NF-κB functions together with another transcription factor, IRF3, to induce type-I interferon production upon viral infection. After viral infection, viral RNA is detected by the RIG-I family of RNA helicases. In 2005, we identified the protein MAVS (also known as IPS-1, VISA or CARDIF) as a key adaptor for RIG-I signaling. MAVS is localized on the mitochondrial outer membrane and this localization is indispensible for its function, as underscored by the finding that viral protease NS3/4A of hepatitis C virus efficiently cleaves MAVS off the mitochondrial membrane to suppress interferon induction. The essential role of MAVS in defense against RNA virus is further confirmed in MAVS knock out mice. Recently, we found that the RIG-I pathway is also important for detecting commensal bacterial RNA and maintaining intestinal homeostasis.
Mechanism of signal transduction in the RIG-I–MAVS pathway
Our recent work has focused on the mechanism of signal transduction in the RIG-I pathway. Through the development of a cell-free system that recapitulates the RIG-I pathway from detection of viral RNA to activation of IRF3, we found that ubiquitination also plays a key role in RIG-I activation. Specifically, after binding to RNA, RIG-I undergoes a conformational change that exposes its N-terminal CARD domains, which then binds to unanchored K63 polyubiquitin chains. This binding promotes the formation of RIG-I tetramer, which interacts with MAVS and promotes MAVS aggregation. Strikingly, MAVS aggregates catalyze the polymerization of other MAVS on the mitochondria through a prion-like mechanism. These prion-like fibers of MAVS recruits multiple ubiquitin E3 ligases, including TRAF2 TRAF5 and TRAF6, to potently activate the cytosolic kinases IKK and TBK1, leading to the activation of NF-κB and IRF3, respectively.
Interferon induction by cytosolic DNA
We are also interested in how cytosolic DNA induces type-I interferons, which are important for immune defense against DNA viruses and intracellular bacteria. Furthermore, inappropriate presence of cytosolic self-DNA could trigger autoimmune diseases. We have found that AT-rich DNAs are transcribed by RNA polymerase III into RNAs bearing 5’-triphosphates, which then induce interferons through the RIG-I pathway. However, most DNA induces interferons in a sequence-independent manner that depends on the endoplasmic reticulum protein STING (also known as MITA). We have recently shown that after stimulation, STING recruits both TBK1 and IRF3, thereby specifying IRF3 phosphorylation by TBK1.
Discovery of cGAS-cGAMP pathway and its role in immune defense
We identified cGAS, which directly binds to cytosolic DNA and synthsizes cGAMP from GTP and ATP. cGAMP with mixed phosphodiester linkages binds to STING with high-affinity and induces STING conformational change and interferon expression. We generated cGAS knockout mice and cell lines. Using these reagents we found that cGAS-cGAMP pathway is important for immune response against DNA viruses and retroviruses.
Novel functions of TLRs
We maintain a considerable degree of flexibility in venturing into other areas of interest in cell signaling and host defense. For example, our recent work on commensal bacteria led us to discover the role of TLR13 in detecting the bacterial 23S ribosomal RNA (rRNA). Remarkably, we found that TLR13 recognizes a specific sequence of about 13 nucleotides near the active site of the 23S rRNA, which catalyzes peptide bond synthesis. Thus, unlike other innate immune sensors that detect a ‘pattern’ of microbial components, TLR13 detects bacterial RNA with exquisite sequence specificity.
- Seth, R.B., Sun, L., Ea, C., and Chen, Z.J. (2005) Identification and characterization of MAVS: a mitochondrial antiviral signaling protein that activates NF-κB and IRF3. Cell 122, 669-682.
- Li, X-D., Sun, L., Seth, R.B., Pineda, G., and Chen, Z.J. (2005) The Hepatitis C Virus Protease NS3/4A Cleaves MAVS off the Mitochondria to Evade Innate Immunity. Proc. Natl. Acad. Sci. U S A102, 17717-17722.
