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:1.6.5.2 (NQO1)
6,196 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Deletion mutagenesis and transfection studies into hepatic (mouse hepatoma (Hepa-1) and human hepatoblastoma (Hep-G2)) and nonhepatic (HeLa) cells indicated that high levels of expression of the human NAD(P)H:quinone oxidoreductase gene in tumor cells and its induction by beta-naphthoflavone and 3-(2)-tert-butyl-4-hydroxyanisole are mediated by human antioxidant response element (hARE) located in the region between -470 and -445. The hARE, when attached to the thymidine kinase promoter and transfected into several mammalian cells, expressed high levels of the chloramphenicol acetyltransferase gene that was inducible by beta-naphthoflavone and 3-(2)-tert-butyl-4-hydroxyanisole. Nucleotide sequence analysis of the hARE revealed the presence of a recognition site for binding to the AP1 protein. Mutation of the AP1 binding site located within the hARE resulted in the loss of expression and induction upon transfection into various cell types. Band shift and competition assays with hARE and nuclear extracts from control and beta-naphthoflavone-treated Hepa-1, Hep-G2 and HeLa cells indicated specific interaction of regulatory protein(s) to the hARE. The supershift assays using antibodies against specific proteins of the AP1 family identified Jun-D and c-Fos as two members in the hARE-protein complex observed in band shift assays.
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PMID:Regulation of human NAD(P)H:quinone oxidoreductase gene. Role of AP1 binding site contained within human antioxidant response element. 840 91

NAD(P)H:quinone oxidoreductase (NQO1) is a flavoprotein which catalyzes the two-electron reduction of quinones and azo-dyes and thus prevents the formation of free radicals and toxic oxygen metabolites that may be generated by the one-electron reductions catalyzed by cytochrome P450 reductase. Analysis of RNA indicated 20- to 50-fold higher levels of NQO1 gene expression in the liver tumors and in the tissue surrounding the tumors of patients with hepatocarcinoma than in normal individuals. An approximately 50-fold higher level of NQO1 mRNA was also observed in human hepatoblastoma (Hep-G2) cells than in normal liver. By deletion mutagenesis in the human NQO1 gene promoter and subsequent transfection into hepatic and nonhepatic cell lines, a 1.42 kb DNA segment has been identified to contain cis-acting elements responsible for high levels of expression of the NQO1 gene in tumor cells.
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PMID:High levels of expression of the NAD(P)H:quinone oxidoreductase (NQO1) gene in tumor cells compared to normal cells of the same origin. 165 29

The human NAD(P)H:quinone oxidoreductase (NQO1) gene, 1850 base pairs (bp) of the 5' flanking region, and 67 bp of the 3' flanking region have been sequenced. The human NQO1 gene is approximately 20 kb in length and has six exons interrupted by five introns. The start site of transcription was determined by primer extension analysis. The first exon is 118 bp in length and codes for two amino acids including the initiating methionine and one G for the first codon of the second exon. The sixth exon is the largest among the exons and is 1833 bp in length. The sequence analysis of the sixth exon revealed the presence of four potential polyadenylation signal sequences (AATAAA) and a single copy of human Alu repetitive sequence. The second intron is the smallest of all the introns (116 bp). Nuclear run-on experiments performed using nuclei isolated from 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) treated and untreated human hepatoblastoma (Hep-G2) cells demonstrated that TCDD treatment increases the rate of transcription of endogenous NQO1 gene by 3-fold.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Human NAD(P)H:quinone oxidoreductase (NQO1) gene structure and induction by dioxin. 165 51

