Identification and expression analysis on bactericidal permeability-increasing protein/lipopolysaccharide-binding protein of blunt snout bream, Megalobrama amblycephala
Identification and expression analysis on bactericidal permeability-increasing protein/lipopolysaccharide-binding protein of blunt snout bream, Megalobrama amblycephala
复制标题
团头鲂杀菌增透蛋白/脂多糖结合蛋白的鉴定及表达分析
DOI：
10.1016/j.fsi.2015.05.013
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发表时间：
2015-08-01
影响因子：
4.7
通讯作者：
Yan, Jinpeng
中科院分区：
文献类型：
--
作者：
Tang, Leilei;Liang, Yinhua;Yan, Jinpeng
杀菌通透性增加蛋白(BPI)和脂多糖结合蛋白(LBP)属于脂转移蛋白/脂多糖结合蛋白家族，在革兰氏阴性菌的天然免疫应答中发挥重要作用。本研究利用RACE技术从团头团头鱼(Megalobrama Amblycephala)中分离到一种新的BPI/LBP(maBPI/LBP)。MaBPI/LBP基因的开放阅读框(ORF)编码一个由474个氨基酸组成的多肽，其中含有一个18-aa的疏水信号肽。在结构上，maBPI/LBP与无脊椎动物和硬骨鱼的BPI/LBPS、LBPS和哺乳动物的BPI非常相似，它含有一个N-端的BPI/LBP/CETP结构域BPI1和一个C-端的BPI/LBP/CETP结构域BPI2和富含脯氨酸的结构域。推导的氨基酸序列同源性分析表明，maBPI/LBP与草鱼和鲤鱼的同源性分别为96.61%和90.07%。重组蛋白maBPI/LBP显示了对革兰氏阴性菌的有效杀灭。MaBPI/LBP基因在广泛的正常受试组织中表达，其中肾脏的表达水平最高。结果表明，脾组织中maBPI/LBP的mRNA表达在内毒素刺激后2~8h显著上调，并于2 h迅速达到峰值(7.40倍，P<；0.05)，证实了maBPI/LBP对内毒素刺激的绝对敏感性。此外，在内毒素刺激后24 h，maBPI/LBP mRNA的表达水平再次达到最高。这些结果表明，maBPI/LBP是一种结构性和可诱导性的急性时相蛋白，参与了Amblycephala对病原菌感染的宿主免疫防御。本研究将进一步了解硬骨鱼BPI/LBP的功能和天然免疫的分子机制。(C)2015爱思唯尔有限公司。保留所有权利。
Bactericidal permeability-increasing protein (BPI) and lipopolysaccharide-binding protein (LBP) belong to the lipid transfer protein/lipopolysaccharide-binding protein family and play a critical role in the innate immune response to Gram-negative bacteria. In the present study, a novel BPI/LBP from blunt snout bream, Megalobrama amblycephala (maBPI/LBP) was isolated by RACE techniques. The open reading frame (ORF) of maBPI/LBP gene encoded a polypeptide of 474 amino acids with a putative 18-aa hydrophobic signal peptide. Structurally, the maBPI/LBP showed highly similar to those of BPI/LBPs from invertebrate and teleost, LBPs and BPIs from mammal, which contained an N-terminal BPI/LBP/CETP domain BPI1 with a LPS-binding domain, a C-terminal BPI/LBP/CETP domain BPI2, and proline-rich domain. The homologous identities of deduced amino acid sequences displayed that the maBPI/LBP possessed significant similarity (96.61% and 90.07%) with those of grass carp and common carp, respectively. The recombinant protein of maBPI/LBP showed effectively kill Gram-negative bacteria. The maBPI/LBP gene was expressed in a wide range of normal tested tissues, with the highest expression levels in the kidney. The experiments revealed that the mRNA expression of maBPI/LBP in spleen considerably up-regulated from 2 h to 8 h post LPS stimulation, and peaked rapidly at 2 h (7.40-fold, P < 0.05), which confirmed that maBPI/LBP was the absolute sensitive to LPS stimulation. Furthermore, the level of maBPI/LBP mRNA expression reached the maximum for a second time at 24 h after LPS stimulation. These results suggested that maBPI/LBP was a constitutive and inducible acute-phase protein contributing to the host immune defense against pathogenic bacterial infection in M. amblycephala. This study will further our understanding of the function of BPI/LBP and the molecular mechanism of innate immunity in teleost. (C) 2015 Elsevier Ltd. All rights reserved.