Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.25.1 (deoxyribonuclease)
 1,471 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Ultraviolet (UV)-induced unscheduled DNA synthesis of xeroderma pigmentosum cells, belonging to complementation groups A, B, C, D, and E, was restored to the normal level by concomitant treatment of the cells with T4 endonuclease V and UV-inactivated HVJ (Sendai virus). The present results suggest that (1) T4 endonuclease molecules were inserted effectively into the cells by the interaction of HVJ with the cell membranes, (2) the enzyme was functional on human chromosomal DNA which had been damaged by UV irradiation in the viable cells, (3) all the studied groups of xeroderma pigmentosum ("variant" was not tested) were defective in the first step (incision) of excision repair.
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PMID:Restoration of ultraviolet-induced unscheduled DNA synthesis of xeroderma pigmentosum cells by the concomitant treatment with bacteriophage T4 endonuclease V and HVJ (Sendai virus). 17 93

The specific action of T4 endonuclease V on damaged DNA in xeroderma pigmentosum cells was examined using an in vivo assay system with hemagglutinating virus of Japan (Sendai virus) inactivated by UV light. A clear dose response was observed between the level of UV-induce unscheduled DNA synthesis of xeroderma pigmentosum cells and the amount of T4 endonuclease V activity added. The T4 enzyme was unstable in human cells, and its half-life was 3 hr. Fractions derived from an extract of Escherichia coli infected with T4V1, a mutant defective in the endonuclease V gene, showed no ability to restore the UV-induced unscheduled DNA synthesis of xeroderma pigmentosum cells. However, fractions derived from an extract of T4D-infected E. coli with endonuclease V activity were effective. The T4 enzyme was effective in xeroderma pigmentosum cells on DNA damaged by UV light but not in cells damaged by 4-nitroquinoline 1-oxide. The results of these experiments show that the T4 enzyme has a specific action on human cell DNA in vivo. Treatment with the T4 enzyme increased the survival of group A xeroderma pigmentosum cells after UV irradiation.
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PMID:Specific action of T4 endonuclease V on damaged DNA in xeroderma pigmentosum cells in vivo. 19 27

Osmotic disruption of confluent cultured human fibroblasts that have been irradiated or exposed to chemical carcinogens allows the specific measurement of repair DNA synthesis using dTTP as a precursor. Fibroblasts similarly prepared from various xeroderma pigmentosum cell lines show the deficiencies of UV-induced DNA synthesis predicted from in vivo studies, while giving normal responses to methyl methanesulfonate. A pyrimidine-dimer-specific enzyme, T4 endonuclease V, stimulated the rate of UV-induced repair synthesis with normal and xeroderma pigmentosum cell lines. This system should prove useful for identifying agents that induce DNA repair, and cells that respond abnormally to such induction. It should also be applicable to an in vitro complementation assay with repair-defective cells and proteins obtained from repair-proficient cells. Finally, by using actively growing fibroblasts and thymidine in the system, DNA replication can be measured and studied in vitro.
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PMID:A cell-free assay measuring repair DNA synthesis in human fibroblasts. 27 17

The repair mode of DNA replication has been demonstrated in isolated nuclei from UV-irradiated human cells. Nuclei are incubated in a mixture containing [(3)H]thymidine triphosphate and bromodeoxyuridine triphosphate in a 1:5 ratio. The (3)H at the density of parental DNA in alkaline CsCl density gradients is then a measure of repair. In nuclei prepared from WI38 cells 30 min after irradiation, repair replication is UV dependent and proceeds at approximately the in vivo rate for 5 min. Repair replication is reduced in irradiated nuclei or in nuclei prepared immediately after irradiation. It is Mg(2+)-dependent and stimulated by added ATP and deoxyribonucleoside triphosphates. No repair replication is observed in nuclei from xeroderma pigmentosum (complementation group A) cells. However, upon addition of coliphage T4 endonuclease V, which specifically nicks DNA containing pyrimidine dimers, repair replication is observed in nuclei from irradiated xeroderma pigmentosum cells and is stimulated in WI38 nuclei. The reaction then persists for an hour and is dependent upon added ATP and deoxyribonucleoside triphosphates. The repair label is in stretches of roughly 35 nucleotides, as it is in intact cells. Added pancreatic DNase does not promote UV-dependent repair synthesis. Our results support the view that xeroderma pigmentosum (group A) cells are defective in the incision step of the DNA excision repair pathway, and demonstrate the utility of this system for probing DNA repair mechanisms.
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PMID:Phage T4 endonuclease V stimulates DNA repair replication in isolated nuclei from ultraviolet-irradiated human cells, including xeroderma pigmentosum fibroblasts. 27 29

