Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Review Article
  • Published:

Revolutionizing DNA repair research and cancer therapy with CRISPR–Cas screens

Nature Reviews Molecular Cell Biology volume 24, pages 477–494 (2023)Cite this article

Subjects

Abstract

All organisms possess molecular mechanisms that govern DNA repair and associated DNA damage response (DDR) processes. Owing to their relevance to human disease, most notably cancer, these mechanisms have been studied extensively, yet new DNA repair and/or DDR factors and functional interactions between them are still being uncovered. The emergence of CRISPR technologies and CRISPR-based genetic screens has enabled genome-scale analyses of gene–gene and gene–drug interactions, thereby providing new insights into cellular processes in distinct DDR-deficiency genetic backgrounds and conditions. In this Review, we discuss the mechanistic basis of CRISPR–Cas genetic screening approaches and describe how they have contributed to our understanding of DNA repair and DDR pathways. We discuss how DNA repair pathways are regulated, and identify and characterize crosstalk between them. We also highlight the impacts of CRISPR-based studies in identifying novel strategies for cancer therapy, and in understanding, overcoming and even exploiting cancer-drug resistance, for example in the contexts of PARP inhibition, homologous recombination deficiencies and/or replication stress. Lastly, we present the DDR CRISPR screen (DDRcs) portal, in which we have collected and reanalysed data from CRISPR screen studies and provide a tool for systematically exploring them.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Fig. 1: Overview of CRISPR–Cas strategies and tools.
Fig. 2: High-throughput CRISPR–Cas9 screening approaches.
Fig. 3: Overview of novel DNA repair factors and associations discovered using CRISPR screens.

Similar content being viewed by others

References

  1. Friedberg, E. C. A brief history of the DNA repair field. Cell Res. 18, 3–7 (2008).

    Article  CAS  PubMed  Google Scholar 

  2. Lindahl, T. Instability and decay of the primary structure of DNA. Nature 362, 709–715 (1993).

    Article  CAS  PubMed  Google Scholar 

  3. Lindahl, T. & Nyberg, B. Rate of depurination of native deoxyribonucleic acid. Biochemistry 11, 3610–3618 (1972).

    Article  CAS  PubMed  Google Scholar 

  4. Lindahl, T. & Barnes, D. E. Repair of endogenous DNA damage. Cold Spring Harb. Symp. Quant. Biol. 65, 127–134 (2000).

    Article  CAS  PubMed  Google Scholar 

  5. Jackson, S. P. & Bartek, J. The DNA-damage response in human biology and disease. Nature 461, 1071–1078 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Doudna, J. A. & Charpentier, E. The new frontier of genome engineering with CRISPR–Cas9. Science 346, 1258096 (2014).

    Article  PubMed  Google Scholar 

  7. Barrangou, R. et al. CRISPR provides acquired resistance against viruses in prokaryotes. Science 315, 1709–1712 (2007).

    Article  CAS  PubMed  Google Scholar 

  8. Makarova, K. S. et al. Evolution and classification of the CRISPR–Cas systems. Nat. Rev. Microbiol. 9, 467–477 (2011).

    Article  CAS  PubMed  Google Scholar 

  9. Przybyla, L. & Gilbert, L. A. A new era in functional genomics screens. Nat. Rev. Genet. 23, 89–103 (2022).

    Article  CAS  PubMed  Google Scholar 

  10. Baskar, R., Lee, K. A., Yeo, R. & Yeoh, K.-W. Cancer and radiation therapy: current advances and future directions. Int. J. Med. Sci. 9, 193–199 (2012).

    Article  PubMed  PubMed Central  Google Scholar 

  11. Tchounwou, P. B., Dasari, S., Noubissi, F. K., Ray, P. & Kumar, S. Advances in our understanding of the molecular mechanisms of action of cisplatin in cancer therapy. J. Exp. Pharmacol. 13, 303–328 (2021).

    Article  PubMed  PubMed Central  Google Scholar 

  12. Zagnoli-Vieira, G. & Caldecott, K. W. Untangling trapped topoisomerases with tyrosyl-DNA phosphodiesterases. DNA Repair. 94, 102900 (2020).

    Article  CAS  PubMed  Google Scholar 

  13. Hanahan, D. Hallmarks of cancer: new dimensions. Cancer Discov. 12, 31–46 (2022).

    Article  CAS  PubMed  Google Scholar 

  14. Pilié, P. G., Tang, C., Mills, G. B. & Yap, T. A. State-of-the-art strategies for targeting the DNA damage response in cancer. Nat. Rev. Clin. Oncol. 16, 81–104 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  15. Dias, M. P., Moser, S. C., Ganesan, S. & Jonkers, J. Understanding and overcoming resistance to PARP inhibitors in cancer therapy. Nat. Rev. Clin. Oncol. 18, 773–791 (2021).

    Article  PubMed  Google Scholar 

  16. Farmer, H. et al. Targeting the DNA repair defect in BRCA mutant cells as a therapeutic strategy. Nature 434, 917–921 (2005).

    Article  CAS  PubMed  Google Scholar 

  17. Bryant, H. E. et al. Specific killing of BRCA2-deficient tumours with inhibitors of poly(ADP-ribose) polymerase. Nature 434, 913–917 (2005). Together with Farmer et al. (2005), this paper establishes that PARP inhibitors are strikingly and selectively toxic to cells bearing BRCA1 or BRCA2 deficiency, highlighting the potential for their use in HR-deficient cancers.

