Miras Manuel, Rodríguez-Hernández Ana M, Romero-López Cristina, Berzal-Herranz Alfredo, Colchero Jaime, Aranda Miguel A, Truniger Verónica
Centro de Edafología y Biología Aplicada del Segura, Consejo Superior de Investigaciones Científicas (CEBAS-CSIC), Murcia, Spain.
Centro de Investigación en Química Aplicada, Consejo Nacional de Ciencia y Tecnología (CONACYT), Saltillo, Mexico.
Front Plant Sci. 2018 May 9;9:625. doi: 10.3389/fpls.2018.00625. eCollection 2018.
In eukaryotes, the formation of a 5'-cap and 3'-poly(A) dependent protein-protein bridge is required for translation of its mRNAs. In contrast, several plant virus RNA genomes lack both of these mRNA features, but instead have a 3'-CITE (for cap-independent translation enhancer), a RNA element present in their 3'-untranslated region that recruits translation initiation factors and is able to control its cap-independent translation. For several 3'-CITEs, direct RNA-RNA long-distance interactions based on sequence complementarity between the 5'- and 3'-ends are required for efficient translation, as they bring the translation initiation factors bound to the 3'-CITE to the 5'-end. For the carmovirus melon necrotic spot virus (MNSV), a 3'-CITE has been identified, and the presence of its 5'-end has been shown to be required for its activity. Here, we analyze the secondary structure of the 5'-end of the MNSV RNA genome and identify two highly conserved nucleotide sequence stretches that are complementary to the apical loop of its 3'-CITE. In cap-independent translation assays with mutant constructs, by disrupting and restoring sequence complementarity, we show that the interaction between the 3'-CITE and at least one complementary sequence in the 5'-end is essential for virus RNA translation, although efficient virus translation and multiplication requires both connections. The complementary sequence stretches are invariant in all MNSV isolates, suggesting that the dual 5'-3' RNA:RNA interactions are required for optimal MNSV cap-independent translation and multiplication.
在真核生物中，其mRNA的翻译需要形成5'-帽和3'-多聚（A）依赖性蛋白质-蛋白质桥。相比之下，几种植物病毒RNA基因组缺乏这两种mRNA特征，而是具有一个3'-CITE（帽依赖性翻译增强子），这是一种存在于其3'-非翻译区的RNA元件，可募集翻译起始因子并能够控制其帽依赖性翻译。对于几种3'-CITE，基于5'-端和3'-端之间序列互补性的直接RNA-RNA长距离相互作用是有效翻译所必需的，因为它们将与3'-CITE结合的翻译起始因子带到5'-端。对于香石竹坏死斑点病毒（MNSV）这种香石竹斑驳病毒属病毒，已鉴定出一个3'-CITE，并且已证明其活性需要其5'-端的存在。在这里，我们分析了MNSV RNA基因组5'-端的二级结构，并鉴定出两个与其3'-CITE的顶端环互补的高度保守的核苷酸序列片段。在使用突变体构建体的帽依赖性翻译测定中，通过破坏和恢复序列互补性，我们表明3'-CITE与5'-端中至少一个互补序列之间的相互作用对于病毒RNA翻译至关重要，尽管有效的病毒翻译和增殖需要两者的连接。互补序列片段在所有MNSV分离株中都是不变的，这表明5'-3' RNA：RNA双重相互作用对于最佳的MNSV帽依赖性翻译和增殖是必需的。
