Activation of the cGAS-STING-IRF3 Axis by Type I and II Interferons Contributes to Host Defense.

Interferons (IFNs) activate JAK-STAT pathways to induce downstream effector genes for host defense against invaded pathogens and tumors. Here both type I (ß) and II (γ) IFNs are shown that can activate the transcription factor IRF3 in parallel with STAT1. IRF3-deficiency impairs transcription of a subset of downstream effector genes induced by IFN-ß and IFN-γ. Mechanistically, IFN-induced activation of IRF3 is dependent on the cGAS-STING-TBK1 axis. Both IFN-ß and IFN-γ cause mitochondrial DNA release into the cytosol. In addition, IFNs induce JAK1-mediated tyrosine phosphorylation of cGAS at Y214/Y215, which is essential for its DNA binding activity and signaling. Furthermore, deficiency of cGAS, STING, or IRF3 impairs IFN-ß- or IFN-γ-mediated antiviral and antitumor activities. The findings reveal a novel IRF3 activation pathway parallel with the canonical STAT1/2 activation pathways triggered by IFNs and provide an explanation for the pleiotropic roles of the cGAS-STING-IRF3 axis in host defense.

Entanglement signatures for quantum synchronization with single-ion phonon laser.

The entanglement properties of quantum synchronization, based on a single-ion phonon laser subjected to an external drive, have been studied. It is found that the maximum value of steady-state entanglement between the ion's internal and external states occurs near the noiseless boundary from synchronization to unsynchronization, accompanied by noticeable oscillatory behaviors during the corresponding time evolution of entanglement. In addition, the later time dynamics of entanglement also indicates the occurrence of frequency entrainment, as evidenced by the strong consistency between the bending of the observed frequency and the emergence of Liouvillian exceptional points (LEPs) in the first two eigenvalues of the Liouvillian eigenspectrum. Moreover, the emergence of LEPs, which is intimately associated with frequency entrainment, should be widely observed in quantum synchronization and can be explored in LEPs-based applications.

LDLR is an entry receptor for Crimean-Congo hemorrhagic fever virus.

Crimean-Congo hemorrhagic fever virus (CCHFV) is the most widespread tick-born zoonotic bunyavirus that causes severe hemorrhagic fever and death in humans. CCHFV enters the cell via clathrin-mediated endocytosis which is dependent on its surface glycoproteins. However, the cellular receptors that are required for CCHFV entry are unknown. Here we show that the low density lipoprotein receptor (LDLR) is an entry receptor for CCHFV. Genetic knockout of LDLR impairs viral infection in various CCHFV-susceptible human, monkey and mouse cells, which is restored upon reconstitution with ectopically-expressed LDLR. Mutagenesis studies indicate that the ligand binding domain (LBD) of LDLR is necessary for CCHFV infection. LDLR binds directly to CCHFV glycoprotein Gc with high affinity, which supports virus attachment and internalization into host cells. Consistently, a soluble sLDLR-Fc fusion protein or anti-LDLR blocking antibodies impair CCHFV infection into various susceptible cells. Furthermore, genetic knockout of LDLR or administration of an LDLR blocking antibody significantly reduces viral loads, pathological effects and death following CCHFV infection in mice. Our findings suggest that LDLR is an entry receptor for CCHFV and pharmacological targeting of LDLR may provide a strategy to prevent and treat Crimean-Congo hemorrhagic fever.

BLK positively regulates TLR/IL-1R signaling by catalyzing TOLLIP phosphorylation.

TLR/IL-1R signaling plays a critical role in sensing various harmful foreign pathogens and mounting efficient innate and adaptive immune responses, and it is tightly controlled by intracellular regulators at multiple levels. In particular, TOLLIP forms a constitutive complex with IRAK1 and sequesters it in the cytosol to maintain the kinase in an inactive conformation under unstimulated conditions. However, the underlying mechanisms by which IRAK1 dissociates from TOLLIP to activate TLR/IL-1R signaling remain obscure. Herein, we show that BLK positively regulates TLR/IL-1R-mediated inflammatory response. BLK-deficient mice produce less inflammatory cytokines and are more resistant to death upon IL-1ß challenge. Mechanistically, BLK is preassociated with IL1R1 and IL1RAcP in resting cells. IL-1ß stimulation induces heterodimerization of IL1R1 and IL1RAcP, which further triggers BLK autophosphorylation at Y309. Activated BLK directly phosphorylates TOLLIP at Y76/86/152 and further promotes TOLLIP dissociation from IRAK1, thereby facilitating TLR/IL-1R-mediated signal transduction. Overall, these findings highlight the importance of BLK as an active regulatory component in TLR/IL-1R signaling.

