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. 2012 Jul 15;26(14):1533–1545. doi: 10.1101/gad.184911.111

Figure 6.

Figure 6.

E2F7 is anti-proliferative and binds specifically to the E2F1 and DHFR promoters. (A) U2OS cells were transfected with increasing amounts of either empty pcDNA3 vector or a Flag-tagged E2F7-expressing pcDNA3 vector. Immunoblot analysis of transfected U2OS cells shows the relative expression levels of Flag-E2F7 in cells. (B) Colony formation assay of U2OS cells cotransfected with pBabe-Puro and either empty pcDNA3 vector or Flag-tagged E2F7-expressing vector. pBabe-Puro was used as a selection marker. The bar graph shows the average of three independent colony formation experiments. (C) Colony formation assay of H1299 (p53-null) cells cotransfected with pBabe-Puro and either empty pcDNA3 vector, Flag-tagged E2F7-expressing vector, or Flag-tagged wild-type p53-expressing vector. pBabe-puro was used as a selection marker. The bar graph shows the average of three independent colony formation experiments. (D) H1299 (p53-null) cells were transfected with 2 μg of pcDNA3-Flag E2F7 or empty pcDNA3. Cells were harvested 24 h after transfection and processed for ChIP using an anti-Flag antibody. Immunopurified genomic DNA fragments were analyzed for Flag-E2F7 enrichment at the indicated promoters by qPCR. (B–D) Error bars indicate the standard error of the mean. Statistical significance is shown using the Student's t-test analysis; (**) P < 0.01; (*) P < 0.05; (n.s.) not significant. The average of three independent experiments is shown. (E) HCT116 (p53+/+) cells were treated with doxorubicin for 24 h and processed for ChIP using a polyclonal E2F7 antibody or no antibody as a control. Immunopurified genomic DNA fragments were analyzed for endogenous E2F7 enrichment at the indicated promoters by qPCR. Primers in the albumin promoter were used as a negative control. (NT) Nontreated. The average of two independent experiments is shown.