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首页> 外文期刊>Molecular and Cellular Biology >The U1 snRNP Base Pairs with the 5′ Splice Site within a Penta-snRNP Complex
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The U1 snRNP Base Pairs with the 5′ Splice Site within a Penta-snRNP Complex

机译：在Penta-snRNP复合物中的U1 snRNP碱基与5'剪接位点配对

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Recognition of the 5′ splice site is an important step in mRNA splicing. To examine whether U1 approaches the 5′ splice site as a solitary snRNP or as part of a multi-snRNP complex, we used a simplified in vitro system in which a short RNA containing the 5′ splice site sequence served as a substrate in a binding reaction. This system allowed us to study the interactions of the snRNPs with the 5′ splice site without the effect of other cis-regulatory elements of precursor mRNA. We found that in HeLa cell nuclear extracts, five spliceosomal snRNPs form a complex that specifically binds the 5′ splice site through base pairing with the 5′ end of U1. This system can accommodate RNA-RNA rearrangements in which U5 replaces U1 binding to the 5′ splice site, a process that occurs naturally during the splicing reaction. The complex in which U1 and the 5′ splice site are base paired sediments in the 200S fraction of a glycerol gradient together with all five spliceosomal snRNPs. This fraction is functional in mRNA spliceosome assembly when supplemented with soluble nuclear proteins. The results argue that U1 can bind the 5′ splice site in a mammalian preassembled penta-snRNP complex.
机译：5'剪接位点的识别是mRNA剪接中的重要步骤。为了检查U1是作为单独的snRNP还是作为多snRNP复合体的一部分接近5'剪接位点，我们使用了简化的体外系统，其中包含5'剪接位点序列的短RNA作为结合物中的底物反应。该系统使我们能够研究snRNPs与5'剪接位点的相互作用，而不受前体mRNA其他 cis 调控元件的影响。我们发现，在HeLa细胞核提取物中，五个剪接体snRNPs形成一个复合物，该复合物通过与U1的5'末端碱基配对而特异性结合5'剪接位点。该系统可适应RNA-RNA重排，其中U5取代了U1与5'剪接位点的结合，这一过程在剪接反应过程中自然发生。 U1和5'剪接位点是甘油梯度的200S部分中碱基配对的沉积物以及所有五个剪接体snRNP的复合体。补充可溶性核蛋白后，该馏分在mRNA剪接体组装中起作用。结果表明，U1可以结合哺乳动物预组装的penta-snRNP复合物中的5'剪接位点。

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