Fig. 2

UBE2O ubiquitinates PTRF. A UBE2O-Myc-cs expression plasmid is a UBE2O mutation expression plasmid. UBE2O expressed by the UBE2O-Myc-cs expression plasmid could not bind to ubiquitin effectively. Flag-PTRF, His-HA-ubiquitin and UBE2O-Myc or UBE2O-Myc-cs expression plasmids were co-transfected into HEK293T cells. After cells were collected, His-HA-ubiquitin in cell lysates was immunoprecipitated with Ni–NTA superflow. The immunoprecipitates and the total cell lysates were immunoblotted for the indicated antibodies. B Flag-PTRF, UBE2O-Myc and His-HA-ubiquitin-KO or His-HA-ubiquitin wild type vectors were co-transfected into HEK293T cells. Cells were harvested after 40 h transfection and were analyzed by His immunoprecipitation ubiquitination assays and western blot