Cellular pathways that repair chromosomal double-strand breaks (DSBs) have pivotal roles in cell growth, development and cancer. These DSB repair pathways have been the target of intensive investigation, but one pathway — alternative end joining (a-EJ) — has long resisted elucidation. In this Review, we highlight recent progress in our understanding of a-EJ, especially the assignment of DNA polymerase theta (Polθ) as the predominant mediator of a-EJ in most eukaryotes, and discuss a potential molecular mechanism by which Polθ-mediated end joining (TMEJ) occurs. We address possible cellular functions of TMEJ in resolving DSBs that are refractory to repair by non-homologous end joining (NHEJ), DSBs generated following replication fork collapse and DSBs present owing to stalling of repair by homologous recombination. We also discuss how these context-dependent cellular roles explain how TMEJ can both protect against and cause genome instability, and the emerging potential of Polθ as a therapeutic target in cancer.
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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).
Boulton, S. J. & Jackson, S. P. Saccharomyces cerevisiae Ku70 potentiates illegitimate DNA double-strand break repair and serves as a barrier to error-prone DNA repair pathways. EMBO J. 15, 5093–5103 (1996).
Kabotyanski, E. B., Gomelsky, L., Han, J.-O., Roth, D. B. & Stamato, T. D. Double-strand break repair in Ku86-and XRCC4-deficient cells. Nucleic acids Res. 26, 5333–5342 (1998).
Liang, F. & Jasin, M. Ku80-deficient cells exhibit excess degradation of extrachromosomal DNA. J. Biol. Chem. 271, 14405–14411 (1996).
Bothmer, A. et al. 53BP1 regulates DNA resection and the choice between classical and alternative end joining during class switch recombination. J. Exp. Med. 207, 855–865 (2010).
Deriano, L., Stracker, T. H., Baker, A., Petrini, J. H. & Roth, D. B. Roles for NBS1 in alternative nonhomologous end-joining of V(D)J recombination intermediates. Mol. Cell 34, 13–25 (2009).
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).
Ma, J. L., Kim, E. M., Haber, J. E. & Lee, S. E. Yeast Mre11 and Rad1 proteins define a Ku-independent mechanism to repair double-strand breaks lacking overlapping end sequences. Mol. Cell Biol. 23, 8820–8828 (2003).
Rahal, E. A. et al. ATM regulates Mre11-dependent DNA end-degradation and microhomology-mediated end joining. Cell Cycle 9, 2866–2877 (2010).
Truong, L. N. et al. Microhomology-mediated end joining and homologous recombination share the initial end resection step to repair DNA double-strand breaks in mammalian cells. Proc. Natl Acad. Sci. USA 110, 7720–7725 (2013).
Xie, A., Kwok, A. & Scully, R. Role of mammalian Mre11 in classical and alternative nonhomologous end joining. Nat. Struct. Mol. Biol. 16, 814–818 (2009).
Yun, M. H. & Hiom, K. CtIP-BRCA1 modulates the choice of DNA double-strand-break repair pathway throughout the cell cycle. Nature 459, 460–463 (2009).
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).
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).
Beall, E. L. & Rio, D. C. Drosophila P-element transposase is a novel site-specific endonuclease. Genes Dev. 11, 2137–2151 (1997).
Foster, S. S., Balestrini, A. & Petrini, J. H. Functional interplay of the Mre11 nuclease and Ku in the response to replication-associated DNA damage. Mol. Cell Biol. 31, 4379–4389 (2011).
Wyatt, D. W. et al. Essential roles for polymerase theta-mediated end joining in the repair of chromosome breaks. Mol. Cell 63, 662–673 (2016).
Yousefzadeh, M. J. et al. Mechanism of suppression of chromosomal instability by DNA polymerase POLQ. PLoS Genet. 10, e1004654 (2014).
Thyme, S. B. & Schier, A. F. Polq-mediated end joining is essential for surviving DNA double-strand breaks during early zebrafish development. Cell Rep. 15, 707–714 (2016).
van Schendel, R., Roerink, S. F., Portegijs, V., van den Heuvel, S. & Tijsterman, M. Polymerase theta is a key driver of genome evolution and of CRISPR/Cas9-mediated mutagenesis. Nat. Commun. 6, 7394 (2015).
Mateos-Gomez, P. A. et al. Mammalian polymerase theta promotes alternative NHEJ and suppresses recombination. Nature 518, 254–257 (2015).
Saito, S., Maeda, R. & Adachi, N. Dual loss of human POLQ and LIG4 abolishes random integration. Nat. Commun. 8, 16112 (2017).
