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The molecular basis and disease relevance of non-homologous DNA end joining

Nature Reviews Molecular Cell Biology volume 21, pages 765–781 (2020)Cite this article

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Abstract

Non-homologous DNA end joining (NHEJ) is the predominant repair mechanism of any type of DNA double-strand break (DSB) during most of the cell cycle and is essential for the development of antigen receptors. Defects in NHEJ result in sensitivity to ionizing radiation and loss of lymphocytes. The most critical step of NHEJ is synapsis, or the juxtaposition of the two DNA ends of a DSB, because all subsequent steps rely on it. Recent findings show that, like the end processing step, synapsis can be achieved through several mechanisms. In this Review, we first discuss repair pathway choice between NHEJ and other DSB repair pathways. We then integrate recent insights into the mechanisms of NHEJ synapsis with updates on other steps of NHEJ, such as DNA end processing and ligation. Finally, we discuss NHEJ-related human diseases, including inherited disorders and neoplasia, which arise from rare failures at different NHEJ steps.

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Fig. 1: Overview of the non-homologous DNA end joining process.
Fig. 2: DSB repair pathway choice.
Fig. 3: Mechanisms of NHEJ synapsis.
Fig. 4: Various NHEJ end processing mechanisms.
Fig. 5: Disease-related NHEJ hypomorphic protein variants identified in humans.

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References

  1. Daley, J. M., Niu, H., Miller, A. S. & Sung, P. Biochemical mechanism of DSB end resection and its regulation. DNA Repair 32, 66–74 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  2. Kaniecki, K., De Tullio, L. & Greene, E. C. A change of view: homologous recombination at single-molecule resolution. Nat. Rev. Genet. 19, 191–207 (2018).

    CAS  PubMed  Google Scholar 

  3. Wright, W. D., Shah, S. S. & Heyer, W.-D. Homologous recombination and the repair of DNA double-strand breaks. J. Biol. Chem. 293, 10524–10535 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Piazza, A. & Heyer, W.-D. Homologous recombination and the formation of complex genomic rearrangements. Trends Cell Biol. 29, 135–149 (2019).

    CAS  PubMed  Google Scholar 

  5. Bell, J. C. & Kowalczykowski, S. C. Mechanics and single-molecule interrogation of DNA recombination. Annu. Rev. Biochem. 85, 193–226 (2016).

    CAS  PubMed  Google Scholar 

  6. Scully, R., Panday, A., Elango, R. & Willis, N. A. DNA double-strand break repair-pathway choice in somatic mammalian cells. Nat. Rev. Mol. Cell Biol. 20, 698–714 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  7. Chang, H. H. Y. et al. Different DNA end configurations dictate which NHEJ components are most important for joining efficiency. J. Biol. Chem. 291, 24377–24389 (2016). This article describes a comprehensive in vitro reconstitution of joining by NHEJ of diverse DNA end structures.

    CAS  PubMed  PubMed Central  Google Scholar 

  8. Waters, C. A. et al. The fidelity of the ligation step determines how ends are resolved during nonhomologous end joining. Nat. Commun. 5, 4286 (2014). This article provides evidence for a ligation-first hierarchy of attempted NHEJ factor engagement at DSBs, resulting in parsimony in the number of processing steps required for resolution.

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Reid, D. A. et al. Organization and dynamics of the nonhomologous end-joining machinery during DNA double-strand break repair. Proc. Natl Acad. Sci. USA 112, E2575–E2584 (2015). This article reports the first reconstituted NHEJ synapsis using purified NHEJ proteins and using a single-molecule Förster resonance energy transfer method.

    CAS  PubMed  Google Scholar 

  10. Zhao, B. et al. The essential elements for the noncovalent association of two DNA ends during NHEJ synapsis. Nat. Commun. 10, 3588 (2019). This article reports that two kinds of synaptic complexes (flexible synapsis and close synapsis) are formed for NHEJ synapsis, and that DNA-PKcs is not required for synapsis.

    PubMed  PubMed Central  Google Scholar 

  11. Zhao, B., Watanabe, G. & Lieber, M. R. Polymerase μ in non-homologous DNA end joining: importance of the order of arrival at a double-strand break in a purified system. Nucleic Acids Res. 48, 3605–3618 (2020). This article reports that Polμ can mediate NHEJ synapsis by itself, and describes for the first time that synapsis can be flexible.

    PubMed  PubMed Central  Google Scholar 

  12. Reid, D. A. et al. Bridging of double-stranded breaks by the nonhomologous end-joining ligation complex is modulated by DNA end chemistry. Nucleic Acids Res. 45, 1872–1878 (2017).

    CAS  PubMed  Google Scholar 

  13. Conlin, M. P. et al. DNA ligase IV guides end-processing choice during nonhomologous end joining. Cell Rep. 20, 2810–2819 (2017). This article determines that tolerance of DNA ends with nucleotide damage requires XRCC4–LIG4-dependent remodelling of the ends within a synaptic complex, and that this is important for cell resistance to ionizing radiation.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Pannunzio, N. R., Watanabe, G. & Lieber, M. R. Nonhomologous DNA end joining for repair of DNA double-strand breaks. J. Biol. Chem. 293, 10512–10523 (2018).

    CAS  PubMed  Google Scholar 

  15. Chang, H. H. Y., Pannunzio, N. R., Adachi, N. & Lieber, M. R. Non-homologous DNA end joining and alternative pathways to double-strand break repair. Nat. Rev. Mol. Cell Biol. 18, 495 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  16. Lieber, M. R. The mechanism of double-strand DNA break repair by the nonhomologous DNA end-joining pathway. Annu. Rev. Biochem. 79, 181–211 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  17. Beucher, A. et al. ATM and Artemis promote homologous recombination of radiation-induced DNA double-strand breaks in G2. EMBO J. 28, 3413–3427 (2009).

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Rothkamm, K., Krüger, I., Thompson, L. H. & Löbrich, M. Pathways of DNA double-strand break repair during the mammalian cell cycle. Mol. Cell. Biol. 23, 5706–5715 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Karanam, K., Kafri, R., Loewer, A. & Lahav, G. Quantitative live cell imaging reveals a gradual shift between DNA repair mechanisms and a maximal use of HR in mid S phase. Mol. Cell 47, 320–329 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Shibata, A. et al. Factors determining DNA double-strand break repair pathway choice in G2 phase. EMBO J. 30, 1079–1092 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Schatz, D. G. & Swanson, P. C. V(D)J recombination: mechanisms of initiation. Annu. Rev. Genet. 45, 167–202 (2011).

    CAS  PubMed  Google Scholar 

  22. Wang, X. S., Lee, B. J. & Zha, S. The recent advances in non-homologous end-joining through the lens of lymphocyte development. DNA Repair 94, 102874 (2020).

    CAS  PubMed  Google Scholar 

  23. Taccioli, G. E. et al. Impairment of V(D)J recombination in double-strand break repair mutants. Science 260, 207–210 (1993).

    CAS  PubMed  Google Scholar 

  24. Chaudhuri, J. & Alt, F. W. Class-switch recombination: interplay of transcription, DNA deamination and DNA repair. Nat. Rev. Immunol. 4, 541–552 (2004).

