Cloning and characterization of cDNA clones encoding CD9 from Atlantic salmon (Salmo salar) and rainbow trout (Oncorhynchus mykiss)

Abstract.

In comparison to mammals, relatively few of the molecules involved in teleost immune responses have been isolated and characterized. A rapid method of isolating molecules important for immune function is subtractive hybridization. One such experiment using infectious hematopoietic necrosis virus-infected Atlantic salmon produced several cDNA clones with similarity to mammalian immune-specific genes, including granzyme M (accession no. AF434669) and CD9. After cloning the rainbow trout version of CD9, sequence analysis showed that both salmonid sequences contained many features of the tetraspanin receptor family to which CD9 belongs. Phylogenetic analysis revealed a close association of the trout and salmon sequences to known CD9 and CD81 receptors. Southern blotting demonstrated that the rainbow trout gene is single copy. Reverse transcriptase PCR showed strong expression of this clone in many tissues, but liver expression was very low – an observation consistent with the clone being a CD9, not a CD81, equivalent. The evidence suggests that the sequences reported here are bona fide teleost CD9 homologues and we are currently producing recombinant proteins and polyclonal antisera for use in functional studies.