Zelensky, A. N., Schimmel, J., Kool, H., Kanaar, R. & Tijsterman, M. Inactivation of Pol theta and C-NHEJ eliminates off-target integration of exogenous DNA. Nat. Commun. 8, 66 (2017).
Roerink, S. F., van Schendel, R. & Tijsterman, M. Polymerase theta-mediated end joining of replication-associated DNA breaks in C. elegans. Genome Res. 24, 954–962 (2014).
Seki, M., Marini, F. & Wood, R. D. POLQ (Pol theta), a DNA polymerase and DNA-dependent ATPase in human cells. Nucleic Acids Res. 31, 6117–6126 (2003).
Takata, K. I. et al. Analysis of DNA polymerase nu function in meiotic recombination, immunoglobulin class-switching, and DNA damage tolerance. PLoS Genet. 13, e1006818 (2017).
Yousefzadeh, M. J. & Wood, R. D. DNA polymerase POLQ and cellular defense against DNA damage. DNA Repair 12, 1–9 (2013).
Black, S. J. et al. Molecular basis of microhomology-mediated end-joining by purified full-length Poltheta. Nat. Commun. 10, 4423 (2019).
Hogg, M., Seki, M., Wood, R. D., Doublie, S. & Wallace, S. S. Lesion bypass activity of DNA polymerase theta (POLQ) is an intrinsic property of the pol domain and depends on unique sequence inserts. J. Mol. Biol. 405, 642–652 (2011).
Wood, R. D. & Doublie, S. DNA polymerase theta (POLQ), double-strand break repair, and cancer. DNA Repair 44, 22–32 (2016).
Zahn, K. E., Averill, A. M., Aller, P., Wood, R. D. & Doublie, S. Human DNA polymerase theta grasps the primer terminus to mediate DNA repair. Nat. Struct. Mol. Biol. 22, 304–311 (2015).
Zahn, K. E., Jensen, R. B., Wood, R. D. & Doublie, S. Human DNA polymerase theta harbors DNA end-trimming activity critical for DNA repair. Mol. Cell 81, 1534–1547 (2021).
Ceccaldi, R. et al. Homologous-recombination-deficient tumours are dependent on Poltheta-mediated repair. Nature 518, 258–262 (2015).
Newman, J. A., Cooper, C. D. O., Aitkenhead, H. & Gileadi, O. Structure of the helicase domain of DNA polymerase theta reveals a possible role in the microhomology-mediated end-joining pathway. Structure 23, 2319–2330 (2015).
Ozdemir, A. Y., Rusanov, T., Kent, T., Siddique, L. A. & Pomerantz, R. T. Polymerase theta-helicase efficiently unwinds DNA and RNA-DNA hybrids. J. Biol. Chem. 293, 5259–5269 (2018).
Bazzano, D., Lomonaco, S. & Wilson, T. E. Mapping yeast mitotic 5’ resection at base resolution reveals the sequence and positional dependence of nucleases in vivo. Nucleic Acids Res. https://doi.org/10.1093/nar/gkab597 (2021).
Mimitou, E. P. & Symington, L. S. Sae2, Exo1 and Sgs1 collaborate in DNA double-strand break processing. Nature 455, 770–774 (2008).
Zhu, Z., Chung, W. H., Shim, E. Y., Lee, S. E. & Ira, G. Sgs1 helicase and two nucleases Dna2 and Exo1 resect DNA double-strand break ends. Cell 134, 981–994 (2008).
Eccleston, J., Yan, C., Yuan, K., Alt, F. W. & Selsing, E. Mismatch repair proteins MSH2, MLH1, and EXO1 are important for class-switch recombination events occurring in B cells that lack nonhomologous end joining. J. Immunol. 186, 2336–2343 (2011).
Howard, S. M., Yanez, D. A. & Stark, J. M. DNA damage response factors from diverse pathways, including DNA crosslink repair, mediate alternative end joining. PLoS Genet. 11, e1004943 (2015).
Koole, W. et al. A Polymerase theta-dependent repair pathway suppresses extensive genomic instability at endogenous G4 DNA sites. Nat. Commun. 5, 3216 (2014).
Wang, Z. et al. DNA polymerase theta (POLQ) is important for repair of DNA double-strand breaks caused by fork collapse. J. Biol. Chem. 294, 3909–3919 (2019).
Kais, Z. et al. FANCD2 maintains fork stability in BRCA1/2-deficient tumors and promotes alternative end-joining DNA repair. Cell Rep. 15, 2488–2499 (2016).
