Fig. 6

Upregulation of eIF5A1 SUMO2 modification upon stress. A HEK-293 cells were cotransfected with HA–eIF5A1–WT, Ubc9, and His6–SUMO2–K11RK48R or His6–SUMO2–K0. 36 h after transfection cells were treated with DMSO or MG132 for 16 h and whole-protein extracts and Histidine-tagged purified proteins were analyzed by western blot with the indicated antibodies. B HEK-293 cells were cotransfected with HA–eIF5A1–WT, pcDNA, Ubc9, and His6–SUMO2 or Ubc9 and His6–SUMO2–K0. Thirty-six hours after transfection, cells were incubated at 43 °C for 2 h, as indicated, and whole-protein extracts and Histidine-tagged purified proteins were analyzed by western blot with the indicated antibodies. C HEK-293 cells were cotransfected with HA–eIF5A1–WT and pcDNA or Ubc9 and His6–SUMO2–K0. Twenty-four hours after transfection, cells were treated with DMSO or TAK-243, as indicated, and at 12 h after treatment, cells were incubated at 43 °C for 2 h. Whole-protein extracts and Histidine-tagged purified proteins were analyzed by western blot with the indicated antibodies