    Article  CAS  PubMed  Google Scholar 

  18. Lord, C. J. & Ashworth, A. PARP inhibitors: synthetic lethality in the clinic. Science 355, 1152–1158 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Pilger, D., Seymour, L. W. & Jackson, S. P. Interfaces between cellular responses to DNA damage and cancer immunotherapy. Genes Dev. 35, 602–618 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Gourley, C. et al. Moving from poly(ADP-ribose) polymerase inhibition to targeting DNA repair and DNA damage response in cancer therapy. J. Clin. Oncol. 37, 2257–2269 (2019).

    Article  CAS  PubMed  Google Scholar 

  21. Lord, C. J., McDonald, S., Swift, S., Turner, N. C. & Ashworth, A. A high-throughput RNA interference screen for DNA repair determinants of PARP inhibitor sensitivity. DNA Repair. 7, 2010–2019 (2008).

    Article  CAS  PubMed  Google Scholar 

  22. Bartz, S. R. et al. Small interfering RNA screens reveal enhanced cisplatin cytotoxicity in tumor cells having both BRCA network and TP53 disruptions. Mol. Cell. Biol. 26, 9377–9386 (2006).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Smogorzewska, A. et al. A genetic screen identifies FAN1, a Fanconi anemia-associated nuclease necessary for DNA interstrand crosslink repair. Mol. Cell 39, 36–47 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  24. O’Connell, B. C. et al. A genome-wide camptothecin sensitivity screen identifies a mammalian MMS22L–NFKBIL2 complex required for genomic stability. Mol. Cell 40, 645–657 (2010).

    Article  PubMed  PubMed Central  Google Scholar 

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

    Article  CAS  PubMed  Google Scholar 

  26. 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).

    Article  CAS  PubMed  Google Scholar 

  27. Paulsen, R. D. et al. A genome-wide siRNA screen reveals diverse cellular processes and pathways that mediate genome stability. Mol. Cell 35, 228–239 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. López-Saavedra, A. et al. A genome-wide screening uncovers the role of CCAR2 as an antagonist of DNA end resection. Nat. Commun. 7, 12364 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  29. Adamson, B., Smogorzewska, A., Sigoillot, F. D., King, R. W. & Elledge, S. J. A genome-wide homologous recombination screen identifies the RNA-binding protein RBMX as a component of the DNA-damage response. Nat. Cell Biol. 14, 318–328 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Acevedo-Arozena, A. et al. ENU mutagenesis, a way forward to understand gene function. Annu. Rev. Genom. Hum. 9, 49–69 (2008).

    Article  CAS  Google Scholar 

  31. Blomen, V. A. et al. Gene essentiality and synthetic lethality in haploid human cells. Science 350, 1092–1096 (2015).

    Article  CAS  PubMed  Google Scholar 

  32. O’Loughlin, T. A. & Gilbert, L. A. Functional genomics for cancer research: applications in vivo and in vitro. Annu. Rev. Cancer Biol. 3, 345–363 (2019).

    Article  Google Scholar 

  33. Forment, J. V. et al. Genome-wide genetic screening with chemically mutagenized haploid embryonic stem cells. Nat. Chem. Biol. 13, 12–14 (2017).

    Article  CAS  PubMed  Google Scholar 

  34. Hsu, P. D., Lander, E. S. & Zhang, F. Development and applications of CRISPR–Cas9 for genome engineering. Cell 157, 1262–1278 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Shalem, O., Sanjana, N. E. & Zhang, F. High-throughput functional genomics using CRISPR–Cas9. Nat. Rev. Genet. 16, 299–311 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Shalem, O. et al. Genome-scale CRISPR–Cas9 knockout screening in human cells. Science 343, 84–87 (2014).

    Article  CAS  PubMed  Google Scholar 

  37. Wang, T. et al. Identification and characterization of essential genes in the human genome. Science 350, 1096–1101 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Wang, T., Wei, J. J., Sabatini, D. M. & Lander, E. S. Genetic screens in human cells using the CRISPR–Cas9 system. Science 343, 80–84 (2014).

    Article  CAS  PubMed  Google Scholar 

  39. Koike-Yusa, H., Li, Y., Tan, E.-P., del Castillo Velasco-Herrera, M. & Yusa, K. Genome-wide recessive genetic screening in mammalian cells with a lentiviral CRISPR-guide RNA library. Nat. Biotechnol. 32, 267–273 (2014).

    Article  CAS  PubMed  Google Scholar 

  40. Hart, T. et al. High-resolution CRISPR screens reveal fitness genes and genotype-specific cancer liabilities. Cell 163, 1515–1526 (2015).

