IDEAS home Printed from https://ideas.repec.org/a/nat/natcom/v12y2021i1d10.1038_s41467-021-27004-1.html
   My bibliography  Save this article

Mechanistic and genetic basis of single-strand templated repair at Cas12a-induced DNA breaks in Chlamydomonas reinhardtii

Author

Listed:
  • Aron Ferenczi

    (University of Edinburgh)

  • Yen Peng Chew

    (University of Edinburgh)

  • Erika Kroll

    (University of Edinburgh
    Rothamsted Research)

  • Charlotte Koppenfels

    (University of Edinburgh)

  • Andrew Hudson

    (University of Edinburgh)

  • Attila Molnar

    (University of Edinburgh)

Abstract

Single-stranded oligodeoxynucleotides (ssODNs) are widely used as DNA repair templates in CRISPR/Cas precision genome editing. However, the underlying mechanisms of single-strand templated DNA repair (SSTR) are inadequately understood, constraining rational improvements to precision editing. Here we study SSTR at CRISPR/Cas12a-induced DNA double-strand breaks (DSBs) in the eukaryotic model green microalga Chlamydomonas reinhardtii. We demonstrate that ssODNs physically incorporate into the genome during SSTR at Cas12a-induced DSBs. This process is genetically independent of the Rad51-dependent homologous recombination and Fanconi anemia pathways, is strongly antagonized by non-homologous end-joining, and is mediated almost entirely by the alternative end-joining enzyme polymerase θ. These findings suggest differences in SSTR between C. reinhardtii and animals. Our work illustrates the promising potentially of C. reinhardtii as a model organism for studying nuclear DNA repair.

Suggested Citation

  • Aron Ferenczi & Yen Peng Chew & Erika Kroll & Charlotte Koppenfels & Andrew Hudson & Attila Molnar, 2021. "Mechanistic and genetic basis of single-strand templated repair at Cas12a-induced DNA breaks in Chlamydomonas reinhardtii," Nature Communications, Nature, vol. 12(1), pages 1-12, December.
    • Handle: RePEc:nat:natcom:v:12:y:2021:i:1:d:10.1038_s41467-021-27004-1
    DOI: 10.1038/s41467-021-27004-1
    as

    Download full text from publisher

    File URL: https://www.nature.com/articles/s41467-021-27004-1
    File Function: Abstract
    Download Restriction: no
    File URL: https://libkey.io/10.1038/s41467-021-27004-1?utm_source=ideas
    LibKey link: if access is restricted and if your library uses this service, LibKey will redirect you to where you can use your library subscription to access this item
    ---><---

    References listed on IDEAS

    as
    1. Quentin Geissmann, 2013. "OpenCFU, a New Free and Open-Source Software to Count Cell Colonies and Other Circular Objects," PLOS ONE, Public Library of Science, vol. 8(2), pages 1-10, February.
    2. Donghyun Lim & Vedagopuram Sreekanth & Kurt J. Cox & Benjamin K. Law & Bridget K. Wagner & Jeffrey M. Karp & Amit Choudhary, 2020. "Engineering designer beta cells with a CRISPR-Cas9 conjugation platform," Nature Communications, Nature, vol. 11(1), pages 1-11, December.
    3. Samuel H. Sternberg & Sy Redding & Martin Jinek & Eric C. Greene & Jennifer A. Doudna, 2014. "DNA interrogation by the CRISPR RNA-guided endonuclease Cas9," Nature, Nature, vol. 507(7490), pages 62-67, March.
    4. Zhuobin Liang & Tzvi Tzfira, 2013. "In vivo formation of double-stranded T-DNA molecules by T-strand priming," Nature Communications, Nature, vol. 4(1), pages 1-8, October.
    5. Molly C. Kottemann & Agata Smogorzewska, 2013. "Fanconi anaemia and the repair of Watson and Crick DNA crosslinks," Nature, Nature, vol. 493(7432), pages 356-363, January.
    6. Anne Bothmer & Tanushree Phadke & Luis A. Barrera & Carrie M Margulies & Christina S. Lee & Frank Buquicchio & Sean Moss & Hayat S. Abdulkerim & William Selleck & Hariharan Jayaram & Vic E. Myer & Cec, 2017. "Characterization of the interplay between DNA repair and CRISPR/Cas9-induced DNA lesions at an endogenous locus," Nature Communications, Nature, vol. 8(1), pages 1-12, April.
    Full references (including those not matched with items on IDEAS)

