Innate sensing of picornavirus infection involves cGAS-STING-mediated antiviral responses triggered by mitochondrial DNA release

Cyclic GMP-AMP synthase (cGAS) plays a key role in the innate immune responses to both DNA and RNA virus infection. Here, we found that enterovirus 71 (EV-A71), Seneca Valley virus (SVV), and foot-and-mouth disease virus (FMDV) infection triggered mitochondria damage and mitochondrial DNA (mtDNA) release in vitro and vivo. These responses were mediated by picornavirus 2B proteins which induced mtDNA release during viral replication. SVV infection caused the opening of mitochondrial permeability transition pore (mPTP) and led to voltage-dependent anion channel 1 (VDAC1)- and BCL2 antagonist/killer 1 (Bak) and Bak/BCL2-associated X (Bax)-dependent mtDNA leakage into the cytoplasm, while EV-A71 and FMDV infection induced mPTP opening and resulted in VDAC1-dependent mtDNA release. The released mtDNA bound to cGAS and activated cGAS-mediated antiviral immune response. cGAS was essential for inhibiting EV-A71, SVV, and FMDV replication by regulation of IFN-β production. cGAS deficiency contributed to higher mortality of EV-A71- or FMDV-infected mice. In addition, we found that SVV 2C protein was responsible for decreasing cGAS expression through the autophagy pathway. The 9th and 153rd amino acid sites in 2C were critical for induction of cGAS degradation. Furthermore, we also show that EV-A71, CA16, and EMCV 2C antagonize the cGAS-stimulator of interferon genes (STING) pathway through interaction with STING, and highly conserved amino acids Y155 and S156 were critical for this inhibitory effect. In conclusion, these data reveal novel mechanisms of picornaviruses to block the antiviral effect mediated by the cGAS-STING signaling pathway, which will provide insights for developing antiviral strategies against picornaviruses.

[1]  L. Laimins,et al.  Human papillomaviruses sensitize cells to DNA damage induced apoptosis by targeting the innate immune sensor cGAS , 2022, PLoS pathogens.

[2]  Vivek V. Thacker,et al.  The cGAS–STING pathway drives type I IFN immunopathology in COVID-19 , 2022, Nature.

[3]  C. Zheng,et al.  The crosstalk between viral RNA- and DNA-sensing mechanisms , 2021, Cellular and Molecular Life Sciences.

[4]  Hiroki Sato,et al.  Downregulation of mitochondrial biogenesis by virus infection triggers antiviral responses by cyclic GMP-AMP synthase , 2021, PLoS pathogens.

[5]  Yong Huang,et al.  PCV2 targets cGAS to inhibit type I interferon induction to promote other DNA virus infection , 2021, PLoS pathogens.

[6]  C. Zheng,et al.  Editorial: Sensing DNA in Antiviral Innate Immunity , 2021, Frontiers in Immunology.

[7]  QUAN LIU,et al.  SARS-CoV-2 Membrane Protein Inhibits Type I Interferon Production Through Ubiquitin-Mediated Degradation of TBK1 , 2021, Frontiers in Immunology.

[8]  Haiping Chen,et al.  MiR-103/miR-107 inhibits enterovirus 71 replication and facilitates type I interferon response by regulating SOCS3/STAT3 pathway , 2021, Biotechnology Letters.

[9]  Huanchun Chen,et al.  Selective autophagy receptor SQSTM1/ p62 inhibits Seneca Valley virus replication by targeting viral VP1 and VP3 , 2021, Autophagy.

[10]  Tianlong Liu,et al.  Mitochondrial dysfunction and mitophagy pathway activation in hepatitis E virus-infected livers of Mongolian gerbils. , 2021, Virus research.

[11]  Zixiang Zhu,et al.  Foot-and-Mouth Disease Virus Inhibits RIP2 Protein Expression to Promote Viral Replication , 2021, Virologica Sinica.

[12]  Juan Du,et al.  The Structure, Function, and Mechanisms of Action of Enterovirus Non-structural Protein 2C , 2020, Frontiers in Microbiology.

[13]  Susu He,et al.  Enterovirus 2C Protein Suppresses IKKα Phosphorylation by Recruiting IKKβ and IKKα into Viral Inclusion Bodies. , 2020, Viral immunology.

[14]  Christopher J. Tonkin,et al.  TDP-43 Triggers Mitochondrial DNA Release via mPTP to Activate cGAS/STING in ALS , 2020, Cell.

[15]  S. Aguirre,et al.  Chikungunya virus antagonizes cGAS-STING mediated type-I interferon responses by degrading cGAS , 2020, PLoS pathogens.

