Identification and molecular characterization of a Spätzle-like protein from Chinese shrimp ( Fenneropenaeus chinensis)

• Published in: Fish & shellfish immunology 2009-11, Vol.27 (5), p.610-617
• Main Authors: Shi, Xiu-Zhen, Zhang, Ran-Ran, Jia, Yu-Ping, Zhao, Xiao-Fan, Yu, Xiao-Qiang, Wang, Jin-Xing
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Animals; Base Sequence; Blotting, Western - veterinary; Brackish; Cambaridae; Chinese shrimp; Cloning, Molecular; DNA, Complementary - genetics; Escherichia coli; Expressed Sequence Tags; Expression pattern; Fenneropenaeus chinensis; Freshwater; Hemocytes - metabolism; Hemocytes - microbiology; Innate immunity; Lepeophtheirus salmonis; Marine; Molecular Sequence Data; Penaeidae - genetics; Penaeidae - metabolism; Penaeidae - microbiology; Phylogeny; Procambarus clarkii; Proteins - genetics; Proteins - metabolism; Reverse Transcriptase Polymerase Chain Reaction - veterinary; RNA, Messenger - biosynthesis; RNA, Messenger - genetics; Salmonidae; Sequence Alignment; Spätzle-like protein; Vibrio - immunology; Vibrio anguillarum; White spot syndrome virus
• ISSN: 1050-4648; 1095-9947
• DOI: 10.1016/j.fsi.2009.07.005

In invertebrates, the Toll signaling pathway is important for activation of antimicrobial peptides in the innate immune system. Activation of the Toll pathway requires binding of Toll with its ligand Spätzle. Here we described a Spätzle-like protein, designated as Fc-Spz, from hemocytes of Chinese shrimp, Fenneropenaeus chinensis. The deduced amino acid sequence of Fc-Spz shares 54% identity with Spätzle-like protein of salmon louse ( Lepeophtheirus salmonis). Semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR), quantitative real time PCR, and Western blot analyses were carried out to analyze the expression pattern and distribution profile of Fc-Spz in shrimp after challenged with bacteria and virus. The results showed that Fc-Spz mRNA was up-regulated in all the tissues tested in shrimp injected with Vibrio anguillarum and white spot syndrome virus (WSSV). The C-terminal active Fc-Spz domain (114 residues) was expressed in Escherichia coli and purified by affinity chromatography. The recombinant Fc-Spz C-114 was injected into crayfish ( Procambarus clarkii) to determine the expression levels of several antimicrobial peptide genes. The results showed that recombinant Fc-Spz C-114 could up-regulate crustin 2 expression in crayfish. These results suggest that Fc-Spz may play a role in the innate immune defence of Chinese shrimp and crayfish.