Innate immune sensing of coronavirus and viral evasion strategies

Coronavirus

Coronaviruses belong to the Orthocoronavirinae subfamily of the Coronaviridae family. Viruses in the Orthocoronavirinae subfamily can be further classified into four genera: Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus². Among these viruses, seven strains of alpha- and betacoronaviruses are known to cause human illness. HCoV-229E, HCoV-OC43, HCoV-NL63, and HCoV-HKU1 cause mild upper respiratory symptoms that are typically recognized as the seasonal common cold². In contrast, the other three strains, SARS-CoV, MERS-CoV, and SARS-CoV-2, which are transmitted through zoonotic transmission, can lead to severe respiratory symptoms with unique pathogenesis, including severe lymphopenia and extensive pneumonia caused by aberrant host antiviral responses³.

Coronaviruses are enveloped positive-sense RNA viruses that possess the largest viral RNA genome, ~27–32 kb in size. Upon coronavirus infection, the virus life cycle initiates with the specific binding of virus spike (S) protein to the target cell surface receptors, causing viral fusion into the target cell⁴. During the intracellular viral life cycle, the genomic RNA of coronavirus is uncoated from the nucleocapsid (N) protein, leading to the translation of two open reading frames (ORFs), ORF1a and ORF1b. The translated ORFs produce two large polyproteins, pp1a and pp1ab, which are further cleaved by viral proteases encoded by the Nsp3 (papain-like protease) and Nsp5 (3C-like protease) genes to generate functional nonstructural proteins (Nsp1-Nsp16)⁵. The viral replication and transcription complex (RTC) formed by Nsp2-Nsp16 further promotes viral genomic RNA replication and subgenomic mRNA transcription. Among these proteins, Nsp3, Nsp4, and Nsp6 are involved in double-membrane vesicle (DMV) formation, together with two other viral replication organelles, namely, convoluted membranes (CMs) and small open double-membrane spherules (DMSs), providing a protective microenvironment for the replication of viral genomic RNA and transcription of subgenomic mRNAs⁶. Nsp7 and Nsp8 are two cofactors of RNA-dependent RNA polymerase (RdRp) that reside in Nsp12⁷, and together with RNA-modifying enzymes residing in Nsp13-Nsp16, they promote viral RNA synthesis and modification⁸. Notably, a 3′–5′ exonuclease residing in Nsp14 has an RNA proofreading function during RNA synthesis⁹. The RNA capping machinery during the viral life cycle is formed by Nsp10 (cofactor), Nsp13 (RNA 5′ triphosphatase activity), Nsp14 (N7-methyltransferase activity), and Nsp16 (2′-O-methyltransferase activity), while the mechanism remains to be elucidated^10–12. Moreover, additional accessory proteins, including ORF3, 6, 7, 8, 9, and 10, with various host-modulatory functions are also necessary for completion of the viral life cycle¹³. The exact number, location, and size of accessory proteins vary in different coronaviruses, and their specific functions remain to be further elucidated. Structural proteins such as S, envelope (E), membrane (M), and N, translated from subgenomic mRNAs, translocate from the endoplasmic reticulum (ER) to the ER-to-Golgi intermediate compartment (ERGIC) and assemble with newly generated genomic RNA encapsidated by the N protein. Mature virions are released by the host exocytosis pathway to target the next host cell¹⁴.

Host innate immune sensors

Immediate cellular responses to pathogen invasion are crucial for maintaining cell homeostasis and survival for all living organisms. Host responses are triggered by germline-encoded cellular receptors, known as ‘pattern recognition receptors’ (PRRs), that recognize specific patterns of ‘non-self’ and ‘danger’ molecules, termed ‘pathogen-associated molecular patterns’ (PAMPs) and ‘danger-associated molecular patterns’ (DAMPs). In mammals, activation of PRRs by PAMPs or DAMPs triggers innate immune responses and produces multiple IFNs and proinflammatory cytokines. In recent decades, various PRRs, such as Toll-like receptors (TLRs), nucleotide-binding oligomerization domain (NOD)-like receptors (NLRs), C-type lectin receptors (CLRs), AIM2-like receptors (ALRs), cyclic GMP-AMP synthase (cGAS), and retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs), have been discovered^15–19. Among these receptors, TLRs and RLRs are two major receptors responsible for sensing RNA virus infection and triggering antiviral IFN programs.

