Innate Immune Response to Cytoplasmic DNA: Mechanisms and Diseases.

DNA has been known to be a potent immune stimulus for more than half a century. However, the underlying molecular mechanisms of DNA-triggered immune response have remained elusive until recent years. Cyclic GMP-AMP synthase (cGAS) is a major cytoplasmic DNA sensor in various types of cells that detect either invaded foreign DNA or aberrantly located self-DNA. Upon sensing of DNA, cGAS catalyzes the formation of cyclic GMP-AMP (cGAMP), which in turn activates the ER-localized adaptor protein MITA (also named STING) to elicit the innate immune response. The cGAS-MITA axis not only plays a central role in host defense against pathogen-derived DNA but also acts as a cellular stress response pathway by sensing aberrantly located self-DNA, which is linked to the pathogenesis of various human diseases. In this review, we summarize the spatial and temporal mechanisms of host defense to cytoplasmic DNA mediated by the cGAS-MITA axis and discuss the association of malfunctions of this axis with autoimmune and other diseases.

Cytoplasmic Mechanisms of Recognition and Defense of Microbial Nucleic Acids.

Microbial nucleic acids are major signatures of invading pathogens, and their recognition by various host pattern recognition receptors (PRRs) represents the first step toward an efficient innate immune response to clear the pathogens. The nucleic acid-sensing PRRs are localized at the plasma membrane, the cytosol, and/or various cellular organelles. Sensing of nucleic acids and signaling by PRRs involve recruitment of distinct signaling components, and PRRs are intensively regulated by cellular organelle trafficking. PRR-mediated innate immune responses are also heavily regulated by posttranslational modifications, including phosphorylation, polyubiquitination, sumoylation, and glutamylation. In this review, we focus on our current understanding of recognition of microbial nucleic acid by PRRs, particularly on their regulation by organelle trafficking and posttranslational modifications. We also discuss how sensing of self nucleic acids and dysregulation of PRR-mediated signaling lead to serious human diseases.

A mitochondrial EglN1-AMPKα axis drives breast cancer progression by enhancing metabolic adaptation to hypoxic stress.

Mitochondria play essential roles in cancer cell adaptation to hypoxia, but the underlying mechanisms remain elusive. Through mitochondrial proteomic profiling, we here find that the prolyl hydroxylase EglN1 (PHD2) accumulates on mitochondria under hypoxia. EglN1 substrate-binding region in the ß2ß3 loop is responsible for its mitochondrial translocation and contributes to breast tumor growth. Furthermore, we identify AMP-activated protein kinase alpha (AMPKα) as an EglN1 substrate on mitochondria. The EglN1-AMPKα interaction is essential for their mutual mitochondrial translocation. After EglN1 prolyl-hydroxylates AMPKα under normoxia, they rapidly dissociate following prolyl-hydroxylation, leading to their immediate release from mitochondria. In contrast, hypoxia results in constant EglN1-AMPKα interaction and their accumulation on mitochondria, leading to the formation of a Ca2+ /calmodulin-dependent protein kinase 2 (CaMKK2)-EglN1-AMPKα complex to activate AMPKα phosphorylation, ensuring metabolic homeostasis and breast tumor growth. Our findings identify EglN1 as an oxygen-sensitive metabolic checkpoint signaling hypoxic stress to mitochondria through its ß2ß3 loop region, suggesting a potential therapeutic target for breast cancer.

The Zinc-Finger Protein ZCCHC3 Binds RNA and Facilitates Viral RNA Sensing and Activation of the RIG-I-like Receptors.

