Figure 6.

S109 is ineffective in cells expressing Cys528 mutant CRM1. (A and B) HCT-15 cells were transiently transfected with plasmids coding for Flag-tagged wild or C528S mutant CRM1 as indicated. After treatment with 2 μM S109 for 2 h, the cells were fixed and stained with antibodies against RanBP1 and the Flag tag. (C) HCT-15 cells that stably expressed wild type or C528S mutant CRM1 were seeded in 96-well plates and incubated with S109 at the indicated concentrations for 72 h. Growth inhibition was analyzed with the CCK-8 assay. (D) HCT-15 cells stably expressing wild type or C528S mutant CRM1 were treated with S109 at the indicated concentrations for 12 h, and then subjected to a Western blot analyses using anti-Flag antibody