The U1 antisense morpholino oligonucleotide (AMO) disrupts U1 snRNP structure to promote intronic PCPA modification of pre-mRNAs.
The U1 antisense morpholino oligonucleotide (AMO) disrupts U1 snRNP structure to promote intronic PCPA modification of pre-mRNAs.
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DOI：
10.1016/j.jbc.2023.104854
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发表时间：
2023-07
影响因子：
4.8
通讯作者：
Yao, Chengguo
中科院分区：
文献类型：
--
作者：
Feng, Qiumin;Lin, Zejin;Deng, Yanhui;Ran, Yi;Yu, Rui;Xiang, Andy Peng;Ye, Congting;Yao, Chengguo
用25 nt U1 AMO（反义吗啉代寡核苷酸）对U1小核核糖核蛋白（snRNP）进行功能性缺失可能导致数千个基因的内含子过早切割和多聚腺苷酸化，这种现象称为U1 snRNP转录;然而，其潜在机制仍然难以捉摸。在这项研究中，我们证明了U1 AMO可以在体外和体内破坏U1 snRNP结构，从而影响U1 snRNP-RNAP聚合酶II的相互作用。通过对RNAP聚合酶II的最大亚基RPB 1的C-末端结构域的Ser 2和Ser 5的磷酸化进行染色质免疫沉淀测序，我们发现转录延伸在U1 AMO处理后受到干扰，在内含子隐蔽聚腺苷酸化位点（PAS）处具有特别高的Ser 2信号磷酸化。此外，我们还发现核心3 '加工因子CPSF/CstF参与了内含子隐蔽PAS的加工。如染色质免疫沉淀测序和单个核苷酸解析交联和免疫沉淀测序分析所示，在U1 AMO处理后，它们的募集向隐蔽的PAS积累。总之，我们的数据表明，破坏U1 snRNP结构介导的U1 AMO提供了一个关键的理解U1转录机制。
Functional depletion of the U1 small nuclear ribonucleoprotein (snRNP) with a 25 nt U1 AMO (antisense morpholino oligonucleotide) may lead to intronic premature cleavage and polyadenylation of thousands of genes, a phenomenon known as U1 snRNP telescripting; however, the underlying mechanism remains elusive. In this study, we demonstrated that U1 AMO could disrupt U1 snRNP structure both in vitro and in vivo, thereby affecting the U1 snRNP–RNAP polymerase II interaction. By performing chromatin immunoprecipitation sequencing for phosphorylation of Ser2 and Ser5 of the C-terminal domain of RPB1, the largest subunit of RNAP polymerase II, we showed that transcription elongation was disturbed upon U1 AMO treatment, with a particular high phosphorylation of Ser2 signal at intronic cryptic polyadenylation sites (PASs). In addition, we showed that core 3′processing factors CPSF/CstF are involved in the processing of intronic cryptic PAS. Their recruitment accumulated toward cryptic PASs upon U1 AMO treatment, as indicated by chromatin immunoprecipitation sequencing and individual-nucleotide resolution CrossLinking and ImmunoPrecipitation sequencing analysis. Conclusively, our data suggest that disruption of U1 snRNP structure mediated by U1 AMO provides a key for understanding the U1 telescripting mechanism.
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影响因子：
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作者：
Kondo Y;Oubridge C;van Roon AM;Nagai K
通讯作者：
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3'RNA聚合酶II CTD上Ser2的前MRNA的末端形成和在人类细胞中相互耦合。
影响因子：
10.5
作者：
Davidson L;Muniz L;West S
通讯作者：
West S