MicroRNAs (miRNAs), a group of small molecule non-encoding RNAs, are key post-transcriptional regulators of gene expression that are implicated in many biological processes. In the current study, miR-217 from Eriocheir sinensis was selected for studying its roles during host–virus interaction. Overexpression or silencing of miR-217 led to considerable effects on white spot syndrome virus (WSSV) replication, implying that miR-217 played a positive role in WSSV infection. In insect High Five cells, miR-217 significantly inhibited Tube gene expression by binding to the 3′-untranslated region of the Tube. Overexpression of miR-217 in crab led to downregulation of tube expression. Knockdown of Tube in vivo led to significant enhancement of WSSV infection and inhibited the expression of five antimicrobial peptide (AMP) genes (Anti-lipopolysaccharide factor ALF1, ALF2, ALF3; Crustin Crus1, Crus2) in WSSV-challenged crabs. Overexpression of miR-217 also led to downregulation of these AMP genes in WSSV-challenged crabs. Our results showed that host miRNA played positive roles in virus infection by regulation of host tube gene, which is the key component of Toll signaling pathway.
Toll receptors are involved in innate immunity of invertebrates. In this study, we identify and characterize two Toll genes (named HcToll4 and HcToll5) from triangle sail mussel Hyriopsis cumingii. HcToll4 has complete cDNA sequence of 3,162 bp and encodes a protein of 909 amino acids. HcToll5 cDNA is 4,501 bp in length and encodes a protein of 924 amino acids. Both deduced HcToll4 and HcToll5 protein contain signal peptide, extracellular leucine rich repeats (LRRs), and intracellular Toll/interleukin-1 (IL-1) receptor domains. Quantitative real-time PCR analysis revealed that HcToll4 and HcToll5 were largely distributed in the hepatopancreas and could be detected in the gills and mantle. HcToll4 and HcToll5 expression could respond to Staphylococcus aureus, Vibrio parahaemolyticus, White spot syndrome virus (WSSV) or Poly I:C challenge. RNA interference by siRNA results showed that HcToll4 and HcToll5 were involved in the regulation of theromacin (HcThe) and whey acidic protein (HcWAP) expression. Based on these results, HcToll4 and HcToll5 might play pivotal function in H. cumingii innate immune response.
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