    Most related items

    These are the items that most often cite the same works as this one and are cited by the same works as this one.
    1. Jun Huang & David Rowe & Pratima Subedi & Wei Zhang & Tyler Suelter & Barbara Valent & David E. Cook, 2022. "CRISPR-Cas12a induced DNA double-strand breaks are repaired by multiple pathways with different mutation profiles in Magnaporthe oryzae," Nature Communications, Nature, vol. 13(1), pages 1-18, December.
    2. Burcu Bestas & Sandra Wimberger & Dmitrii Degtev & Alexandra Madsen & Antje K. Rottner & Fredrik Karlsson & Sergey Naumenko & Megan Callahan & Julia Liz Touza & Margherita Francescatto & Carl Ivar Möl, 2023. "A Type II-B Cas9 nuclease with minimized off-targets and reduced chromosomal translocations in vivo," Nature Communications, Nature, vol. 14(1), pages 1-15, December.
    3. Yi-Li Feng & Qian Liu & Ruo-Dan Chen & Si-Cheng Liu & Zhi-Cheng Huang & Kun-Ming Liu & Xiao-Ying Yang & An-Yong Xie, 2022. "DNA nicks induce mutational signatures associated with BRCA1 deficiency," Nature Communications, Nature, vol. 13(1), pages 1-15, December.
    4. Jessica D. Tischler & Hiroshi Tsuchida & Rosevalentine Bosire & Tommy T. Oda & Ana Park & Richard O. Adeyemi, 2024. "FLIP(C1orf112)-FIGNL1 complex regulates RAD51 chromatin association to promote viability after replication stress," Nature Communications, Nature, vol. 15(1), pages 1-18, December.
    5. Michael Kosicki & Felicity Allen & Frances Steward & Kärt Tomberg & Yangyang Pan & Allan Bradley, 2022. "Cas9-induced large deletions and small indels are controlled in a convergent fashion," Nature Communications, Nature, vol. 13(1), pages 1-11, December.
    6. Xu Feng & Ruyi Xu & Jianglan Liao & Jingyu Zhao & Baochang Zhang & Xiaoxiao Xu & Pengpeng Zhao & Xiaoning Wang & Jianyun Yao & Pengxia Wang & Xiaoxue Wang & Wenyuan Han & Qunxin She, 2024. "Flexible TAM requirement of TnpB enables efficient single-nucleotide editing with expanded targeting scope," Nature Communications, Nature, vol. 15(1), pages 1-14, December.
    7. Sébastien Levesque & Diana Mayorga & Jean-Philippe Fiset & Claudia Goupil & Alexis Duringer & Andréanne Loiselle & Eva Bouchard & Daniel Agudelo & Yannick Doyon, 2022. "Marker-free co-selection for successive rounds of prime editing in human cells," Nature Communications, Nature, vol. 13(1), pages 1-14, December.
    8. J. Ferreira da Silva & G. P. Oliveira & E. A. Arasa-Verge & C. Kagiou & A. Moretton & G. Timelthaler & J. Jiricny & J. I. Loizou, 2022. "Prime editing efficiency and fidelity are enhanced in the absence of mismatch repair," Nature Communications, Nature, vol. 13(1), pages 1-11, December.
    9. Karl-Heinz Tomaszowski & Sunetra Roy & Carolina Guerrero & Poojan Shukla & Caezaan Keshvani & Yue Chen & Martina Ott & Xiaogang Wu & Jianhua Zhang & Courtney D. DiNardo & Detlev Schindler & Katharina , 2023. "Hypomorphic Brca2 and Rad51c double mutant mice display Fanconi anemia, cancer and polygenic replication stress," Nature Communications, Nature, vol. 14(1), pages 1-15, December.
    10. Wei Liu & Hongchao Cao & Jing Wang & Areeg Elmusrati & Bing Han & Wei Chen & Ping Zhou & Xiyao Li & Stephen Keysar & Antonio Jimeno & Cun-Yu Wang, 2024. "Histone-methyltransferase KMT2D deficiency impairs the Fanconi anemia/BRCA pathway upon glycolytic inhibition in squamous cell carcinoma," Nature Communications, Nature, vol. 15(1), pages 1-18, December.
    11. Lin Zhao & Sabrina R. T. Koseki & Rachel A. Silverstein & Nadia Amrani & Christina Peng & Christian Kramme & Natasha Savic & Martin Pacesa & Tomás C. Rodríguez & Teodora Stan & Emma Tysinger & Lauren , 2023. "PAM-flexible genome editing with an engineered chimeric Cas9," Nature Communications, Nature, vol. 14(1), pages 1-8, December.
    12. Jian Wang & Ke Wang & Zhe Deng & Zhiyu Zhong & Guo Sun & Qing Mei & Fuling Zhou & Zixin Deng & Yuhui Sun, 2024. "Engineered cytosine base editor enabling broad-scope and high-fidelity gene editing in Streptomyces," Nature Communications, Nature, vol. 15(1), pages 1-16, December.
