Research Article

Identification of Sp1 as a Transcription Activator to Regulate Fibroblast Growth Factor 21 Gene Expression

Figure 3

Specific binding of Sp1 to the FGF21 promoter. (a) EMSA assay. Nuclear extracts from adipose tissue were incubated with biotin-labeled FGF21 probe encompassing the putative Sp1-B binding site, and Sp1C-probe containing the consensus site for Sp1. Competition experiments were performed using 100-fold and 200-fold excess of the unlabeled FGF21 oligonucleotide. Cross competition was performed using a 100-fold excess of the unlabeled Sp1C oligonucleotide. The specific DNA-protein complexes and free probes are indicated by the arrow. (b) ChIP analysis. Formaldehyde-crosslinked chromatin from adipose tissue or liver was incubated with anti-Sp1 antibody. As the negative control, the chromatin was incubated with nonspecific IgG. DNA immunoprecipitated using the antibody against Sp1 was analyzed by PCR with primers specific for the FGF21. Input DNA and diethylpyrocarbonate-treated water (DEPC H2O) were used as a positive or negative control for the PCR reaction, respectively.