trans splicing of polycistronic Caenorhabditis elegans Pre-mRNAs:: Analysis of the SL2 RNA
trans splicing of polycistronic Caenorhabditis elegans Pre-mRNAs:: Analysis of the SL2 RNA
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DOI：
10.1128/mcb.20.18.6659-6667.2000
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发表时间：
2000-09-01
影响因子：
5.3
通讯作者：
Blumenthal, T
中科院分区：
文献类型：
--
作者：
Evans, D;Blumenthal, T
秀丽隐杆线虫操纵子中的基因被转录为多顺反子前mRNA，其中下游基因产物被反式剪接成专门的剪接前导序列SL2。SL2是由110个核苷酸的RNA，SL2 RNA，作为Sm结合的snRNP存在于细胞中。SL2 RNA在概念上可以折叠成一个遗传上保守的三茎环二级结构。在这里，我们报告的SL2 RNA的体内突变分析。一些序列可以改变而没有后果，而其他变化导致反式剪接的大量损失。有趣的是，剪接的前导序列本身可以被显著改变，使得第一个茎环不能形成，只有相对较小的反式剪接效率损失。然而，茎II的一级序列对于SL2反式剪接至关重要。类似地，第三茎环的保守一级序列在反式剪接中起关键作用。虽然茎环III中的突变允许snRNP形成，但环中的单核苷酸取代阻止反式剪接。相反，SL1 RNA的类似区域不是高度保守的，并且其突变不废除功能。因此，茎环III似乎赋予SL2 RNA特异性功能。最后，一个上游序列，以前预测是一个近端序列元件，显示需要SL2 RNA表达。
Genes in Caenorhabditis elegans operons are transcribed as polycistronic pre-mRNAs in which downstream gene products are trans spliced to a specialized spliced leader, SL2. SL2 is donated by a 110-nucleotide RNA, SL2 RNA, present in the cell as an Sm-bound snRNP. SL2 RNA can be conceptually folded into a phylogenetically conserved three-stem-loop secondary structure. Here we report an in vivo mutational analysis of the SL2 RNA. Some sequences can be changed without consequence, while other changes result in a substantial loss of trans splicing. Interestingly, the spliced leader itself can be dramatically altered, such that the first stem-loop cannot form, with only a relatively small loss in trans-splicing efficiency. However, the primary sequence of stem II is crucial for SL2 trans splicing. Similarly, the conserved primary sequence of the third stem-loop plays a key role in trans splicing. While mutations in stem-loop III allow snRNP formation, a single nucleotide substitution in the loop prevents trans splicing. In contrast, the analogous region of SL1 RNA is not highly conserved, and its mutation does not abrogate function. Thus, stem-loop III appears to confer a specific function to SL2 RNA. Finally, an upstream sequence, previously predicted to be a proximal sequence element, is shown to be required for SL2 RNA expression.