Molecular characterization of tumor necrosis factor receptors 1 and 2 of the goldfish (Carassius aurutus L.).
Tumor necrosis factor alpha (TNFalpha) is a pleotropic cytokine that mediates its effects by binding to one of two TNF receptors, TNF-R1 or TNF-R2. We have recently identified the cDNA sequences of both goldfish TNF-R1 and TNF-R2. In silico analyses revealed conserved cysteine rich domains, predicted docking sites for TNFR-specific downstream signaling factors for both receptors and a conserved death domain for TNF-R1. The expression of these receptors in tissues and various immune cell types was investigated by Q-PCR. The TNF-R2 expression was substantially higher than that of TNF-R1 in all tissues. Both receptors were most robustly expressed in monocytes where as the mRNA levels of TNF-R1 were the lowest in mature macrophages and those of TNF-R2 in peripheral blood leukocytes. Treatment of goldfish macrophages with recombinant goldfish (rg) TNFalpha-2, rgIFN gamma or rgTGF beta differentially altered the expression of both TNF receptors. The rgTNF alpha-2 up-regulated the expression of TNF-R2 but down-regulated the expression of TNF-R1. The rgIFN gamma increased the expression of both TNF receptors while rgTGF beta caused a time-dependent decreases in mRNA of goldfish TNF-R1 and TNF-R2. In vitro binding studies using recombinant TNFalpha-1 and TNFalpha-2 revealed that either isoform was capable of interacting with the recombinant forms of the extracellular domains of either TNF-R1 or TNF-R2. Functional significance of these ligand-receptor interactions was confirmed by experiments showing that goldfish TNF-R1 and TNF-R2 down-regulated the rgTNF alpha-1 or rgTNF alpha-2-primed respiratory burst response of goldfish macrophages.
Grayfer
Leon
L
Department of Biological Sciences, University of Alberta, Edmonton, Alberta T6G 2E9, Canada.
Belosevic
Miodrag
M
D000595:Amino Acid Sequence; D000818:Animals; D006054:Goldfish / Q000235:genetics / Q000378:metabolism*; D007371:Interferon-gamma / Q000494:pharmacology; D007962:Leukocytes / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000378:metabolism*; D047888:Receptors, Tumor Necrosis Factor, Type I / Q000235:genetics / Q000378:metabolism*; D047889:Receptors, Tumor Necrosis Factor, Type II / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000494:pharmacology; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism / Q000494:pharmacology
D012333:RNA, Messenger; D047888:Receptors, Tumor Necrosis Factor, Type I; D047889:Receptors, Tumor Necrosis Factor, Type II; D011994:Recombinant Proteins; D016212:Transforming Growth Factor beta; D014409:Tumor Necrosis Factor-alpha; D007371:Interferon-gamma