Fig. 4

SENP1 interacted with USP28 both in vivo and in vitro. a 293T cells were co-transfected with Flag-USP28 and GFP-SENP1 plasmids for 36 h. Cells were harvested and immunoprecipitation was performed with Flag M2 beads. The input and immunoprecipitates were subjected to western blot with indicated antibodies. b HCT116 cells were lysed and subjected to immunoprecipitation with mouse IgG or anti-USP28 antibody. The input and immunoprecipitates were subjected to western blot with indicated antibodies. c Bacterial purified GST-SENP1 proteins or GST alone were incubated with the purified Flag-USP28 protein for 1 h at 30 °C. Then the reaction was terminated by 2Χ SDS loading buffer and followed by western blot with indicated antibody