, Volume 51, Issue 12, pp 1002-1011

Cloning, sequencing, and analysis of expression of a second IL-1 β gene in rainbow trout (Oncorhynchus mykiss)

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Abstract.

The full-length sequence of a second IL-1 β gene (IL-1 β 2) in rainbow trout (Oncorhynchus mykiss) has been obtained. As with the first IL-1 β gene, IL-1 β 2 is organized into six exons/five introns. There are only small differences in their intron/exon sizes, with the exception of intron 3, which is 334 bp smaller in IL-1 β 2. The transcript encoded by the IL-1 β 2 gene contains a 5′ untranslated region (UTR) of 121 bp, followed by a 762-bp open reading frame and a 518-bp 3′UTR. The 3′UTR contains seven instability attta motifs, typical of inflammatory genes, and a polyadenylation site 11 bp upstream of a 17-bp poly(A) tail. The predicted 254 amino acid sequence of the second IL-1 β gene has 82% similarity to the first gene, 45% similarity to carp IL-1β, and 40% similarity to human IL-1β. Comparison of the two trout genes reveals that the IL-1 β 2 gene has a deletion of 9 bases in exon 3 and an altered splicing site at the 5′ end of exon 4 giving rise to a further 9-bp deletion in the resulting cDNA. As with other nonmammalian IL-1 β genes, no interleukin-converting enzyme (ICE) cut site has been found but the alignment of the amino acid sequence with other species shows a possible cut site between Arg89 and Ala90 that would give arise to a 165-amino acid mature peptide. Expression studies performed by RT-PCR using primers specific for the IL-1 β 2 transcript revealed a clear dose-dependent induction of this gene in cultured trout leukocytes by stimulation with lipopolysaccharide.

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