    CAS  PubMed  Google Scholar 

  25. Mimitou, E. P. & Symington, L. S. Ku prevents Exo1 and Sgs1-dependent resection of DNA ends in the absence of a functional MRX complex or Sae2. EMBO J. 29, 3358–3369 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  26. van Schendel, R., Roerink, S. F., Portegijs, V., van den Heuvel, S. & Tijsterman, M. Polymerase Θ is a key driver of genome evolution and of CRISPR/Cas9-mediated mutagenesis. Nat. Commun. 6, 7394 (2015).

    PubMed  PubMed Central  Google Scholar 

  27. van Kregten, M. et al. T-DNA integration in plants results from polymerase-θ-mediated DNA repair. Nat. Plants 2, 16164 (2016).

    PubMed  Google Scholar 

  28. Chan, S. H., Yu, A. M. & McVey, M. Dual roles for DNA polymerase theta in alternative end-joining repair of double-strand breaks in Drosophila. PLoS Genet. 6, e1001005 (2010).

    PubMed  PubMed Central  Google Scholar 

  29. Saito, S., Maeda, R. & Adachi, N. Dual loss of human POLQ and LIG4 abolishes random integration. Nat. Commun. 8, 16112 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Schimmel, J., Kool, H., van Schendel, R. & Tijsterman, M. Mutational signatures of non-homologous and polymerase theta-mediated end-joining in embryonic stem cells. EMBO J. 36, 3634–3649 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Carvajal-Garcia, J. et al. Mechanistic basis for microhomology identification and genome scarring by polymerase theta. Proc. Natl Acad. Sci. USA 117, 8476–8485 (2020).

    CAS  PubMed  Google Scholar 

  32. Wyatt, D. W. et al. Essential roles for polymerase θ-mediated end joining in the repair of chromosome breaks. Mol. Cell 63, 662–673 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  33. He, P. & Yang, W. Template and primer requirements for DNA Pol θ-mediated end joining. Proc. Natl Acad. Sci. USA 115, 7747–7752 (2018).

    CAS  PubMed  Google Scholar 

  34. Zhang, Y. & Jasin, M. An essential role for CtIP in chromosomal translocation formation through an alternative end-joining pathway. Nat. Struct. Mol. Biol. 18, 80–84 (2011).

    CAS  PubMed  Google Scholar 

  35. Lee-Theilen, M., Matthews, A. J., Kelly, D., Zheng, S. & Chaudhuri, J. CtIP promotes microhomology-mediated alternative end joining during class-switch recombination. Nat. Struct. Mol. Biol. 18, 75–79 (2011).

    CAS  PubMed  Google Scholar 

  36. Mateos-Gomez, P. A. et al. Mammalian polymerase θ promotes alternative NHEJ and suppresses recombination. Nature 518, 254–257 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  37. Bhargava, R., Onyango, D. O. & Stark, J. M. Regulation of single-strand annealing and its role in genome maintenance. Trends Genet. 32, 566–575 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Kelso, A. A., Lopezcolorado, F. W., Bhargava, R. & Stark, J. M. Distinct roles of RAD52 and POLQ in chromosomal break repair and replication stress response. PLoS Genet. 15, e1008319 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Motycka, T. A., Bessho, T., Post, S. M., Sung, P. & Tomkinson, A. E. Physical and functional interaction between the XPF/ERCC1 endonuclease and hRad52. J. Biol. Chem. 279, 13634–13639 (2004).

    CAS  PubMed  Google Scholar 

  40. Lisby, M. & Rothstein, R. Cell biology of mitotic recombination. Cold Spring Harb. Perspect. Biol. 7, a016535 (2015).

    PubMed  PubMed Central  Google Scholar 

  41. Moore, J. K. & Haber, J. E. Cell cycle and genetic requirements of two pathways of nonhomologous end-joining repair of double-strand breaks in Saccharomyces cerevisiae. Mol. Cell. Biol. 16, 2164–2173 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  42. Gauss, G. H. & Lieber, M. R. Mechanistic constraints on diversity in human V(D)J recombination. Mol. Cell. Biol. 16, 258–269 (1996).

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Chapman, J. R., Taylor, Martin, R. G. & Boulton, Simon J. Playing the end game: DNA double-strand break repair pathway choice. Mol. Cell 47, 497–510 (2012).

    CAS  PubMed  Google Scholar 

  44. Symington, L. S. & Gautier, J. Double-strand break end resection and repair pathway choice. Annu. Rev. Genet. 45, 247–271 (2011).

    CAS  PubMed  Google Scholar 

  45. Xu, G. T. et al. REV7 counteracts DNA double-strand break resection and affects PARP inhibition. Nature 521, 541–U308 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  46. Boersma, V. et al. MAD2L2 controls DNA repair at telomeres and DNA breaks by inhibiting 5′ end resection. Nature 521, 537–540 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  47. Zimmermann, M., Lottersberger, F., Buonomo, S. B., Sfeir, A. & de Lange, T. 53BP1 regulates DSB repair using Rif1 to Control 5′ end resection. Science 339, 700–704 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  48. Escribano-Diaz, C. et al. A cell cycle-dependent regulatory circuit composed of 53BP1-RIF1 and BRCA1-CtIP controls DNA repair pathway choice. Mol. Cell 49, 872–883 (2013).

    CAS  PubMed  Google Scholar 

  49. Di Virgilio, M. et al. Rif1 prevents resection of DNA breaks and promotes immunoglobulin class switching. Science 339, 711–715 (2013).

    PubMed  Google Scholar 

  50. Daley, J. M. & Sung, P. RIF1 in DNA break repair pathway choice. Mol. Cell 49, 840–841 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Chapman, J. R. et al. RIF1 is essential for 53BP1-dependent nonhomologous end joining and suppression of DNA double-strand break resection. Mol. Cell 49, 858–871 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  52. Bunting, S. F. et al. 53BP1 inhibits homologous recombination in Brca1-deficient cells by blocking resection of DNA breaks. Cell 141, 243–254 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  53. Setiaputra, D. & Durocher, D. Shieldin – the protector of DNA ends. EMBO Rep. 20, e47560 (2019).

    PubMed  PubMed Central  Google Scholar 

  54. Tomida, J. et al. FAM35A associates with REV7 and modulates DNA damage responses of normal and BRCA1-defective cells. EMBO J. 37, e99543 (2018).

    PubMed  PubMed Central  Google Scholar 

  55. Noordermeer, S. M. et al. The shieldin complex mediates 53BP1-dependent DNA repair. Nature 560, 117–121 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  56. Mirman, Z. et al. 53BP1-RIF1-shieldin counteracts DSB resection through CST- and Polalpha-dependent fill-in. Nature 560, 112–116 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  57. Gupta, R. et al. DNA repair network analysis reveals shieldin as a key regulator of NHEJ and PARP inhibitor sensitivity. Cell 173, 972–988 e923 (2018).

    CAS  PubMed  Google Scholar 

  58. Ghezraoui, H. et al. 53BP1 cooperation with the REV7–shieldin complex underpins DNA structure-specific NHEJ. Nature 560, 122–127 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  59. Dev, H. et al. Shieldin complex promotes DNA end-joining and counters homologous recombination in BRCA1-null cells. Nat. Cell Biol. 20, 954–965 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  60. Gao, S. et al. An OB-fold complex controls the repair pathways for DNA double-strand breaks. Nat. Commun. 9, 3925 (2018). Together with references 54–59, this article describes the discovery of the shieldin complex, which is the effector of 53BP1–RIF1.