Audebert, M., Salles, B. & Calsou, P. Involvement of poly(ADP-ribose) polymerase-1 and XRCC1/DNA ligase III in an alternative route for DNA double-strand breaks rejoining. J. Biol. Chem. 279, 55117–55126 (2004).
Mansour, W. Y., Rhein, T. & Dahm-Daphi, J. The alternative end-joining pathway for repair of DNA double-strand breaks requires PARP1 but is not dependent upon microhomologies. Nucleic Acids Res. 38, 6065–6077 (2010).
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).
Caldecott, K. W. XRCC1 protein; form and function. DNA Repair 81, 102664 (2019).
Yu, W. et al. Repair of G1 induced DNA double-strand breaks in S-G2/M by alternative NHEJ. Nat. Commun. 11, 5239 (2020).
Zatreanu, D. et al. Poltheta inhibitors elicit BRCA-gene synthetic lethality and target PARP inhibitor resistance. Nat. Commun. 12, 3636 (2021).
Zhou, J. et al. A first-in-class polymerase theta inhibitor selectively targets homologous-recombination-deficient tumors. Nat. Cancer 2, 598–610 (2021).
Mateos-Gomez, P. A. et al. The helicase domain of Poltheta counteracts RPA to promote alt-NHEJ. Nat. Struct. Mol. Biol. 24, 1116–1123 (2017).
Deng, S. K., Gibb, B., de Almeida, M. J., Greene, E. C. & Symington, L. S. RPA antagonizes microhomology-mediated repair of DNA double-strand breaks. Nat. Struct. Mol. Biol. 21, 405–412 (2014).
Shukla, V. et al. HMCES functions in the alternative end-joining pathway of the DNA DSB repair during class switch recombination in B cells. Mol. Cell 77, 384–394 e384 (2020).
Hussmann, J. A. et al. Mapping the genetic landscape of DNA double-strand break repair. bioRxiv https://doi.org/10.1101/2021.06.14.448344 (2021).
van Schendel, R., Romeijn, R., Buijs, H. & Tijsterman, M. Preservation of lagging strand integrity at sites of stalled replication by Pol alpha-primase and 9-1-1 complex. Sci Adv 7, eabf2278 (2021).
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).
He, P. & Yang, W. Template and primer requirements for DNA Pol theta-mediated end joining. Proc. Natl Acad. Sci. USA 115, 7747–7752 (2018).
Zhang, Y., Davis, L. & Maizels, N. Pathways and signatures of mutagenesis at targeted DNA nicks. PLoS Genet. 17, e1009329 (2021).
Kamp, J. A., van Schendel, R., Dilweg, I. W. & Tijsterman, M. BRCA1-associated structural variations are a consequence of polymerase theta-mediated end-joining. Nat. Commun. 11, 3615 (2020).
van Schendel, R., van Heteren, J., Welten, R. & Tijsterman, M. Genomic scars generated by polymerase theta reveal the versatile mechanism of alternative end-joining. PLoS Genet. 12, e1006368 (2016).
Feng, W. et al. Genetic determinants of cellular addiction to DNA polymerase theta. Nat. Commun. 10, 4286 (2019).
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).
Kosicki, M., Allen, F. & Bradley, A. Cas9-induced large deletions and small indels are controlled in a convergent fashion. bioRxiv https://doi.org/10.1101/2020.08.05.216739 (2020).
Hwang, T. et al. Defining the mutation signatures of DNA polymerase theta in cancer genomes. NAR Cancer 2, zcaa017 (2020).
Khodaverdian, V. Y. et al. Secondary structure forming sequences drive SD-MMEJ repair of DNA double-strand breaks. Nucleic Acids Res. 45, 12848–12861 (2017).
van Kregten, M. et al. T-DNA integration in plants results from polymerase-theta-mediated DNA repair. Nat. Plants 2, 16164 (2016).
Schimmel, J., van Schendel, R., den Dunnen, J. T. & Tijsterman, M. Templated insertions: a smoking gun for polymerase theta-mediated end joining. Trends Genet. 35, 632–644 (2019).
Morton, L. M. et al. Radiation-related genomic profile of papillary thyroid cancer after the Chernobyl accident. Science 372, eabg2538 (2021).
Yu, A. M. & McVey, M. Synthesis-dependent microhomology-mediated end joining accounts for multiple types of repair junctions. Nucleic Acids Res. 38, 5706–5717 (2010).
