The distal enhancer region of the rat prolactin gene contains elements conferring response to multiple hormones

The ability of an upstream element of the rat PRL gene to permit transcriptional regulation in response to several different hormones has been examined. To test the ability of specific DNA sequences to mediate hormone responsiveness, DNA fragments were subcloned upstream of a thymidine kinase-chloramphenicol acetyltransferase fusion gene and transferred into GH₃ pituitary tumor cells. Initially, fragments representing a distal enhancer element (positions -1713 to -1495) and a more proximal element (positions -292 to -38) were tested. The results demonstrate that the distal enhancer permits cAMP, TRH, epidermal growth factor (EGF), and estradiol to stimulate expression of the thymidine kinase-chloramphenicol acetyltransferase gene. The proximal element permitted fusion gene regulation in response to cAMP, TRH, EGF, and phorbol esters. For the cAMP, TRH, and EGF responses, the distal element permitted responses approximately equal to or greater than responses conferred by the proximal PRL gene fragment. The response of the distal element to cAMP and TRH was more than additive with the response to estradiol, suggesting that the estrogen response element is distinct but may interact cooperatively with the other hormone response elements. Mutation of the estrogen-responsive element abolished both the response to estrogen and the cooperative response with cAMP, but not the response to cAMP itself. Mutation of a sequence involved in basal enhancer activity of the distal element reduced both basal transcription and the response to cAMP. These results suggest that the distal enhancer sequence of the PRL gene contains, in addition to an estrogen response element, elements that confer responsiveness to cAMP, TRH, and EGF. The similarities in the ability of the upstream and proximal elements of the rat PRL gene to respond to multiple hormones suggest that a common mechanism may underlie these similar responses.