    Article  CAS  PubMed  Google Scholar 

  41. Gilbert, L. A. et al. CRISPR-mediated modular RNA-guided regulation of transcription in eukaryotes. Cell 154, 442–451 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Gilbert, L. A. et al. Genome-scale CRISPR-mediated control of gene repression and activation. Cell 159, 647–661 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Perez-Pinera, P. et al. RNA-guided gene activation by CRISPR–Cas9-based transcription factors. Nat. Methods 10, 973–976 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  44. Kearns, N. A. et al. Functional annotation of native enhancers with a Cas9–histone demethylase fusion. Nat. Methods 12, 401–403 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Thakore, P. I. et al. Highly specific epigenome editing by CRISPR–Cas9 repressors for silencing of distal regulatory elements. Nat. Methods 12, 1143–1149 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  46. Sanson, K. R. et al. Optimized libraries for CRISPR–Cas9 genetic screens with multiple modalities. Nat. Commun. 9, 5416 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Iyer, V. S. et al. Designing custom CRISPR libraries for hypothesis-driven drug target discovery. Comput. Struct. Biotechnol. J. 18, 2237–2246 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Wong, A. S. L. et al. Multiplexed barcoded CRISPR–Cas9 screening enabled by CombiGEM. Proc. Natl Acad. Sci. USA 113, 2544–2549 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Shen, J. P. et al. Combinatorial CRISPR–Cas9 screens for de novo mapping of genetic interactions. Nat. Methods 14, 573–576 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Najm, F. J. et al. Orthologous CRISPR–Cas9 enzymes for combinatorial genetic screens. Nat. Biotechnol. 36, 179–189 (2018).

    Article  CAS  PubMed  Google Scholar 

  51. Han, K. et al. Synergistic drug combinations for cancer identified in a CRISPR screen for pairwise genetic interactions. Nat. Biotechnol. 35, 463–474 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. DeWeirdt, P. C. et al. Optimization of AsCas12a for combinatorial genetic screens in human cells. Nat. Biotechnol. 39, 94–104 (2021).

    Article  CAS  PubMed  Google Scholar 

  53. Gier, R. A. et al. High-performance CRISPR–Cas12a genome editing for combinatorial genetic screening. Nat. Commun. 11, 3455 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  54. Dede, M., McLaughlin, M., Kim, E. & Hart, T. Multiplex enCas12a screens detect functional buffering among paralogs otherwise masked in monogenic Cas9 knockout screens. Genome Biol. 21, 262 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Carleton, J. B., Berrett, K. C. & Gertz, J. Multiplex enhancer interference reveals collaborative control of gene regulation by estrogen receptor α-bound enhancers. Cell Syst. 5, 333–344.e5 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  56. Gasperini, M. et al. A genome-wide framework for mapping gene regulation via cellular genetic screens. Cell 176, 1516 (2019).

    Article  CAS  PubMed  Google Scholar 

  57. Yeh, C. D., Richardson, C. D. & Corn, J. E. Advances in genome editing through control of DNA repair pathways. Nat. Cell Biol. 21, 1468–1478 (2019).

    Article  CAS  PubMed  Google Scholar 

  58. DeWeirdt, P. C. et al. Genetic screens in isogenic mammalian cell lines without single cell cloning. Nat. Commun. 11, 752 (2020). This study provides a strategy to create isogenic pairs of cells while avoiding single-cell cloning.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  59. Bowden, A. R. et al. Parallel CRISPR–Cas9 screens clarify impacts of p53 on screen performance. eLife 9, e55325 (2020).

    Article  PubMed  PubMed Central  Google Scholar 

  60. Su, D. et al. CRISPR/CAS9-based DNA damage response screens reveal gene–drug interactions. DNA Repair. 87, 102803 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Hussmann, J. A. et al. Mapping the genetic landscape of DNA double-strand break repair. Cell 184, 5653–5669.e25 (2021). In this study, the authors develop a high-throughput screening approach called Repair-seq to map the genetic dependencies of DNA repair outcomes.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  62. Zimmermann, M. et al. CRISPR screens identify genomic ribonucleotides as a source of PARP-trapping lesions. Nature 559, 285–289 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Fugger, K. et al. Targeting the nucleotide salvage factor DNPH1 sensitizes BRCA-deficient cells to PARP inhibitors. Science 372, 156–165 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  64. Guo, J. U., Su, Y., Zhong, C., Ming, G. & Song, H. Hydroxylation of 5-methylcytosine by TET1 promotes active DNA demethylation in the adult brain. Cell 145, 423–434 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  65. Verma, P. et al. ALC1 links chromatin accessibility to PARP inhibitor response in homologous recombination-deficient cells. Nat. Cell Biol. 23, 160–171 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  66. Hewitt, G. et al. Defective ALC1 nucleosome remodeling confers PARPi sensitization and synthetic lethality with HRD. Mol. Cell 81, 767–783.e11 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  67. Juhász, S. et al. The chromatin remodeler ALC1 underlies resistance to PARP inhibitor treatment. Sci. Adv. 6, eabb8626 (2020).

    Article  PubMed  Google Scholar 

  68. Blessing, C. et al. The oncogenic helicase ALC1 regulates PARP inhibitor potency by trapping PARP2 at DNA breaks. Mol. Cell 80, 862–875.e6 (2020).

    Article  CAS  PubMed  Google Scholar 

  69. Ahel, D. et al. Poly(ADP-ribose)-dependent regulation of DNA repair by the chromatin remodeling enzyme ALC1. Science 325, 1240–1243 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  70. He, Y. J. et al. DYNLL1 binds to MRE11 to limit DNA end resection in BRCA1-deficient cells. Nature 563, 522–526 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  71. Barazas, M. et al. The CST complex mediates end protection at double-strand breaks and promotes PARP inhibitor sensitivity in BRCA1-deficient cells. Cell Rep. 23, 2107–2118 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  72. 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).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  73. Noordermeer, S. M. et al. The shieldin complex mediates 53BP1-dependent DNA repair. Nature 560, 117–121 (2018). Together with Dev et al. (2018), this paper demonstrates that loss of the shieldin complex causes PARP1 resistance in BRCA1-null cells.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  PubMed  PubMed Central  Google Scholar 

  75. Markiewicz-Potoczny, M. et al. TRF2-mediated telomere protection is dispensable in pluripotent stem cells. Nature 589, 110–115 (2021).

