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)

Ara-C has been shown to induce apoptosis of human acute myelogenous leukemia HL-60 cells. The DNA repair enzyme poly(ADP-ribose) polymerase (PARP) is known to be degraded during apoptosis. PARP as a substrate is cleaved by the Yama protease, encoded by the CPP32beta/Yama gene. Yama belongs to the interleukin 1beta converting enzyme/ced-3 family of cysteine proteases that are activated as a cascade, producing proteolytic cleavage of specific substrates that results in the morphological and biochemical features of apoptosis. In the present studies, we determined the effect of high intracellular levels of the antiapoptosis Bcl-2 or Bcl-xL protein on Yama protease activation and PARP degradation during Ara-C-induced apoptosis. For this, we utilized HL-60/Bcl-2, HL-60/Bcl-xL, or control HL-60/neo cells, which were created by transfection of the cDNA of the bcl-2, bcl-xL, or the neomycin-resistant genes, respectively. As compared to HL-60/neo, HL-60/Bcl-2 and HL-60/Bcl-xL cells have 5-fold greater expression of Bcl-2 and Bcl-xL, respectively. However, these cell lines have similar levels of p32Yama and PARP. Treatment with 10 or 100 microM Ara-C for 4 h produced DNA fragmentation and morphological features of apoptosis in HL-60/neo cells. This was associated with the cleavage and activation of p32Yama and PARP degradation but not with the induction of Yama mRNA. In contrast, in HL-60/Bcl-2 and HL-60/ Bcl-xL cells, Ara-C-induced p32Yama activation by its cleavage, PARP degradation and apoptosis were significantly inhibited. High Bcl-2 and Bcl-xL levels in these cells also inhibited Yama protease activity, PARP degradation, and apoptosis due to clinically relevant concentrations of etoposide and mitoxantrone. These results suggest that the activation of the Yama protease and PARP degradation are involved in Ara-C-, etoposide-, or mitoxantrone-induced apoptosis. In addition, they suggest that Bcl-2 and Bcl-xL antagonize drug-induced apoptosis by a mechanism that interferes in the activity of a key cysteine protease that is involved in the execution of apoptosis.
Cancer Res 1996 Oct 15
PMID:Overexpression of Bcl-2 or Bcl-xL inhibits Ara-C-induced CPP32/Yama protease activity and apoptosis of human acute myelogenous leukemia HL-60 cells. 884 Sep 93

Ultraviolet B (UVB) component of the sunlight is the major cause of nonmelanoma skin cancer (NMSC) in humans. UVB is absorbed directly by cellular DNA and produces lesions that may cause mutation(s) in target gene(s) ultimately leading to cancer. Early detection of these lesions, therefore, may help to identify individuals at a high risk to develop NMSC, and devise approaches for the prevention of this common malignancy. Employing mouse skin as a model, we applied a 32P postlabelling method to detect UVB-induced DNA lesions in the epidermis in nanomole quantities. Autoradiography maps showed that epidermal DNA from UVB exposed mice at 24 h contain up to five DNA lesions; the quantitation of these lesions showed that their formation increased in a UVB dose-dependent manner. Treatment of DNA samples with the bacteriophage DNA repair enzyme T4 endonuclease V confirmed that four of these lesions are pyrimidine dimers. While, some of these lesions were repaired 18 h after UVB irradiation, 30% of them persisted even 48 h post-irradiation. Application of a sunscreen containing ethylhexyl-p-methoxycinnamate or chemopreventive agent green tea polyphenols or silymarin to the skin of the mice prior to UVB exposure was found to prevent the formation of pyrimidine dimers.
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PMID:Ultraviolet B radiation-induced DNA lesions in mouse epidermis: an assessment using a novel 32P-postlabelling technique. 895 42

