Estrogens are able to increase plasma levels of several procoagulant and fibrinolytic factors, including Hageman factor XII (FXII). These effects may be responsible for the multiple alterations of hemostasis observed in the course of estrogen therapy. The regulation of FXII gene expression by estrogens was therefore investigated in ovariectomized rats, treated or not with 17?-estradiol. Run-on experiments with isolated liver nuclei demonstrated that FXII gene transcription was about six time higher in treated than in untreated animals, indicating that estrogens are able to stimulate FXII gene expression in vivo. Computerized analysis of the cloned FXII promoter showed the presence of an imperfect palindrome (GGGCAnnnTGACC), resembling the consensus estrogen response element (ERE), at position -43/-31, as well as four additional upstream 5'-GGTCA-3' half-sites. Truncated portions of the human FXII promoter were fused to the chloramphenicol acetyl transferase (CAT) coding sequence and transiently co-transfected, together with the human estrogen receptor, into NIH3T3 and HepG2 cells, either in the presence or absence of 17?-estradiol (10-7 M). A 230 bp fragment (-181/+49) of the FXII promoter conferred a strong estrogen responsiveness (about 15-fold) to the CAT-reporter gene, confirming the finding that a functional ERE resides in this region. The estrogen effect was highly specific since related receptors, such as those for thyroid hormone or retinoic acid, were not able to stimulate CAT activity in the presence of their ligands. Gel mobility and DNase I footprinting assays demonstrated a specific interaction between the FXII promoter and estrogen receptor, in a region encompassing the palindromic FXII-ERE. Insertion of the putative FXII-ERE sequence into the heterologous thymidine kinase promoter conferred a strong estrogen responsiveness. These results represent the first demonstration, at molecular level, of the regulation of a blood coagulation factor gene by estrogens and the first identification of a functional ERE within this class of genes.
FUNCTIONAL CHARACTERIZATION AND RECEPTOR BINDING STUDIES OF THE FACTOR XII ESTROGEN RESPONSE ELEMENT.
A Farsetti;F Moretti;
1995
Abstract
Estrogens are able to increase plasma levels of several procoagulant and fibrinolytic factors, including Hageman factor XII (FXII). These effects may be responsible for the multiple alterations of hemostasis observed in the course of estrogen therapy. The regulation of FXII gene expression by estrogens was therefore investigated in ovariectomized rats, treated or not with 17?-estradiol. Run-on experiments with isolated liver nuclei demonstrated that FXII gene transcription was about six time higher in treated than in untreated animals, indicating that estrogens are able to stimulate FXII gene expression in vivo. Computerized analysis of the cloned FXII promoter showed the presence of an imperfect palindrome (GGGCAnnnTGACC), resembling the consensus estrogen response element (ERE), at position -43/-31, as well as four additional upstream 5'-GGTCA-3' half-sites. Truncated portions of the human FXII promoter were fused to the chloramphenicol acetyl transferase (CAT) coding sequence and transiently co-transfected, together with the human estrogen receptor, into NIH3T3 and HepG2 cells, either in the presence or absence of 17?-estradiol (10-7 M). A 230 bp fragment (-181/+49) of the FXII promoter conferred a strong estrogen responsiveness (about 15-fold) to the CAT-reporter gene, confirming the finding that a functional ERE resides in this region. The estrogen effect was highly specific since related receptors, such as those for thyroid hormone or retinoic acid, were not able to stimulate CAT activity in the presence of their ligands. Gel mobility and DNase I footprinting assays demonstrated a specific interaction between the FXII promoter and estrogen receptor, in a region encompassing the palindromic FXII-ERE. Insertion of the putative FXII-ERE sequence into the heterologous thymidine kinase promoter conferred a strong estrogen responsiveness. These results represent the first demonstration, at molecular level, of the regulation of a blood coagulation factor gene by estrogens and the first identification of a functional ERE within this class of genes.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
