Molecular cloning and expression analysis of a hepcidin antimicrobial peptide gene from turbot (Scophthalmus maximus)

Antimicrobial peptides (AMPs) are regarded as important components of the host innate immune system and play crucial roles in host defence against microbial invasion. A small number of hepcidin AMPs have been isolated from teleosts. Here, we report the isolation of a hepcidin gene from the liver of turbot (Scophthalmus maximus) (GenBank accession numbers: AY994074 and AY994075). In the 1037 bp-long genomic sequence, three exons and two introns were identified. The full-length cDNA is 778 bp long and contains an ORF of 273 bp encoding a prepropeptide of 90 amino acid residues. The predicted prepropeptide consists of three domains: a signal peptide (24 amino acids), a prodomain (40 amino acids) and a mature peptide (26 amino acids). RT-PCR demonstrated that hepcidin transcripts were highly abundant in liver, abundant in heart, head kidney, spleen, skin and gill, less abundant in blood cell, gonad and intestine, and undetectable level in muscle. The level of the hepcidin mRNA in embryos gradually increases during embryogenesis from 2 h (2 cell stage) to 95 h (larva stage) after fertilisation. Challenge of turbot with pathogenic bacteria, Listonella anguillarum, significantly elevated hepcidin mRNA levels in liver and spleen in a time-dependent fashion. The hepcidin transcripts were detected in turbot embryonic cell line (TEC). Challenge of TEC cells with the pathogenic bacteria significantly elevated hepcidin mRNA levels.