Repair of alkylated bases in DNA is performed by O6-methylguanine-DNA methyltransferase (MGMT) and a set of enzymes of the base excision repair pathway involving N-methylpurine-DNA glycosylase (MPG), apurinic endonuclease (APE), DNA polymerase beta (Pol beta) and DNA ligase. The level of expression of these enzymes may exert a profound effect on resistance of cells towards alkylating drugs. We have comparatively analyzed the expression of MGMT and the different base excision repair genes in rat hepatoma cells (line H4IIE) after exposure to alkylating agents, X-rays and the glucocorticoid hormone dexamethasone. Furthermore, the effect of these agents on the activity of the cloned human MGMT promoter was assayed. Exposure of cells to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or ionizing radiation increased MGMT mRNA levels up to 4.5-fold. Under the same conditions of treatment, exerting only a weak toxic effect, MPG and DNA ligase I mRNA levels were not enhanced, whereas the amounts of APE and Pol beta mRNA transiently increased by approximately 2-fold after X-ray and MNNG treatment, respectively. Dexamethasone induced both MGMT, APE and Pol beta mRNA and the induction paralleled the increase in mRNA of the glucocorticoid-dependent gene tyrosine aminotransferase. The observed increase in MGMT mRNA was due to promoter activation, which was shown in transient transfection assays with MGMT promoter-CAT reporter constructs in H4IIE cells. In these assays, the human MGMT promoter was found to be induced by methylating agents (MNNG and methyl methanesulfonate), ionizing radiation and dexamethasone. Weak induction of the promoter was observed after UV irradiation. Treatment with the tumor promoter 12-O-tetradecanoyl-phorbol-13-acetate was ineffective in promoter activation. The transfected MGMT promoter was not inducible by mutagens in HeLa S3 cells, which do not respond with induction of the endogenous MGMT gene. This is the first report showing hormone induction of a DNA repair gene (MGMT). The induction of MGMT and other genes encoding enzymes involved in DNA alkylation damage repair may be relevant in cancer therapy by causing resistance of tumor cells to alkylating drugs.
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PMID:Induction of the alkyltransferase (MGMT) gene by DNA damaging agents and the glucocorticoid dexamethasone and comparison with the response of base excision repair genes. 896 45

The aim of this study was to determine whether stable differences in apoptosis sensitivity were selected for in nonmetastatic and metastatic variants of the LNCaP human prostate carcinoma line that had been isolated from tumors grown orthotopically in the prostate glands and regional lymph nodes of nude mice. The nonmetastatic LNCaP-Pro5 cells were significantly more sensitive to thapsigargin-induced apoptosis than were the metastatic LNCaP-LN3 cells, as measured by viability, DNA fragmentation, and interleukin 1beta-converting enzyme family-mediated cleavage of the DNA repair enzyme, poly(ADP-ribose) polymerase. Apoptosis resistance in the metastatic cells was associated with higher levels of expression of the cell death suppressor BCL-2 and lower levels of the death promoters BAX and BAK than were detected in the nonmetastatic LNCaP-Pro5 cells, whereas levels of two other BCL-2 family members (BCL-X(L) and BAD) were indistinguishable. Our data support the hypothesis that apoptosis resistance contributes to prostate cancer metastasis and that elevated expression of BCL-2 is involved.
Cancer Res 1996 Dec 15
PMID:Apoptosis resistance increases with metastatic potential in cells of the human LNCaP prostate carcinoma line. 897 Nov 61

