Human DEAD Box Helicase 3 Couples IκB Kinase ε to Interferon Regulatory Factor 3 Activation

Abstract

The human DEAD box protein 3 (DDX3) has been implicated in different processes contributing to gene expression. Interestingly, DDX3 is required as an essential host factor for the replication of HIV and hepatitis C virus (HCV) and is therefore considered a potential drug target. On the other hand, DDX3 interacts with IB kinase (IKK) and TANK-binding kinase 1 (TBK1) and contributes to the induction of antiviral type I interferons (IFNs). However, the molecular mechanism by which DDX3 contributes to IFN induction remains unclear. Here we show that DDX3 mediates phosphorylation of interferon regulatory factor 3 (IRF3) by the kinase IKK. DDX3 directly interacts with IKK and enhances its autophosphorylation and activation. IKK then phosphorylates several serine residues in the N terminus of DDX3. Phosphorylation of DDX3 at serine 102 (S102) is required for recruitment of IRF3 to DDX3, facilitating its phosphorylation by IKK. Mutation of S102 to alanine disrupted the interaction between DDX3 and IRF3 but not that between DDX3 and IKK. The S102A mutation failed to enhance ifnb promoter activation, suggesting that the DDX3-IRF3 interaction is crucial for this effect. Our data implicates DDX3 as a scaffolding adaptor that directly facilitates phosphorylation of IRF3 by IKK. DDX3 might thus be involved in pathway-specific activation of IRF3. The human DEAD box RNA helicase DDX3 is a multifunctional cellular protein which has been implicated in various processes linked to gene expression (1). Its RNA helicase activity is coopted by viruses that require DDX3 as an essential host factor for their replication, such as HIV and HCV (2-4). In contrast, we and others have previously demonstrated that DDX3 contributes to antiviral innate immune signaling pathways leading to ifnb induction (5-9). This function of DDX3 is independent of its ATPase and helicase activity (6, 7). ifnb promoter induction requires activation of the transcription factors IRF3 and IRF7, which occurs only downstream of certain, mostly antiviral, pattern recognition receptors (PRRs). Antiviral PRRs comprise the endosomal Toll-like receptors TLR3, TLR7, TLR8, and TLR9, cytosolic RIG-like helicases (RLHs), and several newly discovered cytosolic DNA receptors (10). These receptors recognize different species of viral nucleic acids and induce type I IFNs in response (11). TLR3 and TLR4 engage the TRIF (TIR domain-containing adaptor inducing IFN-␤)-dependent pathway for activation of IRF3 (12). In this pathway, phosphorylation-dependent activation of IRF3 is mediated by the IKK-related kinases TBK1 and IKKε (13,14). The RLHs also utilize TBK1 and IKKε for IRF3 activation after engaging the adaptor molecule MAVS (mitochondrial antiviral signaling) (15, 16). DNA receptors activate IRF3 through TBK1 and the adaptor molecule STING (stimulator of IFN genes).While these signaling pathways converge on IKKε and TBK1 for IRF3 activation, not all receptors that activate IKKε and/or TBK1 lead to IRF3 activation (17). This suggests that additional factors are required for li...

CIKS/DDX3X Interaction Controls the Stability of the Zc3h12a mRNA Induced by IL-17

“…It is possible to speculate that after IL-17 stimulation IKKε is first recruited to CIKS and then DDX3X is recruited to the complex to regulate its biological function. To this regard, it has been recently reported that IKKε phosphorylates DDX3X and that this phosphorylation modulated the interaction between DDX3X and IRF3 (41). Therefore, it is likely that the phosphorylation of DDX3X may affect its function also in the IL-17 pathway.…”

The ATP-Dependent RNA Helicase DDX3X Modulates Herpes Simplex Virus 1 Gene Expression

“…Not surprisingly, DDX3X was found to promote genome replication of many RNA viruses (33). However, DDX3X binds TBK1/IKK, which stimulate both IRF3 activation and interferon type I production, thereby promoting an antiviral state (10,11). This implies that DDX3X has dual functions that can either facilitate or hamper viruses.…”

“…Furthermore, DDX3X also contributes to innate immunity against these viruses. For instance, DDX3X stimulates interferon (IFN) type I production by binding IKK (IB kinase epsilon) and TBK1 (tank-binding kinase 1), leading to IRF3 phosphorylation and activation (10,11). These findings position DDX3X as a critical player for the replication and immunity against RNA-based viruses.…”

Intracellular detection of viral nucleic acids

“…Notably, many of these molecules regulate RLR signaling, and also have other functions in RNA metabolism and gene transcription. DEAD-box protein 3 (DDX3) was first proposed to function as vRNA receptor, activating MAVS-dependent signaling and IFN induction [39]; however recent research indicates that DDX3 exerts a regulatory role in innate signaling by augmenting the activation of TBK1 or IKKε via direct interaction [40]. Moreover, it has been recently reported that DDX3 functions as an antiviral effector protein that restricts HBV transcription [41].…”