The dengue virus (DENV) replication complex is made up of its non-structural (NS) proteins and yet-to-be identified host proteins, but the molecular interactions between these proteins are not fully elucidated. In this work, we sought to uncover the interactions between DENV NS1 and its fellow NS proteins using a yeast two-hybrid (Y2H) approach, and found that domain II of NS1 binds to an N-terminal cytoplasmic fragment of NS4A. Mutations in amino acid residues 41 and 43 in this cytoplasmic region of NS4A disrupted the interaction between NS1 and the NS4A-2K-4B precursor protein. When the NS4A Y41F mutation was introduced into the context of the virus via a DENV2 infectious clone, this mutant virus exhibited impaired viral fitness and decreased infectious virus production. The NS4A Y41F mutant virus triggered a significantly muted transcriptional activation of interferon-stimulated genes compared to wild-type virus that is independent of NS4A’s ability to antagonize type I interferon signalling. Taken together, we have identified a link between DENV NS1 and the cytoplasmic domain in NS4A that is important for its cellular and viral functions.

中文翻译：

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登革热病毒NS4A胞质结构域的突变会影响病毒适应性以及与其他非结构蛋白的相互作用。

登革热病毒（DENV）复制复合物由其非结构性（NS）蛋白和尚待鉴定的宿主蛋白组成，但这些蛋白之间的分子相互作用尚未完全阐明。在这项工作中，我们试图使用酵母双杂交（Y2H）方法揭示DENV NS1及其同系NS蛋白之间的相互作用，并发现NS1的结构域II与NS4A的N端胞质片段结合。NS4A的这个细胞质区域中氨基酸残基41和43的突变破坏了NS1和NS4A-2K-4B前体蛋白之间的相互作用。当通过DENV2感染性克隆将NS4A Y41F突变引入病毒时，这种突变型病毒显示出病毒适应性受损，感染性病毒产生减少。与野生型病毒相比，NS4A Y41F突变病毒触发了干扰素刺激基因的显着沉默转录激活，而这与NS4A拮抗I型干扰素信号传导的能力无关。两者合计，我们已经确定了DENV NS1和NS4A中的胞质结构域之间的联系，这对于其细胞和病毒功能很重要。