ITRAQ proteome analysis of insight into TLR20.2 functions through IFN1 signaling exerts regulatory effects on GCRV replication

Abstract

Maternal immunization is an efficacious way to protect offspring from pathogen infections. To clarify the immune-enhancing mechanism of maternal immunization in grass carp, the iTRAQ assay was applied to screen differentially expressed proteins (DEPs) among eggs produced from maternal-immunized and non-immunized female grass carp. A total of 138 DEPs were obtained and TLR20.2 was the sole identified consensus DEP among the maternal-immunized and non-immunized groups. In general, the gene and protein expressions of TLR20.2 in the egg cell and early developmental stage embryos could be enhanced by maternal-immunized treatment. The mRNA levels of tlr20.2 were up-regulated in the liver, head kidney, and spleen in response to grass carp reovirus (GCRV) infection. Mechanistically, TLR20.2 functions through the IFN1 signaling and displayed inhibitory functions on GCRV replication by remarkably down-regulation of vp2 and vp7 expressions. The HSP90 was identified to be one of the proteins that interacted with TLR20.2, which coexpressed with TLR20.2 and mainly concentrated around the central veins. HSP90 promotes GCRV replication by inhibiting the irf3, irf7, and ifn1 expressions, which resulted in remarkably increased vp2 and vp7 gene expressions. The HMGB1a and HMGB1b could bind to the TLR20–2 promoter, and showed positive effects on tlr20.2 expression by significantly promoting its promoter activity (P < 0.01). Moreover, HMGB1a and HMGB1b exerted negative effects on HSP90 expression and thus the GCRV replication. Overall, this study provided molecular evidence that maternal immunization treatment protect against GCRV infection through the enhancement of TLR20.2 expression, and the HMGB1-TLR20.2-HSP90-IFN axis confers regulatory effects on GCRV replication. The results are novel findings to help elucidate the resistance-enhancing mechanism in fish species.