Toll was first identified as an antifungal gene responsible for the Drosophila immune system, and later studies further elucidated the fundamental role of TLRs in innate immune sensing in mammals²⁰. To date, multiple TLRs have been discovered, and their functions have been clarified. For example, 10 TLR members were identified (TLR1-TLR10) in humans, and 12 mouse TLRs (TLR1-TLR9 and TLR11-TLR13) were discovered, and their functions were examined^15,21. Each TLR can recognize common or distinct PAMPs that are typically derived from components of microbes such as nucleic acids, lipoproteins, and lipids. Among the TLRs, TLR 3, 7, and 8 are responsible for recognizing RNA viruses entering through endocytosis by sensing single- or double-stranded RNA (ssRNA: TLR7/TLR8; dsRNA: TLR3) in endosomal compartments (Fig. ​(Fig.11).

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Fig. 1

Sensing of RNA virus invasion by RLRs and TLRs.

Invasion by RNA viruses is sensed by cytosolic and endosomal RNA sensors, RLRs and TLRs. Both RLRs and TLRs can detect viral RNA species such as viral genomic RNA and dsRNA produced during viral replication. While RIG-I and MDA5 are responsible for sensing cytoplasmic viral RNAs such as 5′-ppp RNA with a secondary structure (RIG-I) or dsRNA (short dsRNA; RIG-I and long dsRNA; MDA5), TLRs can detect endosomal ssRNA (TLR7/8) or dsRNA (TLR3). Activated RLRs undergo a conformational change that allows RLRs to expose CARDs and trigger the IFN signaling pathway through the CARD-CARD interaction between RLRs and MAVS. On the other hand, activated TLR3 and TLR7/8 initiate the antiviral IFN program by recruiting the signal adapter molecules TRIF and MyD88, respectively. Then, subsequent activation of the shared downstream kinases and transcription factors elicits the production of IFNs and proinflammatory cytokines.

Unlike TLRs, RLRs are essential cytoplasmic viral sensors that recognize intracellular non-self RNAs possessing distinct patterns of secondary structures or biochemical modifications^18,22. RLRs are Asp-Glu-Ala-Asp (DEAD) box containing RNA helicases composed of three members, RIG-I, melanoma differentiation-associated gene 5 (MDA5), and laboratory of genetics and physiology 2 (LGP2). Structurally, all RLRs possess an RNA helicase domain with a C-terminal domain (CTD) that is responsible for RNA binding. While RIG-I and MDA5 have tandem caspase activation and recruitment domains (CARDs) for downstream signal transduction, LGP2 lacks a CARD domain, thereby functioning as a modulator of RIG-I and MDA5²³. RIG-I and MDA5 can recognize non-self RNAs through common and distinct mechanisms. dsRNA is a classical non-self RNA that is not produced in uninfected cells due to a lack of RNA-dependent RNA polymerase in mammalian cells. Both RIG-I and MDA5 can be activated by an artificial dsRNA, polyinosinic:polycytidylic (poly I:C) acid. Intriguingly, activation of RIG-I and MDA5 by dsRNA is differentially regulated in a dsRNA length-dependent manner²⁴. In addition to the double-stranded structure, recent studies have identified several biochemical properties of RLR-activating RNA species, such as (1) 5′-triphosphate with secondary structured RNAs²⁵, (2) 5′-diphosphate uncapped RNAs²⁶, and (3) RNAs with an unmethylated 5′-end nucleotide at the 2′-O position^27,28, which are typically generated during replication of RNA viruses, including coronaviruses (Fig. ​(Fig.11).

Upon RNA ligand binding, RLRs and TLRs immediately initiate antiviral defense programs. TLRs initiate downstream signal cascades by recruiting adapter proteins, such as myeloid differentiation primary response 88 (MyD88) (for TLR7 and TLR8) and TIR-domain-containing adapter-inducing IFN-β (TRIF) (for TLR3). On the other hand, RLRs undergo conformational changes from inactive ‘closed’ to active ‘open’ structures mediated by ATPase/helicase activity. Activated RLRs liberate CARDs to bind to the signaling adapter molecule mitochondria antiviral signaling protein (MAVS) via the CARD-CARD interaction. The signaling adapters MyD88, TRIF, and MAVS then coordinate downstream signaling pathways by recruiting several ubiquitin ligases, such as TNF receptor-associated factor (TRAF) 3 and TRAF6, that associate with antiviral kinases, such as TANK-binding kinase 1 (TBK1), I-kappa-B kinase ε (IKKε), and the IKKα/β/γ complex. Consequently, activation of the transcription factors IRF3 and IRF7 and NF-κB leads to the production of type I IFN and proinflammatory cytokines to operate the host antiviral IFN programs²⁹ (Fig. ​(Fig.11).

