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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. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Cyclic GMP-AMP synthase (cGAS) senses double-stranded DNA and synthesizes the second messenger cyclic GMP-AMP (cGAMP), which binds to mediator of IRF3 activation (MITA) and initiates MITA-mediated signaling, leading to induction of type I interferons (IFNs) and other antiviral effectors. Human cytomegalovirus (HCMV), a widespread and opportunistic pathogen, antagonizes the host antiviral immune response to establish latent infection. Here, we identified HCMV tegument protein UL94 as an inhibitor of the cGAS-MITA-mediated antiviral response. Ectopic expression of UL94 impaired cytosolic double-stranded DNA (dsDNA)- and DNA virustriggered induction of type I IFNs and enhanced viral replication. Conversely, UL94 deficiency potentiated HCMV-induced transcription of type I IFNs and downstream antiviral effectors and impaired viral replication. UL94 interacted with MITA, disrupted the dimerization and translocation of MITA, and impaired the recruitment of TBK1 to the MITA signalsome. These results suggest that UL94 plays an important role in the immune evasion of HCMV. IMPORTANCE Human cytomegalovirus (HCMV), a large double-stranded DNA (dsDNA) virus, encodes more than 200 viral proteins. HCMV infection causes irreversible abnormalities of the central nervous system in newborns and severe syndromes in organ transplantation patients or AIDS patients. It has been demonstrated that HCMV has evolved multiple immune evasion strategies to establish latent infection. Previous studies pay more attention to the mechanism by which HCMV evades immune response in the early phase of infection. In this study, we identified UL94 as a negative regulator of the innate immune response, which functions in the late phase of HCMV infection. [ABSTRACT FROM AUTHOR]

STING plays central roles in the innate immune response to pathogens that contain DNA. Sensing cytoplasmic DNA by cyclic GMP-AMP synthase produces cyclic GMP-AMP, which binds to and activates STING and induces STING translocation from the endoplasmic reticulum to the perinuclear microsome. However, this trafficking process has not been fully elucidated yet. In this study, we identified YIPF5 as a positive regulator of STING trafficking. YIPF5 is essential for DNA virus- or intracellular DNA-triggered production of type I IFNs. Consistently, knockdown of YIPF5 impairs cellular antiviral responses to DNA virus. Mechanistically, YIPF5 interacts with both STING and components of COPII, facilitating STING recruitment to COPII in the presence of cytoplasmic dsDNA. Furthermore, knockdown of components of COPII inhibits DNA virus-triggered production of type I IFNs, suggesting that COPII is involved in innate immune responses to DNA viruses. Collectively, our findings demonstrate that YIPF5 positively regulates STING-mediated innate immune responses by recruiting STING to COPII-coated vesicles and facilitating STING trafficking from the endoplasmic reticulum to Golgi, providing important insights into the molecular mechanisms of intracellular DNA-stimulated STING trafficking and activation. [ABSTRACT FROM AUTHOR]

Innate immunity is the first line of host defense against viral invasion. The inductions of type I interferons (IFNs) and inflammatory cytokine are essential to host antiviral immune responses, which are also key targets of viral immune evasion. Human cytomegalovirus (HCMV) can establish long-term latent infection, in which immune evasion is a pivotal step. In this study, we identified HCMV protein UL44, a DNA polymerase processivity factor, as an inhibitor of IRF3- and NF-ĸB-dependent antiviral response. Ectopic expression of UL44 inhibited HCMV-triggered induction of downstream effector genes and enhanced viral replication. Conversely, knockdown of UL44 potentiated HCMV-triggered induction of downstream antiviral genes. UL44 interacted with IRF3 and p65, and inhibited binding of IRF3 and NF-ĸB to the promoters of their downstream antiviral genes. These findings reveal an important mechanism of immune evasion by HCMV at the transcriptional level. Importance Inductions of Type I IFNs and inflammatory cytokines play pivotal roles in host antiviral innate immune responses. Viruses have evolved various mechanisms to interfere with these processes. HCMV causes severe ailments in immunodeficient populations and is a major cause of birth defects. It has been shown that HCMV antagonizes host innate immune defenses, which is important for establishing immune evasion and latent infection. In this study, we identified the HCMV DNA polymerase subunit UL44 as a suppressor of antiviral innate immune responses. Overexpression of UL44 impaired HCMV-triggered induction of type I IFNs and other antiviral genes thus potentiated viral replication, whereas UL44-deficiency showed opposite effects. Mechanistic studies indicated that UL44 acts by inhibiting the binding of IRF3 and NF-ĸB to the promoters of downstream antiviral genes. These findings defined an important mechanism of HCMV immune evasion at the transcriptional level, which may provide a therapeutic target for the treatment of HCMV infection. [ABSTRACT FROM AUTHOR]

STAT3 is a transcription factor that plays central roles in various physiological processes, including differentiation of Th cells. Its deregulation results in serious diseases, including inflammatory diseases and cancer. The mechanisms related to how STAT3 activity is regulated remain enigmatic. Here we show that overexpression of FAM64A potentiates IL-6-induced activation of STAT3 and expression of downstream target genes, whereas deficiency of FAM64A has the opposite effects. FAM64A interactswith STAT3 in the nucleus and regulates binding of STAT3 to the promoters of its target genes. Deficiency of Fam64a significantly impairs differentiation of Th17 but not Th1 or induced regulatory T cells (iTreg). In addition, Fam64a deficiency attenuates experimental autoimmune encephalomyelitis (EAE) and dextran sulfate sodium (DSS)-induced colitis, which is correlated with decreased differentiation of Th17 cells and production of proinflammatory cytokines. Furthermore, Fam64a deficiency suppresses azoxymethane (AOM)/DSS-induced colitis-associated cancer (CAC) in mice. These findings suggest that FAM64A regulates Th17 differentiation and colitis and inflammation-associated cancer by modulating transcriptional activity of STAT3. [ABSTRACT FROM AUTHOR]

Ginsenoside Rg3 (Rg3) is one of the primary constituents isolated from ginseng, and has been found to exhibit cytotoxic effects against cancer cells. The present study aimed to investigate the effects of Rg3 on human multiple myeloma cell proliferation and apoptosis, and to examine its underlying molecular mechanisms. Cell viability was detected using a Cell Counting kit-8 assay, and cell cycle arrest and cell apoptosis were analyzed using flow cytometry. In addition, the expression levels of cell cycle-associated markers and apoptosis-associated proteins, and the release of cytochrome C were determined using western blot analysis. The effects of Rg3 on the insulin-like growth factor (IGF)-1/AKT/mammalian target of rapamycin (mTOR) and mitogen-activated protein kinase signaling pathways were also investigated using western blot analysis. The results showed that Rg3 inhibited cell viability in U266, RPMI8226 and SKO-007 cells in a time- and dose-dependent manner, and caused cell cycle arrest in the G1 phase by regulating the cyclin-dependent kinase pathway. Furthermore, Rg3 induced multiple myeloma cell apoptosis, and was involved in B cell lymphoma-2 (Bcl2)/Bcl2-associated X protein imbalance, caspase activation and the release of cytochrome C from the mitochondria into the cytoplasm. Mechanistically, it was found that the inhibitory effects of Rg3 on multiple myeloma cell proliferation were essential for secretion of IGF-1 and inactivation of the Akt/mTOR pathway. Collectively, these findings demonstrated that Rg3 effectively inhibited cell proliferation and induced apoptosis of multiple myeloma cells. These data broaden the clinical investigation of Rg3 in the treatment of multiple myeloma, associated with the inactivation of IGF-1/AKT/mTOR signaling. [ABSTRACT FROM AUTHOR]