[16]  N. Melamed-Book,et al.  Mitochondrial Targeting of the Enteropathogenic Escherichia coli Map Triggers Calcium Mobilization, ADAM10-MAP Kinase Signaling, and Host Cell Apoptosis , 2020, mBio.

[17]  Zixiang Zhu,et al.  Innate immune evasion by picornaviruses. , 2020, European journal of immunology.

[18]  Navid B. Saleh,et al.  Hydroxyl functionalized multi-walled carbon nanotubes modulate immune responses without increasing 2009 pandemic influenza A/H1N1 virus titers in infected mice. , 2020, Toxicology and Applied Pharmacology.

[19]  J. Cui,et al.  HFE inhibits type I IFNs signaling by targeting the SQSTM1-mediated MAVS autophagic degradation , 2020, Autophagy.

[20]  C. Zheng.  Protein Dynamics in Cytosolic DNA-Sensing Antiviral Innate Immune Signaling Pathways , 2020, Frontiers in Immunology.

[21]  D. Komander,et al.  Dissecting distinct proteolytic activities of FMDV Lpro implicates cleavage and degradation of RLR signaling proteins, not its deISGylase/DUB activity, in type I interferon suppression , 2020, PLoS pathogens.

[22]  Zixiang Zhu,et al.  Seneca Valley Virus 3Cpro Cleaves PABPC1 to Promote Viral Replication , 2020, Pathogens.

[23]  E. Shekhova.  Mitochondrial reactive oxygen species as major effectors of antimicrobial immunity , 2020, PLoS pathogens.

[24]  Geng-fu Xiao,et al.  SFTSV Infection Induces BAK/BAX-Dependent Mitochondrial DNA Release to Trigger NLRP3 Inflammasome Activation. , 2020, Cell reports.

[25]  Y. Kook,et al.  Mycobacterium abscessus infection leads to enhanced production of type 1 interferon and NLRP3 inflammasome activation in murine macrophages via mitochondrial oxidative stress , 2020, PLoS pathogens.

[26]  J. Ajani,et al.  An improved strategy for CRISPR/Cas9 gene knockout and subsequent wildtype and mutant gene rescue , 2020, PloS one.

[27]  Huanchun Chen,et al.  Influenza A virus protein PB1-F2 impairs innate immunity by inducing mitophagy , 2020, Autophagy.

[28]  D. Lombard,et al.  Sirtuin 1 regulates mitochondrial function and immune homeostasis in respiratory syncytial virus infected dendritic cells , 2020, PLoS pathogens.

[29]  Zixiang Zhu,et al.  Genetic Determinants of Altered Virulence of Type O Foot-and-Mouth Disease Virus , 2020, Journal of Virology.

[30]  R. Klausen,et al.  Selective , 2020, Encyclopedia of the UN Sustainable Development Goals.

[31]  W. Sha,et al.  Targeting cyclophilin-D by miR-1281 protects human macrophages from Mycobacterium tuberculosis-induced programmed necrosis and apoptosis , 2019, Aging.

[32]  Wu Tong,et al.  BST2 suppresses porcine epidemic diarrhea virus replication by targeting and degrading virus nucleocapsid protein with selective autophagy , 2019, Autophagy.

[33]  V. Shoshan-Barmatz,et al.  VDAC oligomers form mitochondrial pores to release mtDNA fragments and promote lupus-like disease , 2019, Science.

[34]  S. Hur,et al.  Regulation of cGAS- and RLR-mediated immunity to nucleic acids , 2019, Nature Immunology.

[35]  N. Ben-Tal,et al.  ConSurf‐DB: An accessible repository for the evolutionary conservation patterns of the majority of PDB proteins , 2019, Protein science : a publication of the Protein Society.

[36]  Jianguo Wu,et al.  EV71 infection induces neurodegeneration via activating TLR7 signaling and IL-6 production , 2019, PLoS pathogens.

[37]  T. Ichinohe,et al.  Influenza A virus M2 protein triggers mitochondrial DNA-mediated antiviral immune responses , 2019, Nature Communications.

[38]  Y. Qian,et al.  Avian oncogenic herpesvirus antagonizes the cGAS-STING DNA-sensing pathway to mediate immune evasion , 2019, PLoS pathogens.

[39]  Huanchun Chen,et al.  Seneca Valley virus 2C and 3C inhibit type I interferon production by inducing the degradation of RIG-I. , 2019, Virology.

[40]  Shanshan Zhu,et al.  Seneca valley virus activates autophagy through the PERK and ATF6 UPR pathways. , 2019, Virology.

[41]  M. H. Fernandes,et al.  Persistent Infection and Transmission of Senecavirus A from Carrier Sows to Contact Piglets , 2019, Journal of Virology.

