Open AccessHuman Glucocorticoid Receptor β Binds RU-486 and Is TranscriptionallyActive
Author(s) -
Laura J. LewisTuffin,
Christine M. Jewell,
Rachelle J. Bienstock,
Jennifer B. Collins,
John A. Cidlowski
Publication year - 2007
Publication title -
molecular and cellular biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.14
H-Index - 327
eISSN - 1067-8824
pISSN - 0270-7306
DOI - 10.1128/mcb.01439-06
Subject(s) - biology , glucocorticoid receptor , microbiology and biotechnology , gene isoform , receptor , cytoplasm , yellow fluorescent protein , ligand (biochemistry) , confocal microscopy , glucocorticoid , gene , biochemistry , endocrinology
Humanglucocorticoid receptor (hGR) is expressed as two alternately splicedC-terminal isoforms, α and β. In contrast to thecanonical hGRα, hGRβ is a nucleus-localized orphanreceptor thought not to bind ligand and not to affect genetranscription other than by acting as a dominant negative tohGRα. Here we used confocal microscopy to examine the cellularlocalization of transiently expressed fluorescent protein-taggedhGRβ in COS-1 and U-2 OS cells. Surprisingly, yellowfluorescent protein (YFP)-hGRβ was predominantly located in thecytoplasm and translocated to the nucleus following application of theglucocorticoid antagonist RU-486. This effect of RU-486 was confirmedwith transiently expressed wild-type hGRβ. Confocal microscopyof coexpressed YFP-hGRβ and cyan fluorescentprotein-hGRα in COS-1 cells indicated that the receptors moveinto the nucleus independently. Using a ligand binding assay, weconfirmed that hGRβ bound RU-486 but not the hGRαligand dexamethasone. Examination of the cellular localization ofYFP-hGRβ in response to a series of 57 related compoundsindicated that RU-486 is thus far the only identified ligand thatinteracts with hGRβ. The selective interaction of RU-486 withhGRβ was also supported by molecular modeling and computationaldocking studies. Interestingly, microarray analysis indicates thathGRβ, expressed in the absence of hGRα, can regulategene expression and furthermore that occupation of hGRβ withthe antagonist RU-486 diminishes that capacity despite the lack ofhelix 12 in the ligand bindingdomain.