    PubMed  PubMed Central  Google Scholar 

  61. Panier, S. & Boulton, S. J. Double-strand break repair: 53BP1 comes into focus. Nat. Rev. Mol. Cell Biol. 15, 7–18 (2014).

    CAS  PubMed  Google Scholar 

  62. Botuyan, M. V. et al. Structural basis for the methylation state-specific recognition of histone H4-K20 by 53BP1 and Crb2 in DNA repair. Cell 127, 1361–1373 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  63. Fradet-Turcotte, A. et al. 53BP1 is a reader of the DNA-damage-induced H2A Lys 15 ubiquitin mark. Nature 499, 50–54 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  64. Ochs, F. et al. 53BP1 fosters fidelity of homology-directed DNA repair. Nat. Struct. Mol. Biol. 23, 714–721 (2016).

    CAS  PubMed  Google Scholar 

  65. Tsai, L. J. et al. RNF8 has both KU-dependent and independent roles in chromosomal break repair. Nucleic Acids Res. 48, 6032–6052 (2020).

    PubMed  PubMed Central  Google Scholar 

  66. Guirouilh-Barbat, J. et al. 53BP1 protects against CtIP-dependent capture of ectopic chromosomal sequences at the junction of distant double-strand breaks. PLoS Genet. 12, e1006230 (2016).

    PubMed  PubMed Central  Google Scholar 

  67. Clairmont, C. S. et al. TRIP13 regulates DNA repair pathway choice through REV7 conformational change. Nat. Cell Biol. 22, 87–96 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  68. Callen, E. et al. 53BP1 enforces distinct pre- and post-resection blocks on homologous recombination. Mol. Cell 77, 26–38.e27 (2020).

    CAS  PubMed  Google Scholar 

  69. Ling, A. K. et al. SHLD2 promotes class switch recombination by preventing inactivating deletions within the Igh locus. EMBO Rep. 21, e49823 (2020).

    CAS  Google Scholar 

  70. Slavoff, S. A., Heo, J., Budnik, B. A., Hanakahi, L. A. & Saghatelian, A. A human short open reading frame (sORF)-encoded polypeptide that stimulates DNA end joining. J. Biol. Chem. 289, 10950–10957 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  71. Arnoult, N. et al. Regulation of DNA repair pathway choice in S and G2 phases by the NHEJ inhibitor CYREN. Nature 549, 548–552 (2017).

    PubMed  PubMed Central  Google Scholar 

  72. Hung, P. J. et al. MRI is a DNA damage response adaptor during classical non-homologous end joining. Mol. Cell 71, 332–342 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  73. Castañeda-Zegarra, S. et al. Generation of a mouse model lacking the non-homologous end-joining factor Mri/Cyren. Biomolecules 9, 798 (2019).

    PubMed Central  Google Scholar 

  74. Tadi, Satish K. et al. PAXX is an accessory c-NHEJ factor that associates with Ku70 and has overlapping functions with XLF. Cell Rep. 17, 541–555 (2016).

    Google Scholar 

  75. Ahnesorg, P., Smith, P. & Jackson, S. P. XLF interacts with the XRCC4-DNA ligase IV complex to promote DNA nonhomologous end-joining. Cell 124, 301–313 (2006).

    CAS  PubMed  Google Scholar 

  76. Lu, H., Pannicke, U., Schwarz, K. & Lieber, M. R. Length-dependent binding of human XLF to DNA and stimulation of XRCC4.DNA ligase IV activity. J. Biol. Chem. 282, 11155–11162 (2007).

    CAS  PubMed  Google Scholar 

  77. Hammel, M., Yu, Y. P., Fang, S. J., Lees-Miller, S. P. & Tainer, J. A. XLF regulates filament architecture of the XRCC4·ligase IV complex. Structure 18, 1431–1442 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  78. Yano, K.-i., Morotomi-Yano, K., Lee, K.-J. & Chen, D. J. Functional significance of the interaction with Ku in DNA double-strand break recognition of XLF. FEBS Lett. 585, 841–846 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  79. Nemoz, C. et al. XLF and APLF bind Ku80 at two remote sites to ensure DNA repair by non-homologous end joining. Nat. Struct. Mol. Biol. 25, 971–980 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  80. Ochi, T. et al. DNA repair. PAXX, a paralog of XRCC4 and XLF, interacts with Ku to promote DNA double-strand break repair. Science 347, 185–188 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  81. Xing, M. et al. Interactome analysis identifies a new paralogue of XRCC4 in non-homologous end joining DNA repair pathway. Nat. Commun. 6, 6233 (2015). Together with reference 81, this srticle describes the discovery of PAXX.

    CAS  PubMed  PubMed Central  Google Scholar 

  82. Kumar, V., Alt, F. W. & Frock, R. L. PAXX and XLF DNA repair factors are functionally redundant in joining DNA breaks in a G1-arrested progenitor B-cell line. Proc. Natl Acad. Sci. USA 113, 10619–10624 (2016).

    CAS  PubMed  Google Scholar 

  83. Li, W. et al. The nucleoskeleton protein IFFO1 immobilizes broken DNA and suppresses chromosome translocation during tumorigenesis. Nat. Cell Biol. 21, 1273–1285 (2019).

    CAS  PubMed  Google Scholar 

  84. Graham, T. G., Walter, J. C. & Loparo, J. J. Two-stage synapsis of DNA ends during non-homologous end joining. Mol. Cell 61, 850–858 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  85. Graham, T. G. W., Carney, S. M., Walter, J. C. & Loparo, J. J. A single XLF dimer bridges DNA ends during nonhomologous end joining. Nat. Struct. Mol. Biol. 25, 877–884 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  86. Stinson, B. M., Moreno, A. T., Walter, J. C. & Loparo, J. J. A mechanism to minimize errors during non-homologous end joining. Mol. Cell 77, 1–12 (2020).

    Google Scholar 

  87. Wang, J. L. et al. Dissection of DNA double-strand-break repair using novel single-molecule forceps. Nat. Struct. Mol. Biol. 25, 482–487 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  88. Kulesza, P. & Lieber, M. R. DNA-PK is essential only for coding joint formation in V(D)J recombination. Nucleic Acids Res. 26, 3944–3948 (1998).

    CAS  PubMed  PubMed Central  Google Scholar 

  89. Gao, Y. et al. A targeted DNA-PKcs-null mutation reveals DNA-PK-independent functions for KU in V(D)J recombination. Immunity 9, 367–376 (1998).

    CAS  PubMed  Google Scholar 

  90. Jiang, W. et al. Differential phosphorylation of DNA-PKcs regulates the interplay between end-processing and end-ligation during nonhomologous end-joining. Mol. Cell 58, 172–185 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  91. Davis, B. J., Havener, J. M. & Ramsden, D. A. End-bridging is required for pol μ to efficiently promote repair of noncomplementary ends by nonhomologous end joining. Nucleic Acids Res. 36, 3085–3094 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  92. Andrade, P., Martín, M. J., Juárez, R., López de Saro, F. & Blanco, L. Limited terminal transferase in human DNA polymerase μ defines the required balance between accuracy and efficiency in NHEJ. Proc. Natl Acad. Sci. USA 106, 16203–16208 (2009).

