Synergism between a half-site and an imperfect estrogen-responsive element, and cooperation with COUP-TFI are required for estrogen receptor (ER) to achieve a maximal estrogen-stimulation of rainbow trout ER gene

In all oviparous, liver represents one of the main E-2-target tissues where estrogen receptor (ER) constitutes the key mediator of estrogen action. Th e rainbow trout estrogen receptor (rtER) gene expression is markedly up-reg ulated by estrogens and the sequences responsible for this autoregulation h ave been located in a 0.2 kb upstream transcription start site within - 40/ - 248 enhancer region. Absence of interference with steroid hormone recepto rs and tissue-specific factors and a conserved basal transcriptional machin ery between yeast and higher eukaryotes, make yeast a simple assay system t hat will enable determination of important cia-acting regulatory sequences within rtER gene promoter and identification of transcription factors impli cated in the regulation of this gene. Deletion analysis allowed to show a s ynergistic effect between an imperfect estrogen-responsive element (ERE) an d a consensus half-ERE to achieve a high hormone-dependent transcriptional activation of the rtER gene promoter in the presence of stably expressed rt ER. As in mammalian cells, here we observed a positive regulation of the rt ER gene promoter by the chicken ovalbumin upstream promoter-transcription f actor I (COUP-TFI) through enhancing autoregulation. Using a point mutation COUP-TFI mutant unable to bind DNA demonstrates that enhancement of rtER g ene autoregulation requires the interaction of COUP-TFI to the DNA. Moreove r, this enhancement of transcriptional activation by COUP-TFI requires spec ifically the AF-1 transactivation function of ER and can be observed in the presence of E-2 or 4-hydroxytamoxifen but not ICI 164384. Thus, this paper describes the reconstitution of a hormone-responsive transcription unit in yeast in which the regulation of rtER gene promoter could be enhanced by t he participation of cis-elements and/or trans-acting factors, such as ER it self or COUP-TF.