Ation Capacity Research Articles

Inhibition of PARP Sensitizes Chondrosarcoma Cell Lines to Chemo- and Radiotherapy Irrespective of the IDH1 or IDH2 Mutation Status.

Chondrosarcomas are chemo- and radiotherapy resistant and frequently harbor mutations in isocitrate dehydrogenase (IDH1 or IDH2), causing increased levels of D-2-hydroxyglutarate (D-2-HG). DNA repair defects and synthetic lethality with poly(ADP-ribose) polymerase (PARP) inhibition occur in IDH mutant glioma and leukemia models. Here we evaluated DNA repair and PARP inhibition, alone or combined with chemo- or radiotherapy, in chondrosarcoma cell lines with or without endogenous IDH mutations. Chondrosarcoma cell lines treated with the PARP inhibitor talazoparib were examined for dose–response relationships, as well as underlying cell death mechanisms and DNA repair functionality. Talazoparib was combined with chemo- or radiotherapy to evaluate potential synergy. Cell lines treated long term with an inhibitor normalizing D-2-HG levels were investigated for synthetic lethality with talazoparib. We report that talazoparib sensitivity was variable and irrespective of IDH mutation status. All cell lines expressed Ataxia Telangiectasia Mutated (ATM), but a subset was impaired in poly(ADP-ribosyl)ation (PARylation) capacity, homologous recombination, and O-6-methylguanine-DNA methyltransferase (MGMT) expression. Talazoparib synergized with temozolomide or radiation, independent of IDH1 mutant inhibition. This study suggests that talazoparib combined with temozolomide or radiation are promising therapeutic strategies for chondrosarcoma, irrespective of IDH mutation status. A subset of chondrosarcomas may be deficient in nonclassical DNA repair pathways, suggesting that PARP inhibitor sensitivity is multifactorial in chondrosarcoma.

Arsenite binding-induced zinc loss from PARP-1 is equivalent to zinc deficiency in reducing PARP-1 activity, leading to inhibition of DNA repair

Inhibition of DNA repair is a recognized mechanism for arsenic enhancement of ultraviolet radiation-induced DNA damage and carcinogenesis. Poly(ADP-ribose) polymerase-1 (PARP-1), a zinc finger DNA repair protein, has been identified as a sensitive molecular target for arsenic. The zinc finger domains of PARP-1 protein function as a critical structure in DNA recognition and binding. Since cellular poly(ADP-ribosyl)ation capacity has been positively correlated with zinc status in cells, we hypothesize that arsenite binding-induced zinc loss from PARP-1 is equivalent to zinc deficiency in reducing PARP-1 activity, leading to inhibition of DNA repair. To test this hypothesis, we compared the effects of arsenite exposure with zinc deficiency, created by using the membrane-permeable zinc chelator TPEN, on 8-OHdG formation, PARP-1 activity and zinc binding to PARP-1 in HaCat cells. Our results show that arsenite exposure and zinc deficiency had similar effects on PARP-1 protein, whereas supplemental zinc reversed these effects. To investigate the molecular mechanism of zinc loss induced by arsenite, ICP-AES, near UV spectroscopy, fluorescence, and circular dichroism spectroscopy were utilized to examine arsenite binding and occupation of a peptide representing the first zinc finger of PARP-1. We found that arsenite binding as well as zinc loss altered the conformation of zinc finger structure which functionally leads to PARP-1 inhibition. These findings suggest that arsenite binding to PARP-1 protein created similar adverse biological effects as zinc deficiency, which establishes the molecular mechanism for zinc supplementation as a potentially effective treatment to reverse the detrimental outcomes of arsenic exposure.

Plant invasiveness is not linked to the capacity of regeneration from small fragments: an experimental test with 39 stoloniferous species

Fragmentation and vegetative regenera- tion from small fragments may contribute to popula- tion expansion, dispersal and establishment of new populations of introduced plants. However, no study has systematically tested whether a high capacity of vegetative regeneration is associated with a high degree of invasiveness. For small single-node frag- ments, the presence of internodes may increase regen- eration capacity because internodes may store carbohydrates and proteins that can be used for regeneration. We conducted an experiment with 39 stoloniferous plant species to examine the regeneration capacity of small, single-node fragments with or without attached stolon internodes. We asked (1) whether the presence of stolon internodes increases regeneration from single-node fragments, (2) whether regeneration capacity differs between native and introduced species in China, and (3) whether regener- ation capacity is positively associated with plant invasiveness at a regional scale (within China) and at a global scale. Most species could regenerate from single-node fragments, and the presence of internodes increased regeneration rate and subsequent growth and/or asexual reproduction. Regeneration capacity varied greatly among species, but showed no relation- ship to invasiveness, either in China or globally. High regeneration capacity from small fragments may contribute to performance of clonal plants in general, but it does not appear to explain differences in invasiveness among stoloniferous clonal species.

