A cDNA clone, designated sbIL-6 (seabream interleukin-6), was obtained from a cDNA library of enriched immune-stimulated sequences from gilthead seabream. The deduced sbIL-6 protein corresponds to a 225-amino acid protein with a putative 24-amino acid signal peptide, four conserved alpha helices and one N-linked glycosylation site. At the amino acid level sbIL-6 shares 23-26% identity with mammalian IL-6 sequences and 30-51% identity with other fish IL-6 sequences. The structure of the sbIL-6 gene consisted of 5 exons and 4 introns, spanning 2.4 kb. Healthy fish expressed sbIL-6 in white muscle, skin, spleen, anterior intestine and stomach, while no expression was detected in brain, gill, head kidney, posterior intestine and adipose tissue. A significant up-regulation of sbIL-6 expression was observed after lipopolysaccharide (LPS), Vibrio anguillarum DNA (VaDNA) and peptidoglycan treatment in cultured seabream head kidney leukocytes. Using purified immune cells, sbIL-6 expression was induced similarly in macrophages and acidophilic granulocytes by VaDNA but LPS was more effective in inducing sbIL-6 expression in acidophilic granulocytes than in macrophages. Furthermore, in vivo infection of seabream with live V. anguillarum caused significant increases in sbIL-6 mRNA expression in the thymus, peritoneal exudate, head kidney and gills. In summary, our study provides further evidence for the existence of distinct IL-6 genes in lower vertebrates and for the strong induction of their expression by immune stimuli, supporting the notion of a potentially important role for this cytokine in fish. © 2008 Elsevier Ltd. All rights reserved.
- mRNA expression