Homologous recombination (HR) requires nuclease activities at multiple steps, but the contribution of individual nucleases to the processing of double-strand DNA ends at different stages of HR has not been clearly defined. We used chicken DT40 cells to investigate the role of flap endonuclease 1 (Fen-1) in HR. FEN-1-deficient cells exhibited a significant decrease in the efficiency of immunoglobulin gene conversion while being proficient in recombination between sister chromatids, suggesting that Fen-1 may play a role in HR between sequences of considerable divergence. To clarify whether sequence divergence at DNA ends is truly the reason for the observed HR defect in FEN-1(-/-) cells we inserted a unique I-SceI restriction site in the genome and tested various donor and recipient HR substrates. We found that the efficiency of HR-mediated DNA repair was indeed greatly diminished when divergent sequences were present at the DNA break site. We conclude that Fen-1 eliminates heterologous sequences at DNA damage site and facilitates DNA repair by HR.

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Keywords *DNA damage, *Recombination, Genetic, DNA repair, DNA, Complementary/metabolism, DNA/chemistry, Deoxyribonucleases, Type II Site-Specific/pharmacology, Flap Endonucleases/metabolism/*physiology, Flow Cytometry, Gamma Rays, Models, Genetic, Molecular Sequence Data, Plasmids/metabolism, Sequence Homology, Nucleic Acid, Sister Chromatid Exchange, animals, base sequence, cell cycle, kinetics, mutation, time factors, transfection
Persistent URL dx.doi.org/10.1128/MCB.25.16.6948-6955.2005, hdl.handle.net/1765/9511
Kikuchi, K., Koyama, H., Jasin, M., van Gent, D.C., Takeda, S., Taniguchi, Y., … Adachi, N.. (2005). Fen-1 facilitates homologous recombination by removing divergent sequences at DNA break ends. Molecular and Cellular Biology, 25(16), 6948–6955. doi:10.1128/MCB.25.16.6948-6955.2005