Title: A Balancing Act: MDA5 in Antiviral Immunity and Autoinflammation
Abstract: MDA5 is a pattern-recognition receptor for RNA and induces a type I interferon response. MDA5 is activated in a variety of clinically relevant settings. This includes infection with ssRNA, dsRNA, and dsDNA viruses; several autoimmune and autoinflammatory diseases, such as type 1 diabetes and Aicardi–Goutières syndrome; and some forms of cancer treatment. Synthetic, viral, and cellular RNAs can all activate MDA5. The latter may include transcripts from endogenous retroelements such as Alu repeats. Length and secondary structure are important features that determine whether an RNA molecule is detected by MDA5. Indeed, long, base-paired RNA molecules potently activate MDA5 in the test tube. Induction of interferons during viral infection is mediated by cellular proteins that recognise viral nucleic acids. MDA5 is one such sensor of virus presence and is activated by RNA. MDA5 is required for immunity against several classes of viruses, including picornaviruses. Recent work showed that mutations in the IFIH1 gene, encoding MDA5, lead to interferon-driven autoinflammatory diseases. Together with observations made in cancer cells, this suggests that MDA5 detects cellular RNAs in addition to viral RNAs. It is therefore important to understand the properties of the RNAs which activate MDA5. New data indicate that RNA length and secondary structure are features sensed by MDA5. We review these developments and discuss how MDA5 strikes a balance between antiviral immunity and autoinflammation. Induction of interferons during viral infection is mediated by cellular proteins that recognise viral nucleic acids. MDA5 is one such sensor of virus presence and is activated by RNA. MDA5 is required for immunity against several classes of viruses, including picornaviruses. Recent work showed that mutations in the IFIH1 gene, encoding MDA5, lead to interferon-driven autoinflammatory diseases. Together with observations made in cancer cells, this suggests that MDA5 detects cellular RNAs in addition to viral RNAs. It is therefore important to understand the properties of the RNAs which activate MDA5. New data indicate that RNA length and secondary structure are features sensed by MDA5. We review these developments and discuss how MDA5 strikes a balance between antiviral immunity and autoinflammation. Mammalian cells use pattern-recognition receptors (PRRs) to detect the presence of infectious microorganisms [1Medzhitov R. Approaching the asymptote: 20 years later.Immunity. 2009; 30: 766-775Abstract Full Text Full Text PDF PubMed Scopus (201) Google Scholar]. These receptors are activated by pathogen-associated molecular patterns (PAMPs). In the case of virus infection, PAMPs are often nucleic acids [2Barrat F.J. et al.Importance of nucleic acid recognition in inflammation and autoimmunity.Annu. Rev. Med. 2016; 67: 323-336Crossref PubMed Google Scholar, 3Hartmann G. Nucleic acid immunity.Adv. Immunol. 2017; 133: 121-169Crossref PubMed Scopus (18) Google Scholar]. For example, viral RNAs trigger PRRs, including the endosomal toll-like receptors (TLRs) 3 and 7 and the cytosolic RIG-I-like receptors (RLRs) [2Barrat F.J. et al.Importance of nucleic acid recognition in inflammation and autoimmunity.Annu. Rev. Med. 2016; 67: 323-336Crossref PubMed Google Scholar, 3Hartmann G. Nucleic acid immunity.Adv. Immunol. 2017; 133: 121-169Crossref PubMed Scopus (18) Google Scholar]. The RLR family comprises three members: retinoic acid inducible gene I (RIG-I), melanoma differentiation-associated gene 5 (MDA5), and laboratory of genetics and physiology 2 (LGP2) [2Barrat F.J. et al.Importance of nucleic acid recognition in inflammation and autoimmunity.Annu. Rev. Med. 2016; 67: 323-336Crossref PubMed Google Scholar, 3Hartmann G. Nucleic acid immunity.Adv. Immunol. 2017; 133: 121-169Crossref PubMed Scopus (18) Google Scholar]. These proteins are ubiquitously expressed at low levels. All RLRs contain a DExD/H-box RNA helicase domain and a C terminal domain (CTD), both responsible for RNA binding [4Goubau D. et al.Cytosolic sensing of viruses.Immunity. 