Figure 3

KAI1 increases VHL protein levels. (A) PC3 cells stably transfected with KAI1 (PC3-KAI1 #6) or vector control (PC3-GFP #8) cells were treated with MG132. After 12 hours of treatment, cell lysates were prepared and the levels of CDCP1 were measured by immunoblotting. (B) Cell lysates were prepared from PC3 cells transiently transfected with KAI1 and from stable transfectants. VHL expression was analyzed by immunoprecipitation. Densitometric analysis of VHL was shown after normalization to beta-actin. (C) PC3 cells stably transfected with KAI1 (PC3-KAI1 #5 and PC3-KAI1 #6) or vector control (PC3-GFP #8) were transiently transfected with HA-tagged VHL. Twenty-four hours after transfection, cells were harvested, and the levels of CDCP1 and VHL protein were analyzed by immunoblotting. Densitometric analysis of HA-VHL was shown after normalization to beta-actin. (D) PC3 cells were transfected with HA-tagged VHL, and then treated with siRNA against VHL. The protein levels of CDCP1 and HA-VHL were assessed by immunoblotting.