Fig. 4
Cytosolic death effector ___domain-containing DNA-binding protein (DEDD) promotes retinoblastoma (Rb)/p107 proteasome degradation. a Western blots show Rb protein expression levels 6 h after abolishing synchronization by fetal bovine serum (FBS) in DEDD knockdown triple-negative breast cancer (TNBC) cell lines (HCC1806, HCC38, Rb intact). b Quantitative PCR showing both DEDD and Rb1 gene expression in DEDD-knockdown TNBC cell lines. c Western blots showing Rb protein expression in DEDD-knockdown HCC1806 cells under the MG-132 (proteasome inhibitor) treatment for indicated hours. d Western blots showing Rb protein expression levels in the 293FT cells treated with different dosages of 3×-FLAG-WT-DEDD- or 3×-FLAG-ΔNLS-DEDD-expressing plasmids with or without MG-132 treatment. e Western blots showing Rb protein expression levels in KTB-21 cells with or without the overexpression of cytosolic DEDD under the treatment of MG-132. f Immunoprecipitation assay showing the interaction between cytosolic DEDD and Rb protein after overexpressing 3×-FLAG-ΔNLS-DEDD in 293FT cells. g Immunoprecipitation assay showing Rb protein ubiquitination level after co-transfection of HA-ubiquitin and 3×-FLAG-ΔNLS-DEDD in 293FT cells. h Western blots showing Rb protein expression levels 6 h after abolishing synchronization by FBS in DEDD-knockdown TNBC cell lines (MDA-MB-468, Rb deficient). i Western blots showing Rb protein expression in DEDD-knockdown MDA-MB-468 cells under MG-132 treatment for indicated hours. All quantitative data were generated from a minimum of three replicates. P values were derived from one-way analysis of variance (ANOVA) with multiple comparison test. Error bars represent means ± s.e.m.
