The binding of rat uterine cytosol oestrogen receptors to oligodeoxythymidylate--cellulose. Its relationship to a stable form of receptor complex with separate ligand- and oligonucleotide-binding sites.
The interaction of rat uterine cytosol oestrogen-receptor complexes with the synthetic acceptor oligo(dT)--cellulose was studied. Differences in the stability of receptor complexes and their ability to bind to oligo(dT)--cellulose on storage at 4 degrees C or when exposed to increased temperatures indicated heterogeneity of steroid- and oligonucleotide-binding sites. Dilution, dialysis and (NH4)2SO4 precipitation increased the interaction of receptor complexes with oligo(dT)--cellulose (a step termed activation). This increase may be the result of the removal of low-molecular-weight cytosol components which inhibit receptor activation, dimerization to the 5 S form, which binds to oligo(dT)--cellulose, or interaction of 5 S receptor with the oligonucleotide. Cytosol oestradiol--receptor complexes exhibited biphasic dissociation kinetics. All these manipulations resulted in an increase in the proportion of the slow-dissociating component equivalent to the increase in receptor binding to oligo(dT)--cellulose. In contrast, addition of 10mM-sodium molybdate to cytosol decreased both oligo(dT)--cellulose binding and the proportion of receptor with slow dissociation kinetics. The inclusion of proteinase inhibitors did not affect interactions of receptor with oligo(dT)--cellulose nor the dissociation kinetics. These results suggest that oligo(dT)--cellulose binding may serve to quantify the proportion of cytosol receptor in an active form capable of nuclear interaction and to help to ascertain whether a receptor system is fully functional. This binding procedure could prove useful in the evaluation of oestrogen responsivity under normal and pathological conditions.
D000818:Animals; D001665:Binding Sites; D002482:Cellulose / Q000031:analogs & derivatives* / Q000378:metabolism; D003600:Cytosol / Q000378:metabolism; D004958:Estradiol / Q000378:metabolism; D005260:Female; D066298:In Vitro Techniques; D007700:Kinetics; D008024:Ligands; D008982:Molybdenum / Q000494:pharmacology; D009838:Oligodeoxyribonucleotides / Q000378:metabolism*; D009841:Oligonucleotides / Q000378:metabolism*; D051381:Rats; D011960:Receptors, Estrogen / Q000187:drug effects / Q000378:metabolism*; D014599:Uterus / Q000378:metabolism*
D008024:Ligands; D009838:Oligodeoxyribonucleotides; D009841:Oligonucleotides; D011960:Receptors, Estrogen; C021722:oligo(dT)-cellulose; D004958:Estradiol; D008982:Molybdenum; D002482:Cellulose; C024687:sodium molybdate(VI)