Recent evidence suggests that members of the interleukin-1-beta-converting enzyme (ICE)/Ced-3 family are key mediators of mammalian apoptosis. The known members of the ICE/Ced-3 cysteine protease family are synthesized as proenzymes and require proteolytic processing to produce active, heterodimeric enzymes. The baculovirus protein P35 has recently been shown to inhibit several members of the ICE/Ced-3 cysteine protease family. The importance of ICE/Ced-3 cysteine proteases in programmed cell death prompted us to investigate the role of the apoptotic mediator, CPP32, in the glucocorticoid-mediated cell death pathway. Glucocorticoids induce growth inhibition and apoptosis in sensitive leukemic cell lines, immature thymocytes, and eosinophils. In this report, we demonstrate the enzymatic cleavage of proCPP32 to its active subunits in cells undergoing glucocorticoid-induced apoptotic cell death. Concurrently, in apoptotic cells, PARP, a 116-kilodalton (kDa) human poly(ADP-ribose) polymerase, is proteolytically cleaved to its signature 85-kDa fragment. The proteolytic processing of PARP (the nuclear DNA repair enzyme known to be cleaved in association with apoptosis) is catalyzed by members of the ICE/Ced-3 family. Importantly, stable transfection of the antiapoptotic baculovirus P35 inhibits glucocorticoid-induced apoptotic cell death, proteolytic processing of proCPP32, and cleavage of the 116kDa PARP. We conclude that activation of CPP32 is a critical event in glucocorticoid-induced apoptosis and that this pathway is inhibited at or upstream of CPP32 by baculovirus P35. These data demonstrate that PARP cleavage occurs during glucocorticoid-induced apoptotic cell death and show that this proteolytic process is blocked by the expression of baculovirus P35, supporting a role for activation of the ICE/Ced-3-like cysteine protease during glucocorticoid-induced apoptosis.
Cancer Res 1997 Jan 01
PMID:Baculovirus P35 inhibits the glucocorticoid-mediated pathway of cell death. 898 38

Mammalian DNA polymerase beta is a DNA repair enzyme expressed constitutively at a low level. In vitro, purified DNA polymerase (Pol) beta incorporates the nucleotide analogues 2'-3' deoxycytidine (ddC)-triphosphate and 3'-azido-3'-deoxythymidine (AZT)-triphosphate into DNA, causing chain termination. We have tested the possibility of enhancing the cytotoxicity of these chain terminators against mammalian cells by increasing the level of Pol beta. Chinese hamster ovary AA8 and murine melanoma B16 cell lines were stably transfected with rat pol beta cDNA under the control of a viral enhancer/promoter. We found that overexpression of Pol beta sensitized the cells to ddC and AZT. To confirm the role of this polymerase in this process, we prepared cell extracts from the control and Pol beta overexpressing Chinese hamster ovary cell lines and tested in vitro their capacity to incorporate ddC-triphosphate and AZT-triphosphate into DNA. We found that inhibition of DNA replication by both chain terminators was more pronounced when extracts from pol beta-transfected cells were used, providing a direct evidence of the involvement of Pol beta in the sensitization process. In addition, we showed that cotransfection with bacterial or viral thymidine/thymidylate kinase genes enhanced the Pol beta-mediated cytotoxicity of AZT, suggesting that phosphorylation and polymerization activities might be combined to potentiate their respective effects. These observations may be useful for improving therapeutic efficiency of DNA chain terminators.
Cancer Res 1997 Jan 01
PMID:Overexpression of DNA polymerase beta sensitizes mammalian cells to 2',3'-deoxycytidine and 3'-azido-3'-deoxythymidine. 898 50

Administration of hepatocarcinogens aflatoxin B1 and N-nitrosodimethylamine to rats caused single-strand breaks in nuclear DNA. Inclusion in the diet of rutin, a naturally occurring phenolic flavonoid glycoside, significantly reduced the appearance of such breaks. The protection against DNA damage was found to be reduction in the induction of repair enzymes polymerase, DNA polymerase beta and DNA ligase. Even associated with poly(ADP-ribose) a marginal dose of rutin was effective in this regard. Since DNA damage and inefficient repair are expected to initiate the process of carcinogenesis, modulation by rutin of these parameters emphasizes the protective role of this flavonoid against carcinogenesis induced by chemical carcinogens.
Cancer Lett 1996 Dec 03
PMID:Protective effect of rutin, a flavonol glycoside, on the carcinogen-induced DNA damage and repair enzymes in rats. 902 Sep 19

