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| 菲律宾蛤仔脂多糖诱导的肿瘤坏死因子α VpLITAF基因克隆及其对微生物侵染的应答研究 |
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张林宝,蔡文贵,宁璇璇,张喆,陈海刚,马胜伟,贾晓平
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中国水产科学研究院南海水产研究所,中国水产科学研究院南海水产研究所,国家海洋局烟台海洋环境监测中心站,中国水产科学研究院南海水产研究所,中国水产科学研究院南海水产研究所,中国水产科学研究院南海水产研究所,中国水产科学研究院南海水产研究所
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| 摘要: |
| 脂多糖诱导的肿瘤坏死因子α(LITAF)是一种重要的转录因子, 在调节LPS 诱导炎症因子TNF-α 的表达过程中发挥重要作用。采用表达序列标签(expressed sequence tags, ESTs)结合cDNA 末端快速克隆技术(rapid amplification of cDNA Ends, RACE), 获得了菲律宾蛤仔LITAF 基因 cDNA 全长(VpLITAF), 并利用实时荧光定量PCR 技术研究VpLITAF 的组织分布以及鳗弧菌、藤黄微球菌刺激下的时序表达特征。VpLITAF 基因序列包含393bp 的开放阅读框, 编码130 个氨基酸, 预测分子量为14.39kD, 理论等电点7.47。序列分析表明, VpLITAF 具有高度保守的LITAF 结构域, 与其它贝类LITAF 具有高度同源性, 并在系统进化树中与海洋软体动物LITAF 聚为一簇。VpLITAF 基因主要表达于蛤仔肝胰腺, 肌肉中的表达量则相对较低。鳗弧菌和藤黄微球菌刺激在某些时段可显著诱导VpLITAF 基因的表达水平, 最高可达对照组表达量的3.5 倍, 且鳗弧菌对VpLITAF 基因表达的诱导作用明显强于藤黄微球菌。以上结果表明, VpLITAF 在菲律宾蛤仔的固有免疫反应中发挥着重要作用。 |
| 关键词: 菲律宾蛤仔 脂多糖诱导的肿瘤坏死因子α 微生物侵染 基因表达 |
| DOI：10.11693/hyhz20130400020 |
| 分类号: |
| 基金项目:中央级公益性科研院所基本科研业务费专项资金项目, 2012TS28 号 (中国水产科学研究院南海水产研究所), 2012A0203 号,2013A0301 号(中国水产科学研究院); 海南省社会发展科技专项资金项目, XH201312 号 |
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| MOLECULAR CLONING OF LIPOPOLYSACCHARIDEINDUCED TUMOR NECROSIS FACTOR ALPHA FACTOR FROM VENERUPIS PHILIPPINARUM (VpLITAF), AND ITS TRANSCRIPTIONAL RESPONSE FOLLOWING BACTERIAL CHALLENGE |
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zhanglinbao,Caiwengui,Ningxuanxuan,zhangzhe,chenhaigang,mashengwei and jiaxiaoping
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South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences,South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences,Yantai Oceanic Environmental Monitoring Central Station,State Oceanic Administration,South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences,South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences,South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences,South China Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences
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| Abstract: |
| Lipopolysaccharide (LPS)-induced tumor necrosis factor alpha factor (LITAF) is a novel transcription factor responsible for LPS-induced transcription of tumor necrosis factor-alpha, and it plays an important role in innate immunity system. We identified a full-length cDNA encoding a LITAF from Venerupis philippinarum (designated as VpLITAF) with expressed sequence tag (EST) and rapid amplification of cDNA ends (RACE) approaches, and investigated its temporal expression patterns post Vibrio anguillarum and Micrococcus luteus challenges. The full-length VpLITAF cDNA contains an open reading frame (ORF) of 393 bp encoding a polypeptide of 130 amino acids with a predicted molecular weight of 14.39 kDa, and theoretical isoelectric point of 7.49. VpLITAF possessed a conserved LITAF domain at C-terminal and shared high sequence similarity with the LITAFs from other marine mollusks, indicating that VpLITAF should be a member of the LITAF family. VpLITAF mRNA was found expressed most abundantly in hepatopancreas but least in adductor muscles. Bacterial challenging could significantly up-regulate the mRNA expression of VpLITAF at different time intervals, in which the highest expression level was 3.5-fold of the control. Moreover, V. anguillarum displayed stronger induction of VpLITAF expression than that of M. luteus. All these results suggested that VpLITAF was a potent factor in the regulation of genes that are involved in innate defence mechanism. |
| Key words: Venerupis philippinarum Lipopolysaccharide induced tumor necrosis factor alpha factor (LITAF) bacterial challenge gene expression |