Find in Library

Crustins are cysteine-rich cationic antimicrobial peptides with diverse biological functions including antimicrobial and proteinase inhibitory activities in crustaceans. Although a few crustins reportedly respond to white spot syndrome virus (WSSV) infection, the detailed antiviral mechanisms of crustins remain largely unknown. Our previous research has shown that <i>Sp</i>Crus2, from mud crab <i>Scylla paramamosain</i>, is a type II crustin containing a glycine-rich region (GRR) and a cysteine-rich region (CRR). In the present study, we found that <i>SpCrus2</i> was upregulated in gills after WSSV challenge. Knockdown of <i>SpCrus2</i> by injecting double-stranded RNA (<i>dsSpCrus2</i>) resulted in remarkably increased virus copies in mud crabs after infection with WSSV. These results suggested that <i>SpCrus2</i> played a critical role in the antiviral immunity of mud crab. A GST pull-down assay showed that recombinant <i>Sp</i>Crus2 interacted specifically with WSSV structural protein VP26, and this result was further confirmed by a co-immunoprecipitation assay with <i>Drosophila</i> S2 cells. As the signature sequence of type II crustin, <i>Sp</i>Crus2 GRR is a glycine-rich cationic polypeptide with amphipathic properties. Our study demonstrated that the GRR and CRR of <i>Sp</i>Crus2 exhibited binding activities to VP26, with the former displaying more potent binding ability than the latter. Interestingly, pre-incubating WSSV particles with recombinant <i>Sp</i>Crus2 (r<i>Sp</i>Crus2), rGRR, or rCRR inhibited virus proliferation in vivo; moreover, r<i>Sp</i>Crus2 and rGRR possessed similar antiviral abilities, which were much stronger than those of rCRR. These findings indicated that <i>Sp</i>Crus2 GRR contributed largely to the antiviral ability of <i>Sp</i>Crus2, and that the stronger antiviral ability of GRR might result from its stronger binding activity to the viral structural protein. Overall, this study provided new insights into the antiviral mechanism of <i>Sp</i>Crus2 and the development of new antiviral drugs.