Figure 5
KAI1 decreases VEGF expression. (A) PC3 cells were cotransfected with a VEGF promoter (2.2 kb)-luciferase reporter plasmid and HIF-1α and GFP-KAI1 expression plasmids. Forty-eight hours after transfection, cell lysates were analyzed for luciferase expression. (B) PC3 cells stably transfected with KAI1 (PC3-KAI1 #5 and PC3-KAI1 #6) or vector control (PC3-GFP #8) were cotransfected with a VEGF promoter-luciferase reporter plasmid and a HIF-1α expression vector. Forty-eight hours after transfection, cell lysates were analyzed for luciferase activity. (C) PC3 cells were cotransfected with a VEGF promoter-luciferase reporter plasmid and a GFP-KAI1 expression plasmid. Twenty-four hours after transfection, cells were treated with 100 μM CoCl2 for 24 hours, and then luciferase activity was measured. (D) Total RNA from PC3 cells stably transfected with KAI1 (PC3-KAI1 #5 and PC3-KAI1 #6) or vector control (PC3-GFP #8) was isolated, and VEGF mRNA expression was analyzed by RT-PCR. (E) After treatment of si-CDCP1, PC3 cells were harvested and VEGF mRNA expression was analyzed by RT-PCR. (F). PC3 cells stably transfected with KAI1 (#6) or vector control (PC3-GFP #8) were exposed to CoCl2 (100 μM) for 24 hours. Total RNA was extracted and mRNA levels were quantified by RT-PCR.