Fig. 4

Genetical and pharmacological inhibition of CHK1 impairs UBE2T-induced DDR and radioresistance in HCC. a UBE2T stably overexpressing were transfected with siRNA targeting CHK1 and treated with IR (4 Gy). Cells were prepared for immunofluorescence analysis of γH2AX foci. b γH2AX level of lysates from cells treated as that in panel a was determined by immunoblotting. c Comet assay was conducted in cells with same treatment as that in panel a at indicated timepoints after IR. The quantification of the tail moment is shown. d Colony formation assays were conducted in UBE2T stably overexpressing MHCC-97H cells transduced with lentivirus shRNA targeting CHK1. The representative images and survival curve are shown. e UBE2T stably overexpression MHCC-97H cells were treated with 2 μM MK-8776 1 h before IR, and analyzed for γH2AX foci by immunofluorescence staining. f γH2AX level of lysates from cells treated as that in panel e was determined by immunoblotting. g Comet assay was conducted in cells with same treatment as that in panel e at indicated timepoints after IR. h Colony formation assays were conducted in UBE2T stably overexpressing MHCC-97H cells treated with 2 μM MK-8776. The representative images and survival curve are shown. i Transplanted xenografts were established with UBE2T overexpressing (UBE2T) and vector transduced (Control) MHCC-97H cells, and treated with IR and injected with MK-8776 (50 mg/kg) intraperitoneally for 3 days. Tumor volumes from each group were tracked for 21 days, the representative tumor samples from each group are shown. j Tumor volumes of the xenografts from panel i were measured. k Tumor weights of the removed xenografts from panel i were measured. Data represent the mean ± SD. In (a), (c), (e), and (g), ns, not significant, *P < 0.05 and **P < 0.01, by one-way ANOVA. In (j), **P < 0.01, by two-way ANOVA