Function analysis of fish Tollip gene in response to virus infection

Toll-interacting protein (Tollip) is one of the important regulatory proteins of Toll-like receptor (TLR) signaling pathways. In previous studies, a Tollip sequence of grouper (Epinephelus coioides) was identified and the signal transduction functions of Tollip were studied. However, the response of Tollip to virus infection has not been characterized from grouper. In the present paper, the Tollip homolog (EtTollip) from grouper (Epinephelus tauvina) was cloned and its immune response to Singapore grouper iridovirus (SGIV) was investigated. EtTollip shares significant similarities to other mammalian Tollips, which contain a centrally localized protein kinase C conserved region 2 (C2) domain and a C-terminal CUE domain. After challenging with SGIV, the expression levels of EtTollip were altered in the spleen and head kidney of grouper. EtTollip mainly aggregated in the cytoplasm in a condensed state and was also distributed on the membranes of GS cells. EtTollip significantly inhibited the activities of NF-κB and IFN-β luciferase reporter when transfected into grouper spleen (GS) cells. SGIV can increase the activities of NF-κB and IFN-β luciferase reporter, especially to IFN-β. When transfected EtTollip with EcMyd88, the activity of NF-κB was increased, while transfected EtTollip with EcIRF3, the activity of IFN-β was significantly increased. Over-expressed EtTollip inhibited the transcription of SGIV genes significantly in GS cells, and silencing of EtTollip with siRNA led to increase of SGIV genes loads. Taken together, the results provide new insights in to the importance of Tollip as evolutionarily conserved molecule for grouper innate immunity against virus infection.