A novel piscine vitellogenin gene: structural and functional analyses of estrogen-inducible promoter.

The Oreochromis aureus vitellogenin, OaVtg, gene spans 9 kb and contains 34 exons. Its transcription start site is located 15 bp upstream of the translational start codon. Although the OaVtg promoter has a nonconsensus TATA, transient transfection assay showed that this promoter is capable of driving basal transcription. Two imperfect estrogen response elements: EREp (proximal) and EREd (distal) are located in the promoter at - 532 and - 1352, respectively. In competition gel mobility-shift assays, only EREp exhibited specific binding of the recombinant estrogen receptor protein, GST-C/D OaER. Another imperfect ERE (EREexon2) was detected within exon 2 of the OaVtg gene. This is a novel finding for a vitellogenin (Vtg) gene. EREexon2 similarly showed specific recognition of GST-C/D OaER. Both EREp and EREexon2 showed comparable binding affinities as consensus ERE. In transient transfections, the OaVtg promoter, EREp and EREd elicited significant increase in estrogen-dependent synthesis of CAT protein. Hence, we propose that the non-consensus OaVtg EREs contribute to the estrogen-dependent regulation of the OaVtg gene in vivo.