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:2.4.2.30 (PARP)
13,611 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Folate is an essential cofactor in the generation of endogenous methionine, and there is evidence that folate deficiency exacerbates the effects of a diet low in choline and methionine, including alterations in poly(ADP-ribose) polymerase (PARP) activity, an enzyme associated with DNA replication and repair. Because PARP requires NAD as its substrate, we postulated that a deficiency of both folate and niacin would enhance the development of liver cancer in rats fed a diet deficient in methionine and choline. In two experiments, rats were fed choline- and folate-deficient, low methionine diets containing either 12 or 8% casein (12% MCFD, 8% MCFD) or 6% casein and 6% gelatin with niacin (MCFD) or without niacin (MCFND) and were compared with folate-supplemented controls. Liver NAD concentrations were lower in all methyl-deficient rats after 2-17 mo. At 17 mo, NAD concentrations in other tissues of rats fed these diets were also lower than in controls. Compared with control values, liver PARP activity was enhanced in rats fed the 12% MCFD diet but was lower in MCFND-fed rats following a further reduction in liver NAD concentration. These changes in PARP activity associated with lower NAD concentrations may slow DNA repair and enhance DNA damage. Only rats fed the MCFD and MCFND diets developed hepatocarcinomas after 12-17 mo. In Experiment 2, hepatocarcinomas were found in 100% of rats fed the MCFD and MCFND diets. These preliminary results indicate that folic acid deficiency enhances tumor development. Because tumors developed in 100% of the MCFD-fed rats and because tissue concentrations of NAD in these animals were also low, further studies are needed to clearly define the role of niacin in methyl-deficient rats.
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PMID:Male rats fed methyl- and folate-deficient diets with or without niacin develop hepatic carcinomas associated with decreased tissue NAD concentrations and altered poly(ADP-ribose) polymerase activity. 904 May 40

The biochemical death cascade of apoptosis is separate from, although induced by, the anticancer drug-target interaction. The failure of many of our chemotherapeutic agents reflects an inability of anticancer drugs to induce apoptosis. Understanding the basic cellular mechanisms that control apoptosis will greatly increase our ability to treat cancer. Identification of the components of the apoptotic biochemical cascade will present new targets for complementary enhancement of chemotherapeutically induced cancer cell death. One factor that has been directly implicated in apoptosis is adenosine triphosphate (ATP). Nevertheless, in this regard, ATP is controversial. This commentary takes issue with dogma, and points to the need for additional thought and research in this field. ATP-depleting therapy of tumor-bearing mice has been shown to induce a marked therapeutic result with minimal mortality, and this effect can be further enhanced when combined with chemotherapy. The definitive mechanism of action is still controversial, although several mechanisms for ATP depletion have been implicated in the process. These include reduction in the mitochondrial transmembrane potential, activation of poly (ADP-ribose) polymerase (PARP) and depletion of the coenzyme nicotinamide adenine dinucleotide (NAD+). Even though the definitive experiments have yet to be carried out, the identification of ATP depletion as a critical determinant in apoptosis should allow for the development of new therapeutic strategies in the treatment of human cancer.
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PMID:Chemotherapeutically induced DNA damage, ATP depletion, and the apoptotic biochemical cascade. 911 54

