Phospho-EZH2 (Thr487) Antibody - #AF7400

Fig. 6. PP7080 was required to promote EZH2 degradation to regulate ANKRD1 in GC cells. (A-B) FISH(A) and subcellular fractionation assays(B) were used to determine the distribution of lncRNA PP7080 in BGC823 and AGS cells. (C) Bioinformatics prediction of the interaction probability of PP7080 and RNA-binding proteins on a random forest (RF) or support vector machine (SVM) basis. (D) Level of PP7080 detected by qRT-PCR after RIP for EZH2 and AGO2 in BGC823 and AGS cells. (E) The expression levels of EZH2 in BGC823 and AGS cells transfected with PP7080 plasmid were detected by qRT-PCR and western blot assays. (F-G) The EZH2 protein expression was detected by western blot assay in transfected BGC823 and AGS cells after treatment with CHX (F) and MG132 (G). (H) Western blotting of endogenous EZH2-associated ubiquitination in transfected BGC823 and AGS cells. (I) Western blotting of EZH2-Thr-345 and EZH2-Thr-487 phosphorylation in transfected BGC823 and AGS cells. (J) The expression levels of ANKRD1 in BGC823 and AGS cells transfected with si-EZH2 were detected by qRT-PCR and western blot assays. (K) ChIP–qPCR analysis of EZH2 occupancy and H3K27me3 binding on the ANKRD1 promoter region after overexpression of PP7080 in BGC823 and AGS cells. IgG was used as a negative control.