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)

Diacylglycerol (DG) and its analogue phorbol 12-myristate 13-acetate (PMA) activate the ubiquitous phospholipid/Ca2(+)-dependent protein kinase, protein kinase C (PKC), and cause it to become tightly associated with membranes. DG is produced transiently as it is rapidly metabolized by DG kinase (DGK) to phosphatidic acid. Phorbol esters such as PMA are not metabolized and induced a prolonged membrane association of PKC. Until recently, PKC was the only known phorbol ester receptor. We have shown that a novel brain-specific cDNA, neuronal chimaerin (NC), expressed in Escherichia coli, binds phorbol ester with high affinity, stereospecificity and a phospholipid requirement [Ahmed, Kozma, Monfries, Hall, Lim, Smith & Lim (1990) Biochem. J. 272, 767-773]. The proteins NC, PKC and DGK possess a cysteine-rich domain with the motif HX11/12CX2CXnCX2CX4HX2CX6/7C (where n varies between 12 and 14). The partial motif, CX2CX13CX2C, is present in a number of transcription factors including the steroid hormone receptors and the yeast protein, GAL4, in which zinc plays a structural role of co-ordinating cysteine residues and is essential for DNA binding (protein-nucleic acid interactions). The cysteine-rich domain of NC and PKC is required for phospholipid-dependent phorbol is required for phospholipid-dependent phorbol ester binding, suggesting an involvement of this domain in protein-lipid interactions. We have expressed recombinant NC, PKC and DGK glutathione S-transferase and TrpE fusion proteins in E. coli to investigate the relationship between the cysteine-rich motif, HX11/12CX2CX10-14CX2CX4HX2CX6/7C, zinc and phorbol ester binding. The cysteine-rich domain of NC, PKC and DGK bound 65Zn2+ but only NC and PKC bound [3H]phorbol 12,13-dibutyrate. When NC and PKC were subjected to treatments known to remove metal ions from GAL4 and the human glucocorticoid receptor, phorbol ester binding was inhibited. These data provide evidence for the role of a zinc-dependent structure in phorbol ester binding.
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PMID:The cysteine-rich domain of human proteins, neuronal chimaerin, protein kinase C and diacylglycerol kinase binds zinc. Evidence for the involvement of a zinc-dependent structure in phorbol ester binding. 166 Feb 66

The Ah receptor (AhR) and Ah receptor nuclear translocator (Arnt) heterodimer bind the xenobiotic-responsive element (XRE) sequence in the upstream region of the genes for some drug-metabolizing enzymes, such as P4501A1 and glutathione S-transferase Ya, to activate their transcription. This paper describes transcriptional activation domains of the AhR and Arnt as examined in vivo by DNA transfection experiments using GAL4-AhR or GAL4-Arnt chimeric plasmids and a reporter plasmid containing five GAL4 DNA binding sites. The major activation domain of Arnt was localized in a short segment of the C-terminal 34 amino acids, while the glutamine-rich domain of Arnt showed no transcriptional activity. This activation domain of Arnt could be further divided into two subdomains with some sequence similarity. Point mutation analysis of one of the subdomains revealed that bulky hydrophobic amino acids and neighboring acidic amino acids were necessary for the transcription-enhancing activity of Arnt. The C-terminal half of the AhR showed a strong transcription-stimulating activity, apparently five times as strong as that of Arnt. Further analysis of the activity revealed that the C-terminal transcriptional activity was distributed in several activation domains, one of which is rich in glutamine residues. These results indicate that the glutamine-rich domains of the AhR and Arnt function differently in the heterodimer regulatory complex. Previously, we showed that the enhancer activity of XRE was repressed by E1A proteins, especially the 12S form of E1A. Cotransfection experiments using an E1A12S expression plasmid and a GAL4-AhR or GAL4-Arnt expression plasmid demonstrated that E1A protein rather predominantly inhibited the transcriptional activity of Arnt.
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PMID:Transcriptional activation domains of the Ah receptor and Ah receptor nuclear translocator. 755 46