    Article  CAS  PubMed  Google Scholar 

  76. Wang, C. et al. Genetic vulnerabilities upon inhibition of DNA damage response. Nucleic Acids Res. 49, 8214–8231 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  77. Pinzaru, A. M. et al. Replication stress conferred by POT1 dysfunction promotes telomere relocalization to the nuclear pore. Genes Dev. 34, 1619–1636 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  78. Adam, S. et al. The CIP2A–TOPBP1 axis safeguards chromosome stability and is a synthetic lethal target for BRCA-mutated cancer. Nat. Cancer 2, 1357–1371 (2021).

    Article  CAS  PubMed  Google Scholar 

  79. Álvarez-Quilón, A. et al. Endogenous DNA 3′ blocks are vulnerabilities for BRCA1 and BRCA2 deficiency and are reversed by the APE2 nuclease. Mol. Cell 78, 1152–1165.e8 (2020).

    Article  PubMed  PubMed Central  Google Scholar 

  80. Mengwasser, K. E. et al. Genetic screens reveal FEN1 and APEX2 as BRCA2 synthetic lethal targets. Mol. Cell 73, 885–899.e6 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  81. Ceccaldi, R. et al. Homologous-recombination-deficient tumours are dependent on Pol-mediated repair. Nature 518, 258–262 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  83. Feng, W. et al. Genetic determinants of cellular addiction to DNA polymerase θ. Nat. Commun. 10, 4286 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  84. Ramsden, D. A., Carvajal-Garcia, J. & Gupta, G. P. Mechanism, cellular functions and cancer roles of polymerase-θ-mediated DNA end joining. Nat. Rev. Mol. Cell Biol. 23, 125–140 (2022).

    Article  CAS  PubMed  Google Scholar 

  85. Tang, M. et al. Genome-wide CRISPR screens reveal cyclin C as synthetic survival target of BRCA2. Nucleic Acids Res. 49, 7476–7491 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  86. Hustedt, N. et al. A consensus set of genetic vulnerabilities to ATR inhibition. Open. Biol. 9, 190156 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  87. Wang, C. et al. C17orf53 is identified as a novel gene involved in inter-strand crosslink repair. DNA Repair. 95, 102946 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  88. Huang, J.-W. et al. MCM8IP activates the MCM8-9 helicase to promote DNA synthesis and homologous recombination upon DNA damage. Nat. Commun. 11, 2948 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  89. Tucker, E. J. et al. Meiotic genes in premature ovarian insufficiency: variants in HROB and REC8 as likely genetic causes. Eur. J. Hum. Genet. 30, 219–228 (2022).

    Article  CAS  PubMed  Google Scholar 

  90. Balmus, G. et al. ATM orchestrates the DNA-damage response to counter toxic non-homologous end-joining at broken replication forks. Nat. Commun. 10, 87 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  91. Mazouzi, A., Velimezi, G. & Loizou, J. I. DNA replication stress: causes, resolution and disease. Exp. Cell Res. 329, 85–93 (2014).

    Article  CAS  PubMed  Google Scholar 

  92. Zeman, M. K. & Cimprich, K. A. Causes and consequences of replication stress. Nat. Cell Biol. 16, 2–9 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  93. Blackford, A. N. & Jackson, S. P. ATM, ATR, and DNA-PK: the trinity at the heart of the DNA damage response. Mol. Cell 66, 801–817 (2017).

    Article  CAS  PubMed  Google Scholar 

  94. Cimprich, K. A. & Cortez, D. ATR: an essential regulator of genome integrity. Nat. Rev. Mol. Cell Biol. 9, 616–627 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  95. Lloyd, R. L. et al. Loss of cyclin C or CDK8 provides ATR inhibitor resistance by suppressing transcription-associated replication stress. Nucleic Acids Res. 49, 8665–8683 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  96. Wang, C. et al. Genome-wide CRISPR screens reveal synthetic lethality of RNASEH2 deficiency and ATR inhibition. Oncogene 38, 2451–2463 (2019).

    Article  CAS  PubMed  Google Scholar 

  97. Ruiz, S. et al. A genome-wide CRISPR screen identifies CDC25A as a determinant of sensitivity to ATR inhibitors. Mol. Cell 62, 307–313 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  98. Schleicher, E. M. et al. Dual genome-wide CRISPR knockout and CRISPR activation screens identify mechanisms that regulate the resistance to multiple ATR inhibitors. PLoS Genet. 16, e1009176 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  99. Benslimane, Y. et al. Genome-wide screens reveal that resveratrol induces replicative stress in human cells. Mol. Cell 79, 846–856.e8 (2020).