Identification of the processing/activation of multiple interleukin-1beta converting enzyme (ICE)-like proteases and their target substrates in the intact cell is critical to our understanding of the apoptotic process. In this study we demonstrate processing/activation of at least four ICE-like proteases during the execution phase of apoptosis in human monocytic tumor THP.1 cells. Apoptosis was accompanied by processing of Ich-1, CPP32, and Mch3alpha to their catalytically active subunits, and lysates from these cells displayed a proteolytic activity with kinetics, characteristic of CPP32/Mch3alpha but not of ICE. Fluorescence-activated cell sorting was used to obtain pure populations of normal and apoptotic cells. In apoptotic cells, extensive cleavage of Ich-1, CPP32, and Mch3alpha. was observed together with proteolysis of the ICE-like protease substrates, poly (ADP-ribose) polymerase (PARP), the 70-kD protein component of U1 small nuclear ribonucleoprotein (U1-70K), and lamins A/B. In contrast, no cleavage of CPP32, Mch3alpha or the substrates was observed in normal cells. In cells exposed to an apoptotic stimulus, some processing of Ich-1 was detected in morphologically normal cells, suggesting that cleavage of Ich-1 may occur early in the apoptotic process. The ICE-like protease inhibitor, benzyloxycarbonyl-Val-Ala-Asp (OMe) fluoromethyl ketone (Z-VAD.FMK), inhibited apoptosis and cleavage of Ich-1, CPP32, Mch3alpha, Mch2alpha, PARP, U1-70K, and lamins. These results suggest that Z-VAD.FMK inhibits apoptosis by inhibiting a key effector protease upstream of Ich-1, CPP32, Mch3alpha, and Mch2alpha. Together these observations demonstrate that processing/activation of Ich-1, CPP32, Mch3alpha, and Mch2alpha accompanies the execution phase of apoptosis in THP.1 cells. This is the first demonstration of the activation of at least four ICE-like proteases in apoptotic cells, providing further evidence for a requirement for the activation of multiple ICE-like proteases during apoptosis.
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PMID:Processing/activation of at least four interleukin-1beta converting enzyme-like proteases occurs during the execution phase of apoptosis in human monocytic tumor cells. 912 56

7-hydroxystaurosporine (UCN-01) is a more selective protein kinase C inhibitor than staurosporine. UCN-01 exhibits antitumor activity in experimental tumor models and is presently in clinical trials. Our study reveals that human myeloblastic leukemia HL60 and K562 and colon carcinoma HT29 cells undergo internucleosomal DNA fragmentation and morphological changes characteristic of apoptosis after UCN-01 treatment. These three cell lines lack functional p53, and K562 and HT29 cells are usually resistant to apoptosis. DNA fragmentation in HT29 and K562 cells occurred after 1 day of treatment while it took less than 4 h in HL60 cells. Cycloheximide prevented UCN-01-induced DNA fragmentation in HT-29 cells, but not in HL60 and K562 cells, suggesting that macromolecular synthesis is selectively required for apoptotic DNA fragmentation in HT29 cells. UCN-01-induced DNA fragmentation was preceded by activation of cyclin B1/cdc2 kinase. Further studies in HL60 cells showed that UCN-01-induced apoptosis was associated with degradation of CPP32, PARP, and lamin B and that the inhibitor of caspases (ICE/CED-3 cysteine proteases), Z-VAD-FMK, and the serine protease inhibitor, DCI, protected HL60 cells from UCN-01-induced DNA fragmentation. However, only DCI and TPCK, but not Z-VAD-FMK, inhibited DNA fragmentation in the HL60 cell-free system, suggesting that serine protease(s) may play a role in the execution phase of apoptosis in HL60 cells treated with UCN-01. Z-VAD-FMK and DCI also inhibited apoptosis in HT29 cells. These data demonstrate that the protein kinase C inhibitor and antitumor agent, UCN-01 is a potent apoptosis inducer in cell lines that are usually resistant to apoptosis and lack p53 and that caspases and probably serine proteases are activated during UCN-01-induced apoptosis.
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PMID:7-Hydroxystaurosporine (UCN-01) induces apoptosis in human colon carcinoma and leukemia cells independently of p53. 926 Sep 9

