Fig. 4: PSMD14 promotes estrogen signaling in breast cancer.

A–D qRT-PCR and immunoblot analysis showing PSMD14 depletion decreases ERα protein stability but not ERα mRNA expression. MCF-7 and T47D cells were transfected with 50 nM siControl or 50 nM PSMD14. Cell lysates were immunoblotted with the indicated antibodies. β-Actin was used as internal control. E, F Immunoblot analysis showing MCF-7 and T47D cells in charcoal-stripped FBS and phenol red-free DMEM were transiently transfected with 50 nM siControl or 50 nM PSMD14 and then treated with 10 nM estradiol or vehicle for 6 h. Cell lysates were immunoblotted with the indicated antibodies. β-Actin was used as internal control. Luciferase assays showing PSMD14 depletion affects ERE-luciferase activity in MCF-7 (G) and T47D (H) cells. I, J qRT-PCR analysis of ERα target genes (GREB1, TFF1, IL20) expression in MCF-7 and T47D cells in charcoal-stripped FBS and phenol red-free DMEM were transfected with PSMD14 siRNA or Negative control for 48 h. Then treated with either ethanol or 10 nM estradiol for 6 h. Total RNA was extracted for gene expression analysis. Each group was analyzed in triplicate. *P < 0.05; **P < 0.01; ***P < 0.001 for target gene expression comparison. K Immunoblot analysis showing PSMD14 depletion increases ERα protein stability. MCF-7 cells were transfected with 1 μg Myc vector or Myc-PSMD14, cell lysates were immunoblotted with the indicated antibodies. β-Actin was used as internal control. L qRT-PCR analysis of ERα target genes (GREB1, TFF1, IL20) expression in MCF-7 showed that overexpression of PSMD14 increased the expression of ERα target genes (GREB1, TFF1, IL20). M Luciferase assays showing PSMD14 overexpression affects ERE-luciferase activity in MCF-7 cells. Data are shown as mean ± SD, N = 3. *P < 0.05; **P < 0.01; ***P < 0.001.