Cloning and characterisation of cytokine and cytokine receptor genes in rainbow trout, Oncorhynchus mykiss
Convincing molecular evidence for the existence of both cytokines and their receptors in teleost fish is presented. TGF-β is present in O. mykiss encoding a 112 amino acid mature peptide. An integrin binding site (RGD) and a characteristic tetrabasic cleavage site (RKKR) are present, as is the TGF-β superfamily motif. The mature peptide has 9 conserved cysteine residues (8 of which occur in pairs) as well as two additional conserved TGF-β superfamily residues (Pro36 and Gly46). TGF-β exhibits a wide range tissue distribution including head-kidney macrophages, PBL, brain, gill and spleen tissue, and is encoded by a 2.5Kb mRNA. The trout TGF-β gene is spread over 7 exons, with an additional intron in exon 7 when compared to mammalian and avian models. Isolation of a partial sequence also reveals the presence of TGF-β in a cyprinid species. Phylogenetic analysis suggests trout TGF-β to cluster with mammalian TGF-β1 isoforms, and the avian (TGF-β4) and amphibian (TGF-β5) homologs. Neither TNF-α or TNF receptors were detected in O. mykiss at either the cellular or molecular level. The use of degenerate primers in PCR lead to the isolation of a partial sequence for O. mykiss MHC class I. A full length CXC-R gene of 1.6Kb isolated from O. mykiss displays approximately 65% identity to mammalian CXC-R4 receptors, exhibits the seven-transmembrane domain structure of the G-protein coupled receptors and a tissue-specific distribution. Characteristic superfamily motifs and a putative glycosylation site are present in the sequence. Along with the major features of the adaptive immune response such as the major histocompatibility complex (MHC), T-cell receptor (TCR) and immunoglobulin (Ig), cytokines are now shown to be present at the level of teleost fish.