title: Preventing over-resection by DNA2 helicase/nuclease suppresses repair defects in Fanconi anemia cells.
abstract: FANCD2 is required for the repair of DNA damage by the FA (Fanconi anemia) pathway, and, consequently, FANCD2-deficient cells are sensitive to compounds such as cisplatin and formaldehyde that induce DNA:DNA and DNA:protein crosslinks, respectively. The DNA2 helicase/nuclease is required for RNA/DNA flap removal from Okazaki fragments during DNA replication and for the resection of DSBs (double-strand breaks) during HDR (homology-directed repair) of replication stress-induced damage. A knockdown of DNA2 renders normal cells as sensitive to cisplatin (in the absence of EXO1) and to formaldehyde (even in the presence of EXO1) as FANCD2(-/-) cells. Surprisingly, however, the depletion of DNA2 in FANCD2-deficient cells rescues the sensitivity of FANCD2(-/-) cells to cisplatin and formaldehyde. We previously showed that the resection activity of DNA2 acts downstream of FANCD2 to insure HDR of the DSBs arising when replication forks encounter ICL (interstrand crosslink) damage. The suppression of FANCD2(-/-) by DNA2 knockdowns suggests that DNA2 and FANCD2 also have antagonistic roles: in the absence of FANCD2, DNA2 somehow corrupts repair. To demonstrate that DNA2 is deleterious to crosslink repair, we used psoralen-induced ICL damage to trigger the repair of a site-specific crosslink in a GFP reporter and observed that "over-resection" can account for reduced repair. Our work demonstrates that excessive resection can lead to genome instability and shows that strict regulatory processes have evolved to inhibit resection nucleases. The suppression of FANCD2(-/-) phenotypes by DNA2 depletion may have implications for FA therapies and for the use of ICL-inducing agents in chemotherapy.
authors.affiliation: Braun Laboratories; California Institute of Technology; Pasadena, CA USA.
authors.forename: Kenneth K
authors.initials: KK
authors.lastname: Karanja
authors.affiliation: Department of Biochemistry, Molecular Biology, and Biophysics; University of Minnesota; Minneapolis, MN USA.
authors.forename: Eu Han
authors.initials: EH
authors.lastname: Lee
authors.affiliation: Department of Biochemistry, Molecular Biology, and Biophysics; University of Minnesota; Minneapolis, MN USA.
authors.forename: Eric A
authors.initials: EA
authors.lastname: Hendrickson
authors.affiliation: Braun Laboratories; California Institute of Technology; Pasadena, CA USA.
authors.forename: Judith L
authors.initials: JL
authors.lastname: Campbell
mesh_terms: D045744:Cell Line, Tumor; D002945:Cisplatin / Q000494:pharmacology; D053903:DNA Breaks, Double-Stranded; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism*; D004260:DNA Repair / Q000187:drug effects / Q000235:genetics*; D005199:Fanconi Anemia / Q000235:genetics* / Q000473:pathology; D052236:Fanconi Anemia Complementation Group D2 Protein / Q000235:genetics* / Q000378:metabolism; D005363:Ficusin / Q000737:chemistry / Q000494:pharmacology; D005557:Formaldehyde / Q000494:pharmacology; D042822:Genomic Instability; D006801:Humans
chemical_list: D052236:Fanconi Anemia Complementation Group D2 Protein; D005557:Formaldehyde; D004265:DNA Helicases; C509908:DNA2 protein, human; D005363:Ficusin; D002945:Cisplatin
keywords: DNA recombination; DNA replication; DNA2; FANCD2; Fanconi anemia; formaldehyde DNA damage; interstrand crosslinks