PMID- 12569182 OWN - NLM STAT- MEDLINE DCOM- 20040617 LR - 20131121 IS - 0888-8809 (Print) IS - 0888-8809 (Linking) VI - 17 IP - 5 DP - 2003 May TI - Dissociation of steroid receptor coactivator 1 and nuclear receptor corepressor recruitment to the human glucocorticoid receptor by modification of the ligand-receptor interface: the role of tyrosine 735. PG - 845-59 AB - Within the human glucocorticoid receptor (GR) steroid binding pocket, tyrosine 735 makes hydrophobic contact with the steroid D ring. Substitution of tyrosine735 selectively impairs glucocorticoid transactivation but not transrepression. We now show, using both mammalian two-hybrid and glutathione-S-transferase pull downs, that such substitutions reduce interaction with steroid receptor coactivator 1, both basally and in response to agonist binding. Using a yeast two-hybrid screen we identified one of the three nuclear receptor interacting domains (NCoR-N1) of nuclear receptor corepressor (NCoR) as interacting with the GR C terminus in an RU486-specific manner. This was confirmed in mammalian two-hybrid experiments, and so we used the NCoR-N1 peptide to probe the GR C-terminal conformation. Substitution of Tyr735phe, Tyr735val, and Tyr735 ser, which impaired steroid receptor coactivator 1 (SRC1) interaction, enhanced NCoR-N1 recruitment, basally and after RU486. RU486 did not direct SRC1 recruitment to any of the GR constructs, and dexamethasone did not allow NCoR-N1 recruitment. Using a glutathione-S-transferase pull-down approach, the NCoR-N1 peptide was found to bind the full-length GR constitutively, and no further induction was seen with RU486, but it was reduced by dexamethasone. As both SRC1 and NCoR are predicted to recognize a common hydrophobic cleft in the GR, it seems that changes favorable to one interaction are detrimental to the other, thus identifying a molecular switch. FAU - Stevens, Adam AU - Stevens A AD - Endocrine Sciences Research Group, Faculty of Medicine, University of Manchester, United Kingdom. fras@fs1.ser.man.ac.uk FAU - Garside, Helen AU - Garside H FAU - Berry, Andrew AU - Berry A FAU - Waters, Charlotte AU - Waters C FAU - White, Anne AU - White A FAU - Ray, David AU - Ray D LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20030130 PL - United States TA - Mol Endocrinol JT - Molecular endocrinology (Baltimore, Md.) JID - 8801431 RN - 0 (Hormone Antagonists) RN - 0 (Ligands) RN - 0 (NCOR1 protein, human) RN - 0 (Nuclear Proteins) RN - 0 (Nuclear Receptor Co-Repressor 1) RN - 0 (Receptors, Glucocorticoid) RN - 0 (Receptors, Steroid) RN - 0 (Repressor Proteins) RN - 0 (Transcription Factors) RN - 320T6RNW1F (Mifepristone) RN - 42HK56048U (Tyrosine) RN - 7S5I7G3JQL (Dexamethasone) RN - EC 2.3.1.48 (Histone Acetyltransferases) RN - EC 2.3.1.48 (NCOA1 protein, human) RN - EC 2.3.1.48 (Nuclear Receptor Coactivator 1) RN - EC 2.5.1.18 (Glutathione Transferase) SB - IM MH - Amino Acid Sequence MH - Animals MH - Binding Sites MH - COS Cells MH - Dexamethasone/metabolism/pharmacology MH - Glutathione Transferase/genetics/metabolism MH - Histone Acetyltransferases MH - Hormone Antagonists/metabolism/pharmacology MH - Humans MH - Ligands MH - Mifepristone/metabolism/pharmacology MH - Molecular Sequence Data MH - Nuclear Proteins/genetics/*metabolism MH - Nuclear Receptor Co-Repressor 1 MH - Nuclear Receptor Coactivator 1 MH - Point Mutation MH - Receptors, Glucocorticoid/drug effects/genetics/*metabolism MH - Receptors, Steroid/genetics/*metabolism MH - Repressor Proteins/genetics/*metabolism MH - Structure-Activity Relationship MH - Transcription Factors/genetics/*metabolism MH - Two-Hybrid System Techniques MH - Tyrosine/chemistry/genetics/*metabolism EDAT- 2003/02/06 04:00 MHDA- 2004/06/18 05:00 CRDT- 2003/02/06 04:00 PHST- 2003/02/06 04:00 [pubmed] PHST- 2004/06/18 05:00 [medline] PHST- 2003/02/06 04:00 [entrez] AID - me.2002-0320 [pii] AID - 10.1210/me.2002-0320 [doi] PST - ppublish SO - Mol Endocrinol. 2003 May;17(5):845-59. doi: 10.1210/me.2002-0320. Epub 2003 Jan 30.