The embryonic lethality in DNA ligase IV-deficient mice is rescued by deletion of Ku: implications for unifying the heterogeneous phenotypes of NHEJ mutants

Karanjawala, Zarir E.; Adachi, Noritaka; Irvine, Ryan A.; Oh, Eui K; Shibata, Darryl; Schwarz, Klaus; Hsieh, Chih‐Lin; Lieber, Michael R.

doi:10.1016/s1568-7864(02)00151-9

Cited by 89 publications

(76 citation statements)

References 43 publications

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“…According to this hypothesis, KU-alt and XRCC4-alt would correspond to the same pathway, despite the apparent difference in efficiency. This hypothesis is consistent with the knockout of KU restoring the viability of Lig4-null mice (38).…”

Section: Discussionsupporting

confidence: 85%

Defects in XRCC4 and KU80 differentially affect the joining of distal nonhomologous ends

Guirouilh‐Barbat

Rass

Plo

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

152

183

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XRCC4-null mice have a more severe phenotype than KU80-null mice. Here, we address whether this difference in phenotype is connected to nonhomologous end-joining (NHEJ). We used intrachromosomal substrates to monitor NHEJ of two distal doublestrand breaks (DSBs) targeted by I-SceI, in living cells. In xrcc4-defective XR-1 cells, a residual but significant end-joining process exists, which primarily uses microhomologies distal from the DSB. However, NHEJ efficiency was strongly reduced in xrcc4-defective XR-1 cells versus complemented cells, contrasting with KU-deficient xrs6 cells, which showed levels of end-joining similar to those of complemented cells. Nevertheless, sequence analysis of the repair junctions indicated that the accuracy of end-joining was strongly affected in both xrcc4-deficient and KU-deficient cells. More specifically, these data showed that the KU80/XRCC4 pathway is conservative and not intrinsically error-prone but can accommodate non-fully complementary ends at the cost of limited mutagenesis.double-strand break repair ͉ genome rearrangements D NA double-strand breaks (DSBs) are harmful lesions generated by a variety of endogenous or exogenous stresses, potentially leading to genomic rearrangement. Nonhomologous endjoining (NHEJ) is a prominent pathway for DSB repair (1). Canonical NHEJ involves the successive intervention of the KU80-KU70 heterodimer, DNA-PKcs-Artemis, and, finally, ligase IV (Lig4) associated with its cofactors XRCC4 and Cernunnos/Xlf (2, 3). KU-independent NHEJ (KU-alt) has been described in vitro in both acellular extracts and cultured cells (1, 4-6).Alternative, XRCC4-independent DSB repair pathways (XRCC4-alt) have also been described, using episomic plasmid in cultured cells, using pulse field gel electrophoresis, or in in vitro biochemical experiments (5-10). One hypothesis could propose that XRCC4 and KU are implicated in the same canonical NHEJ pathway, whereas the alternate pathway is independent of both KU and XRCC4. However, in transgenic mice, the inactivation of XRCC4 or Lig4 results in a more severe phenotype than the inactivation of KU (11), suggesting that XRCC4 might have an additional essential function; however, studies show that XRCC4 and Lig4 do not have roles outside of NHEJ, whereas in contrast, KU acts in other processes such as transcription, apoptosis, and responses to the cell microenvironment (12)(13)(14).Alternatively, these varying phenotypes in mice may actually result from differences in DSB repair efficiencies, indicating that defects in XRCC4 might be more deleterious for DSB repair than defects in KU. What challenges this hypothesis, however, is that substantial class switch recombination (CSR) has recently been shown to occur in mouse B cells without XRCC4, whereas no CSR was recorded in cells devoid of KU (15-18).The relative contributions of XRCC4 and KU80 versus the XRCC4-alt and KU-alt pathways, respectively, to DSB repair remain unclear in wild-type cells. Contrasting results were obtained in living cells, using an episomic plasm...

