Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.7.49 (reverse transcriptase)
 31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Specific cytochrome P450 enzymes show tissue-specific induction, and different regulatory units for expression of these enzymes have been identified. The regulation of the phenobarbital (PB)-inducible P450 genes has been relatively well characterized in terms of PB induction, but less so with regard to tissue-specific expression. CYP2B2 is not expressed in the rat lung, whereas cytochrome P450 2B1 (CYP2B1) is a dominating enzyme in the same tissue. The constitutive expression of CYP2B1 and CYP2B2 in liver is low, but inducible by PB, whereas the pulmonary expression of CYP2B1 is not induced by PB. This indicates utilization of different regulating mechanisms in the two organs. A gene construct consisting of the structural gene for LacZ coupled to a 1.3-kb 5' fragment of the rat CYP2B1 gene was used to generate transgenic mice in order to further elucidate the mechanism behind tissue-specific expression and PB induction of the CYP2B1 gene. Using reverse transcriptase-polymerase chain reaction on total RNA extracted from lung and liver tissue, a lung-specific transcription of the transgene was observed. Transcription of the construct was also observed in livers from PB-treated transgenic animals. By histochemical staining of lung sections with 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal), we demonstrated expression at the protein level in bronchiolar cells. In conclusion, our results revealed that the region extending to -1. 3 kb in the 5' flanking region of the CYP2B1 gene included sequences that could partly account for the lung-specific transcription of CYP2B1 and the hepatic induction of CYP2B1 transcription by PB.
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PMID:Cis-acting sequences from the rat cytochrome P450 2B1 gene confer pulmonary and phenobarbital-inducible expression in transgenic mice. 1042 99

A DNA polymerase beta (pol. beta) inhibitor has been isolated independently from two organisms; a red perilla, Perilla frutescens, and a mugwort, Artemisia vulgaris. These molecules were determined by spectroscopic analyses to be the cyanogenic glucoside, D-mandelonitrile-beta-D-glucoside, prunasin. The compound inhibited the activity of rat pol. beta at 150 microM, but did not influence the activities of calf DNA polymerase alpha and plant DNA polymerases, human immunodefficiency virus type 1 reverse transcriptase, calf terminal deoxynucleotidyl transferase, or any prokaryotic DNA polymerases, or DNA and RNA metabolic enzymes examined. The compound dose-dependently inhibited pol. beta activity, the IC(50) value being 98 microM with poly dA/oligo dT(12-18) and dTTP as the DNA template and substrate, respectively. Inhibition of pol. beta by the compound was competitive with the substrate, dTTP. The inhibition was enhanced in the presence of fatty acid, and the IC(50) value decreased to approximately 40 microM. In the presence of C(10)-decanoic acid, the K(i) value for substrate dTTP decreased by 28-fold, suggesting that the fatty acid allowed easier access of the compound to the substrate-binding site.
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PMID:The cyanogenic glucoside, prunasin (D-mandelonitrile-beta-D-glucoside), is a novel inhibitor of DNA polymerase beta. 1042 40

Adenovirus vector system is expected to be useful for direct gene therapy for joint disease. This study first sought to confirm that foreign genes can be transferred to articular chondrocytes in primary culture. Next, recombinant adenovirus vectors harbouring beta-galactosidase gene (LacZ) was injected directly into the temporomandibular joints of Hartley guinea-pigs to clarify the in vivo transfer availability of the adenovirus vectors. Specifically, recombinant adenovirus harbouring LacZ gene (AxlCALacZ) was injected into the upper joint cavities of both mandibular joints of four male 6-week-old Hartley guinea-pigs. Either the same amount of recombinant adenovirus without LacZ gene (Axlw) suspension (placebo) or the same amount of phosphate-buffered saline solution (control) were injected into the upper joint cavities of both joints of another four male guinea-pigs. At 1, 2, 3 and 4 weeks after injection, the joints were dissected and the expression of delivered LacZ was examined by 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal) staining and reverse transcriptase-polymerase chain reaction (RT-PCR). To investigate the expression of transferred gene in other organs, total RNA was extracted from liver, kidney, heart and brain and the expression of LacZ mRNA and 18 S ribosomal RNA were analysed by RT-PCR. Clear expression of LacZ was observed in the articular surfaces of the temporal tubercle, articular disc and synovium of the temporomandibular joints even 4 weeks after injection in the AxlCALacZ-injected group, while no expression was detected in placebo and control groups. Histological examination confirmed that LacZ activity was clearly detected in a few cell layers of the articular surface tissues, which is much more efficient than in a previously study of the knee joint. In the other organs, expression of the delivered transgene was not observed. Based on these findings, direct gene delivery into the articular surface of the temporomandibular joint using the adenovirus vector is feasible as an effective in vivo method.
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PMID:Direct adenovirus-mediated gene delivery to the temporomandibular joint in guinea-pigs. 1047 Nov 54

