Skip to main content
SpringerLink
Log in

DEtail-seq is an ultra-efficient and convenient method for meiotic DNA break profiling in multiple organisms

  • Research Paper
  • Published:
Science China Life Sciences Aims and scope Submit manuscript

Abstract

Programmed DNA double-strand break (DSB) formation is a crucial step in meiotic recombination, yet techniques for high-efficiency and precise mapping of the 3′ ends of DSBs are still in their infancy. Here, we report a novel technique, named DNA End tailing and sequencing (DEtail-seq), which can directly and ultra-efficiently characterize the 3′ ends of meiotic DSBs with near single-nucleotide resolution in a variety of species, including yeast, mouse, and human. We find that the 3′ ends of meiotic DSBs are stable without significant resection in budding yeast. Meiotic DSBs are strongly enriched in de novo H3K4me3 peaks in the mouse genome at leptotene stage. We also profile meiotic DSBs in human and find DSB hotspots are enriched near the common fragile sites during human meiosis, especially at CCCTC-binding factor (CTCF)-associated enhancers. Therefore, DEtail-seq provides a powerful method to detect DSB ends in various species, and our results provide new insights into the distribution and regulation of meiotic DSB hotspots.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

Data availability

All the data of this study are available from the corresponding author upon any reasonable request. The GEO accession numbers for the DEtail-seq data used in this paper are GSE154226, GSE154227 and GSE154289. The detailed information of software used in this study is listed in Table S4 in Supporting Information.

References

  • Bae, S., and Lesch, B.J. (2020). H3K4me1 distribution predicts transcription state and poising at promoters. Front Cell Dev Biol 8, 289.

    Article  PubMed  PubMed Central  Google Scholar 

  • Baudat, F., Buard, J., Grey, C., Fledel-Alon, A., Ober, C., Przeworski, M., Coop, G., and de Massy, B. (2010). PRDM9 is a major determinant of meiotic recombination hotspots in humans and mice. Science 327, 836–840.

    Article  CAS  PubMed  Google Scholar 

  • Baudat, F., Imai, Y., and de Massy, B. (2013). Meiotic recombination in mammals: localization and regulation. Nat Rev Genet 14, 794–806.

    Article  CAS  PubMed  Google Scholar 

  • Blat, Y., Protacio, R.U., Hunter, N., and Kleckner, N. (2002). Physical and functional interactions among basic chromosome organizational features govern early steps of meiotic chiasma formation. Cell 111, 791–802.

    Article  CAS  PubMed  Google Scholar 

  • Boekhout, M., Karasu, M.E., Wang, J., Acquaviva, L., Pratto, F., Brick, K., Eng, D.Y., Xu, J., Camerini-Otero, R.D., Patel, D.J., et al. (2019). REC114 partner ANKRD31 controls number, timing, and location of meiotic DNA breaks. Mol Cell 74, 1053–1068.e8.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Borde, V., and de Massy, B. (2013). Programmed induction of DNA double strand breaks during meiosis: setting up communication between DNA and the chromosome structure. Curr Opin Genet Dev 23, 147–155.

    Article  CAS  PubMed  Google Scholar 

  • Brar, G.A., Yassour, M., Friedman, N., Regev, A., Ingolia, N.T., and Weissman, J.S. (2012). High-resolution view of the yeast meiotic program revealed by ribosome profiling. Science 335, 552–557.

    Article  CAS  PubMed  Google Scholar 

  • Brick, K., Thibault-Sennett, S., Smagulova, F., Lam, K.W.G., Pu, Y., Pratto, F., Camerini-Otero, R.D., and Petukhova, G.V. (2018). Extensive sex differences at the initiation of genetic recombination. Nature 561, 338–342.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Canela, A., Maman, Y., Jung, S., Wong, N., Callen, E., Day, A., Kieffer-Kwon, K.R., Pekowska, A., Zhang, H., Rao, S.S.P., et al. (2017). Genome organization drives chromosome fragility. Cell 170, 507–521.e18.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Canela, A., Sridharan, S., Sciascia, N., Tubbs, A., Meltzer, P., Sleckman, B. P., and Nussenzweig, A. (2016). DNA breaks and end resection measured genome-wide by end sequencing. Mol Cell 63, 898–911.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chen, Y., Lyu, R., Rong, B., Zheng, Y., Lin, Z., Dai, R., Zhang, X., Xie, N., Wang, S., Tang, F., et al. (2020). Refined spatial temporal epigenomic profiling reveals intrinsic connection between PRDM9-mediated H3K4me3 and the fate of double-stranded breaks. Cell Res 30, 256–268.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Chen, Y., Zheng, Y., Gao, Y., Lin, Z., Yang, S., Wang, T., Wang, Q., Xie, N., Hua, R., Liu, M., et al. (2018). Single-cell RNA-seq uncovers dynamic processes and critical regulators in mouse spermatogenesis. Cell Res 28, 879–896.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Consortium, E.P. (2012). An integrated encyclopedia of DNA elements in the human genome. Nature 489, 57–74.

