Expression and cellular localization of naturally occurring beta estrogen receptors in uterine and mammary cell lines.
The protein ER-alpha has been exhaustively characterized in estrogen-sensitive tissues and cell lines. However, little is known regarding the expression and cellular distribution of the newly identified ER-beta protein. We first quantified the specific estradiol binding site content in the estrogen-responsive cell lines MCF-7 (mammary) and SHM (myometrial). In the two cell types, these sites were associated to the expression of both ER-alpha and -beta isoforms. Native ER-beta was visualized to reside inside the nucleus by means of conventional indirect immunofluorescence. The cells expressed ER-beta as a tight approximately 50 kDa triplet when resolved by sodium dodecyl sulfate-polyacrylamide gels (SDS-PAGE) and blotted using antibodies mapping different domains of the cloned ER-beta version. When the cells were subjected to homogenization and differential centrifugation, a substantial proportion of ER-beta immunolabeling was localized at membrane subfractions. ER-beta expression and partitioning was confirmed by Ligand blotting assays using estrogen derivatives coupled to different macromolecular tags. However, ER-alpha was expressed as the major estrogen binding protein in both cell lines. Similar localization experiments were performed on HeLa cells (cervix). Though usually considered ER-negative, this cell line displayed basal significant estrogen binding capacity and co-expression of both ER isoforms. Taken as a whole, the results indicate that ER-beta could be expressed as functional estrogen binding proteins among a dominant population of ER-alpha sites in the cell lines under study.
Departamento de Biología, Bioquímica y Farmacia, Universidad Nacional del Sur. 8000 Bahía Blanca, Argentina.
Paula
P
Monje
Ricardo
R
Boland
D001940:Breast / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D047629:Estrogen Receptor beta; D004967:Estrogens / Q000378:metabolism; D005260:Female; D015870:Gene Expression; D006801:Humans; D008024:Ligands; D008856:Microscopy, Fluorescence; D020033:Protein Isoforms / Q000032:analysis / Q000378:metabolism; D021381:Protein Transport; D011960:Receptors, Estrogen / Q000032:analysis* / Q000378:metabolism; D014407:Tumor Cells, Cultured; D014599:Uterus / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism
D047629:Estrogen Receptor beta; D004967:Estrogens; D008024:Ligands; D020033:Protein Isoforms; D011960:Receptors, Estrogen