The glucocorticoid receptor and c-jun promoters contain AP-1 sites that bind different AP-1 transcription factors

The glucocorticoid receptor (GR) promoter contains several potential transcription factor recognition sites, including a putative AP-1 site. The GR promoter AP-1 site differs from the consensus AP-1 site by a nucleotide substitution, while the c-jun promoter contains a functionally characterized AP-1 site that varies from the consensus AP-1 site by a nucleotide insertion. Electrophoretic mobility shift assays were performed using nuclear extracts from a mouse pituitary tumor cell line (AtT-20) to test the binding capability of the AP-1 proteins, Jun and Fos, to the putative glucocorticoid receptor and the c-jun AP-1 sites. In addition, a comparison of the complexes formed at the GR AP-1 and c-jun AP-1 sites was done using antibodies specific for the Jun and Fos family members. The complexes formed with the GR AP-1 and c-jun AP-1 sites revealed striking differences. The GR AP-1 site formed complexes with both Jun and Fos family members. JunD was the most abundant Jun family member present, followed by JunB. cJun was absent from the complex. The amount of Fra-2 was greater than FosB in GR promoter AP-1 site complexes while Fra-1 was absent. A small amount of cFos may bind to the GR AP-1 site. In contrast to the GR promoter AP-1 site, only Jun family members were involved with complex formation on the c-jun promoter using AtT-20-cell nuclear extract, with JunD binding exceeding that of cJun. These results confirm previous studies suggesting that the c-jun promoter is stimulated solely by Jun family members. They also show preferential binding of Jun family members to different AP-1 sites present in different promoters. Finally, this study supports the hypothesis that the coordinate regulation of GR and c-jun gene regulation is mediated by crosstalk involving a Jun protein.