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)

Poly(ADP-ribose) polymerase (PARP) is an element of the DNA damage surveillance network evolved by eukaryotic cells to cope with numerous environmental and endogenous genotoxic agents. PARP has been found to be involved in vivo in both cell proliferation and base excision repair of DNA. In this study the interaction between PARP and the DNA polymerase alpha-primase tetramer has been examined. We provide evidence that in proliferating cells: (i) PARP is physically associated with the catalytic subunit of the DNA polymerase alpha-primase tetramer, an association confirmed by confocal microscopy, demonstrating that both enzymes are co-localized at the nuclear periphery of HeLa cells; (ii) this interaction requires the integrity of the second zinc finger of PARP and is maximal during the S and G2/M phases of the cell cycle; (iii) PARP-deficient cells derived from PARP knock-out mice exhibited reduced DNA polymerase activity, compared with the parental cells, a reduction accentuated following exposure to sublethal doses of methylmethanesulfonate. Altogether, the present results strongly suggest that PARP participates in a DNA damage survey mechanism implying its nick-sensor function as part of the control of replication fork progression when breaks are present in the template.
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PMID:Functional association of poly(ADP-ribose) polymerase with DNA polymerase alpha-primase complex: a link between DNA strand break detection and DNA replication. 951 81

Poly(ADP-ribose) polymerase (PARP) is a nuclear enzyme which is catalytically activated by DNA strand interruptions. The involvement of PARP has been implicated in different cellular responses to genotoxic damage, including cell survival, DNA repair, transformation, and cell death. However, the exact contribution of PARP polypeptide or its enzymatic product has remained ill defined. Recent studies with two different PARP knock out mice have demonstrated the beneficial role of PARP in maintaining genomic integrity and in survival responses after exposure to whole body gamma-irradiation. Other studies have demonstrated the instrumental role of PARP in death of the neuronal cells after ischemia-reperfusion injury. The recombination inhibiting function of PARP at DNA strand breaks was more evident in a model system deficient in activities of two major DNA strand break binding proteins, PARP and DNA-dependent protein kinase. The present review summarizes similarities and differences obtained with the two PARP knock out mice and reanalyzes the role of PARP in various cellular responses to DNA damage.
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PMID:Cellular responses to DNA damage in the absence of Poly(ADP-ribose) polymerase. 953 73

Poly(ADP-ribose) polymerase (PARP; EC 2.4.2.30) is a zinc-finger DNA-binding protein that detects and signals DNA strand breaks generated directly or indirectly by genotoxic agents. In response to these breaks, the immediate poly(ADP-ribosyl)ation of nuclear proteins involved in chromatin architecture and DNA metabolism converts DNA damage into intracellular signals that can activate DNA repair programs or cell death options. To have greater insight into the physiological function of this enzyme, we have used the two-hybrid system to find genes encoding proteins putatively interacting with PARP. We have identified a physical association between PARP and the base excision repair (BER) protein XRCC1 (X-ray repair cross-complementing 1) in the Saccharomyces cerevisiae system, which was further confirmed to exist in mammalian cells. XRCC1 interacts with PARP by its central region (amino acids 301 to 402), which contains a BRCT (BRCA1 C terminus) module, a widespread motif in DNA repair and DNA damage-responsive cell cycle checkpoint proteins. Overexpression of XRCC1 in Cos-7 or HeLa cells dramatically decreases PARP activity in vivo, reinforcing the potential protective function of PARP at DNA breaks. Given that XRCC1 is also associated with DNA ligase III via a second BRCT module and with DNA polymerase beta, our results provide strong evidence that PARP is a member of a BER multiprotein complex involved in the detection of DNA interruptions and possibly in the recruitment of XRCC1 and its partners for efficient processing of these breaks in a coordinated manner. The modular organizations of these interactors, associated with small conserved domains, may contribute to increasing the efficiency of the overall pathway.
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PMID:XRCC1 is specifically associated with poly(ADP-ribose) polymerase and negatively regulates its activity following DNA damage. 958 96

