Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0019204 (hepatocellular carcinoma)
 71,386 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The cytochrome P4501 gene family consists of two members, CYP1A1 and CYP1A2, that are induced by halogenated hydrocarbons and polycyclic aromatic hydrocarbons. The human CYP1 promoters and 5'-flanking sequences were cloned into luciferase expression vectors to develop cell lines that stably express luciferase activity in response to CYP1 gene induction. Plasmids were initially tested in transient transfection assays. Transient transfections resulted in high-level expression of luciferase by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) from both the CYP1 expression vectors, pLUC1A1 and pLUC1A2. In dose-response experiments, 10 nM TCDD caused a maximal induction of pLUC1A2-directed luciferase activity that was 10-fold over control. Maximal pLUC1A1-directed luciferase activity was 65-fold over control in cells treated with 10 nM TCDD. Stable integration of CYP1-luciferase-neo plasmids, pL1A1N and pL1A2N, into the human hepatoma cell line, HepG2, was achieved by selection with G418. G418-resistant colonies were isolated for both pL1A1N and pL1A2N plasmids. The pL1A2N transfectants showed basal-level luciferase activity, but were nonresponsive to treatment with 10 nM TCDD. These results are in contrast to the observed induction of pLUC1A2-mediated luciferase expression in transient transfection experiments. Stable integration of the human CYP1A2 gene sequences appears to silence the transcriptional activation by TCDD. The pL1A1N transfectants showed inducible luciferase activity and one cell line, referred to as 101L, was used to establish dose-response relationships for TCDD and various polycyclic aromatic hydrocarbons. Maximal induction occurred after treatment with 100 nM TCDD, 10 microM 3-methylcholanthrene, 50 microM benz[a]anthracene, and 50 microM benzo[a]pyrene. These studies illustrate the use of the CYP1A1-luciferase cell line for the study of structure-activity relationships.
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PMID:Response of human CYP1-luciferase plasmids to 2,3,7,8-tetrachlorodibenzo-p-dioxin and polycyclic aromatic hydrocarbons. 844 4

We have isolated genomic clones containing the complete exon 1 and the promoter-regulatory region of the E1 alpha gene (BCKDHA) of human branched-chain alpha-keto acid dehydrogenase complex. The cloning was achieved by amplification of a genomic library in the SRB (P2) host strain that allowed the replication of nonstandard DNA structures. The results of this and previous (Dariush, N., Fisher, C. W., Cox, R. P., and Chuang, D. T. (1991) FEBS Lett. 284, 34-38) studies showed that the human E1 alpha gene contains 9 exons, and spans at least 55 kilobases (kb). Exon 1 is 135 bp in length, and contains multiple transcription initiation sites at bases +1, +18, and +22. The complete human E1 alpha cDNA is, therefore, 1,758 bp in length excluding the poly(A)+ tail, and has 27 nucleotides in the 5'-untranslated region. Sequencing of the 5'-flanking region disclosed the absence of a canonical TATA-box in the vicinity of base -30. Several sets of "CAAT" box-like sequences and Sp1 binding-sites are present. Also present are copies of potential AP-2 binding, fat-specific element 1, fat-specific element 2, glucocorticoid-responsive element, and cAMP-responsive element sequences, as well as multiple sets of direct and inverted repeats. The promoter-regulatory region was characterized using deletion constructs and the luciferase reporter assay. The human hepatoma cells (Hep-G2) and Chinese hamster ovary (CHO) cell lines were used as hosts. The results obtained with Hep-G2 cells indicate that the region for high level transcription is located between bases -320 and -115. Extension of the 5'-end of the insert to beyond base -320 markedly reduces promoter activity. The results strongly suggest the presence of inhibitory elements in the region upstream of base -320. Assays in CHO cells show that the region for high level transcription lies between bases -909 and -115. The variation in the region for high level transcription in Hep-G2 and CHO cells may represent cell-type specific differences in the E1 alpha gene promoter function.
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PMID:Characterization of the promoter-regulatory region and structural organization of E1 alpha gene (BCKDHA) of human branched-chain alpha-keto acid dehydrogenase complex. 846 40