A human protein that is 92% identical and 97% homologous at the amino acid level to RanBP1 from mouse was identified by the two-hybrid method, using two types of target cDNAs fused to sequences encoding the GAL4 DNA-binding domain. The target cDNAs encoded the human Ran/TC4 and human RCC1 proteins, respectively. An in vitro binding experiment showed that RanBP1 binds to RCC1 with the aid of Ran. Partially purified, GST-fused RanBP1 inhibited RCC1-stimulated guanine nucleotide release from Ran in vitro. Consistent with this in vitro finding, overproduction of human RanBP1 was detrimental to growth of tsBN2, a temperature-sensitive BHK21 hamster cell line defective in the RCC1 gene, and inhibited the growth of the Saccharomyces cerevisiae rcc1 mutants prp20, mtr1 and srm1. The specific effect of RanBP1 on rcc1- cells was confirmed by the finding that overproduction of RanBP1 induces significant levels of expression of a FUS1-lacZ gene and an increase in mating efficiencies in a ste3, pheromone receptor-deficient yeast mutant. This phenotype is similar to the srm1, a mutant isolated as a suppressor that restores mating to receptorless mutants. These findings indicate that RanBP1 negatively regulates RCC1.
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PMID:RanBP1, a Ras-like nuclear G protein binding to Ran/TC4, inhibits RCC1 via Ran/TC4. 761 57

We previously showed that v-Rel, the oncoprotein of the avian retrovirus Rev-T, can increase expression from promoters containing binding sites for the cellular transcription factor Sp1 in chicken embryo fibroblasts (S. Sif, A.J. Capobianco, and T.D. Gilmore, Oncogene 8:2501-2509, 1993). In those experiments, v-Rel appeared to increase the transactivating function of Sp1; that is, v-Rel stimulated transactivation by a GAL4-Sp1 protein that lacked the Sp1 DNA-binding domain. We have now shown that in vitro-synthesized v-Rel and GAL4-Sp1 form a complex that can be immunoprecipitated with either anti-Sp1 or anti-v-Rel antiserum. We have also shown that a glutathione S-transferase (GST)-Sp1 fusion protein can specifically interact with in vitro-translated v-Rel and with in vivo-synthesized v-Rel from transformed chicken spleen cells. In addition, we have found that the abilities of wild-type and two mutant forms of v-Rel to increase transactivation by Sp1 in vivo correlate with their abilities to interact with Sp1 in vitro. The sequences important for the interaction of v-Rel with Sp1 in vitro have been mapped to the first 147 amino acids of v-Rel. Other Rel proteins, such as c-Rel, RelA, p52, and p50, were also able to form a complex with Sp1 in vitro. These results suggest that v-Rel increases expression from Sp1 site-containing promoters by functionally interacting with Sp1 and that cellular Rel proteins and Sp1 are likely to interact to influence transcription from natural promoters.
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PMID:Interaction of the v-Rel oncoprotein with cellular transcription factor Sp1. 793 95

We have used the two-hybrid system to identify proteins that interact with the product of RAD7, a gene involved in DNA repair. A screen of a yeast genomic DNA-GAL4 activation domain (GAD) fusion gene library allowed the isolation of plasmids containing sequences corresponding to the 3' end of the SIR3 gene. This gene is known to be involved in the production of transcriptionally silent DNA at the cryptic mating-type cassettes and at telomeres. The cloned sequences coded for amino acids 307-979 of the Sir3 protein. A sir3 deletion allele, constructed in an isogenic rad7-deletion strain, rescued approximately one-quarter of the UV sensitivity associated with the rad7 deletion, indicating that the two genes interact genetically. Radiolabeled fusion proteins, made with the glutathione S-transferase (GST) gene in the vector pGEX-2T, were purified from Escherichia coli and shown to interact in vitro. This evidence suggests that the Sir3 protein interacts with the Rad7 protein to allow the nucleotide excision repair complex access to transcriptionally inactive chromatin. The proportions of 5-FOA-resistant cells in cultures from isogenic RAD+ and rad7-delta strains containing a telomeric URA3 gene were similar, suggesting that the RAD7 gene is not involved in the production or structure of transcriptionally silent chromatin at the telomeres. RAD7-dependent DNA repair of transcriptionally silent chromatin was shown not to induce expression of a telomeric copy of the URA3 gene, suggesting that repair of transcriptionally silent chromatin differs from transcriptionally active chromatin. Expression of a telomeric copy of the URA3 gene was stimulated in a rad7-delta mutant, suggesting that repair of lesions in the absence of Rad7 can result in the activation of transcriptionally silenced genes.
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PMID:Interaction of the yeast RAD7 and SIR3 proteins: implications for DNA repair and chromatin structure. 795 76

