Biochemical Dissection of the RIG-I Antiviral Pathway

Summary

Principal Investigator: Zhijian J Chen
Abstract: DESCRIPTION (provided by applicant): RIG-I like receptors (RLRs) detect viral infections and triggers appropriate immune responses to limit viral infection and spread. RIG-I, the best studied member of RLRs, binds to viral RNA in the cytosol and triggers a signaling cascade that leads to the production of type-I interferons and other antiviral molecules. We have previously identified a mitochondrial antiviral signaling protein termed MAVS (also known as IPS-1, VISA or CARDIF), which plays a pivotal role in the RIG-I signaling cascade. MAVS resides on the mitochondrial outer membrane and it activates cytosolic protein kinases including IKK and TBK1, which phosphorylate the transcription factors NF-?B and IRF3, respectively. NF-?B, IRF3 and other transcription factors function together in the nucleus to turn on the expression of antiviral genes. In order to understand the mechanism of signal transduction in the RIG-I pathway, we have recently established a cell-free system that faithfully recapitulates the activation of IRF3 by viral RNA. Initial dissection of this system leads to the discovery that RIG-I activation requires not only viral RNA, but also a unique form of polyubiquitination involving lysine-63 (K63) of ubiquitin. Further investigation shows that unanchored K63 polyubiquitin chains, which are not conjugated to any cellular protein, binds to the N-terminal CARD domains of RIG-I, resulting in RIG-I activation. These studies provide a strong foundation for further dissection of the RIG-I signaling pathway. In this application, we propose to elucidate the mechanism of RIG-I regulation by RNA and K63 ubiquitin chains (Aim 1). We will also investigate how RIG-I activates MAVS on the mitochondrial membrane (Aim 2). Finally, we will delineate the pathways through which MAVS activate TBK1 and IKK in the cytosol (Aim 3). Together, these lines of investigation should fill some of the major gaps in our current understanding of the RIG-I signaling cascade, and provide mechanistic insights into antiviral innate immunity. PUBLIC HEALTH RELEVANCE: The RIG-I pathway is largely responsible for the first line of immune defense against viral infection in human and many other organisms. In this application, we propose to elucidate the mechanism of signal transduction in key steps of the RIG-I pathway using an innovative biochemical approach. If successful, our research should significantly advance the fields of immunology and virology, and provide new approaches to harness the host innate immunity to combat viral and autoimmune diseases.
Funding Period: 2011-02-15 - 2016-01-31
more information: NIH RePORT

Top Publications

  1. pmc Viperin links lipid bodies to immune defense
    Xiaomo Jiang
    Department of Molecular Biology, University of Texas Southwestern Medical Center Dallas, TX 75390 9148, USA
    Immunity 34:285-7. 2011
  2. pmc Cyclic GMP-AMP containing mixed phosphodiester linkages is an endogenous high-affinity ligand for STING
    Xu Zhang
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Mol Cell 51:226-35. 2013
  3. pmc Cyclic GMP-AMP synthase is an innate immune sensor of HIV and other retroviruses
    Daxing Gao
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Science 341:903-6. 2013
  4. ncbi MAVS recruits multiple ubiquitin E3 ligases to activate antiviral signaling cascades
    Siqi Liu
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, United States
    elife 2:e00785. 2013
  5. pmc Pivotal roles of cGAS-cGAMP signaling in antiviral defense and immune adjuvant effects
    Xiao Dong Li
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Science 341:1390-4. 2013
  6. ncbi A novel mitochondrial MAVS/Caspase-8 platform links RNA virus-induced innate antiviral signaling to Bax/Bak-independent apoptosis
    Souhayla El Maadidi
    Institute of Molecular Medicine and Cell Research, Albert Ludwigs University Freiburg, D 79104 Freiburg, Germany
    J Immunol 192:1171-83. 2014
  7. pmc The cytosolic DNA sensor cGAS forms an oligomeric complex with DNA and undergoes switch-like conformational changes in the activation loop
    Xu Zhang
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA Electronic address
    Cell Rep 6:421-30. 2014
  8. pmc Prion-like polymerization underlies signal transduction in antiviral immune defense and inflammasome activation
    Xin Cai
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
    Cell 156:1207-22. 2014
  9. ncbi Innate immune sensing and signaling of cytosolic nucleic acids
    Jiaxi Wu
    Department of Molecular Biology and
    Annu Rev Immunol 32:461-88. 2014
  10. pmc Regulation of NF-κB by ubiquitination
    Jueqi Chen
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, United States
    Curr Opin Immunol 25:4-12. 2013

