MOLECULAR-BASIS OF ESTROGEN REGULATION OF HAGEMAN-FACTOR-XII GENE-EXPRESSION

Estrogen therapy has been reported to cause multiple alterations in he mostasis and to increase blood levels of several procoagulants, includ ing Hageman factor [factor XII (FXII)]. Liver FXII gene expression has been investigated in ovariectomized rats, treated or not with 17 beta -estradiol. A 6-fold stimulation of FXII gene transcription was observ ed in treated compared to untreated animals, indicating that 17 beta-e stradiol is able to induce FXII gene expression in vivo. We have recen tly shown that human FXII promoter contains an imperfect palindrome, 5 '-GGGCAnnnTGACC-3' at position -43/-31 resembling the consensus estrog en-responsive element (ERE). Portions of different length of the FXII promoter were fused to the chloramphenicol acetyltransferase (CAT) cod ing sequence and transiently cotransfected with human estrogen recepto r (ER) into NIH3T3 and HepGa cells in the presence or absence of 17 be ta-estradiol. A 230-base pair fragment of FXII promoter, spanning nucl eotides -181/49, conferred a strong estrogen responsiveness to the CAT reporter gene, suggesting that a functional ERE resides in this regio n. Cognate receptors, such as those for thyroid hormone or retinoic ac id, did not stimulate CAT activity. Gel mobility assays demonstrated a specific interaction between ER and the 230-bp FXII promoter fragment containing the putative ERE palindrome. Similar results were obtained when an oligonucleotide spanning the consensus ERE was used; the comp lex between ER and FXII promoter sequences was supershifted after the addition of an anti-ER monoclonal antibody. Insertion of FXII-ERE into the heterologous thymidine kinase promoter conferred a strong estroge n responsiveness that was abolished by mutations of the 5'-half of the palindrome. These results represent the first demonstration at the mo lecular level of the regulation of a blood coagulation factor gene by 17 beta-estradiol as well as the first identification of a functional ERE within this class of genes.