2013; 38: 855-869Abstract Full Text Full Text PDF PubMed Scopus (321) Google Scholar]. In addition, RIG-I and MDA5 have two N terminal caspase activation and recruitment domains (CARDs). Upon activation of RLRs by RNA binding, the CARDs interact with the adaptor mitochondrial antiviral signalling protein (MAVS), ultimately leading to the transcription of the genes encoding type I interferons (IFNs) (see Glossary) [2Barrat F.J. et al.Importance of nucleic acid recognition in inflammation and autoimmunity.Annu. Rev. Med. 2016; 67: 323-336Crossref PubMed Google Scholar, 3Hartmann G. Nucleic acid immunity.Adv. Immunol. 2017; 133: 121-169Crossref PubMed Scopus (18) Google Scholar, 4Goubau D. et al.Cytosolic sensing of viruses.Immunity. 2013; 38: 855-869Abstract Full Text Full Text PDF PubMed Scopus (321) Google Scholar] (Figure 1). Autocrine and paracrine IFN stimulation subsequently induces transcription of hundreds of IFN-stimulated genes (ISGs), several of which encode proteins with direct antiviral functions [5Yan N. Chen Z.J. Intrinsic antiviral immunity.Nat. Immunol. 2012; 13: 214-222Crossref PubMed Scopus (216) Google Scholar]. RLRs themselves are encoded by ISGs, constituting a feed-forward loop. Type I IFNs further coordinate cellular immune responses to virus infection and are thus essential for antiviral immunity [6McNab F. et al.Type I interferons in infectious disease.Nat. Rev. Immunol. 2015; 15: 87-103Crossref PubMed Scopus (400) Google Scholar]. In addition to inducing type I IFNs, RLRs and MAVS also activate apoptosis, leading to the elimination of the infected cell [7Orzalli M.H. Kagan J.C. Apoptosis and necroptosis as host defense strategies to prevent viral infection.Trends Cell Biol. 2017; 27: 800-809Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar]. Given the abundance of nucleic acids in healthy cells, a key question is to understand the mechanisms by which nucleic acid-sensing PRRs become activated specifically following virus infection. The features of RIG-I-stimulatory RNAs are well understood and include the presence of two or three phosphate groups at the 5′-end, the absence of 5′-cap methylation, and base-pairing adjacent to the 5′-end [8Goubau D. et al.Antiviral immunity via RIG-I-mediated recognition of RNA bearing 5′-diphosphates.Nature. 2014; 514: 372-375Crossref PubMed Scopus (183) Google Scholar, 9Rehwinkel J. et al.RIG-I detects viral genomic RNA during negative-strand RNA virus infection.Cell. 2010; 140: 397-408Abstract Full Text Full Text PDF PubMed Scopus (321) Google Scholar, 10Pichlmair A. et al.RIG-I-mediated antiviral responses to single-stranded RNA bearing 5′-phosphates.Science. 2006; 314: 997-1001Crossref PubMed Scopus (1284) Google Scholar, 11Hornung V. et al.5′-Triphosphate RNA is the ligand for RIG-I.Science. 2006; 314: 994-997Crossref PubMed Scopus (1430) Google Scholar, 12Baum A. et al.Preference of RIG-I for short viral RNA molecules in infected cells revealed by next-generation sequencing.Proc. 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These features are characteristic of viral RNAs produced by some viruses, such as influenza A virus or reovirus [8Goubau D. et al.Antiviral immunity via RIG-I-mediated recognition of RNA bearing 5′-diphosphates.Nature. 2014; 514: 372-375Crossref PubMed Scopus (183) Google Scholar, 9Rehwinkel J. et al.RIG-I detects viral genomic RNA during negative-strand RNA virus infection.Cell. 2010; 140: 397-408Abstract Full Text Full Text PDF PubMed Scopus (321) Google Scholar, 12Baum A. et al.Preference of RIG-I for short viral RNA molecules in infected cells revealed by next-generation sequencing.Proc. Natl. Acad. Sci. U. S. A. 2010; 107: 16303-16308Crossref PubMed Scopus (218) Google Scholar], but are not typically found in cellular RNAs present in the cytosol of healthy cells, explaining selective activation of RIG-I in virus-infected cells. It is also possible that, in some settings, mislocalised cellular RNAs activate RIG-I and other nucleic acid sensors of the innate immune system [16Chiang J.J. et al.Viral unmasking of cellular 5S rRNA pseudogene transcripts induces RIG-I-mediated immunity.Nat. Immunol. 