Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.5.1.18 (glutathione S-transferase)
 22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Transcription factors, such as nuclear receptors, often exist in various forms that are generated by highly conserved splicing events. Whereas the functional significance of these splicing variants is often not known, it is known that nuclear receptors activate transcription through interaction with coactivators. The parameters, other than ligands, that might modulate those interactions, however, are not well characterized, nor is the role of splicing variants. In this study, transient transfection, yeast two-hybrid, and GST pulldown assays are used to show not only that nuclear receptor hepatocyte nuclear factor 4 alpha1 (HNF4alpha1, NR2A1) interacts with GRIP1, and other coactivators, in the absence of ligand but also that the uncommonly large F domain in the C terminus of the receptor inhibits that interaction. In vitro, the F domain was found to obscure an AF-2-independent binding site for GRIP1 that did not map to nuclear receptor boxes II or III. The results also show that a natural splicing variant containing a 10-amino-acid insert in the middle of the F domain (HNF4alpha2) abrogates that inhibition in vivo and in vitro. A series of protease digestion assays indicates that there may be structural differences between HNF4alpha1 and HNF4alpha2 in the F domain as well as in the ligand binding domain (LBD). The data also suggest that there is a direct physical contact between the F domain and the LBD of HNF4alpha1 and -alpha2 and that that contact is different in the HNF4alpha1 and HNF4alpha2 isoforms. Finally, we propose a model in which the F domain of HNF4alpha1 acts as a negative regulatory region for transactivation and in which the alpha2 insert ameliorates the negative effect of the F domain. A conserved repressor sequence in the F domains of HNF4alpha1 and -alpha2 suggests that this model may be relevant to other nuclear receptors as well.
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PMID:Modulation of transcriptional activation and coactivator interaction by a splicing variation in the F domain of nuclear receptor hepatocyte nuclear factor 4alpha1. 1049 May 91

The human estrogen receptor-related receptor (ERRalpha1, NR3B1a) was shown to bind a steroidogenic factor binding element (SFRE), TCAAGGTCATC, 26 base pairs upstream from the estrogen response element (ERE) of the human lactoferrin gene promoter. A mutation made at SFRE significantly reduced estrogen-dependent transcription from the lactoferrin ERE in human endometrial cells. In this study, we demonstrated that ERRalpha1 binds both SFRE and ERE elements and constitutively transactivates the lactoferrin gene promoter. In DNase I footprinting protection analysis, both SFRE and ERE regions were protected by glutathione S-transferase-ERRalpha1 fusion protein. The receptor formed two protein-DNA complexes with either SFRE or ERE in electrophoresis mobility shift assay. Homodimerization of ERRalpha1 was confirmed with the mammalian two-hybrid system. ERRalpha1 activates reporter constructs containing various types of estrogen response elements in endometrial and non-endometrial cells in transient transfection experiments. Overexpressing the coactivator, SRC1a or GRIP1, further enhances ERRalpha1-induced transcriptional activity. We demonstrated that the AF2 domain of ERRalpha1 is essential for the transactivation function and that deletion or mutation at this region abrogates the activation capability. Protein-protein interaction between the SRC1a and ERRalpha1 C terminus was confirmed with a GST glutathione S-transferase "pull-down" assay. When comparing ERRalpha1 and the estrogen receptor alpha (ERalpha) in many of the experiments, we found that ERalpha can also bind SFRE of the lactoferrin gene and transactivate the promoter activity in a ligand-dependent manner. The present study demonstrated that ERRalpha1 binds similar DNA elements as ERalpha and confers its transactivation function constitutively. Therefore, ERRalpha1 may actively modulate the estrogen response of lactoferrin gene as well as other estrogen-responsive genes.
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PMID:Estrogen receptor-related receptor alpha 1 interacts with coactivator and constitutively activates the estrogen response elements of the human lactoferrin gene. 1077 8

