Molecular identification and expression analysis of tumor necrosis factor in channel catfish ( Ictalurus punctatus)

Abstract

A tumor necrosis factor (TNF) α-like gene, encoding a propeptide of 230 amino acids and a mature (soluble) peptide of 162 amino acids, was identified in channel catfish ( Ictalurus punctatus). While the catfish protein shared features in common with both mammalian TNFα and TNFβ homologs, overall sequence identity/similarity was slightly higher vs. TNFα genes when mature TNF sequences were compared. Phylogenetic analysis placed catfish and other fish TNF sequences within their own cluster apart from mammalian TNFα and β genes, and supported the suggestion that TNFα and β genes separated after the divergence of mammals and teleosts. In contrast to trout and carp, but similar to flounder, catfish TNF was present as a single copy gene. Expression studies demonstrated that catfish TNFα mRNA was present in all tested tissues (i.e. liver, spleen, head kidney, mesonephros, gill, thymus, and PBLs) from an unstimulated fish. Moreover, catfish TNF was constitutively expressed in actively proliferating, but otherwise unstimulated, macrophage (42TA) and T cell (G14D; TS32.17) lines, but not in B cell (1G8 or 3B11) or fibroblast lines. TNF expression was upregulated in PBLs, and in G14D and 42TA cells, but not in 3B11 cells, by PMA/calcium ionophore treatment. These results demonstrate that a catfish homolog of TNFα has been identified, and indicate that catfish TNFα is expressed in catfish in a manner similar to that seen in mammals.