Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: 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)

Coactivators such as TIF2 and SRC-1 modulate the positioning of the dose-response curve for agonist-bound glucocorticoid receptors (GRs) and the partial agonist activity of antiglucocorticoid complexes. These properties of coactivators differ from their initially defined activities of binding to, and increasing the total levels of transactivation by, agonist-bound steroid receptors. We now report that constructs of TIF2 and SRC-1 lacking the two activation domains (AD1 and AD2) have significantly less ability to increase transactivation but retain most of the activity for modulating the dose-response curve and partial agonist activity. Mammalian two-hybrid experiments show that the minimum TIF2 segment with modulatory activity (TIF2.4) does not interact with p300, CREB-binding protein, or PCAF, which also modulates GR activities. DRIP150 and DRIP205 have been implicated in coactivator actions but are unable to modulate GR activities. The absence of synergism by PCAF or DRIP150 with SRC-1 or TIF2, respectively, further suggests that these other factors are not involved. The ability of a TIF2.4 fragment (i.e. TIF2.37), which is not known to interact with proteins, to block the actions of TIF2.4 suggests that an unidentified binder mediates the modulatory activity of TIF2. Pull-down experiments with GST/TIF2.4 demonstrate a direct interaction of TIF2 with GR in a hormone-dependent fashion that requires the receptor interaction domains of TIF2 and is equally robust with agonists and most antiglucocorticoids. These observations, which are confirmed in mammalian two-hybrid assays, suggest that the capacity of coactivators such as TIF2 to modulate the partial agonist activity of antisteroids is mediated by the binding of coactivators to GR-antagonist complexes. In conclusion, the modulatory activity of coactivators with GR-agonist and -antagonist complexes is mechanistically distinct from the ability of coactivators to augment the total levels of transactivation and appears to involve the binding to both GR-steroid complexes and an unidentified TIF2-associated factor(s).
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PMID:Modulation of induction properties of glucocorticoid receptor-agonist and -antagonist complexes by coactivators involves binding to receptors but is independent of ability of coactivators to augment transactivation. 1237 47

p68 RNA helicase has been implicated in a variety of processes, including rearrangement of RNA secondary structures, RNA splicing, gene transcription and tumor development, yet its mechanisms of action are not well understood. In this study, we show that p68 is predominantly localized to the cell nucleus, where it partially colocalizes with the transcriptional coactivator p300. Accordingly, p68 and p300, or the paralogous CREB-binding protein (CBP), coimmunoprecipitate. Similarly, p68 and RNA polymerase II (Pol II) are able to interact in vivo. GST pull-down assays confirmed these interactions in vitro, demonstrating that p68 can interact with several domains of CBP, while CBP/p300 bind to amino acids 176-388 of p68 and RNA Pol II binds to the N-terminal 80 amino acids of p68. Furthermore, p68 stimulates transcription mediated by the C-terminal transactivation domain of CBP. p68 is also able to stimulate TPA oncogene responsive unit (TORU) promoter activity, and p300 acts in synergy with p68. On the other hand, suppression of CBP/p300 function by the adenoviral protein E1A abolishes TORU promoter activation by p68. Altogether, our results suggest the existence of a multiprotein complex in which p68 RNA helicase, CBP/p300 and RNA Pol II jointly promote gene expression.
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PMID:Synergism between p68 RNA helicase and the transcriptional coactivators CBP and p300. 1252 17

Cdc25B is a dual-specific phosphatase that mediates cell cycle progression by activating the cyclin-dependent kinases. It has been shown to possess oncogenic potential. To elucidate its potential contribution to human prostate cancer development, the expression profile of Cdc25B protein in human patients was analysed by immunohistocytochemistry. Cdc25B is frequently overexpressed in human prostate cancer tissues (29 of 30; 97%). In addition, the overexpression is more profound in the tumors of high combined Gleason scores and in late stages. Subsequently, we demonstrated that Cdc25B acts as a coactivator for AR in a hormone-dependent manner in the prostate cancer cell line, LNCaP. This coactivator function, surprisingly, is independent of its cell cycle functions. Cdc25B, on the other hand, directly interacts with AR as evidenced in GST-pull down and mammalian two-hybrid assays. In addition, it is also able to enhance AR-mediated transcription in synergy with other coactivators, including CREB-binding protein (CBP) and p300/CBP associated factor. Therefore, upregulation of Cdc25B in human prostate cancer and its interplay with AR may contribute to prostate cancer development.
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PMID:Overexpression of Cdc25B, an androgen receptor coactivator, in prostate cancer. 1256 65

