scholarly journals Evolution of human, chicken, alligator, frog and zebrafish mineralocorticoid receptors: Allosteric influence on steroid specificity

2017 ◽  
Author(s):  
Yoshinao Katsu ◽  
Kaori Oka ◽  
Michael E. Baker
Keyword(s):  
Dna Binding ◽  
Ligand Binding ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Full Length ◽  
Dna Binding Domain ◽  
Mineralocorticoid Receptors ◽  
Terminal Domain ◽  
Terrestrial Vertebrate ◽  
Hinge Domain

AbstractWe studied the response to aldosterone, 11-deoxycorticosterone, 11-deoxycortisol, cortisol, corticosterone, progesterone, 19-norprogesterone and spironolactone of human, chicken, alligator, frog and zebrafish full-length mineralocorticoid receptors (MRs) and truncated MRs, lacking the N-terminal domain (NTD) and DNA-binding domain (DBD), in which the hinge domain and ligand binding domain (LBD) were fused to a GAL4-DBD. Compared to full-length MRs, some vertebrate MRs required higher steroid concentrations to activate GAL4-DBD-MR-hinge/LBD constructs. For example, 11-deoxycortisol activated all full-length vertebrate MRs, but did not activate truncated terrestrial vertebrate MRs and was an agonist for truncated zebrafish MR. Progesterone, 19-norProgesterone and spironolactone did not activate full-length and truncated human, alligator and frog MRs. However, at 10 nM, these steroids activated full-length chicken and zebrafish MRs; at 100 nM, these steroids had little activity for truncated chicken MRs, while retaining activity for truncated zebrafish MRs, evidence that regulation of progestin activation of chicken MR resides in NTD/DBD and of zebrafish MR in hinge-LBD. Zebrafish and chicken MRs contain a serine corresponding to Ser810 in human MR, required for its antagonism by progesterone, suggesting novel regulation of progestin activation of chicken and zebrafish MRs. Progesterone may be a physiological activator of chicken and zebrafish MRs.

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 Evidence for DNA-Binding Domain-Ligand-Binding Domain Communications in the Androgen Receptor

2012 ◽  
Vol 32 (15) ◽  
pp. 3033-3043 ◽  
Author(s):  
C. Helsen ◽  
V. Dubois ◽  
A. Verfaillie ◽  
J. Young ◽  
M. Trekels ◽  
...  
Keyword(s):  
Androgen Receptor ◽  
Dna Binding ◽  
Ligand Binding ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Dna Binding Domain

scholarly journals Interaction of the phosphorylated DNA-binding domain in nuclear receptor CAR with its ligand-binding domain regulates CAR activation

2017 ◽  
Vol 293 (1) ◽  
pp. 333-344 ◽  
Author(s):  
Ryota Shizu ◽  
Jungki Min ◽  
Mack Sobhany ◽  
Lars C. Pedersen ◽  
Shingo Mutoh ◽  
...  
Keyword(s):  
Dna Binding ◽  
Ligand Binding ◽  
Nuclear Receptor ◽  
Binding Domain ◽  
Ligand Binding Domain ◽  
Dna Binding Domain

scholarly journals Evolution of corticosteroid specificity for human, chicken, alligator and frog glucocorticoid receptors

2016 ◽  
Author(s):  
Yoshinao Katsu ◽  
Satomi Kohno ◽  
Kaori Oka ◽  
Michael E. Baker
Keyword(s):  
Ligand Binding ◽  
Transcriptional Activation ◽  
Glucocorticoid Receptors ◽  
Binding Domain ◽  
Full Length ◽  
Dna Binding Domain ◽  
Terrestrial Vertebrates ◽  
Steroid Sensitivity ◽  
Hinge Domain

AbstractWe investigated the evolution of the response of human, chicken, alligator and frog glucocorticoid receptors (GRs) to dexamethasone, cortisol, corticosterone, 11-deoxycorticosterone, 11-deoxycortisol and aldosterone. We find significant differences among these vertebrates in the transcriptional activation of their full length GRs by these steroids, indicating that there were changes in the specificity of the GR for steroids during the evolution of terrestrial vertebrates. To begin to study the role of interactions between different domains on the GR in steroid sensitivity and specificity for terrestrial GRs, we investigated transcriptional activation of truncated GRs containing their hinge domain and ligand binding domain (LBD) fused to a GAL4 DNA binding domain (GAL4 DBD). Compared to corresponding full length GRs, transcriptional activation of GAL4 DBD-GR hinge/LBD constructs required higher steroid concentrations and displayed altered steroid specificity, indicating that interactions between the hinge/LBD and other domains are important in glucocorticoid activation of these terrestrial GRs.

