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The nucleotide-binding oligomerization domain protein 2 (NOD2) senses bacterial peptidoglycan to induce proinflammatory and antimicrobial responses. Dysregulation of NOD2 signaling is involved in multiple inflammatory disorders. Recently, S-palmitoylation, a novel type of post-translational modification, is reported to play a crucial role in membrane association and ligand-induced signaling of NOD2, yet its influence on the stability of NOD2 is unclear. Here we show that inhibition of S-palmitoylation facilitates the SQSTM1/p62-mediated autophagic degradation of NOD2, while S-palmitoylation of NOD2 by ZDHHC5 promotes the stability of NOD2. Furthermore, we identify a gain-of-function R444C variant of NOD2 short isoform (NOD2s-R444C) in autoinflammatory disease, which induces excessive inflammation through its high S-palmitoylation level. Mechanistically, the NOD2s-R444C variant possesses a stronger binding ability to ZDHHC5, which promotes its S-palmitoylation, and restricts its autophagic degradation by reducing its interaction with SQSTM1/p62. Taken together, our study reveals the regulatory role of S-palmitoylation in controlling NOD2 stability through the crosstalk with autophagy, and provides insights into the association between dysfunctional S-palmitoylation and the occurrence of inflammatory diseases.

The intracellular pattern recognition receptor NOD2 senses bacterial peptidoglycan to drive proinflammatory and antimicrobial responses. Dysregulation of NOD2 signaling confers susceptibility to several immunological and inflammatory diseases. Although palmitoylation of NOD2 is required for its membrane recruitment and activation, whether palmitoylation can modulate the stability of NOD2 to orchestrate inflammation remains unclear. Recently, we have revealed that S-palmitoylation restricts SQSTM1-mediated selective macroautophagic/autophagic degradation of NOD2, and identified a gain-of-function R444C variant of NOD2 short isoform (NOD2s

Deubiquitination plays an important role in the regulation of the crosstalk between macroautophagy/autophagy and innate immune signaling, yet its regulatory mechanisms are not fully understood. Here we identify the deubiquitinase OTUD7B as a negative regulator of antiviral immunity by targeting IRF3 (interferon regulatory factor 3) for selective autophagic degradation. Mechanistically, OTUD7B interacts with IRF3, and activates IRF3-associated cargo receptor SQSTM1/p62 (sequestosome 1) by removing its K63-linked poly-ubiquitin chains at lysine 7 (K7) to enhance SQSTM1 oligomerization. Moreover, viral infection increased the expression of OTUD7B, which forms a negative feedback loop by promoting IRF3 degradation to balance type I interferon (IFN) signaling. Taken together, our study reveals a specific role of OTUD7B in mediating the activation of cargo receptors in a substrate-dependent manner, which could be a potential target against excessive immune responses. Abbreviations: Baf A1: bafilomycin A1; CGAS: cyclic GMP-AMP synthase; DDX58/RIG-I: DExD/H-box helicase 58; DSS: dextran sodium sulfate; DUBs: deubiquitinating enzymes; GFP: green fluorescent protein; IFN: interferon; IKKi: IKBKB/IkappaB kinase inhibitor; IRF3: interferon regulatory factor 3; ISGs: interferon-stimulated genes; MAVS: mitochondrial antiviral signaling protein; MOI: multiplicity of infection; PAMPs: pathogen-associated molecular patterns; SeV: Sendai virus; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1; Ub: ubiquitin; WT: wild-type; VSV: vesicular stomatitis virus.

