Department of Biomedical Sciences, Chang Gung University; Tao Yuan 33302, Taiwan, ROC.
Virol J. 2011 Oct 14;8:471. doi: 10.1186/1743-422X-8-471.
Japanese encephalitis virus (JEV) is a member of the mosquito-borne Flaviviridae family of viruses that causes human encephalitis. Upon infection of a new host, replication of viral RNA involves not only the viral RNA-dependent RNA polymerase (RdRp), but also host proteins. Host factors involved in JEV replication are not well characterized.
We identified Hdj2, a heat-shock protein 40 (Hsp40)/DnaJ homolog, from a mouse brain cDNA library interacting with JEV nonstructural protein 5 (NS5) encoding viral RdRp using yeast two-hybrid system. Specific interaction of Hdj2 with NS5 was confirmed by coimmunoprecipitation and colocalization in JEV-infected cells. Overexpression of Hdj2 in JEV-infected cells led to an increase of RNA synthesis, and the virus titer was elevated approximately 4.5- to 10-fold. Knocking down of Hdj2 by siRNA reduced the virus production significantly.
We conclude that Hdj2 directly associates with JEV NS5 and facilitates viral replication. This study is the first to demonstrate Hdj2 involved in JEV replication, providing insight into a potential therapeutic target and cell-based vaccine development of JEV infection.
日本脑炎病毒(JEV)是黄病毒科的一种蚊媒病毒,可引起人类脑炎。在感染新宿主时,病毒 RNA 的复制不仅涉及病毒 RNA 依赖性 RNA 聚合酶(RdRp),还涉及宿主蛋白。参与 JEV 复制的宿主因子尚未得到很好的描述。
我们使用酵母双杂交系统从小鼠脑 cDNA 文库中鉴定出与 JEV 非结构蛋白 5(NS5)编码病毒 RdRp 相互作用的热休克蛋白 40(Hsp40)/DnaJ 同源物 Hdj2。通过共免疫沉淀和共定位在 JEV 感染细胞中证实了 Hdj2 与 NS5 的特异性相互作用。在 JEV 感染的细胞中过表达 Hdj2 会导致 RNA 合成增加,病毒滴度升高约 4.5 至 10 倍。通过 siRNA 敲低 Hdj2 会显著降低病毒产量。
我们得出结论,Hdj2 直接与 JEV NS5 结合并促进病毒复制。本研究首次证明 Hdj2 参与 JEV 复制,为 JEV 感染的潜在治疗靶点和基于细胞的疫苗开发提供了新的见解。