Fig. 5: ZNF768 interacts with and represses p53 transcriptional activity.

A U87 cells expressing a doxycycline-inducible shRNA targeting ZNF768 were treated or not with doxycycline (20 ng/µl) for 48 h. Cells were next treated with doxorubicin (0.5 µM). Gene expression was measured by RT-qPCR in at least three biological replicates per condition (n =  3/condition). Results are presented as fold change over controls (doxorubicin time 0). B Proteins were extracted from the cells described in (A) and western blots were performed. C U87 were transduced to overexpress Flag-ZNF768. Cells were treated for 24 h with doxorubicin (0.5 µM) and western blots were performed. D Venn diagram presenting the results of four independents rounds of immunoprecipitation (IP). 293T cells were transfected with V5-ACTA or V5-ZNF768 and V5-IPs were performed. Immunoprecipitates were analyzed by mass spectrometry. E 293T cells were transfected and V5 proteins were immunoprecipitated and western blots were performed. F 293T cells were transfected as described in (E) and endogenous p53 was immunoprecipitated and western blots were performed. G 293T cells were transfected with the indicated vectors. V5 proteins were immunoprecipitated and western blots were performed. H, I 293T cells were transfected with V5-ZNF768, V5-ZNF768ΔZincFinger (ZF)1 (missing amino acids 400 to 540), V5-ZNF768ΔZF2 (missing amino acids 252–540), or V5-ZNF768ΔCTD (missing amino acids 1–162). V5 proteins were immunoprecipitated and western blots were performed. J HCT116 cells were transduced to overexpress Flag-ZNF768 or V5-ZNF768ΔZF2. Cells were treated for 24 h with doxorubicin (0.5 µM). Proteins were extracted and western blots were performed. K RT-qPCR were performed from cells treated as described in (J) in at least four independent biological samples per condition (n = 4). In all panels, data represent the mean ± SEM. In panel A, significance was determined by two-way ANOVA with Tukey’s multiple-comparisons test. In panel K, significance was determined by one-way ANOVA with Tukey’s multiple-comparisons test. Details about reproducibility are provided in the Statistics and reproducibility included in the Methods section. Source data are provided as a Source data file.