Structural Insights into Branch Site Proofreading by Human Spliceosome

• Published in: bioRxiv
• Main Authors: Zhang, Xiaofeng, Zhan, Xiechao, Bian, Tong, Yang, Fenghua, Pan, Li, Lu, Yichen, Xing, Zhihan, Qiangfeng Cliff Zhang, Shi, Yigong
• Format: Paper
• Language: English
• Published: Cold Spring Harbor Cold Spring Harbor Laboratory Press 07.11.2022
• Subjects: Conformation; DNA helicase; Editing; mRNA; Proofreading; Ribonucleoproteins (small nuclear); Ribonucleoproteins (U2 small nuclear); RNA helicase; snRNA; Splicing factors
• DOI: 10.1101/2022.11.07.515429

Selection of the pre-mRNA branch site (BS) by U2 snRNP is crucial to prespliceosome (A complex) assembly. The RNA helicase PRP5 proofreads BS selection; but the underlying mechanism remains unclear. Here we report the atomic structures of two sequential complexes leading to prespliceosome assembly: human 17S U2 snRNP and a cross-exon pre-A complex. PRP5 is anchored on 17S U2 snRNP mainly through occupation of the RNA path of SF3B1 by an acidic loop of PRP5; the helicase domain of PRP5 associates with U2 snRNA; the BS-interacting stem loop (BSL) of U2 snRNA is shielded by the splicing factor TAT-SF1, unable to engage the BS. In the pre-A complex, an initial U2/BS duplex is formed; the translocated helicase domain of PRP5 stays with U2 snRNA; the acidic loop still occupies the RNA path. The pre-A conformation is specifically stabilized by the splicing factors SF1, DNAJC8 and SF3A2. Cancer-derived mutations in SF3B1 damage its association with PRP5, compromising BS proofreading. Together, these findings reveal key insights into prespliceosome assembly and BS selection/proofreading by PRP5.Competing Interest StatementThe authors have declared no competing interest.