Cloning and expression analysis of rainbow trout Oncorhynchus mykiss tumour necrosis factor-alpha.
A rainbow trout (Oncorhynchus mykiss) gene for tumor necrosis factor (TNF) has been cloned and sequenced. The cDNA contains an open reading frame of 738 nucleotides that translate into a 246 amino-acid putative peptide, with a 5' untranslated region (UTR) of 140 bp and a 3' UTR of 506 bp. Two potential N-linked glycosylation sites exist in the translation. The genomic sequence measures 2007 bp and contains three introns that intercept four coding exons. Expression studies using RT-PCR have shown that the trout TNF gene is constitutively expressed in the gill and kidney of unstimulated fish. Trout TNF expression could be up-regulated by stimulation of isolated head kidney leucocytes with lipopolysaccharide (LPS). Similarly, stimulation of a trout macrophage cell line (RTS11) with LPS resulted in an increased transcript level, as did incubation with recombinant trout interleukin (IL)-1 beta. The optimal timing for induction of TNF expression in trout macrophages was determined using recombinant trout IL-1 beta, where a clear induction was apparent by 2 h and peaked at 4 h. Evidence that this TNF gene is equivalent to mammalian TNF-alpha is discussed.
Department of Zoology, University of Aberdeen, Aberdeen, UK.
K J
KJ
Laing
T
T
Wang
J
J
Zou
J
J
Holland
S
S
Hong
N
N
Bols
I
I
Hirono
T
T
Aoki
C J
CJ
Secombes
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D005091:Exons / Q000235:genetics; D020869:Gene Expression Profiling; D015723:Gene Library; D005880:Gills / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology; D007438:Introns / Q000235:genetics; D007668:Kidney / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism; D008969:Molecular Sequence Data; D017683:Oncorhynchus / Q000097:blood / Q000235:genetics* / Q000276:immunology*; D016366:Open Reading Frames / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D015533:Transcriptional Activation / Q000187:drug effects; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry / Q000235:genetics*
D007375:Interleukin-1; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D014409:Tumor Necrosis Factor-alpha