Post-translational modifications (PTMs) of proteins are crucial to guarantee the proper biological functions in immune responses. Although protein phosphorylation has been extensively studied, our current knowledge of protein pyrophosphorylation, which occurs based on phosphorylation, is very limited. Protein pyrophosphorylation is originally considered to be a non-enzymatic process, and its function in immune signaling is unknown. Here, we identify a metabolic enzyme, UDP-N-acetylglucosamine pyrophosphorylase 1 (UAP1), as a pyrophosphorylase for protein serine pyrophosphorylation, by catalyzing the pyrophosphorylation of interferon regulatory factor 3 (IRF3) at serine (Ser) 386 to promote robust type I interferon (IFN) responses. Uap1 deficiency significantly impairs the activation of both DNA- and RNA-viruse-induced type I IFN pathways, and the Uap1-deficient mice are highly susceptible to lethal viral infection. Our findings demonstrate the function of protein pyrophosphorylation in the regulation of antiviral responses and provide insights into the crosstalk between metabolism and innate immunity.

Macroautophagy/autophagy is a conserved eukaryotic process to mediate the degradation of cell organelles and protein aggregates, which participates in a variety of cellular responses, including immune signal transduction. KDM4D functions as an important histone demethylase to regulate gene transcription by inhibiting histone H3K9 trimethylation. Whether autophagy epigenetically regulates the immune response via modulating the stability and activity of KDM4D remains largely unclear. Recently, we identified TRIM14 (tripartite motif-containing 14) as an epigenetic regulator, which recruits USP14 and BRCC3 to form a regulatory complex, and promotes an inflammation response through inhibiting OPTN-mediated autophagic degradation of KDM4D.

As a key component of the inflammasome, NLRP3 is a critical intracellular danger sensor emerging as an important clinical target in inflammatory diseases. However, little is known about the mechanisms that determine the kinetics of NLRP3 inflammasome stability and activity to ensure effective and controllable inflammatory responses. Here, we show that S-palmitoylation acts as a brake to turn NLRP3 inflammasome off. zDHHC12 is identified as the S-acyltransferase for NLRP3 palmitoylation, which promotes its degradation through the chaperone-mediated autophagy pathway. Zdhhc12 deficiency in mice enhances inflammatory symptoms and lethality following alum-induced peritonitis and LPS-induced endotoxic shock. Notably, several disease-associated mutations in NLRP3 are associated with defective palmitoylation, resulting in overt NLRP3 inflammasome activation. Thus, our findings identify zDHHC12 as a repressor of NLRP3 inflammasome activation and uncover a previously unknown regulatory mechanism by which the inflammasome pathway is tightly controlled by the dynamic palmitoylation of NLRP3.

In addition to investigating the virology of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), discovering the host–virus dependencies are essential to identify and design effective antiviral therapy strategy. Here, we report that the SARS-CoV-2 entry receptor, ACE2, conjugates with small ubiquitin-like modifier 3 (SUMO3) and provide evidence indicating that prevention of ACE2 SUMOylation can block SARS-CoV-2 infection. E3 SUMO ligase PIAS4 prompts the SUMOylation and stabilization of ACE2, whereas deSUMOylation enzyme SENP3 reverses this process. Conjugation of SUMO3 with ACE2 at lysine (K) 187 hampers the K48-linked ubiquitination of ACE2, thus suppressing its subsequent cargo receptor TOLLIP-dependent autophagic degradation. TOLLIP deficiency results in the stabilization of ACE2 and elevated SARS-CoV-2 infection. In conclusion, our findings suggest selective autophagic degradation of ACE2 orchestrated by SUMOylation and ubiquitination as a potential way to combat SARS-CoV-2 infection.

As a core kinase of antiviral immunity, the activity and stability of TANK-binding kinase 1 (TBK1) is tightly controlled by multiple post-translational modifications. Although it has been demonstrated that TBK1 stability can be regulated by ubiquitin-dependent proteasome pathway, it is unclear whether another important protein degradation pathway, autophagosome pathway, can specifically affect TBK1 degradation by cargo receptors. Here we report that E3 ubiquitin ligase NEDD4 functions as a negative regulator of type I interferon (IFN) signaling by targeting TBK1 for degradation at the late stage of viral infection, to prevent the host from excessive immune response. Mechanically NEDD4 catalyzes the K27-linked poly-ubiquitination of TBK1 at K344, which serves as a recognition signal for cargo receptor NDP52-mediated selective autophagic degradation. Taken together, our study reveals the regulatory role of NEDD4 in balancing TBK1-centered type I IFN activation and provides insights into the crosstalk between selective autophagy and antiviral signaling.