scholarly journals An Inhibitory Region of the DNA-Binding Domain of Thyroid Hormone Receptor Blocks Hormone-Dependent Transactivation

1998 ◽  
Vol 12 (1) ◽  
pp. 34-44 ◽  
Author(s):  
Ying Liu ◽  
Akira Takeshita ◽  
Takashi Nagaya ◽  
Aria Baniahmad ◽  
William W. Chin ◽  
...  
Keyword(s):  
Thyroid Hormone ◽  
Dna Binding ◽  
Ligand Binding ◽  
Transcriptional Activation ◽  
Thyroid Hormone Receptor ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Chimeric Receptor ◽  
Receptor System ◽  
Inhibitory Region

Abstract We have employed a chimeric receptor system in which we cotransfected yeast GAL4 DNA-binding domain/retinoid X receptor β ligand-binding domain chimeric receptor (GAL4RXR), thyroid hormone receptor-β (TRβ), and upstream activating sequence-reporter plasmids into CV-1 cells to study repression, derepression, and transcriptional activation. In the absence of T3, unliganded TR repressed transcription to 20% of basal level, and in the presence of T3, liganded TRβ derepressed transcription to basal level. Using this system and a battery of TRβ mutants, we found that TRβ/RXR heterodimer formation is necessary and sufficient for basal repression and derepression in this system. Additionally, an AF-2 domain mutant (E457A) mediated basal repression but not derepression, suggesting that interaction with a putative coactivator at this site may be critical for derepression. Interestingly, a mutant containing only the TRβ ligand binding domain (LBD) not only mediated derepression, but also stimulated transcriptional activation 10-fold higher than basal level. Studies using deletion and domain swap mutants localized an inhibitory region to the TRβ DNA-binding domain. Titration studies further suggested that allosteric changes promoting interaction with coactivators may account for enhanced transcriptional activity by LBD. In summary, our findings suggest that TR heterodimer formation with RXR is important for repression and derepression, and coactivator interaction with the AF-2 domain may be needed for derepression in this chimeric system. Additionally, there may be an inhibitory region in the DNA-binding domain, which reduces TR interaction with coactivators, and prevents full-length wild-type TRβ from achieving transcriptional activation above basal level in this chimeric receptor system.

scholarly journals Mad proteins contain a dominant transcription repression domain.

1996 ◽  
Vol 16 (10) ◽  
pp. 5772-5781 ◽  
Author(s):  
D E Ayer ◽  
C D Laherty ◽  
Q A Lawrence ◽  
A P Armstrong ◽  
R N Eisenman
Keyword(s):  
Dna Binding ◽  
Transcriptional Repression ◽  
Binding Domain ◽  
Full Length ◽  
Dna Binding Domain ◽  
Interaction Domain ◽  
Transcription Repression ◽  
Helix Loop Helix ◽  
Repression Domain

Transcription repression by the basic region-helix-loop-helix-zipper (bHLHZip) protein Mad1 requires DNA binding as a ternary complex with Max and mSin3A or mSin3B, the mammalian orthologs of the Saccharomyces cerevisiae transcriptional corepressor SIN3. The interaction between Mad1 and mSin3 is mediated by three potential amphipathic alpha-helices: one in the N terminus of Mad (mSin interaction domain, or SID) and two within the second paired amphipathic helix domain (PAH2) of mSin3A. Mutations that alter the structure of the SID inhibit in vitro interaction between Mad and mSin3 and inactivate Mad's transcriptional repression activity. Here we show that a 35-residue region containing the SID represents a dominant repression domain whose activity can be transferred to a heterologous DNA binding region. A fusion protein comprising the Mad1 SID linked to a Ga14 DNA binding domain mediates repression of minimal as well as complex promoters dependent on Ga14 DNA binding sites. In addition, the SID represses the transcriptional activity of linked VP16 and c-Myc transactivation domains. When fused to a full-length c-Myc protein, the Mad1 SID specifically represses both c-Myc's transcriptional and transforming activities. Fusions between the GAL DNA binding domain and full-length mSin3 were also capable of repression. We show that the association between Mad1 and mSin3 is not only dependent on the helical SID but is also dependent on both putative helices of the mSin3 PAH2 region, suggesting that stable interaction requires all three helices. Our results indicate that the SID is necessary and sufficient for transcriptional repression mediated by the Mad protein family and that SID repression is dominant over several distinct transcriptional activators.

