Characterization and bacterial-binding activity of a novel C-type lectin from the red-spotted grouper, Epinephelus akaara

State Key Laboratory of Marine Environmental Science, Xiamen University, Xiamen, Fujian, China.
Genetics and molecular research: GMR (Impact Factor: 0.78). 05/2012; 11(3):2958-71. DOI: 10.4238/2012.May.11.1
Source: PubMed


Because of their specific binding to carbohydrates, lectins play a crucial role in pathogen recognition and clearance in vertebrate animals. Previously, only two types of collectins had been isolated from bony fish: mannan-binding lectin (MBL) and galactose-binding lectin (GalBL). We sequenced a novel collectin (designated EALec1) from the red-spotted grouper, Epinephelus akaara. The gene structure of EALec1 and the alignment of the carbohydrate recognition domain of the three collectins demonstrated that EALec1 is a new type of collectin derived from MBL. We examined the expression pattern of the EALec1 transcripts in 12 tissues of the red-spotted grouper. The EALec1 gene was found to have multiple copies; their transcripts were detected in all 12 tissues. EALec1 was also recombined and expressed in Escherichia coli to investigate its immune functions and carbohydrate-binding characterization. We concluded that EALec1 belongs to the mannan-binding lectin group, despite its different Ca(2+)-dependent sites in the carbohydrate recognition domain, and that it is involved in the recognition and clearance of invaders in the red-spotted grouper.

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    ABSTRACT: C-type lectin-like receptors (CLRs) are important pathogen pattern recognition molecules that recognize carbohydrate structures. However, the functions of these receptors in fish keep less known. In this study, we characterized a novel CLR from a teleost fish, Plecoglossus altivelis (ayu), tentatively named PaCD209L. The cDNA of PaCD209L is 1464 nucleotides (nts) in length, encoding a polypeptide of 281 amino acid residues with a calculated molecular weight of 31.5 kDa. Multiple alignment of the deduced amino acid sequences of PaCD209L and other related fish CLRs revealed that the PaCD209L sequence had typical characteristics of fish CLRs, but without Ca(2+)-binding sites. Sequence comparison and phylogenetic tree analysis showed that PaCD209L shared the highest amino acid identity (44%) with rainbow trout (Oncorhynchus mykiss) CD209aE. PaCD209L transcripts were detected in all of the tissues examined, mainly expressed in the brain and heart. Upon Vibrio anguillarum infection, PaCD209L transcripts were upregulated in all tested tissues and in monocytes/macrophages (MO/MΦ). We prepared recombinant PaCD209L (rPaCD209L) by prokaryotic expression and raised antiserum against PaCD209L. Western blot analysis revealed that native PaCD209L was glycosylated, and its protein expression significantly increased in ayu MO/MΦ upon V. anguillarum infection. In addition, rPaCD209L was able to bind Gram-positive and Gram-negative bacteria in the absence of Ca(2+). After PaCD209L was blocked by anti-PaCD209L IgG, the phagocytosis and bacterial killing activity of MO/MΦ significantly decreased. These results suggest that PaCD209L plays an important role in the regulation of MO/MΦ functions in ayu. Copyright © 2015. Published by Elsevier Ltd.
    Full-text · Article · Apr 2015 · Fish & Shellfish Immunology