    CAS  PubMed  Google Scholar 

  93. Kaminski, A. M. et al. Structural snapshots of human DNA polymerase μ engaged on a DNA double-strand break. Nat. Commun. 11, 4784 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  94. Loc’h, J. et al. Structural evidence for an in trans base selection mechanism involving Loop1 in polymerase μ at an NHEJ double-strand break junction. J. Biol. Chem. 294, 10579–10595 (2019).

    PubMed  PubMed Central  Google Scholar 

  95. Nick McElhinny, S. A. et al. A gradient of template dependence defines distinct biological roles for family X polymerases in nonhomologous end joining. Mol. Cell 19, 357–366 (2005).

    CAS  PubMed  Google Scholar 

  96. Loc’h, J. & Delarue, M. Terminal deoxynucleotidyltransferase: the story of an untemplated DNA polymerase capable of DNA bridging and templated synthesis across strands. Curr. Opin. Struct. Biol. 53, 22–31 (2018).

    PubMed  Google Scholar 

  97. Gouge, J. et al. Structural basis for a novel mechanism of DNA bridging and alignment in eukaryotic DSB DNA repair. EMBO J. 34, 1126–1142 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  98. Loc’h, J., Rosario, S. & Delarue, M. Structural basis for a new templated activity by terminal deoxynucleotidyl transferase: implications for V(D)J recombination. Structure 24, 1452–1463 (2016).

    PubMed  Google Scholar 

  99. Ma, Y. et al. A biochemically defined system for mammalian nonhomologous DNA end joining. Mol. Cell 16, 701–713 (2004).

    CAS  PubMed  Google Scholar 

  100. Ma, Y., Lu, H., Schwarz, K. & Lieber, M. R. Repair of double-strand DNA breaks by the human nonhomologous DNA end joining pathway: the iterative processing model. Cell Cycle 4, 1193–2000 (2005).

    CAS  PubMed  Google Scholar 

  101. Simsek, D. & Jasin, M. Alternative end-joining is suppressed by the canonical NHEJ component Xrcc4–ligase IV during chromosomal translocation formation. Nat. Struct. Mol. Biol. 17, 410–416 (2010).

    CAS  PubMed  PubMed Central  Google Scholar 

  102. Shen, M. W. et al. Predictable and precise template-free CRISPR editing of pathogenic variants. Nature 563, 646–651 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  103. Li, S. et al. Evidence that the DNA endonuclease ARTEMIS also has intrinsic 5′-exonuclease activity. J. Biol. Chem. 289, 7825–7834 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  104. Ma, Y., Pannicke, U., Schwarz, K. & Lieber, M. R. Hairpin opening and overhang processing by an Artemis:DNA-PKcs complex in V(D)J recombination and in nonhomologous end joining. Cell 108, 781–794 (2002).

    CAS  PubMed  Google Scholar 

  105. Gerodimos, C. A., Chang, H. H. Y., Watanabe, G. & Lieber, M. R. Effects of DNA end configuration on XRCC4:DNA ligase IV and its stimulation of Artemis activity. J. Biol. Chem. 292, 13914–13924 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  106. Chang, H. H., Watanabe, G. & Lieber, M. R. Unifying the DNA end-processing roles of the Artemis nuclease: Ku-dependent Artemis resection at blunt DNA ends. J. Biol. Chem. 290, 24036–24050 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  107. Lu, H. et al. A biochemically defined system for coding joint formation in human V(D)J recombination. Mol. Cell 31, 485–497 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  108. Henner, W. D., Grunberg, S. M. & Haseltine, W. A. Enzyme action at 3′ termini of ionizing radiation-induced DNA strand breaks. J. Biol. Chem. 258, 15198–15205 (1983).

    CAS  PubMed  Google Scholar 

  109. Henner, W. D., Rodriguez, L. O., Hecht, S. M. & Haseltine, W. A. Gamma ray induced deoxyribonucleic acid strand breaks. 3′ glycolate termini. J. Biol. Chem. 258, 711–713 (1983).

    CAS  PubMed  Google Scholar 

  110. Valerie, K. & Povirk, L. F. Regulation and mechanisms of mammalian double-strand break repair. Oncogene 22, 5792–5812 (2003).

    CAS  PubMed  Google Scholar 

  111. Zhou, T. et al. Deficiency in 3′-phosphoglycolate processing in human cells with a hereditary mutation in tyrosyl-DNA phosphodiesterase (TDP1). Nucleic Acids Res. 33, 289–297 (2005).

    CAS  PubMed  PubMed Central  Google Scholar 

  112. Povirk, L. F., Zhou, T., Zhou, R., Cowan, M. J. & Yannone, S. M. Processing of 3′-phosphoglycolate-terminated DNA double strand breaks by Artemis nuclease. J. Biol. Chem. 282, 3547–3558 (2007).

    CAS  PubMed  Google Scholar 

  113. Yannone, S. M. et al. Coordinate 5′ and 3′ endonucleolytic trimming of terminally blocked blunt DNA double-strand break ends by Artemis nuclease and DNA-dependent protein kinase. Nucleic Acids Res. 36, 3354–3365 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  114. Anne Esguerra, Z., Watanabe, G., Okitsu, C. Y., Hsieh, C.-L. & Lieber, M. R. DNA-PKcs chemical inhibition versus genetic mutation: impact on the junctional repair steps of V(D)J recombination. Mol. Immunol. 120, 93–100 (2020).

    CAS  PubMed  Google Scholar 

  115. Moshous, D. et al. Artemis, a novel DNA double-strand break repair/V(D)J recombination protein, is mutated in human severe combined immune deficiency. Cell 105, 177–186 (2001).

    CAS  PubMed  Google Scholar 

  116. Blunt, T. et al. Defective DNA-dependent protein kinase activity is linked to V(D)J recombination and DNA repair defects associated with the murine scid mutation. Cell 80, 813–823 (1995).

    CAS  PubMed  Google Scholar 

  117. Woodbine, L. et al. PRKDC mutations in a SCID patient with profound neurological abnormalities. J. Clin. Invest. 123, 2969–2980 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  118. Li, S. et al. Polynucleotide kinase and aprataxin-like forkhead-associated protein (PALF) acts as both a single-stranded DNA endonuclease and a single-stranded DNA 3′ exonuclease and can participate in DNA end joining in a biochemical system. J. Biol. Chem. 286, 36368–36377 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  119. Pryor, J. M. et al. Essential role for polymerase specialization in cellular nonhomologous end joining. Proc. Natl Acad. Sci. USA 112, E4537–4545 (2015). This article demonstrates that in NHEJ, Polµ or Polλ is partially redundant; loss of both ablates almost all synthesis during NHEJ and severely impairs the pathway.

    CAS  PubMed  Google Scholar 

  120. Mahajan, K. N., Nick McElhinny, S. A., Mitchell, B. S. & Ramsden, D. A. Association of DNA polymerase μ (pol μ) with Ku and ligase IV: role for pol μ in end-joining double-strand break repair. Mol. Cell. Biol. 22, 5194–5202 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  121. Martin, M. J., Juarez, R. & Blanco, L. DNA-binding determinants promoting NHEJ by human Polµ. Nucleic Acids Res. 40, 11389–11403 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  122. Gilfillan, S., Dierich, A., Lemeur, M., Benoist, C. & Mathis, D. Mice lacking TdT: mature animals with an immature lymphocyte repertoire. Science 261, 1175–1178 (1993).

