Key Laboratory of Animal Virology of Ministry of Agriculture and College of Life Sciences, Zhejiang University, Hangzhou, China.
J Virol. 2012 Dec;86(23):12997-3004. doi: 10.1128/JVI.01702-12. Epub 2012 Sep 26.
Growing evidence from mammals suggests that host microRNAs (miRNAs) play important roles in the antiviral immune response. However, the roles of invertebrate miRNAs in response to virus infection remain to be investigated. Based on our previous studies, the shrimp miR-7 was found to be upregulated in response to white spot syndrome virus (WSSV) infection. In this study, the results showed that shrimp miR-7 could target the 3'-untranslated region (3'UTR) of the WSSV early gene wsv477, implying that miR-7 was involved in viral DNA replication. In insect High Five cells, the synthesized miR-7 significantly decreased the expression level of the fluorescent construct bearing the 3'UTR of wsv477 compared with the expression of the control constructs. When the activity of transfected miR-7 was blocked by locked-nucleic-acid (LNA)-modified anti-miR-7 oligonucleotide (AMO-miR-7), the repression of luciferase gene expression by miR-7 was relieved. In vivo, when the synthesized miR-7 was injected into shrimp, the numbers of WSSV genome copies/mg gills were 1,000-fold lower than those of WSSV only at 72 and 96 h postinfection. The results indicated that the blocking of endogenous miR-7 by AMO-miR-7 led to about a 10-fold increase of WSSV genome copies/mg gills in WSSV-infected shrimp compared with the control WSSV only. Further, it was revealed that the host Dicer1 was an important component for the biogenesis of miR-7, which had a large effect on virus infection. Therefore, our study revealed a novel regulatory function for an invertebrate miRNA in host-virus interactions by targeting the viral early gene.
越来越多的哺乳动物研究证据表明，宿主 microRNAs（miRNAs）在抗病毒免疫反应中发挥着重要作用。然而，无脊椎动物 miRNAs 对病毒感染的反应作用仍有待研究。根据我们之前的研究，发现虾 miR-7 对白斑综合征病毒（WSSV）感染呈上调表达。在本研究中，结果表明虾 miR-7 可以靶向 WSSV 早期基因 wsv477 的 3'非翻译区（3'UTR），这意味着 miR-7 参与了病毒 DNA 的复制。在昆虫 High Five 细胞中，与对照构建体相比，携带 wsv477 3'UTR 的合成 miR-7 显著降低了荧光构建体的表达水平。当转染 miR-7 的活性被锁核酸（LNA）修饰的抗 miR-7 寡核苷酸（AMO-miR-7）阻断时，miR-7 对荧光素酶基因表达的抑制作用得到缓解。在体内，当合成的 miR-7 被注射到虾体内时，WSSV 基因组拷贝数/mg 鳃中的数量在感染后 72 和 96 小时比仅感染 WSSV 时低 1000 倍。结果表明，与仅感染 WSSV 的对照相比，AMO-miR-7 阻断内源性 miR-7 导致感染 WSSV 的虾中 WSSV 基因组拷贝数/mg 鳃增加了约 10 倍。此外，还揭示了宿主 Dicer1 是 miR-7 生物发生的重要组成部分，它对病毒感染有很大影响。因此，我们的研究通过靶向病毒早期基因，揭示了一种无脊椎动物 miRNA 在宿主-病毒相互作用中的新的调控功能。
