Leiden Institute of Chemistry, University of Leiden, P.O. Box 9502, 2300 RA, Leiden, The Netherlands.
Department of Sustainable Agriculture, Mediterranean Agronomic Institute of Chania, Alsylio Agrokepion, 73100, Chania, Crete, Greece.
Sci Rep. 2022 Jul 7;12(1):11532. doi: 10.1038/s41598-022-15598-5.
Pepino mosaic virus (PepMV) is a potexvirus of the family Alphaflexiviridae within the order of Tymovirales that threatens tomato production worldwide. PepMV possesses a positive-strand RNA genome with a 5'-methylguanosine cap and a 3'-polyA tail. Previously, using partially-purified viral RNA polymerase important secondary structures within the 3'-untranslated region (UTR) of PepMV RNA were identified. Here we show that an RNA pseudoknot can be formed in the 3'-UTR that includes part of the polyA tail. Using protoplasts, we demonstrate that the pseudoknot is required for replication of PepMV RNA. Mutational analysis and native gel electrophoresis further show that the pseudoknot is stabilized by UAU base triples, as is the human telomerase RNA pseudoknot. The presence of a pseudoknot in several other members of the Alpha- and Betaflexiviridae is supported by covariance analysis and native gel electrophoresis of other potexvirus, capillovirus and trichovirus RNAs. The ubiquitous presence of the pseudoknot in viruses of the Betaflexiviridae, suggests that the pseudoknot is a typical trait of the Betaflexiviridae that may have been adopted by many potexviruses during evolution.
西瓜镶嵌病毒（ PepMV ）是一种马铃薯 Y 病毒科的正链 RNA 病毒，在弹状病毒目内。该病毒威胁着全世界的番茄生产。 PepMV 拥有一个 5'-甲基鸟苷帽和 3'-多聚 A 尾的正链 RNA 基因组。先前，使用部分纯化的病毒 RNA 聚合酶，鉴定了 PepMV RNA 3'-非翻译区（UTR ）内的重要二级结构。在这里，我们证明在 3'-UTR 中可以形成一个 RNA 假结，其中包括多聚 A 尾的一部分。通过原生质体实验，我们证明假结对于 PepMV RNA 的复制是必需的。突变分析和 native 凝胶电泳进一步表明，假结由 UAU 碱基三链稳定，正如人类端粒酶 RNA 假结一样。通过共变分析和其他马铃薯 Y 病毒、毛管病毒和绒毛病毒 RNA 的 native 凝胶电泳，支持了在 Alpha 和 Betaflexiviridae 中的几个其他成员中存在假结。假结在 Betaflexiviridae 病毒中的普遍存在表明，假结是 Betaflexiviridae 的一个典型特征，可能在进化过程中被许多马铃薯 Y 病毒采用。