High levels of expression of the DNA repair enzyme O6-alkylguanine DNA-alkyltransferase (OGAT) (EC 2.1.1.63) account for tumor cell resistance to methylating agents. Previous studies suggested that methylating triazenes might have a potential role for the treatment of acute leukemias with low levels of OGAT. In the current study, we transduced the human OGAT cDNA in OGAT-deficient leukemia cell clones. OGAT-transduced cells were more resistant than their OGAT-deficient counterparts to apoptosis triggered by the methylating triazene temozolomide (TZM), as indicated by the results of flow cytometry, terminal deoxynucleotidyl transferase assay, and analysis of DNA fragmentation. Depletion of OGAT activity by O6-benzylguanine increased leukemia cell sensitivity to TZM-mediated apoptosis. Moreover, combined treatment of cells with TZM and benzamide, an inhibitor of the poly(ADP-ribose) polymerase (EC 2.4.2.30), increased the apoptosis induced by the methylating agent. These results demonstrate for the first time that methyl adducts at the O6 position of guanine, which are specifically removed by OGAT, are the principal DNA lesions responsible for the induction of apoptosis on treatment of leukemic cells with the methylating triazene TZM. This study also supports the possible use of TZM for the treatment of acute leukemias and suggests new strategies to increase the susceptibility of tumor cells to methylating triazenes in the clinic.
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PMID:Inhibition of O6-alkylguanine DNA-alkyltransferase or poly(ADP-ribose) polymerase increases susceptibility of leukemic cells to apoptosis induced by temozolomide. 927 47

Mice lacking the gene encoding poly(ADP-ribosyl) transferase (PARP or ADPRT) display no phenotypic abnormalities, although aged mice are susceptible to epidermal hyperplasia and obesity in a mixed genetic background. Whereas embryonic fibroblasts lacking PARP exhibit normal DNA excision repair, they grow more slowly in vitro. Here we investigated the putative roles of PARP in cell proliferation, cell death, radiosensitivity, and DNA recombination, as well as chromosomal stability. We show that the proliferation deficiency in vitro and in vivo is most likely caused by a hypersensitive response to environmental stress. Although PARP is specifically cleaved during apoptosis, cells lacking this molecule apoptosed normally in response to treatment with anti-Fas, tumor neurosis factor alpha, gamma-irradiation, and dexamethasone, indicating that PARP is dispensable in apoptosis and that PARP-/- thymocytes are not hypersensitive to ionizing radiation. Furthermore, the capacity of mutant cells to carry out immunoglobulin class switching and V(D)J recombination is normal. Finally, primary PARP mutant fibroblasts and splenocytes exhibited an elevated frequency of spontaneous sister chromatid exchanges and elevated micronuclei formation after treatment with genotoxic agents, establishing an important role for PARP in the maintenance of genomic integrity.
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PMID:PARP is important for genomic stability but dispensable in apoptosis. 930 63

The tumor suppressor gene p53 is expressed in the contrasting cell fates apoptosis and proliferation. We examined whether the transactivation of the p53 target genes, waf1 and mdm2, is dependent on the cause of p53 induction in human peripheral blood mononuclear cells (PBMC). Both apoptosis triggered by the purine analog 2-chlorodeoxyadenosine (CdA) and growth stimulation by the mitogen phytohemagglutinin (PHA) induced a comparable level and time course of p53 mRNA expression. Both stimuli led also to an increase of p53 protein levels. The cytotoxic agent, but not the mitogen, led to transactivation of waf1 and mdm2 within 18 h. Transactivation was followed by apoptosis of 89% of the PBMC within 48 h. The c-myc oncogene and poly(ADP-ribose)polymerase (PARP), which also have a dual function in proliferation and apoptosis, showed an early induction by both CdA and PHA. These results add further evidence that growth stimulation and DNA damage-induced apoptosis share early gene activation pathways in normal cells. However, since p53 does selectively translate into transactivation of target genes depending on the cause of induction, this function of p53 seems to be regulated by additional factors, which are closely related to the ultimate fate of the cell.
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PMID:Type of inducing signal regulates transactivation by p53. 936 63

