Wang Liu, Xie Jiahao, Zhang Chong, Zou Jian, Huang Zirui, Shang Sitong, Chen Xingyu, Yang Yang, Liu Jianquan, Dong Haohao, Huang Dingming, Su Zhaoming
The State Key Laboratory of Biotherapy, National Clinical Research Center for Geriatrics, West China Hospital; The State Key Laboratory of Oral Diseases, National Clinical Research Center for Oral Diseases, National Center for Stomatology, West China Hospital of Stomatology, Sichuan University, Chengdu, China.
Department of Cariology and Endodontics, West China Hospital of Stomatology, Sichuan University, Chengdu, China.
Nat Struct Mol Biol. 2025 Jan 31. doi: 10.1038/s41594-025-01484-x.
Circularly permuted group II introns (CP introns) consist of rearranged structural domains separated by two tethered exons, generating branched introns and circular exons via back-splicing. Structural and mechanistic understanding of circular RNA (circRNA) generation by CP introns remains elusive. We resolve cryo-electron microscopy structures of a natural CP intron in different states during back-splicing at a resolution of 2.5-2.9 Å. Domain 6 (D6) undergoes a conformational change of 65° after branching, to facilitate 3'-exon recognition and circularization. Previously unseen tertiary interactions compact the catalytic triad and D6 for splicing without protein, whereas a metal ion, M, is observed to stabilize the 5'-exon during splicing. While these unique features were not observed in canonical group II introns and spliceosomes, they are common in CP introns, as demonstrated by the cryo-EM structure of another CP intron discovered by comparative genomics analysis. Our results elucidate the mechanism of CP intron back-splicing dynamics, with potential applications in circRNA research and therapeutics.
环形排列的II组内含子（CP内含子）由被两个相连外显子隔开的重排结构域组成，通过反向剪接产生分支内含子和环状外显子。目前，对于CP内含子产生环状RNA（circRNA）的结构和机制仍不清楚。我们以2.5-2.9埃的分辨率解析了天然CP内含子在反向剪接不同状态下的冷冻电镜结构。分支后，结构域6（D6）发生65°的构象变化，以促进3'-外显子的识别和环化。以前未见过的三级相互作用使催化三联体和D6紧密结合，以便在没有蛋白质的情况下进行剪接，而观察到一个金属离子M在剪接过程中稳定5'-外显子。虽然在经典的II组内含子和剪接体中未观察到这些独特特征，但正如通过比较基因组学分析发现的另一个CP内含子的冷冻电镜结构所表明的那样，它们在CP内含子中很常见。我们的结果阐明了CP内含子反向剪接动力学的机制，在circRNA研究和治疗中具有潜在应用。