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Section: Discussionsupporting

confidence: 85%

Defects in XRCC4 and KU80 differentially affect the joining of distal nonhomologous ends

Guirouilh‐Barbat

Rass

Plo

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

152

183

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“…Interestingly, a Ku70 knockout rescues this synthetic lethality (105). Similar to a study showing that a Ku80 deletion recues the lethality of a Lig4 knockout (106), this demonstrates that several NHEJ factors are epistatic to Ku. Loss of both XLF and DNA-PKcs must severely impair the ability to repair a DSB by NHEJ.…”

Section: Xlf and Paxx Stimulate Ligation By The Xrcc4·lig4 Complexsupporting

confidence: 80%

Nonhomologous DNA end-joining for repair of DNA double-strand breaks

Pannunzio

Watanabe

Lieber

2018

Journal of Biological Chemistry

390

360

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Nonhomologous DNA end joining (NHEJ) is the predominant DSB repair pathway throughout the cell cycle and accounts for nearly all DSB repair outside of the S and G2 phases. NHEJ relies on Ku to thread onto DNA termini and thereby improve the affinity of the NHEJ enzymatic components consisting of polymerases (Pol m and Pol l), a nuclease (the Artemis·DNA-PKcs complex), and a ligase (XLF·XRCC4·Lig4 complex). Each of the enzymatic components is distinctive for its versatility in acting on diverse incompatible DNA end configurations coupled with a flexibility in loading order, resulting in many possible junctional outcomes from one DSB. DNA ends can either be directly ligated or, if the ends are incompatible, processed until a ligatable configuration is achieved that is often stabilized by up to 4 bp of terminal microhomology. Processing of DNA ends results in nucleotide loss or addition, explaining why DSBs repaired by NHEJ are rarely restored to their original DNA sequence. Thus, NHEJ is a single pathway with multiple enzymes at its disposal to repair DSBs, resulting in a diversity of repair outcomes.

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“…Deficiency for DNA ligase IV in a chicken cell line resulted in a high level of IR-induced lethality that was partially rescued by mutation of KU70, possibly because the DSB-binding Ku heterodimer may prevent the HR machinery from processing unrepaired DSBs in the absence of DNA ligase IV (Adachi et al 2001). Further, deficiency for DNA ligase IV in mice results in embryonic lethality, which is rescued by mutation of Ku86 (Karanjawala et al 2002). However, the Gro phenotype of late-stage lig-4;cku-80 double-mutant embryos irradiated with 50 Gy was not significantly different from that of either single mutant (lig-4;cku-80 ¼ 44 6 5% L4; cku-80 ¼ 36 6 18% L4; lig-4 ¼ 32 6 7% L4; wild type ¼ 100% L4; n ¼ 100 each), suggesting that the two genes may act in the same pathway of DSB repair.…”

Section: Resultsmentioning

confidence: 99%

Developmental Modulation of Nonhomologous End Joining in Caenorhabditis elegans

2006

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Homologous recombination and nonhomologous end joining (NHEJ) are important DNA doublestrand break repair pathways in many organisms. C. elegans strains harboring mutations in the cku-70, cku-80, or lig-4 NHEJ genes displayed multiple developmental abnormalities in response to radiationinduced DNA damage in noncycling somatic cells. These phenotypes did not result from S-phase, DNA damage, or mitotic checkpoints, apoptosis, or stress response pathways that regulate dauer formation. However, an additional defect in him-10, a kinetochore component, synergized with NHEJ mutations for the radiation-induced developmental phenotypes, suggesting that they may be triggered by missegregation of chromosome fragments. Although NHEJ was an important DNA repair pathway for noncycling somatic cells in C. elegans, homologous recombination was used to repair radiation-induced DNA damage in cycling somatic cells and in germ cells at all times. Noncycling germ cells that depended on homologous recombination underwent cell cycle arrest in G 2 , whereas noncycling somatic cells that depended on NHEJ arrested in G 1 , suggesting that cell cycle phase may modulate DNA repair during development. We conclude that error-prone NHEJ plays little or no role in DNA repair in C. elegans germ cells, possibly ensuring homology-based double-strand break repair and transmission of a stable genome from one generation to the next.

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The embryonic lethality in DNA ligase IV-deficient mice is rescued by deletion of Ku: implications for unifying the heterogeneous phenotypes of NHEJ mutants

Cited by 89 publications

References 43 publications

Defects in XRCC4 and KU80 differentially affect the joining of distal nonhomologous ends

Defects in XRCC4 and KU80 differentially affect the joining of distal nonhomologous ends

Nonhomologous DNA end-joining for repair of DNA double-strand breaks

Developmental Modulation of Nonhomologous End Joining in Caenorhabditis elegans

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