Adherence of yeast cells of Candida albicans to human oesophageal cells is greater when cells are grown in 500 mM D-galactose in comparison to D-glucose at the same concentration. Moreover, a 190 kDa mannoprotein (MP190) from a yeast cell wall preparation is highly expressed when cells are grown in the presence of galactose but less so in glucose. We now report on the identification of the MP190 and the isolation of its encoding gene. MP190 was purified, and three internal peptides were isolated and sequenced. Each of the three peptides showed significant homology (65-85%) with a glucoamylase (GAM1) from the yeast, Schwanniomyces occidentalis. In order to isolate the C. albicans homologue of GAM1 (GCA1), we probed a genomic library with a 0.9-kb internal fragment of the S. occidentalis GAM1 and isolated a 2.3-kb clone that corresponded to the 5' region of the gene. Polymerase chain reaction (PCR) amplification was used to isolate the remainder of the open reading frame. GCA1 encodes a 946 amino acid protein containing three putative hydrophobic, membrane-spanning domains and 15 potential N-glycosylation sites. Both Gca1p and GAM1 are novel to the family of glycosyl hydrolases. Northern analysis indicated that GCA1 is transcribed to a greater extent in galactose than in sucrose or glucose. Also, using reverse transcriptase (RT)-PCR, we observed expression of GCA1 in a rat model of oral candidiasis, indicating that Gca1p is expressed during disease development.
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PMID:Identification and cloning of GCA1, a gene that encodes a cell surface glucoamylase from Candida albicans. 1052 Jan 61

Exposure of pancreatic islet B-cells to D-glucose and many other insulinotropic agents results in an increase of cytoplasmic calcium concentration, which triggers the exocytosis of secretroy granules. Previous studies have demonstrated that calcium itself, at concentrations ranging from 2 to 18 mM, is able to induce a dose-related stimulation of insulin secretion, even in the absence of any other secretagogue. It was recently demonstrated that parathyroid cells and several other cell types, whether involved or not in calcium homeostasis, sense extracellular calcium through a G-protein coupled calcium-sensing receptor (CaSR). In the present study, the presence of the receptor in islet pancreatic B-cells was scrutinized. Using reverse transcriptase-polymerase chain reaction and Northern blot analysis, we demonstrate the expression of the CaSR in purified rat pancreatic islet B-cells. The nucleotide sequences of the rt-PCR products demonstrated more than 99% homology with the rat kidney CaSR complementary DNA. A specific 5.3 kb transcript of the CaSR was expressed in normal pancreatic B-cells as well as in tumoral insulin-secreting cells. In pancreatic islets, the physiological role of the CaSR in the regulation of insulin release could involve the sensing of endogenous ligands other than calcium.
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PMID:Expression of the calcium-sensing receptor in pancreatic islet B-cells. 1054 80

