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Nucleic Acids Res. 2014 Jun;42(10):6337-51. doi: 10.1093/nar/gku288. Epub 2014 Apr 25.

Suicidal cross-linking of PARP-1 to AP site intermediates in cells undergoing base excision repair.

Author information

1
Laboratory of Structural Biology, NIEHS, National Institutes of Health, 111 T.W. Alexander Drive, Research Triangle Park, NC 27709, USA.
2
William Carey University College of Osteopathic Medicine, 498 Tuscan Avenue, Hattiesburg, MS 39401, USA.
3
Laboratory of Structural Biology, NIEHS, National Institutes of Health, 111 T.W. Alexander Drive, Research Triangle Park, NC 27709, USA wilson5@niehs.nih.gov.

Abstract

Poly(ADP-ribose) polymerase-1 (PARP-1) is an abundant nuclear enzyme in mammalian cells. The enzyme synthesizes polymers of ADP-ribose from the coenzyme NAD(+) and plays multifaceted roles in cellular responses to genotoxic stress, including DNA repair. It had been shown that mouse fibroblasts treated with a DNA methylating agent in combination with a PARP inhibitor exhibit higher cytotoxicity than cells treated with methylating agent alone. This lethality of the PARP inhibitor is dependent on apurinic/apyrimidinic (AP) sites in the DNA and the presence of PARP-1. Here, we show that purified PARP-1 is capable of forming a DNA-protein cross-link (DPC) by covalently attaching to the AP site. This DPC formation is specific to the presence of the natural AP site in DNA and is accompanied by a single-strand DNA incision. Cellular studies confirm the formation of PARP-1 DPCs during alkylating agent-induced base excision repair (BER) and formation of DPCs is enhanced by a PARP inhibitor. Using an N-terminal and C-terminal truncated PARP-1 we show that a polypeptide fragment comprising the zinc 3 and BRCT sub-domains is sufficient for DPC formation. The covalent attachment of PARP-1 to AP site-containing DNA appears to be a suicidal event when BER is overwhelmed or disrupted.

PMID:
24771347
PMCID:
PMC4041460
DOI:
10.1093/nar/gku288
[Indexed for MEDLINE]
Free PMC Article

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