Wei Jingguang, Gao Pin, Zhang Ping, Guo Minglan, Xu Meng, Wei Shina, Yan Yang, Qin Qiwei
Key Laboratory of Tropical Marine Bio-resources and Ecology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China; Guangdong Provincial Key Laboratory of Applied Marine Biology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou 510301, PR China.
State Key Laboratory Breeding Base for Sustainable Exploitation of Tropical Biotic Resources, College of Marine Science, Hainan University, Haikou 570228, PR China.
Fish Shellfish Immunol. 2015 Apr;43(2):396-404. doi: 10.1016/j.fsi.2015.01.006. Epub 2015 Jan 19.
Apolipoproteins, synthesized mainly in liver and intestine and bounded to lipids, play important roles in lipid transport and uptake through the circulation system. In this study, an apolipoprotein A-I gene homologue was cloned from orange-spotted grouper Epinephelus coioides (designed as Ec-ApoA-I) by rapid amplification of cDNA ends (RACE) PCR. The full-length cDNA of Ec-ApoA-I was comprised of 1278 bp with a 792 bp open reading frame (ORF) that encodes a putative protein of 264 amino acids. Quantitative real-time PCR (qPCR) analysis revealed that Ec-ApoA-I was abundant in liver and intestine, and the expression in liver was significantly (P < 0.01) up-regulated after the stimulation of LPS, Poly(I:C), Vibrio alginolyticus, and Singapore grouper iridovirus (SGIV). Recombinant Ec-ApoA-I (rEc-ApoA-I) was produced in Escherichia coli BL21 (DE3) expression system exhibited bacteriolyticactivity against Microcococcus lysodeikticus and Aeromonas hydrophila. Intracellular localization revealed that Ec-ApoA-I distributed in both cytoplasm and nucleus, and predominantly in the cytoplasm. Overexpression of Ec-ApoA-I in grouper Brain (GB) cells could inhibit the replication of SGIV. These results together indicated that Ec-ApoA-I perhaps is involved in the responses to bacterial and viral challenge.
载脂蛋白主要在肝脏和肠道合成，并与脂质结合，在通过循环系统进行的脂质运输和摄取过程中发挥重要作用。在本研究中，通过cDNA末端快速扩增（RACE）PCR从点带石斑鱼（Epinephelus coioides）中克隆了一个载脂蛋白A-I基因同源物（命名为Ec-ApoA-I）。Ec-ApoA-I的全长cDNA由1278 bp组成，其中有一个792 bp的开放阅读框（ORF），编码一个推定的含264个氨基酸的蛋白质。实时定量PCR（qPCR）分析显示，Ec-ApoA-I在肝脏和肠道中含量丰富，在受到脂多糖、聚肌胞苷酸、溶藻弧菌和新加坡石斑鱼虹彩病毒（SGIV）刺激后，肝脏中的表达显著上调（P < 0.01）。在大肠杆菌BL21（DE3）表达系统中产生的重组Ec-ApoA-I（rEc-ApoA-I）对溶壁微球菌和嗜水气单胞菌具有溶菌活性。细胞内定位显示，Ec-ApoA-I分布于细胞质和细胞核中，且主要分布在细胞质中。在石斑鱼脑（GB）细胞中过表达Ec-ApoA-I可抑制SGIV的复制。这些结果共同表明，Ec-ApoA-I可能参与了对细菌和病毒攻击的应答。