The ongoing 2019 novel coronavirus disease (COVID-19) caused by SARS-CoV-2 has posed a worldwide pandemic and a major global public health threat. The severity and mortality of COVID-19 are associated with virus-induced dysfunctional inflammatory responses and cytokine storms. However, the interplay between host inflammatory responses and SARS-CoV-2 infection remains largely unknown. Here, we demonstrate that SARS-CoV-2 nucleocapsid (N) protein, the major structural protein of the virion, promotes the virus-triggered activation of NF-κB signaling. After binding to viral RNA, N protein robustly undergoes liquid–liquid phase separation (LLPS), which recruits TAK1 and IKK complex, the key kinases of NF-κB signaling, to enhance NF-κB activation. Moreover, 1,6-hexanediol, the inhibitor of LLPS, can attenuate the phase separation of N protein and restrict its regulatory functions in NF-κB activation. These results suggest that LLPS of N protein provides a platform to induce NF-κB hyper-activation, which could be a potential therapeutic target against COVID-19 severe pneumonia.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the ongoing coronavirus disease 2019 (COVID-19) pandemic. Alongside investigations into the virology of SARS-CoV-2, understanding the host–virus dependencies are vital for the identification and rational design of effective antiviral therapy. Here, we report the dominant SARS-CoV-2 entry receptor, ACE2, conjugates with small ubiquitin-like modifier 3 (SUMO3) through a proteome-wide protein interaction analysis. We further demonstrate that E3 SUMO ligase PIAS4 prompts the SUMOylation and stabilization of ACE2, whereas deSUMOylation enzyme SENP3 reverses this process. Conjugation of SUMO3 with ACE2 at lysine (K) 187 hampers the K48-linked ubiquitination of ACE2, thus suppressing its subsequent cargo receptor TOLLIP-dependent autophagic degradation. Pharmacological intervention of ACE2 SUMOylation blocks the entry of SARS-CoV-2 and viral infection-triggered immune responses. Collectively, our findings suggest selective autophagic degradation of ACE2 orchestrated by SUMOylation and ubiquitination can be targeted to future antiviral therapy of SARS-CoV-2.

IRF3 (interferon regulatory factor 3) is one of the most critical transcription factors in antiviral innate immune signaling, which is ubiquitously expressed in a variety of cells. Although it has been demonstrated that IRF3 can provoke multiple cellular processes during viral infection, including type I interferon (IFN) production, the mechanisms underlying the precise regulation of IRF3 activity are still not completely understood. Here, we report that selective macroautophagy/autophagy mediated by cargo receptor CALCOCO2/NDP52 promotes the degradation of IRF3 in a virus load-dependent manner. Deubiquitinase PSMD14/POH1 prevents IRF3 from autophagic degradation by cleaving the K27-linked poly-ubiquitin chains at lysine 313 on IRF3 to maintain its basal level and IRF3-mediated type I IFN activation. The autophagic degradation of IRF3 mediated by PSMD14 or CALCOCO2 ensures the precise control of IRF3 activity and fine-tunes the immune response against viral infection. Our study reveals the regulatory role of PSMD14 in balancing IRF3-centered IFN activation with immune suppression and provides insights into the crosstalk between selective autophagy and type I IFN signaling. ATG5: autophagy related gene 5; Baf A1: bafilomycin A1; BECN1: beclin 1; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CGAS: cyclic GMP-AMP synthase; DDX58/RIG-I: DExD/H-box helicase 58; DUBs: deubiquitinating enzymes; IFN: interferon; IRF3: interferon regulatory factor 3; MAVS: mitochondrial antiviral signaling protein; MOI: multiplicity of infection; PAMPs: pathogen-associated molecule patterns; PBMC: peripheral blood mononuclear cell; PSMD14/POH1: proteasome 26S subunit, non-ATPase 14; RIPA: RLR-induced IRF3-mediated pathway of apoptosis; SeV: Sendai virus; SQSTM1/p62: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1; Ub: ubiquitin; WT: wild type

