Nucleic Acids Research Advance Access originally published online on September 17, 2009
Nucleic Acids Research 2009 37(20):6737-6745; doi:10.1093/nar/gkp703
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Nucleic Acids Research, 2009, Vol. 37, No. 20 6737-6745
© The Author(s) 2009. Published by Oxford University Press.
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Genome Integrity, Repair and Replication |
Translesion DNA synthesis-assisted non-homologous end-joining of complex double-strand breaks prevents loss of DNA sequences in mammalian cells
1Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100 Israel, 2Inserm, U768, Hôpital Necker-Enfants Malades, Paris, France and 3Genome Damage and Stability Centre, University of Sussex, East Sussex BN1 9RQ, UK
*To whom correspondence should be addressed. Tel: +972 8 934 3203; Fax: +972 8 934 4169; Email: zvi.livneh{at}weizmann.ac.il
Received March 26, 2009. Revised August 10, 2009. Accepted August 10, 2009.
Double strand breaks (DSB) are severe DNA lesions, and if not properly repaired, may lead to cell death or cancer. While there is considerable data on the repair of simple DSB (sDSB) by non-homologous end-joining (NHEJ), little is known about the repair of complex DSBs (cDSB), namely breaks with a nearby modification, which precludes ligation without prior processing. To study the mechanism of cDSB repair we developed a plasmid-based shuttle assay for the repair of a defined site-specific cDSB in cultured mammalian cells. Using this assay we found that repair efficiency and accuracy of a cDSB with an abasic site in a 5' overhang was reduced compared with a sDSB. Translesion DNA synthesis (TLS) across the abasic site located at the break prevented loss of DNA sequences, but was highly mutagenic also at the template base next to the abasic site. Similar to sDSB repair, cDSB repair was totally dependent on XrccIV, and altered in the absence of Ku80. In contrast, Artemis appears to be specifically involved in cDSB repair. These results may indicate that mammalian cells have a damage control strategy, whereby severe deletions are prevented at the expense of the less deleterious point mutations during NHEJ.
Present address: Shay Covo, Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, National Institutes of Health, Research Triangle Park, NC 27709, USA.
Incumbent of The Maxwell Ellis Professorial Chair in Biomedical Research.