Molecular identification and functional characterization of a WAP domain-containing protein in Apostichopus japonicus

Abstract

WAP domain-containing proteins are important immune effectors that play vital roles in regulating host immune response, such as anti-microbial invasion and tissue repair at mucosal surfaces. In our study, the whole-length cDNA of a protein with a single WAP domain was cloned from Apostichopus japonicus (termed AjWFDC-like), and its roles in immune defense were further investigated. The AjWFDC-like protein shared a 17 aa signal peptide, a typical WAP domain (23–69 aa), a KAZAL domain (76–22 aa), and a thyroglobulin type I domain (156 aa-204 aa). AjWFDC-like mRNA was found in all examined tissues, and its abundance was the highest in the body wall. AjWFDC-like mRNAs in coelomocytes, body walls, and skin mucus were all induced in large quantities after Vibrio splendidus treatment. Indirect immunofluorescence assay showed that AjWFDC-like was mainly distributed in the epithelial cells of the body wall. Moreover, recombinant AjWFDC-like (rAjWFDC-like) showed higher binding activities to lipopolysaccharide, peptidoglycan and mannan and obvious agglutinating effects on Vibrio harveyi, Vibrio anguillarum, V. splendidus, and Vibrio parahaemolyticus. Furthermore, rAjWFDC-like also displayed strongly antimicrobial abilities against V. harveyi and V. anguillarum. Scanning electron microscopy revealed that the membrane integrity of these two bacteria was destroyed by rAjWFDC-like administration, which caused the leakage of cell contents and death. Together, our data showed that AjWFDC-like plays a vital role in the immune defense of A. japonicus by eliminating invasive microorganisms.