Poor folate status may be important in the aetiology of several epithelial cell malignancies including cancer of the uterine cervix. Folic acid is essential in the synthesis of purine nucleotides and the pyrimidine nucleoside thymidine and it is probable that imbalances in these DNA precursors negatively effect DNA stability and may ultimately lead to malignant transformation. The development of a modified 'comet assay' using the bacterial DNA repair enzyme uracil DNA glycosylase, to detect misincorporated uracil in human DNA is reported here. The effect of perturbing folic acid and deoxyuridine levels on uracil misincorporation in normal human lymphocytes and cultured human tumour cells was investigated using this assay. HeLa cells and peripheral human lymphocytes incubated as agarose-embedded nucleoids, with 1 unit of uracil DNA glycosylase per microg of DNA, contained low levels of uracil in their DNA. Both HeLa cells and stimulated human lymphocytes cultured in folate-deficient medium were growth arrested. Incubating human lymphocytes in folate-deficient medium significantly increased the level of uracil detected compared with control cells. HeLa cells showed an increase in non-specific DNA damage (strand breaks). Deoxyuridine (100 microM) significantly increased the level of uracil detected in the DNA of both folate-deficient and control HeLa cells. It appears that this modified comet assay specifically detects misincorporated uracil in single human cells. It should, therefore, prove valuable in determining the role of folic acid status in DNA instability and cancer.
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PMID:Uracil misincorporation in human DNA detected using single cell gel electrophoresis. 932 65

The issue of toxins produced by Helicobacter pylori (H. pylori) urgently requires clarification given that the bacterium causes gastric epithelial cell damage which may lead to precancerous and cancerous changes. During an investigation of the possibility of mono(adenosine 5'-diphosphate (ADP)-ribosyl)ation by H. pylori products, as observed for other bacterial toxins, we found that radioactivity of [adenylate-32P]nicotinamide adenine dinucleotide (NAD) is incorporated into an H. pylori protein of 80 kDa after incubation with crude bacterial extract. In contrast, [carbonyl-14C]NAD did not show any radioactivity incorporation. Unexpectedly, treatment of the modified protein with 0.1 N HCl, but not 0.1 N NaOH, released the AMP moiety. Such chemical properties are characteristic of bacterial DNA ligase-AMP complexes. We found that an antibody raised against Escherichia coli DNA ligase [EC 6.5.1.2] immunoprecipitated the modified 80 kDa protein. Our results indicate that incorporation of radioactivity derived from NAD into the 80 kDa protein was due to adenylation, but not mono(ADP-ribosyl)ation, of the DNA ligase of H. pylori.
Jpn J Cancer Res 1997 Oct
PMID:Helicobacter pylori extracts exhibit nicotinamide adenine dinucleotide-derived adenylation but not mono(adenosine 5'-diphosphate-ribosyl)ation of DNA ligase. 941 50

The multifunctional mammalian apurinic/apyrimidinic (AP) endonuclease (APE) is responsible for the repair of AP sites in DNA. In addition, this enzyme has been shown to function as a redox factor facilitating the DNA-binding capability of JUN and FOS, HeLa AP-1, and numerous other transcription factors, including Myb, members of the CREB family and nuclear factor-kappa B. Although previously presumed to be ubiquitously expressed at comparable levels in all tissues and cell types, recent evidence has shown APE to vary significantly in its expression between tissues and even within tissues. To further characterize APE expression at various stages of cervical neoplasia, we investigated the levels of APE protein expression using immunohistochemistry in normal cervix, pre-invasive and invasive squamous lesions of the cervix, as well as in cervical cancer cell lines. We report here that the APE protein is predominantly expressed in the nuclei of cells from both primary tumors and cervical cell lines, but the level of APE protein is significantly and dramatically elevated in cervical cancer tissue. These results implicate the use of anti-APE antibodies as an effective reagent in the early detection of premalignant and malignant cancer of the cervix. These findings are suggestive that the increase of a DNA repair enzyme in cancerous cells may allow these cells to be refractive to chemotherapy.
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PMID:The apurinic/apyrimidinic endonuclease (APE/ref-1) DNA repair enzyme is elevated in premalignant and malignant cervical cancer. 942 67


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