Recognition of coronavirus by RLRs

Induction of type I IFN by mouse hepatitis virus (MHV), a murine coronavirus, is regulated by both RIG-I and MDA5 in brain cells⁴⁰. In addition, MDA5 plays a pivotal role in controlling the pathogenesis of MHV⁴¹. HCoV-229E and MHV can evade MDA5 sensing through viral 2-O-methyltransferase, which sequesters viral RNAs²⁷. Moreover, a recent study elucidated that MHV Nsp15, a viral endonuclease, removes a polyuridine-rich region in the negative-stranded viral RNA (PUN RNA) that would otherwise trigger an MDA5-mediated antiviral response⁴². Therefore, these studies indicate that MDA5 may be a predominant cytoplasmic sensor of coronavirus RNAs. Although RLRs seem to be dispensable for IFN production against MERS-CoV, both RIG-I and MDA5 contribute to proinflammatory responses in MERS-CoV-infected macrophages⁴³.

It is still unclear whether RLRs are responsible for sensing SARS-CoV-2 infection. Recent studies have shown that although SARS-CoV-2 infection abolishes the type I and III IFN signaling pathways, proinflammatory responses are still robustly induced during SARS-CoV-2 infection^44,45. Given that coronaviruses replicate in the cytoplasm and generate cytoplasmic PAMP RNAs such as replicate intermediate dsRNAs⁴⁶, it is reasonable to assume that RLRs may contribute to the recognition of SARS-CoV-2 infection and the induction of proinflammatory responses. However, it needs to be elucidated whether SARS-CoV-2 indeed produces cytoplasmic PAMP RNAs during replication and, if so, which cytoplasmic RNA sensor(s) are responsible for recognizing PAMP RNAs generated by SARS-CoV-2.

Recognition of coronaviruses by TLRs

In addition to RLRs, several TLRs play essential roles in coronavirus-induced innate immune responses. A recent study showed that inborn errors of TLR3- and IRF7-mediated type I IFN immunity were associated with disease severity and mortality in COVID-19 patients⁴⁷. Moreover, mice lacking TRIF and translocating chain-associated membrane protein (TRAM), downstream adapter molecules of TLR3 and TLR4, showed more severe viral pathogenesis accompanied by increased mortality induced by SARS-CoV infection⁴⁸. Another study also showed that TLR4-deficient mice were more susceptible to MHV1 infection than wild-type mice⁴⁹, suggesting essential roles of TLR3 and TLR4 in antiviral responses against coronavirus infection.

Studies have also shown that TLR7 plays a critical role in the sensing of coronavirus infection in certain types of immune cells. In conventional dendritic cells (cDCs) and plasmacytoid dendritic cells (pDCs), the production of type I IFN and proinflammatory cytokines upon MHV and SARS-CoV infection is TLR7 dependent⁵⁰. Furthermore, a subsequent study showed that type I and type III induction by MERS-CoV are regulated by TLR7 in pDCs⁵¹. Most importantly, a recent clinical study discovered that several natural mutations in the TLR7 gene that cause ‘loss-of-function’ are associated with the severity and mortality of young COVID-19 patients, demonstrating that TLR7 sensing of SARS-CoV-2 is critical for the control of COVID-19 pathogenesis⁵². These results suggest that activation of TLR sensing pathways could be a potential therapeutic approach for COVID-19.

Interestingly, several coronavirus proteins appear to induce host innate immune responses via TLR-derived signaling pathways. It has been shown that the spike protein of SARS-CoV could induce IL-8 production by activating the mitogen-activated protein kinase (MAPK)-AP1 axis signaling pathway in lung epithelial and fibroblast cell lines⁵³. Another group also showed that the SARS-CoV spike protein can be recognized by TLR2 and triggers proinflammatory responses in PBMCs⁵⁴. Furthermore, it was also reported that the SARS-CoV membrane (M) protein could induce type-I IFN production through an unknown cytosolic sensing system that may be involved in a noncanonical TLR signaling pathway⁵⁵. Although the exact mechanisms underlying these regulatory effects are still unclear, these reports suggest that coronavirus proteins can serve as potential agonists of TLRs.