Assessment of the risk of human exposure to man-made chemicals that bind to hormone receptors has emerged as a major public health issue. Among hormone receptors, nuclear receptors tend to be targets of xenobiotics because their endogenous ligands are small, fat-soluble molecules. Nuclear receptors are ligand-inducible transcriptional factors and regulate the transcriptional activity of various target genes. At the start of the initiation step of transcription, nuclear receptors interact with coactivators (TIF2, SRC1, ACTR, CBP/p300, etc.) in an agonist-dependent manner. Using the interaction of the nuclear receptor with a coactivator, we have developed a novel rapid ligand in vitro screening method that is easy to use and has high sensitivity. This method, called by us the CoA-BAP system, is applicable to most nuclear receptors and is suitable for high-throughput screening because the entire experimental operation can be carried out on a microplate. We used human TIF2 as a coactivator including LXXLL motifs expressed in Escherichia coli as a fusion protein with BAP and nuclear receptor LBD expressed in E. coli as a fusion protein with GST. On a GSH-coupled microplate these proteins were incubated with chemicals and the protein-protein interactions were detected as alkaline phosphatase activity. To date we have examined seven nuclear receptors (ERalpha/beta, TRalpha, RARalpha/gamma, RXRalpha,and VDR) and confirmed that the method works well.
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PMID:Basis of a high-throughput method for nuclear receptor ligands. 1286 36

Hypoxia inducible factor 1 (HIF-1) is a heterodimeric transcriptional complex that plays pivotal role in the regulation of cellular utilization of oxygen as well as glucose and is an essential regulator of angiogenesis in solid tumor and ischemic disorders. Recently HIF-1alpha, a subunit of HIF-1 complex, was characterized as a potential target for S-nitrosation, providing no information about the impact of this posttranslational modification on the protein transactivation. Cys-800 of HIF-1alpha protein has reactive SH-group, which is critical for the recruitment of p300 co-activator that is necessary for transcriptional activity of HIF-1 complex. Here we report that S-nitrosation of Cys-800 activates HIF-1alpha-p300 interaction and stimulates protein transactivation. We have found that S-nitrosation of the HIF-1alpha C-terminal domain by nitric oxide derived from donors and nitric oxide synthase increases protein transcriptional activity. The increase of HIF-1 transcriptional activity was not observed in the case of Cys-800 substitution to Ala, though other protein thiol groups were nitrosated. Experiments with GST pull-down assay suggest that S-nitrosation of Cys-800 stimulates the recruitment of p300 co-activator protein to the HIF-1alpha C-terminal domain.
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PMID:S-nitrosation of Cys-800 of HIF-1alpha protein activates its interaction with p300 and stimulates its transcriptional activity. 1291 34

