title: Glucocorticoid receptor identified on nuclear envelopes of male rat livers by affinity labeling and immunochemistry.
abstract: To exert their action at the genome, steroids must traverse the nuclear envelope, either alone or complexed to their receptor. Our previous studies identified two classes of dexamethasone-binding sites on male rat liver nuclear envelopes: a low capacity, high affinity site and a high capacity, low affinity site. The affinity reagent, [3H]dexamethasone mesylate, labeled peptides at 35-85 kDa, which may be the low affinity glucocorticoid-binding peptides, as these peptides showed the same response to hormonal manipulation as the low affinity [3H]dexamethasone-binding sites previously characterized. With dexamethasone mesylate and a monoclonal antibody against the glucocorticoid receptor, we have confirmed that the high affinity binding site on the nuclear envelope is the glucocorticoid receptor. Affinity labeling revealed the presence of a doublet of peptides at 85 and 110 kDa, in the same mol wt range as that reported for the glucocorticoid receptor. Furthermore, these affinity-labeled peptides responded to hormonal manipulation like nuclear glucocorticoid receptors. The monoclonal antibody identified a doublet of peptides, a major component of 92-94 kDa and a minor component of 98 kDa. Again, both peptides responded to hormonal manipulation like nuclear glucocorticoid receptors. The nuclear envelope-associated glucocorticoid receptor is not extracted by 0.1 M NaCl or 1% Triton X-100. These results show that glucocorticoid hormone interacts with the nuclear envelope via binding to the transformed glucocorticoid receptor, lending support to the two-step model of steroid hormone action.
authors.affiliation: Department of Medicine, University of Ottawa, Moses and Rose Loeb Research Institute, Ottawa Civic Hospital, Canada.
authors.forename: G M
authors.initials: GM
authors.lastname: Howell
authors.forename: J A
authors.initials: JA
authors.lastname: Gustafsson
authors.forename: Y A
authors.initials: YA
authors.lastname: Lefebvre
mesh_terms: D000345:Affinity Labels; D000818:Animals; D001667:Binding, Competitive; D003907:Dexamethasone / Q000031:analogs & derivatives* / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D008099:Liver / Q000648:ultrastructure*; D008297:Male; D008970:Molecular Weight; D009685:Nuclear Envelope / Q000187:drug effects / Q000378:metabolism*; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D011965:Receptors, Glucocorticoid / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology
chemical_list: D000345:Affinity Labels; D011965:Receptors, Glucocorticoid; D011092:Polyethylene Glycols; D012965:Sodium Chloride; D003907:Dexamethasone; D017830:Octoxynol; C031710:dexamethasone 21-methanesulfonate; D004229:Dithiothreitol