Secondary siRNAs from Medicago NB-LRRs modulated via miRNA-target interactions and their abundances
Secondary siRNAs from Medicago NB-LRRs modulated via miRNA-target interactions and their abundances
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DOI：
10.1111/tpj.12900
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发表时间：
2015-08-01
期刊：
影响因子：
7.2
通讯作者：
Meyers, Blake C.
中科院分区：
文献类型：
--
作者：
Fei, Qili;Li, Pingchuan;Meyers, Blake C.
小分子RNA是一类在基因表达调控网络中起重要作用的非编码RNA。不同家族的小RNA通过不同的生物合成途径产生。植物特有的一个这样的家族是定相的二级siRNA（phasiRNA）;这些家族需要RDR 6、DCL 4和（通常）microRNA（miRNA）触发其生物发生。蛋白质编码基因是phasiRNA的重要来源。模式豆科植物蒺藜苜蓿（Medicago truncatula）从许多PHAS位点产生phasiRNA，我们的目的是研究它们的生物起源和miRNAs触发这些分子的机制。我们调节了转基因组织中的miRNA丰度，表明phasiRNA的丰度与miRNA触发物和靶前体转录物的水平相关。我们鉴定了响应于miRNA过表达而显著增加的phasiRNA或PHAS基因座的集合。在验证来自miRNA过表达组织的靶标的过程中，我们发现在miRNA-mRNA靶标配对中，第30个末端核苷酸（第22位）而不是第10位对于phasiRNA的产生是重要的。突变单个30末端核苷酸显著减少了phasiRNA的产生。拟南芥中苜蓿NB-LRR靶向miRNA的异位表达表明，只有少数NB-LRR能够产生phasiRNA;我们的数据表明，这可能是由于靶位点侧翼序列决定的靶不可及性。我们的结果表明，靶点可及性是miRNA与靶点相互作用的重要组成部分，可用于靶点预测，并且miRNA靶点侧翼mRNA序列的进化可能会受到影响。
Small RNAs are a class of non-coding RNAs that are of great importance in gene expression regulatory networks. Different families of small RNAs are generated via distinct biogenesis pathways. One such family specific to plants is that of phased, secondary siRNAs (phasiRNAs); these require RDR6, DCL4, and (typically) a microRNA (miRNA) trigger for their biogenesis. Protein-encoding genes are an important source of phasiRNAs. The model legume Medicago truncatula generates phasiRNAs from many PHAS loci, and we aimed to investigate their biogenesis and mechanism by which miRNAs trigger these molecules. We modulated miRNA abundances in transgenic tissues showing that the abundance of phasiRNAs correlates with the levels of both miRNA triggers and the target, precursor transcripts. We identified sets of phasiRNAs or PHAS loci that predominantly and substantially increase in response to miRNA overexpression. In the process of validating targets from miRNA overexpression tissues, we found that in the miRNA-mRNA target pairing, the 30 terminal nucleotide (the 22nd position), but not the 10th position, is important for phasiRNA production. Mutating the single 30 terminal nucleotide dramatically diminishes phasiRNA production. Ectopic expression of Medicago NB-LRR-targeting miRNAs in Arabidopsis showed that only a few NB-LRRs are capable of phasiRNA production; our data indicate that this might be due to target inaccessibility determined by sequences flanking target sites. Our results suggest that target accessibility is an important component in miRNA-target interactions that could be utilized in target prediction, and the evolution of mRNA sequences flanking miRNA-target sites may be impacted.