Inhibition of ER alpha-mediated trans-activation of human coagulation factor XII gene by heteromeric transcription factor NF-Y

Human coagulation factor XII promoter contains an estrogen response element that mediates ligand-activated ER alpha induction of coagulation factor XI I gene expression. The 3'-half of coagulation factor XII-estrogen response element overlaps a putative CCAAT box, the widespread regulatory element sp ecifically recognized by the heteromeric transcription factor NF-Y. Transie nt cotransfection of NF-Y and ER alpha results in strong inhibition of estr ogen stimulation of coagulation factor XII promoter activity. NF-Y antagoni sm is primarily exerted by the NF-YA subunit and does not require binding t o the CCAAT element, as NF-YA mutants with impaired DNA binding capacity re tain the ability to inhibit Ella trans-activation. EMSAs with increasing co ncentrations of recombinant NF-Y do not detect the formation of NF-Y-DNA co mplexes or show impairment of ER alpha binding to estrogen response element . Immunoprecipitation of whole cell extracts with anti-ER alpha antibody re veals an in vivo association between the two transcription factors, which i s abolished by deletion of the NF-YA carboxyl-terminus. In functional exper iments with sequential NF-YA deletion mutants the HAP2-homology region appe ars essential in eliciting NF-YA antagonistic activity. In conclusion, our results demonstrate that heteromeric transcription factor NF-Y inhibits est rogen induction of coagulation factor XII promoter in a DNA binding-indepen dent fashion and suggest a novel role for NF-Y as a partner for the ER alph a transcription complex.