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. 2011 Jul 27;438(Pt 1):111–119. doi: 10.1042/BJ20101939

Figure 2. FPP induces the recruitment of co-activators to PPARγ in vitro.

Figure 2

(A) GST pull-down assay using full-length PPARγ protein. Recombinant SRC-1 fragment (SRC-1-S2) containing a PPAR-binding region was incubated with GST–human PPARγ and vehicle control, 50 μM Pio or 50 μM/100 μM FPP. After washing, the bound recombinant SRC-1 protein was detected by immunoblotting. Total GST–PPARγ protein was detected by Coomassie Brilliant Blue staining. The results are representative of three independent blots. (B) The densitometric analysis of immunoblotting membranes normalized by the amount of GST–PPARγ of (A) is shown. The density of the vehicle control was set at 1 and the relative densities were presented as the fold induction relative to that of the vehicle control. (C) Scatchard plot analysis of the binding of a PPARγ triangles ligand complex to the TAMRA–SRC-1 in the presence of pioglitazone (Pro; closed triangles) or FPP (closed circles). As the control experiment, TAMRA–SRC-1 was incubated together with GST protein. Ligand concentrations (in μM) at all data points are 0.15, 1.0, 2.0 and 3.8 for FPP and 0.1, 0.4, 1.2 and 3.8 for Pio from the left data point respectively. The results are representative of five independent blots. (D) The recruitment of CBP to PPARγ in the presence or absence of FPP (50 or 100 μM) and GW9662 (25 μM) was determined by ELISA. All the values are means±S.E.M. for three to five tests. *P<0.05, **P<0.01.