Enzyme characteristics of recombinant poly(ADP-ribose) polymerases-1 of rat and human origin mirror the correlation between cellular poly(ADP-ribosyl)ation capacity and species-specific life span

Poly(ADP-ribosyl)ation is a posttranslational modification, which is involved in many cellular functions, including DNA repair and maintenance of genomic stability, and has also been implicated in cellular and organismal ageing. We have previously reported that maximum poly(ADP-ribosyl)ation capacity in mononuclear blood cells is correlated with mammalian life span. Here we show that the difference between a long-lived and a short-lived species tested (i.e. man and rat) is directly mirrored by the enzymatic parameters of recombinant poly(ADP-ribose) polymerase-1 (PARP-1), i.e. substrate affinity and reaction velocity. In addition, we have characterized two human PARP-1 alleles and assign their activity difference to their respective initial velocity and not substrate affinity.

Effect of zinc on cellular poly(ADP-ribosyl)ation capacity

Poly(ADP-ribosyl)ation is a posttranslational protein modification, which is catalyzed by poly(ADP-ribose) polymerase-1 (PARP-1) and plays a role in DNA repair and maintenance of genomic stability. A decrease in cellular poly(ADP-ribosyl)ation has been implicated in the aging process. As PARP-1 is a zinc finger protein its decreased function might be related to age-related zinc deficiency. To test this hypothesis we assessed cellular poly(ADP-ribosyl)ation capacity in 29 donors from Greece, Italy and Poland as function of age and nutritional zinc status. Our results reveal a positive correlation between cellular poly(ADP-ribosyl)ation capacity and zinc status in human peripheral blood mononuclear cells (PBMC) (p<0.05). We could also confirm a decrease of PARP-1 activity with donor age, highlighting the role of poly(ADP-ribosyl)ation in the aging process. The results demonstrate that zinc supplementation in elderly people can increase the cellular poly(ADP-ribosyl)ation capacity of their PBMC. We speculate that this may help maintain integrity and stability of the genome more efficiently and thus contribute to an extension of healthspan.

Oxidative DNA Damage Repair andparp 1andparp 2Expression in Epstein-Barr Virus-Immortalized B Lymphocyte Cells from Young Subjects, Old Subjects, and Centenarians

Oxidative DNA damage has been implicated in the aging process and in some of its features such as telomere shortening and replicative senescence. Poly(ADP-ribosyl)ation is involved in many molecular and cellular processes, including DNA damage detection and repair, chromatin modification, transcription, and cell death pathways. We decided to examine the behavior of poly(ADP-ribosyl)ation in centenarians, i.e., those subjects who represent the best example of longevity having reached a very advanced age avoiding the main age-associated diseases. In this study we investigated the relationship between DNA repair capacity and poly(ADP-ribose) polymerase activity in Epstein-Barr virus-immortalized B lymphocyte cell lines from subjects of three different groups of age, including centenarians. Our data show that cells from centenarians have characteristics typical of cells from young people both in their capability of priming the mechanism of repair after H(2)O(2) sublethal oxidative damage and in poly(ADP-ribosyl)ation capacity, while in cells from old subjects these phenomena are delayed or decreased. Moreover, cells from old subjects show a constitutive expression level of both parp 1 and parp 2 genes reduced by a half, together with a reduced presence of modified PARP 1 and other poly(ADP-ribosyl)ated chromatin proteins in comparison to cells from young subjects and centenarians. Our data support the hypothesis that this epigenetic modification is an important regulator of the aging process in humans and it appears to be rather preserved in healthy centenarians, the best example of successful aging.