Poly(ADP-ribose) polymerase (PARP) is a component of the multiprotein DNA replication complex (MRC, DNA synthesome) that catalyzes replication of viral DNA in vitro. PARP poly(ADP-ribosyl)ates 15 of the approximately 40 proteins of the MRC, including DNA polymerase alpha (DNA pol alpha), DNA topoisomerase I (topo I), and proliferating-cell nuclear antigen (PCNA). Although about equal amounts of MRC-complexed and free forms of PCNA were detected by immunoblot analysis of HeLa cell extracts, only the complexed form was poly(ADP-ribosyl)ated, suggesting that poly(ADP-ribosyl)ation of PCNA may regulate its function within the MRC. NAD inhibited the activity of DNA pol delta in the MRC in a dose-dependent manner, whereas the PARP inhibitor, 3-AB, reversed this inhibitory effect. The roles of PARP in modulating the composition and enzyme activities of the DNA synthesome were further investigated by characterizing the complex purified from 3T3-L1 cells before and 24 h after induction of a round of DNA replication required for differentiation of these cells; at the latter time point, approximately 95% of the cells are in S phase and exhibit a transient peak of PARP expression. The MRC was also purified from similarly treated 3T3-L1 cells depleted of PARP by antisense RNA expression; these cells do not undergo DNA replication nor terminal differentiation. Both PARP protein and activity and essentially all of the DNA pol alpha and delta activities exclusively cosedimented with the MRC fractions from S phase control cells, and were not detected in the MRC fractions from PARP-antisense or uninduced control cells. Immunoblot analysis further revealed that, although PCNA and topo I were present in total extracts from both control and PARP-antisense cells, they were present in the MRC fraction only from induced control cells, indicating that PARP may play a role in their assembly into an active DNA synthesome. In contrast, expression of DNA pol alpha, DNA primase, and RPA was down-regulated in PARP-antisense cells, suggesting that PARP may be involved in the expression of these proteins. Depletion of PARP also prevented induction of the expression of the transcription factor E2F-1, which positively regulates transcription of the DNA pol alpha and PCNA genes; thus, PARP may be necessary for expression of these genes when quiescent cells are stimulated to proliferate.
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PMID:Regulation of the expression or recruitment of components of the DNA synthesome by poly(ADP-ribose) polymerase. 964 17

Poly(ADP-ribose) polymerase (PARP) is an abundant nuclear enzyme which is responsible for synthesis of poly(ADP-ribose) in response to DNA damage caused by numerous agents and during DNA base excision repair. After DNA damage, the enzyme binds to nicks in DNA through its N-terminal zinc fingers and catalyzes the formation of poly(ADP-ribose) on various nuclear acceptors including itself. When DNA damage is extensive, cells induce their own demise by activating the proteases that induce apoptosis (caspases) which cleave PARP and other death substrates. Here we report the development of a new approach to investigate the sensitivity of mono(ADP-ribosyl)ated and DNA-bound PARP to cleavage during apoptosis. The development of a stoichiometric labeling procedure of the enzyme has allowed us to evaluate the catalytic properties of caspase 3 toward mono(ADP-ribosyl)ated PARP at various enzyme:substrate molar ratios. We show that low levels of automodification (< or = 3 U of ADP-ribose per chain) do not inhibit the proteolysis of the substrate. In addition, we demonstrate that binding of unmodified PARP to DNA influences the kinetics of its cleavage by caspase 3.
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PMID:Proteolysis of poly(ADP-ribose) polymerase by caspase 3: kinetics of cleavage of mono(ADP-ribosyl)ated and DNA-bound substrates. 965 May 95

Poly(ADP-ribose) polymerase (PARP) is a DNA-binding protein involved in cellular response to various genotoxic agents. To understand the role of PARP in the mechanisms which lead from specific DNA damage to cell death, we studied the effects of PARP inhibition in human lymphoblasts damaged with bleomycin (BLM) and VP16. These agents can induce DNA breakage but through different mechanisms, enabling the study of the different effects of PARP in inducing apoptosis in damaged cells. We demonstrate that in lymphoblasts VP16 treatment induces apoptosis to a greater extent than BLM treatment, and that PARP inhibition reduces VP16-induced apoptosis whereas it has no effect on BLM-induced apoptosis. After VP16 treatment with PARP inhibition, a reduction in the depletion of the proliferative compartment and a G2/M phase arrest are observed. Therefore, the increase in cell viability and the reduction in chromosome damage may both be the result of a prolonged DNA repair time. Hence, PARP appears to play a significant role in VP16-induced apoptosis and not in BLM-induced apoptosis. Since apoptosis is important in tumor treatment these findings might be useful when considering the combined employment of PARP inhibition with antineoplastic drugs.
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PMID:Effects of poly(ADP-ribose) polymerase inhibition on cell death and chromosome damage induced by VP16 and bleomycin. 970 99

