RAG2 and XLF/Cernunnos interplay reveals a novel role for the RAG complex in DNA repair

Lescale, Chloé; Abramowski, Vincent; Bedora-Faure, Marie; Murigneux, Valentine; Vera, Gabriella; Roth, David B.; Revy, Patrick; Villartay, Jean‐Pierre de; Deriano, Ludovic

doi:10.1038/ncomms10529

Cited by 62 publications

(142 citation statements)

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“…The difference in effect on DNA repair and V(D)J recombination in XLF deficiency might be explained by the 2-tier synapse model that has recently been proposed by Lescale et al, showing that both the RAG-PCC and XLF ensure stabilization of DNA ends after RAG1/2 cleavage. 24 They showed that the RAG-PCC has functional redundancy with XLF in RAG DSB stabilization, which indicates that in the presence of RAG, DNA end repair can be achieved during V(D)J recombination.…”

Section: Discussionmentioning

confidence: 99%

“…Since its discovery in 2005, several functions have been described for XLF. It stimulates LIG4 activity, 21,22 and it can form long filaments with XRCC4 that keep the DNA ends together in a ligation synapse, 24,[48][49][50][51] and XLF is essential for gap-filling by polymerase (pol) l and polm during NHEJ. The latter function is interesting, as poll and polm belong to the same polX family of polymerases as For personal use only.…”

Section: Discussionmentioning

confidence: 99%

“…23 However, recently, Lescale et al have shown that the RAG postcleavage complex (RAG-PCC) and XLF have overlapping functions and have suggested that both the RAG-PCC and XLF ensure stabilization of DNA ends after DNA cleavage by RAG. 24 To further understand the function of XLF during V(D)J recombination and NHEJ, the characteristics of the immunoglobulin heavy chain (IgH), Igk (IgK), Igl (IgL), TR b (TRB), and TR d (TRD) gene rearrangements were studied in 6 XLF-deficient patients, using nextgeneration sequencing. We identified an additional role for XLF in the contribution to junctional diversity during V(D)J recombination.…”

Section: Introductionmentioning

confidence: 99%

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XLF deficiency results in reduced N-nucleotide addition during V(D)J recombination

IJspeert

Rozmus

Schwarz

et al. 2016

Blood

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Key Points• XLF belongs to the NHEJ ligation complex and has a dual role in DNA doublestrand break repair and V(D)J recombination.• XLF is involved in Nnucleotide addition, and thereby contributes to junctional diversity of the antigen receptors.Repair of DNA double-strand breaks (DSBs) by the nonhomologous end-joining pathway (NHEJ) is important not only for repair of spontaneous breaks but also for breaks induced in developing lymphocytes during V(D)J (variable [V], diversity [D], and joining [J] genes) recombination of their antigen receptor loci to create a diverse repertoire. Mutations in the NHEJ factor XLF result in extreme sensitivity for ionizing radiation, microcephaly, and growth retardation comparable to mutations in LIG4 and XRCC4, which together form the NHEJ ligation complex. However, the effect on the immune system is variable (mild to severe immunodeficiency) and less prominent than that seen in deficiencies of NHEJ factors ARTEMIS and DNA-dependent protein kinase catalytic subunit, with defects in the hairpin opening step, which is crucial and unique for V(D)J recombination. Therefore, we aimed to study the role of XLF during V(D)J recombination. We obtained clinical data from 9 XLF-deficient patients and performed immune phenotyping and antigen receptor repertoire analysis of immunoglobulin (Ig) and T-cell receptor (TR) rearrangements, using next-generation sequencing in 6 patients. The results were compared with XRCC4 and LIG4 deficiency. Both Ig and TR rearrangements showed a significant decrease in the number of nontemplated (N) nucleotides inserted by terminal deoxynucleotidyl transferase, which resulted in a decrease of 2 to 3 amino acids in the CDR3. Such a reduction in the number of N-nucleotides has a great effect on the junctional diversity, and thereby on the total diversity of the Ig and TR repertoire. This shows that XLF has an important role during V(D)J recombination in creating diversity of the repertoire by stimulating N-nucleotide insertion. (Blood. 2016;128(5):650-659)

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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XLF deficiency results in reduced N-nucleotide addition during V(D)J recombination

IJspeert

Rozmus

Schwarz

et al. 2016

Blood

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“…To help explain these findings, functional redundancy with other factors was suggested and, in due course, demonstrated by XLF loss being shown to display nonepistatic or synthetic-lethal relationships with loss of components of the ATM-H2AX-53BP1 pathway or with C-terminal mutants of the V(D)J recombinase protein RAG2 (Zha et al 2011;Liu et al 2012;Oksenych et al 2012;Lescale et al 2016a). Our data reveal that PAXX and XLF also share Synthetic lethality between PAXX and XLF one or more redundant functions, as, unlike single mutants, mice lacking both proteins display severe growth defects, extensive cell death in the CNS, and an almost complete block to lymphocyte development similar to that of Xrcc4 −/− or Lig4 −/− mice.…”

