{"id":{"repo_id":"uiuc","oai_identifier":"oai:www.ideals.illinois.edu:2142/20261"},"canonical_url":"https://search.dev.ndltd.org/etd/uiuc/oai:www.ideals.illinois.edu:2142/20261","repository":{"repo_id":"uiuc","name":"University of Illinois - Urbana-Champaign","base_url":"https://www.ideals.illinois.edu/oai-pmh"},"display":{"title":"Molecular actions of estrogen agonists and antagonists and their interaction with the human estrogen receptor","abstract":"Studies were undertaken to examine the interaction of estrogen agonists and antagonists with the human estrogen receptor (ER) and to examine the consequences of this interaction on the DNA binding and transactivating ability of the receptor. The functional role of cysteines in the hormone binding domain (HBD) of the receptor was examined by in vitro mutagenesis, followed by analysis of the hormone binding, covalent labeling and transactivating ability of the mutant receptors. Mutagenesis revealed that Cys530 is the preferential site of labeling by covalent affinity labels, however, in the absence of a cysteine residue at postion 530, Cys 381 is labeled, and Cys381 lies near the ligand binding pocket. Receptors bearing single or multiple cysteine to alanine substitutions were not altered in their response to reversibly binding ligands. However, the effectiveness of covalently attaching ligands were altered in receptors containing an alanine at position 530, indicating an important role of Cys530 in the function of these ligands. One receptor mutant (C447A), displayed an estradiol dose response shift in transactivation assays of about 30-50 fold. Analysis of the C447A mutant revealed that this receptor is a temperature-sensitive mutant whose DNA binding is ligand-dependent at elevated temperatures.","abstract_html":"Studies were undertaken to examine the interaction of estrogen agonists and antagonists with the human estrogen receptor (ER) and to examine the consequences of this interaction on the DNA binding and transactivating ability of the receptor. The functional role of cysteines in the hormone binding domain (HBD) of the receptor was examined by in vitro mutagenesis, followed by analysis of the hormone binding, covalent labeling and transactivating ability of the mutant receptors. Mutagenesis revealed that Cys530 is the preferential site of labeling by covalent affinity labels, however, in the absence of a cysteine residue at postion 530, Cys 381 is labeled, and Cys381 lies near the ligand binding pocket. Receptors bearing single or multiple cysteine to alanine substitutions were not altered in their response to reversibly binding ligands. However, the effectiveness of covalently attaching ligands were altered in receptors containing an alanine at position 530, indicating an important role of Cys530 in the function of these ligands. One receptor mutant (C447A), displayed an estradiol dose response shift in transactivation assays of about 30-50 fold. Analysis of the C447A mutant revealed that this receptor is a temperature-sensitive mutant whose DNA binding is ligand-dependent at elevated temperatures.","abstract_has_math":false,"creators":["Reese, Joseph Charles, Jr."],"institution":"University of Illinois at Urbana-Champaign","degree_name":"Ph.D.","degree_level":"Dissertation","degree_discipline":"Molecular and Integrative Physiology","degree_department":null,"school":null,"contributors":["Katzenellenbogen, Benita S."],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2011,"date_issued":"2011-05-07T12:34:05Z","date_published":"2011-05-07T12:34:05Z","updated_at":"2026-07-22T22:25:15Z","subjects":["Biology, Animal Physiology"],"languages":["eng"],"rights":["Copyright 1992 Reese, Joseph Charles, Jr"],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["AAI9236575","(UMI)AAI9236575"],"render_values":[{"text":"AAI9236575","href":null,"code":true},{"text":"(UMI)AAI9236575","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/2142/20261","outbound_label":"Handle","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Katzenellenbogen, Benita S."]},{"key":"dc:creator","label":"Author","values":["Reese, Joseph Charles, Jr."]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date","label":"Dc Date","values":["2011-05-07T12:34:05Z","10000-01-01","1992"]},{"key":"dc:type","label":"Dc Type","values":["text"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Molecular and Integrative Physiology"]},{"key":"thesis:degree_level","label":"Degree Level","values":["Dissertation"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Ph.D."]