Fig. 4

dsP21-322-directed p21 expression occurs at the transcriptional level. a The p21 intron, p21 protein-encoding mRNA and 18S rRNA expression levels were assessed using RT-qPCR in PC-3 and T-24 cells after dsP21-322, dsControl, or mock transfections. The p21 intron and p21 protein-encoding mRNA expression levels were normalized to that of 18S rRNA and plotted as the fold change relative to the mock treatment. b Schematic representation of the locations of the PCR primers that were used in ChIP to amplify the p21 promoter or transcriptional start site. c A ChIP assay was performed using an anti-RNA Pol II antibody to pull down dsP21-322-targeted promoters or d the p21 transcriptional start site associated with RNA Pol II in T-24 and U2-OS cells. The resulting DNA was amplified using qPCR and normalized to input levels. e Schematic representation showing the dsP21-322 binding site (dsP21BS) on the p21 promoter and the nonspecific binding site (NSBS) luciferase reporter. f A luciferase reporter assay for p21 in PC-3 and T-24 cells transfected with the indicated dsP21-322 and luciferase reporter genes. The input consists of nuclear extract prior to treatment with the antibody. IgG serves as the negative control antibody. The results are expressed as the mean ± S.D. calculated from triplicate independent transfection experiments with triplicate qPCR measurements for each