Wang Huidong, Gong Liang, Qi Jiangwei, Hu Meiying, Zhong Guohua, Gong Liang
Key Laboratory of Pesticide and Chemical Biology, Ministry of Education, South China Agricultural University, Guangzhou, PR China.
Arch Insect Biochem Physiol. 2014 Nov;87(3):164-76. doi: 10.1002/arch.21189. Epub 2014 Aug 18.
RNA interference (RNAi) signal can spread from the point where the double-stranded RNA (dsRNA) was initially applied to other cells or tissues. SID-related genes in Caenorhabditis elegans help in the spreading of this signal. However, the mechanisms of systemic RNAi are still not unveiled in insects. In this study, we cloned a full-length cDNA of sid-1-like gene, Pxylsid-1, from Plutella xylostella that contains 1,047 bp opening reading frame encoding a putative protein of 348 amino acids. This transcript is very much similar to the sil-1 in Bombyx mori (68.8%). The higher expression levels of Pxylsid-1 were found at the adult and fourth-instar stages compared to the second-instar stage with 21.48- and 10.36-fold increase, respectively. Its expression levels in different tissues were confirmed with the highest expression in the hemolymph, which showed 21.09-fold increase than the midgut; however it was lower in other tissues. The result of RNAi by feeding bacterially expressed dsRNA targeting Pxylace-1, which showed that the mRNA level of Pxylace-1 decreased by 34.52 and 64.04% after 36- and 72-h treatment, respectively. However, the mRNA level of Pxylsid-1 was not significantly induced when the Pxylace-1 was downregulated. Furthermore, we found that downregulation of Pxylsid-1 did not affect the RNAi effect of Pxylace-1. Hence, the Pxylsid-1 may not be involved in absorption of dsRNA from the midgut fluid. A further study is needed to uncover the function of Pxylsid-1.
RNA干扰（RNAi）信号能够从双链RNA（dsRNA）最初作用的位点扩散至其他细胞或组织。秀丽隐杆线虫中与SID相关的基因有助于这种信号的扩散。然而，昆虫体内系统性RNAi的机制仍未明确。在本研究中，我们从小菜蛾中克隆了一个类sid-1基因Pxylsid-1的全长cDNA，其开放阅读框为1047 bp，编码一个含有348个氨基酸的假定蛋白。该转录本与家蚕中的sil-1非常相似（相似度为68.8%）。与二龄幼虫期相比，Pxylsid-1在成虫期和四龄幼虫期的表达水平更高，分别增加了21.48倍和10.36倍。其在不同组织中的表达水平得到确认，血淋巴中的表达量最高，比中肠高21.09倍；然而在其他组织中表达量较低。通过喂食靶向Pxylace-1的细菌表达dsRNA进行RNAi实验，结果显示，处理36小时和72小时后，Pxylace-1的mRNA水平分别下降了34.52%和64.04%。然而，当Pxylace-1表达下调时，Pxylsid-1的mRNA水平并未显著诱导升高。此外，我们发现下调Pxylsid-1并不影响Pxylace-1的RNAi效果。因此，Pxylsid-1可能不参与从中肠液中吸收dsRNA。需要进一步研究以揭示Pxylsid-1的功能。
