THE NUCLEAR ORPHAN RECEPTORS COUP-TF AND ARP-1 POSITIVELY REGULATE THE TROUT ESTROGEN-RECEPTOR GENE THROUGH ENHANCING AUTOREGULATION

The rainbow trout estrogen receptor (rtER) is a positively autoregulat ed gene in liver cells, In a previous report, we showed that upregulat ion is mediated by an estrogen response element (ERE) located in the p roximal promoter of the gene and that a half binding site for nuclear receptors (5'-TGACCT-3') located 15 bp upstream of the ERE is involved in the magnitude of the estrogen response, We now report that the hum an orphan receptor COUP-TF and a COUP-TF-like protein from trout liver are able to bind to the consensus half-site, When cotransfected with the rtER gene proximal promoter, COUP-TF had no regulatory functions o n its own, Interestingly, COUP-TF enhanced rtER transactivation proper ties in the presence of estradiol in a dose-dependent manner when cotr ansfected with the rtER gene promoter, Unliganded retinoid receptor he terodimers had the same helper function as COUP-TF in the presence of estradiol but were switched to repressors when the ligand all-trans-re tinoic acid was added. Mutation of the consensus half-site only slight ly reduced COUP-TF helper function, suggesting that it actually result s from a complex mechanism that probably involves both DNA binding of COUP-TF to the promoter and protein-protein interaction with another t ranscription factor bound to the promoter, Nevertheless, a DNA-binding -defective mutant of COUP-TF was also defective in ER helper function, Competition footprinting analysis suggested that COUP-TF actually est ablishes contacts with the consensus upstream half-site and the downst ream ERE half-site that would form a DR-24-like response element, Inte raction of COUP-TF with the DR-24 element was confirmed in footprintin g assays by using nuclear extracts from Saccharomyces cerevisiae expre ssing COUP-TF, Finally, interaction of COUP-TF with mutants of the rtE R gene promoter showed that COUP-TF recognizes the ERE when the upstre am half-site is mutated, These data show that COUP-TF may activate tra nscription through interaction with other nuclear receptors. This cros s talk between liganded nuclear receptors and orphan receptors is like ly to modulate the spectrum of action of a particular ligand-receptor complex and may participate in the cell-type specificity of the ligand effect.