Nsp1

Nsp1 of coronaviruses has been reported to impede the host innate immune system by targeting multiple biological pathways. Nsp1 can inhibit protein translation by either blocking the assembly of the translation machinery or inducing the cleavage of 5′-capped host mRNAs. SARS-CoV Nsp1 blocks the translation of cap-dependent and internal ribosome entry site (IRES)-driven mRNA by binding to ribosomal subunits⁶⁵. Similarly, another study also demonstrated that SARS-CoV-2 Nsp1 inhibits the translation of host antiviral proteins by targeting the 40S ribosomal subunit⁶⁶.

Other studies have also shown that Nsp1 suppresses protein translation by inducing 5′-capped host mRNA cleavage. Intriguingly, SARS-CoV viral mRNAs, which contain an intact cap structure and poly-A tail, were not susceptible to Nsp1-mediated RNA cleavage⁶⁷. Further analysis revealed that the presence of the 5′-end leader sequence in the viral mRNA could prevent Nsp1-mediated endonucleolytic RNA cleavage, indicating a unique immune evasion strategy by Nsp1^65,67. Another novel inhibitory mechanism suggested is that SARS-CoV Nsp1 blocks mRNA nuclear export by directly inhibiting nuclear pore complex protein 93 (Nup93), thereby subsequently suppressing protein synthesis⁶⁸.

In addition to its inhibitory roles in protein synthesis, Nsp1 can also block the IFN signaling pathway (Fig. ​(Fig.3).3). Both virus- and IFN-induced antiviral responses were hampered by SARS-CoV Nsp1⁶⁹. Moreover, recent studies have also shown that SARS-CoV-2 Nsp1 can strongly suppress the promoter activities of IFN-stimulated response elements (ISREs)^70,71. Therefore, these reports provide evidence that Nsp1 has multiple antagonizing functions targeting host antiviral programs.

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Fig. 3

Antagonizing IFN-mediated immune responses by coronavirus.

Secreted type I IFNs, IFN-α, and IFN-β can activate IFNAR1/2 in bystander cells to provoke the host antiviral program. On the other hand, coronavirus suppresses host antiviral IFN responses by targeting multiple components of the IFN signaling pathway through the inhibitory functions of viral proteins.

Nsp3

Nsp3 is the largest protein among the genes encoded by the coronavirus genome and has multiple functions. Nsp3 plays a critical role in virus replication by processing the ORF1ab polyprotein via its ‘papain-like protease’ activity. Additionally, protease enzymatic activity appears to be involved in antagonizing the host innate immune response. SARS-CoV Nsp3 can bind to IRF3 and prevent the phosphorylation, dimerization, and nuclear translocation of IRF3, resulting in the suppression of the IFN signaling pathway⁷². Moreover, Nsp3 can inhibit the NF-κB signaling pathway by stabilizing IκBα, an NF-κB inhibitor⁷³, suggesting that Nsp3 can inhibit host antiviral responses in a protease activity-independent manner.

Recently, accumulating evidence has suggested that the deubiquitinating (DUB) activity of Nsp3 is critical for inhibiting host IFN signaling pathways. Several studies have shown that Nsp3 antagonizes ubiquitination and ISGylation of host antiviral proteins, such as IRF3 and p53, resulting in suppression of antiviral responses^74–76. Interestingly, the regulatory mechanism of the Nsp3 deubiquitinating function seems to be distinct between SARS-CoV and SARS-CoV-2. While SARS-CoV Nsp3 preferentially cleaves the ubiquitin chain of target proteins, ISGylated proteins are predominantly processed by SARS-CoV-2 Nsp3^75,77, suggesting shared but distinct inhibitory strategies via Nsp3 of different coronaviruses. Thus, targeting Nsp3 may serve as a potential therapeutic approach for controlling coronavirus-induced diseases.

DMV formation by Nsp3, Nsp4, and Nsp6

During virus replication, coronaviruses form a double-membrane compartment by hijacking the host endoplasmic reticulum (ER) membrane and construct a ‘shelter’ where the viral components can escape cytosolic PRR sensing¹⁴. Several coronavirus nonstructural proteins, such as Nsp3, Nsp4, and Nsp6, are known to play pivotal roles in double-membrane vesicle (DMV) formation. Nsp4, along with Nsp3 and Nsp6, hijacks the ER membrane and forms DMVs by inducing membrane rearrangement. Viral replicase complexes that contain PAMP RNA species are then sequestered into the DMVs, thus preventing activation of cytosolic RNA sensors⁷⁸.

