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Fig. 7 | Journal of Hematology & Oncology

Fig. 7

From: Bmi-1 regulates stem cell-like properties of gastric cancer cells via modulating miRNAs

Fig. 7

Bmi-1 regulates the expression of miR-21 and miR-34a via NF-kB. a Bmi-1 overexpression increases NF-kB transcriptional activity in SGC7901 cells, and Bmi-1 knockdown reduces NF-kB transcriptional activity in MKN45 cells. Gastric cancer cells were co-transfected with pNF-kB-luciferase plasmid and pRL-TK renilla plasmid, and transcriptional activity of NF-kB was tested by luciferase reporter activity assays. b Bmi-1 overexpression induces NF-kB aggregation in nuclear of SGC7901 cells, and Bmi-1 knockdown decreases NF-kB aggregation in nuclear of MKN45 cells. Cytoplasmic(C) and nuclear (N) proteins of cells were extracted and detected by Western blot. Nuclear protein TBP was used as a nuclear protein internal control and β-actin was as a loading control. c Bmi-1 overexpression promotes the binding of p65 to miR-21 promoter region in SGC7901 cells, and Bmi-1 interference reduces the binding of p65 to miR-21 promoter region in MKN45 cells. The binding of p65 to miR-21 promoter was analyzed by ChIP in gastric cancer cells with changed Bmi-1 expression level. The ChIP-enriched DNA analyzed by QRT-PCR was normalized to input DNA, followed by subtracting nonspecific binding determined by control IgG. d Bmi-1 overexpression promotes the binding of p65 to miR-34a promoter region in SGC7901 cells, and Bmi-1 interference reduces the binding of p65 to miR-34a promoter region in MKN45 cells. The binding of p65 to miR-34a promoter was analyzed by ChIP in gastric cancer cells with changed Bmi-1 expression level. The ChIP-enriched DNA analyzed by QRT-PCR was normalized to input DNA, followed by subtracting nonspecific binding determined by control IgG. e NF-kB inhibitor PDTC treatment inhibits the increase of expression of miR-21 and miR-34a induced by Bmi-1 overexpression in SGC7901 gastric cancer cells. Fold changes of miR-21 and miR-34a in control vector transfected cells (vector), Bmi-1 overexpressing cells (Bmi-1), and Bmi-1 overexpressing cells treated with NF-kB inhibitor PDTC (Bmi-1+PDTC) was analyzed by QRT-PCR. Error bars in all panels represent the mean ± SD (*P < 0.05, **P < 0.01)

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