Macroautophagy/autophagy, a eukaryotic homeostatic process that sequesters cytoplasmic constituents for lysosomal degradation, is orchestrated by a number of autophagy-related (ATG) proteins tightly controlled by post-translational modifications. However, the involvement of reversible ubiquitination in the regulation of autophagy remains largely unclear. Here, we performed a single-guide RNA-based screening assay to investigate the functions of deubiquitinating enzymes (DUBs) in regulating autophagy. We identified previously unrecognized roles of several DUBs in modulating autophagy at multiple levels by targeting various ATG proteins. Mechanistically, we demonstrated that STAMBP/AMSH (STAM-binding protein) promotes the stabilization of ULK1 by removing its lysine 48 (K48)-linked ubiquitination, whereas OTUD7B mediates the degradation of PIK3 C3 by enhancing its K48-linked ubiquitination, thus positively or negatively affects autophagy flux, respectively. Together, our study elaborated on the broad involvement of DUBs in regulating autophagy and uncovered the critical roles of the reversible ubiquitination in the modification of ATG proteins. ATG: autophagy-related; Baf A1: bafilomycin A1; DUB: deubiquitinating enzyme; EBSS: Earle’s balanced salt solution; KO: knockout; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; OTUD7B: OTU domain-containing protein 7B; PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3; sgRNA: single-guide RNA; SQSTM1/p62: sequestosome 1; STAMBP/AMSH: STAM-binding protein; ULK1: unc-51 like autophagy activating kinase 1; USP: ubiquitin specific peptidase.

Macroautophagy/autophagy, an evolutionarily conserved eukaryotic bioprocess, plays an important role in the bulk degradation of intracellular macromolecules, organelles, and invading pathogens. PIK3C3/VPS34 (phosphatidylinositol 3-kinase catalytic subunit type 3) functions as a key protein in autophagy initiation and progression. The activity of PIK3C3 is tightly regulated by multiple post-translational modifications, including ubiquitination, however, the regulatory mechanisms underpinning the reversible deubiquitination of PIK3C3 remain poorly understood. Recently, we identified the E3 ubiquitin ligase NEDD4/NEDD4-1 as a positive regulator of autophagy through decreasing the K48-linked ubiquitination of PIK3C3 by recruiting USP13.

The production of type I interferons (IFNs) is one of the hallmarks of intracellular antimicrobial program. Typical type I IFN response activates the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway, which results in the transcription of plentiful IFN-stimulated genes (ISGs) to establish the comprehensive antiviral states. Type I IFN signaling should initiate timely to provoke innate and adaptive immune responses for effective elimination of the invading pathogens. Meanwhile, a precise control must come on the stage to restrain the persistent activation of type I IFN responses to avoid attendant toxicity. Autophagy, a conserved eukaryotic degradation system, mediated by a number of autophagy-related (ATG) proteins, plays an essential role in the clearance of invading microorganism and manipulation of type I responses. Autophagy modulates type I IFN responses through regulatory integration with innate immune signaling pathways, and by removing endogenous ligands of innate immune sensors. Moreover, selective autophagy governs the choice of innate immune factors as specific cargoes for degradation, thus tightly monitoring the type I IFN responses. This review will focus on the cross-regulation between autophagy and type I IFN signaling in host defense.