Polycyclic aromatic hydrocarbons such as benzo(a)pyrene diol-epoxide (BPDE-I) cause hepatocellular carcinoma. To identify short-term carcinogen effects, we studied hepatocytes transfected with nonreplicating plasmids, adducted covalently with BPDE-I, varying in promoter structure and encoded reporter gene (beta-galactosidase or luciferase). BPDE inactivated gene expression as a first-order function of BPDE concentration in adduction reactions. No evidence of cytotoxicity, diminished coprecipitation and availability, enhanced nicking of supercoiled forms and reduced cellular uptake, or instability of adducted plasmids was observed. At low BPDE:plasmid ratios, inactivation occurred with 1 adduct/plasmid within a target 23-27% of plasmid bases. Using nuclear extracts and BPDE-adducted G-free cassette-encoding plasmids, the fraction of full-length RNA polymerase II-initiated transcripts also declined as a first-order function of BPDE concentration when approximately 3 adducts were distributed among 48% of plasmid bases. These observations suggest that carcinogens such as BPDE block mRNA transcription along DNA templates by forming limited numbers of persistent adducts at coding or noncoding sites.
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PMID:Inactivation of plasmid reporter gene expression by one benzo(a)pyrene diol-epoxide DNA adduct in adult rat hepatocytes. 848 14

To develop a molecular tool for tissue-specific targeting of gene expression in immature and differentiated epithelial cells of the small and large intestinal mucosa, we have isolated the 2-kb 5'-flanking region of the human villin gene. This region contains numerous short sequences that are conserved among other tissue-specific promoters of genes expressed in differentiated enterocytes. This DNA fragment promotes the transcription and expression of the luciferase reporter gene in villin-positive intestinal, renal, and hepatoma cell lines but not in a villin-negative keratinocyte cell line. The pattern of expression corresponds that of the endogenous gene, indicating that this sequence can direct intestine-specific transcription. In the differentiating HT29 intestinal cell line, expression of the reporter gene is already detectable in undifferentiated cells, and dramatically increases when terminal differentiation is induced. Thus, as previously reported for the endogenous gene the isolated 5'-flanking region of the villin gene responds positively to conditions known to stimulate terminal differentiation of these cultured epithelial intestinal cells. The reported results indicate that this genomic fragment contains sufficient regulatory elements to recapitulate the expression pattern of the villin promoter during intestinal differentiation.
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PMID:Regulatory sequences on the human villin gene trigger the expression of a reporter gene in a differentiating HT29 intestinal cell line. 849 91

Peroxisome proliferators cause rapid and coordinated transcriptional activation of genes encoding peroxisomal beta-oxidation system enzymes by activating peroxisome proliferator-activated receptor (PPAR) isoform(s). Since the thyroid hormone (T3; 3,3',5-triiodothyronine) receptor (TR), another member of the nuclear hormone receptor superfamily, regulates a subset of fatty acid metabolism genes shared with PPAR, we examined the possibility of interplay between peroxisome proliferator and T3 signaling pathways. T3 inhibited ciprofibrate-induced luciferase activity as well as the endogenous peroxisomal beta-oxidation enzymes in transgenic mice carrying a 3.2-kb 5'-flanking region of the rat peroxisomal enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase gene fused to the coding region of luciferase. Transfection assays in hepatoma H4-II-E-C3 and CV-1 cells indicated that this inhibition is mediated by TR in a ligand-dependent fashion. Gel shift assays revealed that modulation of PPAR action by TR occurs through titration of limiting amounts of retinoid X receptor (RXR) required for PPAR activation. Increasing amounts of RXR partially reversed the inhibition in a reciprocal manner; PPAR also inhibited TR activation. Results with heterodimerization-deficient TR and PPAR mutants further confirmed that interaction between PPAR and TR signaling systems is indirect. These results suggest that a convergence of the peroxisome proliferator and T3 signaling pathways occurs through their common interaction with the heterodimeric partner RXR.
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PMID:Thyroid hormone (T3) inhibits ciprofibrate-induced transcription of genes encoding beta-oxidation enzymes: cross talk between peroxisome proliferator and T3 signaling pathways. 852 10