An in vitro system has been employed to study the apoptotic mechanisms in the AK-5 tumor which is a spontaneously regressing rat histiocytoma. Cytosolic extracts of tumor cells primed for apoptosis using dexamethasone and immune serum from tumor-regressing animals were able to induce apoptosis in intact nuclei and reproduce the classical morphological and biochemical features typical of apoptotic cells. The cleavage of lamin A and PARP to signature fragments by these extracts and the inhibition of the same using peptide inhibitors signify the pivotal role of ICE and ICE-related proteases in apoptosis. Lamin A cleavage was insensitive to YVAD but PARP cleavage was blocked by both YVAD and DEVD. Cell extracts derived from cells overexpressing the Bcl-2 gene and Nedd-2 antisense gene, respectively, failed to induce apoptosis in exogenously added nuclei, suggesting that Bcl-2 gene product is downregulating a key event in apoptotic cascade. The study also demonstrates the coherent action of different ICE-related proteases in apoptosis and their functional redundancy. This system may prove useful for analyzing complex molecular mechanisms underlying apoptosis in tumor cells.
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PMID:Caspase-mediated apoptosis in AK-5 tumor cells: a cell-free study using peptide inhibitors and antisense strategy. 936 20

The molecular mechanisms for sensitivity and resistance of tumor cells towards chemotherapy are only partially understood. In chemosensitive leukemias and solid tumors, anticancer drugs have been shown to induce apoptosis. We previously identified activation of the CD95 (APO-1/Fas) receptor/CD95 ligand (CD95/CD95-L) system as a key mechanism for drug-induced apoptosis. Here, we show that therapeutic concentrations of doxorubicin, methotrexate and cytarabine also induce apoptosis via activation of the CD95 system in primary leukemia cells in vivo. CD95-resistant and doxorubicin-resistant leukemia and neuroblastoma cells display cross-resistance for induction of cell death. Down-regulation of CD95 expression was found in drug-resistant and CD95-resistant cell lines. Furthermore, up-regulation of CD95-L, previously shown to mediate drug-induced apoptosis in a variety of tumor cells, was completely blocked in doxorubicin-resistant cells. The prototype caspase (ICE/Ced-3 protease) substrate, poly(ADP-ribose)polymerase (PARP), was cleaved in sensitive, but not in resistant tumor cells following CD95 triggering or drug treatment. Since failure to activate CD95-L was not due to decreased drug uptake or increased drug efflux, non-multi-drug resistance (non-MDR) mechanisms are involved in this type of resistance. These findings suggested that an intact CD95 system plays a key role in determining sensitivity or resistance towards anticancer therapy.
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PMID:Deficient activation of the CD95 (APO-1/Fas) system in drug-resistant cells. 936 15

The activation of the caspase family of cysteine proteases is a key step in the implementation of apoptotic cell death leading to further downstream effects such as DNA fragmentation. In cultured tumor cells, caspase activity appears only when cells are undergoing apoptosis. Here we show that human and murine T lymphocytes acquire high intracellular activities of cell death-specific caspases upon activation by mitogens and IL-2 without evidence that apoptosis is proceeding. The highest activity is seen when cells are mitogen activated for 3 days. On a per cell basis, caspase activity in activated T cells is much higher than in tumor cells induced to undergo apoptosis. In the presence of exogenously added IL-2 cells stay alive and maintain a high level of caspase activity while IL-2 withdrawal results in cell death and decline of caspase activity. Caspase activity can also be measured in extracts from spleen and lymph nodes from mice injected with superantigen. While in tumor cell lines caspase activity correlates with cleavage of poly(ADP)-ribose polymerase (PARP) and DNA fragmentation, in activated T cells cleavage products of cellular PARP can be detected whereas DNA fragmenting activity appears only upon IL-2 withdrawal which coincides with cell death. These data show that caspase activation in intact cells does not necessarily lead to cell death and argue for a checkpoint in the apoptotic pathway downstream of caspases. Furthermore, they provide a molecular correlate for the high susceptibility of activated T cells for apoptosis.
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PMID:Activation of caspase-3-like enzymes in non-apoptotic T cells. 954 84


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