Upla Paula, Marjomäki Varpu, Nissinen Liisa, Nylund Camilla, Waris Matti, Hyypiä Timo, Heino Jyrki
Department of Biochemistry and Food Chemistry, University of Turku, FI-20520 Turku, Finland.
J Virol. 2008 Feb;82(3):1581-90. doi: 10.1128/JVI.01375-07. Epub 2007 Nov 21.
Calpains are calcium-dependent cysteine proteases that degrade cytoskeletal and cytoplasmic proteins. We have studied the role of calpains in the life cycle of human echovirus 1 (EV1). The calpain inhibitors, including calpeptin, calpain inhibitor 1, and calpain inhibitor 2 as well as calpain 1 and calpain 2 short interfering RNAs, completely blocked EV1 infection in the host cells. The effect of the inhibitors was not specific for EV1, because they also inhibited infection by other picornaviruses, namely, human parechovirus 1 and coxsackievirus B3. The importance of the calpains in EV1 infection also was supported by the fact that EV1 increased calpain activity 3 h postinfection. Confocal microscopy and immunoelectron microscopy showed that the EV1/caveolin-1-positive vesicles also contain calpain 1 and 2. Our results indicate that calpains are not required for virus entry but that they are important at a later stage of infection. Calpain inhibitors blocked the production of EV1 particles after microinjection of EV1 RNA into the cells, and they effectively inhibited the synthesis of viral RNA in the host cells. Thus, both calpain 1 and calpain 2 are essential for the replication of EV1 RNA.
钙蛋白酶是一种依赖钙的半胱氨酸蛋白酶，可降解细胞骨架和细胞质蛋白。我们研究了钙蛋白酶在人肠道病毒1型（EV1）生命周期中的作用。钙蛋白酶抑制剂，包括钙肽素、钙蛋白酶抑制剂1和钙蛋白酶抑制剂2以及钙蛋白酶1和钙蛋白酶2的短发夹RNA，完全阻断了宿主细胞中的EV1感染。这些抑制剂的作用并非EV1所特有，因为它们也抑制其他小核糖核酸病毒的感染，即人帕里病毒1型和柯萨奇病毒B3型。钙蛋白酶在EV1感染中的重要性还得到了以下事实的支持：感染后3小时，EV1会增加钙蛋白酶的活性。共聚焦显微镜和免疫电子显微镜显示，EV1/小窝蛋白-1阳性囊泡中也含有钙蛋白酶1和2。我们的结果表明，病毒进入不需要钙蛋白酶，但它们在感染后期很重要。将EV1 RNA显微注射到细胞中后，钙蛋白酶抑制剂阻断了EV1颗粒的产生，并且它们有效地抑制了宿主细胞中病毒RNA的合成。因此，钙蛋白酶1和钙蛋白酶2对于EV1 RNA的复制都是必不可少的。
