Identification and analysis of a Sciaenops ocellatus ISG15 homologue that is involved in host immune defense against bacterial infection.

ISG15 is an interferon-stimulated gene that encodes a ubiquitin-like protein. ISG15 homologues have been identified in a number of fish species, some of which are known to be regulated at expression level by virus infection and lipopolysaccharide (LPS) treatment. However, the relationship between ISG15 and live bacterial infection has not been investigated in piscine models. In this study, an ISG15 homologue, SoISG15, was identified from red drum Sciaenops ocellatus and analyzed at expression and functional levels. The open reading frame of SoISG15 is 477 base pairs (bp) and intronless, with a 5'-untranslated region (UTR) of 91 bp and a 3'-UTR of 415 bp. The deduced amino acid sequence of SoISG15 shares 60-67% overall identities with the ISG15 of several fish species. SoISG15 possesses two conserved ubiquitin-like domains and the canonical ubiquitin conjugation motif, LRGG, at the C-terminus. Expressional analysis showed that constitutive expression of SoISG15 was highest in blood and lowest in kidney. Experimental challenges with LPS and bacterial pathogens induced significant SoISG15 expression in the kidney but not in the liver. Similar differential induction was also observed at cellular level with primary hepatocytes and head kidney (HK) lymphocytes. Poly(I:C), however, effected drastic induction of SoISG15 expression in kidney and liver at both tissue and cellular levels. Immunoblot analysis showed that SoISG15 was secreted by cultured HK lymphocytes into the extracellular milieu. Recombinant SoISG15 expressed in and purified from Escherichia coli was able to enhance the respiratory burst activity, acid phosphatase activity, and bactericidal activity of HK macrophages. Taken together, the results of this study indicated that SoISG15 possesses apparent immunological property and is likely to be involved in host immune defense against bacterial infection.