In a study published in Nature, researchers led by Prof. TONG Minghan from the Center for Excellence in Molecular Cell Science (Shanghai Institute of Biochemistry and Cell Biology) of the Chinese Academy of Sciences, collaborating with Prof. HUANG Ying from Xinhua Hospital, Shanghai Jiaotong University School of Medicine, reported the successful in vitro reconstitution of meiotic DNA double-strand breaks (DSBs) formation.
Meiosis, a specialized form of cell division in most sexually reproducing organisms, serves as the basis of sexual reproduction. Homologous recombination-a crucial meiotic event-not only enhances genetic diversity through exchanging parental genetic substance but also establishes physical connections between homologous chromosomes to ensure their precise segregation.
This process is initiated by the formation of programmed DSBs, catalyzed by Spo11, a discovery dating back to 1997. However, despite nearly three decades of efforts, reconstituting DSB formation in vitro has remained an unsolved challenge.
In this study, researchers expressed and purified the SPO11-TOP6BL complex using Expi293F cells. They demonstrated that this complex cleaves DNA and covalently attaches to the 5' terminus of DNA breaks in vitro, successfully reconstituting meiotic DSB formation in vitro. This resolved the long-standing question in meiotic recombination.
Using a point-mutation strategy, researchers revealed that Mg2+ is essential for the complex's DNA cleavage activity. Knock-in mice carrying a SPO11 point-mutation that disrupts Mg2+ binding exhibited a complete loss of DSB formation. Interestingly, the activity of the SPO11 complex functions independently of ATP, distinct from its ancestral enzyme, topoisomerase VI.
This study represents a major breakthrough in understanding meiotic recombination. It, provides a powerful platform for dissecting the molecular mechanisms of meiotic homologous recombination.
