Extra-erythrocytic Expression of Antimicrobial Peptides Derived from the beta-subunit of Hemoglobin is Associated with a Potent Anti-parasitic Defense in Fish.

Abstract

Innate immunity plays a crucial role in the defense against prokaryotic and eukaryotic pathogens. Major components of this defense are antimicrobial peptides (AMPs). Some AMPs are derived from larger proteins with other recognized functions (e.g., lactoferrin, histones). In this study, we demonstrate the expression of peptides homologous to the β-chain of hemoglobin (Hb-β), one of the two major subunits of this respiratory protein. These Hb-β peptides (HbβP-1, -2 and -3), isolated from gill of the economically important channel catfish (Ictalurus punctatus), had antibacterial activity and were upregulated in gill and skin epithelium in response to parasitic (Ichthyophthirius multifiliis, ich) infection. One peptide (HbβP-1), while having relatively weak antibacterial activity, had antiparasitic activity comparable to that of other potently antiparasitic AMPs. Also, this cidal activity was specifically directed against the trophozoite (trophont) stage of ich at a low concentration (6.2 ug⁄ml, 1.7 mM) but had no apparent effect on the disseminative (theront) stage or the reproductive (tomont) stage at the highest concentration tested (400 mg⁄ml, 108 mM). In addition, HbβP-1 was not lytic to channel catfish erythrocytes at the highest concentration tested (400 mg⁄ml, 108 mM). Immunohistochemistry and in situ hybridization of skin and gill from fish experimentally challenged with ich indicated that the HbβP-1 sequence was both synthesized and expressed in epithelial tissues of skin and gill, which are the target tissues for ich. "Bug blots" of gill extracts from fish recovering from a bacterial infection suggested that upregulation of these Hb-β related peptides might also occur with other infections. These findings, along with the recent discovery by others that Hb-β is expressed in mammalian macrophages and alveolar epithelium, suggest that hemoglobin-derived AMPs might play a significant role in the non-specific immune response of vertebrates.