Nsp5

Together with Nsp3, Nsp5 is responsible for processing viral polyproteins through its 3C-like protease activity and contributes to virus replication. Intriguingly, porcine deltacoronavirus (PDCoV) Nsp5 has been shown to suppress host antiviral responses by processing host antiviral proteins, such as STAT2⁷⁹, NF-kappa-B essential modulator (NEMO)⁸⁰, and mRNA decapping protein 1a (DCP1a)⁸¹. Given that the inhibitory effect of Nsp5 is dependent on protease activity, protease inhibitors specifically targeting Nsp5 may be a new therapeutic approach.

Nsp8

Nsp8, together with Nsp7, are components of the viral RdRp complex and play a role in facilitating RdRp activity⁸². In addition, a recent result from the preprint manuscript suggested that SARS-CoV-2 Nsp8 also plays an inhibitory role in host innate immunity. It was shown that Nsp8 could directly bind to MDA5 CARD and block K63-linked polyubiquitination, thereby suppressing the MDA5-derived IFN signaling pathway⁸³.

RNA modification by Nsp13, 14, 15, and 16

Coronavirus genomes encode multiple RNA-modifying enzymes that alter the biochemical properties of both host and viral RNAs. Via these enzymes, coronaviruses have evolved several means to evade the host immune system by modifying their RNAs to avoid being sensed by antiviral receptors. It has been shown that coronavirus Nsp13, Nsp14, and Nsp16 are associated with RNA modification. Nsp13 of various coronaviruses was shown to regulate the removal of the 5′-ppp moiety from viral RNAs, a molecular signature of RIG-I ligand^10,84,85.

The addition of a cap structure to the 5′-end of viral RNA by coronavirus capping enzymes is a smart strategy. For example, SARS-CoV Nsp14 plays a pivotal role in the 5′-capping of viral mRNA via its guanine-N7-methyltransferase activity (N7-MTase)^12,86. Furthermore, Nsp16 of SARS-CoV¹¹ and SARS-CoV-2⁸⁷ have been shown to regulate the 2′-O-methylation of viral RNA, which is essential for 5′ capping, thereby avoiding activation of antiviral sensors such as MDA5 and IFIT family proteins²⁷.

In addition to regulating 5′ capping, coronaviruses possess another strategy to modify viral RNA by viral endoribonuclease. As described in the previous section, recent studies have elucidated that Nsp15 can eliminate the 5′-polyuridine region from the virus PUN RNA to prevent being sensed by cytosolic dsRNA sensors, including MDA5, PKR, and OAS/RNase L⁴². Therefore, these coronavirus enzymes involved in RNA modification play a critical role in blocking the early events of host antiviral sensing.

ORF3a

ORF3a localizes to the plasma membrane of the ER and Golgi and induces ER stress by activating the PKR-like ER kinase (PERK) pathway. The activated PERK pathway can induce the phosphorylation and cause the degradation of IFNAR1⁸⁸, evading host antiviral IFN programs (Fig. ​(Fig.3).3). In addition, ORF3a also regulates apoptotic pathways. Recent studies have shown that ORF3a of SARS-CoV and SARS-CoV-2 can trigger host cell apoptosis by inducing caspase activation or Golgi fragmentation^89,90, indicating that ORF3a targets multiple cellular pathways to hamper host antiviral responses.

On the other hand, ORF3a appears to be involved in activating inflammatory pathways. It has been reported that SARS-CoV ORF3a can induce NLRP3-dependent inflammasome activation by directly interacting with ASC⁹¹. Moreover, ORF3a upregulates the NF-κB pathway by interacting with TRAF3, contributing to the inflammasome pathway. However, it is unclear whether ORF3a-mediated proinflammatory responses are beneficial for the viral life cycle.

ORF3b

ORF3b was shown to have a unique shuttling behavior, translocating from the nucleus to the outer membrane of mitochondria. Localization of SARS-CoV ORF3b in mitochondria is critical for its inhibition of the MAVS-mediated IRF3 activation pathway, leading to reductions in IFN and ISG expression⁹². In addition, a recent report showed that SARS-CoV-2 ORF3b inhibits the IFN signaling pathway by blocking the nuclear translocation of IRF3. Interestingly, SARS-CoV-2 produces a smaller ORF3b than SARS-CoV with a shortened C-terminal region. Further analysis performed with several ORF3bs from various CoV strains showed that ORF3b with a shorter C-terminus region showed a more potent inhibitory function than its longer form. Notably, the cytoplasmic localization of ORF3b seems to be associated with its antagonism of the IFN signaling pathway. The authors found that the longer ORF3b found in SARS-CoV-related strains possesses a putative nuclear localization signal (NLS) in its C-terminal region and facilitates its nuclear localization. However, shorter ORF3b from SARS-CoV-2-related strains is predominantly localized in the cytoplasm due to the loss of NLS in the shortened C′ terminus region, resulting in stronger antagonistic function⁹³. However, the underlying mechanism of SARS-CoV-2 ORF3b-mediated inhibition of IRF3 nuclear translocation to the cytoplasm still needs to be clarified.

