title: The expression of two novel orange-spotted grouper (Epinephelus coioides) TNF genes in peripheral blood leukocytes, various organs, and fish larvae.
abstract: The tumour necrosis factor (TNF) super-family is a group of important cytokines involved in inflammation, apoptosis, cell proliferation, and the general stimulation of the immune system. The TNF gene has been cloned in some bony fish; however, its counterparts are still unidentified in the majority of fish species. In this study, we cloned gTNF-1 and gTNF-2 from the orange-spotted grouper (Epinephelus coioides), an economically important farmed fish. Both genes include 4 exons and 3 introns and encoded 253 and 241 amino acid proteins with a molecular weight of approximately 27 and 26 kDa, respectively. The identity of the putative amino acid sequences between gTNF-1 and gTNF-2 was only 38%. The positions of cysteine residues, a protease cleavage site, and a transmembrane domain sequence derived from gTNF-1 and gTNF-2 were similar to those in other fish and mammalian TNF-α. The mRNA expression levels of the 2 gTNF molecules were evaluated in unstimulated/stimulated peripheral blood leukocytes, various organs, and fish larvae. Following lipopolysaccharide (LPS) treatment, gTNF-2 was expressed at higher levels, was up-regulated more quickly, and was more sensitive to the immune response than gTNF-1. gTNF-1 was constitutively expressed in the thymus, brain, and spleen, but it was also expressed in the heart, head kidney, and trunk kidney after LPS stimulation. gTNF-2 was constitutively expressed in the thymus, head kidney, trunk kidney, spleen, and intestine; further, gTNF-2 was highly expressed in all organs post-LPS stimulation. Finally, the gTNF expression levels were evaluated at various developmental stages in grouper larvae. A higher variation of gTNF expression levels was observed in fish larvae from a contaminated hatchery. This study revealed the different expression patterns of gTNF-1 and gTNF-2. In addition, gTNF-2 was more sensitive to pathogens than gTNF-1; therefore, it may be an appropriate marker for pathogen invasion and the evaluation of the larval rearing environment.
authors.affiliation: Institute of Biotechnology, National Cheng Kung University, Tainan 701, Taiwan, ROC.
authors.forename: Freda Wai-San
authors.initials: FW
authors.lastname: Lam
authors.forename: Szu-Yin
authors.initials: SY
authors.lastname: Wu
authors.forename: Shih-Jie
authors.initials: SJ
authors.lastname: Lin
authors.forename: Chin-Chou
authors.initials: CC
authors.lastname: Lin
authors.forename: Yi-Ming
authors.initials: YM
authors.lastname: Chen
authors.forename: Han-Ching
authors.initials: HC
authors.lastname: Wang
authors.forename: Tzong-Yueh
authors.initials: TY
authors.lastname: Chen
authors.forename: Han-Tso
authors.initials: HT
authors.lastname: Lin
authors.forename: John Han-You
authors.initials: JH
authors.lastname: Lin
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005393:Fish Diseases / Q000276:immunology; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation / Q000276:immunology*; D007962:Leukocytes / Q000276:immunology*; D008969:Molecular Sequence Data; D010473:Perciformes / Q000145:classification / Q000235:genetics* / Q000276:immunology*; D010802:Phylogeny; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry / Q000235:genetics* / Q000276:immunology*; D014733:Vibrio / Q000276:immunology; D014735:Vibrio Infections / Q000276:immunology / Q000662:veterinary
chemical_list: D014409:Tumor Necrosis Factor-alpha