Fig. 6.

Stimulation of HIF-1α transcriptional activity and augmentation of PGK1 expression by EGLN1 siRNA in VHL-deficient cells. A, C2 cells were transfected with C-ODD to analyze the effect of EGLN1 siRNA on HIF-1α N-terminal transcriptional activity. Mock siRNA transfection, together with siRNAs targeting the respective firefly luciferase gene (luc), EGLN2, and EGLN3, served as controls. Relative luciferase units (RLU) of Gal4-luc in y-axis were plotted as means ± SD from three independent experiments in duplicate. *p < 0.01, **p < 0.001. B, PGK1 mRNA levels in C2 cells were determined by real-time RT-PCR after transfection with the indicated siRNAs. PGK1 mRNA levels in reference to those of endogenous ACTB are presented in means ± SE. C, Western blot analyses were performed to examine the specific effect of each siRNA as indicated. Endogenous β-actin was used as a loading control. None of the siRNAs increased HIF-1α protein levels under either normoxic or hypoxic conditions. Arrowhead denotes nonspecific detection.