半滑舌鳎抗哈维氏弧菌病关键基因的鉴定及功能分析

The half smooth tongue sole (Cynoglossus semilaevis) is a marine fish which is economically important in China. However, high frequency of diseases restricts the sustainable and healthy development of the aquaculture of C. semilaevis. Based on our previous study on the identification of 11 resistance-related QTL in the highly dese genetic linkage map by using C. semilaevis family resistant to Vibrio harveyi, major histocompatibility complex class II transactivator (CsCIITA), which was located in one QTL, would be a good immune gene for this study. The study will be processed in the following three aspects: 1) the identification of CIITA alleles that are related to the resistance against V. harveyi; 2) the detection of CIITA gene expression and localization in different tissues of C. semilaevis and after the infection of V. harveyi, and the examination of the characters of the recombinant protein of CIITA against V. harveyi; and 3) the analysis of the relationship between CIITA gene and its downstream MHC gene in the cells stimulated with antigens. In this study, the immunologic function of CsCIITA gene and its relationship with MHC genes will be explored during C. semilaevis resists V. harveyi, thereby being helpful to understand molecular mechanisms beyond disease resistance in C. semilaevis and providing theoretical guidance for the culture of disease resistant breed of C. semilaevis.

半滑舌鳎是我国重要的海水养殖鱼类，经济价值极高。然而，频发的病害严重制约了半滑舌鳎养殖业的可持续健康发展。在前期工作中，申请人所在课题组利用半滑舌鳎抗哈维氏弧菌病家系，开展QTL精细定位研究，筛选了11个与抗病性状相关区间，鉴定到一个与抗哈维氏弧菌病相关的基因：MHCII类的反式激活因子CIITA。本课题将以CIITA基因为研究对象，计划从以下3个方面开展研究，探索半滑舌鳎分子免疫机制：1）鉴定与抗哈维氏弧菌病相关的CIITA等位基因；2）阐明CIITA基因的时空表达模式，以及CIITA基因对哈维氏弧菌刺激的免疫响应，检测CIITA重组蛋白的病原结合特性；3）揭示病原刺激下，CIITA基因与MHC基因的相关性。本研究将解析在半滑舌鳎抗哈维氏弧菌病家系抗病力形成过程中，CIITA基因的基因功能及其与MHC基因的关系，有助于理解半滑舌鳎抗病分子机理，并为半滑舌鳎抗病品种的培育提供理论基础。

半滑舌鳎是我国重要的海水养殖鱼类，具有较高的经济价值。然而，频发的病害严重制约了半滑舌鳎养殖业的可持续健康发展。本研究基于前期课题组对于半滑舌鳎抗哈维氏弧菌病家系的QTL定位研究，以CIITA基因为研究对象，进行克隆、时空表达模式研究，以及在病原刺激下，CIITA基因与MHC II基因表达的相关性。结果表明，CIITA基因在半滑舌鳎的各个组织中广泛分布。在易感家系的皮肤组织中，CIITA、MHC IIA和MHC IIB基因表达量均较抗病家系中的表达量有显著上调；而在抗病家系的肝脏组织中，CIITA和MHC IIB基因表达量较易感家系中的表达量有显著上调。三个基因表达趋势在感染的组织中表现一致，均为感染48至72小时后显著上调。原位杂交结果显示，三个基因在鳃、皮肤和肠的表皮黏液细胞中共表达，这些结果表明CIITA参与半滑舌鳎抗哈维氏弧菌的免疫反应，并参与MHC II的调控表达。此外，本研究也是第一次基于半滑舌鳎全基因组数据，鉴定、分析NLR基因家族的分子特征和表达模式，结果表明，半滑舌鳎NLR家族参与半滑舌鳎抗哈维氏弧菌的免疫反应，但不同的基因在半滑舌鳎抗病力形成过程中可能起到了不同的作用。蛋白互作网络分析揭示了半滑舌鳎NLR基因可能参与下游NF-kB信号通路的调控。该研究有助于理解半滑舌鳎抗病分子机理，并为半滑舌鳎抗病品种的培育提供理论基础。