Role of Heparan Sulfate 2-O-Sulfotransferase in Prostate Cancer Cell Proliferation, Invasion, and Growth Factor Signaling
Figure 5
HIF1α activates 2OST expression in model of prostate cancer progression. (a) Schematic of 2OST promoter with regions of HREs (H1 and H2). (b) C4-2B cells were transfected with vector expressing stHIF1α. Real-time PCR for 2OST levels normalized to 18S levels in each sample. Black bars represent control vector alone transfected samples and gray bars represent stHIF1α transfected samples. Error bars indicate standard deviation. (c) Western blot for HIF1α shows increased accumulation of stabilized transcription factor in cells transfected with stHIF1α vector. (d) Inhibition of endogenous HIF1α expression by siRNA results in decreased levels of 2OST mRNA. Real-time PCR analysis of HIF1α and 2OST normalized to levels of 18S. Black bars represent samples treated with scrambled siRNA-treated samples, white bars represent levels of either HIF1α or 2OST in HIF1α siRNA-treated cells. Error bars indicate standard deviation. Asterisk indicates . (e) HIF1α binds directly to the 2OST promoter at predicted HREs. Chromatin Immunoprecipitation analysis of C4-2B total chromatin, HIF1α IP, no antibody (NoAB), and Mock samples. Samples were analyzed by PCR with primers flanking each HRE site. Positive control primers were used from the VEGF promoter [33].
