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:6.5.1.2 (DNA ligase)
2,749 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Several hepatotoxic agents damage Ca++ regulation and produce toxic cell death in a manner consistent with a cause-and-effect relationship; however, vital targets of Ca++ remain unidentified. Recent results show that DNA may be the chief Ca++ target during apoptosis, a form of cell death considered distinct from toxic cell death or necrosis. The present studies explored whether nuclear Ca++ regulation is lost before dimethylnitrosamine-induced necrosis, whether DNA is attacked by Ca(++)-dependent endonucleases and whether inhibitors of Ca(++)-endonuclease activity and the DNA repair enzyme poly(ADP-ribose)polymerase affect necrosis. Adult male ICR mice received 100 mg/kg of dimethylnitrosamine i.p. By 2 to 4 hr, total nuclear Ca++ reached 150 to 180% of control and DNA fragmentation was 140 to 170% of control. Electrophoresis of DNA revealed a sharp decline in genomic DNA with the appearance of DNA fragments in a ladder-like pattern. Ca++ elevation and DNA fragmentation preceded toxic cell death by 4 hr or more and reached peak values at 18 to 24 hr, coincident with maximal alanine aminotransferase leakage. Aurintricarboxylic acid, a Ca(++)-endonuclease inhibitor, reduced toxicity 67%. 3-Aminobenzamide, nicotinamide adenine dinucleotide and theophylline, inhibitors of poly(ADP-ribose)polymerase-mediated DNA repair, potentiated liver damage 2-fold. These results support the hypothesis that DNA fragmentation plays a contributing role in toxic cell death induced by dimethylnitrosamine. Furthermore, the findings suggest that new opportunities may exist to moderate the toxicity of alkylating hepatotoxins by altering DNA regulation.
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PMID:Ca(++)-activated DNA fragmentation and dimethylnitrosamine-induced hepatic necrosis: effects of Ca(++)-endonuclease and poly(ADP-ribose) polymerase inhibitors in mice. 132 12

Both H2O2 (IC50 = 70 microM) and HOCl (IC50 = 8.5 microM) inhibited mitogen-induced MNL proliferation in a dose-dependent manner. This was found to be due to a depletion of intracellular ATP by at least two distinct mechanisms. HOCl and high concentrations (greater than 100 microM) of H2O2 inhibit ATP generation via sulfhydryl group oxidation on the active site of the glyceraldehyde-3-phosphate dehydrogenase (G3PDH) enzyme of the glycolytic pathway. On the other hand, low H2O2 concentrations cause ATP depletion by an activation of the DNA repair enzyme, poly(ADP-ribose)polymerase (pADPRP), leading to consumption of NAD+, an essential cofactor for G3PDH. The anti-oxidants ascorbate and cysteine protected MNL against the anti-proliferative effects of HOCl. Similar results were achieved with the HOCl-mediated inhibition of ATP production and G3PDH activity. However, ascorbate was unable to protect against H2O2-mediated inhibition of MNL functions, while cysteine protected against the inhibitory effects on ATP production and G3PDH activity, induced by this oxidant.
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PMID:Biochemical mechanisms of hydrogen peroxide- and hypochlorous acid-mediated inhibition of human mononuclear leukocyte functions in vitro: protection and reversal by anti-oxidants. 132 47

The influence of poly(ADP-ribose) polymerase (PARP) on the replication of DNA containing the SV40 origin of replication has been examined. Extensive replication of SV40 DNA can be carried out in the presence of T antigen, topoisomerase I, the multimeric human single strand DNA-binding protein (HSSB), and DNA polymerase alpha-DNA primase (pol alpha-primase) complex (the monopolymerase system). In the monopolymerase system, both small products (Okazaki fragments), arising from lagging strand synthesis, and long products, arising from leading strand synthesis, are formed. The synthesis of long products requires the presence of relatively high levels of pol alpha-primase complex. In the presence of PARP, the synthesis of long products was blocked and only small Okazaki fragments accumulated, arising from the replication of the lagging strand template. The inhibition of leading strand synthesis by PARP can be effectively reversed by supplementing the monopolymerase system with the multimeric activator 1 protein (A1), the proliferating cell nuclear antigen (PCNA) and PCNA-dependent DNA polymerase delta (the dipolymerase system). The inhibition of leading strand synthesis in the monopolymerase system was caused by the binding of PARP to the ends of DNA chains, which blocked their further extension by pol alpha. The selective accumulation of Okazaki fragments was shown to be due to the coupled synthesis of primers by DNA primase and their immediate extension by pol alpha complexed to primase. PARP had little effect on this coupled reaction, but did inhibit the subsequent elongation of products, presumably after pol alpha dissociated from the 3'-end of the DNA fragments. PARP inhibited several other enzymatic reactions which required free ends of DNA chains. PARP inhibited exonuclease III, DNA ligase, the 5' to 3' exonuclease, and the elongation of primed DNA templates by pol alpha. In contrast, PARP only partly competed with the elongation of primed DNA templates by the pol delta elongation system which required SSB, A1, and PCNA. These results suggest that the binding of PARP at the ends of nascent DNA chains can be displaced by the binding of A1 and PCNA to primer ends. HSSB can be poly(ADP-ribosylated) in vivo as well as in vitro. However, the selective effect of PARP in blocking leading strand synthesis in the monopolymerase system was shown to depend primarily on its DNA binding property rather than on its ability to synthesize poly(ADP-ribose).
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PMID:Influence of poly(ADP-ribose) polymerase on the enzymatic synthesis of SV40 DNA. 167 70

