Cloning and characterization of estrogen receptor α in mummichog, Fundulus heteroclitus

Abstract

Developmental exposure to 17β-estradiol (E 2) induced the death of embryos and fry, malformations, sex reversal, and incomplete ossification of vertebrae and cranial bones in the cyprinodont fish, the mummichog ( Fundulus heteroclitus). To clarify the mechanism by which exogenous estrogens caused these developmental effects, we determined the sequence of an estrogen receptor (ER) coding region, encoded by 620 amino acid residues. This region shared 80% identity to that of ERα of medaka ( Oryzias latipes). Northern blot analysis showed that two ERα mRNAs with 5.5 and 4 kb were expressed in the liver. These mRNAs were strongly induced by E 2 stimulation. The 4 kb mRNA was expressed 8 h after treatment, whereas the 5.5 kb mRNA was not induced until 12 h after E 2 stimulation. Vitellogenin (VTG) was expressed 8 h after E 2 stimulation in the male liver. Receptor binding assays using the protein of F. heteroclitus ERα (fhERα) ligand binding domain showed that alkylphenols bind to fhERα with a higher affinity (50 times or more) as compared with the human ERα. The present results demonstrate that the fhERα has a sequence very similar to that of medaka, and the mRNA for this receptor was induced by E 2-stimulation, followed subsequently by VTG expression. Furthermore, alkylphenols bind to fhERα more efficiently than to human ERα.