Kent, T., Chandramouly, G., McDevitt, S. M., Ozdemir, A. Y. & Pomerantz, R. T. Mechanism of microhomology-mediated end-joining promoted by human DNA polymerase theta. Nat. Struct. Mol. Biol. 22, 230–237 (2015).
Seki, M. & Wood, R. D. DNA polymerase theta (POLQ) can extend from mismatches and from bases opposite a (6-4) photoproduct. DNA Repair 7, 119–127 (2008).
Bennardo, N., Cheng, A., Huang, N. & Stark, J. M. Alternative-NHEJ is a mechanistically distinct pathway of mammalian chromosome break repair. PLoS Genet. 4, e1000110 (2008).
Ahmad, A. et al. ERCC1-XPF endonuclease facilitates DNA double-strand break repair. Mol. Cell Biol. 28, 5082–5092 (2008).
Arana, M. E., Seki, M., Wood, R. D., Rogozin, I. B. & Kunkel, T. A. Low-fidelity DNA synthesis by human DNA polymerase theta. Nucleic Acids Res. 36, 3847–3856 (2008).
Osia, B. et al. Cancer cells are uniquely susceptible to accumulation of MMBIR mutations. bioRxiv https://doi.org/10.1101/2020.07.19.209445 (2020).
Layer, J. V. et al. Polymerase delta promotes chromosomal rearrangements and imprecise double-strand break repair. Proc. Natl Acad. Sci. USA 117, 27566–27577 (2020).
Meyer, D., Fu, B. X. & Heyer, W. D. DNA polymerases delta and lambda cooperate in repairing double-strand breaks by microhomology-mediated end-joining in Saccharomyces cerevisiae. Proc. Natl Acad. Sci. USA 112, E6907–E6916 (2015).
Lydeard, J. R., Jain, S., Yamaguchi, M. & Haber, J. E. Break-induced replication and telomerase-independent telomere maintenance require Pol32. Nature 448, 820–823 (2007).
Mengwasser, K. E. et al. Genetic screens reveal FEN1 and APEX2 as BRCA2 synthetic lethal targets. Mol. Cell 73, 885–899 e886 (2019).
Simsek, D. et al. DNA ligase III promotes alternative nonhomologous end-joining during chromosomal translocation formation. PLoS Genet. 7, e1002080 (2011).
Chen, X. et al. Distinct kinetics of human DNA ligases I, IIIalpha, IIIbeta, and IV reveal direct DNA sensing ability and differential physiological functions in DNA repair. DNA Repair 8, 961–968 (2009).
Della-Maria, J. et al. Human Mre11/human Rad50/Nbs1 and DNA ligase IIIalpha/XRCC1 protein complexes act together in an alternative nonhomologous end joining pathway. J. Biol. Chem. 286, 33845–33853 (2011).
Wang, H. et al. DNA ligase III as a candidate component of backup pathways of nonhomologous end joining. Cancer Res. 65, 4020–4030 (2005).
Boboila, C. et al. Robust chromosomal DNA repair via alternative end-joining in the absence of X-ray repair cross-complementing protein 1 (XRCC1). Proc. Natl Acad. Sci. USA 109, 2473–2478 (2012).
Masani, S., Han, L., Meek, K. & Yu, K. Redundant function of DNA ligase 1 and 3 in alternative end-joining during immunoglobulin class switch recombination. Proc. Natl Acad. Sci. USA 113, 1261–1266 (2016).
Harris, P. V. et al. Molecular cloning of Drosophila mus308, a gene involved in DNA cross-link repair with homology to prokaryotic DNA polymerase I genes. Mol. Cell Biol. 16, 5764–5771 (1996).
Muzzini, D. M., Plevani, P., Boulton, S. J., Cassata, G. & Marini, F. Caenorhabditis elegans POLQ-1 and HEL-308 function in two distinct DNA interstrand cross-link repair pathways. DNA Repair 7, 941–950 (2008).
Shima, N., Munroe, R. J. & Schimenti, J. C. The mouse genomic instability mutation chaos1 is an allele of Polq that exhibits genetic interaction with Atm. Mol. Cell Biol. 24, 10381–10389 (2004).
van Bostelen, I. & Tijsterman, M. Combined loss of three DNA damage response pathways renders C. elegans intolerant to light. DNA Repair 54, 55–62 (2017).
Ira, G., Malkova, A., Liberi, G., Foiani, M. & Haber, J. E. Srs2 and Sgs1-Top3 suppress crossovers during double-strand break repair in yeast. Cell 115, 401–411 (2003).
