College of Fisheries, Henan Normal University, Xinxiang, 453007, PR China.
College of Life Science, Henan Normal University, Xinxiang, 453007, PR China.
Fish Shellfish Immunol. 2019 Apr;87:829-838. doi: 10.1016/j.fsi.2019.02.034. Epub 2019 Feb 18.
Myeloid differentiation factor 88 (MyD88) and tumor necrosis factor receptor-associated factor 6 (TRAF6) are two critical signal transducers in toll-like receptor (TLR) pathway. In the present study, we identified and characterized the homologues of MyD88 and TRAF6 in Qihe crucian carp Carassius auratus, termed as CaMyD88 and CaTRAF6, respectively, and examined their roles during pathogenic infection. Full-length cDNA of CaMyD88 was 2463 bp, including a 191 bp 5'-untranslated region (UTR), a 1417 bp 3'-UTR, and an 855 bp open reading frame (ORF) encoding for a putative protein with 284 amino acids. Full-length cDNA of CaTRAF6 was identified to be 2555 bp, consisting of a 52 bp 5'-UTR, an 871 bp 3'-UTR, and a 1632 bp ORF encoding a protein of 543 amino acids. Deduced amino acid sequences of CaMyD88 and CaTRAF6 contained the typical domains (CaMyD88: death domain and TIR domain; CaTRAF6: one RING-type zinc finger domain, two TRAF-type zinc finger domains, one coiled-coil region, and one conserved C-terminal meprin and TRAF homology domain) as in other fish. Quantitative Real-Time PCR (qRT-PCR) analysis revealed that both CaMyD88 and CaTRAF6 were ubiquitously expressed throughout the development stages and appeared to be developmentally regulated. In addition, CaMyD88 and CaTRAF6 had a broadly distribution of expression in all examined eleven tissues of healthy fish, although the transcript levels varied among the different tissues. Moreover, it was found that mRNA expressions of CaMyD88 and CaTRAF6 were generally up-regulated after stimulation by polyI:C, flagellin, and Aeromonas hydrophila in spite of the down-regulation appeared at some time points or tissues. These results indicated that CaMyD88 and CaTRAF6 play the critical roles in the immune defense of Qihe crucian carp against pathogenic invasion. The present findings will provide the valuable information for understanding the innate immune responses of Qihe crucian carp and contribute to develop the preventive way against pathogens.
髓样分化因子 88（MyD88）和肿瘤坏死因子受体相关因子 6（TRAF6）是 Toll 样受体（TLR）途径中的两个关键信号转导因子。在本研究中，我们鉴定并表征了鲤鱼（Carassius auratus）齐河亚种的 MyD88 和 TRAF6 同源物，分别称为 CaMyD88 和 CaTRAF6，并研究了它们在致病感染过程中的作用。CaMyD88 的全长 cDNA 为 2463bp，包括一个 191bp 的 5'-非翻译区（UTR）、一个 1417bp 的 3'-UTR 和一个 855bp 的开放阅读框（ORF），编码一个具有 284 个氨基酸的假定蛋白。CaTRAF6 的全长 cDNA 被鉴定为 2555bp，由一个 52bp 的 5'-UTR、一个 871bp 的 3'-UTR 和一个 1632bp 的 ORF 组成，编码一个具有 543 个氨基酸的蛋白。CaMyD88 和 CaTRAF6 的推导氨基酸序列包含了典型的结构域（CaMyD88：死亡结构域和 TIR 结构域；CaTRAF6：一个 RING 型锌指结构域、两个 TRAF 型锌指结构域、一个卷曲螺旋区和一个保守的 C 末端 Meprin 和 TRAF 同源结构域），与其他鱼类中的结构域相同。实时定量 PCR（qRT-PCR）分析表明，CaMyD88 和 CaTRAF6 在整个发育阶段都广泛表达，并且似乎受到发育调控。此外，在健康鱼的所有 11 种检测组织中，CaMyD88 和 CaTRAF6 都具有广泛的表达分布，尽管不同组织的转录水平存在差异。此外，尽管在某些时间点或组织中出现下调，但发现在 polyI:C、鞭毛蛋白和嗜水气单胞菌刺激后，CaMyD88 和 CaTRAF6 的 mRNA 表达通常上调。这些结果表明，CaMyD88 和 CaTRAF6 在鲤鱼对病原体入侵的免疫防御中发挥关键作用。本研究结果将为了解鲤鱼先天免疫反应提供有价值的信息，并有助于开发针对病原体的预防方法。
