Recently, the research team led by Wei Li and Chao Liu from Guangzhou Medical University Affiliated Women and Children's Medical Center, in collaboration with Fei Gao's team from the Institute of Zoology, Chinese Academy of Sciences, and Meijia Zhang's team from South China University of Technology, published a paper titled "Dynamic R-loops at centromeres ensure chromosome alignment during oocyte meiotic divisions in mice" in Science Bulletin. The study unveiled the critical role of centromeric R-loop homeostasis in completing oocyte meiotic divisions. Furthermore, the study also clarified the mechanism by which centromeric R-loops regulate spindle assembly and chromosome alignment during oocyte meiotic divisions.
The study revealed that R-loops and their regulator RNase H1 are present at centromeres during oocyte meiotic divisions. Both centromeric R-loops and RNase H1 exhibited dynamic changes during this process, peaking at the first meiotic metaphase (MI), suggesting their critical roles in oocyte meiotic maturation. Oocyte-specific knockout of Rnaseh1 in mice caused severe maturation defects, characterized by abnormal spindle assembly and chromosome misalignment. Furthermore, Rnaseh1-knockout oocytes failed to progress through early embryonic development after in vitro fertilization, ultimately resulting in female infertility.
During oocyte maturation, Aurora B kinase regulates the stability of kinetochore-microtubule attachments by phosphorylating kinetochore proteins such as HEC1. Dysregulation of Aurora B activity leads to abnormal spindle assembly and chromosome misalignment. Both knockout and overexpression of RNase H1 disrupt centromeric R-loop homeostasis and aberrant centromeric R-loop levels significantly impair Aurora B recruitment and activation at centromeres. As a result, the p-HEC1 S55 level is dysregulated on the kinetochores, ultimately leading to spindle assembly defects and chromosomal misalignment.
Further studies revealed that the levels of ATR-ATRIP complexes at centromeres are correlated with R-loop levels. ATR is recruited to centromeres through its interaction with the ATRIP-RPA complex and activates the catalytic activity of CHK1 by phosphorylating its Ser317 and Ser345 residues. Activated CHK1 subsequently promotes the activation of Aurora B, which in turn regulates kinetochore-microtubule attachments. Treatment of Rnaseh1-knockout oocytes with specific inhibitors targeting ATR, CHK1, and Aurora B partially rescued the meiotic defects. Thus, R-loops facilitate spindle assembly and chromosome alignment by modulating the centromeric ATR-CHK1-Aurora B pathway during oocyte meiotic divisions.
These results indicate that centromeric R-loops are crucial for spindle assembly and chromosome alignment during oocyte meiotic divisions. This study first elucidates the function and mechanism of centromeric R-loop homeostasis during oocyte meiotic divisions. It reveals an R-loop-driven signaling pathway, RPA-ATRIP-ATR-CHK1-Aurora B, which promotes spindle assembly and chromosome alignment during oocyte meiotic maturation in mice. This study expands the current understanding of the biological functions of centromeric R-loops, deepens the knowledge of the regulatory mechanisms of oocyte meiotic maturation, and provides new insights for the diagnosis and treatment of clinical disorders related to oocyte maturation.
Professor Wei Li and Professor Chao Liu from Guangzhou Medical University Affiliated Women and Children's Medical Center, Professor Fei Gao from the Institute of Zoology, Chinese Academy of Sciences, and Professor Meijia Zhang from South China University of Technology are the co-corresponding authors of this article. Dr. Yinghong Chen (graduated), a former Ph.D. student at the Institute of Zoology, Chinese Academy of Sciences, Dr. Liying Wang, a postdoctoral fellow at Guangzhou Medical University, and Qiuxing Zhou, a master's student at Guangzhou Medical University, are the co-first authors. This work was supported by the Key Program of the National Natural Science Foundation of China and the National Science Fund for Distinguished Young Scholars of China.