[42]  A. Oberst,et al.  STING is required for host defense against neuropathological West Nile virus infection , 2019, PLoS pathogens.

[43]  P. Fisher,et al.  Mitochondria in Health and Disease , 2019, Cells.

[44]  Huanchun Chen,et al.  Seneca Valley Virus 2C and 3Cpro Induce Apoptosis via Mitochondrion-Mediated Intrinsic Pathway , 2019, Front. Microbiol..

[45]  D. Walsh,et al.  mTOR Dysregulation by Vaccinia Virus F17 Controls Multiple Processes with Varying Roles in Infection , 2019, Journal of Virology.

[46]  M. Gale,et al.  Interleukin-1β Induces mtDNA Release to Activate Innate Immune Signaling via cGAS-STING. , 2019, Molecular cell.

[47]  Zhengfan Jiang,et al.  Apoptotic Caspases Suppress Type I Interferon Production via the Cleavage of cGAS, MAVS, and IRF3. , 2019, Molecular cell.

[48]  Zixiang Zhu,et al.  Foot-and-Mouth Disease Virus Antagonizes NOD2-Mediated Antiviral Effects by Inhibiting NOD2 Protein Expression , 2019, Journal of Virology.

[49]  Zhijian J. Chen,et al.  Autophagy Induction via STING Trafficking Is a Primordial Function of the cGAS Pathway , 2019, Nature.

[50]  P. Jiang,et al.  Encephalomyocarditis virus 2C protein antagonizes interferon‐&bgr; signaling pathway through interaction with MDA5 , 2019, Antiviral research.

[51]  Zixiang Zhu,et al.  Seneca Valley Virus 3C protease negatively regulates the type I interferon pathway by acting as a viral deubiquitinase , 2018, Antiviral Research.

[52]  B. Damania,et al.  cGAS and STING: At the intersection of DNA and RNA virus-sensing networks , 2018, PLoS pathogens.

[53]  J. Cui,et al.  Zika virus elicits inflammation to evade antiviral response by cleaving cGAS via NS1‐caspase‐1 axis , 2018, The EMBO journal.

[54]  Q. Ding,et al.  Species-specific disruption of STING-dependent antiviral cellular defenses by the Zika virus NS2B3 protease , 2018, Proceedings of the National Academy of Sciences.

[55]  Wenjie Liu,et al.  Foot‐and‐mouth disease virus nonstructural protein 2B interacts with cyclophilin A, modulating virus replication , 2018, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[56]  Zixiang Zhu,et al.  Seneca Valley Virus 3Cpro abrogates the IRF3- and IRF7-mediated innate immune response by degrading IRF3 and IRF7. , 2018, Virology.

[57]  S. Whelan,et al.  STING-dependent translation inhibition restricts RNA virus replication , 2018, Proceedings of the National Academy of Sciences.

[58]  Zixiang Zhu,et al.  Immunogenicity and protective efficacy of an inactivated cell culture-derived Seneca Valley virus vaccine in pigs. , 2018, Vaccine.

[59]  Mingzhou Chen,et al.  Picornavirus 2A protease regulates stress granule formation to facilitate viral translation , 2018, PLoS pathogens.

[60]  L. Daley-Bauer,et al.  Growing Murine Bone Marrow-Derived Macrophages. , 2018, Methods in molecular biology.

[61]  Jing Cheng,et al.  Suppression of Stim1 reduced intracellular calcium concentration and attenuated hypoxia/reoxygenation induced apoptosis in H9C2 cells , 2017, Bioscience reports.

[62]  P. Zhang,et al.  Involvement of inducible nitric oxide synthase and mitochondrial dysfunction in the pathogenesis of enterovirus 71 infection , 2017, Oncotarget.

[63]  K. Zhang,et al.  Emergence of novel Seneca Valley virus strains in China, 2017 , 2017, Transboundary and emerging diseases.

[64]  Y. Qiu,et al.  Human Virus‐Derived Small RNAs Can Confer Antiviral Immunity in Mammals , 2017, Immunity.

[65]  E. Ooi,et al.  Dengue virus activates cGAS through the release of mitochondrial DNA , 2017, Scientific Reports.

[66]  Huanchun Chen,et al.  Seneca Valley Virus Suppresses Host Type I Interferon Production by Targeting Adaptor Proteins MAVS, TRIF, and TANK for Cleavage , 2017, Journal of Virology.

[67]  Bing Yang,et al.  YAP antagonizes innate antiviral immunity and is targeted for lysosomal degradation through IKKɛ-mediated phosphorylation , 2017, Nature Immunology.

