IDEAS home Printed from https://ideas.repec.org/a/nat/natcom/v15y2024i1d10.1038_s41467-024-49385-9.html
   My bibliography  Save this article

Characterization and regulation of cell cycle-independent noncanonical gene targeting

Author

Listed:
  • Shinta Saito

    (Yokohama City University)

  • Noritaka Adachi

    (Yokohama City University)

Abstract

Homology-dependent targeted DNA integration, generally referred to as gene targeting, provides a powerful tool for precise genome modification; however, its fundamental mechanisms remain poorly understood in human cells. Here we reveal a noncanonical gene targeting mechanism that does not rely on the homologous recombination (HR) protein Rad51. This mechanism is suppressed by Rad52 inhibition, suggesting the involvement of single-strand annealing (SSA). The SSA-mediated gene targeting becomes prominent when DSB repair by HR or end-joining pathways is defective and does not require isogenic DNA, permitting 5% sequence divergence. Intriguingly, loss of Msh2, loss of BLM, and induction of a target-site DNA break all significantly and synergistically enhance SSA-mediated targeted integration. Most notably, SSA-mediated integration is cell cycle-independent, occurring in the G1 phase as well. Our findings provide unequivocal evidence for Rad51-independent targeted integration and unveil multiple mechanisms to regulate SSA-mediated targeted as well as random integration.

Suggested Citation

  • 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.
    • Handle: RePEc:nat:natcom:v:15:y:2024:i:1:d:10.1038_s41467-024-49385-9
    DOI: 10.1038/s41467-024-49385-9
    as