Luo, G. et al. Cancer predisposition caused by elevated mitotic recombination in Bloom mice. Nat. Genet. 26, 424–429 (2000).
Verma, P. & Greenberg, R. A. Noncanonical views of homology-directed DNA repair. Genes Dev. 30, 1138–1154 (2016).
LaFave, M. C. & Sekelsky, J. Mitotic recombination: why? when? how? where? PLoS Genet. 5, e1000411 (2009).
LaRocque, J. R. et al. Interhomolog recombination and loss of heterozygosity in wild-type and Bloom syndrome helicase (BLM)-deficient mammalian cells. Proc. Natl Acad. Sci. USA 108, 11971–11976 (2011).
Wechsler, T., Newman, S. & West, S. C. Aberrant chromosome morphology in human cells defective for Holliday junction resolution. Nature 471, 642–646 (2011).
Carvajal-Garcia, J., Crown, K. N., Ramsden, D. A. & Sekelsky, J. DNA polymerase theta suppresses mitotic crossing over. PLoS Genet. 17, e1009267 (2021).
Davis, L., Khoo, K. J., Zhang, Y. & Maizels, N. POLQ suppresses interhomolog recombination and loss of heterozygosity at targeted DNA breaks. Proc. Natl Acad. Sci. USA 117, 22900–22909 (2020).
Chandramouly, G. et al. Poltheta promotes the repair of 5’-DNA-protein crosslinks by microhomology-mediated end-joining. Cell Rep. 34, 108820 (2021).
Lemmens, B., van Schendel, R. & Tijsterman, M. Mutagenic consequences of a single G-quadruplex demonstrate mitotic inheritance of DNA replication fork barriers. Nat. Commun. 6, 8909 (2015).
Yoon, J. H. et al. Error-prone replication through UV lesions by DNA polymerase theta protects against skin cancers. Cell 176, 1295–1309 e1215 (2019).
Roy, S. et al. p53 orchestrates DNA replication restart homeostasis by suppressing mutagenic RAD52 and POLtheta pathways. Elife 7, e31723 (2018).
Deng, L. et al. Mitotic CDK promotes replisome disassembly, fork breakage, and complex DNA rearrangements. Mol. Cell 73, 915–929 e916 (2019).
Leibowitz, M. L., Zhang, C. Z. & Pellman, D. Chromothripsis: a new mechanism for rapid karyotype evolution. Annu. Rev. Genet. 49, 183–211 (2015).
Minocherhomji, S. et al. Replication stress activates DNA repair synthesis in mitosis. Nature 528, 286–290 (2015).
Corneo, B. et al. Rag mutations reveal robust alternative end joining. Nature 449, 483–486 (2007).
Cui, X. & Meek, K. Linking double-stranded DNA breaks to the recombination activating gene complex directs repair to the nonhomologous end-joining pathway. Proc. Natl Acad. Sci. USA 104, 17046–17051 (2007).
Li, Y., Gao, X. & Wang, J. Y. Comparison of two POLQ mutants reveals that a polymerase-inactive POLQ retains significant function in tolerance to etoposide and gamma-irradiation in mouse B cells. Genes. Cell 16, 973–983 (2011).
Martomo, S. A., Saribasak, H., Yokoi, M., Hanaoka, F. & Gearhart, P. J. Reevaluation of the role of DNA polymerase theta in somatic hypermutation of immunoglobulin genes. DNA Repair 7, 1603–1608 (2008).
Bosma, G. C. et al. DNA-dependent protein kinase activity is not required for immunoglobulin class switching. J. Exp. Med. 196, 1483–1495 (2002).
Manis, J. P., Dudley, D., Kaylor, L. & Alt, F. W. IgH class switch recombination to IgG1 in DNA-PKcs-deficient B cells. Immunity 16, 607–617 (2002).
Yan, C. T. et al. IgH class switching and translocations use a robust non-classical end-joining pathway. Nature 449, 478–482 (2007).
Han, L. & Yu, K. Altered kinetics of nonhomologous end joining and class switch recombination in ligase IV-deficient B cells. J. Exp. Med. 205, 2745–2753 (2008).
Kumar, R. J. et al. Dual inhibition of DNA-PK and DNA polymerase theta overcomes radiation resistance induced by p53 deficiency. NAR. Cancer 2, zcaa038 (2020).
Panier, S. & Boulton, S. J. Double-strand break repair: 53BP1 comes into focus. Nat. Rev. Mol. Cell Biol. 15, 7–18 (2014).