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["University of Illinois at Urbana-Champaign"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["Biology, Animal Physiology"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["eng"]},{"key":"dc:rights","label":"Dc Rights","values":["Copyright 1992 Reese, Joseph Charles, Jr"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["AAI9236575","(UMI)AAI9236575","http://hdl.handle.net/2142/20261"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["Studies were undertaken to examine the interaction of estrogen agonists and antagonists with the human estrogen receptor (ER) and to examine the consequences of this interaction on the DNA binding and transactivating ability of the receptor. The functional role of cysteines in the hormone binding domain (HBD) of the receptor was examined by in vitro mutagenesis, followed by analysis of the hormone binding, covalent labeling and transactivating ability of the mutant receptors. Mutagenesis revealed that Cys530 is the preferential site of labeling by covalent affinity labels, however, in the absence of a cysteine residue at postion 530, Cys 381 is labeled, and Cys381 lies near the ligand binding pocket. Receptors bearing single or multiple cysteine to alanine substitutions were not altered in their response to reversibly binding ligands. However, the effectiveness of covalently attaching ligands were altered in receptors containing an alanine at position 530, indicating an important role of Cys530 in the function of these ligands. One receptor mutant (C447A), displayed an estradiol dose response shift in transactivation assays of about 30-50 fold. Analysis of the C447A mutant revealed that this receptor is a temperature-sensitive mutant whose DNA binding is ligand-dependent at elevated temperatures.","The molecular actions of estrogen agonists and antagonists were examined using DNA binding assays in vitro and in whole cells. Estrogen receptor unoccupied by ligand or bound by trans-OH-tamoxifen were capable of binding to DNA in both assays. However, differences were observed with receptor exposed to the antiestrogen ICI 164,384 (ICI) in vivo when the receptor source was from transfected COS-1 cells. Receptor isolated from cells exposed to ICI lost its DNA binding ability through a cell-mediated process. COS cells treated with ICI did not have reduced levels of ER protein, however, treatment of breast cancer cells with ICI resulted in a rapid reduction of ER protein. Studies demonstrated that while ER is reduced in these cells, there are functional levels of ER remaining within the cells. ICI still behaved as a pure antagonist, indicating that neither the rapid downregulation of ER protein or the elimination of the DNA binding ability of receptor in target cells can fully explain the pure antagonistic nature of the antiestrogen ICI.","Made available in DSpace on 2011-05-07T12:34:05Z (GMT). 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The functional role of cysteines in the hormone binding domain (HBD) of the receptor was examined by in vitro mutagenesis, followed by analysis of the hormone binding, covalent labeling and transactivating ability of the mutant receptors. Mutagenesis revealed that Cys530 is the preferential site of labeling by covalent affinity labels, however, in the absence of a cysteine residue at postion 530, Cys 381 is labeled, and Cys381 lies near the ligand binding pocket. Receptors bearing single or multiple cysteine to alanine substitutions were not altered in their response to reversibly binding ligands. However, the effectiveness of covalently attaching ligands were altered in receptors containing an alanine at position 530, indicating an important role of Cys530 in the function of these ligands. One receptor mutant (C447A), displayed an estradiol dose response shift in transactivation assays of about 30-50 fold. Analysis of the C447A mutant revealed that this receptor is a temperature-sensitive mutant whose DNA binding is ligand-dependent at elevated temperatures.","The molecular actions of estrogen agonists and antagonists were examined using DNA binding assays in vitro and in whole cells. Estrogen receptor unoccupied by ligand or bound by trans-OH-tamoxifen were capable of binding to DNA in both assays. However, differences were observed with receptor exposed to the antiestrogen ICI 164,384 (ICI) in vivo when the receptor source was from transfected COS-1 cells. Receptor isolated from cells exposed to ICI lost its DNA binding ability through a cell-mediated process. COS cells treated with ICI did not have reduced levels of ER protein, however, treatment of breast cancer cells with ICI resulted in a rapid reduction of ER protein. Studies demonstrated that while ER is reduced in these cells, there are functional levels of ER remaining within the cells. ICI still behaved as a pure antagonist, indicating that neither the rapid downregulation of ER protein or the elimination of the DNA binding ability of receptor in target cells can fully explain the pure antagonistic nature of the antiestrogen ICI.","Made available in DSpace on 2011-05-07T12:34:05Z (GMT). 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