scholarly journals Structural Analysis on the Pathologic Mutant Glucocorticoid Receptor Ligand-Binding Domains

2016 ◽  
Vol 30 (2) ◽  
pp. 173-188 ◽  
Author(s):  
Darrell E. Hurt ◽  
Shigeru Suzuki ◽  
Takafumi Mayama ◽  
Evangelia Charmandari ◽  
Tomoshige Kino
Keyword(s):  
Glucocorticoid Receptor ◽  
Ligand Binding ◽  
Gene Mutations ◽  
Binding Pocket ◽  
Activation Function ◽  
Binding Activity ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Structural Defects ◽  
Mutant Receptors

Abstract Glucocorticoid receptor (GR) gene mutations may cause familial or sporadic generalized glucocorticoid resistance syndrome. Most of the missense forms distribute in the ligand-binding domain and impair its ligand-binding activity and formation of the activation function (AF)-2 that binds LXXLL motif-containing coactivators. We performed molecular dynamics simulations to ligand-binding domain of pathologic GR mutants to reveal their structural defects. Several calculated parameters including interaction energy for dexamethasone or the LXXLL peptide indicate that destruction of ligand-binding pocket (LBP) is a primary character. Their LBP defects are driven primarily by loss/reduction of the electrostatic interaction formed by R611 and T739 of the receptor to dexamethasone and a subsequent conformational mismatch, which deacylcortivazol resolves with its large phenylpyrazole moiety and efficiently stimulates transcriptional activity of the mutant receptors with LBP defect. Reduced affinity of the LXXLL peptide to AF-2 is caused mainly by disruption of the electrostatic bonds to the noncore leucine residues of this peptide that determine the peptide's specificity to GR, as well as by reduced noncovalent interaction against core leucines and subsequent exposure of the AF-2 surface to solvent. The results reveal molecular defects of pathologic mutant receptors and provide important insights to the actions of wild-type GR.

scholarly journals A Ligand Binding Domain Mutation in the Mouse Glucocorticoid Receptor Functionally Links Chromatin Remodeling and Transcription Initiation

1999 ◽  
Vol 19 (12) ◽  
pp. 8146-8157 ◽  
Author(s):  
Lynn A. Sheldon ◽  
Catharine L. Smith ◽  
Jack E. Bodwell ◽  
Allan U. Munck ◽  
Gordon L. Hager
Keyword(s):  
Glucocorticoid Receptor ◽  
Ligand Binding ◽  
Chromatin Remodeling ◽  
Transcriptional Activation ◽  
Transcription Initiation ◽  
Activation Function ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Activation Process ◽  
Activation Domain

ABSTRACT We utilized the mouse mammary tumor virus (MMTV) long terminal repeat (LTR) in vivo to understand how the interaction of the glucocorticoid receptor (GR) with a nucleosome-assembled promoter allows access of factors required for the transition from a repressed promoter to a derepressed, transcriptionally competent promoter. A mutation (C644G) in the ligand binding domain (LBD) of the mouse GR has provided information regarding the steps required in the derepression/activation process and in the functional significance of the two major transcriptional activation domains, AF1 and AF2. The mutant GR activates transcription from a transiently transfected promoter that has a disordered nucleosomal structure, though significantly less well than the wild-type GR. With an integrated, replicated promoter, which is assembled in an ordered nucleosomal array, the mutant GR does not activate transcription, and it fails to induce chromatin remodeling of the MMTV LTR promoter, as indicated by nuclease accessibility assays. Together, these findings support a two-step model for the transition of a nucleosome-assembled, repressed promoter to its transcriptionally active, derepressed form. In addition, we find that the C-terminal GR mutation is dominant over the transcription activation function of the N-terminal GR activation domain. These findings suggest that the primary activation function of the C-terminal activation domain is different from the function of the N-terminal activation domain and that it is required for derepression of the chromatin-repressed MMTV promoter.

scholarly journals The Distinct Agonistic Properties of the Phenylpyrazolosteroid Cortivazol Reveal Interdomain Communication within the Glucocorticoid Receptor

2005 ◽  
Vol 19 (5) ◽  
pp. 1110-1124 ◽  
Author(s):  
Noritada Yoshikawa ◽  
Keiko Yamamoto ◽  
Noriaki Shimizu ◽  
Sachiko Yamada ◽  
Chikao Morimoto ◽  
...  
Keyword(s):  
Glucocorticoid Receptor ◽  
Ligand Binding ◽  
Conformational Changes ◽  
Binding Pocket ◽  
Activation Function ◽  
Receptor Activation ◽  
Binding Domain ◽  
Fine Tuning ◽  
Docking Analysis ◽  
A Charge