We show here that the OxyR response element (ORE) in the bacterial oxyR promoter can also function as a redox-dependent enhancer in mammalian cells. Fusion of ORE to an SV40 basal promoter driving chloramphenicol acetyltransferase (CAT) expression confers H2O2 inducibility to expression of the cat gene in mouse Hepa-1 hepatoma cells. Nuclear extracts from these cells contain DNA-binding proteins that specifically interact with ORE DNA, cannot be completed by cognate oligonucleotides to AP-1 or NF kappa B, and are constitutively expressed, since treatment with H2O2 causes no detectable changes in binding activity or DNA-protein interaction. Recombinant cDNA clones that express ORE-binding proteins were isolated from a mouse hepatoma expression library and found to be representatives of two different members of the murine Y-box family of transcription factors. Canonical Y-box and ORE oligonucleotides compete with each other for binding to Y-box proteins in gel shift assays and antibodies to FRGY2, a Xenopus Y-box protein, supershift both Y-box and ORE DNA-protein complexes. In addition, antisense oligonucleotides to mouse YB-1 mRNA abolish induction of ORE-mediated cat expression by H2O2, and luciferase reporter constructs containing ORE, or the Y-box from the human MHC class II HLA-DQ gene, exhibit identical dose-dependent H2O2 inducibilities, which can be abolished by addition of 2-mercaptoethanol to the culture medium. These results suggest that the Y-box proteins may be an integral component of a eukaryotic redox signaling pathway.
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PMID:The Y-box motif mediates redox-dependent transcriptional activation in mouse cells. 853 Apr 81

Hepatitis B virus (HBV) DNA contains consensus elements for transactivating proteins whose binding activity in other systems is regulated by inflammatory cytokines. Because HBV replicates within an environment of provoked inflammation, we speculated that the HBV core/pregenomic promoter may be regulated by cytokines produced in response to infection. To evaluate this hypothesis, the HBV core/pregenomic (C/P) promoter and associated cis-acting elements were placed upstream of a luciferase-encoding plasmid. This reporter construct was transfected into cytokine-sensitive hepatoma cells permissive for HBV replication, which were exposed to stimulated mononuclear cell-conditioned medium or human recombinant cytokines. Conditioned medium reduced luciferase expression by 80%. Tumor necrosis factor alpha (TNF-alpha), interferon gamma (IFN-gamma), and interferon alfa (IFN-alpha) each reduced luciferase activity by 40%. Combinations of TNF-alpha and interferons mimicked the extent of conditioned medium inhibition. Non-specific effects from diminished cellular viability or growth were not responsible for decreased luciferase activity. Retention of HBV DNA 330 basepairs upstream of the C/P transcription start site was required to maintain the TNF-alpha effect. A 60% reduction in HBV replicative forms within intracellular core particles was demonstrated with TNF-alpha treatment of Hep G2 cells stably transfected with HBV DNA. The inhibitory action of these cytokines implicates a noncytolytic mechanism by which antigen-nonspecific immune responses in part regulate HBV replication in infected hepatocytes. This function may be beneficial in accelerating viral clearance, but in alternative circumstances could contribute to viral persistence by attenuating immunogen recognition.
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PMID:Cytokine inhibition of the hepatitis B virus core promoter. 855 37