The immune system plays a critical role in defense against invading pathogens, and its function must be strictly controlled to maintain intracellular homeostasis. Once suffering microbial invasion or receiving danger signals, the immune system initiates the responses timely. After the threat removal, the immune system should be shut down to avoid the harm caused by excessive immune activation. Additionally, the immune system needs to be internally adjusted so that it does not respond to self-antigens to avoid autoimmune diseases. The states of nonresponse in immunity are termed as immune tolerance. Numerous studies indicated that macroautophagy (hereafter named as autophagy) is involved in T cells and B cells related immune tolerance. Recently, more and more researches demonstrated that autophagy is not only capable of nonselective degradation of cellular macromolecular components but also responsible for sorting and transporting autophagic substrates through a group of cargo receptors for selective degradation, which is called as selective autophagy. Recent studies indicated that selective autophagy can effectively regulate the immune tolerance and avoid over-activation of immune response by targeting multiple receptors and effectors of immune cells. In this chapter, we will focus on how autophagy participates explicitly in the adaptive and innate immune tolerance.

The production of type I interferons (IFNs) is one of the hallmarks of intracellular antimicrobial program. Typical type I IFN response activates the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway, which results in the transcription of plentiful IFN-stimulated genes (ISGs) to establish the comprehensive antiviral states. Type I IFN signaling should initiate timely to provoke innate and adaptive immune responses for effective elimination of the invading pathogens. Meanwhile, a precise control must come on the stage to restrain the persistent activation of type I IFN responses to avoid attendant toxicity. Autophagy, a conserved eukaryotic degradation system, mediated by a number of autophagy-related (ATG) proteins, plays an essential role in the clearance of invading microorganism and manipulation of type I responses. Autophagy modulates type I IFN responses through regulatory integration with innate immune signaling pathways, and by removing endogenous ligands of innate immune sensors. Moreover, selective autophagy governs the choice of innate immune factors as specific cargoes for degradation, thus tightly monitoring the type I IFN responses. This review will focus on the cross-regulation between autophagy and type I IFN signaling in host defense.

DDX58/RIG-I, is a critical pattern recognition receptor for viral RNA, which plays an essential role in antiviral immunity. Its posttranslational modifications and stability are tightly regulated to mediate the moderate production of type I IFN to maintain the immune homeostasis. Recently, we reported that macroautophagy/autophagy balances type I IFN signaling through selective degradation of ISG15-associated DDX58 via LRRC25. However, the regulatory mechanism about the autophagic degradation of DDX58 remains largely undefined. Here, we identified LRRC59 as a vital positive regulator of DDX58-mediated type I IFN signaling. Upon virus infection, LRRC59 specifically interacted with ISG15-associated DDX58 and blocked its association with LRRC25, the secondary receptor to deliver DDX58 to autophagosomes for SQSTM1/p62-dependent degradation, leading to the stronger antiviral immune responses. Thus, our study reveals a novel regulatory role of selective autophagy in innate antiviral responses mediated by the cross-regulation of LRRC family members. These data further provide insights into the crosstalk between autophagy and innate immune responses. ATG: Autophagy-related; Baf A1: Bafilomycin A1; DDX58/RIG-I: DEAD [Asp-Glu-Ala-Asp] box polypeptide 58; EV: Empty vector; IC poly[I:C]: Intracellular polyriboinosinic polyribocytidylic acid; IFIH1/MDA5: Interferon induced with helicase C domain 1; IFN: Interferon; ISG15: ISG15 ubiquitin like modifier; IKBKE: Inhibitor of nuclear factor kappa B kinase subunit epsilon; IRF3: Interferon regulatory factor 3; KO: Knockout; LRRC: Leucine rich repeat containing; MAVS: Mitochondrial antiviral signaling protein; CGAS/MB21D1: Cyclic GMP-AMP synthase; SeV: Sendai virus; siRNA: small interfering RNA; SQSTM1/p62: Sequestosome 1; TBK1: TANK binding kinase 1; TLR: Toll like receptor; TMEM173/STING: Transmembrane protein 173; VSV: Vesicular stomatitis virus; WT: Wild type