- Sun, Q., Sun, L., Liu, H-H., Chen, X., Seth, R.B., Forman, J. and Chen, Z.J. (2006) The specific and essential role of MAVS in antiviral innate immune responses. Immunity 24, 633-642.
- Bhoj, V.G., Sun, Q., Bhoj, E., Somers, C., Chen, X., Torres, J-P., Mejias, A., Gomez., A., Jafri, H., Ramilo, O., Chen, Z.J. (2008). MAVS and MyD88 are essential for innate immunity but not cytotoxic T lymphocyte response against respiratory syncytial virus. Proc. Natl. Acad. Sci. U S A 105, 14046-14051.
- Li, X.D., Chiu, Y.H., Ismail, A.S., Behrendt, C.L., Wight-Carter, M., Hooper, L.V., and Chen, Z.J. (2011). Mitochondrial antiviral signaling protein (MAVS) monitors commensal bacteria and induces an immune response that prevents experimental colitis. Proc Natl Acad Sci U S A108, 17390-17395.
- Zeng, W., Xu, M., Liu, S., Sun, L., Chen, Z.J. (2009) Key role of Ubc5 and K63 polyubiquitination in viral activation of IRF3. Molecular Cell 36, 302-314.
- Zeng, W., Sun, L., Jiang, X., Chen, X., Hou, F., Adhikari, A., Xu, M., and Chen, Z.J. (2010) Reconstitution of the RIG-I pathway reveals a signaling role of unanchored polyubiquitin chains in innate immunity. Cell 141, 315-330.
- Hou, F., Sun, L., Zheng, H., Skaug, B., Jiang, Q.X., and Chen, Z.J. (2011). MAVS Forms Functional Prion-like Aggregates to Activate and Propagate Antiviral Innate Immune Response. Cell 146, 448-461.
- Jiang, X., Kinch, L., Brautigam, C.A., Chen, X., Du, F., Grishin, N., Chen, Z.J. (2012) Ubiquitin-induced oligomerization of RIG-I and MDA5 activates antiviral inate immune response. Immunity 36, 959-973.
- Liu, S., Chen, J., Cai, X., Wu, J., Chen, X., Wu, Y-T., Sun, L., and Chen, Z.J. (2013) MAVS recruits mutliple ubiquitin E3 ligases to activate antiviral signaling cascades. eLife 2:e00785
- Chiu, Y.H., Macmillan, J.B., and Chen, Z.J. (2009). RNA polymerase III detects cytosolic DNA and induces type I interferons through the RIG-I pathway. Cell 138, 576-591.
- Tanaka, Y., and Chen, Z.J. (2012). STING Specifies IRF3 Phosphorylation by TBK1 in the Cytosolic DNA Signaling Pathway. Science Signaling 5, ra20.
- Wu, J., Sun, L., Chen, X., Du, F., Shi, H., Chen, C., Chen Z.J. (2012) Cyclic GMP-AMP Is an Endogenous Second Messenger in Innate Immune Signaling by Cytosolic DNA. Science 339, 826-830.
- Sun, L., Wu, J., Du, F., Chen, X., Chen, Z.J. (2012) Cyclic GMP-AMP Synthase Is a Cytosolic DNA Sensor That Activates the Type I Interferon Pathway. Science 339, 786-791.
- Zhang, X., Shi, H., Wu, J., Zhang, X., Sun, L., Chen, C., and Chen, Z.J. (2013) Cyclic GMP-AMP containing mixed phosphodiester linkages is an endogenous high-affinity ligand for STING. Mol. Cell 51:226-235.
- Li, X.D., Wu, J., Gao, D., Wang, H., Sun, L., and Chen, Z.J. (2013) Pivotal Roles of cGAS-cGAMP Signaling in Antiviral Defense and Immune Adjuvant Effects. Science in press.
- Li, X. and Chen, Z.J. (2012). Sequence specific detection of bacterial 23S ribosomal RNA by TLR13. eLife 1, e00102