To investigate whether target cell DNA injury participates in cytolysis by human neutrophil defensins (HNP), we analyzed HNP-treated cells for single strand breaks by the alkaline unwinding assay and the activation of ADPribose polymerase, a DNA repair enzyme. Strand breaks and ADP-ribosylation were first detected in K562 and Raji targets 6-8 hr after incubation with HNP and increased to maximal levels by 18 hr. DNA was not degraded into nucleosome-sized fragments. To assess the impact of DNA injury on cytolysis, we increased strand breakage by coincubating targets with HNP and two inhibitors of ADPribose polymerase, 3-aminobenzamide, or nicotinamide. Concurrently with inhibiting polymerase activity and increasing DNA injury, these agents significantly enhanced HNP-mediated cytolysis. Enhancement occurred only at time points (over 6 hr) and in targets (only nucleated targets) where HNP-induced DNA injury could be occurring. These data indicate that neutrophil defensins can induce DNA injury in targets and suggest such injury may be involved in target cell death.
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PMID:Human neutrophil peptide defensins induce single strand DNA breaks in target cells. 191 32

Variants of mouse leukaemia L1210 cells have been isolated in which cytotoxicity to dimethyl sulphate is not fully potentiated by ADP-ribosyl transferase inhibitor 3-aminobenzamide, as occurs in normal L1210 cells. These variants were selected after mutagenesis by growing the cells in dimethyl sulphate and 3-aminobenzamide. The characterisation of one of these variants is described. Variant 3 cells repair low doses of DNA damage in the presence of ADP-ribosyl transferase inhibitors. The Vmax of the ADP-ribosyl transferase enzyme in these cells is only increased 35% compared to normal wild-type L1210 cells. The basal DNA ligase I activity is increased 66% above wild-type whereas DNA ligase II activity appears to be unchanged. The most striking observation, however, is that the DNA ligase II activity is not increased after dimethyl sulphate treatment as occurs in wild-type L1210 cells. It seems that by increasing DNA ligase I levels these cells can survive DNA damage in the presence of 3-aminobenzamide. This variant (mutant) provides genetic evidence for our previously published hypothesis that (ADP-ribose)n biosynthesis is required for efficient DNA repair after DNA damage by monofunctional alkylating agents, because ADP-ribosyl transferase activity regulates DNA ligase activity. This variant is the first mammalian cell reported in which DNA ligase activity is altered, as far as we are aware. In yeast, a DNA ligase mutant has a cell division cycle (cdc) phenotype. Presumably, DNA ligase is essential for DNA synthesis, repair and recombination. The present variant provides further evidence that in mammalian cells, DNA ligase II activity is related to ADP-ribosyl transferase activity.
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PMID:A mammalian cell variant in which 3-aminobenzamide does not potentiate the cytotoxicity of dimethyl sulphate. 301 97

Previous studies have shown that structural changes in DNA, including the ligation of pre-existing DNA breaks and the opening and closure of new breaks, occur shortly after exposure of granulomonocytic precursors (CFU-GM) to granulocyte-macrophage colony stimulating activity (GM-CSA). Monocytic differentiation of CFU-GM is selectively inhibited by compounds known to inhibit the nuclear enzyme ADP-ribosyl transferase (ADPRT). Since this enzyme, which transfers ADP-ribose units to chromatin proteins, is known to activate DNA ligase, we attempted to determine whether ligation of one or both types of DNA break is required for monocytic differentiation. Breaks in DNA were examined using the nucleoid sedimentation technique in which DNA breaks cause loss of DNA supercoiling in nucleoids and concomitant changes in their sedimentation through neutral sucrose gradients. We here report that two distinct patterns of DNA strand breakage and ligation are associated with differentiation to the granulocyte and monocyte lineages. Monocytic inducers (phorbolester and vitamin D3) predominantly produce closure of pre-existing strand breaks, whereas granulocytic inducers (granulocyte colony stimulating activity, G-CSA; retinoic acid) cause opening and closure of new breaks. Only ligation of the pre-existing breaks is highly sensitive to inhibition by 3-methoxybenzamide (a potent ADPRT inhibitor), and only monocytic differentiation is impaired by addition of this compound. These findings suggest that DNA structural changes may be directly involved in granulocyte-macrophage switching.
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PMID:Contrasting patterns of DNA strand breakage and ADP-ribosylation-dependent DNA ligation during granulocyte and monocyte differentiation. 303 Apr 64