2018; 19: 53-62Crossref PubMed Scopus (10) Google Scholar, 17Dhir A. et al.Mitochondrial double-stranded RNA triggers antiviral signalling in humans.Nature. 2018; 560: 238-242Crossref PubMed Scopus (0) Google Scholar]. Although RIG-I and MDA5 share similar domains, they detect different viral infections [18Kato H. et al.Differential roles of MDA5 and RIG-I helicases in the recognition of RNA viruses.Nature. 2006; 441: 101-105Crossref PubMed Scopus (2089) Google Scholar]. For example, RIG-I detects infection with orthomyxoviruses, such as influenza A virus, while MDA5 senses picornavirus infection. In contrast to RIG-I, the mechanisms that allow MDA5 to recognise viral RNAs while avoiding cellular RNAs are less well understood. Biochemical and structural work using recombinant MDA5 protein demonstrated that MDA5 senses RNA length and secondary structure [19Peisley A. et al.Cooperative assembly and dynamic disassembly of MDA5 filaments for viral dsRNA recognition.Proc. Natl. Acad. Sci. U. S. A. 2011; 108: 21010-21015Crossref PubMed Scopus (118) Google Scholar, 20Peisley A. et al.Kinetic mechanism for viral dsRNA length discrimination by MDA5 filaments.Proc. Natl. Acad. Sci. U. S. A. 2012; 109: E3340-E3349Crossref PubMed Scopus (0) Google Scholar, 21Wu B. et al.Structural basis for dsRNA recognition, filament formation, and antiviral signal activation by MDA5.Cell. 2013; 152: 276-289Abstract Full Text Full Text PDF PubMed Scopus (211) Google Scholar]. Understanding molecular features of MDA5-stimulatory RNAs is important for another reason: in addition to its protective role in antiviral defence, MDA5 has been implicated in autoimmune and autoinflammatory diseases such as type 1 diabetes (T1D), systemic lupus erythematosus (SLE) and Aicardi–Goutières syndrome (AGS) [2Barrat F.J. et al.Importance of nucleic acid recognition in inflammation and autoimmunity.Annu. Rev. Med. 2016; 67: 323-336Crossref PubMed Google Scholar, 3Hartmann G. Nucleic acid immunity.Adv. Immunol. 2017; 133: 121-169Crossref PubMed Scopus (18) Google Scholar, 22Crow Y.J. Manel N. Aicardi–Goutieres syndrome and the type I interferonopathies.Nat. Rev. Immunol. 2015; 15: 429-440Crossref PubMed Scopus (221) Google Scholar]. Moreover, new data indicate that MDA5 is activated during some forms of cancer treatment [23Roulois D. et al.DNA-demethylating agents target colorectal cancer cells by inducing viral mimicry by endogenous transcripts.Cell. 2015; 162: 961-973Abstract Full Text Full Text PDF PubMed Scopus (235) Google Scholar, 24Chiappinelli K.B. et al.Inhibiting DNA. methylation causes an interferon response in cancer via dsRNA including endogenous retroviruses.Cell. 2015; 162: 974-986Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar] or in settings where mitochondrial RNA degradation is compromised [17Dhir A. et al.Mitochondrial double-stranded RNA triggers antiviral signalling in humans.Nature. 2018; 560: 238-242Crossref PubMed Scopus (0) Google Scholar]. These observations highlight that, in some circumstances, cellular RNAs trigger MDA5. Indeed, transcripts from repetitive genome segments, including endogenous retroelements, such as Alu elements, and mitochondrial RNA have been suggested to bind and activate MDA5 [17Dhir A. et al.Mitochondrial double-stranded RNA triggers antiviral signalling in humans.Nature. 2018; 560: 238-242Crossref PubMed Scopus (0) Google Scholar, 23Roulois D. et al.DNA-demethylating agents target colorectal cancer cells by inducing viral mimicry by endogenous transcripts.Cell. 2015; 162: 961-973Abstract Full Text Full Text PDF PubMed Scopus (235) Google Scholar, 24Chiappinelli K.B. et al.Inhibiting DNA. methylation causes an interferon response in cancer via dsRNA including endogenous retroviruses.Cell. 2015; 162: 974-986Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 25Ahmad S. et al.Breaching self-tolerance to Alu duplex RNA underlies MDA5-mediated inflammation.Cell. 2018; 172 (e13): 797-810Abstract Full Text Full Text PDF PubMed Scopus (11) Google Scholar, 26Zhao K. et al.LINE1 contributes to autoimmunity through both RIG-I- and MDA5-mediated RNA sensing pathways.J. Autoimmun. 