Epstein-Barr virus (EBV) nuclear antigen 3C (EBNA3C) is critical for EBV immortalization of infected B lymphocytes and can coactivate the EBV LMP1 promoter with EBNA2. EBNA3C amino acids 365 to 545 are necessary and sufficient for coactivation and are required for SUMO-1 and SUMO-3 interaction. We found that EBNA3C but not EBNA3CDelta343-545 colocalized with SUMO-1 in nuclear bodies and was modified by SUMO-2, SUMO-3, and SUMO-1. EBNA3C amino acids 545 to 628 and amino acids 30 to 365 were also required for EBNA3C sumolation and nuclear body localization but were dispensable for coactivation, indicating that EBNA3C sumolation is not required for coactivation. Furthermore, EBNA3C amino acids 476 to 992 potently coactivated with EBNA2 but EBNA3C amino acids 516 to 922 lacked activity, indicating that amino acids 476 to 515 are critical for coactivation. EBNA3C amino acids 476 to 515 include DDDVIEV(507-513), which are similar to SUMO-1 EEDVIEV(84-90). EBNA3C m1 and m2 point mutations, DDD(507-509) mutated to AAA and DVIEVID(509-513) mutated to AVIAVIA, respectively, diminished SUMO-1 and SUMO-3 interaction in directed yeast two-hybrid and glutathione S-transferase pulldown assays. Furthermore, EBNA3C m1 and m2 did not coactivate the LMP1 promoter with EBNA2. Overexpression of wild-type SUMO-1, SUMO-3, and the SUMO-conjugating enzyme UBC9 coactivated the LMP1 promoter with EBNA2. Since EBNA2 activation is dependent on p300/CBP, the possible effect of EBNA3C on p300-mediated transcription was assayed. EBNA3C potentiated transcription of p300 fused to a heterologous DNA binding domain, whereas EBNA3C m1 and m2 did not. All of these data are consistent with a model in which EBNA3C upregulates EBNA2-mediated gene activation by binding to a sumolated repressor and inhibiting repressive effects on p300/CBP and other transcription factor(s) at EBNA2-regulated promoters.
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PMID:EBNA3C coactivation with EBNA2 requires a SUMO homology domain. 1467 Nov 18

Depletion of peroxisome proliferator-activated receptor gamma (PPARgamma) accompanies myofibroblastic transdifferentiation of hepatic stellate cells (HSC), the primary cellular event underlying liver fibrogenesis. The treatment of activated HSC in vitro or in vivo with synthetic PPARgamma ligands suppresses the fibrogenic activity of HSC. However, it is uncertain whether PPARgamma is indeed a molecular target of this effect, because the ligands are also known to have receptor-independent actions. To test this question, the present study examined the effects of forced expression of PPARgamma via an adenoviral vector on morphologic and biochemical features of culture-activated HSC. The vector-mediated expression of PPARgamma itself is sufficient to reverse the morphology of activated HSC to the quiescent phenotype with retracted cytoplasm, prominent dendritic processes, reduced stress fibers, and accumulation of retinyl palmitate. These effects are abrogated by concomitant expression of a dominant negative mutant of PPARgamma that prevents transactivation of but not binding to the PPAR response element. PPARgamma expression also inhibits the activation markers such as the expression of alpha-smooth muscle actin, type I collagen, and transforming growth factor beta1; DNA synthesis; and JunD binding to the activator protein-1 (AP-1) site and AP-1 promoter activity. Inhibited JunD activity by PPARgamma is not due to reduced JunD expression or JNK activity or to a competition for p300. But it is due to a JunD-PPARgamma interaction as demonstrated by co-immunoprecipitation and glutathione S-transferase pull-down analysis. Further, the use of deletion constructs reveals that the DNA binding region of PPARgamma is the JunD interaction domain. In summary, our results demonstrate that the restoration of PPARgamma reverses the activated HSC to the quiescent phenotype and suppresses AP-1 activity via a physical interaction between PPARgamma and JunD.
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PMID:Peroxisome proliferator-activated receptor gamma induces a phenotypic switch from activated to quiescent hepatic stellate cells. 1470 44

