Yong Ran

ISG56 is a negative-feedback regulator of virus-triggered signaling and cellular antiviral response

IFN-stimulated gene 56 (ISG56) is one of the first identified proteins induced by viruses and type I IFNs. In this study, we identified ISG56 as a virus-induced protein associated with MITA, an adapter protein involved in virus-triggered induction of type I IFNs. Overexpression of ISG56 inhibited Sendai virus-triggered activation of IRF3, NF-κB, and the IFN-β promoter, whereas knockdown of ISG56 had opposite effects. Consistently, overexpression of ISG56 reversed cytoplasmic poly(I:C)-induced inhibition of vesicular stomatitis virus (VSV) replication, whereas knockdown of ISG56 inhibited VSV replication. Competitive coimmunoprecipitation experiments indicated that ISG56 disrupted the interactions between MITA and VISA or TBK1, two components in the virus-triggered IFN signaling pathways. These results suggest that ISG56 is a mediator of negative-feedback regulation of virus-triggered induction of type I IFNs and cellular antiviral responses.

Regulation of virus-triggered signaling by OTUB1- and OTUB2-mediated deubiquitination of TRAF3 and TRAF6.

Ubiquitination and deubiquitination have emerged as critical post-translational regulatory mechanisms for activation or attenuation of the virus-triggered type I interferon (IFN)2 induction pathways. In this study, we identified two deubiquitinating enzymes, OTUB1 and OTUB2, as negative regulators of virus-triggered type I IFN induction. Overexpression of OTUB1 and OTUB2 inhibited virus-induced activation of IRF3 and NF-κB, transcription of the IFNB1 gene as well as cellular antiviral response, whereas knockdown of OTUB1 and OTUB2 had opposite effects. Coimmunoprecipitations indicated OTUB1 and -2 interacted with TRAF3 and TRAF6, two E3 ubiquitin ligases required for virus-triggered IRF3 and NF-κB activation, respectively. Furthermore, we found that OTUB1 and OTUB2 mediated virus-triggered deubiquitination of TRAF3 and -6. These findings suggest that OTUB1 and OTUB2 negatively regulate virus-triggered type I IFN induction and cellular antiviral response by deubiquitinating TRAF3 and -6.

Tripartite motif 8 (TRIM8) modulates TNFα- and IL-1β–triggered NF-κB activation by targeting TAK1 for K63-linked polyubiquitination

The tripartite motif (TRIM)-containing proteins are a family of proteins that have been known to be involved in divergent biological processes, including important roles in immune responses through regulating various signaling pathways. In this study, we identified a member of the TRIM family, TRIM8, as a positive regulator of tumor necrosis factor-α (TNFα) and interleukin-1β (IL-1β)–triggered NF-κB activation. Overexpression of TRIM8 activated NF-κB and potentiated TNFα- and IL-1β–induced activation of NF-κB, whereas knockdown of TRIM8 had opposite effects. Coimmunoprecipitations indicated that TRIM8 interacted with TGFβ activated kinase 1 (TAK1), a serine/threonine kinase essential for TNFα- and IL-β–induced NF-κB activation. Furthermore, we found that TRIM8 mediated K63-linked polyubiquitination of TAK1 triggered by TNFα and IL-1β. Our findings demonstrate that TRIM8 serves as a critical regulator of TNFα- and IL-1β–induced NF-κB activation by mediating K63-linked polyubiquitination of TAK1.

WDR5 is essential for assembly of the VISA-associated signaling complex and virus-triggered IRF3 and NF-κB activation

Viral infection causes activation of the transcription factors NF-κB and IRF3, which collaborate to induce type I interferons (IFNs) and cellular antiviral response. The mitochondrial outer membrane protein VISA acts as a critical adapter for assembling a virus-induced complex that signals NF-κB and IRF3 activation. Using a biochemical purification approach, we identified the WD repeat protein WDR5 as a VISA-associated protein. WDR5 was recruited to VISA in a viral infection dependent manner. Viral infection also caused translocation of WDR5 from the nucleus to mitochondria. Knockdown of WDR5 impaired the formation of virus-induced VISA-associated complex. Consistently, knockdown of WDR5 inhibited virus-triggered activation of IRF3 and NF-κB as well as transcription of the IFNB1 gene. These findings suggest that WDR5 is essential in assembling a virus-induced VISA-associated complex and plays an important role in virus-triggered induction of type I IFNs.

The ER-Associated Protein ZDHHC1 Is a Positive Regulator of DNA Virus-Triggered, MITA/STING-Dependent Innate Immune Signaling

Viral DNA sensing within the cytosol of infected cells activates type I interferon (IFN) expression. MITA/STING plays an essential role in this pathway by acting as both a sensor for the second messenger cGAMP and as an adaptor for downstream signaling components. In an expression screen for proteins that can activate the IFNB1 promoter, we identified the ER-associated protein ZDHHC1 as a positive regulator of virus-triggered, MITA/STING-dependent immune signaling. Zdhhc1(-/-) cells failed to effectively produce IFNs and other cytokines in response to infection with DNA but not RNA viruses. Zdhhc1(-/-) mice infected with the neurotropic DNA virus HSV-1 exhibited lower cytokine levels and higher virus titers in the brain, resulting in higher lethality. ZDHHC1 constitutively associated with MITA/STING and mediates dimerization/aggregation of MITA/STING and recruitment of the downstream signaling components TBK1 and IRF3. These findings support a role for ZDHHC1 in mediating MITA/STING-dependent innate immune response against DNA viruses.

