Direct analysis of nematode cis- and trans-spliceosomes: a functional role for U5 snRNA in spliced leader addition trans-splicing and the identification of novel Sm snRNPs.
Direct analysis of nematode cis- and trans-spliceosomes: a functional role for U5 snRNA in spliced leader addition trans-splicing and the identification of novel Sm snRNPs.
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发表时间：
1996-08
期刊：
影响因子：
4.5
通讯作者：
P. Maroney;Yi-Tao Yu;Yi-Tao Yu;Marzena Jankowska;Timothy W. Nilsen
中科院分区：
文献类型：
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作者：
P. Maroney;Yi-Tao Yu;Yi-Tao Yu;Marzena Jankowska;Timothy W. Nilsen
线虫中的大多数核前mRNA通过顺式和反式剪接进行加工。在反式剪接中，5'末端外显子，剪接的前导序列（SL），衍生自反式剪接特异性Sm snRNP，SL RNP。由于U snRNP是反式剪接所需的辅因子，并且由于该加工反应通过与顺式剪接相同的两步反应途径进行，因此长期以来一直认为反式剪接在类似于顺式剪接体的复合物中被催化。然而，顺式和反式剪接体之间的相似性或差异尚未建立。特别是，U 5 snRNP在反式剪接中的作用尚不清楚。在这里，我们已经使用亲和选择分析U snRNA成分的线虫顺式和反式剪接体。我们发现，U 5 snRNP是一个完整的组成部分的反式剪接体，并使用位点特异性交联，我们表明，U 5 snRNP建立特定的相互作用与SL RNA外显子。我们还确定了两个新的Sm snRNP，丰富的顺式和反式剪接体。最后，我们提供的证据表明，SL RNP含有多snRNP（SL，U4，U 5和U6 RNP）可能是一个功能性前体在trans-spliceosome组装。
Most nuclear pre-mRNAs in nematodes are processed by both cis- and trans-splicing. In trans-splicing, the 5' terminal exon, the spliced leader sequence (SL), is derived from a trans-splicing specific Sm snRNP, the SL RNP. Because U snRNPs are required cofactors for trans-splicing, and because this processing reaction proceeds via a two-step reaction pathway identical to that of cis-splicing, it has long been assumed that trans-splicing is catalyzed in a complex analogous to the cis-spliceosome. However, similarities or differences between cis- and trans-spliceosomes have not been established. In particular, the role of U5 snRNP in trans-splicing has been unclear. Here, we have used affinity selection to analyze the U snRNA constituents of nematode cis- and trans-spliceosomes. We find that U5 snRNP is an integral component of the trans-spliceosome and, using site-specific crosslinking, we show that U5 snRNP establishes specific Interactions with the SL RNA exon. We also identify two novel Sm snRNPs that are enriched in both cis- and trans-spliceosomes. Finally, we provide evidence that a SL RNP-containing multi-snRNP (SL, U4, U5, and U6 RNPs) may be a functional precursor in trans-spliceosome assembly.