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XPF-ERCC1 acts in Unhooking DNA interstrand crosslinks in cooperation with FANCD2 and FANCP/SLX4

Daisy Klein Douwel1, Rick A C M Boonen1, David T Long2

  • 1Hubrecht Institute-KNAW, University Medical Center Utrecht and Cancer Genomics Netherlands, Uppsalalaan 8, 3584 CT Utrecht, the Netherlands.

Molecular Cell
|April 15, 2014
PubMed

Insights

DNA interstrand crosslinks (ICLs) are toxic DNA lesions repaired by the Fanconi anemia pathway. This study reveals XPF-ERCC1 and SLX4 cooperate with FANCD2 ubiquitylation for ICL unhooking, a crucial repair step.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • DNA interstrand crosslinks (ICLs) are highly toxic lesions that covalently link DNA strands.
  • ICLs are repaired by a complex, replication-coupled pathway in eukaryotes, involving the Fanconi anemia (FA) pathway.
  • Activation of the FA pathway critically depends on the ubiquitylation of the FANCD2 protein.

Purpose of the Study:

  • To elucidate the molecular mechanism of DNA interstrand crosslink repair.
  • To identify the proteins involved in the critical DNA unhooking step of ICL repair.
  • To understand the role of FANCD2 ubiquitylation in recruiting repair factors.

Main Methods:

  • Utilized Xenopus egg extracts, a system supporting replication-coupled ICL repair.
  • Investigated the roles of XPF-ERCC1, SLX4/FANCP, and FANCD2 in ICL repair.
  • Analyzed the recruitment of repair factors to ICLs in response to FANCD2 ubiquitylation.

Main Results:

  • Demonstrated that the 3' flap endonuclease XPF-ERCC1 cooperates with SLX4/FANCP to perform unhooking incisions at ICLs.
  • Showed that efficient recruitment of XPF-ERCC1 and SLX4 to ICLs is dependent on FANCD2 and its ubiquitylation.
  • Established a link between FANCD2 ubiquitylation and the initiation of DNA incisions in ICL repair.

Conclusions:

  • The Fanconi anemia pathway utilizes XPF-ERCC1 and SLX4/FANCP to execute unhooking incisions, a key step in ICL repair.
  • FANCD2 ubiquitylation is essential for recruiting these nucleases to the ICL site.
  • These findings clarify the molecular mechanism by which the FA pathway facilitates replication-coupled ICL repair.

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