Department of Biochemistry, Chang Gung University, Kweishan, Taoyuan, Taiwan.
Cell Microbiol. 2010 Jun;12(6):796-813. doi: 10.1111/j.1462-5822.2010.01434.x. Epub 2010 Jan 11.
Picornavirus infection alters the endoplasmic reticulum (ER) membrane but it is unclear whether this induces ER stress. Infection of rhabdomyosarcoma cells with enterovirus 71 (EV71), a picornavirus, caused overexpression of the ER-resident chaperone proteins, BiP and calreticulin, and phosphorylation of eIF2alpha, but infection with UV-inactivated virus did not, indicating that ER stress was induced by viral replication and not by viral attachment or entry. Silencing (si)RNA knockdown demonstrated that phosphorylation of eIF2alpha was dependent on PKR: eIF2alpha phosphorylation was reduced by siPKR but not by siPERK. We provided evidence showing that PERK is upstream of PKR and is thus able to negatively regulate the PKR-eIF2alpha pathway. Pulse-chase experiments revealed that EV71 infection inhibited translation and activation of ATF6. Expression of BiP at the protein level was activated by a virus-dependent, ATF6-independent mechanism. EV71 upregulated XBP1 mRNA level, but neither IRE1-mediated XBP1 splicing nor its active spliced protein was detected, and its downstream gene, EDEM, was not activated. Epigenetic BiP overexpression alleviated EV71-induced ER stress and reduced viral protein expression and replication. Our results suggest that EV71 infection induces ER stress but modifies the outcome to assist viral replication.
微小核糖核酸病毒感染会改变内质网（ER）膜，但尚不清楚这是否会引发内质网应激。肠道病毒 71（EV71）是微小核糖核酸病毒的一种，感染横纹肌肉瘤细胞会导致内质网驻留伴侣蛋白 BiP 和钙网蛋白的过度表达，以及 eIF2alpha 的磷酸化，但感染紫外线失活的病毒则不会，表明 ER 应激是由病毒复制引起的，而不是由病毒附着或进入引起的。siRNA 敲低表明 eIF2alpha 的磷酸化依赖于 PKR：siPKR 可降低 eIF2alpha 的磷酸化，但 siPERK 则不能。我们提供的证据表明 PERK 位于 PKR 的上游，因此能够负调控 PKR-eIF2alpha 通路。脉冲追踪实验表明，EV71 感染抑制了翻译和 ATF6 的激活。依赖于病毒而非 ATF6 的机制激活了 BiP 的蛋白质水平表达。EV71 上调了 XBP1 mRNA 水平，但未检测到 IRE1 介导的 XBP1 剪接及其活性剪接蛋白，其下游基因 EDEM 也未被激活。表观遗传的 BiP 过表达缓解了 EV71 诱导的 ER 应激，降低了病毒蛋白的表达和复制。我们的研究结果表明，EV71 感染会引发内质网应激，但会改变结局以促进病毒复制。