ORF6

ORF6 is a small protein that is mainly localized in the cytoplasm and partially colocalized in the ER and Golgi compartment. Together with Nsp1, ORF6 appears to be the most potent antagonist of host antiviral responses. Multiple groups have shown that ORF6 can suppress IFN and ISG induction by blocking specific nuclear import and export pathways. For example, ORF6 inhibits the nuclear import of innate immune signaling-related transcription factors such as IRF3 and STAT1^70,71, resulting in the shutdown of downstream events (Figs. ​(Figs.22 and ​and3).3). Mechanistically, ORF6 targets the karyopherin-containing importin complex through its acidic region of its C′ terminus to block the nuclear transport of NLS-containing proteins⁹⁴. In addition, ORF6 was shown to block the nuclear export of newly synthesized mRNA by interacting with the nuclear pore complex (NPC) proteins Rae1 and Nup98, thereby suppressing gene expression in infected cells^95,96.

ORF7a

Studies have shown that SARS-CoV ORF7a induces cell cycle arrest and apoptosis via a caspase-dependent pathway⁹⁷. ORF7a interacts with pro-survival factors, such as B cell lymphoma (Bcl)-2 and Bcl-X_L, and directly inhibits the pro-survival function of Bcl-X_L, triggering apoptosis⁹⁸. Furthermore, ORF7a can directly bind to and inhibit bone marrow stromal antigen 2 (BTS-2)/Tetherin, an antiviral protein that restricts the release of enveloped viruses from host cells⁹⁹.

In addition, ORF7a was shown to suppress protein translation by hampering the host stress response pathway. For example, ORF7a specifically targets the p38 MAPK pathway and inhibits cellular protein synthesis, resulting in apoptosis¹⁰⁰. This inhibitory regulation can potentially cause a reduction in antiviral protein synthesis, leading to the pathogenesis of SARS-CoV.

ORF9b

Although the initial study with SARS-CoV suggested that ORF9b is a virion-associated accessory factor, recent studies have further elucidated that ORF9b can inhibit host innate immunity by targeting mitochondria-mediated antiviral immunity. Shi et al.¹⁰¹ showed that SARS-CoV ORF9b could trigger the degradation of MAVS, TRAF3, and TRAF6 by tethering poly(rC)-binding protein 2 (PCBP2) and E3 ligase atrophin 1 interacting protein 4 (AIP4) in mitochondria, thereby suppressing downstream antiviral responses. A recent study with SARS-CoV-2 suggested that ORF9b antagonizes the type I IFN pathway through association with translocator of outer membrane 70 (TOM70), a critical mitochondrial import receptor regulating IFN responses¹⁰². Overexpression of TOM70 rescued IFN-β expression, further confirming the specific inhibition of the TOM70-mediated antiviral signaling pathway by ORF9b.

Nucleocapsid

The nucleocapsid (N) protein is a coronavirus structural protein that plays essential roles in viral transcription and virion assembly. In addition to its contribution to virus replication, N plays pivotal roles in suppressing host innate immunity^103–105. Two studies with SARS-CoV showed that N suppresses IFN production. Lu and colleagues suggested that SARS-CoV N targets the upstream event of RNA sensing to block the IFN signaling pathway¹⁰³. A subsequent study by Hu and colleagues also showed that N could directly bind to tripartite motif-containing 25 (TRIM25) via its C-terminal region and interfere with the TRIM25–RIG-I interaction, thereby suppressing TRIM25-mediated RIG-I ubiquitination and activation¹⁰⁵.

In addition to targeting the RNA sensing pathway, a recent study with SARS-CoV-2 provided evidence that SARS-CoV-2 N also targets the IFN signaling pathway by suppressing the phosphorylation and nuclear translocation of STAT1 and STAT2¹⁰⁴ (Fig. ​(Fig.3).3). However, the exact mechanism underlying N-mediated inhibition of the IFN signaling pathway is still unclear. Nevertheless, these data suggest that the coronavirus N protein can hamper host innate immunity by targeting multiple antiviral responses.

This work was supported by the Japan Society for the Promotion of Science (JSPS) (grant number:　19K23837) and the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (No. 2019R1I1A1A01060399 and 2020R1I1A2073517).