Fig. 5

CDK5 induces apoptosis through direct binding to DP1 and activation of E2F1 signaling. (A) GSEA was performed in the TCGA and a panel of Gene Expression Omnibus (GEO) cohorts using 50 hallmark gene sets from MSigDB. Heatmap showing the NES for each gene set. (B) Western blots (left) of CDK5, E2F1, DP1, apoptotic protease activating factor 1 (APAF1), and p73 in AGS cells with CDK5 overexpression or MKN45 cells with CDK5 knockdown. qRT–PCR analysis (right) of E2F1 and DP1 in AGS cells with CDK5 overexpression or MKN45 cells with CDK5 knockdown. Ns indicates no significance. (C) Representative immunofluorescence images (scale bars, 50 μm) of CDK5 (red), DP1 (green) and 4′,6-diamidino-2-phenylindole (DAPI; blue) in CDK5-overexpressing AGS cells. (D) Immunoprecipitation (IP) assay using an antibody against CDK5 or DP1 in MKN45 cells. IgG was used as a negative control. Western blotting was performed for CDK5 and DP1. (E) Effects of CDK5 depletion on DP1 stability in MKN45 cells. The cells were incubated with cycloheximide (CHX) for the indicated times. (F) Effect of CDK5 on DP1 ubiquitination in MKN45 cells