Herbert Austin, Hatfield Abigail, Randazza Alexandra, Miyamoto Valeria, Palmer Katie, Lackey Lela
Department of Genetics and Biochemistry, Center for Human Genetics, Clemson University.
bioRxiv. 2025 Feb 22:2025.02.19.638873. doi: 10.1101/2025.02.19.638873.
SF3B1 is a core component of the spliceosome involved in branch point recognition and 3' splice site selection. SF3B1 mutation is common in myelodysplastic syndrome and other blood disorders. The most common mutation in SF3B1 is K700E, a lysine to glutamic acid change within the pre-mRNA interacting heat repeat domain. A hallmark of SF3B1 mutation is an increased use of cryptic 3' splice sites; however, the properties distinguishing SF3B1-sensitive splice junctions from other alternatively spliced junctions are unknown. We identify a subset of 192 core splice junctions that are mis-spliced with SF3B1 K700E mutation. We use our core set to test whether SF3B1-sensitive splice sites are different from control cryptic 3' splice sites via RNA structural accessibility. As a comparison, we define a set of SF3B1-resistant splice junctions with cryptic splice site use that does not change with SF3B1 K700E mutation. We find sequence differences between SF3B1-sensitive and SF3B1-resistant junctions, particularly at the cryptic sites. SF3B1-sensitive cryptic 3' splice sites are within an extended polypyrimidine tract and have lower splice site strength scores. We develop experimental RNA structure data for 83 SF3B1-sensitive junctions and 39 SF3B1-resistant junctions. We find that the pattern of structural accessibility at the NAG splicing motif in cryptic and canonical 3' splice sites is similar. In addition, this pattern can be found in both SF3B1-resistant and SF3B1-sensitive junctions. However, SF3B1-sensitive junctions have cryptic splice sites that are less structurally distinct from the canonical splice sites. In addition, SF3B1-sensitive splice junctions are overall more flexible than SF3B1-resistant junctions. Our results suggest that the SF3B1-sensitive splice junctions have unique structure and sequence properties, containing poorly differentiated, weak splice sites that lead to altered 3' splice site recognition in the presence of SF3B1 mutation.
SF3B1是剪接体的核心组成部分，参与分支点识别和3'剪接位点选择。SF3B1突变在骨髓增生异常综合征和其他血液疾病中很常见。SF3B1中最常见的突变是K700E，即前体mRNA相互作用热重复结构域内赖氨酸到谷氨酸的变化。SF3B1突变的一个标志是隐蔽3'剪接位点的使用增加；然而，区分SF3B1敏感剪接连接与其他可变剪接连接的特性尚不清楚。我们鉴定出192个核心剪接连接的子集，它们在SF3B1 K700E突变时发生错误剪接。我们使用我们的核心集通过RNA结构可及性来测试SF3B1敏感剪接位点是否与对照隐蔽3'剪接位点不同。作为比较，我们定义了一组SF3B1抗性剪接连接，其隐蔽剪接位点的使用不会随SF3B1 K700E突变而改变。我们发现SF3B1敏感和抗性连接之间存在序列差异，特别是在隐蔽位点。SF3B1敏感的隐蔽3'剪接位点位于延伸的聚嘧啶序列内，并且具有较低的剪接位点强度分数。我们为83个SF3B1敏感连接和39个SF3B1抗性连接开发了实验性RNA结构数据。我们发现隐蔽和经典3'剪接位点处NAG剪接基序的结构可及性模式相似。此外，这种模式在SF3B1抗性和敏感连接中均可见。然而，SF3B1敏感连接的隐蔽剪接位点在结构上与经典剪接位点的差异较小。此外，SF3B1敏感剪接连接总体上比SF3B1抗性连接更灵活。我们的结果表明，SF3B1敏感剪接连接具有独特的结构和序列特性，包含分化不良、较弱的剪接位点，这些位点在存在SF3B1突变时会导致3'剪接位点识别改变。
