Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UMLS:C0043346 (xeroderma pigmentosum)
 2,924 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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

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

Introduction of the denV gene of phage T4, encoding the pyrimidine dimer-specific endonuclease V, into xeroderma pigmentosum cells XP12RO(M1) was reported to result in partial restoration of colony-forming ability and excision repair synthesis. We have further characterized 3 denV-transformed XP clones in terms of rates of excision of pyrimidine dimers and size of the resulting resynthesized regions following exposure to 100 J/m2 from an FS-40 sunlamp. In the denV-transformed XP cells we observed 50% dimer removal within 3-6 h after UV exposure as compared to no measurable removal in the XP12RO(M1) line and 50% dimer excision after 18 h in the GM637A human, control cells. Dimer removal was assayed with Micrococcus luteus UV-endonuclease in conjunction with sedimentation of treated DNA in alkaline sucrose gradients. The size of the resulting repaired regions was determined by the bromouracil photolysis technique. Based on the photolytic sensitivity of DNA repaired in the presence of bromodeoxyuridine, we calculated that the excision of a dimer in the GM637A cells appears to be accompanied by the resynthesis of a region approximately 95 nucleotides in length. Conversely, the resynthesized regions in the denV-transformed clones were considerably smaller and were estimated to be between 13 and 18 nucleotides in length. These results may indicate that either the endonuclease that initiated dimer repair dictated the size of the resynthesized region or that the long-patch repair observed in the normal cells resulted from the repair of non-dimer DNA lesions.
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PMID:Excision repair characteristics of denV-transformed xeroderma pigmentosum cells. 291 65

A structural gene for T4 endonuclease V was constructed by ligating synthetic oligonucleotides. The endonuclease V was overproduced in E. coli under control of the E. coli tryptophan promoter and purified to apparent homogeneity. The product had comparable DNA glycosylase and apurinic/apyrimidinic (AP) endonuclease activities to the natural enzyme in vitro. When this endonuclease V was microinjected into the cytoplasm of xeroderma pigmentosum (XP) cells of complementation group A, B, C, D, F, G or H, unscheduled DNA synthesis (UDS) above the residual level was detected in all the cells at a dose of about 10(3) molecules following UV irradiation. The gain numbers of UDS in these XP cells increased with increase in the dose of enzyme and reached a plateau at the normal cell level on introduction of about 10(4) molecules. Introduction of more enzyme into either XP cells or normal human cells did not increase the grain number under regular labelling conditions (2.5 h, 37 degrees C). In normal mouse cells, introduction of the enzyme increased the grain number more than 4-fold under the same conditions during at least 8.5 h following UV irradiation. Furthermore, with a labelling time of 30 min, the enzyme more than doubled the grain number even in normal human cells.
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PMID:Microinjection of T4 endonuclease V produced by a synthetic denV gene stimulates unscheduled DNA synthesis in both xeroderma pigmentosum and normal cells. 291 66

The UV-induced unscheduled DNA synthesis (UDS) in cultured human fibroblasts of repair-deficient xeroderma pigmentosum complementation groups A and C was assayed after injection of identical activities of either Uvr excinuclease (UvrA, B, C and D) from Escherichia coli or endonuclease V from phage T4. Under conditions where the T4 enzyme was able to induce repair synthesis in both XP complementation groups in agreement with earlier observations (de Jonge et al., 1985), no effect of the UvrABCD excinuclease could be observed either when the enzymatic complex was injected into the cytoplasm, or when it was delivered directly into the nucleus. In addition, no effect of the E. coli excinuclease was found on the repair ability of normal repair-proficient human fibroblasts. We conclude that the UvrABCD excinuclease may not work on DNA lesions in human chromatin.
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PMID:Microinjection of Escherichia coli UvrA, B, C and D proteins into fibroblasts of xeroderma pigmentosum complementation groups A and C does not result in restoration of UV-induced unscheduled DNA synthesis. 301 26

In this paper we report both transient and stable complementation of pyrimidine dimer repair in xeroderma pigmentosum cells by the denV gene of bacteriophage T4, coding for endonuclease V, a dimer-specific DNA glycosylase. Cotransfection with pRSVdenV in SV40-transformed XP12RO(M1) cells (complementation group A) restored transient expression of an indicator plasmid (pRSVcat) bearing a UV-inactivated chloramphenicol acetyltransferase (cat) gene. In addition, XP12RO(M1) clones stably transformed by pRSVdenV-SVgpt expressed transient chloramphenicol acetyltransferase activity when transfected with UV-inactivated pRSVcat plasmid. These clones also showed partial restoration of colony forming ability and excision repair synthesis after UV irradiation. Immunofluorescence, using an endonuclease V polyclonal antibody, showed the presence of the phage glycosylase in stably transformed xeroderma pigmentosum cells. The cotransfection assay affords a rapid, sensitive procedure to screen for functional cloned DNA repair genes and to test mutant cells for the deficiency of specific steps in DNA repair, such as incision.
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PMID:Transient and stable complementation of ultraviolet repair in xeroderma pigmentosum cells by the denV gene of bacteriophage T4. 356 13

