Cloning and expression analysis of three striped trumpeter (Latris lineata) pro-inflammatory cytokines, TNF-α, IL-1β and IL-8, in response to infection by the ectoparasitic, Chondracanthus goldsmidi

This study reports the cloning and sequencing of three striped trumpeter (Latris lineata Forster) pro-inflammatory cytokines, TNF-α, IL-1β and IL-8, as well as their differential expression in response to an infection by the ectoparasite Chondracanthus goldsmidi. The striped trumpeter TNF-α transcript consisted of 1093 bp, including a 759 bp ORF which translated into a 253 aa transmembrane peptide. The sequence contained a TACE cut site, that would produce a 167 aa soluble peptide containing the TNF ligand family signature. The IL-1β sequence consisted of 963 bp, including a 774 bp ORF which translated into a 258 aa protein. The protein lacked both a signal peptide and an ICE cleavage site, but did contain the IL-1 family signature. The sequence for the chemokine IL-8 contained 906 bp, with an ORF of 297 bp, which translated into a 99 aa protein. The protein lacked an ELR motif as is common with many teleost IL-8 sequences. The differential expression of the three cytokine genes in parasitized fish was investigated via quantitative real-time PCR. A significant up-regulation of all three pro-inflammatory cytokines was found in the gills, which were the site of parasite attachment. Examination of head kidney cells revealed a significant up-regulation of TNF-α, but not IL-1β or IL-8. Conversely, the spleen cells showed significant up-regulation of both IL-1β and IL-8, but not TNF-α. These findings allow for more detailed investigations of the striped trumpeter immune response.