Zhirnov Oleg P, Klenk Hans-Dieter
D.I. Ivanovsky Institute of Virology, Moscow 123098, Russia.
Apoptosis. 2007 Aug;12(8):1419-32. doi: 10.1007/s10495-007-0071-y.
PI3k-Akt and p53 pathways are known to play anti- and pro-apoptotic roles in cell death, respectively. Whether these pathways are recruited in influenza virus infection in highly productive monkey (CV-1) and canine (MDCK) kidney cells was studied here. Phosphorylation of Akt (Akt-pho) was found to occur only early after infection (5-9 h.p.i). Nuclear accumulation and phosphorylation of p53 (p53-pho), and expression of its natural target p21/waf showed low constitutive levels at this period, whereas all three parameters were markedly elevated at the late apoptotic stage (17-20 h.p.i.). Up-regulation of Akt-pho and p53-pho was not induced by UV-inactivated virus suggesting that it required virus replication. Also, mRNAs of p53 and its natural antagonist mdm2 were not increased throughout infection indicating that p53-pho was up-regulated by posttranslational mechanisms. However, p53 activation did not seem to play a leading role in influenza-induced cell death: (i) infection of CV1 and MDCK cells with recombinant NS1-deficient virus provoked accelerated apoptotic death characterized by the lack of p53 activation; (ii) mixed apoptosis-necrosis death developed in influenza-infected human bronchial H1299 cells carrying a tetracycline-regulated p53 gene did not depend on p53 gene activation by tetracycline. Virus-induced apoptosis and signaling of Akt and p53 developed in IFN-deficient VERO cells with similar kinetics as in IFN-competent CV1-infected cells indicating that these processes were endocrine IFN-independent. Apoptosis in influenza-infected CV-1 and MDCK cells was Akt-dependent and was accelerated by Ly294002, a specific inhibitor of PI3k-Akt signaling, and down-regulated by the viral protein NS1, an inducer of host Akt. The obtained data suggest that influenza virus (i) initiates anti-apoptotic PI3k-Akt signaling at early and middle phases of infection to protect cells from fast apoptotic death and (ii) provokes both p53-dependent and alternative p53-independent apoptotic and/or necrotic (in some host systems) cell death at the late stage of infection.
已知PI3k-Akt和p53信号通路分别在细胞死亡中发挥抗凋亡和促凋亡作用。本研究探讨了在高效生产性的猴（CV-1）和犬（MDCK）肾细胞中，这些信号通路是否参与流感病毒感染过程。研究发现，Akt磷酸化（Akt-pho）仅在感染后早期（感染后5-9小时）出现。在此期间，p53的核积累和磷酸化（p53-pho）及其天然靶点p21/waf的表达处于低组成水平，而在凋亡晚期（感染后17-20小时），这三个参数均显著升高。紫外线灭活病毒未诱导Akt-pho和p53-pho上调，表明这需要病毒复制。此外，在整个感染过程中，p53及其天然拮抗剂mdm2的mRNA均未增加，表明p53-pho是通过翻译后机制上调的。然而，p53激活似乎在流感诱导的细胞死亡中不发挥主导作用：（i）用重组NS1缺陷病毒感染CV1和MDCK细胞引发加速凋亡死亡，其特征是缺乏p53激活；（ii）携带四环素调控p53基因的流感感染人支气管H1299细胞中出现的混合凋亡-坏死性死亡不依赖于四环素对p53基因的激活。病毒诱导的凋亡以及Akt和p53信号在IFN缺陷的VERO细胞中的发展动力学与IFN competent的CV1感染细胞相似，表明这些过程不依赖于内分泌IFN。流感感染的CV-1和MDCK细胞中的凋亡是Akt依赖性的，PI3k-Akt信号的特异性抑制剂Ly294002可加速凋亡，而病毒蛋白NS1（宿主Akt的诱导剂）可下调凋亡。所得数据表明，流感病毒（i）在感染的早期和中期启动抗凋亡的PI3k-Akt信号，以保护细胞免于快速凋亡死亡；（ii）在感染后期引发p53依赖性和替代性p53非依赖性凋亡和/或坏死性（在某些宿主系统中）细胞死亡。
