Xiao Jia, Zhou Zhi-Cheng, Chen Chao, Huo Wei-Li, Yin Zhi-Xin, Weng Shao-Ping, Chan Siu-Ming, Yu Xiao-Qiang, He Jian-Guo
State Key Laboratory for Biocontrol, School of Life Sciences, Sun Yat-sen University, 135 Xingang Road West, Guangzhou 510275, PR China.
Mol Immunol. 2007 Jul;44(14):3615-22. doi: 10.1016/j.molimm.2007.03.016. Epub 2007 May 4.
In this report, we cloned a cDNA and the gene of mandarin fish (Siniperca chuatsi) tumor necrosis factor-alpha (TNF-alpha). The length of TNF-alpha cDNA was 1461 bp, contained an open reading frame (ORF) of 762 bp, which encoded 253 amino acids. The positions of cysteine residues, transmembrane sequence, and protease cleavage site were similar with other reported fish TNF-alpha and mammalian TNF-alpha. Mandarin fish TNF-alpha gene has a length of 1940 bp, and consists of four exons and three introns. There are three NF-kappaB/rel regulatory/binding sites in gene's 5' flanking region. Expression of mandarin fish TNF-alpha mRNA was constitutive in all tissues detected. After challenge of infectious spleen and kidney necrosis virus (ISKNV), which results a high mortality rate in mandarin fish and has caused huge economic losses in China, the expression of TNF-alpha mRNA in head kidney and kidney did not show significant changes from 0 day to 15 days post-infection. While in blood and spleen, the gene's mRNA showed up-regulations in 4 days post-infection and down-regulations in 15 days post-infection, which indicated the mRNA expression of TNF-alpha may related to the infection of ISKNV and the survival of mandarin fish. Mature form of recombinant mandarin fish TNF-alpha protein (400-4000 ng/ml) showed an evident apoptosis induction ability of human HeLa cells, in a dose-dependent manner, although to a lesser extent compared to the recombinant human TNF-alpha protein (200 ng/ml).
在本报告中，我们克隆了鳜鱼（Siniperca chuatsi）肿瘤坏死因子-α（TNF-α）的cDNA和基因。TNF-α cDNA长度为1461 bp，包含一个762 bp的开放阅读框（ORF），编码253个氨基酸。半胱氨酸残基、跨膜序列和蛋白酶切割位点的位置与其他已报道的鱼类TNF-α和哺乳动物TNF-α相似。鳜鱼TNF-α基因长度为1940 bp，由四个外显子和三个内含子组成。在基因的5'侧翼区域有三个NF-κB/rel调节/结合位点。鳜鱼TNF-α mRNA在所有检测组织中均组成性表达。在用传染性脾肾坏死病毒（ISKNV）攻击后，该病毒在中国导致鳜鱼高死亡率并造成巨大经济损失，感染后0天至15天，头肾和肾脏中TNF-α mRNA的表达没有显著变化。而在血液和脾脏中，该基因的mRNA在感染后4天出现上调，在感染后15天出现下调，这表明TNF-α的mRNA表达可能与ISKNV感染及鳜鱼的存活有关。重组鳜鱼TNF-α蛋白的成熟形式（400 - 4000 ng/ml）对人HeLa细胞显示出明显的凋亡诱导能力，呈剂量依赖性，尽管与重组人TNF-α蛋白（200 ng/ml）相比程度较小。
