72 h with PMCA4b-specific and pan-PMCA antibodies. Western blots were analyzed by densitometry. Data were normalized to the expression levels of ß-tubulin, and changes in PMCA4b protein level were expressed as fold increase over the untreated controls. Bars represent mean ± SE from three independent experiments. Asterisks indicate significant differences (*p < 0.05) from control, as calculated by ANOVA and Dunnett’s post hoc tests.