The Function of Hypoxia-Inducible Factor (HIF) Is Independent of the Endoplasmic Reticulum Protein OS-9
Figure 2
OS-9 shows no effect on regulation of HIF-1α.
Total cell lysates were used for SDS-PAGE and subsequent Western blotting. To generate nearly anoxic conditions, cells were exposed to an oxygen consuming chemical system or to 1% or 3% O2 for 4 h to generate hypoxia. (A) U2OS, HeLa and Hep3B cells were transiently transfected with the plasmid pOS-9-V5 48 h prior to the experiment. Lamin A and actin were used as loading controls. (B) U2OS cells were subjected to ER stress by incubation either with tunicamycin (1 µg/ml) or thapsigargin (0.5 µg/ml) for 20 h. To detect HIF-1α under normoxia, cells were treated with DMOG (1 mM) for 4 h. A sample of DMSO-only treated cells was loaded to exclude unspecific side effects of the solvent. (C) U2OS cells were transduced with lentiviral construct pLKO.1-shRNA-OS-9 (shOS-9) mediating a stable knockdown of OS-9 expression. Control cells (c) were transduced with plasmid pLKO.1-puro. Representative Western blots are shown for each subfigure.