Abstract：
The Dengue virus (DENV) is the most significant arthropod-borne viral pathogen in humans with 400 million infections annually. DENV comprises four distinct serotypes (DENV-1 to -4) which complicates vaccine development. Any of the four serotypes can cause clinical illness but with distinctive infection dynamics. Variations in sequences identified within the four genomes induce structural differences in crucial RNA motifs that were suggested to be correlated to the degree of pathogenicity among DENV-1 to -4. In particular, the RNA Stem-loop A (SLA) at the 5\'-end of the genome, acts as a key regulator of the viral replication cycle by interacting with the viral NS5 polymerase to initiate the minus-strand viral RNA synthesis and later to methylate and cap the synthesized RNA. The molecular details of this interaction remain not fully described. Here, we report the solution secondary structures of SLA from DENV-1 to -4. Our results highlight that the four SLA exhibit structural and dynamic differences. Secondly, to determine whether SLA RNA contains serotype-specific determinants for the recognition by the viral NS5 protein, we investigated interactions between SLA from DENV -1 to -4 and DENV2 NS5 using combined biophysical approaches. Our results show that NS5 from DENV2 is able to bind SLA from other serotypes, but that other viral or host factors may be necessary to stabilize the complex and promote the catalytically active state of the NS5. By contrast, we show that a serotype-specific binding is driven by specific interactions involving conformational changes within the SLA RNA.
摘要：
登革热病毒（DENV）是人类中最重要的节肢动物传播的病毒病原体，每年感染4亿。DENV包含四种不同的血清型(DENV-1至-4),这使疫苗开发复杂化。四种血清型中的任何一种都可以引起临床疾病，但具有独特的感染动力学。在四个基因组中鉴定的序列的变化诱导关键RNA基序的结构差异，这些结构差异被认为与DENV-1至-4之间的致病性程度相关。特别是,基因组5'末端的RNA茎环A（SLA），通过与病毒NS5聚合酶相互作用以启动负链病毒RNA合成，然后甲基化和封顶合成的RNA，从而充当病毒复制周期的关键调节剂。这种相互作用的分子细节仍未完全描述。这里,我们报告了从DENV-1到-4的SLA的溶液二级结构。我们的结果突出表明，四个SLA表现出结构和动态差异。其次,为了确定SLARNA是否含有血清型特异性决定子，用于病毒NS5蛋白的识别，我们使用联合生物物理方法研究了从DENV-1到-4的SLA与DENV2NS5之间的相互作用。我们的结果表明，来自DENV2的NS5能够结合来自其他血清型的SLA，但可能需要其他病毒或宿主因子来稳定复合物并促进NS5的催化活性状态。相比之下,我们表明血清型特异性结合是由涉及SLARNA内构象变化的特异性相互作用驱动的。