[68]  Meitian Wang,et al.  Crystal structure of 2C helicase from enterovirus 71 , 2017, Science Advances.

[69]  S. Aguirre,et al.  Dengue virus NS2B protein targets cGAS for degradation and prevents mitochondrial DNA sensing during infection , 2017, Nature Microbiology.

[70]  Dan Li,et al.  Hepatitis B virus X protein sensitizes HL-7702 cells to oxidative stress-induced apoptosis through modulation of the mitochondrial permeability transition pore , 2016, Oncology reports.

[71]  V. Hornung,et al.  cGAS-Mediated Innate Immunity Spreads Intercellularly through HIV-1 Env-Induced Membrane Fusion Sites. , 2016, Cell host & microbe.

[72]  J. Cui,et al.  TRIM14 Inhibits cGAS Degradation Mediated by Selective Autophagy Receptor p62 to Promote Innate Immune Responses. , 2016, Molecular cell.

[73]  Huizhi Guo,et al.  Biological function of Foot-and-mouth disease virus non-structural proteins and non-coding elements , 2016, Virology Journal.

[74]  T. de los Santos,et al.  The Pathogenesis of Foot-and-Mouth Disease in Pigs , 2016, Front. Vet. Sci..

[75]  R. Xavier,et al.  Trained immunity: A program of innate immune memory in health and disease , 2016, Science.

[76]  Zheng Xing,et al.  Differential Regulation of TLR Signaling on the Induction of Antiviral Interferons in Human Intestinal Epithelial Cells Infected with Enterovirus 71 , 2016, PloS one.

[77]  Zixiang Zhu,et al.  The VP3 structural protein of foot‐and‐mouth disease virus inhibits the IFN‐β signaling pathway , 2016, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[78]  Elizabeth E Gray,et al.  DNA tumor virus oncogenes antagonize the cGAS-STING DNA-sensing pathway , 2015, Science.

[79]  Zixiang Zhu,et al.  Multifunctional roles of leader protein of foot-and-mouth disease viruses in suppressing host antiviral responses , 2015, Veterinary Research.

[80]  Leiliang Zhang,et al.  Enterovirus 71 2C Protein Inhibits NF-κB Activation by Binding to RelA(p65) , 2015, Scientific Reports.

[81]  Wenhua Li,et al.  Coxsackievirus A16 Elicits Incomplete Autophagy Involving the mTOR and ERK Pathways , 2015, PloS one.

[82]  N. Grishin,et al.  Phosphorylation of innate immune adaptor proteins MAVS, STING, and TRIF induces IRF3 activation , 2015, Science.

[83]  T. Taniguchi,et al.  Apoptotic Caspases Prevent the Induction of Type I Interferons by Mitochondrial DNA , 2014, Cell.

[84]  Y. Liu,et al.  Picornavirus Morphogenesis , 2014, Microbiology and Molecular Reviews.

[85]  Zheng Xing,et al.  Robust antiviral responses to enterovirus 71 infection in human intestinal epithelial cells. , 2013, Virus research.

[86]  E. Cheng,et al.  Bax and Bak function as the outer membrane component of the mitochondrial permeability pore in regulating necrotic cell death in mice , 2013, eLife.

[87]  Xiang-tao Liu,et al.  Engineering Foot-and-Mouth Disease Viruses with Improved Growth Properties for Vaccine Development , 2013, PloS one.

[88]  A. Basu,et al.  STING Mediates Neuronal Innate Immune Response Following Japanese Encephalitis Virus Infection , 2012, Scientific Reports.

[89]  D. Green,et al.  Mitochondria and cell death: outer membrane permeabilization and beyond , 2010, Nature Reviews Molecular Cell Biology.

[90]  Q. Jin,et al.  The 3C Protein of Enterovirus 71 Inhibits Retinoid Acid-Inducible Gene I-Mediated Interferon Regulatory Factor 3 Activation and Type I Interferon Responses , 2010, Journal of Virology.

[91]  Yang Zhang,et al.  I-TASSER: a unified platform for automated protein structure and function prediction , 2010, Nature Protocols.

[92]  M. Montero,et al.  Mitochondrial free [Ca2+] levels and the permeability transition. , 2009, Cell calcium.

[93]  Thomas D. Schmittgen,et al.  Analyzing real-time PCR data by the comparative CT method , 2008, Nature Protocols.

[94]  T. Kadono,et al.  Na+/H+ exchanger inhibitor cariporide attenuates the mitochondrial Ca2+ overload and PTP opening. , 2007, American journal of physiology. Heart and circulatory physiology.

[95]  E. Chávez,et al.  Mitochondrial DNA fragments released through the permeability transition pore correspond to specific gene size. , 2007, Life sciences.