Key Laboratory of Agricultural Animal Genetics, Breeding and Reproduction, Ministry of Education & College of Animal Science and Technology, Huazhong Agricultural University, Wuhan 430070, China.
Virol J. 2013 Aug 12;10:258. doi: 10.1186/1743-422X-10-258.
Japanese encephalitis virus (JEV) is a mosquito-borne flavivirus that causes acute viral encephalitis in humans. Pigs are important amplifiers of JEV. The entry mechanism of JEV into porcine cells remains largely unknown. In this study, we present a study of the internalization mechanism of JEV in porcine kidney epithelial PK15 cells.
We demonstrated that the disruption of the lipid raft by cholesterol depletion with methyl-β-cyclodextrin (MβCD) reduced JEV infection. We also found that the knockdown of clathrin by small interfering RNA (siRNA) significantly reduced JEV-infected cells and JEV E-glycoprotein levels, suggesting that JEV utilizes clathrin-dependent endocytosis. In contrast, the knockdown of caveolin-1, a principal component of caveolae, had only a small (although statistically significant) effect on JEV infection, however, JEV entry was not affected by genistein. These results suggested that JEV entry was independent of caveolae.
Taken together, our results demonstrate that JEV enters porcine kidney epithelial PK15 cells through cholesterol- and clathrin-mediated endocytosis.
日本脑炎病毒（JEV）是一种通过蚊子传播的黄病毒，可导致人类急性病毒性脑炎。猪是 JEV 的重要扩增宿主。JEV 进入猪细胞的进入机制在很大程度上尚不清楚。在这项研究中，我们研究了 JEV 在猪肾上皮 PK15 细胞中的内化机制。
我们证明了通过用甲基-β-环糊精（MβCD）耗尽胆固醇破坏脂筏会降低 JEV 感染。我们还发现，小干扰 RNA（siRNA）敲低网格蛋白可显著减少 JEV 感染的细胞和 JEV E 糖蛋白水平，表明 JEV 利用网格蛋白依赖性内吞作用。相比之下，窖蛋白-1（ caveolin-1）的主要成分小窝蛋白的敲低对 JEV 感染的影响很小（尽管具有统计学意义），但是，JEV 进入不受染料木黄酮的影响。这些结果表明 JEV 的进入不依赖于小窝。
综上所述，我们的结果表明 JEV 通过胆固醇和网格蛋白介导的内吞作用进入猪肾上皮 PK15 细胞。