    CAS  PubMed  Google Scholar 

  123. Lange, S. S., Takata, K. & Wood, R. D. DNA polymerases and cancer. Nat. Rev. Cancer 11, 96–110 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  124. Moon, A. F. et al. Structural insight into the substrate specificity of DNA polymerase μ. Nat. Struct. Mol. Biol. 14, 45–53 (2007).

    CAS  PubMed  Google Scholar 

  125. Domínguez, O. et al. DNA polymerase mu (Pol mu), homologous to TdT, could act as a DNA mutator in eukaryotic cells. EMBO J. 19, 1731–1742 (2000).

    PubMed  PubMed Central  Google Scholar 

  126. Gu, J. et al. XRCC4:DNA ligase IV can ligate incompatible DNA ends and can ligate across gaps. EMBO J. 26, 1010–1023 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  127. Moon, A. F. et al. The X family portrait: structural insights into biological functions of X family polymerases. DNA Repair 6, 1709–1725 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  128. Kato, K. I., Gonçalves, J. M., Houts, G. E. & Bollum, F. J. Deoxynucleotide-polymerizing enzymes of calf thymus gland. II. Properties of the terminal deoxynucleotidyltransferase. J. Biol. Chem. 242, 2780–2789 (1967).

    CAS  PubMed  Google Scholar 

  129. Boulé, J. B., Rougeon, F. & Papanicolaou, C. Terminal deoxynucleotidyl transferase indiscriminately incorporates ribonucleotides and deoxyribonucleotides. J. Biol. Chem. 276, 31388–31393 (2001).

    PubMed  Google Scholar 

  130. Ruiz, J. F. et al. Lack of sugar discrimination by human Pol µ requires a single glycine residue. Nucleic Acids Res. 31, 4441–4449 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  131. Martin, M. J., Garcia-Ortiz, M. V., Esteban, V. & Blanco, L. Ribonucleotides and manganese ions improve non-homologous end joining by human Polµ. Nucleic Acids Res. 41, 2428–2436 (2012).

    PubMed  PubMed Central  Google Scholar 

  132. Nick McElhinny, S. A. & Ramsden, D. A. Polymerase Mu is a DNA-directed DNA/RNA polymerase. Mol. Cell. Biol. 23, 2309–2315 (2003).

    CAS  PubMed  PubMed Central  Google Scholar 

  133. Pryor, J. M. et al. Ribonucleotide incorporation enables repair of chromosome breaks by nonhomologous end joining. Science 361, 1126–1129 (2018). This article shows that polymerases in cells very often incorporate ribonucleotides during NHEJ, and that this facilitates the ligation step.

    CAS  PubMed  PubMed Central  Google Scholar 

  134. Lieber, M. R., Hesse, J. E., Mizuuchi, K. & Gellert, M. Lymphoid V(D)J recombination: nucleotide insertion at signal joints as well as coding joints. Proc. Natl Acad. Sci. USA 85, 8588–8592 (1988).

    CAS  PubMed  Google Scholar 

  135. Duvauchelle, J.-B., Blanco, L., Fuchs, R. P. P. & Cordonnier, A. M. Human DNA polymerase mu (Pol µ) exhibits an unusual replication slippage ability at AAF lesion. Nucleic Acids Res. 30, 2061–2067 (2002).

    CAS  PubMed  PubMed Central  Google Scholar 

  136. Tippin, B., Kobayashi, S., Bertram, J. G. & Goodman, M. F. To slip or skip, visualizing frameshift mutation dynamics for error-prone DNA polymerases. J. Biol. Chem. 279, 45360–45368 (2004).

    CAS  PubMed  Google Scholar 

  137. Maga, G. et al. DNA elongation by the human DNA polymerase λ polymerase and terminal transferase activities are differentially coordinated by proliferating cell nuclear antigen and replication protein A. J. Biol. Chem. 280, 1971–1981 (2005).

    CAS  PubMed  Google Scholar 

  138. Jäger, U. et al. Follicular lymphomas BCL-2/IgH junctions contain templated nucleotide insertions: novel insights into the mechanism of t(14;18) translocation. Blood 95, 3520–3529 (2000).

    PubMed  Google Scholar 

  139. Welzel, N. et al. Templated nucleotide addition and immunoglobulin JH-gene utilization in t(11;14) junctions: implications for the mechanism of translocation and the origin of mantle cell lymphoma. Cancer Res. 61, 1629–1636 (2001).

    CAS  PubMed  Google Scholar 

  140. Grawunder, U. et al. Activity of DNA ligase IV stimulated by complex formation with XRCC4 protein in mammalian cells. Nature 388, 492–495 (1997).

    CAS  PubMed  Google Scholar 

  141. Wilson, T. E., Grawunder, U. & Lieber, M. R. Yeast DNA ligase IV mediates non-homologous DNA end joining. Nature 388, 495–498 (1997).

    CAS  PubMed  Google Scholar 

  142. Schar, P., Herrmann, G., Daly, G. & Lindahl, T. A newly identified DNA ligase of Saccharomyces cerevisiae involved in RAD52-independent repair of DNA double-strand breaks. Genes Dev. 11, 1912–1924 (1997).

    CAS  PubMed  PubMed Central  Google Scholar 

  143. Chen, S.-H. & Yu, X. Human DNA ligase IV is able to use NAD+ as an alternative adenylation donor for DNA ends ligation. Nucleic Acids Res. 47, 1321–1334 (2019).

    CAS  PubMed  Google Scholar 

  144. Liu, X., Shao, Z., Jiang, W., Lee, B. J. & Zha, S. PAXX promotes KU accumulation at DNA breaks and is essential for end-joining in XLF-deficient mice. Nat. Commun. 8, 13816 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  145. Balmus, G. et al. Synthetic lethality between PAXX and XLF in mammalian development. Genes Dev. 30, 2152–2157 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  146. Lescale, C. et al. Specific roles of XRCC4 paralogs PAXX and XLF during V(D)J recombination. Cell Rep. 16, 2967–2979 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  147. Ropars, V. et al. Structural characterization of filaments formed by human Xrcc4-Cernunnos/XLF complex involved in nonhomologous DNA end-joining. Proc. Natl Acad. Sci. USA 108, 12663–12668 (2011).

    CAS  PubMed  Google Scholar 

  148. Brouwer, I. et al. Sliding sleeves of XRCC4-XLF bridge DNA and connect fragments of broken DNA. Nature 535, 566–569 (2016).

    CAS  PubMed  Google Scholar 

  149. Bermudez-Hernandez, K. et al. A method for quantifying molecular interactions using stochastic modelling and super-resolution microscopy. Sci. Rep. 7, 14882 (2017).

    PubMed  PubMed Central  Google Scholar 

  150. Wang, Y., Lamarche, B. J. & Tsai, M. D. Human DNA ligase IV and the ligase IV/XRCC4 complex: analysis of nick ligation fidelity. Biochemistry 46, 4962–4976 (2007).

    CAS  PubMed  Google Scholar 

  151. Gu, J., Lu, H., Tsai, A. G., Schwarz, K. & Lieber, M. R. Single-stranded DNA ligation and XLF-stimulated incompatible DNA end ligation by the XRCC4-DNA ligase IV complex: influence of terminal DNA sequence. Nucleic Acids Res. 35, 5755–5762 (2007).