Isono, M. et al. BRCA1 directs the repair pathway to homologous recombination by promoting 53BP1 dephosphorylation. Cell Rep. 18, 520–532 (2017).
Bouwman, P. et al. 53BP1 loss rescues BRCA1 deficiency and is associated with triple-negative and BRCA-mutated breast cancers. Nat. Struct. Mol. Biol. 17, 688–695 (2010).
Bunting, S. F. et al. 53BP1 inhibits homologous recombination in Brca1-deficient cells by blocking resection of DNA breaks. Cell 141, 243–254 (2010).
Nacson, J. et al. BRCA1 mutation-specific responses to 53BP1 loss-induced homologous recombination and PARP inhibitor resistance. Cell Rep. 24, 3513–3527 e3517 (2018).
Setiaputra, D. & Durocher, D. Shieldin - the protector of DNA ends. EMBO Rep. 20, e47560 (2019).
Clouaire, T. et al. Comprehensive mapping of histone modifications at DNA double-strand breaks deciphers repair pathway chromatin signatures. Mol. Cell 72, 250–262 e256 (2018).
Price, B. D. & D’Andrea, A. D. Chromatin remodeling at DNA double-strand breaks. Cell 152, 1344–1354 (2013).
Schep, R. et al. Impact of chromatin context on Cas9-induced DNA double-strand break repair pathway balance. Mol. Cell 81, 2216–2230 (2021).
Liu, Q. et al. Subjugation of TGFbeta signaling by human papilloma virus in head and neck squamous cell carcinoma shifts DNA repair from homologous recombination to alternative end joining. Clin. Cancer Res. 24, 6001–6014 (2018).
Liu, Q. et al. Loss of TGFbeta signaling increases alternative end-joining DNA repair that sensitizes to genotoxic therapies across cancer types. Sci. Transl. Med. 13, eabc4465 (2021).
Leeman, J. E. et al. Human papillomavirus 16 promotes microhomology-mediated end-joining. Proc. Natl Acad. Sci. USA 116, 21573–21579 (2019).
Knijnenburg, T. A. et al. Genomic and molecular landscape of DNA damage repair deficiency across the cancer genome atlas. Cell Rep. 23, 239–254 e236 (2018).
Lemee, F. et al. DNA polymerase up-regulation is associated with poor survival in breast cancer perturbs DNA replication and promotes genetic instability. Proc. Natl Acad. Sci. USA 107, 13390–13395 (2010).
Higgins, G. S. et al. Overexpression of POLQ confers a poor prognosis in early breast cancer patients. Oncotarget 1, 175–184 (2010).
Allen, F. et al. Predicting the mutations generated by repair of Cas9-induced double-strand breaks. Nat. Biotechnol. 37, 64–72 (2019).
Ata, H. et al. Robust activation of microhomology-mediated end joining for precision gene editing applications. PLoS Genet. 14, e1007652 (2018).
Chen, W. et al. Massively parallel profiling and predictive modeling of the outcomes of CRISPR/Cas9-mediated double-strand break repair. Nucleic Acids Res. 47, 7989–8003 (2019).
Mann, C. M. et al. The Gene Sculpt Suite: a set of tools for genome editing. Nucleic Acids Res. 47, W175–W182 (2019).
Shen, M. W. et al. Predictable and precise template-free CRISPR editing of pathogenic variants. Nature 563, 646–651 (2018).
van Overbeek, M. et al. DNA repair profiling reveals nonrandom outcomes at cas9-mediated breaks. Mol. Cell 63, 633–646 (2016).
Iyer, S. et al. Precise therapeutic gene correction by a simple nuclease-induced double-stranded break. Nature 568, 561–565 (2019).
Alexandrov, L. B. et al. The repertoire of mutational signatures in human cancer. Nature 578, 94–101 (2020).
Alexandrov, L. B. et al. Signatures of mutational processes in human cancer. Nature 500, 415–421 (2013).
Davies, H. et al. HRDetect is a predictor of BRCA1 and BRCA2 deficiency based on mutational signatures. Nat. Med. 23, 517–525 (2017).
Lord, C. J. & Ashworth, A. BRCAness revisited. Nat. Rev. Cancer 16, 110–120 (2016).
Riaz, N. et al. Pan-cancer analysis of bi-allelic alterations in homologous recombination DNA repair genes. Nat. Commun. 8, 857 (2017).
Telli, M. L. et al. Homologous recombination deficiency (HRD) score predicts response to platinum-containing neoadjuvant chemotherapy in patients with triple-negative breast cancer. Clin. Cancer Res. 22, 3764–3773 (2016).