    Article  Google Scholar 

  • Creyghton, M.P., Cheng, A.W., Welstead, G.G., Kooistra, T., Carey, B.W., Steine, E.J., Hanna, J., Lodato, M.A., Frampton, G.M., Sharp, P.A., et al. (2010). Histone H3K27ac separates active from poised enhancers and predicts developmental state. Proc Natl Acad Sci USA 107, 21931–21936.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • d’Adda di Fagagna, F. (2008). Living on a break: cellular senescence as a DNA-damage response. Nat Rev Cancer 8, 512–522.

    Article  PubMed  Google Scholar 

  • de Massy, B. (2013). Initiation of meiotic recombination: how and where? Conservation and specificities among eukaryotes. Annu Rev Genet 47, 563–599.

    Article  CAS  PubMed  Google Scholar 

  • Dujon, B. (1996). The yeast genome project: what did we learn? Trends Genet 12, 263–270.

    Article  CAS  PubMed  Google Scholar 

  • Gansauge, M.T., Gerber, T., Glocke, I., Korlević, P., Lippik, L., Nagel, S., Riehl, L.M., Schmidt, A., and Meyer, M. (2017). Single-stranded DNA library preparation from highly degraded DNA using T4 DNA ligase. Nucleic Acids Res 45, e79.

    CAS  PubMed  PubMed Central  Google Scholar 

  • Garcia, V., Phelps, S.E.L., Gray, S., and Neale, M.J. (2011). Bidirectional resection of DNA double-strand breaks by Mre11 and Exo1. Nature 479, 241–244.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Gel, B., Díez-Villanueva, A., Serra, E., Buschbeck, M., Peinado, M.A., and Malinverni, R. (2016). regioneR: an R/Bioconductor package for the association analysis of genomic regions based on permutation tests. Bioinformatics 32, 289–291.

    Article  CAS  PubMed  Google Scholar 

  • Gittens, W.H., Johnson, D.J., Allison, R.M., Cooper, T.J., Thomas, H., and Neale, M.J. (2019). A nucleotide resolution map of Top2-linked DNA breaks in the yeast and human genome. Nat Commun 10, 4846.

    Article  PubMed  PubMed Central  Google Scholar 

  • Hinch, A.G., Becker, P.W., Li, T., Moralli, D., Zhang, G., Bycroft, C., Green, C., Keeney, S., Shi, Q., Davies, B., et al. (2020). The configuration of RPA, RAD51, and DMC1 binding in meiosis reveals the nature of critical recombination intermediates. Mol Cell 79, 689–701.e10.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ji, F., Liao, H., Pan, S., Ouyang, L., Jia, F., Fu, Z., Zhang, F., Geng, X., Wang, X., Li, T., et al. (2020). Genome-wide high-resolution mapping of mitotic DNA synthesis sites and common fragile sites by direct sequencing. Cell Res 30, 1009–1023.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kaiser, V.B., and Semple, C.A. (2018). Chromatin loop anchors are associated with genome instability in cancer and recombination hotspots in the germline. Genome Biol 19, 101.

    Article  PubMed  PubMed Central  Google Scholar 

  • Kara, N., Krueger, F., Rugg-Gunn, P., and Houseley, J. (2021). Genome-wide analysis of DNA replication and DNA double-strand breaks using TrAEL-seq. PLoS Biol 19, e3000886.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Keeney, S. (2008). Spo11 and the formation of DNA double-strand breaks in meiosis. In: Egel, R., and Lankenau, D.H., eds. Recombination and Meiosis. Genome Dynamics and Stability. Berlin, Heidelberg: Springer. 81–123.

    Chapter  Google Scholar 

  • Keeney, S., Lange, J., and Mohibullah, N. (2014). Self-organization of meiotic recombination initiation: general principles and molecular pathways. Annu Rev Genet 48, 187–214.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Lange, J., Yamada, S., Tischfield, S.E., Pan, J., Kim, S., Zhu, X., Socci, N. D., Jasin, M., and Keeney, S. (2016). The landscape of mouse meiotic double-strand break formation, processing, and repair. Cell 167, 695–708.e16.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Langmead, B., and Salzberg, S.L. (2012). Fast gapped-read alignment with Bowtie 2. Nat Methods 9, 357–359.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Langmead, B., Trapnell, C., Pop, M., and Salzberg, S.L. (2009). Ultrafast and memory-efficient alignment of short DNA sequences to the human genome. Genome Biol 10, R25.

