Aleksashin Nikolay A, Langeberg Conner J, Shelke Rohan R, Yin Tianhao, Cate Jamie H D
Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
Nucleic Acids Res. 2025 Jan 24;53(3). doi: 10.1093/nar/gkae1248.
West Nile virus (WNV) requires programmed -1 ribosomal frameshifting for translation of the viral genome. The efficiency of WNV frameshifting is among the highest known. However, it remains unclear why WNV exhibits such a high frameshifting efficiency. Here, we employed dual-luciferase reporter assays in multiple human cell lines to probe the RNA requirements for highly efficient frameshifting by the WNV genome. We find that both the sequence and structure of a predicted RNA pseudoknot downstream of the slippery sequence-the codons in the genome on which frameshifting occurs-are required for efficient frameshifting. We also show that multiple proposed RNA secondary structures downstream of the slippery sequence are inconsistent with efficient frameshifting. We also find that the base of the pseudoknot structure likely is unfolded prior to frameshifting. Finally, we show that many mutations in the WNV slippery sequence allow efficient frameshifting, but often result in aberrant shifting into other reading frames. Mutations in the slippery sequence also support a model in which frameshifting occurs concurrent with or after ribosome translocation. These results provide a comprehensive analysis of the molecular determinants of WNV-programmed ribosomal frameshifting and provide a foundation for the development of new antiviral strategies targeting viral gene expression.
西尼罗河病毒（WNV）需要程序性-1核糖体移码来翻译病毒基因组。WNV移码的效率在已知的移码效率中是最高的之一。然而，目前尚不清楚WNV为何表现出如此高的移码效率。在此，我们在多种人类细胞系中采用双荧光素酶报告基因检测法，以探究WNV基因组高效移码所需的RNA条件。我们发现，滑序列（基因组中发生移码的密码子）下游预测的RNA假结的序列和结构对于高效移码都是必需的。我们还表明，滑序列下游多个提出的RNA二级结构与高效移码不一致。我们还发现，假结结构的基部在移码之前可能是解开的。最后，我们表明，WNV滑序列中的许多突变允许高效移码，但往往会导致异常移码到其他阅读框。滑序列中的突变也支持一种模型，即移码与核糖体易位同时发生或在核糖体易位之后发生。这些结果对WNV程序性核糖体移码的分子决定因素进行了全面分析，并为开发针对病毒基因表达的新抗病毒策略奠定了基础。
