Complete inhibition of poly(ADP-ribose) polymerase activity prevents the recovery of C3H1OT1/2 cells from oxidative stress

Shah, Girish M.; Poirier, D.; Desnoyers, Serge; Saint-Martin, Sylvie; Hoflack, Jean-Christophe; Rong, Pei-Min; ApSimon, Michéle M.; Kirkland, James B.; Poirier, Guy G.

doi:10.1016/0167-4889(96)00004-3

Cited by 31 publications

(15 citation statements)

References 32 publications

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“…Cells treated with inhibitors of PARP have been reported to become resistant to various genotoxic agents, such as UVB-and ␥-irradiation, tumor necrosis factor (TNF), alkylating agents, and free radicals (Malorni et al 1995;McGowan et al 1996). In contrast, other studies showed that inhibition or a dominantnegative mutation of PARP sensitizes cells to cell death induced by oxidative stress, alkylating agents, ␥-irradiation, or heat shock (Nosseri et al 1994;Kü pper et al 1995;Schreiber et al 1995;Shah et al 1996). Although results obtained from these studies are contradictory and the mechanisms involved in such processes are complex, the function of PARP is well documented in free radicalinduced cell toxicity in neurons and pancreatic islet cells (Zhang et al 1994;Heller et al 1995).…”

Section: Nadmentioning

confidence: 92%

PARP is important for genomic stability but dispensable in apoptosis

Wang¹,

Stingl²,

Morrison³

et al. 1997

Genes Dev.

535

361

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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 ␣, ␥-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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Section: Nadmentioning

confidence: 92%

PARP is important for genomic stability but dispensable in apoptosis

Wang¹,

Stingl²,

Morrison³

et al. 1997

Genes Dev.

535

361

View full text Add to dashboard Cite

show abstract

“…However, NAD levels continued to drop after the polymer peak, even though more than 50% of PARP was cleaved. A concentration of 1 mM of the PARP inhibitor DHQ, known to completely inhibit PARP in vivo (26), only partially prevented the loss of NAD (Fig. 2B).…”

Section: Parp Cleavage By Caspases At the Time Of Its Activation-mentioning

confidence: 99%

“…The automodification reaction was stopped by the addition of 100 M DHQ (26). Upon addition of 0.1% CHAPS and the indicated amount of caspase-3 or -7, the substrate was digested for specific times at 37°C.…”

Section: Materials-[mentioning

confidence: 99%

Cleavage of Automodified Poly(ADP-ribose) Polymerase during Apoptosis

Germain¹,

Affar²,

D’Amours³

et al. 1999

Journal of Biological Chemistry

Self Cite

410

269

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The abundant nuclear enzyme poly(ADP-ribose) polymerase (PARP) synthesizes poly(ADP-ribose) in response to DNA strand breaks. During almost all forms of apoptosis, PARP is cleaved by caspases, suggesting the crucial role of its inactivation. A few studies have also reported a stimulation of PARP during apoptosis. However, the role of PARP stimulation and cleavage during this cell death process remains poorly understood. Here, we measured the stimulation of endogenous poly(ADPribose) synthesis during VP-16-induced apoptosis in HL60 cells and found that PARP was cleaved by caspases at the time of its poly(ADP-ribosyl)ation. In vitro experiments showed that PARP cleavage by caspase-7, but not by caspase-3, was stimulated by its automodification by long and branched poly(ADP-ribose). Consistently, caspase-7 exhibited an affinity for poly(ADP-ribose), whereas caspase-3 did not. In addition, caspase-7 was activated and accumulated in the nucleus of HL60 cells in response to the VP-16 treatment. Furthermore, caspase-7 activation was concommitant with PARP cleavage in the caspase-3-deficient cell line MCF-7 in response to staurosporine treatment. These results strongly suggest that, in vivo, it is caspase-7 that is responsible for PARP cleavage and that poly(ADP-ribosyl)ation of PARP accelerates its proteolysis. Cleavage of the active form of caspase substrates could be a general feature of the apoptotic process, ensuring the rapid inactivation of stress signaling proteins.