The serum response factor (SRF) is a 67-kDa phosphoprotein that, together with auxiliary factors, modulates transcription of immediate early genes containing serum response elements in their promoters. Here we show that the carboxyl-terminal domain of human SRF is phosphorylated in vivo and is recognized in vitro by the double-stranded DNA-activated serine/threonine-specific protein kinase, DNA-PK. SRF phosphorylation by DNA-PK was stimulated by its cognate binding site. Protein microsequence analysis of a 22-amino acid synthetic SRF peptide and phosphopeptide analysis of genetically altered glutathione S-transferase-SRF fusion proteins identified Ser-435 and Ser-446 of human SRF as sites phosphorylated by DNA-PK. Both serines are followed by glutamine. Changing Gln-436 and Gln-447 to other residues reduced or eliminated phosphorylation by DNA-PK, confirming that these glutamines are important determinants for kinase recognition. The carboxyl-terminal transcription activation domain was mapped within a 71-amino acid region that contains both DNA-PK phosphorylation sites. Amino acid substitutions that interfered with phosphorylation by DNA-PK at Ser-435/446 in GAL4-SRF fusion proteins were reduced in transactivation potency. From these data we suggest that DNA-PK phosphorylation may modulate SRF activity in vivo.
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PMID:The carboxyl-terminal transactivation domain of human serum response factor contains DNA-activated protein kinase phosphorylation sites. 840 51

Addition of mitogenic growth factors to quiescent cells triggers complex signal transduction cascades that result in the reprogramming of gene expression and entry into the cell cycle. We have found that an oncogenic variant of the c-Raf-1 protein kinase stimulated the expression of promoters containing NF-kappa B binding sites. In situ immunofluorescence analysis revealed elevated nuclear levels of the p65 subunit of NF-kappa B in v-raf-transformed NIH 3T3 cells. Incubation of HeLa cell cytoplasmic extracts with a purified recombinant glutathione S-transferase-raf fusion protein in the presence of ATP released active NF-kappa B that could be detected by electrophoretic gel mobility shift assay. Coincubation of purified recombinant I kappa B and glutathione S-transferase-raf in the presence of ATP resulted in the phosphorylation of I kappa B. Coexpression of GAL4 (activation domain)-I kappa B and GAL4 (DNA-binding domain)-raf fusion proteins in yeast resulted in stimulation of a GAL4-responsive reporter gene, indicating that I kappa B and Raf interact physically in vivo. These results indicate that the Raf-1 kinase functions in signal transduction in part by activating the NF-kappa B transcription factor by phosphorylating I kappa B in the cytoplasmic I kappa B-NF-kappa B complex to release active NF-kappa B.
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PMID:Raf-1 protein kinase activates the NF-kappa B transcription factor by dissociating the cytoplasmic NF-kappa B-I kappa B complex. 841 86

