State Key Laboratory of Freshwater Ecology and Biotechnology, Institute of Hydrobiology, Chinese Academy of Sciences, Wuhan 430072, China.
Dev Comp Immunol. 2012 Oct;38(2):383-8. doi: 10.1016/j.dci.2012.06.002. Epub 2012 Jun 18.
The mammalian response to lipopolysaccharide (LPS) is mainly mediated by Toll-Like Receptor 4 (TLR4). Fish and mammalian TLR4 vary; fish TLR4 ligands are unknown. Isolation of fish tlr4 genes is difficult due to their complex genomic structure. Three bacterial artificial chromosome (BAC) clones containing grass carp tlr4 were obtained. Four tlr4 genes, with a varied genomic structure and different protein domains were subsequently isolated by constructing a subcloned library and rapid amplification of cDNA ends (RACE). The four tlr4 genes were expressed during development from 12h post-fertilization, in all healthy adult fish tissues tested, and significantly increased in grass carp reovirus (GCRV)-infected liver and muscle, suggesting the tlr4 genes play a role in GCRV infection. This study effectively separated each gene in the tlr4 gene cluster, implies that grass carp TLR4 proteins have different ligand recognition specificities to mammalian TLRs, and provides information on the functional evolution of TLRs.
哺乳动物对脂多糖（LPS）的反应主要由 Toll 样受体 4（TLR4）介导。鱼类和哺乳动物的 TLR4 不同；鱼类 TLR4 配体尚不清楚。由于其复杂的基因组结构，鱼类 tlr4 基因的分离很困难。获得了含有草鱼 TLR4 的三个细菌人工染色体（BAC）克隆。通过构建亚克隆文库和快速扩增 cDNA 末端（RACE），随后分离出四个具有不同基因组结构和不同蛋白结构域的 TLR4 基因。在受精后 12 小时的发育过程中，在所有测试的健康成年鱼类组织中均表达了这四个 TLR4 基因，并在草鱼虹彩病毒（GCRV）感染的肝脏和肌肉中显著增加，表明 TLR4 基因在 GCRV 感染中发挥作用。本研究有效地分离了 TLR4 基因簇中的每个基因，这表明草鱼 TLR4 蛋白对哺乳动物 TLR 具有不同的配体识别特异性，并提供了 TLR 功能进化的信息。