    CAS  PubMed  PubMed Central  Google Scholar 

  152. Tsai, C. J., Kim, S. A. & Chu, G. Cernunnos/XLF promotes the ligation of mismatched and noncohesive DNA ends. Proc. Natl Acad. Sci. USA 104, 7851–7856 (2007).

    CAS  PubMed  Google Scholar 

  153. Roberts, S. A. & Ramsden, D. A. Loading of the nonhomologous end joining factor, Ku, on protein-occluded DNA ends. J. Biol. Chem. 282, 10605–10613 (2007).

    CAS  PubMed  Google Scholar 

  154. Batenburg, N. L. et al. ATM and CDK2 control chromatin remodeler CSB to inhibit RIF1 in DSB repair pathway choice. Nat. Commun. 8, 1921 (2017).

    PubMed  PubMed Central  Google Scholar 

  155. Dos Santos, M. et al. Influence of chromatin condensation on the number of direct DSB damages induced by ions studied using a Monte Carlo code. Radiat. Prot. Dosimetry 161, 469–473 (2014).

    PubMed  Google Scholar 

  156. Falk, M., Lukasova, E. & Kozubek, S. Higher-order chromatin structure in DSB induction, repair and misrepair. Mutat. Res. 704, 88–100 (2010).

    CAS  PubMed  Google Scholar 

  157. Falk, M., Lukasova, E., Gabrielova, B., Ondrej, V. & Kozubek, S. Chromatin dynamics during DSB repair. Biochim. Biophys. Acta 1773, 1534–1545 (2007).

    CAS  PubMed  Google Scholar 

  158. Altmeyer, M. et al. Liquid demixing of intrinsically disordered proteins is seeded by poly(ADP-ribose). Nat. Commun. 6, 8088 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  159. Menolfi, D., Zha, S. & ATM, A. T. R. and DNA-PKcs kinases-the lessons from the mouse models: inhibition not equal deletion. Cell Biosci. 10, 8 (2020).

    PubMed  PubMed Central  Google Scholar 

  160. Stucki, M. & Jackson, S. P. gammaH2AX and MDC1: anchoring the DNA-damage-response machinery to broken chromosomes. DNA Repair 5, 534–543 (2006).

    CAS  PubMed  Google Scholar 

  161. Doil, C. et al. RNF168 binds and amplifies ubiquitin conjugates on damaged chromosomes to allow accumulation of repair proteins. Cell 136, 435–446 (2009).

    CAS  PubMed  Google Scholar 

  162. Stewart, G. S. et al. The RIDDLE syndrome protein mediates a ubiquitin-dependent signaling cascade at sites of DNA damage. Cell 136, 420–434 (2009).

    CAS  PubMed  Google Scholar 

  163. Baranes-Bachar, K. et al. The ubiquitin E3/E4 ligase UBE4A adjusts protein ubiquitylation and accumulation at sites of DNA damage, facilitating double-strand break repair. Mol. Cell 69, 866–878 e867 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  164. Kilic, S. et al. Phase separation of 53BP1 determines liquid-like behavior of DNA repair compartments. EMBO J. 38, e101379 (2019).

    PubMed  PubMed Central  Google Scholar 

  165. Ariumi, Y. et al. Suppression of the poly(ADP-ribose) polymerase activity by DNA-dependent protein kinase in vitro. Oncogene 18, 4616–4625 (1999).

    CAS  PubMed  Google Scholar 

  166. Veuger, S. J., Curtin, N. J., Smith, G. C. & Durkacz, B. W. Effects of novel inhibitors of poly(ADP-ribose) polymerase-1 and the DNA-dependent protein kinase on enzyme activities and DNA repair. Oncogene 23, 7322–7329 (2004).

    CAS  PubMed  Google Scholar 

  167. Hochegger, H. et al. Parp-1 protects homologous recombination from interference by Ku and ligase IV in vertebrate cells. EMBO J. 25, 1305–1314 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  168. Wang, M. et al. PARP-1 and Ku compete for repair of DNA double strand breaks by distinct NHEJ pathways. Nucleic Acids Res. 34, 6170–6182 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  169. Cheng, Q. et al. Ku counteracts mobilization of PARP1 and MRN in chromatin damaged with DNA double-strand breaks. Nucleic Acids Res. 39, 9605–9619 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  170. Frit, P., Barboule, N., Yuan, Y., Gomez, D. & Calsou, P. Alternative end-joining pathway(s): bricolage at DNA breaks. DNA Repair 17, 81–97 (2014).

    CAS  PubMed  Google Scholar 

  171. Yang, G. et al. Super-resolution imaging identifies PARP1 and the Ku complex acting as DNA double-strand break sensors. Nucleic Acids Res. 46, 3446–3457 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  172. Couto, C. A. et al. PARP regulates nonhomologous end joining through retention of Ku at double-strand breaks. J. Cell Biol. 194, 367–375 (2011).

    CAS  PubMed  PubMed Central  Google Scholar 

  173. Caron, M. C. et al. Poly(ADP-ribose) polymerase-1 antagonizes DNA resection at double-strand breaks. Nat. Commun. 10, 2954 (2019).

    PubMed  PubMed Central  Google Scholar 

  174. Lee, K. J. et al. Phosphorylation of Ku dictates DNA double-strand break (DSB) repair pathway choice in S phase. Nucleic Acids Res. 44, 1732–1745 (2016).

    PubMed  Google Scholar 

  175. Kakarougkas, A. & Jeggo, P. A. DNA DSB repair pathway choice: an orchestrated handover mechanism. Br. J. Radiol. 87, 20130685 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  176. Gigi, V. et al. RAG2 mutants alter DSB repair pathway choice in vivo and illuminate the nature of ‘alternative NHEJ’. Nucleic Acids Res. 42, 6352–6364 (2014).

    CAS  PubMed  PubMed Central  Google Scholar 

  177. Delacote, F. & Lopez, B. S. Importance of the cell cycle phase for the choice of the appropriate DSB repair pathway, for genome stability maintenance: the trans-S double-strand break repair model. Cell Cycle 7, 33–38 (2008).

    CAS  PubMed  Google Scholar 

  178. Pellegrino, S., Michelena, J., Teloni, F., Imhof, R. & Altmeyer, M. Replication-coupled dilution of H4K20me2 guides 53BP1 to pre-replicative chromatin. Cell Rep. 19, 1819–1831 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  179. Ochs, F. et al. Stabilization of chromatin topology safeguards genome integrity. Nature 574, 571–574 (2019).

    CAS  PubMed  Google Scholar 

  180. Spies, J. et al. 53BP1 nuclear bodies enforce replication timing at under-replicated DNA to limit heritable DNA damage. Nat. Cell Biol. 21, 487–497 (2019).

    CAS  PubMed  Google Scholar 

  181. Hu, J. et al. Chromosomal loop domains direct the recombination of antigen receptor genes. Cell 163, 947–959 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  182. Lin, S. G., Ba, Z., Alt, F. W. & Zhang, Y. RAG chromatin scanning during V(D)J recombination and chromatin loop extrusion are related processes. Adv. Immunol. 139, 93–135 (2018).

