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. 2011 Aug;179(2):1030–1040. doi: 10.1016/j.ajpath.2011.04.026

Figure 5.

Figure 5

PPARγ binds and recruits corepressors to the GRE site in the Ob promoter. A: Nuclear extracts from MCF-7 cells were incubated with GRE probe (lane 1). A 100-fold molar excess of unlabeled (lane 2) and mutated (Mut) (lane 3) probe was added. MCF-7 nuclear extracts treated with 1000 ng/mL of leptin and/or 10 μmol/L BRL for 6 hours (lanes 4, 5, and 6). Anti-GR (lane 7), anti-PPARγ (lane 8) antibodies (Abs) or IgG (lane 9) were added to the mixture. Lane 10 shows probe alone. B: ChIP with the anti-GR, anti-PPARγ, and anti-Pol II antibodies. ChIP with the anti-GR antibody was re-ChIP with anti-PPARγ antibody. C: ChIP with the anti-GR or anti-PPARγ antibodies was re-ChIP with either anti-NCoR or anti-SMRT antibodies. The leptin (Lep) promoter sequence including the GRE site was detected by real-time-PCR with specific primers (see Materials and Methods). The results are mean ± SE of three separate experiments expressed as the percentage of control. *P < 0.05 versus untreated cells; **P < 0.05 versus leptin. D: MCF-7 cells were treated with 1000 ng/mL of leptin and/or 10 μmol/L BRL for 48 hours. Immunoprecipitation was performed using anti-GR (left panel) or anti-PPARγ (right panel) antibodies and then blotted with anti-PPARγ, GR, NCoR, SMRT, or DAX-1 antibodies.