    Article  CAS  PubMed  Google Scholar 

  100. Wang, R. et al. DNA polymerase ι compensates for Fanconi anemia pathway deficiency by countering DNA replication stress. Proc. Natl Acad. Sci. USA 117, 33436–33445 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  101. Adeyemi, R. O. et al. The Protexin complex counters resection on stalled forks to promote homologous recombination and crosslink repair. Mol. Cell 81, 4440–4456.e7 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  102. Zhao, Y. et al. Applying genome-wide CRISPR to identify known and novel genes and pathways that modulate formaldehyde toxicity. Chemosphere 269, 128701 (2021).

    Article  CAS  PubMed  Google Scholar 

  103. Cai, M.-Y. et al. Cooperation of the ATM and Fanconi anemia/BRCA pathways in double-strand break end resection. Cell Rep. 30, 2402–2415.e5 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  104. Schubert, L. et al. SCAI promotes error-free repair of DNA interstrand crosslinks via the Fanconi anemia pathway. EMBO Rep. 23, e53639 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  105. Maiani, E. et al. AMBRA1 regulates cyclin D to guard S-phase entry and genomic integrity. Nature 592, 799–803 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  106. Chaikovsky, A. C. et al. The AMBRA1 E3 ligase adaptor regulates the stability of cyclin D. Nature 592, 794–798 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  107. Olivieri, M. et al. A genetic map of the response to DNA damage in human cells. Cell 182, 481–496.e21 (2020). This study identifies 890 genes whose loss causes either resistance or sensitivity to various DNA-damaging agents, and builds a genetic map of responses to DNA damage.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  108. van der Weegen, Y. et al. ELOF1 is a transcription-coupled DNA repair factor that directs RNA polymerase II ubiquitylation. Nat. Cell Biol. 23, 595–607 (2021).

    Article  PubMed  PubMed Central  Google Scholar 

  109. Geijer, M. E. et al. Elongation factor ELOF1 drives transcription-coupled repair and prevents genome instability. Nat. Cell Biol. 23, 608–619 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  110. Carnie, C. J. & Jackson, S. P. The ELOF(1)ant in the room of TCR. Nat. Cell Biol. 23, 584–586 (2021).

    Article  CAS  PubMed  Google Scholar 

  111. Liu, X. et al. ERCC6L2 promotes DNA orientation-specific recombination in mammalian cells. Cell Res. 30, 732–744 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  112. Feng, Y. et al. FAM72A antagonizes UNG2 to promote mutagenic repair during antibody maturation. Nature 600, 324–328 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  113. Hundley, F. V. et al. A comprehensive phenotypic CRISPR–Cas9 screen of the ubiquitin pathway uncovers roles of ubiquitin ligases in mitosis. Mol. Cell 81, 1319–1336.e9 (2021).

    Article  CAS  PubMed  Google Scholar 

  114. Sanchez-Burgos, L. et al. Activation of the integrated stress response overcomes multidrug resistance in FBXW7-deficient cells. EMBO Mol. Med. 14, e15855 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  115. Liao, S., Maertens, O., Cichowski, K. & Elledge, S. J. Genetic modifiers of the BRD4-NUT dependency of NUT midline carcinoma uncovers a synergism between BETis and CDK4/6is. Genes Dev. 32, 1188–1200 (2018).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  116. Zhang, Q. et al. The WD40 domain of FBXW7 is a poly(ADP-ribose)-binding domain that mediates the early DNA damage response. Nucleic Acids Res. 47, 4039–4053 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  117. Hanna, R. E. et al. Massively parallel assessment of human variants with base editor screens. Cell 184, 1064–1080.e20 (2021).

    Article  CAS  PubMed  Google Scholar 

  118. Cuella-Martin, R. et al. Functional interrogation of DNA damage response variants with base editing screens. Cell 184, 1081–1097.e19 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  119. Kweon, J. et al. A CRISPR-based base-editing screen for the functional assessment of BRCA1 variants. Oncogene 39, 30–35 (2020).

    Article  CAS  PubMed  Google Scholar 

  120. Huang, C., Li, G., Wu, J., Liang, J. & Wang, X. Identification of pathogenic variants in cancer genes using base editing screens with editing efficiency correction. Genome Biol. 22, 80 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  121. Sangree, A. K. et al. Benchmarking of SpCas9 variants enables deeper base editor screens of BRCA1 and BCL2. Nat. Commun. 13, 1318 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  122. Koblan, L. W. et al. Efficient C•G-to-G•C base editors developed using CRISPRi screens, target-library analysis, and machine learning. Nat. Biotechnol. 39, 1414–1425 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  123. Chen, P. J. et al. Enhanced prime editing systems by manipulating cellular determinants of editing outcomes. Cell 184, 5635–5652.e29 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  124. Fang, P., de Souza, C., Minn, K. & Chien, J. Genome-scale CRISPR knockout screen identifies TIGAR as a modifier of PARP inhibitor sensitivity. Commun. Biol. 2, 335 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  125. Kim, Y. et al. High-throughput functional evaluation of human cancer-associated mutations using base editors. Nat. Biotechnol. 40, 874–884 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  126. Pettitt, S. J. et al. Genome-wide and high-density CRISPR–Cas9 screens identify point mutations in PARP1 causing PARP inhibitor resistance. Nat. Commun. 9, 1849 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  127. Thompson, N. A. et al. Combinatorial CRISPR screen identifies fitness effects of gene paralogues. Nat. Commun. 12, 1302 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  128. Behan, F. M. et al. Prioritization of cancer therapeutic targets using CRISPR–Cas9 screens. Nature 568, 511–516 (2019). This comprehensive study provides a systematic analysis of core-fitness genes and cancer type-specific fitness genes, and the Cancer Dependency Map (DepMap).

