Molecular Identification of C-type Lectin Receptor Gene CD302 from Japanese sea bass (Lateolabrax japonicus) and Its Expression upon Vibrio harveyi Infection
ZHANG Yi-Rong, LI Chang-Hong*, CHEN Jiong
Laboratory of Biochemistry and Molecular Biology, School of Marine Sciences, Ningbo University, Ningbo 315832, China
Abstract：Cluster of differentiation 302 (CD302), also known as DEC-205-associated C-type lectin-1 (DCL-1), is a type I transmembrane C-type lectin receptor. It participates in the processes of cell adhesion, migration, endocytosis and phagocytosis, and plays an important role in resisting pathogenic infection. In this study, the LjCD302 cDNA sequence (GenBank No. MT468568) was obtained from Japanese sea bass (Lateolabrax Japonicus) tissue transcriptome. It spanned 2 280 nucleotides (nts) and comprised an open reading frame of 729 nts, which encoded a polypeptide of 242 amino acids (aa) with a molecular weight of 26.9 kD and an isoelectric point of 4.97. LjCD302 composed of a signal peptide sequence (aa 1-27), a C-type lectin-like domain (CTLD) (aa 30-165), a transmembrane domain (aa 179-201) and a cytoplasmic tail (aa 202-242). The aa sequence homology analysis showed that LjCD302 shared the highest homology with that of the barramundi perch (Lates calcarifer) (82.0%). Phylogenetic tree analysis showed that CD302 from fish, amphibians, reptiles and mammals grouped into different clusters, and LjCD302 belonged to the fish cluster and had the closest evolutionary relationship with the barramundi perch CD302. Real-time quantitative PCR analysis showed that LjCD302 mRNA in healthy Japanese sea bass was mainly expressed in the liver, followed by the head kidney, and its expression was significantly up-regulated in head kidney, gill, intestine, spleen, and liver of Japanese sea bass after Vibrio harveyi infection (P<0.05). The CTLD domain of LjCD302 was prokaryotically expressed, and its antiserum was prepared. Western blot analysis showed that LjCD302 expression was significantly up-regulated in the head kidney of V. harveyi-infected Japanese sea bass (P<0.05). In summary, the expression of LjCD302 mRNA and protein was positively correlated with V. harveyi infection, suggesting that it may play a crucial role in the antibacterial immune response in fish. The results provide a basis for further research on the function of fish CLR and its mechanism of action in antibacterial immunity.
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