Tyrosine phosphorylation of IRF3 by BLK facilitates its sufficient activation and innate antiviral response.

Viral infection triggers the activation of transcription factor IRF3, and its activity is precisely regulated for robust antiviral immune response and effective pathogen clearance. However, how full activation of IRF3 is achieved has not been well defined. Herein, we identified BLK as a key kinase that positively modulates IRF3-dependent signaling cascades and executes a pre-eminent antiviral effect. BLK deficiency attenuates RNA or DNA virus-induced ISRE activation, interferon production and the cellular antiviral response in human and murine cells, whereas overexpression of BLK has the opposite effects. BLK-deficient mice exhibit lower serum cytokine levels and higher lethality after VSV infection. Moreover, BLK deficiency impairs the secretion of downstream antiviral cytokines and promotes Senecavirus A (SVA) proliferation, thereby supporting SVA-induced oncolysis in an in vivo xenograft tumor model. Mechanistically, viral infection triggers BLK autophosphorylation at tyrosine 309. Subsequently, activated BLK directly binds and phosphorylates IRF3 at tyrosine 107, which further promotes TBK1-induced IRF3 S386 and S396 phosphorylation, facilitating sufficient IRF3 activation and downstream antiviral response. Collectively, our findings suggest that targeting BLK enhances viral clearance via specifically regulating IRF3 phosphorylation by a previously undefined mechanism.

Orthodontic-surgical treatment for severe skeletal class II malocclusion with vertical maxillary excess and four premolars extraction: A case report.

BACKGROUND: Patient satisfaction with facial appearance at the end of orthodontic camouflage treatment is very important, especially for skeletal malocclusion. This case report highlights the importance of the treatment plan for a patient initially treated with four-premolar-extraction camouflage, despite indications for orthognathic surgery. CASE SUMMARY: A 23-year-old male sought treatment complaining about his unsatisfactory facial appearance. His maxillary first premolars and mandibular second premolars had been extracted, and a fixed appliance had been used to retract his anterior teeth for two years without improvement. He had a convex profile, a gummy smile, lip incompetence, inadequate maxillary incisor inclination, and almost a class I molar relationship. Cephalometric analysis showed severe skeletal class II malocclusion (A point-nasion-B point = 11.5°) with a retrognathic mandible (sella-nasion-B point = 75.9°), a protruded maxilla (sella-nasion-A point = 87.4°), and vertical maxillary excess (upper incisor to palatal plane = 33.2 mm). The excessive lingual inclination of the maxillary incisors (upper incisor to nasion-A point line = -5.5°) was due to previous treatment attempts to compensate for the skeletal class II malocclusion. The patient was successfully retreated with decompensating orthodontic treatment combined with orthognathic surgery. The maxillary incisors were repositioned and proclined in the alveolar bone, the overjet was increased, and a space was created for orthognathic surgery, including maxillary impaction, anterior maxillary back-setting, and bilateral sagittal split ramus osteotomy to correct his skeletal anteroposterior discrepancy. Gingival display was reduced, and lip competence was restored. In addition, the results remained stable after 2 years. The patient was satisfied with his new profile as well as with the functional malocclusion at the end of treatment. CONCLUSION: This case report provides orthodontists a good example of how to treat an adult with severe skeletal class II malocclusion with vertical maxillary excess after an unsatisfactory orthodontic camouflage treatment. Orthodontic and orthognathic treatment can significantly correct a patient's facial appearance.

African Swine Fever Virus Cysteine Protease pS273R Inhibits Type I Interferon Signaling by Mediating STAT2 Degradation.