    Article  CAS  PubMed  Google Scholar 

  129. Chan, E. M. et al. WRN helicase is a synthetic lethal target in microsatellite unstable cancers. Nature 568, 551–556 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  130. Cui, Y. et al. CRISP-view: a database of functional genetic screens spanning multiple phenotypes. Nucleic Acids Res. 49, D848–D854 (2021).

    Article  CAS  PubMed  Google Scholar 

  131. Feng, X. et al. Genome-wide CRISPR screens using isogenic cells reveal vulnerabilities conferred by loss of tumor suppressors. Sci. Adv. 8, eabm6638 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  132. Tzelepis, K. et al. A CRISPR dropout screen identifies genetic vulnerabilities and therapeutic targets in acute myeloid leukemia. Cell Rep. 17, 1193–1205 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  133. Gallo, D. et al. CCNE1 amplification is synthetic lethal with PKMYT1 kinase inhibition. Nature 604, 749–756 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  134. Fong, P. C. et al. Poly(ADP)-ribose polymerase inhibition: frequent durable responses in BRCA carrier ovarian cancer correlating with platinum-free interval. J. Clin. Oncol. 28, 2512–2519 (2010).

    Article  CAS  PubMed  Google Scholar 

  135. Audeh, M. W. et al. Oral poly(ADP-ribose) polymerase inhibitor olaparib in patients with BRCA1 or BRCA2 mutations and recurrent ovarian cancer: a proof-of-concept trial. Lancet 376, 245–251 (2010).

    Article  CAS  PubMed  Google Scholar 

  136. Edwards, S. L. et al. Resistance to therapy caused by intragenic deletion in BRCA2. Nature 451, 1111–1115 (2008).

    Article  CAS  PubMed  Google Scholar 

  137. Clements, K. E. et al. Identification of regulators of poly-ADP-ribose polymerase inhibitor response through complementary CRISPR knockout and activation screens. Nat. Commun. 11, 6118 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  138. Tchasovnikarova, I. A., Marr, S. K., Damle, M. & Kingston, R. E. TRACE generates fluorescent human reporter cell lines to characterize epigenetic pathways. Mol. Cell 82, 479–491.e7 (2022).

    Article  CAS  PubMed  Google Scholar 

  139. Dixit, A. et al. Perturb-seq: dissecting molecular circuits with scalable single-cell RNA profiling of pooled genetic screens. Cell 167, 1853–1866.e17 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  140. Adamson, B. et al. A multiplexed single-cell CRISPR screening platform enables systematic dissection of the unfolded protein response. Cell 167, 1867–1882.e21 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  141. Norman, T. M. et al. Exploring genetic interaction manifolds constructed from rich single-cell phenotypes. Science 365, 786–793 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  142. Replogle, J. M. et al. Combinatorial single-cell CRISPR screens by direct guide RNA capture and targeted sequencing. Nat. Biotechnol. 38, 954–961 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  143. Pierce, S. E., Granja, J. M. & Greenleaf, W. J. High-throughput single-cell chromatin accessibility CRISPR screens enable unbiased identification of regulatory networks in cancer. Nat. Commun. 12, 2969 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  144. Rubin, A. J. et al. Coupled single-cell CRISPR screening and epigenomic profiling reveals causal gene regulatory networks. Cell 176, 361–376.e17 (2019).

    Article  CAS  PubMed  Google Scholar 

  145. Meyenberg, M., da Silva, J. & Loizou, J. I. Tissue specific DNA repair outcomes shape the landscape of genome editing. Front. Genet. 12, 728520 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  146. Komor, A. C., Kim, Y. B., Packer, M. S., Zuris, J. A. & Liu, D. R. Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage. Nature 533, 420–424 (2016). The authors of this study develop the ‘base editing’ approach to genome editing, which enables the direct, irreversible conversion of a DNA base into another in a programmable manner, without DSB formation or a donor template.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  147. Gaudelli, N. M. et al. Programmable base editing of A•T to G•C in genomic DNA without DNA cleavage. Nature 551, 464–471 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  148. Anzalone, A. V. et al. Search-and-replace genome editing without double-strand breaks or donor DNA. Nature 576, 149–157 (2019).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  149. Konermann, S. et al. Genome-scale transcriptional activation by an engineered CRISPR–Cas9 complex. Nature 517, 583–588 (2015).

    Article  CAS  PubMed  Google Scholar 

  150. Choudhury, S. R., Cui, Y., Lubecka, K., Stefanska, B. & Irudayaraj, J. CRISPR–dCas9 mediated TET1 targeting for selective DNA demethylation at BRCA1 promoter. Oncotarget 7, 46545–46556 (2016).

    Article  PubMed  PubMed Central  Google Scholar 

  151. Olivieri, M. & Durocher, D. Genome-scale chemogenomic CRISPR screens in human cells using the TKOv3 library. Star. Protoc. 2, 100321 (2021).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  152. Chatterjee, N. & Walker, G. C. Mechanisms of DNA damage, repair, and mutagenesis. Environ. Mol. Mutagen. 58, 235–263 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  153. Ciccia, A. & Elledge, S. J. The DNA damage response: making it safe to play with knives. Mol. Cell 40, 179–204 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  154. Hoeijmakers, J. H. J. DNA damage, aging, and cancer. N. Engl. J. Med. 361, 1475–1485 (2009).