To investigate the coagulation system in crustacean decapoda, a homodimeric glycoprotein of 380 kDa was purified from the hemolymph of tiger shrimp (Penaeus monodon) by sequential DEAE anion exchange chromatography. The purified protein was coagulated by the shrimp hemocyte transglutaminase in the presence of Ca2+. The clottable protein contains 44% alpha helices and 26% beta sheets as determined by circular dichroism spectra. Its conformation is stable in buffer of pH 4-9. To solve its primary structure, partial sequences of the purified polypeptides from cyanogen bromide cleavage and endopeptidase digestion were also determined. A shrimp cDNA expression library was constructed. By combination with antibody screening, reverse transcriptase PCR using degenerate primers from determined amino acid sequences and cDNA library screening with digoxigenin-labeled DNA probes, the entire cDNA of 6124 bp was obtained. This cDNA encodes a protein of 1670 amino acids, including a 14-amino acid signal peptide. With four potential N-glycosylation sites, the clottable protein was found to contain 3.8% high-mannose glycan; and Man8GlcNAc and Man9GlcNAc were released upon endo-beta-N-acetylglucosaminidase hydrolysis. Upon conducting a protein sequence database survey, the shrimp clottable protein shows 36% identities to the crayfish clotting protein and lower similarities to members of insect vitellogenins, apolipoprotein B and mammalian von Willebrand factor. Notably, a region rich in Gln residues, a polyGln motif and five Ser-Lys-Thr-Ser repeats are present in the shrimp protein, suggesting this protein might be a transglutaminase substrate. Northern blot analysis revealed that the clottable protein is expressed in most of the shrimp tissues but not in the mature hemocytes.
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PMID:Molecular cloning and characterization of a hemolymph clottable protein from tiger shrimp (Penaeus monodon). 1056 6

The cysteine-rich region of the 170-kDa subunit galactose-adherence lectin (Gal-lectin) of Entamoeba histolytica is a subunit vaccine candidate and a protective antigen in the gerbil model of amebiasis. Macrophage-mediated immunity is important for protection against E. histolytica and is activated by Th1 cytokines. As Th1 differentiation is promoted by IL-12, we investigated what portion of the Gal-lectin could stimulate IL-12 in human THP-1 macrophages. Native Gal-lactin stimulated IL-12 p40 / p35 mRNA expression in a dose- and time-dependent manner as measured by reverse transcriptase-PCR. Human immune serum and Gal-lectin mAb inhibition studies identified amino acids (aa) 596 - 998 as immunogenic and containing the IL-12 inducing domain. IFN-gamma priming augmented Gal-lectin-induced IL-12 mRNA expression independent of TNF-alpha and IL-1beta, and was required for IL-12 p70 protein production from macrophages and human peripheral blood mononuclear cells. Gal-lectin plus IFN-gamma stimulated IL-12 p40 and p35 gene transcription with stable mRNA transcripts and a differential requirement for protein synthesis. These results suggest that aa 596 - 998 of the Gal-lectin can confer Th1-mediated protection against amebiasis through IL-12 induction.
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PMID:A subunit vaccine candidate region of the Entamoeba histolytica galactose-adherence lectin promotes interleukin-12 gene transcription and protein production in human macrophages. 1067 Nov 97

Gene transfer was performed using asialo-oroso-mucoid-polylysine (ASOR-PL) conjugates to allow targeted expression of the gene in cells of hepatic origin. In a gel-electrophoretic analysis, the ASOR-PL conjugate produced a complete DNA retardation effect at the optimal ratio of 222:1 (ASOR-PL conjugate/pCMV beta-gal plasmid). The gene-transfer efficiency of the ASOR-PL conjugate was evaluated in HepG2 cells that express asialoglycoprotein receptor and NIH 3T3 cells that do not. The expression was assayed by 5-bromo-4-chloroindol-3-yl beta-D-galactopyranoside ('X-Gal') staining and Chlorophenol Red beta-D-galactopyranoside. When an expression vector for the tumour-suppressor gene p53, pCMVp53, complexed to ASOR-PL conjugate, was transfected into HepG2 cells, the exogenously provided p53 gene was detected in the HepG2 cells by PCR. To improve the efficiency of DNA delivery and expression of the therapeutic proteins poloxamer 407, a fusogenic peptide, influenza-virus haemagglutinin HA2 and chloroquine were individually incorporated into the system. The expression level of beta-galactosidase in HepG2 cells was increased by about four times by the presence of poloxamer 407, whereas the fusogenic peptide HA2 and chloroquine had no effects. When HepG2 cells were transfected with pCMVp53 in the presence of poloxamer 407, the mRNA of transfected p53 could be detected by reverse transcriptase PCR. The current findings open the possibility that a receptor-mediated gene-delivery system for hepatic gene therapy using ASOR-PL conjugate in combination with poloxamer 407 may be developed in the future.
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PMID:Improvement of receptor-mediated gene delivery to HepG2 cells using an amphiphilic gelling agent. 1091 34