The ICGC/TCGA Pan-Cancer Analysis of Whole Genomes Consortium. Pan-cancer analysis of whole genomes. Nature 578, 82–93 (2020).
Li, Y. et al. Patterns of somatic structural variation in human cancer genomes. Nature 578, 112–121 (2020).
Yang, L. et al. Diverse mechanisms of somatic structural variations in human cancer genomes. Cell 153, 919–929 (2013).
Jones, R. E. et al. Escape from telomere-driven crisis is DNA ligase III dependent. Cell Rep. 8, 1063–1076 (2014).
Nishizawa-Yokoi, A. et al. Agrobacterium T-DNA integration in somatic cells does not require the activity of DNA polymerase theta. New Phytol. 229, 2859–2872 (2021).
Hu, Z. et al. Genome-wide profiling of HPV integration in cervical cancer identifies clustered genomic hot spots and a potential microhomology-mediated integration mechanism. Nat. Genet. 47, 158–163 (2015).
Beagan, K. et al. Drosophila DNA polymerase theta utilizes both helicase-like and polymerase domains during microhomology-mediated end joining and interstrand crosslink repair. PLoS Genet. 13, e1006813 (2017).
Higgins, G. S. et al. A small interfering RNA screen of genes involved in DNA repair identifies tumor-specific radiosensitization by POLQ knockdown. Cancer Res. 70, 2984–2993 (2010).
Russo, M. et al. Adaptive mutability of colorectal cancers in response to targeted therapies. Science 366, 1473–1480 (2019).
Tobalina, L., Armenia, J., Irving, E., O’Connor, M. J. & Forment, J. V. A meta-analysis of reversion mutations in BRCA genes identifies signatures of DNA end-joining repair mechanisms driving therapy resistance. Ann. Oncol. 32, 103–112 (2021).
Waks, A. G. et al. Reversion and non-reversion mechanisms of resistance to PARP inhibitor or platinum chemotherapy in BRCA1/2-mutant metastatic breast cancer. Ann. Oncol. 31, 590–598 (2020).
Weigelt, B. et al. Diverse BRCA1 and BRCA2 reversion mutations in circulating cell-free DNA of therapy-resistant breast or ovarian cancer. Clin. Cancer Res. 23, 6708–6720 (2017).
Ballhausen, A. et al. The shared frameshift mutation landscape of microsatellite-unstable cancers suggests immunoediting during tumor evolution. Nat. Commun. 11, 4740 (2020).
Litchfield, K. et al. Escape from nonsense-mediated decay associates with anti-tumor immunogenicity. Nat. Commun. 11, 3800 (2020).
Turajlic, S. et al. Insertion-and-deletion-derived tumour-specific neoantigens and the immunogenic phenotype: a pan-cancer analysis. Lancet Oncol. 18, 1009–1021 (2017).
Pryor, J. M. et al. Essential role for polymerase specialization in cellular nonhomologous end joining. Proc. Natl Acad. Sci. USA 112, E4537–E4545 (2020).
Zhao, B., Rothenberg, E., Ramsden, D. A. & Lieber, M. R. The molecular basis and disease relevance of non-homologous DNA end joining. Nat. Rev. Mol. Cell Biol. 21, 765–781 (2020).
Lee, K. et al. Microhomology selection for microhomology mediated end joining in Saccharomyces cerevisiae. Genes 10, 284 (2019).
Villarreal, D. D. et al. Microhomology directs diverse DNA break repair pathways and chromosomal translocations. PLoS Genet. 8, e1003026 (2012).
Decottignies, A. Microhomology-mediated end joining in fission yeast is repressed by pku70 and relies on genes involved in homologous recombination. Genetics 176, 1403–1415 (2007).
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).
Lee, K. & Lee, S. E. Saccharomyces cerevisiae Sae2- and Tel1-dependent single-strand DNA formation at DNA break promotes microhomology-mediated end joining. Genetics 176, 2003–2014 (2007).
Deshpande, R. A. et al. ATP-driven Rad50 conformations regulate DNA tethering, end resection, and ATM checkpoint signaling. EMBO J. 33, 482–500 (2014).
Paull, T. T. & Gellert, M. A mechanistic basis for Mre11-directed DNA joining at microhomologies. Proc. Natl Acad. Sci. USA 97, 6409–6414 (2000).
Williams, R. S. et al. Mre11 dimers coordinate DNA end bridging and nuclease processing in double-strand-break repair. Cell 135, 97–109 (2008).
