FOXM1 may not be an upstream mediator of EZH2 ubiquitination. (A) Expression level of FOXM1 in normal NK and NKTL cell lines. (B) IP showing absence of interaction between EZH2 and FOXM1 in NKYS and NK-S1. (C-D) Quantitative reverse transcription PCR assay in (C) NKYS or (D) NK-S1 cells upon FOXM1 knock-down. Data are shown as mean ± SD, N = 3. *P < .05; **P < .01; ***P < .001. NKTL cells were harvested for RNA extraction 48 hours after transfection. (E) Immunoblots of indicated proteins upon FOXM1 knock-down in NKYS and NK-S1 cells. NKTL cells were harvested for immunoblots 48 hours after transfection. (F) Level of EZH2 ubiquitination upon knock-down of FOXM1 in NKYS cells. MG132 was given at 1 µM for 6 hours of treatment 48 hours after transfection. (G-H) Level of EZH2 ubiquitination upon overexpression of indicated proteins in HEK293T cells. MG132 was given at 10 µM for 6 hours of treatment 48 hours after transfection. (F-H) Densitometry analysis was used to quantify average changes in 3 individual experiments. All immunoblots were performed in at least 3 individual experiments and representative images are shown. SD, standard deviation.