Beijing Advanced Innovation Center for Structural Biology & Frontier Research Center for Biological Structure, School of Life Sciences, Tsinghua University, Beijing 100084, China; Tsinghua-Peking Center for Life Sciences, School of Life Sciences, Tsinghua University, Beijing 100084, China.
State Key Laboratory of Stem Cell and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China; University of Chinese Academy of Sciences, Beijing 100049, China.
Cell. 2023 Jun 22;186(13):2865-2879.e20. doi: 10.1016/j.cell.2023.05.032. Epub 2023 Jun 9.
Retroelements are the widespread jumping elements considered as major drivers for genome evolution, which can also be repurposed as gene-editing tools. Here, we determine the cryo-EM structures of eukaryotic R2 retrotransposon with ribosomal DNA target and regulatory RNAs. Combined with biochemical and sequencing analysis, we reveal two essential DNA regions, Drr and Dcr, required for recognition and cleavage. The association of 3' regulatory RNA with R2 protein accelerates the first-strand cleavage, blocks the second-strand cleavage, and initiates the reverse transcription starting from the 3'-tail. Removing 3' regulatory RNA by reverse transcription allows the association of 5' regulatory RNA and initiates the second-strand cleavage. Taken together, our work explains the DNA recognition and RNA supervised sequential retrotransposition mechanisms by R2 machinery, providing insights into the retrotransposon and application reprogramming.
逆转录元件是广泛存在的跳跃元件，被认为是基因组进化的主要驱动因素，也可以被重新用作基因编辑工具。在这里，我们确定了具有核糖体 DNA 靶标和调节 RNA 的真核 R2 逆转录转座子的冷冻电镜结构。结合生化和测序分析，我们揭示了两个必需的 DNA 区域 Drr 和 Dcr，它们是识别和切割所必需的。3' 调节 RNA 与 R2 蛋白的结合加速了第一条链的切割，阻止了第二条链的切割，并从 3'-尾开始启动逆转录。通过逆转录去除 3' 调节 RNA 允许 5' 调节 RNA 的结合，并启动第二条链的切割。总之，我们的工作解释了 R2 机制的 DNA 识别和 RNA 监督的顺序逆转录转座机制，为逆转录转座子和应用程序重新编程提供了深入的了解。