Summary: The class III phosphoinositide 3-kinase vacuolar protein sorting 34 (VPS34) is a core protein of autophagy initiation, yet the regulatory mechanisms responsible for its stringent control remain poorly understood. Here, we report that the E3 ubiquitin ligase NEDD4-1 promotes the autophagy flux by targeting VPS34. NEDD4-1 undergoes lysine 29 (K29)-linked auto-ubiquitination at K1279 and serves as a scaffold for recruiting the ubiquitin-specific protease 13 (USP13) to form an NEDD4-1-USP13 deubiquitination complex, which subsequently stabilizes VPS34 to promote autophagy through removing the K48-linked poly-ubiquitin chains from VPS34 at K419. Knockout of either NEDD4-1 or USP13 increased K48-linked ubiquitination and degradation of VPS34, thus attenuating the formation of the autophagosome. Our results identify an essential role for NEDD4-1 in regulating autophagy, which provides molecular insights into the mechanisms by which ubiquitination regulates autophagy flux. : Xie et al. demonstrate that HECT ubiquitin E3 ligase NEDD4-1 undergoes lysine 29 (K29)-linked auto-ubiquitination and serves as a scaffold to recruit the ubiquitin-specific protease 13 (USP13) to form a deubiquitination complex, which subsequently stabilizes VPS34 to promote autophagy through the removal of K48-linked poly-ubiquitin chains on VPS34. Keywords: autophagy, VPS34, NEDD4-1, USP13, deubiquitination complex, auto-ubiquitination

Viral infection triggers host innate immune responses, which primarily include the activation of type I interferon (IFN) signaling and inflammasomes. Here, we report that Zika virus (ZIKV) infection triggers NLRP3 inflammasome activation, which is further enhanced by viral non‐structural protein NS1 to benefit its replication. NS1 recruits the host deubiquitinase USP8 to cleave K11‐linked poly‐ubiquitin chains from caspase‐1 at Lys134, thus inhibiting the proteasomal degradation of caspase‐1. The enhanced stabilization of caspase‐1 by NS1 promotes the cleavage of cGAS, which recognizes mitochondrial DNA release and initiates type I IFN signaling during ZIKV infection. NLRP3 deficiency increases type I IFN production and strengthens host resistance to ZIKV in vitro and in vivo . Taken together, our work unravels a novel antagonistic mechanism employed by ZIKV to suppress host immune response by manipulating the interplay between inflammasome and type I IFN signaling, which might guide the rational design of therapeutics in the future.

MAVS signalosome plays an important role in RIG-I-like receptor (RLR)-induced antiviral signaling. Upon the recognition of viral RNAs, RLRs activate MAVS, which further recruits TRAF6 and other signaling proteins to initiate type I interferon (IFN) activation. MAVS signalosome also regulates virus-induced apoptosis to limit viral replication. However, the mechanisms that control the activity of MAVS signalosome are still poorly defined. Here, we report NLRP11, a Nod-like receptor, is induced by type I IFN and translocates to mitochondria to interact with MAVS upon viral infection. Using MAVS as a platform, NLRP11 degrades TRAF6 to attenuate the production of type I IFNs as well as virus-induced apoptosis. Our findings reveal the regulatory role of NLRP11 in antiviral immunity by disrupting MAVS signalosome.

Nuclear factor κB (NF-κB) is a family of critical transcription factors that play a critical role in innate immune responses and inflammation, yet the molecular mechanisms responsible for its tight regulation is not fully understood. In this study, we identified LRRC25, a member of leucine-rich repeat (LRR)-containing protein family, as a negative regulator in the NF-κB signaling pathway. Ectopic expression of LRRC25 impaired NF-κB activation, whereas knockout of LRRC25 potentiated NF-κB activation and enhanced the production of inflammatory cytokines. Further study demonstrated that the LRR domain of LRRC25 interacted with the Rel Homology domain (RHD) of p65/RelA and promotes the degradation of p65/RelA. Furthermore, LRRC25 enhanced the interaction between p65/RelA and cargo receptor p62, thus facilitating the degradation of p65/RelA through autophagy pathway. Our study has not only identified LRRC25 as a novel inhibitor of NF-κB signaling pathway, but also uncovers a new mechanism of crosstalk between NF-κB signaling and autophagy pathways.

