Black rockfish C-type lectin, SsCTL4: A pattern recognition receptor that promotes bactericidal activity and virus escape from host immune defense.

C-type lectin (CTL) is an immune receptor and is received extensive attention of its important roles in immune response and immune escape. Some CTL, such as CTL4, has been well characterized in human and several other mammals, but much less documentation exists about the immunological function of CTL4 in lower vertebrates. In the present study, a C-type lectin domain family 4 member, SsCTL4, which is also high homology with CD209 antigen-like protein, from the teleost fish black rockfish (Sebastes schlegelii) was identified and examined at expression and functional levels. The open reading frame of SsCTL4 is 765 bp, and the deduced amino acid sequence of SsCTL4 shares 78%-84% overall identities with the C-type lectin of several fish species. In silico analysis identified several conserved C-type lectin features, including a carbohydrate-recognition domain and four disulfide bond-forming cysteine residues. Expression of SsCTL4 occurred in multiple tissues and was upregulated during bacterial and viral infection. Recombinant SsCTL4 (rSsCTL4) exhibited apparent binding activities against bacteria (Edwardsiella tarda and Vibrio anguillarum) and virus (infectious spleen and kidney necrosis virus, ISKNV). rSsCTL4 was able to agglutinate the Gram-negative and Gram-positive bacteria in a Ca2+-dependent manner. The agglutinating ability of rSsCTL4 was abolished in the absence of calcium or presence of mannose. rSsCTL4 also increased macrophage bactericidal activity. In the presence of rSsCTL4, fish exhibited enhanced resistance against bacterial infection but increased susceptibility to viral infections. Collectively, these results indicate that SsCTL4 serves as a pattern recognition receptor that not only promotes bactericidal activity, but may also serve as targets for virus manipulation of host defense system.

A high-mobility group box 1 that binds to DNA, enhances pro-inflammatory activity, and acts as an anti-infection molecule in black rockfish, Sebastes schlegelii.

High-mobility group box (HMGB) 1 is a chromosomal protein that plays critical roles in DNA transcription, replication and repair. In addition, HMGB1 functions as a pro-inflammatory molecule in many vertebrates and invertebrates. In teleosts, very limited studies of HMGB1 have been reported. In this study, we identified a HMGB1 homologue (SsHMGB1) from black rockfish (Sebastes schlegelii) and analyzed its structure, expression and biological function. The open reading frame of SsHMGB1 is 621 bp, with a 5'-untranslated region (UTR) of 62 bp and a 3'-UTR of 645 bp. SsHMGB1 contains two typical HMG boxes and an acidic C-terminal tail. The deduced amino acid sequence of SsHMGB1 shares the highest overall identity (89.4%) with the HMGB1 of Anoplopoma fimbria. The expression of SsHMGB1 occurred in multiple tissues and was highest in the brain. Moreover, the mRNA level of SsHMGB1 in head kidney (HK) macrophages could be induced by Listonella anguillarum in a time-dependent manner. Recombinant SsHMGB1 purified from Escherichia coli (i) bound DNA fragments in a dose-dependent manner; and (ii) induced the expression of cytokines in HK macrophages, including a significant increase in TNF-α activity and enhanced mRNA level of TNF13B and IL-1 ß, which are known to be involved in antibacterial defense; moreover, (iii) significantly improved the macrophage bactericidal activity together with reduced pathogen dissemination and replication of bacteria in fish kidney. These results indicated that SsHMGB1 is a novel HMGB1 that possesses apparent immunoregulatory properties and is likely to be involved in fighting bacterial infection.