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

Chicken ovalbumin upstream promoter-transcription factors (COUP-TFs) are orphan receptors that belong to the steroid/thyroid hormone receptor (TR) superfamily and can repress the transcriptional activity of several target genes; however, the precise mechanism of this repression is unknown. Transfection of a Gal4 DNA-binding domain fused to the putative ligand-binding domain of COUP-TFI (Gal4-COUP-TFI) significantly represses the basal transcriptional activity of a reporter gene containing Gal4-binding sites. Cotransfection of COUP-TFI can relieve the Gal4-COUP-TFI repression in a dose-dependent manner. In contrast, COUP-TFI delta35, which lacks the repressor domain (the C-terminal 35 amino acids), fails to relieve this repression. This finding suggests that the repressor domain of COUP-TFI may squelch a limiting amount of corepressor in HeLa cells. In addition, increasing concentrations of TRbeta also can relieve the COUP-TFI repression in a hormone-sensitive manner. Similarly, overexpression of increasing concentration of COUP-TFI, but not COUP-TFI delta35, can squelch the silencing activity of the unliganded TRbeta. Collectively, these results indicate that COUP-TFI and TRbeta share a common corepressor(s) for their silencing activity. To determine which corepressor is involved in the COUP-TF-silencing activity, we used a yeast two-hybrid and in vitro GST pull-down assays to demonstrate that COUP-TFI can interact with the fragment of N-CoR (nuclear receptor-corepressor) encoding amino acids 921-2453 and the fragments of SMRT (silencing mediator for retinoic acid receptor and TR) encoding amino acids 29-564 and 565-1289, respectively. Interestingly, the fragment of SMRT encoding amino acids 1192-1495, which strongly interacts with TRbeta, interacts very weakly with COUP-TFI. Furthermore, overexpression of N-CoR or SMRT potentiates the silencing activity of COUP-TFI and can relieve the COUP-TFI-mediated squelching of Gal4-COUP-TFI activity. Therefore, our studies indicate that N-CoR and SMRT act as corepressors for the COUP-TFI silencing activity.
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PMID:Gene silencing by chicken ovalbumin upstream promoter-transcription factor I (COUP-TFI) is mediated by transcriptional corepressors, nuclear receptor-corepressor (N-CoR) and silencing mediator for retinoic acid receptor and thyroid hormone receptor (SMRT). 917 Dec 35

EBNA2 is essential for Epstein-Barr virus (EBV) immortalization of B lymphocytes. EBNA2 functions as a transcriptional activator and targets responsive promoters through interaction with the cellular DNA binding protein CBF1. We have examined the mechanism whereby EBNA2 overcomes CBF1-mediated transcriptional repression. A yeast two-hybrid screen performed using CBF1 as the bait identified a protein, SKIP, which had not previously been recognized as a CBF1-associated protein. Protein-protein interaction assays demonstrated contacts between SKIP and the SMRT, CIR, Sin3A, and HDAC2 proteins of the CBF1 corepressor complex. Interestingly, EBNA2 also interacted with SKIP in glutathione S-transferase affinity and mammalian two-hybrid assays and colocalized with SKIP in immunofluorescence assays. Interaction with SKIP was not affected by mutation of EBNA2 conserved region 6, the CBF1 interaction region, but was abolished by mutation of conserved region 5. Mutation of conserved region 5 also severely impaired EBNA2 activation of a reporter containing CBF1 binding sites. Thus, interaction with both CBF1 and SKIP is necessary for efficient promoter activation by EBNA2. A model is presented in which EBNA2 competes with the SMRT-corepressor complex for contacts on SKIP and CBF1.
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PMID:A role for SKIP in EBNA2 activation of CBF1-repressed promoters. 1064 67

Retinoid receptor-related testis-associated receptor (RTR)/germ cell nuclear factor is a nuclear orphan receptor that plays an important role in the control of gene expression during early embryonic development and gametogenesis. It has been shown to repress transcriptional activation. In this study, we further characterize this repressor function. We demonstrate that RTR can suppress the transcriptional activation induced by the estrogen receptor related-receptor alpha1 through its response element. The latter is at least in part due to competition for binding to the same response element. In addition, RTR inhibits basal transcriptional activation, indicating that it functions as an active repressor. Mammalian two-hybrid analyses showed that RTR interacts with the co-repressor nuclear co-repressor (N-CoR) but is unable to interact with the co-repressor SMRT or RIP140. Pull-down analyses with glutathione S-transferase-RTR fusion protein demonstrated that RTR physically interacts with N-CoR in vitro, suggesting a potential role for N-CoR in the transcriptional repression by RTR. To identify the regions in RTR essential for the binding of RTR to N-CoR, the effect of various deletion and point mutations on this interaction was examined. This analysis revealed that this interaction requires the hinge domain, helix 3 as well as the helix 12 region of RTR. The residues Ser(246)-Tyr(247) in the hinge domain, Lys(318) in helix 3, and Lys(489)-Thr(490) in helix 12 are identified as being critical in this interaction. Our results demonstrate that RTR can function as an active transcriptional repressor and that this repression can be mediated through interactions with the co-repressor N-CoR. We show that this interaction exhibits several characteristics unique to RTR. Through its repressor function, RTR can suppress the induction of transcriptional activation by other nuclear receptors. These repressor activities may provide important mechanisms by which RTR regulates gene expression during development and gametogenesis.
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PMID:Characterization of the repressor function of the nuclear orphan receptor retinoid receptor-related testis-associated receptor/germ cell nuclear factor. 1094 Mar 6

