DNA damage can result from intrinsic cellular processes and from exposure to stressful environments. Such DNA damage generally threatens genome integrity and cell viability1. However, here we report that the transient induction of DNA strand breaks (single-strand breaks, double-strand breaks or both) in the moss Physcomitrella patens can trigger the reprogramming of differentiated leaf cells into stem cells without cell death. After intact leafy shoots (gametophores) were exposed to zeocin, an inducer of DNA strand breaks, the STEM CELL-INDUCING FACTOR 1 (STEMIN1)2 promoter was activated in some leaf cells. These cells subsequently initiated tip growth and underwent asymmetric cell divisions to form chloronema apical stem cells, which are in an earlier phase of the life cycle than leaf cells and have the ability to form new gametophores. This DNA-strand-break-induced reprogramming required the DNA damage sensor ATR kinase, but not ATM kinase, together with STEMIN1 and closely related proteins. ATR was also indispensable for the induction of STEMIN1 by DNA strand breaks. Our findings indicate that DNA strand breaks, which are usually considered to pose a severe threat to cells, trigger cellular reprogramming towards stem cells via the activity of ATR and STEMINs.

Center for Optical Research and Education Utsunomiya University Utsunomiya Japan

College of Life Science and Technology Huazhong Agricultural University Wuhan P R China

Department of Basic Biology The Graduate University for Advanced Studies SOKENDAI Okazaki Japan

Division of Evolutionary Biology National Institute for Basic Biology Okazaki Japan

Institute of Experimental Botany Academy of Sciences of the Czech Republic Prague Czech Republic

Key Laboratory of Horticultural Plant Biology Huazhong Agricultural University Wuhan P R China

School of Engineering Utsunomiya University Utsunomiya Japan

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Yoshiyama, K. O., Sakaguchi, K. & Kimura, S. DNA damage response in plants: conserved and variable response compared to animals. Biology 2, 1338–1356 (2013). PubMed DOI PMC

Ishikawa, M. et al. Physcomitrella STEMIN transcription factor induces stem cell formation with epigenetic reprogramming. Nat. Plants 5, 681–690 (2019). PubMed DOI

Ikeuchi, M., Ogawa, Y., Iwase, A. & Sugimoto, K. Plant regeneration: cellular origins and molecular mechanisms. Development 143, 1442–1451 (2016). PubMed DOI

Pilkington, M. The regeneration of the stem apex. New Phytol. 28, 37–53 (1929). DOI

Reinhardt, D., Frenz, M., Mandel, T. & Kuhlemeier, C. Microsurgical and laser ablation analysis of interactions between the zones and layers of the tomato shoot apical meristem. Development 130, 4073–4083 (2003). PubMed DOI

Heyman, J. et al. ERF115 controls root quiescent center cell division and stem cell replenishment. Science 342, 860–863 (2013). PubMed DOI

Sena, G., Wang, X., Liu, H. Y., Hofhuis, H. & Birnbaum, K. D. Organ regeneration does not require a functional stem cell niche in plants. Nature 457, 1150–1153 (2009). PubMed DOI PMC

Zhou, W. et al. A Jasmonate signaling network activates root stem cells and promotes regeneration. Cell 177, 942–956 (2019). PubMed DOI

Fulcher, N. & Sablowski, R. Hypersensitivity to DNA damage in plant stem cell niches. Proc. Natl Acad. Sci. USA 106, 20984–20988 (2009). PubMed DOI PMC

Johnson, R. A. et al. SUPPRESSOR OF GAMMA RESPONSE 1 links DNA damage response to organ regeneration. Plant Physiol. 176, 1665–1675 (2017). PubMed DOI PMC

Steward, F. C., Mapes, M. O. & Mears, K. Growth and organized development of cultured cells. II. Organization in cultures grown from freely suspended cells. Am. J. Bot. 45, 705–709 (1958). DOI

Kareem, A. et al. PLETHORA genes control regeneration by a two-step mechanism. Curr. Biol. 25, 1017–1030 (2015). PubMed DOI PMC

Ishikawa, M. et al. Physcomitrella cyclin-dependent kinase A links cell cycle reactivation to other cellular changes during reprogramming of leaf cells. Plant Cell 23, 2924–2938 (2011). PubMed DOI PMC

Sato, Y. et al. Cells reprogramming to stem cells inhibit the reprogramming of adjacent cells in the moss Physcomitrella patens. Sci. Rep. 7, 1909 (2017). PubMed DOI PMC

Li, C. et al. A Lin28 homolog reprograms differentiated cells to stem cells in the moss Physcomitrella patens. Nat. Commun. 8, 14242 (2017). PubMed DOI PMC

Kofuji, R. & Hasebe, M. Eight types of stem cells in the life cycle of the moss Physcomitrella patens. Curr. Opin. Plant Biol. 17, 13–21 (2014). PubMed DOI

Chankova, S. G., Dimova, E., Dimitrova, M. & Bryant, P. E. Induction of DNA double-strand breaks by zeocin in Chlamydomonas reinhardtii and the role of increased DNA double-strand breaks rejoining in the formation of an adaptive response. Radiat. Environ. Biophys. 46, 409–416 (2007). PubMed DOI

Chen, J., Ghorai, M. K., Kenney, G. & Stubbe, J. A. Mechanistic studies on bleomycin-mediated DNA damage: multiple binding modes can result in double-stranded DNA cleavage. Nucleic Acids Res. 36, 3781–3790 (2008). PubMed DOI PMC