Detail Information

Publications21

  1. pmc Viperin links lipid bodies to immune defense
    Xiaomo Jiang
    Department of Molecular Biology, University of Texas Southwestern Medical Center Dallas, TX 75390 9148, USA
    Immunity 34:285-7. 2011
    ..In this issue of Immunity, Saitoh et al. (2011) uncovered an unexpected function of Viperin and lipid bodies in interferon induction by Toll-like receptors, specifically in plasmacytoid dendritic cells...
  2. pmc Cyclic GMP-AMP containing mixed phosphodiester linkages is an endogenous high-affinity ligand for STING
    Xu Zhang
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Mol Cell 51:226-35. 2013
    ..The crystal structure of STING bound to 2'3'-cGAMP revealed the structural basis of this high-affinity binding and a ligand-induced conformational change in STING that may underlie its activation. ..
  3. pmc Cyclic GMP-AMP synthase is an innate immune sensor of HIV and other retroviruses
    Daxing Gao
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Science 341:903-6. 2013
    ..These results indicate that cGAS is an innate immune sensor of HIV and other retroviruses. ..
  4. ncbi MAVS recruits multiple ubiquitin E3 ligases to activate antiviral signaling cascades
    Siqi Liu
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, United States
    elife 2:e00785. 2013
    ..DOI:http://dx.doi.org/10.7554/eLife.00785.001. ..
  5. pmc Pivotal roles of cGAS-cGAMP signaling in antiviral defense and immune adjuvant effects
    Xiao Dong Li
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Science 341:1390-4. 2013
    ..cGas(-/-) mice were more susceptible to lethal infection with herpes simplex virus 1 (HSV1) than wild-type mice. We also show that cGAMP is an adjuvant that boosts antigen-specific T cell activation and antibody production in mice. ..
  6. ncbi A novel mitochondrial MAVS/Caspase-8 platform links RNA virus-induced innate antiviral signaling to Bax/Bak-independent apoptosis
    Souhayla El Maadidi
    Institute of Molecular Medicine and Cell Research, Albert Ludwigs University Freiburg, D 79104 Freiburg, Germany
    J Immunol 192:1171-83. 2014
    ..Thus, after RNA virus infection MAVS does not only elicit a type I antiviral response but also recruits caspase-8 to mitochondria to mediate caspase-3 activation and apoptosis in a Bax/Bak-independent manner. ..
  7. pmc The cytosolic DNA sensor cGAS forms an oligomeric complex with DNA and undergoes switch-like conformational changes in the activation loop
    Xu Zhang
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA Electronic address
    Cell Rep 6:421-30. 2014
    ..Functional analyses of cGAS mutants demonstrate that both the protein-protein interface and the two DNA binding surfaces are critical for cGAS activation. These results provide insights into the mechanism of DNA sensing by cGAS. ..
  8. pmc Prion-like polymerization underlies signal transduction in antiviral immune defense and inflammasome activation
    Xin Cai
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
    Cell 156:1207-22. 2014
    ..These results indicate that prion-like polymerization is a conserved signal transduction mechanism in innate immunity and inflammation...
  9. ncbi Innate immune sensing and signaling of cytosolic nucleic acids
    Jiaxi Wu
    Department of Molecular Biology and
    Annu Rev Immunol 32:461-88. 2014
    ..Here, we review the recent advances in understanding the mechanism of nucleic acid sensing and signaling in the cytosol of mammalian cells as well as the emerging role of cytosolic nucleic acids in autoimmunity. ..
  10. pmc Regulation of NF-κB by ubiquitination
    Jueqi Chen
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, United States
    Curr Opin Immunol 25:4-12. 2013
    ..In this review, we will focus on recent advances in understanding the role of ubiquitination in NF-κB regulation in various pathways...
  11. pmc Cyclic GMP-AMP synthase is a cytosolic DNA sensor that activates the type I interferon pathway