Poly(ADP-ribose) polymerase (PARP) is thought to be involved in DNA repair given its ability to recognize and bind to DNA strand breaks. During apoptosis, PARP is proteolytically cleaved into two stable fragments, the N-terminal 25-kDa DNA-binding domain (DBD) and the 85-kDa fragment containing the automodification and catalytic domains. To understand the DNA-binding properties of PARP, we expressed a recombinant hexahistidine tagged protein (His-DBD) in Escherichia coli. We modified expression to facilitate protein folding by including zinc and reducing the induction temperature. Properly folded, the DNA-binding domain of PARP binds to single- and double-stranded DNA in a structure-specific manner. To eliminate contamination with bacterial DNA that occurred during the purification process, a purification procedure was developed to produce DNA-free protein. In addition, to remove the hexahistidine tag from the recombinant protein, thrombin cleavage was carried out while the recombinant protein was bound to a DNA column. This procedure stabilized the recombinant protein and resulted in nearly 100% cleavage with no appreciable loss to unwanted proteolytic degradation. This nondenaturing purification scheme results in high-quality, native PARP-DBD for use in structural and biochemical studies.
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PMID:A nondenaturing purification scheme for the DNA-binding domain of poly(ADP-ribose) polymerase, a structure-specific DNA-binding protein. 975 54

Poly(ADP-ribose) polymerase (PARP) is a highly abundant nuclear enzyme which metabolizes NAD, in response to DNA strand breakage, to produce chains of poly(ADP-ribose) attached to nuclear proteins. PARP activation has been implicated in ischemia/reperfusion injury, but its biological significance is not fully understood. We have modified an existing in situ method for detection of PARP activity by using an NAD analogue in which adenine is modified by an "etheno" (vinyl) bridge. Etheno-NAD serves as a PARP substrate in an initial enzymatic reaction; a specific antibody to ethenoadenosine is then used in an immunohistochemical reaction to detect the production of modified poly(ADP-ribose). The method produces strong and specific labeling of nuclei in which PARP has been activated, i.e., those in which DNA strand breaks have been produced, and the results can be analyzed by microscopy, flow cytometry, or colorimetry. The method is applicable to cultured cells in several formats and to frozen tissue sections. The particular characteristics of the new method may assist in future in situ studies of PARP activation.
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PMID:In situ staining for poly(ADP-ribose) polymerase activity using an NAD analogue. 977 27

Poly(ADP-ribose) polymerase (PARP) is a nuclear enzyme, which is activated by DNA strand breaks. Although PARP is known to be cleaved by the cysteine protease, caspase-3/CPP32, during apoptosis, signal cascade which regulates the PARP activity has not been fully understood. In this study, we investigated post-translational modification of PARP. We found that PARP was phosphorylated by a serine kinase in vivo. PARP was activated temporarily and extensive auto-modification occurred on PARP, possibly by the fragmented DNA during apoptosis induced by etoposide in Jurkat cells. However, the phosphorylation level was not changed for up to 6 h, after PARP cleavage began in apoptosis by the treatment with etoposide. Furthermore, we showed the presence of a PARP-associated kinase in nuclear extracts of the HTLV-I infected T-cell lines but not in uninfected T-cell lines, whereas this kinase did not inhibit the PARP activity even in the presence of ATP. Taken together, in vivo phosphorylation of PARP might be independent of the activation or cleavage of PARP.
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PMID:In vivo phosphorylation of poly(ADP-ribose) polymerase is independent of its activation. 978 97

Poly(ADP-ribose) polymerase (PARP) (EC 2.4.2.30), the only enzyme known to synthesize ADP-ribose polymers from NAD+, is activated in response to DNA strand breaks and functions in the maintenance of genomic integrity. Mice homozygous for a disrupted gene encoding PARP are viable but have severe sensitivity to gamma-radiation and alkylating agents. We demonstrate here that both 3T3 and primary embryo cells derived from PARP-/- mice synthesized ADP-ribose polymers following treatment with the DNA-damaging agent, N-methyl-N'-nitro-N-nitrosoguanidine, despite the fact that no PARP protein was detected in these cells. ADP-ribose polymers isolated from PARP-/- cells were indistinguishable from that of PARP+/+ cells by several criteria. First, they bound to a boronate resin selective for ADP-ribose polymers. Second, treatment of polymers with snake venom phosphodiesterase and alkaline phosphatase yielded ribosyladenosine, a nucleoside diagnostic for the unique ribosyl-ribosyl linkages of ADP-ribose polymers. Third, they were digested by treatment with recombinant poly(ADP-ribose) glycohydrolase, an enzyme highly specific for ADP-ribose polymers. Collectively, these data demonstrate that ADP-ribose polymers are formed in PARP-/- cells in a DNA damage-dependent manner. Because the PARP gene has been disrupted, these results suggest the presence of a previously unreported activity capable of synthesizing ADP-ribose polymers in PARP-/- cells.
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PMID:Poly(ADP-ribose) polymerase null mouse cells synthesize ADP-ribose polymers. 980 57


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