Sakasai, R. & Iwabuchi, K. The distinctive cellular responses to DNA strand breaks caused by a DNA topoisomerase I poison in conjunction with DNA replication and RNA transcription. Genes Genet. Syst. 90, 187–194 (2015). PubMed DOI

Angelis, K. J., Dušinská, M. & Collins, A. R. Single cell gel electrophoresis: detection of DNA damage at different levels of sensitivity. Electrophoresis 20, 2133–2138 (1999). PubMed DOI

Lanier, C., Manier, N., Cuny, D. & Deram, A. The comet assay in higher terrestrial plant model: review and evolutionary trends. Environ. Pollut. 207, 6–20 (2015). PubMed DOI

Jones, K., Kim, D. W., Park, J. S. & Khang, C. H. Live-cell fluorescence imaging to investigate the dynamics of plant cell death during infection by the rice blast fungus Magnaporthe oryzae. BMC Plant Biol. 16, 69 (2016). PubMed DOI PMC

Antosiewicz-Bourget, J. et al. Induced pluripotent stem cell lines derived from human somatic cells. Science 318, 1917–1920 (2007). PubMed DOI

Hu, Z., Cools, T. & De Veylder, L. Mechanisms used by plants to cope with DNA damage. Annu. Rev. Plant Biol. 67, 439–462 (2016). PubMed DOI

Collonnier, C. et al. CRISPR–Cas9-mediated efficient directed mutagenesis and RAD51-dependent and RAD51-independent gene targeting in the moss Physcomitrella patens. Plant Biotechnol. J. 15, 122–131 (2017). PubMed DOI

Hong, J. H. et al. A sacrifice-for-survival mechanism protects root stem cell niche from chilling stress. Cell 170, 102–113 (2017). PubMed DOI

Sharma, P., Jha, A. B., Dubey, R. S. & Pessarakli, M. Reactive oxygen species, oxidative damage, and antioxidative defense mechanism in plants under stressful conditions. J. Bot. 2012, 217037 (2012).

Rodriguez, E., Azevedo, R., Fernandes, P. & Santos, C. Cr(VI) induces DNA damage, cell cycle arrest and polyploidization: a flow cytometric and comet assay study in Pisum sativum. Chem. Res. Toxicol. 24, 1040–1047 (2011). PubMed DOI

Gehring, M., Reik, W. & Henikoff, S. DNA demethylation by DNA repair. Trends Genet. 25, 82–90 (2009). PubMed DOI

Gursoy-Yuzugullu, O., House, N. & Price, B. D. Patching broken DNA: nucleosome dynamics and the repair of DNA breaks. J. Mol. Biol. 428, 1846–1860 (2016). PubMed DOI

Rensing, S. A. et al. The Physcomitrella genome reveals evolutionary insights into the conquest of land by plants. Science 319, 64–69 (2008). PubMed DOI

Nishiyama, T., Hiwatashi, Y., Sakakibara, K., Kato, M. & Hasebe, M. Tagged mutagenesis and gene-trap in the moss, Physcomitrella patens by shuttle mutagenesis. DNA Res. 7, 9–17 (2000). PubMed DOI

Aoyama, T. et al. AP2-type transcription factors determine stem cell identity in the moss Physcomitrella patens. Development 139, 3120–3129 (2012). PubMed DOI

Enciso, M., Sarasa, J., Agarwal, A., Fernández, J. L. & Gosálvez, J. A two-tailed comet assay for assessing DNA damage in spermatozoa. Reprod. Biomed. Online 18, 609–616 (2009). PubMed DOI

Cortés-Gutiérrez, E. I., Fernández, J. L., Dávila-Rodríguez, M. I., López-Fernández, C. & Gosálvez, J. Two-tailed comet assay (2T-Comet): simultaneous detection of DNA single and double strand breaks. Methods Mol. Biol. 1560, 285–293 (2017). PubMed DOI

Gyori, B. M., Venkatachalam, G., Thiagarajan, P. S., Hsu, D. & Clement, M. V. OpenComet: an automated tool for comet assay image analysis. Redox Biol. 2, 457–465 (2014). PubMed DOI PMC

R: A Language and Environment for Statistical Computing v.3.5.1 (R Core Team, 2018); https://doi.org/10.1787/csp-aut-table-2018-1-en

Wickham, H. ggplot2: Elegant graphics for data analysis v.3.2.1 https://doi.org/10.1007/978-3-319-24277-4_2 (2016).

Odell, J., Caimi, P., Sauer, B. & Russell, S. Site-directed recombination in the genome of transgenic tobacco. Mol. Gen. Genet. 223, 369–378 (1990). PubMed DOI

Martin, M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet J. 17, 10–12 (2011). DOI

Bray, N. L., Pimentel, H., Melsted, P. & Pachter, L. Near-optimal probabilistic RNA-seq quantification. Nat. Biotechnol. 34, 525–527 (2016). PubMed DOI

Pimentel, H., Bray, N. L., Puente, S., Melsted, P. & Pachter, L. Differential analysis of RNA-seq incorporating quantification uncertainty. Nat. Methods 14, 687–690 (2017). PubMed DOI

Jari, O. et al. vegan: Community ecology package. R package v.2.5-2 (2018).

Livak, K. J. & Schmittgen, T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2−ΔΔCT method. Methods 25, 402–408 (2001). PubMed DOI

Lang, D. et al. The Physcomitrella patens chromosome-scale assembly reveals moss genome structure and evolution. Plant J. 93, 515–533 (2018). PubMed DOI