Interference of ribosomal frameshifting by antisense peptide nucleic acids suppresses SARS coronavirus replication.
Interference of ribosomal frameshifting by antisense peptide nucleic acids suppresses SARS coronavirus replication.
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DOI：
10.1016/j.antiviral.2011.04.009
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发表时间：
2011-07
影响因子：
7.6
通讯作者：
Oh JW
中科院分区：
文献类型：
--
作者：
Ahn DG;Lee W;Choi JK;Kim SJ;Plant EP;Almazán F;Taylor DR;Enjuanes L;Oh JW
真核RNA病毒利用的程序性-1核糖体移码（-1 PRF）对于基因组复制所需的病毒RNA复制酶多聚蛋白的受控、有限合成起着至关重要的作用。严重急性呼吸综合征冠状病毒（SARS-CoV）的病毒RNA复制酶多聚蛋白由两个重叠的开放阅读框架1a和1b编码，它们通过−1 PRF信号连接。我们评估了针对高度保守的RNA序列的反义肽核酸（PNA）对- PRF信号的抗病毒作用。核糖体移码被PNA抑制，PNA在细胞系中结合−1 PRF信号中的序列特异性假结结构，使用含有−1 PRF信号的基于双内切酶的报告质粒进行评估。用与细胞穿透肽（CPP）融合的PNA处理用表达萤火虫荧光素酶的SARS-CoV复制子转染的细胞，导致SARS-CoV复制子的复制受到抑制，50%抑制浓度为4.4 μM。PNA治疗没有诱导I型干扰素应答，表明PNA的作用不是由于先天免疫应答。我们的研究结果表明，-1 PRF，SARS病毒复制的关键，可以被CPP-PNA抑制，提供了一个有效的反义策略阻断-1 PRF信号。
The programmed −1 ribosomal frameshifting (−1 PRF) utilized by eukaryotic RNA viruses plays a crucial role for the controlled, limited synthesis of viral RNA replicase polyproteins required for genome replication. The viral RNA replicase polyproteins of severe acute respiratory syndrome coronavirus (SARS-CoV) are encoded by the two overlapping open reading frames 1a and 1b, which are connected by a −1 PRF signal. We evaluated the antiviral effects of antisense peptide nucleic acids (PNAs) targeting a highly conserved RNA sequence on the – PRF signal. The ribosomal frameshifting was inhibited by the PNA, which bound sequence-specifically a pseudoknot structure in the −1 PRF signal, in cell lines as assessed using a dual luciferase-based reporter plasmid containing the −1 PRF signal. Treatment of cells, which were transfected with a SARS-CoV-replicon expressing firefly luciferase, with the PNA fused to a cell-penetrating peptide (CPP) resulted in suppression of the replication of the SARS-CoV replicon, with a 50% inhibitory concentration of 4.4 μM. There was no induction of type I interferon responses by PNA treatment, suggesting that the effect of PNA is not due to innate immune responses. Our results demonstrate that −1 PRF, critical for SARS-CoV viral replication, can be inhibited by CPP-PNA, providing an effective antisense strategy for blocking −1 PRF signals.