    Article  PubMed  PubMed Central  Google Scholar 

  • Li, Y., Song, Y., Xu, W., Li, Q., Wang, X., Li, K., Wang, J., Liu, Z., Velychko, S., Ye, R., et al. (2020). R-loops coordinate with SOX2 in regulating reprogramming to pluripotency. Sci Adv 6, eaba0777.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu, T., Wang, L., Chen, H., Huang, Y., Yang, P., Ahmed, N., Wang, T., Liu, Y., and Chen, Q. (2017). Molecular and cellular mechanisms of apoptosis during dissociated spermatogenesis. Front Physiol 8.

  • Luo, Z., Wang, X., Jiang, H., Wang, R., Chen, J., Chen, Y., Xu, Q., Cao, J., Gong, X., Wu, J., et al. (2020). Reorganized 3D genome structures support transcriptional regulation in mouse spermatogenesis. iScience 23, 101034.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mimitou, E.P., and Symington, L.S. (2009). DNA end resection: many nucleases make light work. DNA Repair 8, 983–995.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mimitou, E.P., Yamada, S., and Keeney, S. (2017). A global view of meiotic double-strand break end resection. Science 355, 40–45.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Neale, M.J., Pan, J., and Keeney, S. (2005). Endonucleolytic processing of covalent protein-linked DNA double-strand breaks. Nature 436, 1053–1057.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ohkura, H. (2015). Meiosis: an overview of key differences from mitosis. Cold Spring Harb Perspect Biol 7, a015859.

    Article  PubMed  PubMed Central  Google Scholar 

  • Paiano, J., Wu, W., Yamada, S., Sciascia, N., Callen, E., Paola Cotrim, A., Deshpande, R.A., Maman, Y., Day, A., Paull, T.T., et al. (2020). ATM and PRDM9 regulate SPO11-bound recombination intermediates during meiosis. Nat Commun 11, 857.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pan, J., Sasaki, M., Kniewel, R., Murakami, H., Blitzblau, H.G., Tischfield, S.E., Zhu, X., Neale, M.J., Jasin, M., Socci, N.D., et al. (2011). A hierarchical combination of factors shapes the genome-wide topography of yeast meiotic recombination initiation. Cell 144, 719–731.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Panier, S., and Durocher, D. (2013). Push back to respond better: regulatory inhibition of the DNA double-strand break response. Nat Rev Mol Cell Biol 14, 661–672.

    Article  CAS  PubMed  Google Scholar 

  • Petrosino, G., Zilio, N., Sriramachandran, A.M., and Ulrich, H.D. (2020). Preparation and analysis of GLOE-seq libraries for genome-wide mapping of DNA replication patterns, single-strand breaks, and lesions. STAR Protoc 1, 100076.

    Article  PubMed  PubMed Central  Google Scholar 

  • Pratto, F., Brick, K., Khil, P., Smagulova, F., Petukhova, G.V., and Camerini-Otero, R.D. (2014). Recombination initiation maps of individual human genomes. Science 346, 1256442.

    Article  PubMed  PubMed Central  Google Scholar 

  • Quinlan, A.R., and Hall, I.M. (2010). BEDTools: a flexible suite of utilities for comparing genomic features. Bioinformatics 26, 841–842.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Ramírez, F., Dündar, F., Diehl, S., Grüning, B.A., and Manke, T. (2014). deepTools: a flexible platform for exploring deep-sequencing data. Nucleic Acids Res 42, W187–W191.

    Article  PubMed  PubMed Central  Google Scholar 

  • Robinson, J.T., Thorvaldsdóttir, H., Winckler, W., Guttman, M., Lander, E. S., Getz, G., and Mesirov, J.P. (2011). Integrative genomics viewer. Nat Biotechnol 29, 24–26.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shaha, C., Tripathi, R., and Mishra, D.P. (2010). Male germ cell apoptosis: regulation and biology. Phil Trans R Soc B 365, 1501–1515.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sriramachandran, A.M., Petrosino, G., Méndez-Lago, M., Schäfer, A.J., Batista-Nascimento, L.S., Zilio, N., and Ulrich, H.D. (2020). Genome-wide nucleotide-resolution mapping of DNA Replication patterns, singlestrand breaks, and lesions by GLOE-Seq. Mol Cell 78, 975–985.e7.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Symington, L.S. (2014). End resection at double-strand breaks: mechanism and regulation. Cold Spring Harb Perspect Biol 6, a016436.

    Article  PubMed  PubMed Central  Google Scholar 

  • Tubbs, A., Sridharan, S., van Wietmarschen, N., Maman, Y., Callen, E., Stanlie, A., Wu, W., Wu, X., Day, A., Wong, N., et al. (2018). Dual roles of poly(dA:dT) tracts in replication initiation and fork collapse. Cell 174, 1127–1142.e19.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Weinbauer, G.F., Aslam, H., Krishnamurthy, H., Brinkworth, M.H., Einspanier, A., and Hodges, J.K. (2001). Quantitative analysis of spermatogenesis and apoptosis in the common marmoset (Callithrix jacchus) reveals high rates of spermatogonial turnover and high spermatogenic efficiency. Biol Reprod 64, 120–126.