2018; 90: 105-115Crossref PubMed Scopus (1) Google Scholar]. Here, we review RNA sensing by MDA5 in the context of antiviral immunity and autoinflammation. We discuss how important recent developments in this area set the stage for future exploration of MDA5-RNA interactions in living cell systems, including models of authentic virus infection and autoinflammation. MDA5 was first described in 2002 as a helicase protein in mouse and human cells [27Kovacsovics M. et al.Overexpression of Helicard, a CARD-containing helicase cleaved during apoptosis, accelerates DNA degradation.Curr. Biol. 2002; 12: 838-843Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 28Kang D.C. et al.mda-5: An interferon-inducible putative RNA helicase with double-stranded RNA-dependent ATPase activity and melanoma growth-suppressive properties.Proc. Natl. Acad. Sci. U. S. A. 2002; 99: 637-642Crossref PubMed Scopus (396) Google Scholar]. Interestingly, these initial reports suggested that MDA5 is involved in the execution of apoptosis. In 2004, Rick Randall's group found that overexpression of MDA5 alone induces the expression of IFN-β, a type I IFN [29Andrejeva J. et al.The V proteins of paramyxoviruses bind the IFN-inducible RNA helicase, mda-5, and inhibit its activation of the IFN-beta promoter.Proc. Natl. Acad. Sci. U. S. A. 2004; 101: 17264-17269Crossref PubMed Scopus (703) Google Scholar]. Furthermore, this study showed that the IFN-β response of cells stimulated by transfection of polyriboinosinic:polyribocytidylic acid (poly I:C), a synthetic RNA, is greatly enhanced by MDA5 overexpression [29Andrejeva J. et al.The V proteins of paramyxoviruses bind the IFN-inducible RNA helicase, mda-5, and inhibit its activation of the IFN-beta promoter.Proc. Natl. Acad. Sci. U. S. A. 2004; 101: 17264-17269Crossref PubMed Scopus (703) Google Scholar]. Shortly afterwards, knockouts of the mouse Ifih1 gene encoding MDA5 demonstrated that MDA5 is essential for the type I IFN response to poly I:C [18Kato H. et al.Differential roles of MDA5 and RIG-I helicases in the recognition of RNA viruses.Nature. 2006; 441: 101-105Crossref PubMed Scopus (2089) Google Scholar, 30Gitlin L. et al.Essential role of mda-5 in type I IFN responses to polyriboinosinic:polyribocytidylic acid and encephalomyocarditis picornavirus.Proc. Natl. Acad. Sci. U. S. A. 2006; 103: 8459-8464Crossref PubMed Scopus (749) Google Scholar]. Together, these landmark studies established MDA5 as an RNA sensor inducing type I IFN (Figure 1). Poly I:C is often used as a synthetic mimic of double-stranded RNA (dsRNA). This led to the hypothesis that the PAMP recognised by MDA5 is the double-stranded conformation of RNA. Indeed, structural studies of MDA5 show that the protein adopts a ring-like conformation around dsRNA [21Wu B. et al.Structural basis for dsRNA recognition, filament formation, and antiviral signal activation by MDA5.Cell. 2013; 152: 276-289Abstract Full Text Full Text PDF PubMed Scopus (211) Google Scholar]. Contacts between MDA5 and dsRNA are along the phosphodiester backbone, suggestive of sequence-nonspecific binding [21Wu B. et al.Structural basis for dsRNA recognition, filament formation, and antiviral signal activation by MDA5.Cell. 2013; 152: 276-289Abstract Full Text Full Text PDF PubMed Scopus (211) Google Scholar]. Experiments in the test tube using recombinant MDA5 and in vitro transcribed dsRNAs established that MDA5 forms filaments along dsRNA [19Peisley A. et al.Cooperative assembly and dynamic disassembly of MDA5 filaments for viral dsRNA recognition.Proc. Natl. Acad. Sci. U. S. A. 2011; 108: 21010-21015Crossref PubMed Scopus (118) Google Scholar, 31Berke I.C. Modis Y. MDA5 cooperatively forms dimers and ATP-sensitive filaments upon binding