Sequence-Nonspecific Suppression of Gene Expression by Double-stranded RNA in Silkworm Cultured Cells
Sequence-Nonspecific Suppression of Gene Expression by Double-stranded RNA in Silkworm Cultured Cells
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DOI：
10.11416/jibs.78.1_33
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发表时间：
2009-02
影响因子：
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通讯作者：
K. Sakashita;Tsuneyuki Tatsuke;J. M. Lee;Y. Kawaguchi;T. Kusakabe
中科院分区：
文献类型：
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作者：
K. Sakashita;Tsuneyuki Tatsuke;J. M. Lee;Y. Kawaguchi;T. Kusakabe
双链RNA（dsRNA）在真核细胞中诱导多种生物学反应。序列特异性RNA干扰（RNAi）在真核生物中是高度保守的。在哺乳动物细胞中，已知长dsRNA通过激活几种细胞途径具有序列非特异性基因沉默效应。至于昆虫细胞，迄今为止只报道了最少量的信息。在这里，我们研究了双链RNA对鳞翅目细胞系家蚕BmN 4基因表达的序列非特异性影响。用长dsRNA和表达荧光素酶或β -半乳糖苷酶的报告质粒共转染细胞，并测量每种酶的活性。与靶基因不同源的dsRNA以浓度依赖性的方式降低报告子活性，尽管其有效浓度显著高于由与靶基因同源的dsRNA引起的浓度。使用与来自一些生物体如蚕、果蝇和人的几个基因互补的dsRNA以及合成的dsRNA，聚肌苷酸-聚胞苷酸（poly（I：C）），进一步证实了非同源dsRNA的序列非特异性抑制。由于在另一个鳞翅目细胞系草地贪夜蛾Sf 21中也观察到了长dsRNA的序列非依赖性效应，因此它可能是鳞翅目昆虫中保守的免疫反应之一。
Double-stranded RNA (dsRNA) induces various biological responses in eukaryotic cells. Sequence-specific RNA interference (RNAi) is highly conserved throughout eukaryotic organisms. In mammalian cells, long dsRNA is known to have sequence-nonspecific gene silencing effect by activating several cellular pathways. As for insect cells, only the smallest amount of information has been reported so far. Here, we investigate the sequence-non-specific effect of dsRNA on gene expression in a lepidopteran cell line, silkworm Bombyx mori BmN4. Cells were cotransfected with long dsRNA and a reporter plasmid expressing luciferase or β -galactosidase, and measured for the activities of each enzyme. dsRNAs which are not homologous to the target gene reduced the reporter ac- tivity in a concentration-dependent manner, albeit its effective concentration was considerably higher than that caused by dsRNA homologous to the target gene. The sequence-nonspecific repression by non-homologous dsRNAs was further confirmed using dsRNAs complementary to several genes from some organisms such as silkworm, fruit fly and human, as well as a synthetic dsRNA, polyinosinic-polycytidylic acid (poly(I:C)). Since the sequence-independent effect by long dsRNA was also observed in another lepidopteran cell line, fall armyworm Spodoptera frugiperda Sf21, it may be one of the conserved immune responses among lepidopteran insects.