Fig. 2

Hypoxia and HIF-1α promoted USP28 deSUMOylation. a HCT116 cells stable expressing of His-SUMO2 or vector control was lysed and subjected to western blot with indicated antibodies. b HCT116 cells stable expressing of His-SUMO2 and HCT116 cells were cultured under normoxic (16% O₂) or hypoxic (5% O₂) conditions for 24 h. SUMOylated proteins were pulled down with Ni⁺ beads. The input and SUMOylated proteins were then subjected to western blot with indicated antibodies. c HCT116 cells stable expressing of His-SUMO2 were transfected with vector control or HA-HIF-1α for 36 h, cells were lysed and SUMOylated proteins were pulled down with Ni+ beads and subjected to western blot with indicated antibodies. d HCT116 cells stable expressing of His-SUMO2 were transfected with control shRNA plasmid or shRNA against HIF-1α for 24 h and under hypoxic (5% O₂) conditions for additional 24 h. Cells were harvested and SUMOylated proteins were pulled down with Ni+ beads and subjected to western blot with indicated antibodies