Identification and functional analysis of 20 box H/ACA small nucleolar RNAs (snoRNAs) from Schizosaccharomyces pombe
Identification and functional analysis of 20 box H/ACA small nucleolar RNAs (snoRNAs) from Schizosaccharomyces pombe
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DOI：
10.1074/jbc.m500326200
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发表时间：
2005-04-22
影响因子：
4.8
通讯作者：
Qu, LH
中科院分区：
文献类型：
--
作者：
Li, SG;Zhou, H;Qu, LH
考虑到核仁中富集的所有小核仁RNA（snoRNA），我们从粟酒裂殖酵母中构建了一个专门的小核RNA cDNA文库，并首次分离出20种新型box H/ACA snoRNA。其中13个被表征为预测在18 S和25 S rRNA中指导19个假尿嘧啶化的新向导。其余7种snoRNA被认为是缺乏与rRNA或snRNA的序列互补性的孤儿向导。我们已经通过在裂殖酵母中的基因缺失实验证明了10个新的snoRNA的功能。snoRNA对S. pombe，尽管揭示了snR 94耗尽对酵母生长的影响，特别是在23 ° C下。共精确定位了30个假尿苷酸化位点。粟酒裂殖酵母rRNA，显示出一个独特的假尿苷化模式在芽殖酵母。有趣的是，S. pombe 25 S rRNA指出了Psi 2345在赋予酵母生长优势方面的关键作用。与酵母中内含子编码的box C/D snoRNA相反，所有box H/ACA snoRNA似乎独立于两个蛋白质编码基因之间的基因间区域转录，除了snR 35，snR 35嵌套在一个开放的阅读框中编码一个假设的蛋白质，尽管从相反的链表达。值得注意的是，snR 90与内含子编码的box C/D snoRNA共转录，这是第一次证明非编码RNA基因通过其外显子和内含子编码两种不同类型的snoRNA。详细比较了S.粟酒裂殖酵母snoRNA及其在不同生物体中的功能同源物表明了snoRNA与rRNA协同进化以在远距离真核生物中保留转录后修饰位点的机制。
Considering all small nucleolar RNAs (snoRNAs) enriched in the nucleolus, we generated a specialized cDNA library of small nuclear RNAs from Schizosaccharomyces pombe and isolated, for the first time, 20 novel box H/ACA snoRNAs. Thirteen of these were characterized as novel guides that were predicted to direct 19 pseudouridylations in 18 S and 25 S rRNAs. The remaining seven snoRNAs were considered as orphan guides that lack sequence complementarity to either rRNAs or snRNAs. We have experimentally demonstrated the function of the 10 novel snoRNAs by gene deletion in the fission yeast. The snoRNAs were shown to be dispensable for the viability of S. pombe, although an impact of snR94 depletion on yeast growth, especially at 23 degrees C, was revealed. A total of 30 pseudouridylation sites were precisely mapped in the S. pombe rRNAs, showing a distinctive pseudouridylation pattern in the budding yeast. Interestingly, the absence of pseudouridylation on U2347 in S. pombe 25 S rRNA pointed out a critical role for Psi 2345 in conferring a growth advantage for yeast. In contrast to the intron-encoded box C/D snoRNAs in yeast, all box H/ACA snoRNAs appeared to be transcribed independently from intergenic regions between two protein-coding genes, except for snR35, which was nested in an open reading frame encoding for a hypothetical protein, although expressed from the opposite strand. Remarkably, snR90 was cotranscribed with an intron-encoded box C/D snoRNA, and this is the first demonstration of a non-coding RNA gene that encodes two different types of snoRNAs by its exon and intron. A detailed comparison of the S. pombe snoRNAs, with their functional homologues in diverse organisms, suggests a mechanism by which the snoRNAs have evolved in coordination with rRNAs to preserve the post-transcriptional modification sites among distant eukaryotes.