Figure 4.

PP1α-AR binding is disrupted by androgen-induced AR N/C interaction. (A) 293T cells transfected with Flag-PP1α and HA-AR full length were incubated overnight in CDS medium without or with indicated doses of androgen, followed by co-IP analysis. (B) 293T cells were transfected with Flag-PP1α and HA-AR. Cells were then incubated overnight in CDS medium with bicalutamide (BIC, 10 μM), enzalutamide (ENZ, 10 μM), DHT (10 nM) and combinations as indicated, followed by co-IP analysis. (C) 293T cells were transfected with Flag-PP1α and AR, followed by incubation overnight in CDS medium without or with DHT in combination with indicated AR antagonists (mifepristone: RU486; cyproterone acetate: CPA; and ARN509: ARN; 10 μM each), followed by co-IP analysis. (D) 293T cells were transfected with Flag-PP1α and AR W741C (bicalutamide-activated mutant) or F876L (enzalutamide-activated mutant) constructs. Cells were then incubated overnight in CDS medium without or with indicated ligands (BIC: 10 μM of bicalutamide and ENZ: 10 μM of enzalutamide), followed by co-IP analysis. (E) 293T cells were transfected with Flag-PP1α and indicated HA-AR constructs. Cells were then incubated overnight in CDS medium without or with DHT (10 nM) for co-IP analysis. (F) 293T cells were co-transfected of Flag-PP1α and pBIND-AR-LBD. Cells were then incubated overnight in CDS medium without or with 10 nM DHT as indicated, followed by co-IP analysis. (G) 293T cells were transfected with Flag-PP1α together with HA-AR wild-type (WT) versus its FQNLF motif mutant (FQNAA). The cells were then incubated overnight in CDS medium, followed by treatment with 10 nM DHT for 30 min or 2 h, followed by co-IP.