    CAS  PubMed  Google Scholar 

  183. Zhang, Y. et al. The fundamental role of chromatin loop extrusion in physiological V(D)J recombination. Nature 573, 600–604 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  184. Zhang, X. et al. Fundamental roles of chromatin loop extrusion in antibody class switching. Nature 575, 385–389 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  185. Lottersberger, F., Karssemeijer, R. A., Dimitrova, N. & de Lange, T. 53BP1 and the LINC complex promote microtubule-dependent DSB mobility and DNA repair. Cell 163, 880–893 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  186. Schrank, B. R. et al. Nuclear ARP2/3 drives DNA break clustering for homology-directed repair. Nature 559, 61–66 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  187. Dimitrova, N., Chen, Y. C., Spector, D. L. & de Lange, T. 53BP1 promotes non-homologous end joining of telomeres by increasing chromatin mobility. Nature 456, 524–528 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  188. Raschellà, G., Melino, G. & Malewicz, M. New factors in mammalian DNA repair—the chromatin connection. Oncogene 36, 4673–4681 (2017).

    PubMed  PubMed Central  Google Scholar 

  189. Mirman, Z. & de Lange, T. 53BP1: a DSB escort. Genes Dev. 34, 7–23 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  190. Dutertre, M. & Vagner, S. DNA-damage response RNA-binding proteins (DDRBPs): perspectives from a new class of proteins and their RNA targets. J. Mol. Biol. 429, 3139–3145 (2017).

    CAS  PubMed  Google Scholar 

  191. D’Alessandro, G. & d’Adda di Fagagna, F. Transcription and DNA damage: holding hands or crossing swords? J. Mol. Biol. 429, 3215–3229 (2017).

    PubMed  Google Scholar 

  192. D’Alessandro, G. et al. BRCA2 controls DNA:RNA hybrid level at DSBs by mediating RNase H2 recruitment. Nat. Commun. 9, 5376 (2018).

    PubMed  PubMed Central  Google Scholar 

  193. Chakraborty, A. et al. Classical non-homologous end-joining pathway utilizes nascent RNA for error-free double-strand break repair of transcribed genes. Nat. Commun. 7, 13049 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  194. Zhang, Y. et al. Long noncoding RNA LINP1 regulates repair of DNA double-strand breaks in triple-negative breast cancer. Nat. Struct. Mol. Biol. 23, 522–530 (2016).

    CAS  PubMed  PubMed Central  Google Scholar 

  195. Michelini, F. et al. Damage-induced lncRNAs control the DNA damage response through interaction with DDRNAs at individual double-strand breaks. Nat. Cell Biol. 19, 1400–1411 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  196. Pessina, F. et al. Functional transcription promoters at DNA double-strand breaks mediate RNA-driven phase separation of damage-response factors. Nat. Cell Biol. 21, 1286–1299 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  197. Jaafar, L., Li, Z., Li, S. & Dynan, W. S. SFPQ*NONO and XLF function separately and together to promote DNA double-strand break repair via canonical nonhomologous end joining. Nucleic Acids Res. 45, 1848–1859 (2017).

    CAS  PubMed  Google Scholar 

  198. Li, S. et al. Cell-type specific role of the RNA-binding protein, NONO, in the DNA double-strand break response in the mouse testes. DNA Repair 51, 70–78 (2017).

    CAS  PubMed  PubMed Central  Google Scholar 

  199. Nasmyth, K. & Haering, C. H. Cohesin: its roles and mechanisms. Annu. Rev. Genet. 43, 525–558 (2009).

    CAS  PubMed  Google Scholar 

  200. Uhlmann, F. SMC complexes: from DNA to chromosomes. Nat. Rev. Mol. Cell Biol. 17, 399–412 (2016).

    CAS  PubMed  Google Scholar 

  201. Bauerschmidt, C. et al. Cohesin promotes the repair of ionizing radiation-induced DNA double-strand breaks in replicated chromatin. Nucleic Acids Res. 38, 477–487 (2009).

    PubMed  PubMed Central  Google Scholar 

  202. Kim, J.-S., Krasieva, T. B., LaMorte, V., Taylor, A. M. R. & Yokomori, K. Specific recruitment of human cohesin to laser-induced DNA damage. J. Biol. Chem. 277, 45149–45153 (2002).

    CAS  PubMed  Google Scholar 

  203. Kong, X. et al. Distinct functions of human cohesin-SA1 and cohesin-SA2 in double-strand break repair. Mol. Cell. Biol. 34, 685–698 (2014).

    PubMed  PubMed Central  Google Scholar 

  204. Caron, P. et al. Cohesin protects genes against γH2AX induced by DNA double-strand breaks. PLoS Genet. 8, e1002460 (2012).

    CAS  PubMed  PubMed Central  Google Scholar 

  205. Schär, P., Fäsi, M. & Jessberger, R. SMC1 coordinates DNA double-strand break repair pathways. Nucleic Acids Res. 32, 3921–3929 (2004).

    PubMed  PubMed Central  Google Scholar 

  206. Potts, P. R., Porteus, M. H. & Yu, H. Human SMC5/6 complex promotes sister chromatid homologous recombination by recruiting the SMC1/3 cohesin complex to double-strand breaks. EMBO J. 25, 3377–3388 (2006).

    CAS  PubMed  PubMed Central  Google Scholar 

  207. Enervald, E. et al. A regulatory role for the cohesin loader NIPBL in nonhomologous end joining during immunoglobulin class switch recombination. J. Exp. Med. 210, 2503–2513 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  208. Thomas-Claudepierre, A.-S. et al. The cohesin complex regulates immunoglobulin class switch recombination. J. Exp. Med. 210, 2495–2502 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  209. Gelot, C. et al. The cohesin complex prevents the end joining of distant DNA double-strand ends. Mol. Cell 61, 15–26 (2016).

    CAS  PubMed  Google Scholar 

  210. Davidson, I. F. et al. DNA loop extrusion by human cohesin. Science 366, 1338–1345 (2019).

    CAS  PubMed  Google Scholar 

  211. Kim, Y., Shi, Z., Zhang, H., Finkelstein, I. J. & Yu, H. Human cohesin compacts DNA by loop extrusion. Science 366, 1345–1349 (2019).

    CAS  PubMed  PubMed Central  Google Scholar 

  212. Ba, Z. et al. CTCF orchestrates long-range cohesin-driven V(D)J recombinational scanning. Nature https://doi.org/10.1038/s41586-41020-42578-41580 (2020).

    Article  PubMed  PubMed Central  Google Scholar 

  213. Zhu, C., Bogue, M. A., Lim, D.-S., Hasty, P. & Roth, D. B. Ku86-deficient mice exhibit severe combined immunodeficiency and defective processing of V(D)J recombination intermediates. Cell 86, 379–389 (1996).

    CAS  PubMed  Google Scholar 

  214. Taccioli, G. E. et al. Targeted disruption of the catalytic subunit of the DNA-PK gene in mice confers severe combined immunodeficiency and radiosensitivity. Immunity 9, 355–366 (1998).

    CAS  PubMed  Google Scholar 

  215. Rooney, S. et al. Leaky scid phenotype associated with defective V(D)J coding end processing in Artemis-deficient mice. Mol. Cell 10, 1379–1390 (2002).

    CAS  PubMed  Google Scholar 

  216. Frank, K. M. et al. Late embryonic lethality and impaired V (D)J recombination in mice lacking DNA ligase IV. Nature 396, 173–177 (1998).