Antiestrogens, including tamoxifen and raloxifene, block estrogen receptor (ER) action by blocking the interactions of an estrogen-dependent activation function (AF-2) with p160 coactivators. Although tamoxifen does show some agonist activity in the presence of ERalpha, this stems from a distinct constitutive activation function (AF-1) that lies within the ERalpha N terminus. Previous studies identified a naturally occurring mutation (D351Y) that allows ERalpha to perceive tamoxifen and raloxifene as estrogens. Here, we examine the contributions of ERalpha activation functions to the D351Y phenotype. We find that the AF-2 function of ERalpha D351Y lacks detectable tamoxifen-dependent activity when tested in isolation but does synergize with AF-1 to allow enhanced tamoxifen response. Weak tamoxifen-dependent interactions between the ERalpha D351Y AF-2 function and GRIP1, a representative p160, can be detected in glutathione S-transferase binding assays and mammalian two-hybrid assays. Furthermore, tamoxifen-dependent AF-2 activity can be detected in the presence of ERalpha D351Y and high levels of overexpressed GRIP1. We therefore propose that the D351Y mutation allows weak tamoxifen-dependent AF-2 activity but that this activity is only detectable when AF-1 is strong, and AF-1 and AF-2 synergize, or when p160s are overexpressed. We discuss the possible structural basis of this effect.
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PMID:An antiestrogen-responsive estrogen receptor-alpha mutant (D351Y) shows weak AF-2 activity in the presence of tamoxifen. 1098 90

Activated estrogen receptor alpha (ERalpha) modulates transcription triggered by the transcription factor activator protein-1 (AP-1), which consists of Jun-Jun homodimers and Jun-Fos heterodimers. Previous studies have demonstrated that the interference occurs without binding of ERalpha to DNA but probably results from protein.protein interactions. However, involvement of a direct interaction between ERalpha and AP-1 is still debated. Using glutathione S-transferase pull-down assays, we demonstrated that ERalpha bound directly to c-Jun and JunB but not to FOS family members, in a ligand-independent manner. The interaction could occur when c-Jun was bound onto DNA, as shown in a protein-protein-DNA assay. It implicated the C-terminal part of c-Jun and amino acids 259-302 present in the ERalpha hinge domain. ERalpha but not an ERalpha mutant deleted of amino acids 250-303 (ER241G), also associated with c-Jun in intact cells, in the presence of estradiol, as shown by two-hybrid and coimmunoprecipitation assays. We also show that ERalpha, c-Jun, and the p160 coactivator GRIP1 can form a multiprotein complex in vitro and in intact cells and that the ERalpha.c-Jun interaction could be crucial for the stability of this complex. VP16-ERalpha and c-Jun, which both interact with GRIP1, had synergistic effect on GAL4-GRIP1-induced transcription in the presence of estradiol, and this synergistic effect was not observed with the ERalpha mutant VP16-ER241G or when c-Fos, which bound GRIP1 but not ERalpha, was used instead of c-Jun. Finally, ER241G was inefficient for regulation of AP-1 activity, and an ERalpha truncation mutant encompassing the hinge domain had a dominant negative effect on ERalpha action. These results altogether demonstrate that ERalpha can bind to c-Jun in vitro and in intact cells and that this interaction, by stabilizing a multiprotein complex containing p160 coactivator, is likely to be involved in estradiol regulation of AP-1 responses.
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PMID:Characterization of the physical interaction between estrogen receptor alpha and JUN proteins. 1147 71

Brain natriuretic peptide (BNP) gene expression is a well documented marker of hypertrophy in the cardiac myocyte. Triiodothyronine (T(3)), the bioactive form of thyroid hormone, triggers a unique form of hypertrophy in cardiac myocytes that accompanies the selective activation or suppression of specific gene targets. In this study, we show that the BNP gene is a target of T(3) action. BNP secretion was increased 6-fold, BNP mRNA levels 3-fold, and BNP promoter activity 3-5-fold following T(3) treatment. This was accompanied by an increase in myocyte size, sarcomeric organization, and protein synthesis. Of note, several of the responses to T(3) synergized with those to the conventional hypertrophic agonist endothelin. The response to the liganded thyroid hormone receptor (TR) was mediated by an unusual thyroid hormone response element located between -1000 and -987 relative to the transcription start site. Both TR homodimers and TR.retinoid X receptor heterodimers associated with this element in an electrophoretic mobility shift assay. Protein fragments harboring the LXXLL motifs of the coactivators GRIP1 and SRC1 or TRAP220 interacted predominantly with the TR.retinoid X receptor heterodimeric pair in a ligand-dependent fashion. Both TR homodimers and heterodimers in the unliganded state selectively associated with glutathione S-transferase-nuclear receptor corepressor fragments harboring one of three receptor interaction domains containing the sequence (I/L)XX(I/V)I. These interactions were dissociated following the addition of T(3). Collectively, these findings identify the BNP gene as a potential model for the investigation of TR-dependent gene regulation in the heart.
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PMID:Triiodothyronine increases brain natriuretic peptide (BNP) gene transcription and amplifies endothelin-dependent BNP gene transcription and hypertrophy in neonatal rat ventricular myocytes. 1256 79

