Department of Microbiology and Immunology, Hokkaido University Graduate School of Medicine, Kita-ku, Sapporo, Japan.
Eur J Immunol. 2010 Apr;40(4):940-8. doi: 10.1002/eji.200940203.
Retinoic acid-inducible gene-I (RIG-I)-like receptors (RLR) are members of the DEAD box helicases, and recognize viral RNA in the cytoplasm, leading to IFN-beta induction through the adaptor IFN-beta promoter stimulator-1 (IPS-1) (also known as Cardif, mitochondrial antiviral signaling protein or virus-induced signaling adaptor). Since uninfected cells usually harbor a trace of RIG-I, other RNA-binding proteins may participate in assembling viral RNA into the IPS-1 pathway during the initial response to infection. We searched for proteins coupling with human IPS-1 by yeast two-hybrid and identified another DEAD (Asp-Glu-Ala-Asp) box helicase, DDX3 (DEAD/H BOX 3). DDX3 can bind viral RNA to join it in the IPS-1 complex. Unlike RIG-I, DDX3 was constitutively expressed in cells, and some fraction of DDX3 is colocalized with IPS-1 around mitochondria. The 622-662 a.a DDX3 C-terminal region (DDX3-C) directly bound to the IPS-1 CARD-like domain, and the whole DDX3 protein also associated with RLR. By reporter assay, DDX3 helped IPS-1 up-regulate IFN-beta promoter activation and knockdown of DDX3 by siRNA resulted in reduced IFN-beta induction. This activity was conserved on the DDX3-C fragment. DDX3 only marginally enhanced IFN-beta promoter activation induced by transfected TANK-binding kinase 1 (TBK1) or I-kappa-B kinase-epsilon (IKKepsilon). Forced expression of DDX3 augmented virus-mediated IFN-beta induction and host cell protection against virus infection. Hence, DDX3 is an antiviral IPS-1 enhancer.
维甲酸诱导基因 I（RIG-I）样受体（RLR）是 DEAD 盒解旋酶的成员，它们在细胞质中识别病毒 RNA，通过衔接子 IFN-β 启动子刺激物-1（IPS-1）（也称为 Cardif、线粒体抗病毒信号蛋白或病毒诱导的信号衔接子）诱导 IFN-β 的产生。由于未感染的细胞通常含有微量的 RIG-I，因此在感染的初始反应中，其他 RNA 结合蛋白可能参与将病毒 RNA 组装到 IPS-1 途径中。我们通过酵母双杂交筛选与人类 IPS-1 相互作用的蛋白，并鉴定出另一种 DEAD（天冬氨酸-谷氨酸-丙氨酸-天冬氨酸）盒解旋酶 DDX3（DEAD/H 盒 3）。DDX3 可以结合病毒 RNA 并将其加入 IPS-1 复合物中。与 RIG-I 不同，DDX3 在细胞中持续表达，并且 DDX3 的一部分与 IPS-1 周围的线粒体共定位。DDX3 的 622-662 a.a C 末端区域（DDX3-C）直接与 IPS-1 CARD 样结构域结合，整个 DDX3 蛋白也与 RLR 结合。通过报告基因检测，DDX3 有助于 IPS-1 上调 IFN-β 启动子的激活，siRNA 敲低 DDX3 导致 IFN-β 诱导减少。这种活性在 DDX3-C 片段上是保守的。DDX3 仅略微增强转染的 TANK 结合激酶 1（TBK1）或 I-κB 激酶-ε（IKKepsilon）诱导的 IFN-β 启动子激活。DDX3 的强制表达增强了病毒介导的 IFN-β 诱导和宿主细胞对病毒感染的保护。因此，DDX3 是一种抗病毒 IPS-1 增强子。