The transcriptional co-activator CBP [CREB (cAMP-response-element-binding protein)-binding protein] and its paralogue p300 play a key role in the regulation of both activity and stability of the tumour suppressor p53. Degradation of p53 is mediated by the ubiquitin ligase MDM2 (mouse double minute protein) and is also reported to be regulated by CBP/p300. Direct protein-protein interaction between a central domain of MDM2 and the TAZ1 (transcriptional adaptor zinc-binding domain) [C/H1 (cysteine/histidine-rich region 1)] domain of p300 and subsequent formation of a ternary complex including p53 have been reported previously. We expressed and purified the proposed binding domains of HDM2 (human homologue of MDM2) and CBP, and examined their interactions using CD spectroscopy. The binding studies were extended by using natively purified GST (glutathione S-transferase)-p300 TAZ1 and GST-p53 fusion proteins, together with in vitro translated HDM2 fragments, under similar solution conditions to those in previous studies, but omitting added EDTA, which causes unfolding and aggregation of the zinc-binding TAZ1 domain. Comparing the binding properties of the known TAZ1 interaction partners HIF-1alpha (hypoxia-inducible factor 1), CITED2 (CBP/p300-interacting transactivator with glutamic- and aspartic-rich tail) and STAT2 (signal transducer and activator of transcription 2) with HDM2, our data suggest that TAZ1 in its native state does not serve as a specific recognition domain of HDM2. Rather, unfolded TAZ1 and HDM2 proteins have a high tendency to aggregate, and non-specific protein complexes are formed under certain conditions.
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PMID:The CBP/p300 TAZ1 domain in its native state is not a binding partner of MDM2. 1527 Jul

Expression of human T-cell leukemia virus type 1 (HTLV-1) is regulated by the viral transcriptional activator Tax. Tax activates viral transcription through interaction with the cellular transcription factor CREB and the coactivators CBP/p300. In this study, we have analyzed the role of histone deacetylase 1 (HDAC1) on HTLV-1 gene expression from an integrated template. First we show that trichostatin A, an HDAC inhibitor, enhances Tax expression in HTLV-1-transformed cells. Second, using a cell line containing a single-copy HTLV-1 long terminal repeat, we demonstrate that overexpression of HDAC1 represses Tax transactivation. Furthermore, a chromatin immunoprecipitation assay allowed us to analyze the interaction of transcription factors, coactivators, and HDACs with the basal and activated HTLV-1 promoter. We demonstrate that HDAC1 is associated with the inactive, but not the Tax-transactivated, HTLV-1 promoter. In vitro and in vivo glutathione S-transferase-Tax pull-down and coimmunoprecipitation experiments demonstrated that there is a direct physical association between Tax and HDAC1. Importantly, biotinylated chromatin pull-down assays demonstrated that Tax inhibits and/or dissociates the binding of HDAC1 to the HTLV-1 promoter. Our results provide evidence that Tax interacts directly with HDAC1 and regulates binding of the repressor to the HTLV-1 promoter.
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PMID:Tax relieves transcriptional repression by promoting histone deacetylase 1 release from the human T-cell leukemia virus type 1 long terminal repeat. 1519 48

We have studied enzymes involved in histone arginine methylation in the filamentous fungus Aspergillus nidulans. Three distinct protein arginine methyltransferases (PRMTs) could be identified, which all exhibit intrinsic histone methyltransferase activity when expressed as glutathione S-transferase (GST) fusion proteins. Two of these proteins, termed RmtA (arginine methyltransferase A) and RmtC, reveal significant sequence homology to the well-characterized human proteins PRMT1 and PRMT5, respectively. Native as well as recombinant RmtA is specific for histone H4 with arginine 3 as the methylation site. Furthermore, methylation of histone H4 by recombinant RmtA affects the acetylation by p300/CBP, supporting an interrelation of histone methylation and acetylation in transcriptional regulation. The second methyltransferase, named RmtB, is only distantly related to human/rat PRMT3 and must be considered as a member of a separate group within the PRMT family. The 61 kDa protein, expressed as a GST fusion protein, exhibits a unique substrate specificity in catalyzing the methylation of histones H4, H3, and H2A. Unlike human PRMT3, the Aspergillus enzyme lacks a Zn-finger domain in the amino-terminal part indicating functional differences of RmtB. Furthermore, phylogenetic analysis indicated that RmtB together with other fungal homologues is a member of a separate group within the PRMT proteins. The existence of in vivo arginine methylation on histones as demonstrated by site-specific antibodies and the high level and specificity of PRMTs for individual core histones in A. nidulans suggests an important role of these enzymes for chromatin modulating activities.
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PMID:Histone methyltransferases in Aspergillus nidulans: evidence for a novel enzyme with a unique substrate specificity. 1531 44


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