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Molecular and Cellular Biology, June 2000, p. 4028-4035, Vol. 20, No. 11
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Homologous and Nonhomologous Recombination Resulting in Deletion: Effects of p53 Status, Microhomology, and Repetitive DNA Length and Orientation

Dan Gebow,dagger Nathan Miselis, and Howard L. Liber*

Department of Radiation Oncology, Massachusetts General Hospital, Boston, Massachusetts 02114

Received 29 June 1999/Returned for modification 10 August 1999/Accepted 24 January 2000

Repetitive DNA elements frequently are precursors to chromosomal deletions in prokaryotes and lower eukaryotes. However, little is known about the relationship between repeated sequences and deletion formation in mammalian cells. We have created a novel integrated plasmid-based recombination assay to investigate repeated sequence instability in human cells. In a control cell line, the presence of direct or inverted repeats did not appreciably influence the very low deletion frequencies (2 × 10-7 to 9 × 10-7) in the region containing the repeat. Similar to what has been observed in lower eukaryotes, the majority of deletions resulted from the loss of the largest direct repeat present in the system along with the intervening sequence. Interestingly, in closely related cell lines that possess a mutant p53 gene, deletion frequencies in the control and direct-repeat plasmids were 40 to 300 times higher than in their wild-type counterparts. However, mutant p53 cells did not preferentially utilize the largest available homology in the formation of the deletion. Surprisingly, inverted repeats were approximately 10,000 times more unstable in all mutant p53 cells than in wild-type cells. Finally, several deletion junctions were marked by the addition of novel bases that were homologous to one of the preexisting DNA ends. Contrary to our expectations, only 6% of deletions in all cell lines could be classified as arising from nonhomologous recombination.

* Corresponding author. Mailing address: Department of Radiation Oncology, Massachusetts General Hospital, 100 Blossom St., Cox 302, Boston, MA 02114. Phone: (617) 726-4143. Fax: (617) 724-8320. E-mail: hliber{at}partners.org.

dagger Present address: Department of Molecular, Cellular and Developmental Biology, University of California, Santa Barbara, CA 93015.

Molecular and Cellular Biology, June 2000, p. 4028-4035, Vol. 20, No. 11
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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