Fig. 4

C2CD4A decreased half-life of p53 and promoted ubiquitin degradation of p53 by enhancing the MDM2-p53 interaction a C2CD4A-knochdown HCT116 p53+/+ cells were treated with DMSO and MG132. The treatment of DMSO served as a control. 293 T cells, with stable overexpressed C2CD4A, were treated with MG132. The level of C2CD4A and p53 expression were detected using western blot. b, c C2CD4A-knochdown HCT116 p53^+/+ cells (b), C2CD4A-overexpression (c) and control HCT116 p53^+/+ cells were transfected with CHX and harvested at indicated points after CHX treatment, respectively. The level of C2CD4A and p53 expression were detected using western blot. d 293 T cells were co-transfected with Flag-C2CD4A, HA-p53, His-ubiquitin or vector. His was immunoprecipitated using anti-His antibody, and p53 ubiquitination was detected using anti-HA antibody (left). 293 T cells were co-transfected with His-ubiquitin, sh-C2CD4A-1, sh-C2CD4A-2, HA-p53 or vector. His was immunoprecipitated using anti-His antibody, and P53 ubiquitination was detected using anti-HA antibody (right). co-immunoprecipitation revealed the polyubiquitination of HA-p53. Co-IP assays revealed the polyubiquitination of HA-p53. e 293 T and HCT116 cells were co-transfected with the indicated plasmids, and Co-IP assays were performed using anti-GFP antibody to detect the interaction between p53 and MDM2. f The p53, MDM2, C2CD4A expressions were detected in the tranfected 293 T cells using western blot