Mechanistic analysis of a DNA end processing pathway mediated by the Xenopus Werner syndrome protein.
The first step of homology-dependent repair of DNA double-strand breaks is the strand-specific processing of DNA ends to generate 3' single-strand tails. Despite its importance, the molecular mechanism underlying end processing is poorly understood in eukaryotic cells. We have taken a biochemical approach to investigate DNA end processing in nucleoplasmic extracts derived from the unfertilized eggs of Xenopus laevis. We found that double-strand DNA ends are specifically degraded in the 5' --> 3' direction in this system. The reaction consists of two steps: an ATP-dependent unwinding of double-strand ends and an ATP-independent 5' --> 3' degradation of single-strand tails. We also found that the Xenopus Werner syndrome protein, a member of the RecQ helicase family, plays an important role in DNA end processing. Mechanistically, Xenopus Werner syndrome protein (xWRN) is required for the unwinding of DNA ends but not for the degradation of single-strand tails. The xWRN-mediated end processing is remarkably similar to the end processing that has been proposed for the Escherichia coli RecQ helicase and RecJ single-strand nuclease, suggesting that this mechanism might be conserved in prokaryotes and eukaryotes.
Toczylowski
Thomas
T
Fox Chase Cancer Center, Philadelphia, Pennsylvania 19111, USA.
Yan
Hong
H
D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000235:genetics*; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D000071657:Werner Syndrome Helicase; D029867:Xenopus Proteins / Q000235:genetics / Q000378:metabolism*; D014982:Xenopus laevis / Q000235:genetics* / Q000378:metabolism*
D029867:Xenopus Proteins; D004247:DNA; D004265:DNA Helicases; D000071657:Werner Syndrome Helicase; C507159:WRN protein, Xenopus