Molecular cloning, characterization and expression analysis of a novel wap65-1 gene from Plecoglossus altivelis

Abstract

Warm temperature acclimation associated 65-kDa protein 1 (WAP65-1) is a specific fish plasma glycoprotein that is possibly involved in various physiological or pathological processes. In this study, we obtained the cDNA and genomic DNA sequences of the Plecoglossus altivelis wap65-1 (Pawap65-1) gene. Multiple sequence alignment showed that Pawap65-1 is similar in structure to wap65-1 in fish. Phylogenetic analysis revealed that Pawap65-1 is most closely related to that of a rainbow trout. Pawap65-1 transcripts are present in various tissues and are most abundant in the liver. We expressed recombinant PaWAP65-1 in Escherichia coli and raised antiserum against it in mouse. Western blot analysis revealed that the higher molecular mass of PaWAP65-1 in blood plasma was caused by post-translational N-glycosylation. Quantitative real-time quantitative PCR (qPCR) and Western blot analysis data showed that the hepatic mRNA and blood plasma levels of PaWAP65-1 were both influenced by warm temperature acclimation and cadmium exposure, but not by Listonella anguillarum infection, hypo-osmotic, or cold temperature acclimation. In conclusion, our data reveals that PaWAP65-1 is a stress-related protein, and may play a role in fish acclimation to warm temperature and cadmium exposure.