double-stranded RNA.EMBO J. 2012; 31: 1714-1726Crossref PubMed Scopus (0) Google Scholar]. These filaments are particularly stable on long dsRNA molecules and are mediated by both protein–RNA and protein–protein interactions [20Peisley A. et al.Kinetic mechanism for viral dsRNA length discrimination by MDA5 filaments.Proc. Natl. Acad. Sci. U. S. A. 2012; 109: E3340-E3349Crossref PubMed Scopus (0) Google Scholar, 21Wu B. et al.Structural basis for dsRNA recognition, filament formation, and antiviral signal activation by MDA5.Cell. 2013; 152: 276-289Abstract Full Text Full Text PDF PubMed Scopus (211) Google Scholar]. These biophysical studies used filament formation and ATP hydrolysis by the helicase domain of MDA5 as surrogates for its activation. Consistent with these data is the finding in cells that the IFN response to transfected poly I:C depends on length: long poly I:C molecules preferentially trigger MDA5, whereas short poly I:C activates RIG-I [32Kato H. et al.Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid-inducible gene-I and melanoma differentiation-associated gene 5.J. Exp. Med. 2008; 205: 1601-1610Crossref PubMed Scopus (819) Google Scholar]. Together, these observations have led to the widely held notion that MDA5 detects long dsRNA. However, other observations suggest that dsRNA – that is, two complementary RNA molecules annealed to form a helical A-form duplex – may not always be sufficient to explain activation of MDA5. For example, IFN induction is particularly strong in response to poly I:C, while other dsRNAs such as poly A:U, trigger no response [33Colby C. Chamberlin M.J. The specificity of interferon induction in chick embryo cells by helical RNA.Proc. Natl. Acad. Sci. U. S. A. 1969; 63: 160-167Crossref PubMed Google Scholar, 34Pichlmair A. et al.Activation of MDA5 requires higher-order RNA structures generated during virus infection.J. Virol. 2009; 83: 10761-10769Crossref PubMed Scopus (242) Google Scholar]. It should nevertheless be noted that, compared with G:C and I:C duplexes, A:U duplexes have a lower stability, which could explain their lower signalling activity. It is also noteworthy that poly I:C consists of annealed inosine and cytidine homopolymers. These are produced enzymatically from ribonucleoside diphosphates, using polynucleotide phosphorylase, and are heterogeneous in length [35Grunberg-Manago M. et al.Enzymic synthesis of polynucleotides. I. Polynucleotide phosphorylase of Azotobacter vinelandii. 1956.Biochim. Biophys. Acta. 1989; 1000: 65-81PubMed Google Scholar]. Annealing thus results in double-stranded regions with single-stranded overhangs. These are available for base-pairing with other molecules, potentially resulting in the formation of more complex, branched RNA structures, which have been proposed to play a role in MDA5 activation [34Pichlmair A. et al.Activation of MDA5 requires higher-order RNA structures generated during virus infection.J. Virol. 2009; 83: 10761-10769Crossref PubMed Scopus (242) Google Scholar]. Early studies in MDA5-deficient mice reported that these animals are highly susceptible to infection with encephalomyocarditis virus (EMCV), failing to induce type I IFNs [18Kato H. et al.Differential roles of MDA5 and RIG-I helicases in the recognition of RNA viruses.Nature. 2006; 441: 101-105Crossref PubMed Scopus (2089) Google Scholar, 30Gitlin L. et al.Essential role of mda-5 in type I IFN responses to polyriboinosinic:polyribocytidylic acid and encephalomyocarditis picornavirus.Proc. Natl. Acad. Sci. U. S. A. 2006; 103: 8459-8464Crossref PubMed Scopus (749) Google Scholar]. More recently, MDA5 deficiency in humans has been shown to increase susceptibility to viral infection [36Zaki M. et al.Recurrent and prolonged infections in a child with a homozygous IFIH1 nonsense mutation.Front. Genet. 