Fen-1 facilitates homologous recombination by removing divergent sequences at DNA break ends
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.
|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