    Article  CAS  PubMed  Google Scholar 

  • Wen, F.P., Guo, Y.S., Hu, Y., Liu, W.X., Wang, Q., Wang, Y.T., Yu, H.Y., Tang, C.M., Yang, J., Zhou, T., et al. (2016). Distinct temporal requirements for autophagy and the proteasome in yeast meiosis. Autophagy 12, 671–688.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • White, C.I., and Haber, J.E. (1990). Intermediates of recombination during mating type switching in Saccharomyces cerevisiae. EMBO J 9, 663–673.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xu, W., Li, K., Li, S., Hou, Q., Zhang, Y., Liu, K., and Sun, Q. (2020). The R-loop atlas of Arabidopsis development and responses to environmental stimuli. Plant Cell 32, 888–903.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Xu, W., Xu, H., Li, K., Fan, Y., Liu, Y., Yang, X., and Sun, Q. (2017). The R-loop is a common chromatin feature of the Arabidopsis genome. Nat Plants 3, 704–714.

    Article  CAS  PubMed  Google Scholar 

  • Xu, Z., Song, Z., Li, G., Tu, H., Liu, W., Liu, Y., Wang, P., Wang, Y., Cui, X., Liu, C., et al. (2016). H2B ubiquitination regulates meiotic recombination by promoting chromatin relaxation. Nucleic Acids Res 44, 9681–9697.

    CAS  PubMed  PubMed Central  Google Scholar 

  • Yang, X., Liu, Q.L., Xu, W., Zhang, Y.C., Yang, Y., Ju, L.F., Chen, J., Chen, Y.S., Li, K., Ren, J., et al. (2019). m6A promotes R-loop formation to facilitate transcription termination. Cell Res 29, 1035–1038.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Young, K.A., Ball, G.F., and Nelson, R.J. (2001). Photoperiod-induced testicular apoptosis in European starlings (Sturnus vulgaris). Biol Reprod 64, 706–713.

    Article  CAS  PubMed  Google Scholar 

  • Yuan, W., Zhou, J., Tong, J., Zhuo, W., Wang, L., Li, Y., Sun, Q., and Qian, W. (2019). ALBA protein complex reads genic R-loops to maintain genome stability in Arabidopsis. Sci Adv 5, eaav9040.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhu, Z., Chung, W.H., Shim, E.Y., Lee, S.E., and Ira, G. (2008). Sgs1 helicase and two nucleases Dna2 and Exo1 resect DNA double-strand break ends. Cell 134, 981–994.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was supported by the National Natural Science Foundation of China (91740105, 31822028, 32071437, 31900302) and Central Public-interest Scientific Institution Basal Research Fund (Y2022QC33). We greatly appreciate all the Sun Lab members for useful discussions. The Sun Lab is supported by Tsinghua-Peking Joint Center for Life Sciences.

Author information

Author notes

  1. Contributed equally to this work

Authors and Affiliations

  1. School of Life Sciences, Tsinghua University, Beijing, 100084, China

    Wei Xu, Qin Li, Kuan Li & Qianwen Sun

  2. Guangzhou Women and Children’s Medical Center, Guangzhou Medical University, Guangzhou, 510623, China

    Chao Liu & Wei Li

  3. Department of Urology, Department of Andrology, Department of Reproductive Medicine Center, and Department of Human Sperm Bank, Peking University Third Hospital, Beijing, 100191, China

    Zhe Zhang & Hui Jiang

  4. Tsinghua-Peking Center for Life Sciences, Beijing, 100084, China

    Wei Xu, Qin Li, Kuan Li & Qianwen Sun

  5. Shenzhen Branch, Guangdong Laboratory of Lingnan Modern Agriculture, Genome Analysis Laboratory of the Ministry of Agriculture and Rural Affairs, Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences, Shenzhen, 518120, China

    Wei Xu & Changbin Sun

Authors
  1. Wei Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Chao Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Zhe Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Changbin Sun
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Qin Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Kuan Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Hui Jiang
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Wei Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Qianwen Sun
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Wei Xu, Hui Jiang, Wei Li or Qianwen Sun.

Additional information

Compliance and ethics

The author(s) declare that they have no conflict of interest.

Electronic supplementary material

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Xu, W., Liu, C., Zhang, Z. et al. DEtail-seq is an ultra-efficient and convenient method for meiotic DNA break profiling in multiple organisms. Sci. China Life Sci. 66, 1392–1407 (2023). https://doi.org/10.1007/s11427-022-2277-y

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11427-022-2277-y

Keywords

Search

Navigation