Role of poly(ADP-ribosyl)ation during human spermatogenesis

Genomic stability of cells is known to be linked to their poly(ADP-ribosyl)ation capacity. We aimed to demonstrate, for the first time, the patterns of poly(ADP-ribosyl)ation during human spermatogenesis. Retrospective case-control study. Teaching hospital. Azoospermic men who underwent testicular biopsy for sperm recovery. Testicular biopsy evaluation by immunohistochemistry for the expression of poly(ADP-ribose) polymerase-1 (PARP-1) enzyme and of poly(ADP-ribose) (PAR) (an indicator for PARP activity.) The subcellular localization of both markers in testes with full spermatogenesis (obstructive azoospermia), spermatocyte maturation arrest, or Sertoli cell-only syndrome. Expression of both markers was localized in germ cell nuclei in full spermatogenesis: PAR expression, indicating PARP activity, was exhibited in round and elongating spermatids and in a subpopulation of primary spermatocytes. Strong immunoreactivity for PAR was identified in all of the spermatocytes in maturation arrest at the spermatocyte level. Sertoli cells lacked immunoreactivity for both markers, whereas other somatic testicular cells were rarely immunostained. The detection of PAR expression in germ-line cells and its subcellular localization in meiotic and postmeiotic prophases demonstrates chromatin modifications occurring during spermatogenesis and establishes a key role for poly(ADP-ribosyl)ation in germ cell differentiation, presumably to safeguard DNA integrity.

Determination of cellular poly(ADP-ribosyl)ation capacity by flow cytometry

Determination of cellular poly(ADP-ribosyl)ation capacity by flow cytometry

Flow-cytometric assessment of cellular poly(ADP-ribosyl)ation capacity in peripheral blood lymphocytes

BackgroundPoly(ADP-ribosyl)ation is a posttranslational modification of nuclear proteins catalysed by poly(ADP-ribose) polymerases (PARPs), using NAD+ as a substrate. Activation of PARP-1 is in immediate response to DNA damage generated by endogenous and exogenous damaging agents. It has been implicated in several crucial cellular processes including DNA repair and maintenance of genomic stability, which are both intimately linked with the ageing process. The measurement of cellular poly(ADP-ribosyl)ation capacity, defined as the amount of poly(ADP-ribose) produced under maximal stimulation, is therefore relevant for research on ageing, as well as for a variety of other scientific questions.ResultsThis paper reports a new, robust protocol for the measurement of cellular poly(ADP-ribosyl)ation capacity in PBMC or Jurkat T-cells using flow cytometry, based on a previously established immuno-dot-blot assay. In order to validate the new assay, we determined the dose-response curve of 3-aminobenzamide, a well-known competitive PARP inhibitor, and we derived an IC50 that is very close to the published value. When testing a set of PBMC samples taken from fifteen healthy young human donors, we could confirm the presence of a substantial interindividual variation, as previously observed using a radiometric assay.ConclusionThe methodology described in this paper should be generally useful for the determination of cellular poly(ADP-ribosyl)ation capacity in a wide variety of settings, especially for the comparison of large sets of samples, such as population studies. In contrast to previously published radiometric or immuno-dot-blot assays, the new FACS-based method allows (i) selective analysis of mononuclear cells by gating and (ii) detection of a possible heterogeneity in poly(ADP-ribosyl)ation capacity between cells of the same type.

Misregulation of poly(ADP‐ribose) polymerase‐1 activity and cell type‐specific loss of poly(ADP‐ribose) synthesis in the cerebellum of aged rats

Protein modification by ADP-ribose polymers is a common regulatory mechanism in eukaryotic cells and is involved in several aspects of brain physiology and physiopathology, including neurotransmission, memory formation, neurotoxicity, ageing and age-associated diseases. Here we show age-related misregulation of poly(ADP-ribose) synthesis in rat cerebellum as revealed by: (i) reduced poly(ADP-ribose) polymerase-1 (PARP-1) activation in response to enzymatic DNA cleavage, (ii) altered protein poly(ADP-ribosyl)ation profiles in isolated nuclei, and (iii) cell type-specific loss of poly(ADP-ribosyl)ation capacity in granule cell layer and Purkinje cells in vivo. In particular, although PARP-1 could be detected in virtually all granule cells, only a fraction of them appeared to be actively engaged in poly(ADP-ribose) synthesis and this fraction was reduced in old rat cerebellum. NAD(+), quantified in tissue homogenates, was essentially the same in the cerebellum of young and old rats suggesting that in vivo factors other than PARP-1 content and/or NAD(+) levels may be responsible for the age-associated lowering of poly(ADP-ribose) synthesis. Moreover, PARP-1 expression was substantially down-regulated in Purkinje cells of senescent rats.

