山羊STING和TBK1基因组织特异性表达和分布的研究

Tissue-specific expression pattern and histological distribution of STING and TBK1 in goat

干扰素刺激基因(STING)和TANK结合激酶1(TBK1)是介导宿主Ⅰ型干扰素抗病毒应答的关键因子。为深入研究STING和TBK1在山羊天然免疫应答中的作用,采用RT-qPCR方法进行了STING和TBK1组织特异性表达检测,并进行了其组织分布的研究。由于物种差异较大,缺乏山羊STING的商品化抗体,本试验首先利用原核表达的STING蛋白制备了多克隆抗体。结果显示,利用原核表达的STING重组蛋白免疫新西兰大白兔后,经ELISA和Western-blot证明获得了特异性好的多克隆抗体。组织特异性表达检测结果显示,STING在肠系膜淋巴结、心脏和脾的相对表达水平较高,而TBK1在大脑、肝、心脏和肾的相对表达水平较高。免疫组织化学检测结果显示,在心肌细胞、肝细胞、淋巴细胞、细支气管上皮细胞、肾小管上皮细胞和大脑皮层神经元中均呈STING和TBK1阳性。研究结果为进一步探索STING和TBK1在山羊天然免疫应答中的作用提供了基础数据。

Stimulator of interferon genes(STING) and TANK-binding kinase 1(TBK1) are key factors in antiviral response which induce the host’s response of typeⅠ interferon. In order to further study the role of STING and TBK1 in goat’s innate immune response,tissue-specific expression of STING and TBK1 was investigated by real-time quantitative PCR and histological distribution was detected in various organs of goat.Due to the highly diverse between species,there is no suitable commercial antibody for STING detection in goat and the polyclonal antibody was first generated by using prokaryotic expression of STING protein.The results showed the obtained polyclonal antibody was of high specificity proved by ELISA and Western-blot detection after immunization to New Zealand white rabbits with the prokaryotic expression of STING recombinant protein.Tissue-specific expression pattern results showed the relative expression level of STING was high in the mesenteric lymph nodes,heart and spleen while TBK1 was high in brain,liver,heart,and kidney.Immunohistochemical detection showed that STING and TBK1 were positive in the cardiac muscle cells,hepatocytes,lymphoid cells,bronchiolar epithelial cells,renal tubular epithelial cells and the neurons in cerebral cortex. The results will provide the basic data for further studies of the role of STING and TBK1 in the innate immune response in goat.