FIGURE 4.

A similar motif functions in AR. AR contains an FQNLF motif similar to that of Htt. This motif was mutated by alanine substitution to AQNAA (3A) in ARN127YFP and full-length AR-YFP proteins. These proteins were transiently transfected in HeLa cells, followed by Western blot or direct fluorescence imaging. A, the 3A mutation increases the steady state of both ARN127YFP and AR-YFP. B, 6 h of treatment of cells expressing ARN127YFP with 10 μm proteasome inhibitors (MG132, ALLN, and lactacystin) versus vehicle control (dimethyl sulfoxide (DMSO)) increases the steady state levels of wt protein but not the 3A mutant. Lactacystin (lact) titration also affects the steady state levels of wt ARN127YFP protein but not the 3A mutant. C, full-length AR-YFP localizes to the nucleus and to cytoplasm puncta in the presence of 10 nm DHT. The DHT-induced AR puncta have various appearances depicted in three panels: small versus large, single versus multiple. AR(3A)-YFP never localizes to these puncta and is localized exclusively in the cell nucleus after DHT treatment. All of the gels are representative of at least three experiments. norm, normalized band intensity/actin. Scale bars, 10 μm.