    Article  CAS  PubMed  Google Scholar 

  155. Mehta, A. & Haber, J. E. Sources of DNA double-strand breaks and models of recombinational DNA repair. Cold Spring Harb. Perspect. Biol. 6, a016428 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  156. Tubbs, A. & Nussenzweig, A. Endogenous DNA damage as a source of genomic instability in cancer. Cell 168, 644–656 (2017).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  157. Kunkel, T. A. & Erie, D. A. Eukaryotic mismatch repair in relation to DNA replication. Annu. Rev. Genet. 49, 291–313 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  158. Schärer, O. D. Nucleotide excision repair in eukaryotes. Cold Spring Harb. Perspect. Biol. 5, a012609 (2013).

    Article  PubMed  PubMed Central  Google Scholar 

  159. Caldecott, K. W. Single-strand break repair and genetic disease. Nat. Rev. Genet. 9, 619–631 (2008).

    Article  CAS  PubMed  Google Scholar 

  160. Ronson, G. E. et al. PARP1 and PARP2 stabilise replication forks at base excision repair intermediates through Fbh1-dependent Rad51 regulation. Nat. Commun. 9, 746 (2018).

    Article  PubMed  PubMed Central  Google Scholar 

  161. Liu, T. & Huang, J. DNA end resection: facts and mechanisms. Genomics Proteom. Bioinforma. 14, 126–130 (2016).

    Article  Google Scholar 

  162. Johnson, R. D. Sister chromatid gene conversion is a prominent double-strand break repair pathway in mammalian cells. EMBO J. 19, 3398–3407 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  163. Sfeir, A. & Symington, L. S. Microhomology-mediated end joining: a back-up survival mechanism or dedicated pathway? Trends Biochem. Sci. 40, 701–714 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  164. 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).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  165. Ceccaldi, R., Sarangi, P. & D’Andrea, A. D. The Fanconi anaemia pathway: new players and new functions. Nat. Rev. Mol. 17, 337–349 (2016).

    Article  CAS  Google Scholar 

  166. Kottemann, M. C. & Smogorzewska, A. Fanconi anaemia and the repair of Watson and Crick DNA crosslinks. Nature 493, 356–363 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  167. Zhang, H., Xiong, Y. & Chen, J. DNA–protein cross-link repair: what do we know now? Cell Biosci. 10, 3 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  168. Barker, S., Weinfeld, M. & Murray, D. DNA–protein crosslinks: their induction, repair, and biological consequences. Mutat. Res. 589, 111–135 (2005).

    Article  CAS  PubMed  Google Scholar 

  169. Colic, M. et al. Identifying chemogenetic interactions from CRISPR screens with DrugZ. Genome Med. 11, 52 (2019).

    Article  PubMed  PubMed Central  Google Scholar 

  170. Li, W. et al. MAGeCK enables robust identification of essential genes from genome-scale CRISPR/Cas9 knockout screens. Genome Biol. 15, 554 (2014).

    Article  PubMed  PubMed Central  Google Scholar 

  171. Oughtred, R. et al. The BioGRID database: a comprehensive biomedical resource of curated protein, genetic, and chemical interactions. Protein Sci. 30, 187–200 (2021).

    Article  CAS  PubMed  Google Scholar 

  172. Nambiar, T. S., Baudrier, L., Billon, P. & Ciccia, A. CRISPR-based genome editing through the lens of DNA repair. Mol. Cell 82, 348–388 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  173. Zhao, D. et al. Glycosylase base editors enable C-to-A and C-to-G base changes. Nat. Biotechnol. 39, 35–40 (2021).

    Article  CAS  PubMed  Google Scholar 

  174. Kurt, I. C. et al. CRISPR C-to-G base editors for inducing targeted DNA transversions in human cells. Nat. Biotechnol. 39, 41–46 (2021).

    Article  CAS  PubMed  Google Scholar 

  175. Rees, H. A. & Liu, D. R. Publisher correction: base editing: precision chemistry on the genome and transcriptome of living cells. Nat. Rev. Genet. 19, 801 (2018).

    Article  CAS  PubMed  Google Scholar 

  176. Anzalone, A. V., Koblan, L. W. & Liu, D. R. Genome editing with CRISPR–Cas nucleases, base editors, transposases and prime editors. Nat. Biotechnol. 38, 824–844 (2020).

    Article  CAS  PubMed  Google Scholar 

  177. Komor, A. C. et al. Improved base excision repair inhibition and bacteriophage Mu Gam protein yields C:G-to-T:A base editors with higher efficiency and product purity. Sci. Adv. 3, eaao4774 (2017).