We have used a T4 endonuclease V assay method for UV-induced pryrimidine dimers in cellular DNA in vivo to obtain evidence for recombinational DNA exhanges after UV irradiation of normal human and Xeroderma pigmentosum (XP) cells. Our data indicate that the endonuclease-sensitive sites in excision-defective XP cells are removed very slowly from the irradiated parental strands and appear concomitantly in daughter strands newly synthesized during post-UV incubation. In the defective XP cells, the extent of appearance of sensitive sites in daughter strands synthesized during a period of 24 h after 10 J/m2 appears to be small, probably less than 15% of the initial number of sensitive sites detected in cellular parental strands. Demonstration of such exchanges between normal-density parental and 5-bromodeoxyuridine-labeled daughter strands by alkaline CsCl isopycnic centrifugation was unsuccessful. Further, the extent is much lower in normal human cells because of their efficient excision repair of the dimers before and after exchanges than in the defective XP cells.
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PMID:Low-level DNA exchanges in normal human and xeroderma pigmentosum cells after UV irradiation. 86 96

UV exposure has been linked to skin cancer in humans by epidemiology and the rare genetic disease xeroderma pigmentosum. However, UV produces multiple photoproducts in DNA, and their relative contribution is uncertain. An enzyme which specifically repairs cyclobutane pyrimidine dimers in DNA, T4 endonuclease V, was encapsulated in liposomes for topical delivery into mouse and human skin. In both species, liposomes applied after UV exposure localized in the epidermis and stimulated the removal of cyclobutane pyrimidine dimers. UV-irradiated mice treated with these liposomes had a dose-dependent decrease in the incidence of squamous cell carcinoma compared to controls. The results demonstrate that unrepaired cyclobutane pyrimidine dimers in DNA are a direct cause of cancer in mammalian skin.
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PMID:Pyrimidine dimer removal enhanced by DNA repair liposomes reduces the incidence of UV skin cancer in mice. 163 36

The phage T4 denV gene, coding for the pyrimidine-dimer specific T4 endonuclease V, was transfected into human repair-proficient fibroblasts, repair-deficient xeroderma pigmentosum fibroblasts, and into wild type CHO hamster cells. Transfectants maintained denV DNA and expressed denV mRNA. Purified T4 endonuclease V encapsulated in liposomes was also used to treat repair-proficient and -deficient human cells. The denV transfected clones and liposome-treated cells showed increased unscheduled DNA synthesis and enhanced removal of pyrimidine dimers compared to controls. Both denV gene transfection and endonuclease V liposome treatment enhanced post-UV survival in xeroderma pigmentosum cells but had no effect on survival in repair-proficient human or hamster cells. The results demonstrate that an exogenous DNA repair enzyme can correct the DNA repair defect in xeroderma pigmentosum cells and enhance DNA repair in normal cells.
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PMID:Enhancement of ultraviolet-DNA repair in denV gene transfectants and T4 endonuclease V-liposome recipients. 166 12