Oxidants are generated in vivo by multiple mechanisms, including stimulation of leukocytes, hyperoxia, metabolism of arachidonic acid, and the activation of various oxidases. When the biochemical defences to the oxidants are inadequate, injury of tissues results. This injury was observed in rabbits and rhesus monkeys when pulmonary inflammation was induced with phorbol esters or formylated peptide given intrabronchially. We have recently investigated metabolic changes in various cells exposed to oxidants that are generated from stimulated leukocytes, including H2O2, O2, and HOCl. The target cells used were P388D1 murine macrophage-like tumour cells, human peripheral lymphocytes, GM 1380 human fibroblasts and rabbit alveolar macrophages. The oxidants used were H2O2 and PMA stimulated PMNs or neutroplasts. Lysis could only be prevented when catalase was added within the first 30-40 min of H2O2 exposure indicating that early metabolic changes determined the fate of the cell. Within seconds after the addition of H2O2 to P388D1 cells activation of the hexose monophosphate shunt (HMPS) was observed indicative of increased glutathione cycle activity. At the same time DNA strand breaks (determined by an alkaline unwinding technique) were detected. They resulted in the activation of the DNA repair enzyme poly-ADP-ribose polymerase (pADP-RP) within minutes after the addition of H2O2. At the same time ATP and NAD (the substrate of pADP-RP) concentrations dropped and nicotinamide accumulated extracellularly. 10-15 min after oxidant exposure free intracellular Ca++ concentrations determined by Quin 2 fluorescence started to increase due to release from intracellular stores.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Oxidant and protease injury of the lung. 369 17

Incubation of DNA polymerase alpha, DNA polymerase beta, terminal deoxynucleotidyl transferase, or DNA ligase II in a reconstituted poly(ADP-ribosyl)ating enzyme system markedly suppressed the activity of these enzymes. Components required for poly(ADP-ribose) synthesis including poly(ADP-ribose) polymerase, NAD+, DNA, and Mg2+ were all essential for the observed suppression. Purified poly(ADP-ribose) itself, however, was slightly inhibitory to all of these enzymes. Furthermore, the suppressed activities of DNA polymerase alpha, DNA polymerase beta, and terminal deoxynucleotidyl transferase were largely restored (3 to 4-fold stimulation was observed) by a mild alkaline treatment, a procedure known to hydrolyze alkaline-labile ester linkage between poly(ADP-ribose) and an acceptor protein. All of these results strongly suggest that the four nuclear enzymes were inhibited as a result of poly(ADP-ribosyl)ation of either the enzyme molecule itself or some regulatory proteins of these enzymes.
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PMID:Inhibition of DNA polymerase alpha, DNA polymerase beta, terminal deoxynucleotidyl transferase, and DNA ligase II by poly(ADP-ribosyl)ation reaction in vitro. 392 Oct 27

U.v. damage to the DNA of HeLa cells induces the polymerisation of ADP-ribose, but only if repair synthesis is inhibited so that incomplete repair sites (i.e., DNA breaks) accumulate to abnormally high levels. 3-Aminobenzamide greatly reduces the ADP-ribose polymerisation response. However, 3-aminobenzamide does not reduce the rate of rejoining of the accumulated breaks when the inhibition of repair synthesis is reversed. Therefore, rejoining of these DNA breaks (in contrast to the rejoining of other kinds of break) appears not to depend on activation of polynucleotide ligase by ADP-ribosylation.
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PMID:Poly (ADP-ribose) is not involved in the rejoining of DNA breaks accumulated to high levels in u.v.-irradiated HeLa cells. 401 70

ADP-ribosyl transferase (ADP-RT) is a chromatin-bound nuclear enzyme catalysing the transfer of ADP-ribose from NAD+ to chromatin proteins. The enzyme is activated by DNA strand breaks and has been suggested to have roles in both DNA repair (via its effect on DNA ligase II) and in differentiation. We recently demonstrated that specific inhibitors of ADP-RT preferentially inhibit differentiation of human granulocyte-macrophage progenitor cells to the macrophage lineage and that the specific proliferation/differentiation stimulus granulocyte-macrophage colony stimulating activity (GM-CSA) activates ADP-RT in human marrow cells within 3 h of exposure. The purpose of this study was to investigate the role of ADP-RT in monocyte-macrophage differentiation. By altering the time of addition of ADP-RT inhibitor it was demonstrated that maximal inhibition of macrophage differentiation only occurs when the inhibitor is added within the first 24 h of culture. This suggests that it is an early event during the induced differentiation of granulocyte-macrophage progenitor cells which requires ADP-RT. Fluorometric assay of the level of DNA strand breaks showed that GM-CSA induces DNA strand breaks which are rapidly ligated only if ADP-RT is available. These data and those of our earlier studies suggest that DNA rearrangement may be involved in differentiation of granulocyte-macrophage progenitors to the monocyte-macrophage pathway. Such a DNA rearrangement could provide a molecular basis for commitment of multipotent progenitors to a single lineage.
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PMID:DNA strand breakage and ADP-ribosyl transferase mediated DNA ligation during stimulation of human bone marrow cells by granulocyte-macrophage colony stimulating activity. 608 35


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