The roles of endonucleolytic cleavage and exonucleolytic digestion in the 5′-end processing of S-cerevisiae box C/D snoRNAs
The roles of endonucleolytic cleavage and exonucleolytic digestion in the 5′-end processing of S-cerevisiae box C/D snoRNAs
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DOI：
10.1261/rna.5126203
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发表时间：
2003-11-01
期刊：
影响因子：
4.5
通讯作者：
Chanfreau, G
中科院分区：
文献类型：
--
作者：
Lee, CY;Lee, A;Chanfreau, G
小核仁RNA（snoRNAs）在核糖体RNA代谢中起重要作用。在酿酒酵母中，盒C/D snoRNAs由切除的内含子、多顺反子前体或独立转录单位合成。先前的研究表明，只有少数独立转录的盒C/D snoRNAs在其5'端进行加工。在此我们描述了另外12种独立转录的盒C/D snoRNAs，它们经历5'端加工。在这些snoRNAs的前体中发现的5'延伸包含Rnt1p（酿酒酵母中RNase III的同源物）的切割位点，并且在缺乏Rnt1p时未加工的前体在体内积累。当5'→3'外切核酸酶Xrn1p和Rat1p失活（xrn1Δ rat1 - 1）时在体内鉴定出Rnt1p切割产物，并且使用模型RNA底物和重组Rnt1p在体外也鉴定出该产物。当rnt1Δ缺失与xrn1Δ rat1 - 1突变相结合时，其中一些snoRNAs显示未加工前体水平增加，这表明这些外切核酸酶参与snoRNA前体的5'加工或降解。在缺乏三甲基鸟苷加帽酶Tgs1p时，未加工的前体没有明显的不稳定，这表明5'单甲基帽足以确保这些前体的稳定。这些结果表明，来自酵母基因组的大多数独立转录的盒C/D snoRNAs经历5'端加工，并且Rnt1p内切核酸酶以及Xrn1p和Rat1p 5'→3'外切核酸酶在这些snoRNAs的5'端加工中具有部分冗余功能。
Small nucleolar RNAs (snoRNAs) play important roles in ribosomal RNA metabolism. In Saccharomyces cerevisiae, box C/D snoRNAs are synthesized from excised introns, polycistronic precursors, or independent transcription units. Previous studies have shown that only a few independently transcribed box C/D snoRNAs are processed at their 5' end. Here we describe 12 additional independently transcribed box C/D snoRNAs that undergo 5'-end processing. 5' Extensions found in the precursors of these snoRNAs contain cleavage sites for Rnt1p, the S. cerevisiae homolog of RNase III, and unprocessed precursors accumulate in vivo in the absence of Rnt1p. Rnt1p cleavage products were identified in vivo when the 5' --> 3' exonucleases Xrn1p and Rat1p are inactivated (xrn1Delta rat1-1) and in vitro using model RNA substrates and recombinant Rnt1p. Some of these snoRNAs show increased levels of unprocessed precursors when the rnt1Delta deletion is combined to the xrn1Delta rat1-1 mutation, suggesting that these exonucleases participate in the 5' processing or the degradation of the snoRNA precursors. Unprocessed precursors are not significantly destabilized in the absence of the trimethylguanosine capping enzyme Tgs1p, suggesting that a 5' monomethyl cap is sufficient to ensure stabilization of these precursors. These results demonstrate that the majority of independently transcribed box C/D snoRNAs from the yeast genome undergo 5'-end processing and that the Rnt1p endonuclease and the Xrn1p and Rat1p 5' --> 3' exonucleases have partially redundant functions in the 5'-end processing of these snoRNAs.