    Lijun Sun
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
    Science 339:786-91. 2013
    ..cGAS bound to DNA in the cytoplasm and catalyzed cGAMP synthesis. These results indicate that cGAS is a cytosolic DNA sensor that induces interferons by producing the second messenger cGAMP...
  12. pmc Cyclic GMP-AMP is an endogenous second messenger in innate immune signaling by cytosolic DNA
    Jiaxi Wu
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
    Science 339:826-30. 2013
    ..cGAMP bound to STING, leading to the activation of IRF3 and induction of interferon-β. Thus, cGAMP functions as an endogenous second messenger in metazoans and triggers interferon production in response to cytosolic DNA...
  13. pmc MAVS forms functional prion-like aggregates to activate and propagate antiviral innate immune response
    Fajian Hou
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Cell 146:448-61. 2011
    ..These results suggest that a prion-like conformational switch of MAVS activates and propagates the antiviral signaling cascade...
  14. pmc Mitochondrial antiviral signaling protein (MAVS) monitors commensal bacteria and induces an immune response that prevents experimental colitis
    Xiao Dong Li
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Proc Natl Acad Sci U S A 108:17390-5. 2011
    ..These results unveil a previously unexplored role of MAVS in monitoring intestinal commensal bacteria and maintaining tissue homeostasis...
  15. pmc Intrinsic antiviral immunity
    Nan Yan
    Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, Texas, USA
    Nat Immunol 13:214-22. 2012
    ..This review focuses on recent advances in understanding of the roles of intrinsic antiviral factors that restrict infection by human immunodeficiency virus and influenza virus...
  16. pmc STING specifies IRF3 phosphorylation by TBK1 in the cytosolic DNA signaling pathway
    Yasuo Tanaka
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Sci Signal 5:ra20. 2012
    ..This scaffolding function of STING (and possibly of other adaptor proteins) may explain why IRF3 is activated in only a subset of signaling pathways that activate TBK1...
  17. pmc Ubiquitination in signaling to and activation of IKK
    Zhijian J Chen
    Department of Molecular Biology, Howard Hughes Medical Institute, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Immunol Rev 246:95-106. 2012
    ....
  18. pmc Ubiquitin-induced oligomerization of the RNA sensors RIG-I and MDA5 activates antiviral innate immune response
    Xiaomo Jiang
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390 9148, USA
    Immunity 36:959-73. 2012
    ..These results suggest a unified mechanism of RIG-I and MDA5 activation and reveal a unique mechanism by which ubiquitin regulates cell signaling and immune response...
  19. pmc Sequence specific detection of bacterial 23S ribosomal RNA by TLR13
    Xiao Dong Li
    Department of Molecular Biology
    elife 1:e00102. 2012
    ..Knockout of TLR13 in mice abolished the induction of interleukin-1β and other cytokines by the 23S rRNA sequence. Thus, TLR13 detects bacterial RNA with exquisite sequence specificity.DOI:http://dx.doi.org/10.7554/eLife.00102.001...
  20. ncbi The cGAS-cGAMP-STING pathway of cytosolic DNA sensing and signaling
    Xin Cai
    Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA
    Mol Cell 54:289-96. 2014
    ..In this review, we focus on the mechanism of DNA sensing by the newly discovered cGAS-cGAMP-STING pathway and highlight recent progress in dissecting the in vivo functions of this pathway in immune defense as well as autoimmunity...

Research Grants30

  1. Mechanisms of Protein Kinase Activation by Ubiquitin in the NF-kB Pathways
    Zhijian J Chen; Fiscal Year: 2013
    ..If successful, our research should reveal the fundamental mechanisms of NF-kB activation in immune and inflammatory responses and provide potential new approaches to inhibit aberrant NF-kB activation in human diseases. ..