Fig. 2: NDRG3 inhibits the cap-dependent translation of HIF-1α.

a PC3 and DU145 cells, which had been transfected with NDRG3-silencing siRNA, were incubated under hypoxia for 24 h and subjected to RT-qPCR to measure the mRNA levels of HIF-1α and HIF-2α (means + s.d., n = 3). ns denotes “not significantly different.” b The transfected PC3 or DU145 cells were incubated under hypoxia for 8 h, followed by re-oxygenation at 21% O₂. HIF-1α and HIF-2α proteins were immunoblotted and their blots were quantified using the ImageJ program. Relative band intensities were normalized to the corresponding β-tubulin intensities. Each symbol represents the mean ± s.d. (n = 3), and the linear regression was plotted using the SigmaPlot program. c The transfected PC3 or DU145 cells were incubated with 15 μM MG132 for the indicated times. Western blot intensities were analyzed using ImageJ, and the relative band intensities were normalized to the corresponding β-tubulin intensities. Each symbol represents the mean ± s.d. (n = 3). * denotes P < 0.05 versus the siCtrl group at the corresponding time. d PC3 or DU145 cells were co-transfected with the TK-HIF1A 5′UTR-luciferase plasmid, the CMV-β-galactosidase plasmid, and siCtrl or siNDRG3. After stabilization for 24 h, the transfected cells were incubated under normoxia or hypoxia for 24 h. Luciferase activities were measured and normalized to β-galactosidase activities. Each bar represents the mean + s.d. (n = 3) and * denotes P < 0.05 between the indicated groups