NAD(P)H:quinone oxidoreductases (NQOs) are flavoproteins that catalyze the oxidation of NADH or NADPH by various quinones and oxidation-reduction dyes. We have previously described a complementary DNA that encodes a dioxin-inducible cytosolic form of human NAD(P)H:quinone oxidoreductase (NQO1). In the present report we describe the nucleotide sequence and deduced amino acid sequence for a cDNA clone that is likely to encode a second form of NAD(P)H:quinone oxidoreductase (NQO2) which was isolated by screening a human liver cDNA library by hybridization with a NQO1 cDNA probe. The NQO2 cDNA is 976 nucleotides long and encodes a protein of 231 amino acids (Mr = 25,956). The human NQO2 cDNA and protein are 54% and 49% similar to human liver cytosolic NQO1 cDNA and protein, respectively. COS1 cells transfected with NQO2 cDNA showed a 5-7-fold increase in NAD(P)H:quinone oxidoreductase activity as compared to nontransfected cells when either 2,6-dichlorophenolindophenol or menadione was used as substrate. Western blot analysis of the expressed NQO1 and NQO2 cDNA proteins showed cross-reactivity with rat NQO1 antiserum, indicating that NQO1 and NQO2 proteins are immunologically related. Northern blot analysis shows the presence of one NQO2 mRNA of 1.2 kb in control and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) treated human hepatoblastoma Hep-G2 cells and that TCDD treatment does not lead to enhanced levels of NQO2 mRNA as it does for NQO1 mRNA. Southern blot analysis of human genomic DNA suggests the presence of a single gene approximately 14-17 kb in length. The NQO2 gene locus is highly polymorphic as indicated by several restriction fragment length polymorphisms detected with five different restriction enzymes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Nucleotide and deduced amino acid sequence of a human cDNA (NQO2) corresponding to a second member of the NAD(P)H:quinone oxidoreductase gene family. Extensive polymorphism at the NQO2 gene locus on chromosome 6. 169 23

NAD(P)H: quinone oxidoreductase (NQO1) is believed to be protective against cancer and toxicity caused by exposure to quinones and their metabolic precursors. This enzyme catalyzes the two-electron reduction of compounds, compared with one-electron reduction mediated by NADPH: cytochrome-P450 oxidoreductase which produces toxic and mutagenic free radicals. Recently we cloned and sequenced the cDNA encoding human 2.3,7,8-tetrachlorodibenzo-p-dioxin (dioxin)-inducible cytosolic NQO1 [Jaiswal et al. (1988) J. Biol. Chem. 263, 13572-13578] and provided preliminary evidence that this enzyme may correspond to diaphorase 4, an enzymatic activity present in various tissues that catalyzes the reduction of a variety of quinones by both NADH and NADPH [Edwards et al. (1980) Biochem. J. 187, 429-436]. In the present report we characterize the catalytic properties of the protein encoded by the NQO1 cDNA. The enzyme was synthesized in monkey kidney COS-1 cells transfected with a pMT2-based expression plasmid containing the NQO1 cDNA. Western blot analysis of the transfected cells using an antibody against rat liver cytosolic NQO1 revealed a 31-kDa band that was not detected in nontransfected cells. This band corresponded to a polypeptide with the same electrophoretic mobility as the endogenous NQO1 protein detected in the human hepatoblastoma (Hep-G2) cells with the same antibody. The immunoreactive protein detected in human Hep-G2 cells was induced approximately fourfold by exposure of the cultures to dioxin, an increase commensurate with the increased in quinone oxidoreductase activity. These studies suggest that the protein encoded by NQO1 cDNA is indeed similar, if not identical, to the dioxin-inducible protein band detected in human Hep-G2 cells. Further characterization of the product of NQO1 cDNA, which was present at approximately 20-30-fold higher levels in transfected COS cells than the endogenous product in uninduced human Hep-G2 cells indicated that it had very high capacity (greater than 1000-fold over background) to catalyze the reduction of 2.6-dichloroindophenol and menadione. Besides these two commonly used substrates for quinone reductase, the expressed NQO1 protein also effectively metabolized 2,6-dimethylbenzoquinone, methylene blue, p-benzoquinone, 1,4-naphthoquinone, 2-methyl-1,4-benzoquinone, with the latter being the most potent electron acceptor at 50 microM concentration of the substrate.
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PMID:The human dioxin-inducible NAD(P)H: quinone oxidoreductase cDNA-encoded protein expressed in COS-1 cells is identical to diaphorase 4. 189 80

