Sendai Virus C Protein Plays a Role in Restricting PKR Activation by Limiting the Generation of Intracellular Double-Stranded RNA
Sendai Virus C Protein Plays a Role in Restricting PKR Activation by Limiting the Generation of Intracellular Double-Stranded RNA
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DOI：
10.1128/jvi.00599-08
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发表时间：
2008-10-01
影响因子：
5.4
通讯作者：
Gotoh, Bin
中科院分区：
文献类型：
--
作者：
Takeuchi, Kenji;Komatsu, Takayuki;Gotoh, Bin
仙台病毒（SeV）C蛋白是一种多功能蛋白，在调节病毒基因组复制和转录、拮抗宿主干扰素系统、抑制病毒诱导的细胞凋亡以及促进病毒组装和出芽中发挥重要作用。在这里，我们报告了一个新的作用，SeV C蛋白，限制双链RNA（dsRNA）的产生，以维持蛋白质合成的速度在感染的细胞。发现即使在野生型（wt）SeV感染后，细胞内蛋白质合成速率仍得以维持，但在C-敲除SeV感染后明显受到抑制。这表明C蛋白在感染后维持蛋白质合成的需要。与野生型SeV感染相反，C-敲除SeV感染引起翻译起始因子eIF 2 α和dsRNA依赖性蛋白激酶（PKR）的磷酸化。eIF 2 α的磷酸化主要是由于PKR的作用，因为小干扰RNA敲低PKR限制了eIF 2 α的磷酸化。然而，C蛋白既不能抑制poly（I）：poly（C）激活的PKR和eIF 2 α的磷酸化，也不能抑制纽卡斯尔病病毒诱导的PKR和eIF 2 α的磷酸化，这表明C蛋白不靶向导致PKR激活的共同途径。用特异性识别dsRNA的单克隆抗体的免疫荧光染色实验揭示了在用C-敲除SeV而不是wt SeV感染的细胞中产生大量dsRNA。在组成型表达C蛋白的细胞中，由C-敲除SeV诱导的dsRNA产生以及PKR和eIF 2 α的磷酸化被显著抑制。总之，这些结果表明SeV C蛋白限制dsRNA的产生，从而保持PKR失活以维持细胞内蛋白质合成。
Sendai virus (SeV) C protein is a multifunctional protein that plays important roles in regulating viral genome replication and transcription, antagonizing the host interferon system, suppressing virus-induced apoptosis, and facilitating virus assembly and budding. We here report a novel role of SeV C protein, the limitation of double-stranded RNA (dsRNA) generation for maintaining the rate of protein synthesis in infected cells. It was found that the intracellular protein synthesis rate was maintained even after wild-type (wt) SeV infection, but markedly suppressed following C-knockout SeV infection. This indicates the requirement of C protein for maintaining protein synthesis after infection. In contrast to wt SeV infection, C-knockout SeV infection caused phosphorylation of both the translation initiation factor eIF2 alpha and dsRNA-dependent protein kinase (PKR). Phosphorylation of eIF2 alpha occurred mainly due to the action of PKR, since knockdown of PKR by small interfering RNA limited eIF2 alpha phosphorylation. C protein, however, could inhibit neither poly(I): poly(C)-activated nor Newcastle disease virus-induced phosphorylation of PKR and eIF2 alpha, suggesting that C protein does not target common pathways leading to PKR activation. Immunofluorescent staining experiments with a monoclonal antibody specifically recognizing dsRNA revealed generation of a large amount of dsRNA in cells infected with C-knockout SeV but not wt SeV. The dsRNA generation as well as phosphorylation of PKR and eIF2 alpha induced by C-knockout SeV was markedly suppressed in cells constitutively expressing C protein. Taken together, these results demonstrate that the SeV C protein limits generation of dsRNA, thereby keeping PKR inactive to maintain intracellular protein synthesis.