Fig. 3

USP14 in the cytoplasm is critical to the nuclear import of AR. a MDA-MB-453 cells were treated with DHT (10 nM) for 24 h. Cytosolic and nuclear protein fractions were prepared, which were then immunoprecipitated with AR antibody beads, and immunoblotted for AR and USP14. HSP90 was used as a cytoplasmic control. Lamin B was used as a nuclear control. b MDA-MB-453 cells stably expressing USP14 shRNA or control shRNA were exposed to DHT (10 nM) for indicated days. Cell viability was detected using MTS assay. Error bars correspond to a 95% CI of three independent experiments. *P < 0.05. c Cytosolic and nuclear protein lysates were extracted, respectively, from MDA-MB-453 cells stably expressing USP14 or control shRNA exposed to DHT (10 nM) for 24 h. Western blot assay was used to detect AR expression. d MDA-MB-453 cells stably expressing USP14 shRNA or control shRNA were exposed to vehicle (Veh) or DHT (10 nM) for 24 h. Immunofluorescence microscopy shows endogenous AR (orange) and nucleus (blue). Scale bars represent 20 μm