Identification of an additional two-cysteine containing type I interferon in rainbow trout Oncorhynchus mykiss provides evidence of a major gene duplication event within this gene family in teleosts

Abstract

Multiple type I interferons (IFNs) have recently been identified in salmonids, containing two or four conserved cysteines. In this work, a novel two-cysteine containing (2C) IFN gene was identified in rainbow trout. This novel trout IFN gene (termed IFN5) formed a phylogenetic group that is distinct from the other three salmonid IFN groups sequenced to date and had a close evolutionary relationship with IFNs from advanced fish species. Our data demonstrate that two subgroups are apparent within each of the 2C and 4C type I IFNs, an evolutionary outcome possibly due to two rounds of genome duplication events that have occurred within teleosts. We have examined gene expression of the trout 2C type I IFN in cultured cells following stimulation with lipopolysaccharide, phytohaemagglutinin, polyI:C or recombinant IFN, or after transfection with polyI:C. The kinetics of gene expression was also studied after viral infection. Analysis of the regulatory elements in the IFN promoter region predicted several binding sites for key transcription factors that potentially play an important role in mediating IFN5 gene expression.

Intron-Containing Type I and Type III IFN Coexist in Amphibians: Refuting the Concept That a Retroposition Event Gave Rise to Type I IFNs

“…Two major groups of fish type I IFNs were also revealed, as seen previously, with each duplicated into further subgroups, supporting our previous hypothesis that at least some teleost species appear to have undergone two major IFN duplication events (9).…”

“…It is still possible that type III IFN homologs could be present in fish and awaits further study. Recent studies with salmonids also highlight that it is not necessary to have intronless IFN genes for multiple gene duplications to occur, with the salmon genome consisting of at least 11 intron-containing genes (8,9). IFN-like genes have been found in cartilaginous fish, such as elephant shark (7), and these predicted proteins possess a CAWE motif that is present in four cysteine containing type I IFNs among all vertebrate orders and also have a close evolutionary relationship with vertebrate type I IFNs (data not shown), supporting our previous hypothesis that type I IFNs could have originated from an ancestor containing four conserved cysteines (7).…”

“…Recent identification of the IFN genes from fish, amphibians, and reptiles has provided enormous information on IFN gene evolution (6)(7)(8)(9)(10)(11)(12). It has been generally accepted that the IFNs originated from an ancient progenitor, which gave rise to the IL-10 family cytokines early in evolution (13).…”

The Two Groups of Zebrafish Virus-Induced Interferons Signal via Distinct Receptors with Specific and Shared Chains

“…1B). A further subdivision of each of these groups in two subgroups has been recently proposed by Chang et al (28); however, when we included the new zebrafish IFN4 sequence in this phylogeny, it was not possible to ascribe it with a significant bootstrap value to one or the other subgroup of 2-cysteine IFNs (Fig. 1C).…”

Cloning of two rainbow trout nucleotide-binding oligomerization domain containing 2 (NOD2) splice variants and functional characterization of the NOD2 effector domains

“…RTG-2 cells (5 Â 10 6 cells in 100 ml nucleofector solution) were electroporated with 5 ml polyI:C (1 mg/ml) or 5 ml PBS using an Amaxa Nucleofector II transfection system (Lonza) under Programme T20, washed immediately with 1 ml HBSS buffer and cultured at 20 C for 24 h. The cells were collected for RNA extraction and real time PCR analysis as described previously [21]. Namely after total RNA was extracted using the RNASTAT reagent (AMS Biotechnology (Europe) Ltd) and treated with RNase free DNase I (Fermentas Life Sciences, Germany), the RNA was reverse transcribed into cDNA using a first strand cDNA synthesis kit (Fermentas Life Sciences, Germany).…”