We report here that calreticulin interacts with protein disulfide isomerase (PDI). The PDI-calreticulin complex can be dissociated by Zn(2+)-iminodiacetate-substituted Sepharose-agarose chromatography, suggesting that these interactions may be Zn2+-dependent. Direct interaction between calreticulin and PDI is also documented by calreticulin affinity chromatography. PDI was the only pancreatic microsomal protein retained on the calreticulum affinity column. Calreticulin and PDI were identified by their NH2-terminal amino acid sequence analysis, mobilities in SDS-polyacrylamide gel electrophoresis, binding of 45Ca2+, and their reactivity with specific antibodies. Using glutathione S-transferase-calreticulin fusion proteins, we show that PDI interacts strongly with the P-domain and only weakly with the N-domain of calreticulin. Expression of calreticulin domains and PDI as fusion proteins with GAL4 in the yeast two-hybrid system revealed that calreticulin interacted with PDI also under normal cellular conditions. Interaction with PDI required only the NH2-terminal region of the N-domain (amino acid residues 1-83) and the P-domain (amino acid residues 150-240) of calreticulin. Importantly, interaction between calreticulin and PDI led to the modulation of their activities. In the presence of PDI, calreticulin does not bind Ca2+ with high affinity. Calreticulin or the N-domain of calreticulin inhibited PDI ability to refold scrambled RNase A.
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PMID:Interaction of calreticulin with protein disulfide isomerase. 853 5

Increases in transforming growth factor beta1 (TGF-beta1) mRNA and biological activity in the early phase of human cytomegalovirus (CMV) infection in fibroblasts are paralleled by increased TGF-beta1-chloramphenicol acetyltransferase (CAT) reporter gene activity. To determine how CMV infection transactivates the TGF-beta1 promoter, we examined the effects of the cotransfected IE2 regulatory protein of human CMV on 5'-deleted TGF-beta1 promoter-CAT reporter genes in transient DNA transfection assays. Two upstream TGF-beta1 promoter regions each containing an Egr-1 consensus site were shown to be important for IE2-induced transactivation in a cell type that displayed greatly reduced nonspecific activity. Furthermore, transfer of an Egr-l site from between positions -125 and -98, but not point mutant versions of this site, to a heterologous promoter also conveyed IE2 responsiveness. Addition of an IE2 expression vector or use of the U373 A45 astrocytoma cell line expressing IE2 also produced synergistic stimulation of GAL4-Egr-l-mediated activation of a target promoter containing GAL4 binding sites. The 80-kDa IE2 protein present in A45 cells proved to selectively bind to glutathione S-transferase (GST)-Egr-1 beads. The results of in vitro protein binding assays also revealed that an intact in vitro-translated IE2 protein bound directly to the GST-Egr-1 fusion protein through the zinc finger domain of the Egr-1 protein and that this binding activity was abolished by deletion of parts of the zinc finger DNA-binding domain. Similarly, the Egr-1 protein was found to associate preferentially with a small region within the C-terminal half of the IE2 protein adjacent to the DNA-binding and dimerization domains that are important for both transactivation and downregulation. We conclude from these observations that IE2 may regulate transcription of the TGF-beta1 gene as well as other potential cellular targets by virtue of its ability to interact with the Egr-1 DNA-binding protein.
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PMID:The IE2 regulatory protein of human cytomegalovirus induces expression of the human transforming growth factor beta1 gene through an Egr-1 binding site. 879 51

Since the first report documenting that HIV-1 Vpr was involved in the stimulation of transactivation of several unrelated promoters, little additional information has been reported. By using transient transfection experiments, we confirmed and extended these previously reported data. Further in vivo experiments showed that Vpr can co-operatively stimulate transactivation activity of a minimal promoter containing one GAL4 DNA-binding site, when it is co-expressed with different heterologous activator domains fused to GAL4 DNA-binding domain. Thus, Vpr could transactivate in concert with an activator domain, but has no effect on the transactivation of a minimal promoter in the absence of activator protein. To investigate whether Vpr can interact with components of the basal transcriptional machinery, in vitro protein-protein binding assays were performed using either translated, radiolabeled Vpr or TFIIB proteins and glutathione S-transferase Vpr or TFIIB chimeric proteins. We demonstrated that the portion of Vpr ranging from amino acids 15 to 77 interacts specifically with the basal transcription factor TFIIB. Also, our data indicated that the N-terminal domain of TFIIB is required for the interaction.
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PMID:The human immunodeficiency virus type 1 Vpr transactivator: cooperation with promoter-bound activator domains and binding to TFIIB. 880 Feb 8


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