2017; 8: 130Crossref PubMed Scopus (0) Google Scholar, 37Lamborn I.T. et al.Recurrent rhinovirus infections in a child with inherited MDA5 deficiency.J. Exp. Med. 2017; 214: 1949-1972Crossref PubMed Scopus (9) Google Scholar, 38Asgari S. et al.Severe viral respiratory infections in children with IFIH1 loss-of-function mutations.Proc. Natl. Acad. Sci. U. S. A. 2017; 114: 8342-8347Crossref PubMed Scopus (1) Google Scholar]. EMCV is a widely used model belonging to the Picornaviridae family that includes important human pathogens such as hepatitis A virus, coxsackie B virus, enterovirus, and rhinovirus. As summarised in Table 1, subsequent work by many laboratories revealed that MDA5 is involved in type I IFN induction during infection with several other types of viruses. This includes virus families characterised by genomes consisting of single-stranded (ss), positive- or negative-sense RNA, dsRNA, and dsDNA. However, in contrast to EMCV infection, in which MDA5 is essential for type I IFN induction, MDA5 plays a partial role in other infections (Table 1). For example, dsDNA viruses are also detected by the cytosolic DNA sensing pathway, and some RNA viruses trigger both MDA5 and RIG-I.Table 1Viral Infections Detected by MDA5Virus familyGenomeExamplesRole of MDA5 in type I IFN inductionSelected RefsPicornaviridaessRNA (+)Encephalomyocarditis virus; Rhinovirus; Coxsackie B virusEssential18Kato H. et al.Differential roles of MDA5 and RIG-I helicases in the recognition of RNA viruses.Nature. 2006; 441: 101-105Crossref PubMed Scopus (2089) Google Scholar, 30Gitlin L. et al.Essential role of mda-5 in type I IFN responses to polyriboinosinic:polyribocytidylic acid and encephalomyocarditis picornavirus.Proc. Natl. Acad. Sci. U. S. A. 2006; 103: 8459-8464Crossref PubMed Scopus (749) Google Scholar, 43Feng Q. et al.MDA5 detects the double-stranded RNA replicative form in picornavirus-infected cells.Cell Rep. 2012; 2: 1187-1196Abstract Full Text Full Text PDF PubMed Scopus (93) Google Scholar, 102Wang J.P. et al.MDA5 and MAVS mediate type I interferon responses to coxsackie B virus.J. Virol. 2010; 84: 254-260Crossref PubMed Scopus (0) Google Scholar, 103Slater L. et al.Co-ordinated role of TLR3, RIG-I and MDA5 in the innate response to rhinovirus in bronchial epithelium.PLoS Pathog. 2010; 6e1001178Crossref PubMed Scopus (158) Google ScholarFlaviviridaessRNA (+)West Nile virus; Hepatitis C virus; Zika virusPartial44Hertzog J. et al.Infection with a Brazilian isolate of Zika virus generates RIG-I stimulatory RNA and the viral NS5 protein blocks type I IFN induction and signaling.Eur. J. Immunol. 2018; 48: 1120-1136Crossref PubMed Scopus (4) Google Scholar, 104Cao X. et al.MDA5 plays a critical role in interferon response during hepatitis C virus infection.J. Hepatol. 2015; 62: 771-778Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar, 105Fredericksen B.L. et al.Establishment and maintenance of the innate antiviral response to West Nile virus involves both RIG-I and MDA5 signaling through IPS-1.J. Virol. 2008; 82: 609-616Crossref PubMed Scopus (0) Google ScholarTogaviridaessRNA (+)Sindbis virusPartial34Pichlmair A. et al.Activation of MDA5 requires higher-order RNA structures generated during virus infection.J. Virol. 2009; 83: 10761-10769Crossref PubMed Scopus (242) Google Scholar, 106Burke C.W. et al.Characteristics of alpha/beta interferon induction after infection of murine fibroblasts with wild-type and mutant alphaviruses.Virology. 2009; 395: 121-132Crossref PubMed Scopus (39) Google Scholar, 107Akhrymuk I. et al.Both RIG-I and MDA5 detect alphavirus replication in concentration-dependent mode.Virology. 2016; 487: 230-241Crossref PubMed Scopus (9) Google ScholarCoronaviridaessRNA (+)SARS coronavirusPartial57Zust R. et al.Ribose 2′-O-methylation provides a molecular signature for the distinction of self and non-self mRNA dependent on the RNA sensor Mda5.Nat. Immunol. 2011; 12: 137-143Crossref PubMed Scopus (0) Google Scholar, 108Zalinger Z.B. et al.MDA5 is critical to host defense during infection with murine coronavirus.J. Virol. 2015; 89: 12330-12340Crossref PubMed Scopus (8) Google Scholar, 109Li J. et al.Murine coronavirus induces type I interferon in oligodendrocytes through recognition by RIG-I and MDA5.J. Virol. 