Cloning and characterization of the human oviduct-specific gycoprotein (HuOGP) gene promoter

Abstract

Oviduct-specific glycoprotein (OGP) is a high molecular weight glycoprotein belonging to the chitinase protein family. OGPs are found to associate with the zona pellucida and plasma membrane of the oocyte and developing embryo. Recent studies have shown that OGP plays an important role in pre-fertilization reproductive events (i.e. sperm capacitation, sperm-zona binding and zona penetration). To have a better understanding of human OGP (HuOGP) gene expression, a 3.3 kb DNA fragment containing the 5′-flanking and the intron I region of the HuOGP gene was isolated. DNA sequence analysis of the HuOGP putative promoter revealed little homology with its hamster and mouse ogp gene counterparts. One transcription initiation site was found 12 nucleotides upstream of the first ATG codon of the HuOGP gene. The HuOGP gene promoter lacked typical CAAT or GC boxes, but contained eight half estrogen-responsive elements (ERE) and an imperfect ERE (iERE; 5′-GGTCANNNTGACT-3′) site. Deletion mutants of the 3.3 kb DNA fragment were generated and fused to a promoterless β-galactosidase (β-Gal) gene. Transfection studies revealed that in the presence of 100 nmol/l estradiol-17β (E2), a minimal 0.3 kb promoter construct (pH-298/+25βGal) mediated a high level of β-Gal expression in immortalized human oviductal epithelial OE-E6/E7 cells, but not in MCF-7 and CHO-K1 cells. By electromobility shift assay and specific estrogen receptor antibodies, we demonstrated that estrogen receptor β present in the OE-E6/E7 cells binds to the iERE. These findings allow a better understanding of the regulation of OGP gene expression in the human oviduct.