Octamer transcription factor-1 (Oct-1) is a member of the POU (Pit-1, Oct-1, unc-86) family of transcription factors and is involved in the transcriptional regulation of a variety of gene expressions related to cell cycle regulation, development, and hormonal signals. It has been shown that Oct-1 acts not only as a transcriptional activator but also as a transcriptional repressor for certain genes. The mechanism of the repressive function of Oct-1 has not been well understood. Here we demonstrate by using the glutathione S-transferase pull-down assays and coimmunoprecipitation assays that the POU domain of Oct-1 directly interacts with a silencing mediator for retinoid and thyroid hormone receptors (SMRT). The interaction surfaces are located in the C-terminal region of SMRT, which are different from previously described silencing domains I and II or receptor interacting domains I and II. In transient transfection assays in COS1 cells, overexpression of SMRT attenuated the augmentation of Oct-1 transcriptional activity by OBF-1/OCA-B, activator for Oct-1. In pull-down assays, increasing amounts of SMRT could compete the binding of OCA-B to Oct-1 POU domain. The activity of Oct-1 could be determined by a regulated balance between SMRT and OCA-B. Furthermore, cotransfected unliganded thyroid hormone receptor enhanced the transactivation by Oct-1, and addition of 3,3',5-tri-iodo-l-thyronine obliterated the stimulatory effects. Consequently, in the presence of cotransfected thyroid hormone receptor, the octamer response element acts as an element negatively regulated by 3,3',5-tri-iodo-l-thyronine. The results suggest that the transcriptional activity of Oct-1 can be modulated by interaction through its POU domain by a silencing mediator SMRT resulting in the cross-talk between Oct-1 and nuclear receptors.
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PMID:Silencing mediator for retinoid and thyroid hormone receptors interacts with octamer transcription factor-1 and acts as a transcriptional repressor. 1113 19

Calmodulin-dependent protein kinase IV (CaMKIV) is a key mediator of Ca(2+)-induced gene expression. In this study, CaMKIV was found to directly associate with and phosphorylate the nuclear factor-kappaB (NFkappaB) component p65 both in vitro and in vivo. The phosphorylation of p65 by CaMKIV resulted in recruitment of transcription coactivator cAMP-response element-binding protein-binding protein and concomitant release of corepressor silencing mediator for retinoid and thyroid hormone receptors, as demonstrated by the glutathione S-transferase pull down and mammalian two hybrid assays. In addition, cotransfection of CaMKIV resulted in cytosolic translocation of the silencing mediator for retinoid and thyroid hormone receptors. Consistent with these results, cotransfected CaMKIV dramatically stimulated the NFkappaB transactivation in mammalian cells. From these results, NFkappaB is suggested to be a novel downstream effector molecule of CaMKIV.
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PMID:Ca2+/calmodulin-dependent protein kinase IV stimulates nuclear factor-kappa B transactivation via phosphorylation of the p65 subunit. 1127 68

Nuclear mitotic apparatus protein-retinoic acid receptor alpha (NuMA-RARalpha) is the fourth of five fusion proteins identified in acute promyelocytic leukemia (APL) patients. The molecular basis for its oncogenic activity has not been delineated. In gel-shift assays, NuMA-RARalpha bound to retinoic acid response elements (RAREs) both as a homodimer and as a heterodimer with RXRalpha. The binding profile of NuMA-RARalpha to a panel of RAREs was very similar to PML-RARalpha and PLZF-RARalpha. In transient transfection assays using HepG2 cells, NuMA-RARalpha inhibited wild-type RARalpha transcriptional activity, while it augmented STAT3 transcriptional activity. In GST-pull down experiments, NuMA-RARalpha formed a complex with the corepressor SMRT, was released from the NuMA-RARalpha/SMRT complexes by all-trans retinoic acid (ATRA) at 10(-7)-10(-6) M and became associated with the coactivator TRAM-1 at 10(-8) M ATRA. Studies comparing NuMA-RARalpha with NuMA-RARalpha(deltaCC) demonstrated that the dimerization or alpha-helical coiled-coil domain of NuMA was required for homodimer formation, transcriptional repression of wild-type RARalpha, transcriptional activation of STAT3, and stability of the NuMA-RARalpha/SMRT complex. Confocal fluorescent microscopy of HeLa cells was performed following transient expression of cyan fluorescent protein (CFP)-tagged proteins and incubation of cells with or without ATRA. Within the nucleus, CFP-NuMA-RARalpha exhibited a speckled pattern identical to that observed in cells transfected with CFP-NuMA. Furthermore, CFP-NuMA-RARalpha colocalized with yellow fluorescent protein-tagged (YFP)-NuMA. In contrast, CFP-NuMA-RARalpha(deltaCC) exhibited a diffuse granular pattern within the nucleus, similar to RARalpha. These results indicate that the dimerization domain of NuMA-RARalpha is critical for each of the known oncogenic activities of NuMA fusion proteins as well as its sequestration to nuclear sites normally occupied by NuMA and is distinct from RARalpha.
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PMID:Essential role for the dimerization domain of NuMA-RARalpha in its oncogenic activities and localization to NuMA sites within the nucleus. 1258 66