Poly(ADP-ribosyl)ation and aging

Poly(ADP-ribosyl)ation is a DNA strand break-driven post-translational modification of proteins catalyzed by poly(ADP-ribose) polymerase-1 (PARP-1), with NAD + serving as substrate. Poly(ADP-ribosyl)ation is triggered by DNA strand breaks, is functionally associated with DNA repair pathways and is a survival factor for cells under low to moderate levels of genotoxic stress. We have previously described a positive correlation between poly(ADP-ribosyl)ation capacity of mononuclear blood cells with longevity of mammalian species. Our comparison of purified recombinant human and rat PARP-1 revealed that this correlation might be explained in part by evolutionary sequence divergence. We have also developed molecular genetic approaches to modulate the poly(ADP-ribosyl)ation status in living cells. Our results revealed that PARP-1 acts as a negative regulator of DNA damage-induced genomic instability, the latter being known as an important driving force for carcinogenesis. Our recent data obtained in transgenic mice with selective expression of a dominant negative version of PARP-1 in basal skin keratinocytes indicate that PARP-1 activity suppresses skin papilloma formation in a two-stage skin carcinogenesis protocol. It is tempting to speculate that increased poly(ADP-ribosyl)ation capacity in long-lived species might help retard the accumulation of DNA damage and of mutations and thus slow down the rate of aging and of carcinogenesis more efficiently as compared with short-lived animals.

L-selegiline potentiates the cellular poly(ADP-ribosyl)ation response to ionizing radiation.

DNA strand breaks induced by alkylating agents, oxidants, or ionizing radiation trigger the covalent modification of nuclear proteins with poly(ADP-ribose), which is catalyzed for the most part by poly(ADP-ribose) polymerase-1 and plays a role in DNA base-excision repair. Poly(ADP-ribosyl)ation capacity of mononuclear blood cells correlates positively with life span of mammalian species. Here, we show that l-selegiline, an anti-Parkinson drug with neuroprotective activity and life span-extending effect in laboratory animals, can potentiate gamma-radiation-induced poly(ADP-ribose) formation in intact cells. COR4 hamster cells were incubated with l-selegiline (50 nM) for various time periods, followed by gamma-irradiation (45 Gy). Quantification of cellular poly(ADP-ribose) levels at 10 min after starting the irradiation revealed significant increases (up to 1.8-fold) in cells preincubated with the drug for 8 h to 7 days compared with drug-free irradiated controls. There was no selegiline-induced change in poly(ADP-ribose) levels of unirradiated cells nor in basal or radiation-induced DNA strand breaks, respectively. Surprisingly, poly(ADP-ribose) polymerase-1 protein was down-regulated by l-selegiline treatment. Addition of l-selegiline to purified poly(ADP-ribose) polymerase-1 did not alter enzymatic activity. In conclusion, the results of the present study identify a novel intervention to potentiate the cellular poly(ADP-ribosyl)ation response. We hypothesize that the effect of l-selegiline is due to modulation of accessory proteins regulating poly(ADP-ribose) polymerase-1 activity and that increased cellular poly- (ADP-ribosyl)ation capacity may contribute to the neuroprotective potential and/or life span extension afforded by l-selegiline.

Physiology and pathophysiology of poly(ADP-ribosyl)ation.

One of the immediate eukaryotic cellular responses to DNA breakage is the covalent post-translational modification of nuclear proteins with poly(ADP-ribose) from NAD+ as precursor, mostly catalysed by poly(ADP-ribose) polymerase-1 (PARP-1). Recently several other polypeptides have been shown to catalyse poly(ADP-ribose) formation. Poly(ADP-ribosyl)ation is involved in a variety of physiological and pathophysiological phenomena. Physiological functions include its participation in DNA-base excision repair, DNA-damage signalling, regulation of genomic stability, and regulation of transcription and proteasomal function, supporting the previously observed correlation of cellular poly(ADP-ribosyl)ation capacity with mammalian life. The pathophysiology effects are mediated through PARP-1 overactivity, which can cause cell suicide by NAD+ depletion. It is apparent that the latter effect underlies the pathogenesis of a wide range of disease states including type-1 diabetes, ischaemic infarcts in various organs, and septic or haemorrhagic shock. Therefore pharmacological modulation of poly(ADP-ribosyl)ation may prove to be an exciting option for various highly prevalent, disabling and even lethal diseases.

