RNF4 degrades poly-sumoylated Tax. (A) Endogenous Tax-RNF4 interactions in untreated or AS/IFN-treated HuT-102 cells as detected by Duolink. Nuclei were stained with DAPI (blue). (B) HeLa cells transiently expressing Tax were cotransfected with siCTRL or siRNA against RNF4 (siRNF4). (Upper) After treatment with AS/IFN for up to 48 hours, cell extracts were immune-precipitated with anti-Tax antibodies, followed by western blot with indicated antibodies. (Lower) Tax, RNF4, and actin are shown in WCE. (C) HuT-102 cells were transfected with siCTRL or siRNF4 and treated with AS/IFN for up to 48 hours. Degradation of endogenous Tax was analyzed by western blot. Vinculin serves as a loading control. Quantification in percentages shows the amount of remaining Tax after normalization. (D) HeLa cells transiently expressing Tax were cotransfected with RNF4. Western blot was performed with indicated antibodies 48 hours after transfection.