FIG 1.

USP7 interacts with PRC1 and SCML2. (A) Immunoprecipitation of SCML2, BMI1, RING1B, and USP7 was performed with nuclear extracts from HCT116 cells. Pulldown material was analyzed by Western blotting with specific antibodies against USP7 and SCML2. Nonspecific IgG served as a control, and 2% of the input (In) is shown. (B) USP7 immunoprecipitation in nuclear extracts from HCT116 cells. Pulldown material was analyzed by Western blotting with specific antibodies against RING1B, BMI1, KDM2B, EZH2, and histones H2A and H2B. Nonspecific IgG served as a control, and 2% of the input is shown. (C) USP7 immunoprecipitation in either control nuclear extracts (−Benz) or nuclear extracts treated with benzonase (50 U/mg) (+Benz) for 16 h prior to immunoprecipitation. Nonspecific IgG served as a control, and 2% of the input is shown. (D) Schematic representation of the domains in USP7 and the fragments used in the in vitro pulldown experiments. (E) Western blot analysis of the in vitro pulldown of His-SCML2B with GST alone or GST-fused USP7 fragments. (F) Schematic representation of the two human isoforms of SCML2 showing the different domains of the protein and the fragments employed for the in vitro pulldown experiments. (G) Western blot analysis of the in vitro pulldown of His-USP7-N with GST alone or GST fusions of MBT-DUF, MBT-DUF lacking the RBR, or the RBR alone.