Stratified multisite ubiquitination of RIG-I provides a robust and optimal control of innate antiviral responses., The activation of retinoic acid–inducible gene I (RIG-I), an indispensable viral RNA sensor in mammals, is subtly regulated by ubiquitination. Although multiple ubiquitination sites at the amino terminus of RIG-I have been identified, their functional allocations in RIG-I activation remain elusive. We identified a stratified model for RIG-I amino-terminal ubiquitination, in which initiation at either Lys164 or Lys172 allows subsequent ubiquitination at other lysines, to trigger and amplify RIG-I activation. Experimental and mathematical modeling showed that multisite ubiquitination provides robustness in RIG-I–mediated type I interferon (IFN) signaling. Furthermore, the flexibly controlled ultrasensitivity and IFN activation intensity determine the specificity of the IFN-stimulated gene transcription and manipulate cell fate in antiviral immune response. Our work demonstrates that tunable type I IFN signaling can be regulated through multisite RIG-I ubiquitination and elucidates a new paradigm for dynamic regulation in RIG-I–mediated antiviral signaling.

Summary Tetherin (BST2/CD317) is an interferon-inducible antiviral factor known for its ability to block the release of enveloped viruses from infected cells. Yet its role in type I interferon (IFN) signaling remains poorly defined. Here, we demonstrate that Tetherin is a negative regulator of RIG-I like receptor (RLR)-mediated type I IFN signaling by targeting MAVS. The induction of Tetherin by type I IFN accelerates MAVS degradation via ubiquitin-dependent selective autophagy in human cells. Moreover, Tetherin recruits E3 ubiquitin ligase MARCH8 to catalyze K27-linked ubiquitin chains on MAVS at lysine 7, which serves as a recognition signal for NDP52-dependent autophagic degradation. Taken together, our findings reveal a negative feedback loop of RLR signaling generated by Tetherin-MARCH8-MAVS-NDP52 axis and provide insights into a better understanding of the crosstalk between selective autophagy and optimal deactivation of type I IFN signaling.

Macroautophagy/autophagy is a conserved intracellular degradation system that traffics substrates including protein aggregates, defunct or disused organelles and invading pathogens to lysosomes via double-membrane vesicles called autophagosomes. BECN1/Beclin 1 functions as a key protein in autophagy initiation and progression; however, the role of BECN1 in innate immunity has not been fully investigated. Recently, we have found that USP19 affects the ubiquitination of BECN1, hence promoting the formation of autophagosomes and inhibiting DDX58/RIG-I-mediated type I interferon signaling.

RNA interference (RNAi) has great potential for use in insect pest control. However, some significant challenges must be overcome before RNAi-based pest control can become a reality. One challenge is the proper selection of a good target gene for RNAi. Here, we report that the insect ecdysone receptor (EcR) is a good potential target for RNAi-based pest control in the brown planthopper Nilaparvata lugens, a serious insect pest of rice plants. We demonstrated that the use of a 360 bp fragment (NlEcR-c) that is common between NlEcR-A and NlEcR-B for feeding RNAi experiments significantly decreased the relative mRNA expression levels of NlEcR compared with those in the dsGFP control. Feeding RNAi also resulted in a significant reduction in the number of offspring per pair of N. lugens. Consequently, a transgenic rice line expressing NlEcR dsRNA was constructed by Agrobacterium- mediated transformation. The results of qRT-PCR showed that the total copy number of the target gene in all transgenic rice lines was 2. Northern blot analysis showed that the small RNA of the hairpin dsNlEcR-c was successfully expressed in the transgenic rice lines. After newly hatched nymphs of N. lugens fed on the transgenic rice lines, effective RNAi was observed. The NlEcR expression levels in all lines examined were decreased significantly compared with the control. In all lines, the survival rate of the nymphs was nearly 90%, and the average number of offspring per pair in the treated groups was significantly less than that observed in the control, with a decrease of 44.18-66.27%. These findings support an RNAi-based pest control strategy and are also important for the management of rice insect pests.