African swine fever virus (ASFV) is a large DNA virus that causes African swine fever (ASF), an acute and hemorrhagic disease in pigs with lethality rates of up to 100%. To date, how ASFV efficiently suppress the innate immune response remains enigmatic. In this study, we identified ASFV cysteine protease pS273R as an antagonist of type I interferon (IFN). Overexpression of pS273R inhibited JAK-STAT signaling triggered by type I IFNs. Mechanistically, pS273R interacted with STAT2 and recruited the E3 ubiquitin ligase DCST1, resulting in K48-linked polyubiquitination at K55 of STAT2 and subsequent proteasome-dependent degradation of STAT2. Furthermore, such a function of pS273R in JAK-STAT signaling is not dependent on its protease activity. These findings suggest that ASFV pS273R is important to evade host innate immunity. IMPORTANCE ASF is an acute disease in domestic pigs caused by infection with ASFV. ASF has become a global threat with devastating economic and ecological consequences. To date, there are no commercially available, safe, and efficacious vaccines to prevent ASFV infection. ASFV has evolved a series of strategies to evade host immune responses, facilitating its replication and transmission. Therefore, understanding the immune evasion mechanism of ASFV is helpful for the development of prevention and control measures for ASF. Here, we identified ASFV cysteine protease pS273R as an antagonist of type I IFNs. ASFV pS273R interacted with STAT2 and mediated degradation of STAT2, a transcription factor downstream of type I IFNs that is responsible for induction of various IFN-stimulated genes. pS273R recruited the E3 ubiquitin ligase DCST1 to enhance K48-linked polyubiquitination of STAT2 at K55 in a manner independent of its protease activity. These findings suggest that pS273R is important for ASFV to escape host innate immunity, which sheds new light on the mechanisms of ASFV immune evasion.

Transcription-independent regulation of STING activation and innate immune responses by IRF8 in monocytes.

Sensing of cytosolic DNA of microbial or cellular/mitochondrial origin by cGAS initiates innate immune responses via the adaptor protein STING. It remains unresolved how the activity of STING is balanced between a productive innate immune response and induction of autoimmunity. Here we show that interferon regulatory factor 8 (IRF8) is essential for efficient activation of STING-mediated innate immune responses in monocytes. This function of IRF8 is independent of its transcriptional role in monocyte differentiation. In uninfected cells, IRF8 remains inactive via sequestration of its IRF-associated domain by its N- and C-terminal tails, which reduces its association with STING. Upon triggering the DNA sensing pathway, IRF8 is phosphorylated at Serine 151 to allow its association with STING via the IRF-associated domain. This is essential for STING polymerization and TBK1-mediated STING and IRF3 phosphorylation. Consistently, IRF8-deficiency impairs host defense against the DNA virus HSV-1, and blocks DNA damage-induced cellular senescence. Bone marrow-derived mononuclear cells which have an autoimmune phenotype due to deficiency of Trex1, respond to IRF-8 deletion with reduced pro-inflammatory cytokine production. Peripheral blood mononuclear cells from systemic lupus erythematosus patients are characterized by elevated phosphorylation of IRF8 at the same Serine residue we find to be important in STING activation, and in these cells STING is hyper-active. Taken together, the transcription-independent function of IRF8 we describe here appears to mediate STING activation and represents an important regulatory step in the cGAS/STING innate immune pathway in monocytes.

African swine fever virus I267L acts as an important virulence factor by inhibiting RNA polymerase III-RIG-I-mediated innate immunity.

ASFV is a large DNA virus that is highly pathogenic in domestic pigs. How this virus is sensed by the innate immune system as well as why it is so virulent remains enigmatic. In this study, we show that the ASFV genome contains AT-rich regions that are recognized by the DNA-directed RNA polymerase III (Pol-III), leading to viral RNA sensor RIG-I-mediated innate immune responses. We further show that ASFV protein I267L inhibits RNA Pol-III-RIG-I-mediated innate antiviral responses. I267L interacts with the E3 ubiquitin ligase Riplet, disrupts Riplet-RIG-I interaction and impairs Riplet-mediated K63-polyubiquitination and activation of RIG-I. I267L-deficient ASFV induces higher levels of interferon-ß, and displays compromised replication both in primary macrophages and pigs compared with wild-type ASFV. Furthermore, I267L-deficiency attenuates the virulence and pathogenesis of ASFV in pigs. These findings suggest that ASFV I267L is an important virulence factor by impairing innate immune responses mediated by the RNA Pol-III-RIG-I axis.

Herpes simplex virus protein UL56 inhibits cGAS-Mediated DNA sensing to evade antiviral immunity.

After herpes simplex virus type 1 (HSV-1) infection, the cytosolic sensor cyclic GMP-AMP synthase (cGAS) recognizes DNA and catalyzes synthesis of the second messenger 2'3'-cGAMP. cGAMP binds to the ER-localized adaptor protein MITA (also known as STING) to activate downstream antiviral responses. Conversely, HSV-1-encoded proteins evade antiviral immune responses via a wide variety of delicate mechanisms, promoting viral replication and pathogenesis. Here, we identified HSV-1 envelop protein UL56 as a negative regulator of cGAS-mediated innate immune responses. Overexpression of UL56 inhibited double-stranded DNA-triggered antiviral responses, whereas UL56-deficiency increased HSV-1-triggered induction of downstream antiviral genes. UL56-deficiency inhibited HSV-1 replication in wild-type but not MITA-deficient cells. UL56-deficient HSV-1 showed reduced replication in the brain of infected mice and was less lethal to infected mice. Mechanistically, UL56 interacted with cGAS and inhibited its DNA binding and enzymatic activity. Furthermore, we found that UL56 homologous proteins from different herpesviruses had similar roles in antagonizing cGAS-mediated innate immune responses. Our findings suggest that UL56 is a component of HSV-1 evasion of host innate immune responses by antagonizing the DNA sensor cGAS, which contributes to our understanding of the comprehensive mechanisms of immune evasion by herpesviruses.