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“…NAD ϩ was added to reactions as indicated, and arrows indicate the position of phosphorylated RPA p34 and phosphorylated p53 proteins. dihydroxyisoquinoline (DHQ) (37). In the presence of 1 mM DHQ the automodification activity of PARP in the presense of 200 M NAD ϩ is nearly completely inhibited (Fig.…”

Section: A Lane D)mentioning

confidence: 99%

Stimulation of the DNA-dependent Protein Kinase by Poly(ADP-Ribose) Polymerase

Ruscetti¹,

Lehnert²,

Halbrook³

et al. 1998

Journal of Biological Chemistry

216

145

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The DNA-dependent protein kinase (DNA-PK) is a heterotrimeric enzyme that binds to double-stranded DNA and is required for the rejoining of double-stranded DNA breaks in mammalian cells. It has been proposed that DNA-PK functions in this DNA repair pathway by binding to the ends of broken DNA molecules and phosphorylating proteins that bind to the damaged DNA ends. Another enzyme that binds to DNA strand breaks and may also function in the cellular response to DNA damage is the poly(ADP-ribose) polymerase (PARP). Here, we show that PARP can be phosphorylated by purified DNA-PK, and the catalytic subunit of DNA-PK is ADP-ribosylated by PARP. The protein kinase activity of DNA-PK can be stimulated by PARP in the presence of NAD ؉ in a reaction that is blocked by the PARP inhibitor 1,5-dihydroxyisoquinoline. The stimulation of DNA-PK by PARP-mediated protein ADP-ribosylation occurs independent of the Ku70/80 complex. Taken together, these results show that PARP can modify the activity of DNA-PK in vitro and suggest that these enzymes may function coordinately in vivo in response to DNA damage.Biochemical pathways that function in the recognition and repair of DNA damage are critical for maintaining genomic integrity. Potentially devastating DNA damage in the form of double-strand breaks (DSBs) 1 can occur when cells are exposed to ionizing radiation, oxidative stress and radiomimetic drugs. The DNA-dependent protein kinase (DNA-PK) is a key component of DNA DSB rejoining pathways in mammalian cells. DNA-PK is a heterotrimeric enzyme complex comprised of a 460-kDa catalytic subunit (1) and a regulatory component consisting of the Ku70 and Ku80 proteins (2, 3). The Ku70 and Ku80 proteins form a heterodimeric complex that binds to the ends of double-stranded DNA with high affinity (4 -8). The catalytic subunit of DNA-PK (DNA-PKcs) binds to the Ku70/80 complex in the presence of double-stranded DNA (9) and phosphorylates a wide variety of protein substrates in vitro on serine and threonine residues (10, 11). The protein kinase activity of DNA-PK is autoregulatory; in the absence of a phosphorylation substrate, DNA-PKcs autophosphorylates and dissociates from the Ku70/80-DNA complex (12).Evidence that DNA-PK plays an integral role in the repair of DNA DSB has been provided through the characterization of rodent cell lines that have mutations that disrupt the expression of the Ku80 (13-19) or DNA-PKcs (20 -24). Although it is clear that DNA-PK is an important component of mammalian DNA DSB repair pathways, it is not known how the enzyme participates in these processes. In vitro, DNA-PK preferentially phosphorylates protein substrates that co-localize on the same DNA molecule (3,25). This suggests that the specificity of the phosphorylation reaction may be regulated, in part, via the co-localization of the enzyme and substrate target on DNA. Based on these data, it has been proposed that DNA-PK could participate in the DNA rejoining reaction by phosphorylating DNA repair factors that co-localize with it on broken DNA en...

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Complete inhibition of poly(ADP-ribose) polymerase activity prevents the recovery of C3H1OT1/2 cells from oxidative stress

Cited by 31 publications

References 32 publications

PARP is important for genomic stability but dispensable in apoptosis

PARP is important for genomic stability but dispensable in apoptosis

Cleavage of Automodified Poly(ADP-ribose) Polymerase during Apoptosis

Stimulation of the DNA-dependent Protein Kinase by Poly(ADP-Ribose) Polymerase

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