Section: Xlfmentioning

confidence: 99%

Synthetic lethality between PAXX and XLF in mammalian development

et al. 2016

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PAXX was identified recently as a novel nonhomologous end-joining DNA repair factor in human cells. To characterize its physiological roles, we generated Paxx-deficient mice. Like Xlf−/− mice, Paxx−/− mice are viable, grow normally, and are fertile but show mild radiosensitivity. Strikingly, while Paxx loss is epistatic with Ku80, Lig4, and Atm deficiency, Paxx/Xlf double-knockout mice display embryonic lethality associated with genomic instability, cell death in the central nervous system, and an almost complete block in lymphogenesis, phenotypes that closely resemble those of Xrcc4−/− and Lig4−/− mice. Thus, combined loss of Paxx and Xlf is synthetic-lethal in mammals.

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“…Potential processes in which XLF and DSBR factors may be functionally redundant are not well-characterized but may include tethering ends or facilitating their joining (6,16). Notably, XLF also has functional redundancy with a truncation mutant of RAG2 for CE joining during V(D)J recombination, potentially implicating the RAG2 protein in some aspect of shepherding the V(D)J recombination joining reaction specifically to C-NHEJ (20,21).…”

mentioning

confidence: 99%

PAXX and XLF DNA repair factors are functionally redundant in joining DNA breaks in a G1-arrested progenitor B-cell line

Kumar

Frock

2016

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

105

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Classical nonhomologous end joining (C-NHEJ) is a major mammalian DNA double-strand break (DSB) repair pathway. Core C-NHEJ factors, such as XRCC4, are required for joining DSB intermediates of the G1 phase-specific V(D)J recombination reaction in progenitor lymphocytes. Core factors also contribute to joining DSBs in cycling mature B-lineage cells, including DSBs generated during antibody class switch recombination (CSR) and DSBs generated by ionizing radiation. The XRCC4-like-factor (XLF) C-NHEJ protein is dispensable for V(D)J recombination in normal cells, but because of functional redundancy, it is absolutely required for this process in cells deficient for the ataxia telangiectasia-mutated (ATM) DSB response factor. The recently identified paralogue of XRCC4 and XLF (PAXX) factor has homology to these two proteins and variably contributes to ionizing radiation-induced DSB repair in human and chicken cells. We now report that PAXX is dispensable for joining V(D)J recombination DSBs in G1-arrested mouse pro-B-cell lines, dispensable for joining CSR-associated DSBs in a cycling mouse B-cell line, and dispensable for normal ionizing radiation resistance in both G1-arrested and cycling pro-B lines. However, we find that combined deficiency for PAXX and XLF in G1-arrested pro-B lines abrogates DSB joining during V(D)J recombination and sensitizes the cells to ionizing radiation exposure. Thus, PAXX provides core C-NHEJ factor-associated functions in the absence of XLF and vice versa in G1-arrested pro-B-cell lines. Finally, we also find that PAXX deficiency has no impact on V(D)J recombination DSB joining in ATM-deficient pro-B lines. We discuss implications of these findings with respect to potential PAXX and XLF functions in C-NHEJ.T o repair DNA double-strand breaks (DSBs), mammalian cells use two major DSB repair pathways: homologous recombination (HR) and classical nonhomologous end joining (C-NHEJ). HR requires a long homologous template for repair and is active in S/G2 cell cycle phase (1). In contrast, C-NHEJ, which directly ligates DSBs with short (1-4 bp) or no homologies, functions throughout interphase, including during the G1 cell cycle phase (2-4). C-NHEJ repairs diverse DSBs, including those arising ectopically by ionizing radiation as well as DSBs generated as intermediates during V(D)J recombination in developing progenitor (pro)-B and -T lymphocytes and Ig heavy chain (IgH) class switch recombination (CSR) in activated mature B lymphocytes (5). The C-NHEJ pathway has a number of characterized members, including "core" C-NHEJ factors that were so-named in part based on their requirement for joining known types of ends via C-NHEJ and their evolutionary conservation (6). Such core factors include the Ku70 and Ku80 proteins, which form the "Ku" end recognition complex, and the XRCC4 and DNA Ligase4 proteins, which form the C-NHEJ ligation complex (5, 6).Exons that encode Ig and T-cell receptor (TCR) variable region exons are assembled from germline V, D, and J gene segments. This V(D)J recombinatio...

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RAG2 and XLF/Cernunnos interplay reveals a novel role for the RAG complex in DNA repair

Cited by 62 publications

References 70 publications

XLF deficiency results in reduced N-nucleotide addition during V(D)J recombination

XLF deficiency results in reduced N-nucleotide addition during V(D)J recombination

Synthetic lethality between PAXX and XLF in mammalian development

PAXX and XLF DNA repair factors are functionally redundant in joining DNA breaks in a G1-arrested progenitor B-cell line

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