FAM177A1 Inhibits IL-1ß-Induced Signaling by Impairing TRAF6-Ubc13 Association.

The proinflammatory cytokine IL-1ß is a crucial mediator of inflammatory responses. IL-1ß-induced signaling is finely regulated by various mechanisms, and its imbalance is involved in a variety of diseases. In this study, we identified FAM177A1, a protein of unknown function, as a negative regulator of IL-1ß-induced signaling in human cells. Overexpression of FAM177A1 inhibited IL-1ß-triggered activation of NF-κB and transcription of inflammatory genes, whereas knockdown of FAM177A1 showed the opposite effects. Mechanistically, FAM177A1 competitively bound to the E3 ubiquitin ligase TRAF6 and impaired its interaction with the E2-conjugating enzyme Ubc13; therefore, it inhibited TRAF6-mediated polyubiquitination and recruitment of downstream signaling molecules. These findings reveal a function of FAM177A1 and promote our understanding of the regulatory mechanisms of IL-1ß-induced inflammatory responses.

[Evaluation of Lateral Pterygoid Muscle Contraction in Patients with Temporomandibular Disorders Based on 3D-T2 Weighted Imaging].

Objective To evaluate lateral pterygoid muscle(LPM)contraction in the patients with temporomandibular disorders(TMD)based on 3D-T2 weighted imaging(3D-T2WI).Multiplanar reconstruction(MPR)was employed to measure the length of LPM in the images taken in closed-and open-mouth positions. Methods Seventeen TMD patients [age of(29.82±10.70)years,males/females=8/9] and 13 normal volunteers [control,age of(23.54±3.31)years,males/females=6/7] received 3D-T2WI of the temporomandibular joints in closed-and open-mouth positions from November 2019 to April 2020 in Department of Radiology,Hainan Hospital of Chinese PLA General Hospital.According to the position of the discs,the subjects were classified into the following groups:TMD with disc displacement without reduction(TMD-DDwoR),TMD with disc displacement with reduction(TMD-DDwR),TMD without disc displacement(TMDwoDD),and normal control without disc displacement(NCwoDD).MPR was employed to measure the maximal length of the superior belly of LPM.One-way analysis of variance,receiver operating characteristic curve,and permutation test were employed for the statistical analyses. Results The contraction of LPM was significantly shorter in TMD-DDwoR group [(3.36±1.96)mm] than in TMDwoDD group [(7.90±3.95)mm],NCwoDD group [(8.77±3.13)mm](F=12.891,P=0.000),and TMD-DDwR group[(7.12±3.69)mm](χ2=5.314,P=0.031). Conclusion This study confirmed that the contraction of LPM decreased in patients with TMD-DDwoR,which provided imaging evidence for the study of disc displacement mechanism in TMD patients.

SARS-CoV-2 nucleocapsid protein impairs stress granule formation to promote viral replication.

The newly emerging coronavirus SARS-CoV-2 causes severe lung disease and substantial mortality. How the virus evades host defense for efficient replication is not fully understood. In this report, we found that the SARS-CoV-2 nucleocapsid protein (NP) impaired stress granule (SG) formation induced by viral RNA. SARS-CoV-2 NP associated with the protein kinase PKR after dsRNA stimulation. SARS-CoV-2 NP did not affect dsRNA-induced PKR oligomerization, but impaired dsRNA-induced PKR phosphorylation (a hallmark of its activation) as well as SG formation. SARS-CoV-2 NP also targeted the SG-nucleating protein G3BP1 and impaired G3BP1-mediated SG formation. Deficiency of PKR or G3BP1 impaired dsRNA-triggered SG formation and increased SARS-CoV-2 replication. The NP of SARS-CoV also targeted both PKR and G3BP1 to impair dsRNA-induced SG formation, whereas the NP of MERS-CoV targeted PKR, but not G3BP1 for the impairment. Our findings suggest that SARS-CoV-2 NP promotes viral replication by impairing formation of antiviral SGs, and reveal a conserved mechanism on evasion of host antiviral responses by highly pathogenic human betacoronaviruses.