New polymorphisms in the human poly(ADP-ribose) polymerase-1 coding sequence: lack of association with longevity or with increased cellular poly(ADP-ribosyl)ation capacity

New polymorphisms in the human poly(ADP-ribose) polymerase-1 coding sequence: lack of association with longevity or with increased cellular poly(ADP-ribosyl)ation capacity

New polymorphisms in the human poly(ADP-ribose) polymerase-1 coding sequence: lack of association with longevity or with increased cellular poly(ADP-ribosyl)ation capacity.

Poly(ADP-ribose) polymerase-1 (PARP-1) encoded by the PARP-1 gene, is a ubiquitous and abundant DNA-binding protein involved in the cellular response to various genotoxic agents. In a previous study we showed that maximal oligonucleotide-stimulated poly(ADP-ribosyl)ation was significantly higher in permeabilised lymphoblastoid cell lines from a French population of centenarians compared with controls aged 20-70 years, supporting the notion that longevity is associated with a genetically determined, high poly(ADP-ribosyl)ation capacity. Here, we describe four new genetic polymorphisms, three of which represent silent nucleotide variants (C402T, T1011C, G1215A), and one of which leads to a valine762-to-alanine exchange (T2444C). We undertook an association study between two of these polymorphisms and human longevity or poly(ADP-ribosyl)ation capacity in permeabilised lymphoblastoid cells. By analysing 648 DNA samples from a French population (324 centenarians and 324 controls) by fluorescent-allele-specific PCR, we showed the absence of any significant enrichment of any of the genotypes in the study of centenarians versus controls. Furthermore, we studied genotype distributions from individuals who had previously been tested for poly(ADP-ribosyl)ation capacity. None of the genotype combinations at any polymorphic site studied could be related to a high or low level of poly(ADP-ribosyl)ation capacity. Together, these results strongly suggest that the longevity-related differences in the poly(ADP-ribosyl)ation capacity of human lymphoblastoid cell lines cannot be explained by genetic polymorphisms in the PARP-1 coding sequence and that other mechanisms have to be considered as potential regulators of specific poly(ADP-ribosyl)ation capacity.

Poly(ADP-ribosyl)ation, genomic instability, and longevity.

Poly(ADP-ribosyl)ation is a DNA strandbreak-driven posttranslational modification of nuclear proteins that is catalyzed by poly(ADP-ribose) polymerase-1 (PARP-1), with NAD+ serving as substrate. Recently, additional PARP isoforms were described that seem to account for a minor fraction of cellular poly(ADP-ribose) synthesis. We have previously described a correlation between poly(ADP-ribosyl)ation capacity of mononuclear leukocytes of various mammalian species and species-specific life span. Likewise, lymphoblastoid cell lines derived from human centenarians display a higher poly(ADP-ribosyl)ation capacity than do controls. At the functional level, recent data show that PARP-1 is a key regulator of alkylation-induced sister-chromatid exchange, imposing a negative control commensurate with the enzyme activity. PARP-1 activity may therefore be responsible for tuning the rate of genomic instability events that are provoked by the constant attack of endogenous and exogenous genotoxins to a level appropriate for the longevity potential of a given organism or species.

Poly(ADP-ribosyl)ation: a posttranslational protein modification linked with genome protection and mammalian longevity.

Poly(ADP-ribosyl)ation is a posttranslational modification of nuclear proteins catalysed by the 113-kDa enzyme poly(ADP-ribose) polymerase-1 (PARP-1) and, to a lesser extent, by several other recently described polypeptides. The catalytic function of PARP-1 is directly stimulated by DNA strand breaks, thus making poly(ADP-ribosyl)ation one of the immediate cellular responses to oxidative and other types of DNA damage. Poly(ADP-ribosyl)ation plays an important role in the recovery of proliferating cells from certain types of DNA damage, and this has been linked mechanistically with an involvement in DNA base-excision repair. Furthermore PARP-1 activity is necessary to maintain genomic stability under conditions of genotoxic stress and is actually a key regulator of alkylation-induced sister-chromatid exchange formation, imposing a control that is strictly negative and commensurate with the enzyme activity level. Finally, there is a positive correlation between the poly(ADP-ribosyl)ation capacity of mononuclear leukocytes of various mammalian species and species-specific life span. Likewise, lymphoblastoid cell lines derived from human centenarians display a higher poly(ADP-ribosyl)ation capacity than controls. In conclusion, PARP-1 may be viewed as a factor that is responsible for downregulating the rate of genomic instability events, which are provoked by the constant attack by endogenous and exogenous DNA-damaging agents, in such a way as to tune them to a level which is just appropriate for the life span potential of a given species.