Human NAD(P)H:quinone oxidoreductase2 (NQO2) gene, 1336 base pairs (bp) of the 5'-flanking region and 165 bp of the 3'-flanking region, have been sequenced. NQO2 gene is 20 kilobase pairs in length and have seven exons interrupted by six introns as compared to the previously cloned NQO1 gene which contains six exons. 187 bp of the first exon in the NQO2 gene are noncoding and are absent in the NQO1 gene. 92 bp of the second exon in the NQO2 gene corresponded to the first exon of the NQO1 gene and so on. The sizes and nucleotide sequences of exons 3-6 are highly conserved between NQO2 and NQO1 genes. The last exon in the NQO2 gene is 1603 bp shorter than the last exon of the NQO1 gene and encodes for 58 amino acids as compared to 101 amino acids encoded by the NQO1 gene. This makes NQO2 protein 43 amino acids shorter than the NQO1 protein. The high degree of conservation between NQO2 and NQO1 gene organization and sequence confirmed that NQO2 gene encodes for a second member of the NQO gene family in human. Nucleotide sequence analysis of the 5'-flanking region of the NQO2 gene revealed presence of four SP1 binding sites at positions -214, -170, -106, and -75, a single copy of the antioxidant response element (ARE) at nucleotide -936, and three copies of xenobiotic response element (XRE) at positions -708, -557, and -51. ARE and XRE elements have previously been found in the promoters of the NQO1 and glutathione S-transferase Ya subunit genes and mediate increases in their expression in response to polycyclic aromatic compounds, phenolic antioxidants, and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), respectively. The NQO2 cDNA-derived protein in monkey kidney COS1 cells efficiently catalyzed nitroreduction of anti-tumor compound CB10-200, an analog of nitrophenylaziridine. Northern blot analysis indicates that NQO2 gene is expressed in human heart, brain, lung, liver, and skeletal muscle but does not express in placenta. In contrast, the NQO1 gene was expressed in all human tissues. Large variations were noticed for expression of the NQO2 and NQO1 genes among various tissues, 1336 bp of the 5'-flanking region of the NQO2 gene containing ARE and XRE was found sufficient to increase expression of the CAT gene in response to beta-naphthoflavone and tCDD in transfected human hepatoblastoma (Hep-G2) cells.
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PMID:Human NAD(P)H:quinone oxidoreductase2. Gene structure, activity, and tissue-specific expression. 818 56

NAD(P)H:quinone oxidoreductase 1 (NQO1) is a flavoprotein that catalyzes two-electron reduction and detoxification of quinones. We have shown previously that twenty-four base pairs of the human Antioxidant Response Element (hARE) mediate basal and xenobiotic-induced expression of the NQO1 gene [Li and Jaiswal, J Biol Chem 267: 15097-15104, 1992]. In the present report, we have characterized a second cis-element, AP-2, at nucleotide position -157 of the human NQO1 gene promotor that regulates basal and cAMP-induced transcription of the NQO1 gene. The NQO1 gene AP-2 mediated expression of the chloramphenicol acetyl transferase (CAT) gene and the binding of nuclear proteins to the AP-2 element were observed in HeLa (AP-2 positive) cells but not in human hepatoblastoma Hep-G2 (AP-2 deficient) cells, indicating the involvement of transcription factors AP-2 in the regulation of NQO1 gene expression. Affinity purification of nuclear protein that binds to the NQO1 gene AP-2 DNA element and western analysis revealed that AP-2 indeed binds to the NQO1 gene AP-2 element and regulates its expression HeLa cells. The involvement of AP-2 in the regulation of NQO1 gene expression was confirmed by the observation that cDNA-derived AP-2 protein in Hep-G2 cells increased in NQO1 gene AP-2 but not mutant AP-2 mediated expression of CAT gene in Hep-G2 cells.
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PMID:AP-2-mediated regulation of human NAD(P)H: quinone oxidoreductase 1 (NQO1) gene expression. 860 72

Human breast (MCF-7, HBL 100, T47D, BT20, HS578T), colon (HT29, CACO2, SW620, SW480, COLO320DM) and small cell lung cancer (NCI-N417, OH3, SW2) cell lines were transplanted subcutaneously into severe combined immunodeficient (SCID) mice. When sizeable tumours developed, the mice were sacrificed and the following enzyme activities were detected histochemically: presumed nitric oxide synthase-associated diaphorase (NOSaD), beta-D-glucuronidase (beta-Gluc) and non-specific alkaline phosphatase (alP). Except for HT29 and MCF-7 presumed NOSaD activity was not detected in the tumour itself or in the neo-vasculature of the tumours. beta-Gluc activity was found in all tumour cells (except N417 and COLO 320), in the necrotic parts of the tumours and in stromal cells of the tumour bed. AlP activity was present in all tumours including their necrotic areas. However, the activities of beta-Gluc and alP varied considerably even within one tumour, ranging from very weak to very strong. Principally the results show that the human/SCID mouse tumour model is well suited to test modern applications of tumour therapy involving the enzymes NOSaD, beta-Gluc and alP. In particular, antibody directed enzyme prodrug therapy concepts and activation of prodrugs by enzymes released from tumour cells into the necrotic areas of the tumour can be evaluated in this in vivo model.
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PMID:Histochemistry of therapeutically relevant enzymes in human tumours transplanted into severe combined immunodeficient (SCID) mice: nitric oxide synthase-associated diaphorase, beta-D-glucuronidase and non-specific alkaline phosphatase. 896 Mar 2

