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Nucleic Acids Research, 2000, Vol. 28, No. 19 3771-3778
© 2000 Oxford University Press

Role of the nucleotide excision repair gene ERCC1 in formation of recombination-dependent rearrangements in mammalian cells

R. Geoffrey Sargent, James L. Meservy, Brian D. Perkins, April E. Kilburn, Zsofia Intody, Gerald M. Adair1, Rodney S. Nairn1 and John H. Wilson*

The Verna and Marrs McLean Department of Biochemistry and Molecular Biology, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX 77030, USA and 1The University of Texas M.D. Anderson Cancer Center Science Park Research Division, PO Box 389, Smithville, TX 78957, USA

Spontaneous recombination between direct repeats at the adenine phosphoribosyltransferase (APRT) locus in ERCC1-deficient cells generates a high frequency of rearrangements that are dependent on the process of homologous recombination, suggesting that rearrangements are formed by misprocessing of recombination intermediates. Given the specificity of the structure-specific Ercc1/Xpf endonuclease, two potential recombination intermediates are substrates for misprocessing in ERCC1– cells: heteroduplex loops and heteroduplex intermediates with non-homologous 3' tails. To investigate the roles of each, we constructed repeats that would yield no heteroduplex loops during spontaneous recombination or that would yield two non-homologous 3' tails after treatment with the rare-cutting endonuclease I-SceI. Our results indicate that misprocessing of heteroduplex loops is not the major source of recombination-dependent rearrangements in ERCC1-deficient cells. Our results also suggest that the Ercc1/Xpf endonuclease is required for efficient removal of non-homologous 3' tails, like its Rad1/Rad10 counterpart in yeast. Thus, it is likely that misprocessing of non-homologous 3' tails is the primary source of recombination-dependent rearrangements in mammalian cells. We also find an unexpected effect of ERCC1 deficiency on I-SceI-stimulated rearrangements, which are not dependent on homologous recombination, suggesting that the ERCC1 gene product may play a role in generating the rearrangements that arise after I-SceI-induced double-strand breaks.

* To whom correspondence should be addressed. Tel: +1 713 798 5760; Fax: +1 713 796 9438; Email: jwilson@bcm.tmc.edu Present addresses: R. Geoffrey Sargent, Pangene Corporation, Mountain View, CA 94043, USA Brian D. Perkins, Department of Molecular and Cellular Biology, Harvard University, Cambridge, MA 02138, USA April E. Kilburn, Office of Technology and Licensing, University of Texas, Austin, TX 78759, USA Zsofia Intody, Department of Opthalmology No. 1, Semmelweis University of Medicine, Budapest, Hungary


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