日本乙型脑炎病毒对干扰素诱导基因ISG15表达的影响

为研究机体干扰素诱导基因对日本乙型脑炎(JEV)感染的作用,通过IFN-α和JEV刺激猪睾丸(ST)细胞,实时荧光定量PCR检测Mx1、Mx2、OAS1、OAS2、PKR和干扰素诱导基因-15(ISG15)表达情况,发现除PKR外,其他基因相对mRNA量都有不同程度上升,其中ISG15上升最为显著。ISG15在ST细胞中过表达,检测病毒基因组和滴度,结果表明:ISG15可显著抑制JEV增殖;shRNA靶向干扰ISG15,JEV病毒量上升,说明ISG15在体外对JEV的增殖有抑制作用。将JEV的C、M、E、NS1、NS2A、NS2B、NS3、NS4A、NS4B基因克隆入pcDNA3.1真核表达载体,转染ST细胞后E和NS3基因引起ISG15相对mRNA量显著上升,说明JEV的E和NS3基因参与诱导ISG15的表达。

Effects of Japanese Encephalitis Virus on Expression of Interferon-stimulated Gene 15

To investigate the interaction between Japanese encephalitis virus (JEV) and interferonstimulated genes,swine testicular (ST)cells were first treated with interferon α and infected with Japanese encephalitis virus.The results showed that mRNA levels of Mxl,Mx2,OAS1,OAS2 and interferon-stimulated gene 15 (ISG15) were significantly up-regulated except PKR,and the mRNA level of ISG15 was the highest among these.The results of viral genomes and titers showed that replication of JEV was significantly inhibited by overexpression of ISG15 compared with untreated ST cells,and viral yield was significantly increased by shRNA targeting the ISG15 gene.In the end,eukaryotic expression vector (pcDNA3.1) carrying capsid (C),membrane (M),envelope (E) or6 nonstructural genes (NS1,NS2A,NS2B,NS3,NS4A or NS4B) of JEV were constructed and transfected into ST cells to investigate effect on expression of ISG15.ISG15 mRNA level was significantly increased in ST cells tansfected with E and NS3 genes compared with other genes of JEV and the control.The results implied that a high level of ISG15 was correlated with the expression of JEV E and NS3 gene.In total,this study enables elucidation of the molecular mechanism of ISG15 against JEV,which will be useful for developing a novel treatment to combat JEV infection.