MITA/STING: a central and multifaceted mediator in innate immune response.

Abstract The recognition of nucleic acids is a general strategy used by the host to detect invading pathogens. Many studies have established that MITA/STING is a central component in the innate immune response to cytosolic DNA and RNA derived from pathogens. MITA can act both as a direct sensor of cyclic dinucleotides (CDNs) and as an adaptor for the recruitment of downstream signaling components. In both roles, MITA is part of signaling cascades that orchestrate innate immune defenses against various pathogens, including viruses, bacteria and parasites. Here, we highlight recent studies that have uncovered the molecular mechanisms of MITA-mediated signal transduction and regulation, and discuss some notable issues that remain elusive.

The Dual-specificity Phosphatase DUSP14 Negatively Regulates Tumor Necrosis Factor- and Interleukin-1-induced Nuclear Factor-κB Activation by Dephosphorylating the Protein Kinase TAK1

Background: Activation of TAK1 is an essential step in TNF- and IL-1-induced NF-κB activation pathways. Results: DUSP14 dephosphorylates TAK1 at Thr-187 and negatively regulates TNF- and IL-1-induced NF-κB activation. Conclusion: DUSP14 is a negative regulator of TNF- and IL-1-induced NF-κB activation. Significance: Our study reveals a new post-translational regulatory mechanism of NF-κB activation triggered by the proinflammatory cytokines. The transcription factor NF-κB is critically involved in the inflammatory response triggered by the proinflammatory cytokines TNF and IL-1. Various studies have demonstrated that activation of TAK1 (TGF-β-activated kinase 1) is an essential step in TNF- and IL-1-induced NF-κB activation pathways. In this study, we identified a member of the dual-specificity phosphatase family, DUSP14, as a negative regulator of TNF- and IL-1-triggered NF-κB activation by expression screens. We found that DUSP14 interacted with TAK1 and that this interaction was enhanced by TNF or IL-1 stimulation. Overexpression of DUSP14 dephosphorylated TAK1 at Thr-187, a residue in the activation loop critically involved in TAK1 activation. Knockdown of DUSP14 increased basal as well as TNF- and IL-1-induced TAK1 phosphorylation at Thr-187. Overexpression of DUSP14, but not its phosphatase-deficient mutant, inhibited TNF- and IL-1-induced as well as TAK1-mediated NF-κB activation, whereas knockdown of DUSP14 had opposite effects. These findings suggest that DUSP14 negatively regulates TNF- or IL-1-induced NF-κB activation by dephosphorylating TAK1 at Thr-187. Our study reveals a new post-translational regulatory mechanism of NF-κB activation triggered by the proinflammatory cytokines.

RAVER1 is a coactivator of MDA5-mediated cellular antiviral response

Detection of viral nucleic acids by pattern recognition receptors initiates type I interferon (IFN) induction and innate antiviral response. The RIG-I-like receptors (RLRs), including RIG-I and MDA5, recognize cytoplasmic viral RNA in most cell types and are critically involved in innate antiviral response. RIG-I and MDA5 are structurally related and mediate similar signaling pathways. While the regulation of RIG-I activity has been extensively investigated, little is known about the regulatory mechanisms of MDA5 activity. Here we identified ribonucleoprotein PTB-binding 1 (RAVER1) as a specific MDA5-interacting protein. RAVER1 was associated with MDA5 upon viral infection. Overexpression of RAVER1 at low dosages enhanced MDA5- but not RIG-I-mediated activation of the IFN-β promoter, whereas knockdown of RAVER1 inhibited MDA5- but not RIG-I-mediated induction of downstream antiviral genes. Mechanistically, overexpression of RAVER1 enhanced the binding of MDA5 to its ligand poly(I:C), whereas knockdown of RAVER1 had opposite effect. Our findings suggest that RAVER1 specifically regulates MDA5 activity, revealing a mechanism of differential regulation of MDA5- and RIG-I-mediated innate antiviral response.

Linear ubiquitination of NEMO brakes the antiviral response.

NEMO is a key component of antiviral signaling. Belgnaoui et al. (2012) discover that linear ubiquitination of NEMO at a late phase of virus infection switches NEMO from a positive to a negative regulator of RIG-I-mediated interferon (IFN) induction by disrupting the VISA/MAVS-TRAF3 complex and thus terminating the antiviral response.

LSm14A Plays a Critical Role in Antiviral Immune Responses by Regulating MITA Level in a Cell-Specific Manner

Viral infection triggers induction of antiviral cytokines and effectors, which are critical mediators of innate antiviral immune response. It has been shown that the processing body–associated protein LSm14A is involved in the induction of antiviral cytokines in cell lines but in vivo evidence is lacking. By generating LSm14A-deficient mice, in this study, we show that LSm14A plays a critical and specific role in the induction of antiviral cytokines in dendritic cells (DCs) but not in macrophages and fibroblasts. Induction of antiviral cytokines triggered by the DNA viruses HSV-1 and murid herpesvirus 68 and the RNA virus vesicular stomatitis virus but not Sendai virus was impaired in Lsm14a−/− DCs, which is correlated to the functions of the adaptor protein MITA/STING in the antiviral signaling pathways. LSm14A deficiency specifically downregulated MITA/STING level in DCs by impairing its nuclear mRNA precursor processing and subsequently impaired antiviral innate and adaptive immune responses. Our findings reveal a nuclear mRNA precursor processing and cell-specific regulatory mechanism of antiviral immune responses.