Zorio D A, Lea K, Blumenthal T
Department of Biology, Indiana University, Bloomington 47405, USA.
Mol Cell Biol. 1997 Feb;17(2):946-53. doi: 10.1128/MCB.17.2.946.
The U2 small nuclear ribonucleoprotein particle (snRNP) auxiliary factor, U2AF, is an essential splicing factor required for recognition of the polypyrimidine tract and subsequent U2 snRNP assembly at the branch point. Because Caenorhabditis elegans introns lack both polypyrimidine tract and branch point consensus sequences but have a very highly conserved UUUUCAG/R consensus at their 3' splice sites, we hypothesized that U2AF might serve to recognize this sequence and thus promote intron recognition in C. elegans. Here we report the cloning of the gene for the large subunit of U2AF, uaf-1. Three classes of cDNA were identified. In the most abundant class the open reading frame is similar to that for the U2AF65 from mammals and flies. The remaining two classes result from an alternative splicing event in which an exon containing an in-frame stop codon is inserted near the beginning of the second RNA recognition motif. However, this alternative mRNA is apparently not translated. Interestingly, the inserted exon contains 10 matches to the 3' splice site consensus. To determine whether this feature is conserved, we sequenced uaf-1 from the related nematode Caenorhabditis briggsae. It is composed of six exons, including an alternatively spliced third exon interrupting the gene at the same location as in C. elegans. uaf-1 is contained in an operon with the rab-18 gene in both species. Although the alternative exons from the two species are not highly conserved and would not encode related polypeptides, the C. briggsae alternative exon has 18 matches to the 3' splice site consensus. We hypothesize that the array of 3' splice site-like sequences in the pre-mRNA and alternatively spliced exon may have a regulatory role. The alternatively spliced RNA accumulates at high levels following starvation, suggesting that this RNA may represent an adaption for reducing U2AF65 levels when pre-mRNA levels are low.
U2小核核糖核蛋白颗粒（snRNP）辅助因子U2AF是识别多嘧啶序列以及随后在分支点组装U2 snRNP所必需的剪接因子。由于秀丽隐杆线虫内含子既缺乏多嘧啶序列也缺乏分支点共有序列，但在其3'剪接位点具有非常高度保守的UUUUCAG/R共有序列，我们推测U2AF可能用于识别该序列，从而促进秀丽隐杆线虫中的内含子识别。在此我们报告U2AF大亚基基因uaf-1的克隆。鉴定出三类cDNA。在最丰富的一类中，开放阅读框与哺乳动物和果蝇的U2AF65相似。其余两类是由选择性剪接事件导致的，其中一个含有框内终止密码子的外显子插入到第二个RNA识别基序起始附近。然而，这种选择性mRNA显然未被翻译。有趣的是，插入的外显子与3'剪接位点共有序列有10个匹配。为了确定该特征是否保守，我们对相关线虫秀丽新杆线虫的uaf-1进行了测序。它由六个外显子组成，包括一个选择性剪接的第三个外显子，该外显子在与秀丽隐杆线虫相同的位置中断基因。在这两个物种中，uaf-1都与rab-18基因包含在一个操纵子中。尽管来自两个物种的选择性外显子并非高度保守且不会编码相关多肽，但秀丽新杆线虫的选择性外显子与3'剪接位点共有序列有18个匹配。我们推测前体mRNA和选择性剪接外显子中的3'剪接位点样序列阵列可能具有调节作用。饥饿后，选择性剪接的RNA大量积累，表明当前体mRNA水平较低时，这种RNA可能代表一种降低U2AF65水平的适应机制。
