Modulation of ribosomal frameshifting frequency and its effect on the replication of Rous sarcoma virus.
Modulation of ribosomal frameshifting frequency and its effect on the replication of Rous sarcoma virus.
复制标题
核糖体移码频率的调节及其对劳斯肉瘤病毒复制的影响。
DOI：
10.1128/jvi.01846-12
复制
发表时间：
2012
影响因子：
5.4
通讯作者：
Nikolic EI
中科院分区：
文献类型：
--
作者：
Nikolic EI
程序化-1核糖体移码广泛用于RNA病毒复制酶的表达，是抗病毒干预的潜在靶点。人们对确定在病毒复制受到损害之前可以调节移码效率的程度感兴趣，我们已经使用α逆转录病毒劳斯肉瘤病毒（RSV）作为模型系统解决了这个问题。在RSV中，移码对于从重叠的gagandpol编码序列产生Gag-Pol多蛋白至关重要。移码信号由两个元件组成，一个是七核苷酸滑动序列，另一个是下游的刺激性 RNA 结构，该结构被认为是 RNA 假结。将点突变引入感染性 RSV 克隆的移码信号中，并在转染和随后感染易感细胞后监测病毒复制。引入的突变旨在产生一系列移码效率，但对编码氨基酸的影响最小。我们的结果表明，导致移码效率降低 3 倍的点突变会显着减少病毒复制，并且进一步减少会产生严重的抑制作用。相比之下，3倍移码刺激的耐受性良好。这些观察结果表明，移码的小分子抑制剂可能具有作为抗病毒干预药物的潜力。在这项工作的过程中，我们首次在体内证实了RSV刺激性RNA确实是RNA假结，但假结并不是病毒生存所必需的。
Programmed −1 ribosomal frameshifting is widely used in the expression of RNA virus replicases and represents a potential target for antiviral intervention. There is interest in determining the extent to which frameshifting efficiency can be modulated before virus replication is compromised, and we have addressed this question using the alpharetrovirus Rous sarcoma virus (RSV) as a model system. In RSV, frameshifting is essential in the production of the Gag-Pol polyprotein from the overlappinggagandpolcoding sequences. The frameshift signal is composed of two elements, a heptanucleotide slippery sequence and, just downstream, a stimulatory RNA structure that has been proposed to be an RNA pseudoknot. Point mutations were introduced into the frameshift signal of an infectious RSV clone, and virus replication was monitored following transfection and subsequent infection of susceptible cells. The introduced mutations were designed to generate a range of frameshifting efficiencies, yet with minimal impact on encoded amino acids. Our results reveal that point mutations leading to a 3-fold decrease in frameshifting efficiency noticeably reduce virus replication and that further reduction is severely inhibitory. In contrast, a 3-fold stimulation of frameshifting is well tolerated. These observations suggest that small-molecule inhibitors of frameshifting are likely to have potential as agents for antiviral intervention. During the course of this work, we were able to confirm, for the first timein vivo, that the RSV stimulatory RNA is indeed an RNA pseudoknot but that the pseudoknotper seis not absolutely required for virus viability.