Fig. 6

AC092894.1 promotes de-ubiquitination of AR by recruitment of USP3. A Schematic diagram of RNA pull-down experiment. B Western blotting to detect AR expression after RNA pull down in LoVo-OxR cells. C RT-qPCR detection of AC092894.1 expression after RIP experiments. D RT-qPCR detection of AR mRNA expression in LoVo-OxR (AC092894.1), HCT116-OxR (AC092894.1), and their corresponding control vector cells. E Venn diagrams represent potential AR de-ubiquitinated proteins screened in the RNA-pull down-MS and Ubibrowser database. F Western blotting to detect the expression of USP3 in LoVo-OxR cells after RNA pull down. G RT-qPCR to detect the expression of AC092894.1 after RIP experiments. H Western blotting to assess USP3 expression. I The interaction of USP3 and AR was analyzed by co-immunoprecipitation after transfection of Flag-USP3 into LoVo-OxR (AC098894.1) and its control vector cells. J For USP3 knockdown LoVo-OxR cells and control vector cells, degradation of AR was detected by Western blotting after treatment with MG132 for 8 h followed by treatment with CHX for the indicated times. K His-ub and AR plasmids were co-transfected into LoVo-OxR (AC098894.1) and its control vector cells, and the AR de-ubiquitination was analyzed by immunoblotting with ub antibody