Quantitative Nonisotopic Immuno-Dot-Blot Method for the Assessment of Cellular Poly(ADP-ribosyl)ation Capacity

Quantitative Nonisotopic Immuno-Dot-Blot Method for the Assessment of Cellular Poly(ADP-ribosyl)ation Capacity

Increased poly(ADP-ribose) polymerase activity in lymphoblastoid cell lines from centenarians.

Poly(ADP-ribosyl)ation is a posttranslational modification of nuclear proteins which is catalyzed by poly(ADP-ribose) polymerase and represents an immediate response of eukaryotic cells to oxidative and other types of DNA damage. Previously a strong correlation had been detected between maximal poly(ADP-ribose) polymerase activity in permeabilized mononuclear leukocytes of various mammalian species and species-specific life span. To study a possible relation between longevity and poly(ADP-ribosyl)ation in humans we measured maximal oligonucleotide-stimulated poly(ADP-ribose) polymerase activity in permeabilized, Epstein-Barr virus transformed lymphoblastoid cell lines from a French population of 49 centenarians and 51 controls aged 20-70 years. Maximal enzyme activity was significantly higher in centenarians than in controls [median of controls: 9035 cpm/10(6) cells (lower quartile: 6156; upper quartile: 11,410); median of centenarians: 10,380 cpm/10(6) cells (lower quartile: 7994; upper quartile: 12,991); P=0.031 by Mann-Whitney U test]. In a subset of 16 controls and 24 centenarians, cellular poly(ADP-ribose) polymerase content was determined by quantitative western blotting, thus allowing the calculation of specific enzyme activity. The latter was significantly higher in centenarians (P=0.006), the median value for centenarians being about 1.6-fold that of controls. Specific poly(ADP-ribose) polymerase activity was a more powerful parameter for differentiating between centenarians and controls than enzyme activity relative to cell number. In addition, in a genetic association study we analyzed 437 DNA samples (239 centenarians and 198 controls) by PCR amplification of a polymorphic dinucleotide repeat located in the promoter region of the poly(ADP-ribose) polymerase gene in an attempt to detect an association between this polymorphic marker and variability of enzyme activity or human longevity. However, this genetic analysis revealed no significant enrichment of any of the alleles or genotypes identified among centenarians or controls, but its power was limited by the relatively weak heterozygosity of this polymorphic marker in our population (51%). Viewed together with previous results on poly(ADP-ribose) polymerase activity in various mammalian species, the present data provide further evidence for the notion that longevity is associated with a high poly(ADP-ribosyl)ation capacity.

Functional overexpression of human poly(ADP-ribose) polymerase in transfected rat tumor cells

Poly(ADP-ribose) polymerase (PARP, EC 2.4.2.30) is a nuclear enzyme possibly involved in DNA base excision repair. The presence of single- or double-strand breaks in DNA stimulates this enzyme to covalently modify acceptor proteins with poly(ADP-ribose) in a reaction that uses NAD+ as substrate. To test the hypothesis that increased PARP activity could promote resistance towards DNA-damaging agents and gamma-radiation, we established stable rat cell transfectants that constitutively express human PARP. A number of subclones that showed different levels of PARP activity were isolated from two primary transfectants of different clonal origin. PARP activity was determined in permeabilized cells after maximal stimulation with a short, double-stranded oligonucleotide. Activity in different human PARP-expressing subclones was increased 1.6- to 3.1-fold compared with non-expressing subclones. In vivo labeling of poly(ADP-ribose) was performed in one of these subclones, revealing that the level of poly(ADP-ribose) accumulation after the same treatment with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) was four times higher in the human PARP-expressing subclone compared with both non-expressing transfected control cells and parental cells. Clonal survival assays revealed a sensitization upon treatment with gamma-radiation (up to 1.4-fold) or MNNG (up to 2.7-fold) of several subclones expressing human PARP; in some others survival was not changed. Survival after cisplatin (DDP) treatment remained essentially unchanged. A protective effect against DNA-damage was never observed. We conclude that human PARP overexpression in rodent cells leads to increased poly(ADP-ribosyl)ation capacity and does not promote survival after gamma-radiation or treatment with the DNA-damaging agents MNNG or DDP.