Deep sequencing uncovers commonality in small RNA profiles between transgene-induced and naturally occurring RNA silencing of chalcone synthase-A gene in petunia.
Deep sequencing uncovers commonality in small RNA profiles between transgene-induced and naturally occurring RNA silencing of chalcone synthase-A gene in petunia.
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DOI：
10.1186/1471-2164-14-63
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发表时间：
2013-01-30
期刊：
影响因子：
4.4
通讯作者：
Kanazawa A
中科院分区：
文献类型：
--
作者：
Kasai M;Matsumura H;Yoshida K;Terauchi R;Taneda A;Kanazawa A
在植物基因组中引入转录与内源基因同源的RNA的转基因可以诱导两个基因的沉默，这种现象称为共抑制。共抑制首先在用编码查耳酮合酶的CHS-A基因转化的转基因矮牵牛植物中发现，其中产生花中的无色素部分或完全白色的花。在具有CHS-A共抑制的转基因矮牵牛中观察到的一些花色模式类似于现有的非转基因品种。虽然已知非转基因矮牵牛中产生白色扇形的机制是由于CHS-A基因的RNA沉默，如共抑制，但这些现象中是否涉及相同的触发因子和/或RNA降解模式尚不清楚。在这里，我们使用小RNA的深度测序和生物信息学分析来解决这个问题。我们分析了短干扰RNA（siRNA）产生的非色素部门的花瓣组织在转基因矮牵牛植物，有CHS-A共抑制和非转基因矮牵牛品种红星星，有自然发生的CHS-A RNA沉默。在两个沉默系统中，21-nt和22-nt siRNA分别是最丰富和第二丰富的大小类别。CHS-A siRNA的产生仅限于外显子2，表明通过RNA沉默途径的RNA降解发生在该外显子中。在共抑制和自然发生的RNA沉默中检测到共同的siRNA，并且基于这些植物中siRNA数量的它们的等级彼此相关。值得注意的是，这些系统中普遍存在高度丰富的siRNAs。在多个位点的多个相位中检测到定相siRNA，并且产生定相siRNA的区域的一些末端是保守的。发现共同抑制和天然存在的CHS-A基因沉默的siRNA产生的特征表明这些沉默系统之间的机制相似性，特别是在siRNA的生物合成过程中，包括CHS-A转录物的切割和随后在外显子2中产生二级siRNA。数据还表明，这些事件发生在多个位点，这可能是这些沉默现象的一个特征。
Introduction of a transgene that transcribes RNA homologous to an endogenous gene in the plant genome can induce silencing of both genes, a phenomenon termed cosuppression. Cosuppression was first discovered in transgenic petunia plants transformed with the CHS-A gene encoding chalcone synthase, in which nonpigmented sectors in flowers or completely white flowers are produced. Some of the flower-color patterns observed in transgenic petunias having CHS-A cosuppression resemble those in existing nontransgenic varieties. Although the mechanism by which white sectors are generated in nontransgenic petunia is known to be due to RNA silencing of the CHS-A gene as in cosuppression, whether the same trigger(s) and/or pattern of RNA degradation are involved in these phenomena has not been known. Here, we addressed this question using deep-sequencing and bioinformatic analyses of small RNAs. We analyzed short interfering RNAs (siRNAs) produced in nonpigmented sectors of petal tissues in transgenic petunia plants that have CHS-A cosuppression and a nontransgenic petunia variety Red Star, that has naturally occurring CHS-A RNA silencing. In both silencing systems, 21-nt and 22-nt siRNAs were the most and the second-most abundant size classes, respectively. CHS-A siRNA production was confined to exon 2, indicating that RNA degradation through the RNA silencing pathway occurred in this exon. Common siRNAs were detected in cosuppression and naturally occurring RNA silencing, and their ranks based on the number of siRNAs in these plants were correlated with each other. Noticeably, highly abundant siRNAs were common in these systems. Phased siRNAs were detected in multiple phases at multiple sites, and some of the ends of the regions that produced phased siRNAs were conserved. The features of siRNA production found to be common to cosuppression and naturally occurring silencing of the CHS-A gene indicate mechanistic similarities between these silencing systems especially in the biosynthetic processes of siRNAs including cleavage of CHS-A transcripts and subsequent production of secondary siRNAs in exon 2. The data also suggest that these events occurred at multiple sites, which can be a feature of these silencing phenomena.
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