Litopenaeus vannamei Toll-interacting protein (LvTollip) is a potential negative regulator of the shrimp Toll pathway involved in the regulation of the shrimp antimicrobial peptide gene penaeidin-4 (PEN4).

The Toll-like receptor (TLR)-nuclear factor (NF)-κB signaling pathway is evolutionarily conserved from insects to mammals as a regulator of the expression of immune-related genes. In mammals, TLR-NF-κB signaling is tightly controlled because excessive activation of this pathway can result in severe damage to the host. The mammalian Toll-interacting protein (Tollip) has an important function in the negative regulation of this pathway, but no reports about invertebrate Tollip have been published to date. In this study, we cloned Litopenaeus vannamei Tollip (LvTollip) and investigated its function in the regulation of the NF-κB pathway-controlled antimicrobial peptide genes (AMPs). The LvTollip full-length cDNA is 1231bp long and contains an open reading frame of 813bp that encodes a 270-amino acid protein. LvTollip shares significant similarities to mammalian Tollips, which contain a centrally localized protein kinase C conserved region 2 (C2) domain and a C-terminal CUE domain. After challenges with the white spot syndrome virus (WSSV) or Vibrio alginolyticus, the expression levels of LvTollip were altered in the gill, hemocyte, hepatopancreatic, intestinal, and muscle tissues. In Drosophila S2 cells, LvTollip localized in the membrane and the cytoplasm and significantly inhibited the promoter activities of the NF-κB pathway-controlled AMP penaeidin-4 (PEN4). In LvTollip-knockdown shrimp, the expression level of AMP PEN4 was increased. However, the mortality rates of LvTollip-knockdown shrimp in response to WSSV or V. alginolyticus infections were not significantly different from those of the control group. Our results suggested that LvTollip might be involved in the negative regulation of PEN4 and that LvTollip expression was responsive to microbial infections. Crown