    Article  PubMed  PubMed Central  Google Scholar 

  178. Arbab, M. et al. Determinants of base editing outcomes from target library analysis and machine learning. Cell 182, 463–480.e30 (2020).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  179. Ferreira da Silva, J. et al. Prime editing efficiency and fidelity are enhanced in the absence of mismatch repair. Nat. Commun. 13, 760 (2022).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

The authors thank R. Belotserkovskaya and G. D’Alessandro for critical reading of the manuscript, K. Dry for editorial assistance and other members of the S.P.J. laboratory for advice and discussions. The authors apologize to authors whose work they did not cite owing to space constraints. S.W.A. is supported by the Mark Foundation for Cancer Research and was a recipient of an Outstanding Postdoctoral Women Fellowship from the Israeli Council for Higher Education. A.S.-B. and J.C.T. are supported by European Research Council (ERC) Synergy grant no 855741 (DDREAMM). J.C.T. and V.G. were supported by Wellcome Investigator Award (206388/Z/17/Z). The S.P.J. laboratory is supported by Cancer Research UK and the ERC.

Author information

Author notes

  1. These authors contributed equally: Samah W. Awwad, Almudena Serrano-Benitez.

Authors and Affiliations

  1. Cancer Research UK Cambridge Institute, University of Cambridge, Cambridge, UK

    Samah W. Awwad, Almudena Serrano-Benitez, John C. Thomas & Stephen P. Jackson

  2. The Gurdon Institute and Department of Biochemistry, University of Cambridge, Cambridge, UK

    Samah W. Awwad, Almudena Serrano-Benitez, John C. Thomas, Vipul Gupta & Stephen P. Jackson

Authors
  1. Samah W. Awwad
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Almudena Serrano-Benitez
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. John C. Thomas
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Vipul Gupta
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Stephen P. Jackson
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

S.P.J., S.W.A. and A.S.-B. researched data for the article, substantially contributed to discussion of the content, wrote the article and reviewed the manuscript before submission. J.C.T. and V.G. researched data for the article and substantially contributed to discussion of the content.

Corresponding authors

Correspondence to Almudena Serrano-Benitez, John C. Thomas or Stephen P. Jackson.

Ethics declarations

Competing interests

The authors declare no competing interests.

Peer review

Peer review information

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

Additional information

Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Related links

BioGRID ORCS: https://orcs.thebiogrid.org/

ClinVar: https://www.ncbi.nlm.nih.gov/clinvar/

DDR CRISPR screen (DDRcs) portal: https://stevejacksonlab.org/ddrcs

DepMap Portal: https://depmap.org/

Glossary

6-Thioguanine

A nucleotide (guanine) analogue that, when incorporated into DNA, is toxic to mismatch repair (MMR)-proficient cells, which are unable to effectively repair 6-thioguanine-induced lesions.

BRCA1-A complex

A protein complex (BRCC45–ABRAXAS–MERIT40–RAP80–BRCC36) that has been reported to modify double-strand break (DSB) resection dynamics and limit homologous recombination (HR) repair, somehow counteracting ataxia telangiectasia mutated (ATM) function.

Class-switch recombination

(CSR). A DNA recombination process that occurs in B cells to switch between the production of immunoglobulin isotypes.

Clustered regularly interspaced short palindromic repeat

(CRISPR). A form of immunity against viruses in prokaryotes, comprising genomic loci of short repeats interspersed with DNA sequences of viral origin called ‘spacers’, and the CRISPR-associated (Cas) family of nucleases.

Dropout hits

Genes that, when depleted or lost, affect a specific condition measured in a screen.

Dual-guide

A construct including two single guide RNA (sgRNA) sequences targeting two regions of the same gene.

Homologous recombination

(HR). A conserved type of DNA repair process that relies on use of an extensively homologous template from a sister chromatid, another homologous sequence elsewhere in the genome or an experimentally delivered DNA molecule.

Non-homologous end joining

(NHEJ). The primary pathway in higher eukaryotes that repairs double-strand breaks (DSBs) by directly tethering the break ends without use of a homologous template, and after potential modification of the ends, ligating them in a way that often introduces mutations.

Non-productive HR intermediates

Aberrant intermediates observed in response to unsuccessful homologous recombination (HR) (for example, in POLQ/53BP1 double-knockout cells), which are often associated with larger than usual RecA-like protein (RAD51) foci.

Polθ-mediated end joining

(TMEJ). A double-strand break (DSB) repair mechanism that entails exposure of microhomology sequences internal to the DSB ends before ligation, leading to deletion of the sequence flanking the DSB; sometimes associated with chromosomal rearrangements.

Sanitizer of cellular nucleotide pools

An enzyme involved in preventing the incorporation of aberrant nucleotides into genomic DNA.

Saturation genome editing

Clustered regularly interspaced short palindromic repeat (CRISPR)–Cas9 genome editing aimed at introducing all possible single-nucleotide variants into a targeted genomic region.

Single guide RNAs

(sgRNAs). Artificial fusions of the clustered regularly interspaced short palindromic repeat (CRISPR) RNA, which recognizes the target sequence in DNA, and the scaffold trans-activating CRISPR RNA, which includes secondary structures crucial for its loading onto Cas9.

Rights and permissions

Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Awwad, S.W., Serrano-Benitez, A., Thomas, J.C. et al. Revolutionizing DNA repair research and cancer therapy with CRISPR–Cas screens. Nat Rev Mol Cell Biol 24, 477–494 (2023). https://doi.org/10.1038/s41580-022-00571-x

Download citation

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41580-022-00571-x

This article is cited by

Search

Advanced search

Quick links

Nature Briefing: Cancer

Sign up for the Nature Briefing: Cancer newsletter — what matters in cancer research, free to your inbox weekly.

Get what matters in cancer research, free to your inbox weekly. Sign up for Nature Briefing: Cancer