    Download full text from publisher

    File URL: https://www.nature.com/articles/s41467-024-49385-9
    File Function: Abstract
    Download Restriction: no
    File URL: https://libkey.io/10.1038/s41467-024-49385-9?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. Shinta Saito & Ryo Maeda & Noritaka Adachi, 2017. "Dual loss of human POLQ and LIG4 abolishes random integration," Nature Communications, Nature, vol. 8(1), pages 1-10, December.
    2. Eric Van Dyck & Alicja Z. Stasiak & Andrzej Stasiak & Stephen C. West, 1999. "Binding of double-strand breaks in DNA by human Rad52 protein," Nature, Nature, vol. 401(6751), pages 403-403, September.
    3. Eric Van Dyck & Alicja Z. Stasiak & Andrzej Stasiak & Stephen C. West, 1999. "Binding of double-strand breaks in DNA by human Rad52 protein," Nature, Nature, vol. 398(6729), pages 728-731, April.
    4. Alex N. Zelensky & Joost Schimmel & Hanneke Kool & Roland Kanaar & Marcel Tijsterman, 2017. "Inactivation of Pol θ and C-NHEJ eliminates off-target integration of exogenous DNA," Nature Communications, Nature, vol. 8(1), pages 1-7, December.
    5. 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.
    6. Yaqun Teng & Tribhuwan Yadav & Meihan Duan & Jun Tan & Yufei Xiang & Boya Gao & Jianquan Xu & Zhuobin Liang & Yang Liu & Satoshi Nakajima & Yi Shi & Arthur S. Levine & Lee Zou & Li Lan, 2018. "ROS-induced R loops trigger a transcription-coupled but BRCA1/2-independent homologous recombination pathway through CSB," Nature Communications, Nature, vol. 9(1), pages 1-12, December.
    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. Yuxin Huang & Wenjing Li & Tzeh Foo & Jae-Hoon Ji & Bo Wu & Nozomi Tomimatsu & Qingming Fang & Boya Gao & Melissa Long & Jingfei Xu & Rouf Maqbool & Bipasha Mukherjee & Tengyang Ni & Salvador Alejo & , 2024. "DSS1 restrains BRCA2’s engagement with dsDNA for homologous recombination, replication fork protection, and R-loop homeostasis," Nature Communications, Nature, vol. 15(1), pages 1-17, December.
    2. Alba Muniesa-Vargas & Carlota Davó-Martínez & Cristina Ribeiro-Silva & Melanie van der Woude & Karen L. Thijssen & Ben Haspels & David Häckes & Ülkem U. Kaynak & Roland Kanaar & Jurgen A. Marteijn & A, 2024. "Persistent TFIIH binding to non-excised DNA damage causes cell and developmental failure," Nature Communications, Nature, vol. 15(1), pages 1-16, December.
    3. 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.
    4. 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.
    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. 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.
    8. 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.
    9. 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.
    10. 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.
    11. Duško Lainšček & Vida Forstnerič & Veronika Mikolič & Špela Malenšek & Peter Pečan & Mojca Benčina & Matjaž Sever & Helena Podgornik & Roman Jerala, 2022. "Coiled-coil heterodimer-based recruitment of an exonuclease to CRISPR/Cas for enhanced gene editing," Nature Communications, Nature, vol. 13(1), pages 1-12, December.
    12. Sundaram Acharya & Asgar Hussain Ansari & Prosad Kumar Das & Seiichi Hirano & Meghali Aich & Riya Rauthan & Sudipta Mahato & Savitri Maddileti & Sajal Sarkar & Manoj Kumar & Rhythm Phutela & Sneha Gul, 2024. "PAM-flexible Engineered FnCas9 variants for robust and ultra-precise genome editing and diagnostics," Nature Communications, Nature, vol. 15(1), pages 1-23, December.
    13. Afsaneh Sadremomtaz & Robert F. Glass & Jorge Eduardo Guerrero & Dennis R. LaJeunesse & Eric A. Josephs & Reza Zadegan, 2023. "Digital data storage on DNA tape using CRISPR base editors," Nature Communications, Nature, vol. 14(1), pages 1-10, December.
    14. Manisha Jalan & Aman Sharma & Xin Pei & Nils Weinhold & Erika S. Buechelmaier & Yingjie Zhu & Sana Ahmed-Seghir & Abhirami Ratnakumar & Melody Bona & Niamh McDermott & Joan Gomez-Aguilar & Kyrie S. An, 2024. "RAD52 resolves transcription-replication conflicts to mitigate R-loop induced genome instability," Nature Communications, Nature, vol. 15(1), pages 1-17, December.
    15. Haibo Yang & Emily M. Lachtara & Xiaojuan Ran & Jessica Hopkins & Parasvi S. Patel & Xueping Zhu & Yao Xiao & Laiyee Phoon & Boya Gao & Lee Zou & Michael S. Lawrence & Li Lan, 2023. "The RNA m5C modification in R-loops as an off switch of Alt-NHEJ," Nature Communications, Nature, vol. 14(1), pages 1-15, December.
    16. Xuan Zhang & Jun Xu & Jing Hu & Sitao Zhang & Yajing Hao & Dongyang Zhang & Hao Qian & Dong Wang & Xiang-Dong Fu, 2024. "Cockayne Syndrome Linked to Elevated R-Loops Induced by Stalled RNA Polymerase II during Transcription Elongation," Nature Communications, Nature, vol. 15(1), pages 1-16, December.
    17. 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.
    18. Martin Andrs & Henriette Stoy & Barbora Boleslavska & Nagaraja Chappidi & Radhakrishnan Kanagaraj & Zuzana Nascakova & Shruti Menon & Satyajeet Rao & Anna Oravetzova & Jana Dobrovolna & Kalpana Surend, 2023. "Excessive reactive oxygen species induce transcription-dependent replication stress," Nature Communications, Nature, vol. 14(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. Yumin Wang & Boya Gao & Luyuan Zhang & Xudong Wang & Xiaolan Zhu & Haibo Yang & Fengqi Zhang & Xueping Zhu & Badi Zhou & Sean Yao & Aiko Nagayama & Sanghoon Lee & Jian Ouyang & Siang-Boon Koh & Eric L, 2024. "Meiotic protein SYCP2 confers resistance to DNA-damaging agents through R-loop-mediated DNA repair," Nature Communications, Nature, vol. 15(1), pages 1-17, 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:15:y:2024:i:1:d:10.1038_s41467-024-49385-9. 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.