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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

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

Published by Oxford University Press on behalf of Nucleic Acids Research 2014. This work is written by (a) US Government employee(s) and is in the public domain in the US.

PMID:
24771347
[PubMed - indexed for MEDLINE]
PMCID:
PMC4041460
Free PMC Article
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