Repair kinetics of ultraviolet (UV) light-induced (6-4) photoproducts in xeroderma pigmentosum complementation group A, D, and variant cells were studied by the enzyme-linked immunosorbent assay (ELISA) using a specific monoclonal antibody raised against (6-4) photoproducts, together with unscheduled DNA synthesis (UDS) and loss of T4 endonuclease V-susceptible sites (ESS). Group AXP35KO cells completely failed to repair both ESS (cyclobutane pyrimidine dimers) and antibody-recognizing (6-4) photoproducts until tested 24 h after irradiation, and had 2% early-time UDS. Group DXP43KO cells showed about 10% removal of both (6-4) photoproducts and ESS in 24 h, despite showing a residually higher level of 40% early-time and cumulative UDS. Thus, the results substantiated the extreme UV hypersensitivity of XP group A and D cells. However, XP52KO variant cells exhibited the normal level of UDS and ESS loss, but a slightly reduced repair of antibody-recognizing (6-4) photoproducts at 6 and 12 h after irradiation, which may account for a small UV hypersensitivity of the XP variant cells.
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PMID:Repair of 254 nm ultraviolet-induced (6-4) photoproducts: monoclonal antibody recognition and differential defects in xeroderma pigmentosum complementation groups A, D, and variant. 279 53

Human hereditary diseases such as xeroderma pigmentosum, Fanconi's anemia, ataxia telangiectasia, and Bloom's syndrome are characterized by a proneness for developing cancer associated with abnormalities in the processing of DNA damage. The molecular defects responsible for predisposing human tissues to cancer are still not well understood, despite the fact that a considerable amount of work has already been done on this problem. In this paper, we show that in human tumor cell lines, in cells transformed by DNA tumor viruses, and in cells derived from certain cancer-prone disorders, the level of activity of a 42-kDa deoxyribonuclease is many times higher than in diploid untransformed control cells. This suggests that this activity is linked to, or may play a role in, malignant transformation.
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PMID:Enhanced deoxyribonuclease activity in human transformed cells and in Bloom's syndrome cells. 280 19

To extend our knowledge of the excision repair system in mammalian cells we have focussed on the isolation of genes and proteins involved in this process. For the purification and characterization of human repair proteins the microneedle injection assay technique is utilized. This system is based on the transient correction of the excision repair defect of xeroderma pigmentosum (XP) fibroblasts (scored as increase of ultraviolet (u.v.)-induced unscheduled DNA synthesis (UDS) upon microinjection of crude extracts from complementing XP or normal cells. Specific correction is observed in fibroblasts of all (9) excision-deficient XP complementation groups. The XP-A and G correcting factors were found to be proteins and several purification steps (including (NH4)2SO4 fractionation, chromatography of phosphocellulose, heparin and u.v.-irradiated DNA-cellulose) have been worked out for the XP-A correcting protein. The microinjection system was also used for the introduction of (partially) purified repair enzymes of lower organisms. Micrococcus luteus endonuclease and bacteriophage T4 endonuclease V were able to correct all XP complementation groups tested, in marked contrast to the more sophisticated Escherichia coli uvrABC complex injected with uvrD. Photoreversal of dimers could be registered after introduction of the yeast photoreactivating enzyme in repair-competent, XP-variant, XP-C and XP-I fibroblasts (monitored as decrease of (residual) UDS). Remarkably, no effect was noticed in XP-A, D, E and H, suggesting that something prevents dimers in these cells from being monomerized by the injected enzyme. Using DNA-mediated gene transfer we have cloned a human gene (designated ERCC-1) that compensates for the excision defect of the u.v. and mitomycin C-sensitive Chinese hamster ovary cell (CHO) mutant 43-3B (complementation group 2). Characterization of this gene and its cDNA revealed the following features: (1) ERCC-1 corrects the full spectrum of repair deficiencies in mutants of complementation group 2. No correction is observed in mutants of the other CHO complementation groups. (2) The ERCC-1 gene has a size of 15 X 10(3) base-pairs (bp) and consists of 10 exons, one of which appears to be differentially spliced. (3) It encodes two largely identical mRNAs, which differ in the presence or absence of a 72 bp coding exon, situated in the 3' half of the mRNA. Only the cDNA of the large transcript is able to confer repair proficiency to 43-3B cells. No effect of u.v. treatment is found at the level of ERCC-1 transcription in HeLa cells.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Characterization of genes and proteins involved in excision repair of human cells. 282 Oct 19


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