Feldman, T. et al. Recurrent deletions in clonal hematopoiesis are driven by microhomology-mediated end joining. Nat. Commun. 12, 2455 (2021).
Sakofsky, C. J. & Malkova, A. Break induced replication in eukaryotes: mechanisms, functions, and consequences. Crit. Rev. Biochem. Mol. Biol. 52, 395–413 (2017).
Zhang, F. et al. The DNA replication FoSTeS/MMBIR mechanism can generate genomic, genic and exonic complex rearrangements in humans. Nat. Genet. 41, 849–853 (2009).
Moon, A. F. et al. The X family portrait: structural insights into biological functions of X family polymerases. DNA Repair 6, 1709–1725 (2007).
Abkevich, V. et al. Patterns of genomic loss of heterozygosity predict homologous recombination repair defects in epithelial ovarian cancer. Br. J. Cancer 107, 1776–1782 (2012).
Birkbak, N. J. et al. Telomeric allelic imbalance indicates defective DNA repair and sensitivity to DNA-damaging agents. Cancer Discov. 2, 366–375 (2012).
Popova, T. et al. Ploidy and large-scale genomic instability consistently identify basal-like breast carcinomas with BRCA1/2 inactivation. Cancer Res. 72, 5454–5462 (2012).
Turner, N., Tutt, A. & Ashworth, A. Hallmarks of ‘BRCAness’ in sporadic cancers. Nat. Rev. Cancer 4, 814–819 (2004).
Chopra, N. et al. Homologous recombination DNA repair deficiency and PARP inhibition activity in primary triple negative breast cancer. Nat. Commun. 11, 2662 (2020).
Loibl, S. et al. Survival analysis of carboplatin added to an anthracycline/taxane-based neoadjuvant chemotherapy and HRD score as predictor of response-final results from GeparSixto. Ann. Oncol. 29, 2341–2347 (2018).
Tutt, A. et al. Carboplatin in BRCA1/2-mutated and triple-negative breast cancer BRCAness subgroups: the TNT trial. Nat. Med. 24, 628–637 (2018).
Zhao, E. Y. et al. Homologous recombination deficiency and platinum-based therapy outcomes in advanced breast cancer. Clin. Cancer Res. 23, 7521–7530 (2017).
Tumiati, M. et al. A functional homologous recombination assay predicts primary chemotherapy response and long-term survival in ovarian cancer patients. Clin. Cancer Res. 24, 4482–4493 (2018).
Ahrabi, S. et al. A role for human homologous recombination factors in suppressing microhomology-mediated end joining. Nucleic Acids Res. 44, 5743–5757 (2016).
This work was supported by US NIH grants CA222092 and CA247773 to D.A.R. and G.P.G.
G.P.G. receives research funding from Breakpoint Therapeutics, which is developing inhibitors of polymerase-θ. D.A.R. has a materials transfer agreement with Artios Pharma, and is using an Artios Pharma compound that inhibits polymerase-θ for research purposes, with no financial compensation.
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- A-family polymerases
One of seven groupings of eukaryotic DNA polymerases, consisting in mammals of polymerase-γ, polymerase-ν and polymerase-θ.
- Presynaptic filament
In homologous recombination, resected DNA ends bound by RAD51; a precursor to synapsis of the ends with a sister chromatid or homologous chromosome.
- Non-allelic HR
Recombination between homologous sequences that are not allelic (for example, between repeat sequences on different chromosomes).
Stable secondary structures of DNA generated by guanine-rich sequences; can impede DNA replication and transcription.
- Genomic scar
A recurring pattern of mutagenesis that can be attributed to a specific cause or DNA repair process.
- Mitotic DNA synthesis
DNA replication stress-induced repair process that involves DNA synthesis during mitosis, possibly involving break-induced replication.
Clustered chromosomal rearrangements observed in cancer that involve shattering of a chromosome (portion) into many fragments, most likely during erroneous mitotic progression, followed by mutagenic rejoining of the fragments.
- Telomere crisis
A stage of telomere erosion that is sufficient to cause chromosome instability and cell death.
‘Transfer DNA’ that is transferred from the plasmid genome of some tumour-inducing bacteria into the genome of a plant host.
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Ramsden, D.A., Carvajal-Garcia, J. & Gupta, G.P. Mechanism, cellular functions and cancer roles of polymerase-theta-mediated DNA end joining. Nat Rev Mol Cell Biol 23, 125–140 (2022). https://doi.org/10.1038/s41580-021-00405-2
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