A role for XLF in DNA repair and recombination in human somatic cells

Farjana Jahan Fattah, Junghun Kweon, Yongbao Wang, Eu Han Lee, Yinan Kan, Natalie Lichter, Natalie Weisensel, Eric A. Hendrickson

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Classic non-homologous end-joining (C-NHEJ) is required for the repair of radiation-induced DNA double-strand breaks (DSBs) in mammalian cells and plays a critical role in lymphoid V(D)J recombination. A core C-NHEJ component is the DNA ligase IV co-factor, Cernunnos/XLF (hereafter XLF). In patients, mutations in XLF cause predicted increases in radiosensitivity and deficits in immune function, but also cause other less well-understood pathologies including neural disorders. To characterize XLF function(s) in a defined genetic system, we used a recombinant adeno-associated virus-mediated gene targeting strategy to inactivate both copies of the XLF locus in the human HCT116 cell line. Analyses of XLF-null cells (which were viable) showed that they were highly sensitive to ionizing radiation and a radiomimetic DNA damaging agent, etoposide. XLF-null cells had profound DNA DSB repair defects as measured by in vivo plasmid end-joining assays and were also dramatically impaired in their ability to form either V(D)J coding or signal joints on extrachromosomal substrates. Thus, our somatic XLF-null cell line recapitulates many of the phenotypes expected from XLF patient cell lines. Subsequent structure:function experiments utilizing the expression of wild-type and mutant XLF cDNAs demonstrated that all of the phenotypes of an XLF deficiency could be rescued by the overexpression of a wild-type XLF cDNA. Unexpectedly, mutant forms of XLF bearing point mutations at amino acid positions L115 and L179, also completely complemented the null phenotype suggesting, in contrast to predictions to the contrary, that these mutations do not abrogate XLF function. Finally, we demonstrate that the absence of XLF causes a small, but significant, increase in homologous recombination, implicating XLF in DSB pathway choice regulation. We conclude that human XLF is a non-essential, but critical, C-NHEJ-repair factor.

Original languageEnglish (US)
Pages (from-to)39-53
Number of pages15
JournalDNA Repair
Volume15
Issue number1
DOIs
StatePublished - Mar 1 2014

Fingerprint

Null Lymphocytes
Joining
DNA Repair
Genetic Recombination
Repair
Double-Stranded DNA Breaks
Cells
Phenotype
Cell Line
DNA
Complementary DNA
V(D)J Recombination
HCT116 Cells
Dependovirus
Mutation
Bearings (structural)
Gene Targeting
Radiation Tolerance
Homologous Recombination
Etoposide

Cite this

A role for XLF in DNA repair and recombination in human somatic cells. / Fattah, Farjana Jahan; Kweon, Junghun; Wang, Yongbao; Lee, Eu Han; Kan, Yinan; Lichter, Natalie; Weisensel, Natalie; Hendrickson, Eric A.

In: DNA Repair, Vol. 15, No. 1, 01.03.2014, p. 39-53.

Research output: Contribution to journalArticle

Fattah, FJ, Kweon, J, Wang, Y, Lee, EH, Kan, Y, Lichter, N, Weisensel, N & Hendrickson, EA 2014, 'A role for XLF in DNA repair and recombination in human somatic cells', DNA Repair, vol. 15, no. 1, pp. 39-53. https://doi.org/10.1016/j.dnarep.2013.12.006
Fattah FJ, Kweon J, Wang Y, Lee EH, Kan Y, Lichter N et al. A role for XLF in DNA repair and recombination in human somatic cells. DNA Repair. 2014 Mar 1;15(1):39-53. https://doi.org/10.1016/j.dnarep.2013.12.006
Fattah, Farjana Jahan ; Kweon, Junghun ; Wang, Yongbao ; Lee, Eu Han ; Kan, Yinan ; Lichter, Natalie ; Weisensel, Natalie ; Hendrickson, Eric A. / A role for XLF in DNA repair and recombination in human somatic cells. In: DNA Repair. 2014 ; Vol. 15, No. 1. pp. 39-53.
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AU - Weisensel, Natalie

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