An extended U2AF(65)-RNA-binding domain recognizes the 3' splice site signal.
An extended U2AF(65)-RNA-binding domain recognizes the 3' splice site signal.
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DOI：
10.1038/ncomms10950
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发表时间：
2016-03-08
影响因子：
16.6
通讯作者：
Kielkopf CL
中科院分区：
文献类型：
--
作者：
Agrawal AA;Salsi E;Chatrikhi R;Henderson S;Jenkins JL;Green MR;Ermolenko DN;Kielkopf CL
前体mRNA剪接因子U2 AF 65如何识别人类基因转录本中主要3′剪接位点的多聚嘧啶信号，目前尚不完全清楚。我们确定了四个结构的扩展U2 AF 65-RNA结合域结合Py-tract寡核苷酸在2.0和1.5之间的分辨率。这些结构与RNA结合和剪接分析揭示了U2 AF 65域间残基在识别连续的九核苷酸Py道中的不可预见的作用。双RNA识别基序（RRM）之间的U2 AF 65接头残基识别中心核苷酸，而N-和C-末端RRM延伸识别3′末端和第三个核苷酸。单分子FRET实验表明，构象选择和诱导适合的U2 AF 65 RRM的Py-tract协会的互补机制。总之，这些结果推进了对剪接位点信号的主要类别的分子识别的机械理解。 前体mRNA剪接因子U2 AF 65识别人类基因转录本中的3′剪接位点，但其细节尚不完全清楚。在这里，作者报告了U2 AF 65结构和单分子FRET，揭示了剪接位点识别的机制。
How the essential pre-mRNA splicing factor U2AF65 recognizes the polypyrimidine (Py) signals of the major class of 3′ splice sites in human gene transcripts remains incompletely understood. We determined four structures of an extended U2AF65–RNA-binding domain bound to Py-tract oligonucleotides at resolutions between 2.0 and 1.5 Å. These structures together with RNA binding and splicing assays reveal unforeseen roles for U2AF65 inter-domain residues in recognizing a contiguous, nine-nucleotide Py tract. The U2AF65 linker residues between the dual RNA recognition motifs (RRMs) recognize the central nucleotide, whereas the N- and C-terminal RRM extensions recognize the 3′ terminus and third nucleotide. Single-molecule FRET experiments suggest that conformational selection and induced fit of the U2AF65 RRMs are complementary mechanisms for Py-tract association. Altogether, these results advance the mechanistic understanding of molecular recognition for a major class of splice site signals. The pre-mRNA splicing factor U2AF65 recognizes 3′ splice sites in human gene transcripts, but the details are not fully understood. Here, the authors report U2AF65 structures and single molecule FRET that reveal mechanistic insights into splice site recognition.