Twenty-four base pairs of the human antioxidant response element (hARE) are required for high basal transcription of the NAD(P)H:quinone oxidoreductase1 (NQO1) gene and its induction in response to xenobiotics and antioxidants. hARE is a unique cis-element that contains one perfect and one imperfect AP1 element arranged as inverse repeats separated by 3 bp, followed by a "GC" box. We report here that Jun, Fos, Fra, and Nrf nuclear transcription factors bind to the hARE. Overexpression of cDNA derived combinations of the nuclear proteins Jun and Fos or Jun and Fra1 repressed hARE-mediated chloramphenicol acetyltransferase (CAT) gene expression in transfected human hepatoblastoma (Hep-G2) cells. Further experiments suggested that this repression was due to overexpression of c-Fos and Fra1, but not due to Jun proteins. The Jun (c-Jun, Jun-B, and Jun-D) proteins in all the possible combinations were more or less ineffective in repression or upregulation of hARE-mediated gene expression. Interestingly, overexpression of Nrf1 and Nrf2 individually in Hep-G2 and monkey kidney (COS1) cells significantly increased CAT gene expression from reporter plasmid hARE-thymidine kinase-CAT in transfected cells that were inducible by beta-naphthoflavone and teri-butyl hydroquinone. These results indicated that hARE-mediated expression of the NQO1 gene and its induction by xenobiotics and antioxidants are mediated by Nrf1 and Nrf2. The hARE-mediated basal expression, however, is repressed by overexpression of c-Fos and Fra1.
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PMID:Nrf1 and Nrf2 positively and c-Fos and Fra1 negatively regulate the human antioxidant response element-mediated expression of NAD(P)H:quinone oxidoreductase1 gene. 896 64

Xenobiotics and antioxidants induce expression of detoxifying enzymes including NAD(P)H: quinone oxidoreductase (NQO1), NRH:quinone oxidoreductase (NQO2), and glutathione S-transferase Ya (GST Ya), presumably to provide protection to cells against electrophilic and oxidative stress. Antioxidant response elements (AREs) have been found in the promoter regions of the various detoxifying enzyme genes. An ARE is required for basal expression and induction of the various detoxifying enzyme genes in response to xenobiotics and antioxidants. In this study, we demonstrated that exposure of cells to xenobiotics [e.g. beta-naphthoflavone (beta-NF)] and antioxidants [e.g. tert-butyl hydroquinone (t-BHQ)] also induced the expression of the proto-oncogene c-jun. The induction of c-jun gene expression followed kinetics similar to the induction of NQO1 and NQO2 genes with respect to the level and time of exposure. Sequence analysis of the c-jun gene promoter revealed the presence of an ARE between nucleotides -538 and -514. The c-jun ARE was highly homologous to the AREs from genes encoding NQO1, NQO2, and GST Ya. Constructs containing the c-jun ARE and 1.7 and 4.5 kb of the c-jun promoter ligated to the chloramphenicol acetyltransferase (CAT) gene, upon transfection in human hepatoblastoma (Hep-G2) cells, expressed the CAT gene, which was inducible with beta-NF and t-BHQ. Band shift assays indicated binding of two specific nuclear protein complexes with the c-jun gene ARE. The faster running c-jun gene ARE-nuclear protein complex was specifically competed out by unlabeled NQO1 and GST Ya gene AREs. These results suggest that c-jun gene expression is coordinately induced and regulated with detoxifying enzyme genes in response to xenobiotics and antioxidants. The results also suggest involvement of an ARE-mediated mechanism of induction of c-jun gene expression. However, a comparison of fold induction of endogenous c-jun gene and transfected c-jun promoter/ARE-CAT constructs indicated involvement of another ARE upstream of the 4.5-kb promoter and/or additional mechanisms such as stabilization of c-Jun RNA in response to exposure to xenobiotics and antioxidants.
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PMID:Coordinated induction of the c-jun gene with genes encoding quinone oxidoreductases in response to xenobiotics and antioxidants. 1041 96


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