DNA interference: DNA-induced gene silencing in the appendicularian Oikopleura dioica
DNA interference: DNA-induced gene silencing in the appendicularian Oikopleura dioica
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DOI：
10.1098/rspb.2015.0435
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发表时间：
2015-05-22
影响因子：
4.7
通讯作者：
Nishida, Hiroki
中科院分区：
文献类型：
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作者：
Omotezako, Tatsuya;Onuma, Takeshi A.;Nishida, Hiroki
RNA干扰作为一种基因沉默系统在真核生物中被广泛应用，以防御入侵的核酸。为了响应入侵的双链RNA（dsRNA），mRNA以序列特异性方式降解。然而，到目前为止，DNA干扰（DNAi）仅在植物、纤毛虫和古生菌中有报道，而在后生动物中还没有探索。在这里，我们表明，线性双链DNA促进序列特异性转录阻断和mRNA降解的发育胚胎的apapellularian Oikopleura dioica。引入聚合酶链反应（PCR）产物或编码Brachyury的线性化质粒诱导尾部畸形和mRNA降解。这种畸形也是由假定的5 '侧翼区和内含子的DNA片段而没有编码区所促进的。编码Zic样1和乙酰胆碱酯酶的PCR产物也分别诱导感觉器官和肌肉乙酰胆碱酯酶活性的丧失。共注射编码EGFP和mCherry的mRNA以及编码这些荧光蛋白的PCR产物分别诱导绿色或红色荧光的序列特异性减少。这些结果表明O. dioica具有防御外源DNA和RNA的系统，DNA片段诱导的基因沉默可能通过转录阻断和mRNA降解介导。这是首次在后生动物中发现DNAi。
RNA interference is widely employed as a gene-silencing system in eukaryotes for host defence against invading nucleic acids. In response to invading double-stranded RNA (dsRNA), mRNA is degraded in sequence-specific manner. So far, however, DNA interference (DNAi) has been reported only in plants, ciliates and archaea, and has not been explored in Metazoa. Here, we demonstrate that linear double-stranded DNA promotes both sequence-specific transcription blocking and mRNA degradation in developing embryos of the appendicularian Oikopleura dioica. Introduced polymerase chain reaction (PCR) products or linearized plasmids encoding Brachyury induced tail malformation and mRNA degradation. This malformation was also promoted by DNA fragments of the putative 5'-flanking region and intron without the coding region. PCR products encoding Zic-like1 and acetylcholine esterase also induced loss of sensory organ and muscle acetylcholinesterase activity, respectively. Co-injection of mRNA encoding EGFP and mCherry, and PCR products encoding these fluorescent proteins, induced sequence-specific decrease in the green or red fluorescence, respectively. These results suggest that O. dioica possesses a defence system against exogenous DNA and RNA, and that DNA fragment-induced gene silencing would be mediated through transcription blocking as well as mRNA degradation. This is the first report of DNAi in Metazoa.