Interleukin (IL)-2 Is a Key Regulator of T Helper 1 and T Helper 2 Cytokine Expression in Fish: Functional Characterization of Two Divergent IL2 Paralogs in Salmonids

Abstract

Mammalian interleukin (IL)-2 is a cytokine centrally involved in the differentiation and survival of CD4+ T helper subsets and CD4+ T regulatory cells and in activation of cytotoxic effector lymphocytes. In bony fish, <i>IL2</i> orthologs have been identified with an additional divergent <i>IL2-Like</i> gene on the same locus present in several fish species. We report here two divergent <i>IL2</i> paralogs, <i>IL2A</i> and <i>IL2B</i>, in salmonids that originated from the whole genome duplication event in this fish lineage. The salmonid <i>IL2</i> paralogs differ not only in sequence but also in exon sizes. The IL-2 isoforms that are encoded have disparate pI values and may have evolved to preferentially bind specific IL-2 receptors. Rainbow trout <i>IL2</i> paralogs are highly expressed in thymus, spleen, gills, kidney and intestine, important tissues/organs in fish T cell development and function. Their expression in peripheral blood leukocytes (PBL) is low constitutively but can be upregulated by the mixed leukocyte reaction, by the T cell mitogen phytohemagglutinin and by signal mimics of T cell activation (phorbol 12-myristate 13-acetate and calcium ionophore). Both trout IL-2 isoforms promoted PBL proliferation and sustained high-level expression of <i>CD4</i> and <i>CD8</i>, suggesting that trout IL-2 isoforms are T cell growth/survival factors mainly expressed by activated T cells. The recombinant proteins for these two trout <i>IL2</i> paralogs have been produced in <i>E. coli</i> and possess shared but also distinct bioactivities. IL-2A, but not IL-2B, induced <i>IL12P35A1</i> and <i>CXCR1</i> expression in PBL. IL-2B had a stronger effect on upregulation of the T helper 1 (Th1) cytokine <i>interferon-γ</i> (<i>IFNγ</i>) and could sustain <i>CD8α</i> and <i>CD8β</i> expression levels. Nevertheless, both cytokines upregulated key Th1 (<i>IFNγ1, IFNγ2, TNFα2</i> and <i>IL12</i>) and T helper 2 (Th2) cytokines (<i>IL4/13B1</i> and <i>IL4/13B2</i>), cytokine and chemokine receptors and the antimicrobial peptide <i>cathelicidin-1</i> but had limited effects on T helper 17 cytokines and <i>TGFβ1</i> in PBL. They could also enhance PBL phagocytosis. These results suggest, for the first time in fish, that IL-2 isoforms may have an important role in regulating Th1 and Th2 cell development, and innate and adaptive host defenses in fish, and shed light on lineage-specific expansion, evolution, and functional diversification of <i>IL2</i> in vertebrates.