The shrimp IKK-NF-κB signaling pathway regulates antimicrobial peptide expression and may be subverted by white spot syndrome virus to facilitate viral gene expression

The I kappa B kinases IKK alpha and IKK beta and the IKK-related kinases TANK-binding kinase 1 (TBK1) and IKK epsilon are the master regulators of the NF-kappa B signaling pathway. Although this pathway has been extensively studied in mammals, less attention has been paid in crustaceans, which have significant economic value. Here, we report the cloning and functional studies of two IKK homologs, LvIKK beta and LvIKK epsilon, from Pacific white shrimp, Litopenaeus vannamei. LvIKK beta and LvIKK epsilon mRNAs are widely expressed in different tissues and are responsive to white spot syndrome virus (WSSV) infection. When overexpressed in Drosophila S2 cells, LvIKK beta but not LvIKK epsilon activates the promoters of NF-kappa B pathway-controlled antimicrobial peptide genes (AMPs), such as the Penaeidins (PENs). In HEK 293T cells, both LvIKK beta and LvIKK epsilon activate an NF-kappa B reporter. The silencing of LvIKK beta or LvIKK epsilon using double-stranded RNA (dsRNA)-mediated RNA interference (RNAi) decreases the expression of L. vannamei AMPs, including PENs, lysozyme and crustins. Intriguingly, LvIKK beta- or LvIKK epsilon-silenced L. vannamei are resistant to WSSV infection. We hypothesized that successful infection with WSSV requires the activation of the IKK-NF-kappa B signaling pathway to modulate viral gene expression. We constructed luciferase reporters for 147 WSSV genes. By screening, we found that the WSSV051, WSSV059, WSSV069, WSSV083, WSSV090, WSSV107, WSSV244, WSSV303, WSSV371 and WSSV445 promoters can be activated by LvIKK beta or LvIKK epsilon in Drosophila S2 cells. Taken together, our results reveal that LvIKK beta and LvIKK epsilon may participate in the regulation of shrimp AMPs and that WSSV may subvert the L. vannamei IKK-NF-kappa B signaling pathway to facilitate viral gene expression.