Cells deficient in PARP-1 show an accelerated accumulation of DNA single strand breaks, but not AP sites, over the PARP-1-proficient cells exposed to MMS.

Pachkowski, B; Tano, K; Afonin, V; Elder, RH; Takeda, S; Watanabe, M; Swenberg, J; Nakamura, J

Abstract

Poly(ADP-ribose) polymerase-1 (PARP-1) is a base excision repair (BER) protein that binds to DNA single strand breaks (SSBs) and subsequently synthesizes and transfers poly(ADP-ribose) polymers to various nuclear proteins. Numerous biochemical studies have implicated PARP-1 as a modulator of BER; however, the role of PARP-1 in BER in living cells remains unclear partly due to lack of accurate quantitation of BER intermediates existing in cells. Since DT40 cells, chicken B lymphocytes, naturally lack PARP-2, DT40 cells allow for the investigation of the PARP-1 null phenotype without confounding by PARP-2. To test the hypothesis that PARP-1 is necessary for efficient BER during methylmethane sulfonate (MMS) exposure in vertebrate cells, intact DT40 cells and their isogenic PARP-1 null counterparts were challenged with different exposure scenarios for phenotypic characterization. With chronic exposure, PARP-1 null cells exhibited sensitivity to MMS but with an acute exposure did not accumulate base lesions or AP sites to a greater extent than wild-type cells. However, an increase in SSB content in PARP-1 null cell DNA, as indicated by glyoxal gel electrophoresis under neutral conditions, suggested the presence of BER intermediates. These data suggest that during exposure, PARP-1 impacts the stage of BER after excision of the deoxyribosephosphate moiety from the 5' end of DNA strand breaks by polymerase beta.

• Journal Article Type: Article
• Publication Date: Dec 1, 2009
• Deposit Date: Oct 6, 2011
• Journal: Mutation research -Fundamental and Molecular Mechanisms of Mutagenesis
• Print ISSN: 0027-5107
• Publisher: Elsevier
• Peer Reviewed: Peer Reviewed
• Volume: 671
• Issue: 1-2
• Pages: 93-9
• DOI: https://doi.org/10.1016/j.mrfmmm.2009.09.006
• Keywords: Alkylating agent, N7-methylguanine, AP sites, base excision repair, single strand breaks, PARP-1
• Publisher URL: http://dx.doi.org/ 10.1016/j.mrfmmm.2009.09.006