    CAS  PubMed  Google Scholar 

  217. Altmann, T. & Gennery, A. R. DNA ligase IV syndrome; a review. Orphanet J. Rare Dis. 11, 137–137 (2016).

    PubMed  PubMed Central  Google Scholar 

  218. Saito, S., Kurosawa, A. & Adachi, N. Mutations in XRCC4 cause primordial dwarfism without causing immunodeficiency. J. Hum. Genet. 61, 679–685 (2016).

    CAS  PubMed  Google Scholar 

  219. Woodbine, L., Gennery, A. R. & Jeggo, P. A. The clinical impact of deficiency in DNA non-homologous end-joining. DNA Repair 16, 84–96 (2014).

    CAS  PubMed  Google Scholar 

  220. Li, G. et al. Lymphocyte-specific compensation for XLF/Cernunnos end-joining functions in V(D)J recombination. Mol. Cell 31, 631–640 (2008).

    CAS  PubMed  PubMed Central  Google Scholar 

  221. Grawunder, U., Zimmer, D. & Lieber, M. R. DNA ligase IV binds to XRCC4 via a motif located between rather than within its BRCT domains. Curr. Biol. 8, 873–876 (1998).

    CAS  PubMed  Google Scholar 

  222. Gao, Y. et al. A critical role for DNA end-joining proteins in both lymphogenesis and neurogenesis. Cell 95, 891–902 (1998).

    CAS  PubMed  Google Scholar 

  223. Bee, L. et al. A nonsense mutation of human XRCC4 is associated with adult-onset progressive encephalocardiomyopathy. EMBO Mol. Med. 7, 918–929 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  224. de Villartay, J.-P. When natural mutants do not fit our expectations: the intriguing case of patients with XRCC4 mutations revealed by whole-exome sequencing. EMBO Mol. Med. 7, 862–864 (2015).

    PubMed  PubMed Central  Google Scholar 

  225. Cui, X. et al. Both CpG Methylation and AID are required for the fragility of the human Bcl-2 major breakpoint region: implications for the timing of the breaks in the t(14;18). Mol. Cell Biol. 33, 947–957 (2013).

    CAS  PubMed  PubMed Central  Google Scholar 

  226. Daley, J. M., Laan, R. L., Suresh, A. & Wilson, T. E. DNA joint dependence of pol X family polymerase action in nonhomologous end joining. J. Biol. Chem. 280, 29030–29037 (2005).

    CAS  PubMed  Google Scholar 

  227. Meek, K. Activation of DNA-PK by hairpinned DNA ends reveals a stepwise mechanism of kinase activation. Nucleic Acids Res. 48, 9098–9108 (2020).

    PubMed  PubMed Central  Google Scholar 

  228. Bétermier, M., Bertrand, P. & Lopez, B. S. Is non-homologous end-joining really an inherently error-prone process? PLoS Genet. 10, e1004086 (2014).

    PubMed  PubMed Central  Google Scholar 

  229. Bhargava, R. et al. C-NHEJ without indels is robust and requires synergistic function of distinct XLF domains. Nat. Commun. 9, 2484 (2018).

    PubMed  PubMed Central  Google Scholar 

  230. Felgentreff, K. et al. Functional analysis of naturally occurring DCLRE1C mutations and correlation with the clinical phenotype of ARTEMIS deficiency. J. Allergy Clin. Immunol. 136, 140–150.e7 (2015).

    CAS  PubMed  PubMed Central  Google Scholar 

  231. Ru, H., Zhang, P. & Wu, H. Structural gymnastics of RAG-mediated DNA cleavage in V(D)J recombination. Curr. Opin. Struct. Biol. 53, 178–186 (2018).

    CAS  PubMed  PubMed Central  Google Scholar 

  232. Chen, X. et al. Cutting antiparallel DNA strands in a single active site. Nat. Struct. Mol. Biol. 27, 119–126 (2020).

    CAS  PubMed  PubMed Central  Google Scholar 

  233. Corneo, B. et al. Rag mutations reveal robust alternative end joining. Nature 449, 483–486 (2007).

    CAS  PubMed  Google Scholar 

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Acknowledgements

This work was supported by US National Institutes of Health grants (GM118009, CA196671, CA100504 and P30 CA014089 to M.R.L.; CA097096 and CA84442 to D.A.R; and R01 GM108119 to E.R.), American Cancer Society grant 130304-RSG-16-241-01-DMC (E.R.), and V Foundation for Cancer Research grant D2018-020 (E.R.).

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Authors and Affiliations

  1. Department of Pathology, University of Southern California Keck School of Medicine, Los Angeles, CA, USA

    Bailin Zhao & Michael R. Lieber

  2. Department of Biochemistry & Molecular Medicine, University of Southern California Keck School of Medicine, Los Angeles, CA, USA

    Bailin Zhao & Michael R. Lieber

  3. Department of Molecular Microbiology & Immunology, University of Southern California Keck School of Medicine, Los Angeles, CA, USA

    Bailin Zhao & Michael R. Lieber

  4. USC Norris Comprehensive Cancer Center, University of Southern California Keck School of Medicine, Los Angeles, CA, USA

    Bailin Zhao & Michael R. Lieber

  5. Molecular and Computational Biology Section, Department of Biological Sciences, Dana and David Dornsife College of Letters, Arts and Sciences, University of Southern California, Los Angeles, CA, USA

    Bailin Zhao & Michael R. Lieber

  6. Department of Biochemistry and Molecular Pharmacology, New York University School of Medicine, New York, NY, USA

    Eli Rothenberg

  7. Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC, USA

    Dale A. Ramsden

Authors
  1. Bailin Zhao
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  2. Eli Rothenberg
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  3. Dale A. Ramsden
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  4. Michael R. Lieber
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Contributions

The authors contributed equally to all aspects of the article.

Corresponding authors

Correspondence to Dale A. Ramsden or Michael R. Lieber.

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The authors declare no competing interests.

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Nature Reviews Molecular Cell Biology thanks Sathees Raghavan, Dongyi Xu and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Supplementary information

Glossary

Homologous recombination

(HR). An important DNA double-strand break repair mechanism, which usually requires long homologous sequences.

Class switch recombination

(CSR). Recombination of the immunoglobulin heavy chain locus, which results in a switch of the expressed heavy chain isotype from IgM to IgA, IgE or IgG.

Microhomology

Short stretches of base pairs of complementarity between two broken DNA ends.

Synaptic complex

The complex formed by the two juxtaposed DNA ends of a double-strand break and related non-homologous end joining proteins.

X polymerases

A subtype of DNA polymerases that includes terminal deoxynucleotidyl transferase, polymerase-µ (Polµ), Polλ and Polβ.

N nucleotides

In V(D)J recombination, nucleotides added by the polymerase terminal deoxynucleotidyl transferase to the ends of coding DNA segments independently of a template.

Inverted repeats

Nucleotides that are added at a DNA double-strand break repair junction and are sometimes copied inversely from either of the two broken DNA ends.

T nucleotides

Nucleotides added at a DNA double-strand break repair junction, which are sometimes copied (directly or inversely) from either strand of either of the two broken DNA ends in a template-dependent manner.

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Zhao, B., Rothenberg, E., Ramsden, D.A. et al. The molecular basis and disease relevance of non-homologous DNA end joining. Nat Rev Mol Cell Biol 21, 765–781 (2020). https://doi.org/10.1038/s41580-020-00297-8

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