Fig. 5: Paclitaxel upregulates GBP2, and GBP2 is a critical mediator for paclitaxel-induced cytotoxic effect on CML cells.

a The changes in protein levels of GBP2, BCL-2 members, and AKT following the treatments with paclitaxel (1 μM) or imatinib (1 μM) for 24 h were evaluated by western blot analyses. Representative blots (left panel) and quantitative data (right panel) from three independent experiments are shown. Asterisks indicate statistically significant values (*P < 0.05). b The changes in the relative association of MCL-1 with GBP2 or BAK were assessed by immunoprecipitation followed by immune blot analyses of K562 cells treated with or without paclitaxel (1 µM) for 24 h. Representative (left) and quantified amounts (right) of GBP2 or BAK pulled downed by MCL-1 immunoprecipitants are presented from three independent experiments (*P < 0.05). c, d The effects of GBP2 silencing on paclitaxel- (c) or imatinib- (d) induced death of K562 cells were examined. Data presented are mean ± SEM of three independent experiments performed in triplicate (**P < 0.01; ***P < 0.001). e The protein level changes induced by paclitaxel (1 µM) were examined in GBP2 KO K562 cells by western blot analyses. Representative blots (left panel) and quantitative data (mean ± SEM; right panel) from three independent experiments are presented. Different letters indicate statistically significant differences (P < 0.05). f The changes in cell death activity induced by indicated concentrations of paclitaxel for 24 h were examined in GBP2 KO cell lines. In addition, the effects by knocking-in GBP2 were assessed by transfecting GBP2 KO K562 cells (1 × 10⁴) with empty vector or GBP2 expression plasmid (30 ng) for 24 h. Data were represented as mean ± SEM of three independent experiments performed in triplicate (**P < 0.01; ***P < 0.001). Cell lysates were subjected to western blot to confirm the expression of GBP2. Representative blots are shown, and GAPDH was used as a loading control. g, h The roles of GBP2 in paclitaxel-induced apoptosis (g) and cell cycle progression (h) in WT K562 cells and two independent GBP2 KO K562 cell lines were assessed using flow cytometry. Cells were exposed to DMSO (control) or paclitaxel (1 µM) for 24 h. The percentages of cells in the sub-G1 phase (g) and the G0/G1, S, and G2/M phases (h) are presented. Data are shown as mean ± SEM of three independent datasets. Different letters indicate statistically significant differences (P < 0.001).