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首页> 外文期刊>Molecular and Cellular Biology >Mutational analysis of Saccharomyces cerevisiae U4 small nuclear RNA identifies functionally important domains.
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Mutational analysis of Saccharomyces cerevisiae U4 small nuclear RNA identifies functionally important domains.

机译：酿酒酵母U4小核RNA的突变分析确定功能上重要的域。

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U4 small nuclear RNA (snRNA) is essential for pre-mRNA splicing, although its role is not yet clear. On the basis of a model structure (C. Guthrie and B. Patterson, Annu. Rev. Genet. 22:387-419, 1988), the molecule can be thought of as having six domains: stem II, 5' stem-loop, stem I, central region, 3' stem-loop, and 3'-terminal region. We have carried out extensive mutagenesis of the yeast U4 snRNA gene (SNR14) and have obtained information on the effect of mutations at 105 of its 160 nucleotides. Fifteen critical residues in the U4 snRNA have been identified in four domains: stem II, the 5' stem-loop, stem I, and the 3'-terminal region. These domains have been shown previously to be insensitive to oligonucleotide-directed RNase H cleavage (Y. Xu, S. Petersen-Bj?rn, and J. D. Friesen, Mol. Cell. Biol. 10:1217-1225, 1990), suggesting that they are involved in intra- or intermolecular interactions. Stem II, a region that base pairs with U6 snRNA, is the most sensitive to mutation of all U4 snRNA domains. In contrast, stem I is surprisingly insensitive to mutational change, which brings into question its role in base pairing with U6 snRNA. All mutations in the putative Sm site of U4 snRNA yield a lethal or conditional-lethal phenotype, indicating that this region is important functionally. Only two nucleotides in the 5' stem-loop are sensitive to mutation; most of this domain can tolerate point mutations or small deletions. The 3' stem-loop, while essential, is very tolerant of change. A large portion of the central domain can be removed or expanded with only minor effects on phenotype, suggesting that it has little function of its own. Analysis of conditional mutations in stem II and stem I indicates that although these single-base changes do not have a dramatic effect on U4 snRNA stability, they are defective in RNA splicing in vivo and in vitro, as well as in spliceosome assembly. These results are discussed in the context of current knowledge of the interactions involving U4 snRNA.
机译：U4小核RNA（snRNA）对于mRNA前剪接是必不可少的，尽管其作用尚不清楚。根据模型结构（C. Guthrie和B. Patterson，Annu。Rev. Genet。22：387-419，1988），该分子可被认为具有六个结构域：茎II，5'茎环，茎I，中央区域，3'茎环和3'末端区域。我们已经对酵母U4 snRNA基因（SNR14）进行了广泛的诱变，并获得了有关其160个核苷酸中105个突变的影响的信息。已在四个域中鉴定出U4 snRNA中的15个关键残基：茎II，5'茎环，茎I和3'末端区域。先前已经显示这些结构域对寡核苷酸定向的RNase H裂解不敏感（Y.Xu，S.Petersen-Bjrn，和JDFriesen，Mol.Cell.Biol.10：1217-1225，1990），这表明它们参与分子内或分子间的相互作用。茎II是与U6 snRNA碱基配对的区域，对所有U4 snRNA结构域的突变最敏感。相比之下，茎I对突变变化却不敏感，这使人怀疑它在与U6 snRNA碱基配对中的作用。 U4 snRNA的假定Sm位点中的所有突变均产生致命或条件致死表型，表明该区域在功能上很重要。 5'茎环中只有两个核苷酸对突变敏感。该结构域的大部分可以耐受点突变或小的缺失。 3'茎环虽然必不可少，但却能容忍变化。中央结构域的大部分可以被移除或扩展，而对表型的影响很小，这表明它本身没有什么功能。对茎II和茎I中条件突变的分析表明，尽管这些单碱基变化对U4 snRNA稳定性没有显着影响，但它们在体内和体外RNA剪接以及剪接体组装中均存在缺陷。在涉及U4 snRNA相互作用的当前知识的背景下讨论了这些结果。

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