The Peroxisomal 3-keto-acyl-CoA thiolase B Gene Expression Is under the Dual Control of PPARα and HNF4α in the Liver
Figure 3
TB PPRE3 is a novel binding site for the nuclear receptor HNF4α. (a) Binding of HNF4α to native radiolabelled (32P) TB PPRE3 was determined by gel shift assay. A double strand oligonucleotide containing (32P) TB PPRE3 was incubated with lysates of transfected (by the expression vector HNF4α) COS-7 cells. Fold excess of specific (Sp.) cold probe (PPRE of the peroxisomal ACOX-I gene) was used for data shown lanes 3, 4 and 5. Nonspecific (Non Sp.) cold probe (Sp1) was used for data shown lanes 6, 7 and 8. Binding complexes were resolved on a 6% non-denaturing polyacrylamide gel. The arrow indicates the specific binding of HNF4α. The star indicates nonspecific binding. (b) Supershift assay (lane 2) was performed with an antibody directed against HNF4α. (c) COS-7 cells were transfected with increasing amounts (0 to 250 ng) of expression vectors encoding WT HNF4α with a Luc reporter vector containing TB PPRE3. (d) COS-7 cells were transfected with increasing amounts (0 to 250 ng) of expression vectors encoding wild-type HNF4α with a Luc reporter vector containing one copy of the DR1 localized in intron 5 of the mouse version of Thb (+4083 +4095 bp). Values are mean of three independent experiments ±SEM. Significantly different compared to control (transfection with empty pcDNA3.1 vector) with * by one-way ANOVA test.