Molecular genetics of chromosome translocations involving EWS and related family members

1999 ◽  
Vol 1 (3) ◽  
pp. 127-138 ◽  
Author(s):  
JUNGHO KIM ◽  
JERRY PELLETIER
Keyword(s):  
Transcription Factors ◽  
Dna Binding ◽  
Molecular Genetics ◽  
Family Members ◽  
Binding Domain ◽  
Dna Binding Domain ◽  
Chromosome Translocations ◽  
Amino Terminal ◽  
Terminal Domain ◽  
Related Family

Kim, Jungho, and Jerry Pelletier. Molecular genetics of chromosome translocations involving EWS and related family members. Physiol. Genomics 1: 127–138, 1999.—Many types of sarcomas are characterized by specific chromosomal translocations that appear to result in the production of novel, tumor-specific chimeric transcription factors. Many of these show striking similarities: the emerging picture is that the amino-terminal domain of the fusion product is donated by the Ewing's sarcoma gene ( EWS) or a related member from the same gene family, whereas the carboxy-terminal domain often consists of a DNA-binding domain derived from one of a number of transcription factors. Given the observation that the different translocation partners of the EWS protooncogene are associated with distinct types of sarcomas, the functional consequence of fusing EWS (or a related family member) to a different DNA-binding domain can only be understood in the context of functional studies that define the specificity of action of the different fusion products. An understanding of the molecular structure and function of these translocations provides new methods for diagnosis and novel targets for therapeutics.

scholarly journals Regulation of Cell Cycle Transcription Factor Swi4 through Auto-Inhibition of DNA Binding

1999 ◽  
Vol 19 (10) ◽  
pp. 6729-6741 ◽  
Author(s):  
Kristin Baetz ◽  
Brenda Andrews
Keyword(s):  
Cell Cycle ◽  
Transcription Factors ◽  
Dna Binding ◽  
Binding Domain ◽  
Terminal Region ◽  
Full Length ◽  
Intramolecular Interactions ◽  
Dna Binding Domain ◽  
C Terminus ◽  
Binding Factor

ABSTRACTInSaccharomyces cerevisiae, two transcription factors, SBF (SCB binding factor) and MBF (MCB binding factor), promote the induction of gene expression at the G1/S-phase transition of the mitotic cell cycle. Swi4 and Mbp1 are the DNA binding components of SBF and MBF, respectively. The Swi6 protein is a common subunit of both transcription factors and is presumed to play a regulatory role. SBF binding to its target sequences, the SCBs, is a highly regulated event and requires the association of Swi4 with Swi6 through their C-terminal domains. Swi4 binding to SCBs is restricted to the late M and G1phases, when Swi6 is localized to the nucleus. We show that in contrast to Swi6, Swi4 remains nuclear throughout the cell cycle. This finding suggests that the DNA binding domain of Swi4 is inaccessible in the full-length protein when not complexed with Swi6. To explore this hypothesis, we expressed Swi4 and Swi6 in insect cells by using the baculovirus system. We determined that partially purified Swi4 cannot bind SCBs in the absence of Swi6. However, Swi4 derivatives carrying point mutations or alterations in the extreme C terminus were able to bind DNA or activate transcription in the absence of Swi6, and the C terminus of Swi4 inhibited Swi4 derivatives from binding DNA intrans. Full-length Swi4 was determined to be monomeric in solution, suggesting an intramolecular mechanism for auto-inhibition of binding to DNA by Swi4. We detected a direct in vitro interaction between a C-terminal fragment of Swi4 and the N-terminal 197 amino acids of Swi4, which contain the DNA binding domain. Together, our data suggest that intramolecular interactions involving the C-terminal region of Swi4 physically prevent the DNA binding domain from binding SCBs. The interaction of the carboxy-terminal region of Swi4 with Swi6 alleviates this inhibition, allowing Swi4 to bind DNA.

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