SARS-CoV-2 membrane glycoprotein M antagonizes the MAVS-mediated innate antiviral response.

A novel SARS-related coronavirus (SARS-CoV-2) has recently emerged as a serious pathogen that causes high morbidity and substantial mortality. However, the mechanisms by which SARS-CoV-2 evades host immunity remain poorly understood. Here, we identified SARS-CoV-2 membrane glycoprotein M as a negative regulator of the innate immune response. We found that the M protein interacted with the central adaptor protein MAVS in the innate immune response pathways. This interaction impaired MAVS aggregation and its recruitment of downstream TRAF3, TBK1, and IRF3, leading to attenuation of the innate antiviral response. Our findings reveal a mechanism by which SARS-CoV-2 evades the innate immune response and suggest that the M protein of SARS-CoV-2 is a potential target for the development of SARS-CoV-2 interventions.

SRP54 Negatively Regulates IFN-Beta Production and Antiviral Response by Targeting RIG-I and MDA5.

During virus infection, RIG-I-like receptors (RLRs) recognize viral RNAs and recruit the adaptor protein VISA to activate downstream signaling, leading to activation of transcription factors NF-κB and IRF3, which collaborate to induce type I interferons (IFNs). IFNs further induce expression of hundreds of IFN-stimulated genes (ISGs) that suppress viral replication and facilitate the adaptive immune response. Dysregulated production of IFNs is implicated in various immune diseases. Here we identified Signal Recognition Particle 54 (SRP54) as a negative regulator of RLRs-induced antiviral signaling. Overexpression of SRP54 inhibited RNA virus-triggered induction of IFN-ß and increased viral replication, whereas knockdown of SRP54 had opposite effects. Mechanistically, SRP54 interacted with both RIG-I and MDA5 and impaired their association with VISA. Our findings demonstrate that SRP54 acts as a negative regulator of RLRs-mediated innate immune response by disrupting the recruitment of VISA to RIG-I/MDA5.

Histone deacetylase 3 promotes innate antiviral immunity through deacetylation of TBK1.

TANK-binding kinase 1 (TBK1), a core kinase of antiviral pathways, activates the production of interferons (IFNs). It has been reported that deacetylation activates TBK1; however, the precise mechanism still remains to be uncovered. We show here that during the early stage of viral infection, the acetylation of TBK1 was increased, and the acetylation of TBK1 at Lys241 enhanced the recruitment of IRF3 to TBK1. HDAC3 directly deacetylated TBK1 at Lys241 and Lys692, which resulted in the activation of TBK1. Deacetylation at Lys241 and Lys692 was critical for the kinase activity and dimerization of TBK1 respectively. Using knockout cell lines and transgenic mice, we confirmed that a HDAC3 null mutant exhibited enhanced susceptibility to viral challenge via impaired production of type I IFNs. Furthermore, activated TBK1 phosphorylated HDAC3, which promoted the deacetylation activity of HDAC3 and formed a feedback loop. In this study, we illustrated the roles the acetylated and deacetylated forms of TBK1 play in antiviral innate responses and clarified the post-translational modulations involved in the interaction between TBK1 and HDAC3.

Ubiquitination of TLR3 by TRIM3 signals its ESCRT-mediated trafficking to the endolysosomes for innate antiviral response.

Trafficking of toll-like receptor 3 (TLR3) from the endoplasmic reticulum (ER) to endolysosomes and its subsequent proteolytic cleavage are required for it to sense viral double-stranded RNA (dsRNA) and trigger antiviral response, yet the underlying mechanisms remain enigmatic. We show that the E3 ubiquitin ligase TRIM3 is mainly located in the Golgi apparatus and transported to the early endosomes upon stimulation with the dsRNA analog poly(I:C). TRIM3 mediates K63-linked polyubiquitination of TLR3 at K831, which is enhanced following poly(I:C) stimulation. The polyubiquitinated TLR3 is recognized and sorted by the ESCRT (endosomal sorting complex required for transport) complexes to endolysosomes. Deficiency of TRIM3 impairs TLR3 trafficking from the Golgi apparatus to endosomes and its subsequent activation. Trim3-/- cells and mice express lower levels of antiviral genes and show lower levels of inflammatory response following poly(I:C) but not lipopolysaccharide (LPS) stimulation. These findings suggest that TRIM3-mediated polyubiquitination of TLR3 represents a feedback-positive regulatory mechanism for TLR3-mediated innate immune and inflammatory responses.