Miraglia L, Seiwert S, Igel A H, Ares M
Sinsheimer Laboratories, University of California, Santa Cruz 95064.
Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):7061-5. doi: 10.1073/pnas.88.16.7061.
U2 is a highly conserved small nuclear RNA essential for pre-mRNA splicing in mammals and yeast and for trans-splicing in trypanosomes. To test the function of variant U2 RNA structures from different organisms, we conducted phylogenetic exchanges of U2 domains. Replacing nucleotides 1-120 of yeast U2 with the corresponding region of human U2 generates a U2 RNA that is correctly folded and functions in yeast. In contrast, replacement of the branchpoint interaction region of yeast U2 with the corresponding region from trypanosome is dominant lethal. Using a GAL-U2 promoter fusion, we show that the dominant phenotype can be made conditional and that the accumulation of mutant U2 is followed rapidly by inhibition of nuclear pre-mRNA splicing. The results suggest that U2 small nuclear ribonucleoprotein particles normally participate in stable complexes with a limiting splicing factor prior to formation of U2-intron branchpoint base pairs.
U2是一种高度保守的小核RNA，对哺乳动物和酵母中的前体mRNA剪接以及锥虫中的反式剪接至关重要。为了测试来自不同生物体的变体U2 RNA结构的功能，我们进行了U2结构域的系统发育交换。用人类U2的相应区域替换酵母U2的核苷酸1-120会产生一种在酵母中正确折叠并发挥功能的U2 RNA。相比之下，用锥虫的相应区域替换酵母U2的分支点相互作用区域具有显性致死性。使用GAL-U2启动子融合，我们表明显性表型可以是条件性的，并且突变U2的积累会迅速导致核前体mRNA剪接受到抑制。结果表明，U2小核糖核蛋白颗粒通常在形成U2-内含子分支点碱基对之前与一种有限的剪接因子参与稳定复合物的形成。
