In-depth sequencing of the siRNAs associated with peach latent mosaic viroid infection.
In-depth sequencing of the siRNAs associated with peach latent mosaic viroid infection.
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DOI：
10.1186/1471-2199-11-16
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发表时间：
2010-02-16
影响因子：
--
通讯作者：
Perreault JP
中科院分区：
文献类型：
--
作者：
Bolduc F;Hoareau C;St-Pierre P;Perreault JP
已经观察到类病毒感染后，在感染的植物中存在类病毒衍生的siRNA的积累。一些实验结果表明，这些小RNA可能是由植物防御系统产生的，以保护它免受感染，表明类病毒可以引发RNA沉默途径。本研究的目的是确定在桃潜伏花叶类病毒（PLMVd），一个模型RNA基因组，最容易受到RNA干扰机制的区域。从受感染的树中分离的RNA已被用于平行的类病毒种类和小的非编码RNA种类的测序。具体地，根据常规方法扩增、克隆和测序PLMVd RNA，而通过高通量测序确定小的非编码RNA。第一个导致了18种新型PLMVd变体的分型。第二个提供了一个小RNA文库，包括880 000个对应于PLMVd衍生的siRNA的序列，占感染文库序列的11.2%。这些siRNA主要含有21-22个核苷酸的RNA，并且等价地分布在类病毒的正极性和负极性之间。它们覆盖了完整的类病毒基因组，尽管数量因地区而异。这些区域不一定与作为Dicer样酶底物的双链要求相关。我们注意到，一些序列包含锤头状自切割位点，表明环状构象异构体可以被RNA沉默机制加工。最后，观察到5'核苷酸性质的相对丰度的偏差（A，U >> G，C）。所使用的方法为我们提供了从受感染的桃树中检索到的PLMVd衍生的siRNA的定量表示。这些siRNA占小的非编码RNA的相对大的比例。令人惊讶的是，来自PLMVd基因组的一些区域的siRNA似乎过表达，尽管这些区域不一定形成足够长的双链结构以满足底物特异性的Dicer样标准。重要的是，这个大的siRNA文库给出了关于所涉及的沉默机制的组分的几个提示。
It has been observed that following viroid infection, there is an accumulation of viroid-derived siRNAs in infected plants. Some experimental results suggest that these small RNAs may be produced by the plant defense system to protect it from infection, indicating that viroids can elicit the RNA-silencing pathways. The objective of this study is to identify in the peach latent mosaic viroid (PLMVd), a model RNA genome, the regions that are most susceptible to RNA interference machinery. The RNA isolated from an infected tree have been used to sequence in parallel viroid species and small non-coding RNA species. Specifically, PLMVd RNAs were amplified, cloned and sequenced according to a conventional approach, while small non-coding RNAs were determined by high-throughput sequencing. The first led to the typing of 18 novel PLMVd variants. The second provided a library of small RNAs including 880 000 sequences corresponding to PLMVd-derived siRNAs, which makes up 11.2% of the sequences of the infected library. These siRNAs contain mainly 21-22 nucleotide RNAs and are equivalently distributed between the plus and the minus polarities of the viroid. They cover the complete viroid genome, although the amount varies depending on the regions. These regions do not necessarily correlate with the double-stranded requirement to be a substrate for Dicer-like enzymes. We noted that some sequences encompass the hammerhead self-cleavage site, indicating that the circular conformers could be processed by the RNA-silencing machinery. Finally, a bias in the relative abundance of the nature of the 5' nucleotides was observed (A, U >> G, C). The approach used provided us a quantitative representation of the PLMVd-derived siRNAs retrieved from infected peach trees. These siRNAs account for a relatively large proportion of the small non-coding RNAs. Surprisingly, the siRNAs from some regions of the PLMVd genome appear over-represented, although these regions are not necessarily forming sufficiently long double-stranded structures to satisfy Dicer-like criteria for substrate specificity. Importantly, this large library of siRNAs gave several hints as to the components of the involved silencing machinery.
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影响因子：
3.7
作者：
Di Serio F;Gisel A;Navarro B;Delgado S;Martínez de Alba AE;Donvito G;Flores R
通讯作者：
Flores R
影响因子：
3
作者：
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通讯作者：
Birney E
影响因子：
7
作者：
Morin, Ryan D.;O'Connor, Michael D.;Marra, Marco A.
通讯作者：
Marra, Marco A.
影响因子：
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作者：
Martínez de Alba, AE;Flores, R;Hernández, C
通讯作者：
Hernández, C
影响因子：
14.9
作者：
Papaefthimiou, I;Hamilton, AJ;Tabler, M
通讯作者：
Tabler, M