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In vitro reconstitution of meiotic DNA double-strand-break formation

Author

Listed:
  • Xinzhe Tang

    (University of Chinese Academy of Sciences)

  • Zetao Hu

    (Shanghai Jiao Tong University School of Medicine
    Fudan University)

  • Jian Ding

    (University of Chinese Academy of Sciences)

  • Meixia Wu

    (University of Chinese Academy of Sciences)

  • Pin Guan

    (University of Chinese Academy of Sciences)

  • Yawei Song

    (University of Chinese Academy of Sciences)

  • Yue Yin

    (Chinese Academy of Sciences)

  • Wei Wu

    (University of Chinese Academy of Sciences)

  • Jinbiao Ma

    (Fudan University)

  • Ying Huang

    (Shanghai Jiao Tong University School of Medicine)

  • Ming-Han Tong

    (University of Chinese Academy of Sciences)

Abstract

The Spo11 complex catalyses the formation of DNA double-strand breaks (DSBs), initiating meiotic recombination—a process that is essential for fertility and genetic diversity1,2. Although the function of Spo11 has been known for 27 years, previous efforts to reconstitute DSB formation in vitro have been unsuccessful. Here we biochemically characterize the mouse SPO11–TOP6BL protein complex, and show that this complex cleaves DNA and covalently attaches to the 5′ terminus of DNA breaks in vitro. Using a point-mutation strategy, we reveal that Mg2+ is essential for the DNA-cleavage activity of this complex in vitro, as confirmed by knock-in mice carrying a point mutation in SPO11 that disrupts its binding to Mg2+, thereby abolishing DSB formation. However, the activity of the SPO11 complex is ATP-independent. We also present evidence that the mouse SPO11 complex is biochemically distinct from the ancestral topoisomerase VI. Our findings establish a mechanistic framework for understanding the first steps of meiotic recombination.

Suggested Citation

  • Xinzhe Tang & Zetao Hu & Jian Ding & Meixia Wu & Pin Guan & Yawei Song & Yue Yin & Wei Wu & Jinbiao Ma & Ying Huang & Ming-Han Tong, 2025. "In vitro reconstitution of meiotic DNA double-strand-break formation," Nature, Nature, vol. 639(8055), pages 800-807, March.
    • Handle: RePEc:nat:nature:v:639:y:2025:i:8055:d:10.1038_s41586-024-08551-1
    DOI: 10.1038/s41586-024-08551-1
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