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. Author manuscript; available in PMC: 2013 Sep 5.
Published in final edited form as: Cell Metab. 2013 Mar 5;17(3):423–435. doi: 10.1016/j.cmet.2013.01.016

Figure 7. TLE3 binds to Prdm16 and competes for its interaction with PPARγ.<.

Figure 7

br>A. TLE3 and Prdm16 localize to common target genes. ChIP assays were performed on 10T1/2-CAR cells stably transduced with Flag-tagged Prdm16 as described in Experimental Procedures. Cells were crosslinked on Day 4 after induction with BAT differentiation cocktail, and proteins precipitated with control IgG or antibodies to Flag, endogenous PPARγ, and endogenous TLE3 as indicated. A non-specific region of chromosome 15 was used as a negative control for binding. Promoter occupancy was determined by realtime PCR. B. Expression of TLE3 displaces Prdm16 from BAT- and WAT-selective gene promoters. Stable 10T1/2-CAR cells expressing vector or Flag-Prdm16 were differentiated for 7 days with BAT induction cocktail and then transduced with control or TLE3-expressing adenoviral vectors for 3-days. ChIP assays were performed using control IgG or an antibody to Flag as in A. C. 293 cells were transfected with V5-Prdm16, Flag-PPARγ2 and/or Flag-TLE3. Lysates were immunoprecipitated with anti-V5 antibody, and the precipitates were analyzed by immunoblotting. Similar results were obtained in multiple independent experiments. D. 293 cells were transfected with V5-Prdm16, myc-C/EBPβ and/or Flag-TLE3. Lysates were immunoprecipitated with anti-V5 antibody, and the precipitates were analyzed by immunoblotting. Similar results were obtained in multiple independent experiments.