Understanding of viral replication through the interaction with coronavirus nsp1 and host proteins

严重急性呼吸综合征冠状病毒(SARS-CoV)是一种新出现的疾病的病原体。NSP1与40S核糖体亚基结合，导致表达NSP1的细胞的翻译关闭。NSP1-40S复合体进一步在宿主mRNA的5‘非编码区附近诱导内核裂解RNA，从而促进RNA的衰退。然而，nsp1诱导RNA裂解的机制尚不清楚。通过下拉纯化和质谱分析，我们确定了UPF1是NSP1的结合伙伴。UPF1是一种主要与细胞质RNA结合的蛋白，在无意义介导的RNA衰变途径中是必不可少的。在转基因细胞中的免疫共沉淀分析证实了NSP1和UPF1之间的特异性相互作用。小干扰RNA介导的UPF1基因敲除可抑制NSP1介导的RNA切割。我们的结果提示了SARA冠状病毒NSP1蛋白对宿主mRNA的一种新的切割机制。

Severe acute respiratory syndrome coronavirus (SARS-CoV) is the etiological agent of a newly emerged disease. Nsp1 binds to 40S ribosome subunits, which causes a translational shutoff in the cells expressing nsp1. The nsp1-40S complex further induces an endonucleolytic RNA cleavage near the 5'UTR of host mRNA, resulting in the promotion of RNA decay. However, the mechanism by which nsp1-induced RNA cleavage is unknown. We identified Upf1 as a binding partner of nsp1 by pulldown purification and mass spectrometry. Upf1 is a predominantly cytoplasmic RNA-binding protein that is essential for nonsense-mediated RNA decay pathway. Coimmunoprecipitation analyses in transfected cells confirmed a specific interaction between nsp1 and Upf1. Small interfering RNA-mediated knockdown of Upf1 resulted in suppression of nsp1-mediated RNA cleavage. Our data indicate a novel mechanism of host mRNA cleavage by SARA-CoV nsp1 protein.