COUP-TF II represses the transactivation of AR.

<p>(A) Dose-dependent inhibition of AR transactivation by COUP-TFs. PPC-1 cells were cotransfected with 350 ng of pPBARE×7-tk-Luc reporter and 50 ng of AR expression plasmid along with increasing concentration (100, 250, and 500 ng) of COUP-TF I or COUP-TF II. Cells were treated with or without 3 nM DHT for 24 h. (B) COUP-TF II-mediated repression of AR transactivation on natural AR-target promoters. PPC-1 cells were transfected as in “A”, with PSA-luc or MMTV-luc reporter. (C) COUP-TF II-mediated repression of endogenous AR transactivation. LNCaP cells were transfected as in “A”, without the AR expression plasmid. (D) Repression of androgen-induced PSA mRNA expression by COUP-TF II. LNCaP cells were infected with AdGFP or AdCOUP-TF II. After 24 h of recovery, the cells were treated with 10 nM DHT, and cultured for another 24 h prior to harvesting. Quantitative RT-PCR analysis was conducted using specific primers for PSA and β-actin. The relative PSA mRNA expression was normalized by β-actin expression. At least three independent experiments were combined and values represent the mean±SEM (A–D). **, P<0.01; ***, P<0.001. (E) Repression of androgen-induced PSA protein expression by COUP-TF II. LNCaP cells were infected and processed as in “D”, and cultured for another 48 h prior to harvesting. Western blot analysis of total proteins was conducted using anti-PSA, anti-AR (sc-815), anti-HA, and anti-α-tubulin. Data are representative of three independent experiments (left). The relative PSA protein expression was quantified by normalizing with tubulin expression (rignt).