The prevalence of the alternative alleles of an unusual length polymorphism in the promoter of the human antithrombin III (AT3) gene was determined in a sample of 155 unrelated individuals from the Northern Irish population. The 108bp L allele and the 32bp S allele occurred at frequencies of 0.21 and 0.79 respectively. Some homology was noted between the L-specific sequence and the region immediately downstream. Residual homology was also evident between the L and S sequences, suggesting that the S allele was derived from the L allele during evolution by partial deletion followed by sequence divergence. The functional significance of the polymorphism was investigated by transient transfection of AT3 promoter/luciferase reporter gene constructs into two human hepatoma cell lines in vitro. The promoter strength of the L allele was found to be 1.6-fold higher than the S allele in HepG2 cells whereas in Hep3B cells, the strength of the S allele was 1.7-fold higher than that of the L allele. In order to evaluate the phenotypic consequences of the AT3 promoter polymorphism in vivo, plasma samples from the 155 control individuals were assayed for antithrombin III (ATIII) activity. Mean activities of the different promoter polymorphism genotypes (SS, LL, SL) were not significantly different. These results suggest that the AT3 promoter polymorphism does not contribute to the variation in plasma ATIII activity that occurs in the general population.
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PMID:Functional analysis of an unusual length polymorphism in the human antithrombin III (AT3) gene promoter. 856 37

The gene encoding the rat branched-chain 2-oxo-acid dehydrogenase kinase (EC 2.7.1.115) has been isolated and partially characterized. The entire gene, including the promoter-regulatory region, spans 6 kb and contains 11 exons. The 5'-untranslated region comprising 264 bp is interrupted by intron 1 which is 581 bp in size. The complete in-frame sequence of intron 7 encodes the 49 amino acid insert previously reported to be present in the larger isoform of the rat kinase (Harris, Popov, Shimomura, Zhao, Jaskiewicz, Nanaumi and Suzuki (1992) Adv. Enzyme Regul. 32, 267-284). Sequencing of the 679 bp of the 5'-flanking region showed the absence of a canonical TATA box, similar to other branched-chain 2-oxo-acid dehydrogenase-complex genes. Several candidate cis-acting elements are present. These include CAAT boxes, Sp-1-binding sites, GCN-4 sites, CCAAT enhancer binding-protein sites (C/EBP) and glucocorticoid-responsive element (GRE) sites. Also present are a pair of direct repeats of unknown function. The luciferase-reporter assay showed that promoter activity is markedly higher in normal rat kidney (NRK-52E) cells than in rat hepatoma (FTO-2B) cells, and that the 5'-flanking region between bases -449 and +264 is both necessary and sufficient for basal transcription of the kinase gene.
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PMID:Structural organization of the rat branched-chain 2-oxo-acid dehydrogenase kinase gene and partial characterization of the promoter-regulatory region. 857 99

We have studied the role of the 2047 bp 5'-upstream region and 232 bp first exon sequence of the rat S100 beta gene in glial specific expression. S100 beta luciferase expression vectors carrying serial deletions of the S100 beta 5' upstream sequence were constructed and transiently transfected into the peripheral nervous system (PNS) glial-type cell line RT4-D6, the PNS neuronal-type cell line RT4-E5, and the central nervous system (CNS) glial-type cell line C6. The hepatoma cell line HTC was also transfected as a nonneural control. From this functional analysis, we found a glial-specific positive regulatory sequence(s) mapped between -583 and -106 relative to the transcriptional start site. This region confers a significantly higher level of luciferase expression in the glial-type cell lines RT4-D6 and C6 than in the neuronal cell line RT4-E5 and the hepatoma cell line HTC. Also, a non-cell type specific positive regulatory element was identified in the first exon sequence between +78 and +232. Though non-cell type-specific, it was found to have a predominant effect in glial cells. From these observations, we have concluded that the 2047 bp 5'-upstream region and 232 bp of the first exon sequence confers the high levels of S100 beta expression in glial cells through these two positive elements.
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PMID:Regulation of the rat S100 beta gene expression: the role of the 2 kb 5'-upstream sequence in glial specific expression. 860 Feb 92


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