Figure 2. MUC1-C suppresses AR expression by a miR-135b-mediated mechanism.
A. LNCaP/GFP and LNCaP/MUC1-C cells were transfected with the empty pGL3 vector or 0.1 µg (open bars), 0.2 µg (shaded bars) and 0.5 µg (solid bars) pAR-Luc. As an internal control, the cells were also transfected with the SV-40-Renilla-Luc plasmid. Luciferase activity was measured at 48 h after transfection. The results are expressed as relative luciferase activity (mean±SD from three separate experiments) compared to that obtained from cells transfected with the empty pGL3 vector (assigned a value of 1). B. Total RNA from the indicated LNCaP cells was analyzed for miR-135b and miR-147 levels. U6 expression was included as a control. C and D. LNCaP cells were transfected with a control scrambled siRNA for 48 h or miR-135b for the indicated times. Total RNA was analyzed for AR and GAPDH mRNA levels by RT-PCR (C). Lysates were immunoblotted with the indicated antibodies (D).