Abstract Recent structural analyses of the nuclear receptors establish a paradigm of receptor activation, in which agonist binding induces the ligand binding domain (LBD)/activation function-2 helix to form a charge clamp for coactivator recruitment. However, these analyses have not sufficiently addressed the mechanisms for differential actions of various synthetic steroids in terms of fine tuning of multiple functions of whole receptor molecules. In the present study, we used the glucocorticoid receptor (GR)-specific agonist cortivazol (CVZ) to probe the plasticity and functional modularity of the GR. Structural docking analysis revealed that although CVZ is more bulky than other agonists, it can be accommodated in the ligand binding pocket of the GR by reorientation of several amino acid side chains but without major alterations in the active conformation of the LBD. In this induced fit model, the phenylpyrazole A-ring of CVZ establishes additional contacts with helices 3 and 5 of the LBD that may contribute to a more stable LBD configuration. Structural and functional analysis revealed that CVZ is able to compensate for the deleterious effects of a C-terminal deletion of the LBD in a manner that mimics the stabilizing influence of the F602S point mutation. CVZ-mediated productive recruitment of transcriptional intermediary factor 2 to the C-terminally deleted LBD requires the receptor’s own DNA binding domain and is positively influenced by the N-terminal regions of GR or progesterone receptor. These results support a model where ligand-dependent conformational changes in the LBD play a role in GR-mediated gene regulation via modular interaction with the DBD and activation function-1.

scholarly journals Folding and stability of the ligand-binding domain of the glucocorticoid receptor

2002 ◽  
Vol 11 (8) ◽  
pp. 1926-1936 ◽  
Author(s):  
Stephen H. McLaughlin ◽  
Sophie E. Jackson
Keyword(s):  
Glucocorticoid Receptor ◽  
Ligand Binding ◽  
Binding Domain ◽  
Ligand Binding Domain

scholarly journals Molecular Determinants of Glucocorticoid Receptor Mobility in Living Cells: the Importance of Ligand Affinity

2003 ◽  
Vol 23 (6) ◽  
pp. 1922-1934 ◽  
Author(s):  
Marcel J. M. Schaaf ◽  
John A. Cidlowski
Keyword(s):  
Dna Binding ◽  
Glucocorticoid Receptor ◽  
Ligand Binding ◽  
Conformational Change ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Dependent Manner ◽  
Dna Binding Domain ◽  
Ligand Affinity ◽  
High Affinity

ABSTRACT The actions of glucocorticoids are mediated by the glucocorticoid receptor (GR), which is activated upon ligand binding, and can alter the expression of target genes either by transrepression or transactivation. We have applied FRAP (fluorescence recovery after photobleaching) to quantitatively assess the mobility of the yellow fluorescent protein (YFP)-tagged human GR α-isoform (hGRα) in the nucleus of transiently transfected COS-1 cells and to elucidate determinants of its mobility. Addition of the high-affinity agonist dexamethasone markedly decreases the mobility of the receptor in a concentration-dependent manner, whereas low-affinity ligands like corticosterone decrease the mobility to a much lesser extent. Analysis of other hGRα ligands differing in affinity suggests that it is the affinity of the ligand that is a major determinant of the decrease in mobility. Similar results were observed for two hGRα antagonists, the low-affinity antagonist ZK98299 and the high-affinity antagonist RU486. The effect of ligand affinity on mobility was confirmed with the hGRα mutant Q642V, which has an altered affinity for triamcinolone acetonide, dexamethasone, and corticosterone. Analysis of hGRα deletion mutants indicates that both the DNA-binding domain and the ligand-binding domain of the receptor are required for a maximal ligand-induced decrease in receptor mobility. Interestingly, the mobility of transfected hGRα differs among cell types. Finally, the proteasome inhibitor MG132 immobilizes a subpopulation of unliganded receptors, via a mechanism requiring the DNA-binding domain and the N-terminal part of the ligand-binding domain. Ligand binding makes the GR resistant to the immobilizing effect of MG132, and this effect depends on the affinity of the ligand. Our data suggest that ligand binding induces a conformational change of the receptor which is dependent on the affinity of the ligand. This altered conformation decreases the mobility of the receptor, probably by targeting the receptor to relatively immobile nuclear domains with which it transiently associates. In addition, this conformational change blocks immobilization of the receptor by MG132.

scholarly journals The AF1 and AF2 Domains of the Androgen Receptor Interact with Distinct Regions of SRC1

1999 ◽  
Vol 19 (12) ◽  
pp. 8383-8392 ◽  
Author(s):  
Charlotte L. Bevan ◽  
Sue Hoare ◽  
Frank Claessens ◽  
David M. Heery ◽  
Malcolm G. Parker
Keyword(s):  
Androgen Receptor ◽  
Ligand Binding ◽  
Activation Function ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Rich Region ◽  
Independent Manner ◽  
N Terminus ◽  
Androgen Receptor Activity ◽  
Glutamine Rich Region

ABSTRACT The androgen receptor is unusual among nuclear receptors in that most, if not all, of its activity is mediated via the constitutive activation function in the N terminus. Here we demonstrate that p160 coactivators such as SRC1 (steroid receptor coactivator 1) interact directly with the N terminus in a ligand-independent manner via a conserved glutamine-rich region between residues 1053 and 1123. Although SRC1 is capable of interacting with the ligand-binding domain by means of LXXLL motifs, this interaction is not essential since an SRC1 mutant with no functional LXXLL motifs retains its ability to potentiate androgen receptor activity. In contrast, mutants lacking the glutamine-rich region are inactive, indicating that this region is both necessary and sufficient for recruitment of SRC1 to the androgen receptor. This recruitment is in direct contrast to the recruitment of SRC1 to the estrogen receptor, which requires interaction with the ligand-binding domain.

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