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Molecular and Cellular Biology, October 2006, p. 7645-7657, Vol. 26, No. 20
0270-7306/06/$08.00+0     doi:10.1128/MCB.00672-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

Conservative Repair of a Chromosomal Double-Strand Break by Single-Strand DNA through Two Steps of Annealing{triangledown} ,{dagger}

Francesca Storici,{ddagger} Joyce R. Snipe, Godwin K. Chan, Dmitry A. Gordenin,{ddagger} and Michael A. Resnick{ddagger}*

Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, NIH, Research Triangle Park, North Carolina 27709

Received 18 April 2006/ Returned for modification 18 May 2006/ Accepted 2 August 2006

The repair of chromosomal double-strand breaks (DSBs) is essential to normal cell growth, and homologous recombination is a universal process for DSB repair. We explored DSB repair mechanisms in the yeast Saccharomyces cerevisiae using single-strand oligonucleotides with homology to both sides of a DSB. Oligonucleotide-directed repair occurred exclusively via Rad52- and Rad59-mediated single-strand annealing (SSA). Even the SSA domain of human Rad52 provided partial complementation for a null rad52 mutation. The repair did not involve Rad51-driven strand invasion, and moreover the suppression of strand invasion increased repair with oligonucleotides. A DSB was shown to activate targeting by oligonucleotides homologous to only one side of the break at large distances (at least 20 kb) from the break in a strand-biased manner, suggesting extensive 5' to 3' resection, followed by the restoration of resected DNA to the double-strand state. We conclude that long resected chromosomal DSB ends are repaired by a single-strand DNA oligonucleotide through two rounds of annealing. The repair by single-strand DNA can be conservative and may allow for accurate restoration of chromosomal DNAs with closely spaced DSBs.

* Corresponding author. Mailing address: Head Chromosome Stability Section, Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, NIH, Research Triangle Park, NC 27709. Phone: (919) 541-4480. Fax: (919) 541-7593. E-mail: Resnick{at}niehs.nih.gov.

{triangledown} Published ahead of print on 14 August 2006.

{dagger} Supplemental material for this article may be found at http://mcb.asm.org/.

{ddagger} D.A.G. and M.A.R. contributed equally to this study.

Molecular and Cellular Biology, October 2006, p. 7645-7657, Vol. 26, No. 20
0270-7306/06/$08.00+0     doi:10.1128/MCB.00672-06
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



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