4 research outputs found

    Hypoxia induces the ERRγ gene expression in hepatoma cell lines.

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    <p>(<b>A–B</b>), HepG2 cells were seeded in 60-cm<sup>2</sup> dishes and incubated overnight. Then cells were incubated under hypoxia at indicated time period. The expression of ERRγ was analyzed by Western blot (<b>A</b>) and Q-PCR (<b>B</b>) analysis. (<b>C–D</b>) HepG2 cells were seeded in 60-cm<sup>2</sup> dishes and incubated overnight and then treated with DFO at indicated concentration and time period. The expression of ERRs was analyzed by Western blot (<b>C</b>) and Q-PCR (<b>D</b>) analysis. ERRγ gene expression was normalized to L32 gene expression, and α or β-tubulin expression. All data are representative of at least three independent experiments. Error bars show ± S.E.M. <sup>*</sup><i>P</i><0.05, <sup>**</sup><i>P</i><0.01 by two-tailed Student <i>t</i>-test.</p

    ERRγ directly regulates hypoxia mediated PDK4 gene expression.

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    <p>(<b>A</b>) HepG2 cells were transfected with hPDK4-Luc. After transfection, the cells were exposed to hypoxia for indicated period and lysates were utilized for luciferase and β-galactosidase assay. Experiments were done in triplicate and data are expressed as the fold activation relative to the control. (<b>B</b>) HepG2 cells were seeded in 60-cm<sup>2</sup> dishes and trasnfected siERRγ and control siRNA for 72 hr and then exposed hypoxia for 9 hr. Cells were harvested for analyzing luciferase and β-galactodidase assay. (<b>C–D</b>) HepG2 cells were transfected with several deletion constructs of hPDK4 (−848)-Luc, hPDK4 (−500)-Luc, hPDK4 (−291)-Luc and hPDK4-mtERRE1-Luc with pcDNA3-ERRγ in the presence or absence with hypoxia exposure, respectively. 48 hr after transfection, the cells were harvested and performed luciferase and β-galactodidase assay. Experiments were done in duplicate and data expressed as the fold activation related to control. (<b>E</b>) HepG2 cell were transiently transfected with hPDK4 (−848)-Luc, hPDK4-mtERRE1-Luc, pcDNA3-ERRγ and pcDNA3-HIF-1α. (<b>F</b>) ChIP assay: HepG2 cell was exposed to hypoxia for 9 hr. Input represents 10% of purified DNA in each sample. Cell extracts were immunoprecipitated with anti-ERRα and purified DNA samples were employed for Q-PCR with primers binding to ERRE (−502 to −252) and distal site (−1056 to −886) on the <i>PDK4</i> gene promoter. All data are representative of at least three independent experiments. Error bars show ± S.E.M. <sup>***</sup><i>P</i><0.001 by two-tailed Student <i>t</i>-test.</p
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