Fig. 4: ZFP82 stabilizes p53 by inhibiting HDAC3–p53 interactions.

A Upper panel: Immuno-coprecipitation suggests ZFP82 interacts with HDAC3 and disrupted HDAC3-p53 interaction in a time depended manner, in the presence of chemotherapy reagent 5-FU (10 μg/mL). Lower panel: In the presence of 5-FU (10 μg/mL), ZFP82 induces HDAC3 cleavage, and thus promotes p53 acetylation, down-stream target of p53 signaling PUMA and BAX is also activated. B After demethylation treatment of 5-Azacytidine (Aza), the restored expression of ZFP82 also induced p53 acetylation, HDAC3 cleavage, activates its downstream targets Puma and Bax, and promotes caspase-3 apoptosis pathway. C Flow Cytometry demonstrates in the presence of 5-FU (10 μg/mL), co-overexpression of HDAC3 + ZFP82 leads to partially restored apoptosis function, while overexpression of ZFP82 along significantly enhances the 5-FU induced apoptosis (HDAC3 vs HDAC3 + ZFP82 vs ZFP82: 9.32% vs 18% vs 26.52%, p < 0.05). right panel shows the statistic histogram of the apoptosis results (**, p < 0.01). D Chromatin Immunoprecipitation (ChIP) analyses against the p53 promoter. Compared to vector, whether at the presence of 5-FU or not, ectopic expression of ZFP82 decreased HDAC3-p53 complex on p53 promoter (**, p < 0.01). E ZFP82 increases p53-p300 complex to the promoter region of Bax. Chromatin Immunoprecipitation (ChIP) analyses against the p53 binding sites (p53-RE) of BAX promotor shows, compared to vector, whether at the presence of 5-Fu or not, the re-expressing of ZFP82 enhances 5-FU-induced recruitment of the p53-p300 complex to the p53-RE of Bax promotor (***, p < 0.001). F ChIP and re-ChIP assays showed that ZFP82 overexpression enhanced the efficiency of 5-FU–induced binding of acetylated p53 and p300 to the p53 response element of Bax, while reducing the binding of HDAC3 to p53. (***, p < 0.001).