    13. Yang Liu & Filipe Pinto & Xinyi Wan & Zhugen Yang & Shuguang Peng & Mengxi Li & Jonathan M. Cooper & Zhen Xie & Christopher E. French & Baojun Wang, 2022. "Reprogrammed tracrRNAs enable repurposing of RNAs as crRNAs and sequence-specific RNA biosensors," Nature Communications, Nature, vol. 13(1), pages 1-12, December.
    14. Shinta Saito & Noritaka Adachi, 2024. "Characterization and regulation of cell cycle-independent noncanonical gene targeting," Nature Communications, Nature, vol. 15(1), pages 1-13, December.
    15. Yanbo Wang & W. Taylor Cottle & Haobo Wang & Momcilo Gavrilov & Roger S. Zou & Minh-Tam Pham & Srinivasan Yegnasubramanian & Scott Bailey & Taekjip Ha, 2022. "Achieving single nucleotide sensitivity in direct hybridization genome imaging," Nature Communications, Nature, vol. 13(1), pages 1-12, December.
    16. Carla Umansky & Agustín E. Morellato & Matthias Rieckher & Marco A. Scheidegger & Manuela R. Martinefski & Gabriela A. Fernández & Oleg Pak & Ksenia Kolesnikova & Hernán Reingruber & Mariela Bollini &, 2022. "Endogenous formaldehyde scavenges cellular glutathione resulting in redox disruption and cytotoxicity," Nature Communications, Nature, vol. 13(1), pages 1-15, December.
    17. Adeeb Rahman & Neeti Sanan-Mishra, 2024. "When an Intruder Comes Home: GM and GE Strategies to Combat Virus Infection in Plants," Agriculture, MDPI, vol. 14(2), pages 1-26, February.
    18. Dmitrii Degtev & Jack Bravo & Aikaterini Emmanouilidi & Aleksandar Zdravković & Oi Kuan Choong & Julia Liz Touza & Niklas Selfjord & Isabel Weisheit & Margherita Francescatto & Pinar Akcakaya & Michel, 2024. "Engineered PsCas9 enables therapeutic genome editing in mouse liver with lipid nanoparticles," Nature Communications, Nature, vol. 15(1), pages 1-15, December.
    19. Dixit, Yatika & Yadav, Preeti & Sharma, Arun Kumar & Pandey, Poornima & Kuila, Arindam, 2023. "Multiplex genome editing to construct cellulase engineered Saccharomyces cerevisiae for ethanol production from cellulosic biomass," Renewable and Sustainable Energy Reviews, Elsevier, vol. 187(C).
    20. Haoqing Shen & Omer An & Xi Ren & Yangyang Song & Sze Jing Tang & Xin-Yu Ke & Jian Han & Daryl Jin Tai Tay & Vanessa Hui En Ng & Fernando Bellido Molias & Priyankaa Pitcheshwar & Ka Wai Leong & Ker-Ka, 2022. "ADARs act as potent regulators of circular transcriptome in cancer," Nature Communications, Nature, vol. 13(1), pages 1-16, December.

    More about this item

    Statistics

    Access and download statistics

    Corrections

    All material on this site has been provided by the respective publishers and authors. You can help correct errors and omissions. When requesting a correction, please mention this item's handle: RePEc:nat:natcom:v:12:y:2021:i:1:d:10.1038_s41467-021-27004-1. See general information about how to correct material in RePEc.

    If you have authored this item and are not yet registered with RePEc, we encourage you to do it here. This allows to link your profile to this item. It also allows you to accept potential citations to this item that we are uncertain about.

    If CitEc recognized a bibliographic reference but did not link an item in RePEc to it, you can help with this form .

    If you know of missing items citing this one, you can help us creating those links by adding the relevant references in the same way as above, for each refering item. If you are a registered author of this item, you may also want to check the "citations" tab in your RePEc Author Service profile, as there may be some citations waiting for confirmation.

    For technical questions regarding this item, or to correct its authors, title, abstract, bibliographic or download information, contact: Sonal Shukla or Springer Nature Abstracting and Indexing (email available below). General contact details of provider: http://www.nature.com .

    Please note that corrections may take a couple of weeks to filter through the various RePEc services.

    IDEAS is a RePEc service. RePEc uses bibliographic data supplied by the respective publishers.