2010; 84: 6472-6482Crossref PubMed Scopus (0) Google ScholarParamyxoviridaessRNA (−)Measles virus; human Metapneumovirus; Sendai virusaSome Sendai virus stocks, particularly the Cantell strain, activate mostly RIG-I.Partial110Ikegame S. et al.Both RIG-I and MDA5 RNA helicases contribute to the induction of alpha/beta interferon in measles virus-infected human cells.J. Virol. 2010; 84: 372-379Crossref PubMed Scopus (76) Google Scholar, 111Gitlin L. et al.Melanoma differentiation-associated gene 5 (MDA5) is involved in the innate immune response to Paramyxoviridae infection in vivo.PLoS Pathog. 2010; 6e1000734Crossref PubMed Scopus (0) Google Scholar, 112Banos-Lara Mdel R. et al.Critical role of MDA5 in the interferon response induced by human metapneumovirus infection in dendritic cells and in vivo.J. Virol. 2013; 87: 1242-1251Crossref PubMed Scopus (0) Google ScholarReoviridaedsRNARotavirusPartial8Goubau D. et al.Antiviral immunity via RIG-I-mediated recognition of RNA bearing 5′-diphosphates.Nature. 2014; 514: 372-375Crossref PubMed Scopus (183) Google Scholar, 32Kato H. et al.Length-dependent recognition of double-stranded ribonucleic acids by retinoic acid-inducible gene-I and melanoma differentiation-associated gene 5.J. Exp. Med. 2008; 205: 1601-1610Crossref PubMed Scopus (819) Google Scholar, 113Dou Y. et al.The innate immune receptor MDA5 limits rotavirus infection but promotes cell death and pancreatic inflammation.EMBO J. 2017; 36 (274–2757)Crossref Scopus (1) Google ScholarPoxviridaedsDNAVaccinia virusPartial34Pichlmair A. et al.Activation of MDA5 requires higher-order RNA structures generated during virus infection.J. Virol. 2009; 83: 10761-10769Crossref PubMed Scopus (242) Google Scholar, 114Pham A.M. et al.PKR transduces MDA5-dependent signals for type I IFN induction.PLoS Pathog. 2016; 12e1005489Crossref PubMed Scopus (24) Google ScholarHerpesviridaedsDNAHerpes simplex virus 1Partial115Melchjorsen J. et al.Early innate recognition of herpes simplex virus in human primary macrophages is mediated via the MDA5/MAVS-dependent and MDA5/MAVS/RNA polymerase III-independent pathways.J. Virol. 2010; 84: 11350-11358Crossref PubMed Scopus (0) Google ScholarHepadnaviridaedsDNAHepatitis B virusbHepatitis D virus, a satellite virus that only infects HBV-infected cells and has a circular, ssRNA(−) genome, is also sensed by MDA5 [117].Partial116Lu H.L. Liao F. Melanoma differentiation-associated gene 5 senses hepatitis B virus and activates innate immune signaling to suppress virus replication.J. Immunol. 2013; 191: 3264-3276Crossref PubMed Scopus (0) Google Scholara Some Sendai virus stocks, particularly the Cantell strain, activate mostly RIG-I.b Hepatitis D virus, a satellite virus that only infects HBV-infected cells and has a circular, ssRNA(−) genome, is also sensed by MDA5 117Zhang Z. et al.Hepatitis D virus replication is sensed by MDA5 and induces IFN-beta/lambda responses in hepatocytes.J. Hepatol. 2018; 69: 25-35Abstract Full Text Full Text PDF PubMed Scopus (0) Google Scholar. Open table in a new tab Immunofluorescence experiments using monoclonal antibodies, called J2 and K1, that recognise dsRNA [39Schonborn J. et al.Monoclonal antibodies to double-stranded RNA as probes of RNA structure in crude nucleic acid extracts.Nucleic Acids Res. 1991; 19: 2993-3000Crossref PubMed Google Scholar] revealed strong staining in cells infected with many of the viruses that activate MDA5 [34Pichlmair A. et al.Activation of MDA5 requires higher-order RNA structures generated during virus infection.J. Virol. 2009; 83: 10761-10769Crossref PubMed Scopus (242) Google Scholar, 40Weber F. et al.Double-stranded RNA is produced by positive-strand RNA viruses and DNA viruses but not in detectable amounts by negative-strand RNA viruses.J. Virol. 2006; 80: 5059-5064Crossref PubMed Scopus (486) Google Scholar, 41Triantafilou K. et al.Visualisation of direct interaction of MDA5 and the dsRNA replicative intermediate form of positive strand RNA viruses.J. 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