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Molecular and Cellular Biology, November 2004, p. 9682-9694, Vol. 24, No. 21
0270-7306/04/$08.00+0     DOI: 10.1128/MCB.24.21.9682-9694.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

The Mre11 Nuclease Is Not Required for 5' to 3' Resection at Multiple HO-Induced Double-Strand Breaks

Bertrand Llorente and Lorraine S. Symington*

Department of Microbiology and Institute of Cancer Research, Columbia University Medical Center, New York, New York

Received 29 March 2004/ Returned for modification 28 April 2004/ Accepted 3 August 2004

Current hypotheses suggest the Mre11 nuclease activity could be directly involved in double-strand break (DSB) resection in the presence of a large number of DSBs or limited to processing abnormal DNA ends. To distinguish between these possibilities, we used two methods to create large numbers of DSBs in Saccharomyces cerevisiae chromosomes, without introducing other substrates for the Mre11 nuclease. Multiple DSBs were created either by expressing the HO endonuclease in strains containing several HO cut sites embedded within randomly dispersed Ty1 elements or by phleomycin treatment. Analysis of resection by single-strand DNA formation in these systems showed no difference between strains containing MRE11 or the mre11-D56N nuclease defective allele, suggesting that the Mre11 nuclease is not involved in the extensive 5' to 3' resection of DSBs. We postulate that the ionizing radiation (IR) sensitivity of mre11 nuclease-defective mutants results from the accumulation of IR-induced DNA damage that is normally processed by the Mre11 nuclease. We also report that the processivity of 5' to 3' DSB resection and the yield of repaired products are affected by the number of DSBs in a dose-dependent manner. Finally, we show that the exonuclease Exo1 is involved in the processivity of 5' to 3' resection of an HO-induced DSB at the MAT locus.

* Corresponding author. Mailing address: Department of Microbiology, Columbia University Medical Center, 701 W. 168th St., New York, NY 10032. Phone: (212) 305-4794. Fax: (212) 305-1741. E-mail: lss5{at}columbia.edu.

Molecular and Cellular Biology, November 2004, p. 9682-9694, Vol. 24, No. 21
0022-538X/04/$08.00+0     DOI: 10.1128/MCB.24.21.9682-9694.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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