The orphan nuclear receptor hB1F (also known as NR5A2, LRH-1, FTF or CPF) plays important roles in regulating the expression of several cellular and viral genes actively involved in a wide range of biological processes such as the bile acid biosynthesis, liver specific gene regulatory network and hepatitis B virus replication. The activity of nuclear receptors is regulated by multiple mechanisms, including coactivation and corepression. In this study, it was found that the silencing mediator for retinoic acid receptor and thyroid hormone receptor (SMRT) specifically represses the transcriptional activity of hB1F, on either GAL4 dependent reporter system or the hB1F-responsive HBV enhancer II/core promoter. The repression imposed by SMRT is observed in different cell lines. Interestingly, hB1F couldn t interact with SMRT directly, as demonstrated by mammalian two-hybrid analysis or GST pull-down assay. Taken together, it can be concluded for the first time that the transcriptional activity of hB1F is regulated specifically by the corepressor SMRT via an indirect mechanism.
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PMID:Corepressor SMRT specifically represses the transcriptional activity of orphan nuclear receptor hB1F/hLRH-1. 1451 6

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

This study describes the physical and functional interactions between ICP0 of herpes simplex virus type 1 and class II histone deacetylases (HDACs) 4, 5, and 7. Class II HDACs are mainly known for their participation in the control of cell differentiation through the regulation of the activity of the transcription factor MEF2 (myocyte enhancer factor 2), implicated in muscle development and neuronal survival. Immunofluorescence experiments performed on transfected cells showed that ICP0 colocalizes with and reorganizes the nuclear distribution of ectopically expressed class I and II HDACs. In addition, endogenous HDAC4 and at least one of its binding partners, the corepressor protein SMRT (for silencing mediator of retinoid and thyroid receptor), undergo changes in their nuclear distribution in ICP0-transfected cells. As a result, during infection endogenous HDAC4 colocalizes with ICP0. Coimmunoprecipitation and glutathione S-transferase pull-down assays confirmed that class II but not class I HDACs specifically interacted with ICP0 through their amino-terminal regions. This region, which is not conserved in class I HDACs but homologous to the MITR (MEF2-interacting transcription repressor) protein, is responsible for the repression, in a deacetylase-independent manner, of MEF2 by sequestering it under an inactive form in the nucleus. Consequently, we show that ICP0 is able to overcome the HDAC5 amino-terminal- and MITR-induced MEF2A repression in gene reporter assays. This is the first report of a viral protein interacting with and controlling the repressor activity of class II HDACs. We discuss the putative consequences of such an interaction for the biology of the virus both during lytic infection and reactivation from latency.
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PMID:Functional interaction between class II histone deacetylases and ICP0 of herpes simplex virus type 1. 1519 49

Transcriptional repression is a major regulatory mechanism in cell differentiation, organogenesis, and oncogenesis. Two repressors of ligand-dependent transcription factors, nuclear receptor corepressor (N-CoR) and the related protein SMRT were identified as a silencing mediator for thyroid hormone receptor beta and as a silencing mediator for retinoic acid and thyroid hormone receptors, respectively. Nuclear receptor coactivators such as steroid receptor coactivator-1 (SRC-1) contain multiple LXXLL motifs, which are essential and sufficient for its ligand-dependent interaction with nuclear receptors. N-CoR also has an LXXLL motif, located between repressor domains 1 and 2, and conserved between mouse and man. In contrast, SMRT lacks this motif. This paper describes functional implications of the LXXLL motif in N-CoR. A 57-amino acid portion of N-CoR containing the LDNLL sequence (N-CoR(LDNLL)) fused to GST interacted with retinoic acid receptor alpha (RARalpha) and thyroid hormone receptor beta (TRbeta) in vitro. Similarly, [(35)S-methionine]N-CoR(LDNLL) interacted with a RARalpha fusion protein. N-CoR(LDNLL) also bound to RARalpha in vivo as determined in mammalian one-hybrid system in transfected CV-1 cells and by two-hybrid assays in bacteria. The interaction with RARalpha in vitro and in vivo was specific as determined by mutation of the sequence LDNLL to LDNAA. Our data suggest that the LDNLL motif in N-CoR has functional significance because it mediates interaction with nuclear receptors such as RARalpha and TRbeta.
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PMID:Functional analyses of an LXXLL motif in nuclear receptor corepressor (N-CoR). 1533 96


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