A large variety of natural products have been described as anti-HIV agents, and for a portion thereof the target of interaction has been identified. Cyanovirin-N, a 11-kDa protein from Cyanobacterium (blue-green alga) irreversibly inactivates HIV and also aborts cell-to-cell fusion and transmission of HIV, due to its high-affinity interaction with gp120. Various sulfated polysaccharides extracted from seaweeds (i.e., Nothogenia fastigiata, Aghardhiella tenera) inhibit the virus adsorption process. Ingenol derivatives may inhibit virus adsorption at least in part through down-regulation of CD4 molecules on the host cells. Inhibition of virus adsorption by flavanoids such as (-)epicatechin and its 3-O-gallate has been attributed to an irreversible interaction with gp120 (although these compounds are also known as reverse transcriptase inhibitors). For the triterpene glycyrrhizin (extracted from the licorice root Glycyrrhiza radix) the mode of anti-HIV action may at least in part be attributed to interference with virus-cell binding. The mannose-specific plant lectins from Galanthus, Hippeastrum, Narcissus, Epipac tis helleborine, and Listera ovata, and the N-acetylgl ucosamine-specific lectin from Urtica dioica would primarily be targeted at the virus-cell fusion process. Various other natural products seem to qualify as HIV-cell fusion inhibitors: the siamycins [siamycin I (BMY-29304), siamycin II (RP 71955, BMY 29303), and NP-06 (FR901724)] which are tricyclic 21-amino-acid peptides isolated from Streptomyces spp that differ from one another only at position 4 or 17 (valine or isoleucine in each case); the betulinic acid derivative RPR 103611, and the peptides tachyplesin and polyphemusin which are highly abundant in hemocyte debris of the horseshoe crabs Tachypleus tridentatus and Limulus polyphemus, i.e., the 18-amino-acid peptide T22 from which T134 has been derived. Both T22 and T134 have been shown to block T-tropic X4 HIV-1 strains through a specific antagonism with the HIV corecept or CXCR4. A number of natural products have been reported to interact with the reverse transcriptase, i.e., baicalin, avarol, avarone, psychotrine, phloroglucinol derivatives, and, in particular, calanolides (from the tropical rainforest tree, Calophyllum lanigerum) and inophyllums (from the Malaysian tree, Calophyllum inophyllum). The natural marine substance illimaquinone would be targeted at the RNase H function of the reverse transcriptase. Curcumin (diferuloylmethane, from turmeric, the roots/rhizomes of Curcuma spp), dicaffeoylquinic and dicaffeoylt artaric acids, L-chicoric acid, and a number of fungal metabolites (equisetin, phomasetin, oteromycin, and integric acid) have all been proposed as HIV-1 integrase inhibitors. Yet, we have recently shown that L-c hicoric acid owes its anti-HIV activity to a specific interaction with the viral envelope gp120 rather than integrase. A number of compounds would be able to inhibit HIV-1 gene expression at the transcription level: the flavonoid chrysin (through inhibition of casein kinase II, the antibacter ial peptides melittin (from bee venom) and cecropin, and EM2487, a novel substance produced by Streptomyces. (ABSTRACT TRUNCATED)
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PMID:Current lead natural products for the chemotherapy of human immunodeficiency virus (HIV) infection. 1093 47

Ty3 is a gypsy-type, retrovirus-like element found in the budding yeast Saccharomyces cerevisiae. In cells overexpressing Ty3 under the GAL1 upstream activation sequence, Ty3 RNA, proteins, and DNA are made. Elucidation of the molecular masses and amino-terminal sequences of protease and reverse transcriptase indicated the existence of an additional intervening domain, designated J, in the Ty3 Gag3-Pol3p polyprotein. A region analogous to J can be found in many retrotransposable elements closely related to Ty3; however, J does not correspond to any of the highly conserved retroviral protein domains. Ty3 mutants deleted for the J-coding region showed moderately reduced transposition frequency but greatly reduced levels of Ty3 DNA. These results show that under galactose regulation, the Ty3 J domain is not absolutely essential.
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PMID:Ten-kilodalton domain in Ty3 Gag3-Pol3p between PR and RT is dispensable for Ty3 transposition. 1115 29


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