图注:A,绿色荧光显示在正常培养的细胞中,石斑鱼hepcidin 均匀分布于细胞核和细胞质中;B,在病毒感染细胞后,hepcidin多肽凝聚于细胞边缘(箭头所示),形成屏障,发挥抗病毒作用。
石斑鱼是热带和亚热带地区最重要的海水养殖鱼类品种之一,据联合国粮农组织(FAO)统计,全球石斑鱼年产量2007年已接近20万吨,经济价值巨大。然而,在大规模和高密度人工养殖条件下,石斑鱼的养殖成活率还不到40%,制约石斑鱼养殖业健康可持续发展的主要瓶颈在于各种传染性疾病的暴发和流行。由于大量使用抗生素易造成病原耐药性、环境污染和食品安全等问题,从海洋生物中发现天然抗菌和抗病毒物质,发展可替代抗生素的免疫防控技术,已成为研究发展新趋势。
中科院南海海洋研究所海洋生物资源可持续利用重点实验室(LMB)秦启伟研究员领导的团队,从病毒感染后的赤点石斑鱼脾组织差减文库中成功克隆出2个新的抗菌肽基因EC-hepcidin1和EC-hepcidin2。这两个基因分别由267和263个碱基组成,编码88和87个氨基酸,其中每个多肽分子含有4个半胱氨酸残基和2对二硫键,两个基因之间的相似度为79%。其基因组结构与哺乳动物的相似,包括3个外显子和2个内含子。石斑鱼Hepcidin基因广泛表达分布于鱼体各组织,在病毒、细菌和真菌病原感染后,基因表达显著升高,其中,在肝脏组织中的表达量最高。人工合成包含有2对二硫键的多肽具有很强的杀菌和抗病毒活性,是首次报道成功合成的具有很强生物功能活性的Hepcidin多肽分子,在防治海水鱼类重大病毒病方面具有很好的应用前景。
研究成果已于近日发表在农林水产类国际著名期刊Fish & Shellfish Immunology, 2011,February,30(2):559-568,并被选为该杂志的研究亮点(Research highlight)文章。论文发表后已在国际上引起了关注,韩国和罗马尼亚等国科学家致函通讯作者秦启伟,表示出对该研究的浓厚兴趣。
该研究受到国家杰出青年基金、973计划项目和中科院知识创新工程重要方向项目资助。(生物谷Bioon.com)
生物谷推荐原文出处:
Fish & Shellfish Immunology, 2011,February,30(2):559-568
Molecular cloning and characterization of two novel hepcidins from orange-spotted grouper, Epinephelus coioides.
Zhou JG, Wei JG, Xu D, Cui HC, Yan Y, Ou-Yang ZL, Huang XH, Huang YH, Qin QW.
Orange-spotted grouper, Epinephelus coioides is one of the most important economic species of marine-cultured fish in China and Southeast Asia countries. However, very little information of the innate immune mechanisms against microbial pathogens is available in grouper, Epinephelus sp. Hepcidin, as an antimicrobial peptide (AMP), is a very important component in the innate immune system and widespread in fish. In this study, two novel types of hepcidin gene (designated EC-hepcidin1 and EC-hepcidin2) were cloned from E. coioides. They consist of open reading frames (ORFs) of 267bp and 263bp encoding the putative peptides of 88 and 87 amino acids, respectively. The homologous identity of deduced amino acid sequences between EC-hepcidin1 and EC-hepcidin2 is up to 79%, and predicted mature regions of both them have four cysteines residues. Genomic DNAs of both EC-hepcidin1 and EC-hepcidin2 consist of three exons and two introns. RT-PCR results showed that EC-hepcidin1 transcript was most abundant in liver and less in stomach. However, the transcript of EC-hepcidin2 was only detected in liver. The expressions of both EC-hepcidins were up-regulated by microbial and viral challenges, and iron overload, respectively, and EC-hepcidin1 was more responsive. The growth of Gram-negative bacterium of Vibrio vulnificus and Gram-positive bacterium of Staphylococcus aureus was inhibited by synthetic EC-hepcidins, and EC-hepcidin1 displayed stronger antimicrobial activity. The replication of Singapore grouper iridovirus (SGIV) was inhibited in the EC-hepcidin1 and EC-hepcidin2 over-expressed stable transfected fish cell lines (GS/pcDNA-Hep1, GS/pcDNA-Hep2) indicative of the antiviral activity of EC-hepcidins. These data should offer important information on the antimicrobial and antiviral roles of EC-hepcidins, and will be help to the better understanding of molecular mechanisms of grouper innate immunity.