Roles of poly(ADP-ribose) in repair and replication were studied in UV-irradiated normal, xeroderma pigmentosum (XP), and Cockayne syndrome (CS) fibroblasts. UV radiation reduced cellular NAD with concurrent synthesis of poly-(ADP-ribose) in a dose-dependent manner in normal and CS cells, but not in XP cells. Enzymatic incision of DNA of UV-irradiated XP cells by T4-endonuclease V activated poly(ADP-ribose) polymerase. Methyl methanesulfonate (MMS) also reduced the cellular NAD in all the above strains. However, inhibitors of poly(ADP-ribose) synthesis did not affect UV-induced unscheduled DNA synthesis (UDS) and UV and MMS survivals. Thus, poly(ADP-ribose) synthesis may have a chromatin-stabilizing effect, but not the key role in excision repair of UV damage, unlike in the repair of dimethyl sulfate alkylation. CS cells were deficient in the NAD pool and in the recovery of post-UV DNA synthesis, which were rescued by an exogenous supply of NAD. Such normalization in CS cells was inhibited by excess nicotinamide as in normal cells, suggesting that replicon reinitiation may require specific poly(ADP-ribosyl)ation in UV-irradiated human cells.
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PMID:Roles of poly(ADP-ribose) synthesis in repair and replication in normal human, Cockayne syndrome, and xeroderma pigmentosum fibroblasts after UV irradiation. 641 14

The distribution of ultraviolet-induced DNA repair patches in the genome of xeroderma pigmentosum cells of complementation group C was investigated by determining the molecular weight distribution of repair labeled DNA and prelabeled DNA in alkaline sucrose gradients after treatment with the dimerspecific endonuclease V of bacteriophage T4. The results were consistent with the data reported by Mansbridge and Hanawalt (1983) and suggest that DNA-repair synthesis in xeroderma pigmentosum cells of complementation group C occurs in localized regions of the genome. Analysis of the spatial distribution of ultraviolet-induced repair patches in DNA loops attached to the nuclear matrix revealed that in xeroderma pigmentosum cells of complementation group C repair patches are preferentially situated near the attachment sites of DNA loops at the nuclear matrix. In normal human fibroblasts we observed no enrichment of repair-labeled DNA at the nuclear matrix and repair patches appeared to be distributed randomly along the DNA loops. The enrichment of repair-labeled DNA at the nuclear matrix in xeroderma pigmentosum cells of complementation group C may indicate that the residual DNA-repair synthesis in these cells occurs preferentially in transcribing regions of the genome.
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PMID:Preferential repair of nuclear matrix associated DNA in xeroderma pigmentosum complementation group C. 649 70

Endonuclease V of bacteriophage T4 possesses two enzymatic activities, a DNA N-glycosylase specific for cyclobutane pyrimidine dimers (CBPD) and an associated apurinic/apyrimidinic (AP) lyase. Extensive structural and functional studies of endonuclease V have revealed that specific amino acids are associated with these two activities. Controversy still exists regarding the role of the aromatic amino acid stretch close to the carboxyl terminus, in particular the tryptophan at position 128. We have expressed wild-type and mutant W128S endonuclease V in Escherichia coli from an inducible tac promoter. Purified W128S endonuclease V demonstrated substantially decreased N-glycosylase (approximately 5-fold) and AP lyase (10- to 20-fold) activities in vitro compared to the wild-type enzyme when a UV-irradiated poly(dA)-poly(dT) substrate was used. However, a much smaller difference in AP lyase activity between the two forms was observed with a site-specific abasic oligonucleotide. The difference in enzymatic activity was qualitatively, but not quantitatively, reflected in the survival of UV-irradiated bacteria, that is the W128S cells were slightly less UV resistant than wild-type cells. No difference was observed in the complementation of UV repair using UV-damaged denV- T4 phage. A more pronounced difference between the wild-type and W128S proteins was observed in human xeroderma pigmentosum group A cells by host-cell reactivation of a UV-irradiated reporter gene. The relatively large discrepancy between the in vitro and in vivo results observed with bacteria may be because saturated levels of DNA repair are obtained in vivo with relatively low levels of endonuclease V. However, under limiting in vitro conditions and in human cells in vivo a considerable difference between the W128S mutant and wild-type endonuclease V activities can be detected. Our results demonstrate that tryptophan-128 is important for endonuclease V activity.
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PMID:Replacing tryptophan-128 of T4 endonuclease V with a serine residue results in decreased enzymatic activity in vitro and in vivo. 747 8