The induction of CYP2B gene expression by phenobarbital (PB) is mediated by the translocation of the constitutive androstane receptor (CAR) from the cytoplasm to the nucleus. The CAR/RXR heterodimer binds to two DR-4 sites in a complex phenobarbital responsive unit (PBRU) in the CYP2B gene. The short heterodimer partner (SHP), an orphan nuclear receptor that lacks a conventional DNA binding domain, was initially identified by its interaction with CAR. We have examined the role of SHP in CAR-mediated transactivation of the CYP2B gene. Coexpression of SHP inhibited the transactivation of the CYP2B gene by CAR in cultured hepatoma cells and the p160 coactivator GRIP1 reversed the inhibition. The interaction of CAR with SHP was confirmed by GST pulldown experiments. SHP did not block the binding of either CAR/RXR to the PBRU or binding of GRIP1 to the CAR/RXR complex in gel mobility shift assays, but slightly increased CAR/RXR binding and slightly altered the mobility of the CAR/RXR/GRIP1 complex, suggesting an interaction of SHP with these complexes. The presence of SHP in the complexes, however, could not be detected in an antibody supershift assay. Recombinant corepressors mSin3A, SMRT, and HDAC1, but not NCoR1, interacted with GST-SHP but each of these corepressors in liver nuclear extracts bound to GST-SHP. SMRT and NCoR1 inhibited CAR-mediated activation independent of SHP, but mSin3A and HDAC1 had little effect alone, and were additive with SHP. These studies demonstrate that SHP does not inhibit CAR-mediated trans-activation by interfering with DNA binding or by competition with GRIP1. Instead, SHP may either inhibit recruitment of other coactivators by GRIP1 or actively recruit corepressors directly to the CAR/RXR/PBRU complex.
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PMID:Repression of CAR-mediated transactivation of CYP2B genes by the orphan nuclear receptor, short heterodimer partner (SHP). 1500 Jul 48

SMILE (small heterodimer partner interacting leucine zipper protein) has been identified as a corepressor of the glucocorticoid receptor, constitutive androstane receptor, and hepatocyte nuclear factor 4alpha. Here we show that SMILE also represses estrogen receptor-related receptor gamma (ERRgamma) transactivation. Knockdown of SMILE gene expression increases ERRgamma activity. SMILE directly interacts with ERRgamma in vitro and in vivo. Domain mapping analysis showed that SMILE binds to the AF2 domain of ERRgamma. SMILE represses ERRgamma transactivation partially through competition with coactivators PGC-1alpha, PGC-1beta, and GRIP1. Interestingly, the repression of SMILE on ERRgamma is released by SIRT1 inhibitors, a catalytically inactive SIRT1 mutant, and SIRT1 small interfering RNA but not by histone protein deacetylase inhibitor. In vivo glutathione S-transferase pulldown and coimmunoprecipitation assays validated that SMILE physically interacts with SIRT1. Furthermore, the ERRgamma inverse agonist GSK5182 enhances the interaction of SMILE with ERRgamma and SMILE-mediated repression. Knockdown of SMILE or SIRT1 blocks the repressive effect of GSK5182. Moreover, chromatin immunoprecipitation assays revealed that GSK5182 augments the association of SMILE and SIRT1 on the promoter of the ERRgamma target PDK4. GSK5182 and adenoviral overexpression of SMILE cooperate to repress ERRgamma-induced PDK4 gene expression, and this repression is released by overexpression of a catalytically defective SIRT1 mutant. Finally, we demonstrated that ERRgamma regulates SMILE gene expression, which in turn inhibits ERRgamma. Overall, these findings implicate SMILE as a novel corepressor of ERRgamma and recruitment of SIRT1 as a novel repressive mechanism for SMILE and ERRgamma inverse agonist.
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PMID:Transcriptional corepressor SMILE recruits SIRT1 to inhibit nuclear receptor estrogen receptor-related receptor gamma transactivation. 1969 Jan 66