Immunorecognition of the active form of the oestrogen receptor by using a monoclonal antibody.
In previous studies, two forms (alpha and beta) of the oestrogen receptor, with different immunological characteristics, were observed in the cytosol fraction of fetal guinea-pig uterus, by using a monoclonal antibody to the human oestrogen receptor (D547Sp gamma). Only the alpha form was recognized by the antibody, shifting its sedimentation coefficient in high-salt sucrose gradients. The present work investigated the effect of several factors (time, temperature, high salt concentrations and Na2MoO4) on the interconversion of these two forms. Only the beta form was observed when cytosol was incubated with oestradiol for only 2-3 h, but 20 h later, 40-60% of the total oestradiol-receptor complexes were found as the alpha form. The transformation from the beta to the alpha form was accelerated by temperature (25 degrees C, 15 min) and exposure to high salt concentrations (0.4 M-KCl). On the other hand, Na2MoO4 completely blocked the transformation induced by time and temperature, but had little effect on that induced by KCl. The appearance of the alpha form always correlated with an increase in receptor binding to nuclei and DNA-cellulose. Finally, it was found that the isolated beta form, recovered from the gradient, was transformed into the alpha form after overnight dialysis under reduced pressure. The present data suggest that the alpha form, which is recognized by the monoclonal antibody, is the activated form of the oestrogen receptor.
D000818:Animals; D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex / Q000032:analysis; D002467:Cell Nucleus / Q000378:metabolism; D002482:Cellulose / Q000031:analogs & derivatives / Q000378:metabolism; D002499:Centrifugation, Density Gradient; D004247:DNA / Q000031:analogs & derivatives / Q000378:metabolism; D005260:Female; D006168:Guinea Pigs; D008982:Molybdenum / Q000494:pharmacology; D011189:Potassium Chloride / Q000494:pharmacology; D011960:Receptors, Estrogen / Q000187:drug effects / Q000276:immunology*; D013696:Temperature; D013997:Time Factors
D000911:Antibodies, Monoclonal; D000936:Antigen-Antibody Complex; C033190:DNA-cellulose; D011960:Receptors, Estrogen; D011189:Potassium Chloride; D008982:Molybdenum; D002482:Cellulose; D004247:DNA; C024687:sodium molybdate(VI)