海南黑山羊IFITM3基因组织特异性表达和分布的研究

Tissue-specific expression pattern and histological distribution of IFITM3 in Hainan black goat

干扰素诱导跨膜蛋白3(interferon-induced transmembrane protein 3,IFITM3)在机体抗病毒和抗炎等方面发挥重要作用。为深入研究山羊IFITM3在病毒感染中的分子作用机制,对获得的海南黑山羊IFITM3基因进行了序列分析并采用RT-qPCR方法进行了其组织特异性表达的检测,利用原核表达的IFITM3蛋白制备了多克隆抗体,并进行IFITM3在黑山羊组织中分布的研究。结果显示,该基因表现出种属内保守性和种属间差异性,编码蛋白由147个氨基酸组成,存在2个潜在跨膜区,不含信号肽。成功构建pET-32a(+)-IFITM3表达载体并表达了35 ku重组蛋白,免疫家兔后,经ELISA和Western-blot检测,获得的多克隆抗体特异性好。组织特异性表达的检测结果表明,IFITM3在心脏表达水平最高,大脑和胸腺表达量最低。IFITM3在心肌细胞、肝细胞、脾网状内皮细胞、肺支气管上皮细胞、肾小管上皮细胞、大脑皮质、胸腺髓质、肠系膜淋巴结皮质淋巴细胞、胃底腺细胞和肠腺细胞中均有分布。研究结果为深入探讨IFITM3在山羊疾病中的分子作用机制提供了基础数据。

Interferon-induced transmembrane protein 3（IFITM3） plays an important role in the an- tivirus and anti-inflammation, ln order to study the molecular mechanism of Hainan black goat IFITM3 in the process of virus infection,sequences of IFITM3 were analyzed and relative expression levels were investigated by real-time quantitative PCR. Polyclonal antibodies were generatedby using prokaryotic expression IFITM3 protein,and the distribution was detected in various organs of Hainan black goat. The results showed IFITM3 gene was conserved within the genus and had differences between species. Encoding protein contained 147 amino acids,which had two potential transmembrane domains and without a signals peptide. The pET-32a （^-）-IFITM3 vector was successfully constructed and 35 ku recombinant protein was expressed, Antibodies were of high specificity detected by ELISA and Western-blot after immunization to rabbits. Tissue-specific expression pattern results showed the highest expression of IFITM3 was in the heart and the lowest was in the bruin and thymus. IFITM3 was distributed in myocardial cells,liver cells,reticuloendothelial cells of the spleen,pulmonary bronchial epithelial cells,renal tubular epithelial cells,cerebral cortex,thymus medulla,cortical ]ymphocytes of mesenteric lymph nodes,fundic gland cells and intestinal gland cells. These results provide the basic data for further studies of the molecular mechanisms of IFITM3 in goat diseases.