Fig. 3

β-TrCP regulates the stability and ubiquitination of Dlg5. a Western blot analysis of SMMC-7721 cells transfected with indicated doses of Flag-β-TrCP. b Western blot analysis of SMMC-7721 cells transfected with siRNAs against control or β-TrCP. c SMMC-7721 cells transfected with siRNAs against control or β-TrCP for 24 h, then 20 μg/ml cycloheximide (CHX) was added for the indicated time course. Cell extracts were subjected to western blot with the indicated antibodies. d SMMC-7721 cells transfected with siRNAs against control or β-TrCP for 36 h, cell lysate was subjected to immunoprecipitation by anti-Dlg5 antibody. Immunoprecipitates was detected by western blot using indicated antibodies. e Alignment of amino acids corresponding to the DSGxxxE sequence with Dlg5 orthologs. f 293T cells were co-transfected Flag-β-TrCP with HA-Dlg5 WT or HA-Dlg5 S730A for 36 h, cell lysate was subjected to immunoprecipitation by anti-Flag antibody. Immunoprecipitates was detected by western blot using indicated antibodies. g 293T cells transfected with indicated plasmids for 36 h, cell lysate was subjected to western blot detection. h 293T cells were co-transfected Flag-β-TrCP and his-K48-ub with HA-Dlg5 WT or HA-Dlg5 S730A for 36 h, cell lysate was subjected to immunoprecipitation by Ni+ purification. Immunoprecipitate was detected by western blot using indicated antibodies