Recognition of viral RNA by the retinoic acid-inducible gene-I (RIG-I)-like receptors (RLRs) initiates innate antiviral immune response. How the binding of viral RNA to and activation of the RLRs are regulated remains enigmatic. In this study, we identified ZCCHC3 as a positive regulator of the RLRs including RIG-I and MDA5. ZCCHC3 deficiency markedly inhibited RNA virus-triggered induction of downstream antiviral genes, and ZCCHC3-deficient mice were more susceptible to RNA virus infection. ZCCHC3 was associated with RIG-I and MDA5 and functions in two distinct processes for regulation of RIG-I and MDA5 activities. ZCCHC3 bound to dsRNA and enhanced the binding of RIG-I and MDA5 to dsRNA. ZCCHC3 also recruited the E3 ubiquitin ligase TRIM25 to the RIG-I and MDA5 complexes to facilitate its K63-linked polyubiquitination and activation. Thus, ZCCHC3 is a co-receptor for RIG-I and MDA5, which is critical for RLR-mediated innate immune response to RNA virus.

Sumoylation Promotes the Stability of the DNA Sensor cGAS and the Adaptor STING to Regulate the Kinetics of Response to DNA Virus.

During viral infection, sensing of cytosolic DNA by the cyclic GMP-AMP synthase (cGAS) activates the adaptor protein STING and triggers an antiviral response. Little is known about the mechanisms that determine the kinetics of activation and deactivation of the cGAS-STING pathway, ensuring effective but controlled innate antiviral responses. Here we found that the ubiquitin ligase Trim38 targets cGas for sumoylation in uninfected cells and during the early phase of viral infection. Sumoylation of cGas prevented its polyubiquitination and degradation. Trim38 also sumoylated Sting during the early phase of viral infection, promoting both Sting activation and protein stability. In the late phase of infection, cGas and Sting were desumoylated by Senp2 and subsequently degraded via proteasomal and chaperone-mediated autophagy pathways, respectively. Our findings reveal an essential role for Trim38 in the innate immune response to DNA virus and provide insight into the mechanisms that ensure optimal activation and deactivation of the cGAS-STING pathway.

Endocytosis triggers V-ATPase-SYK-mediated priming of cGAS activation and innate immune response.

The current view of nucleic acid-mediated innate immunity is that binding of intracellular sensors to nucleic acids is sufficient for their activation. Here, we report that endocytosis of virus or foreign DNA initiates a priming signal for the DNA sensor cyclic GMP-AMP synthase (cGAS)-mediated innate immune response. Mechanistically, viral infection or foreign DNA transfection triggers recruitment of the spleen tyrosine kinase (SYK) and cGAS to the endosomal vacuolar H+ pump (V-ATPase), where SYK is activated and then phosphorylates human cGASY214/215 (mouse cGasY200/201) to prime its activation. Upon binding to DNA, the primed cGAS initiates robust cGAMP production and mediator of IRF3 activation/stimulator of interferon genes-dependent innate immune response. Consistently, blocking the V-ATPase-SYK axis impairs DNA virus- and transfected DNA-induced cGAMP production and expression of antiviral genes. Our findings reveal that V-ATPase-SYK-mediated tyrosine phosphorylation of cGAS following endocytosis of virus or other cargos serves as a priming signal for cGAS activation and innate immune response.

KAT5 acetylates cGAS to promote innate immune response to DNA virus.

The DNA sensor cGMP-AMP synthase (cGAS) senses cytosolic microbial or self DNA to initiate a MITA/STING-dependent innate immune response. cGAS is regulated by various posttranslational modifications at its C-terminal catalytic domain. Whether and how its N-terminal unstructured domain is regulated by posttranslational modifications remain unknown. We identified the acetyltransferase KAT5 as a positive regulator of cGAS-mediated innate immune signaling. Overexpression of KAT5 potentiated viral-DNA-triggered transcription of downstream antiviral genes, whereas a KAT5 deficiency had the opposite effects. Mice with inactivated Kat5 exhibited lower levels of serum cytokines in response to DNA virus infection, higher viral titers in the brains, and more susceptibility to DNA-virus-induced death. Mechanistically, KAT5 catalyzed acetylation of cGAS at multiple lysine residues in its N-terminal domain, which promoted its DNA-binding ability. Our findings suggest that KAT5-mediated cGAS acetylation at its N terminus is important for efficient innate immune response to DNA virus.

MARCH3 attenuates IL-1ß-triggered inflammation by mediating K48-linked polyubiquitination and degradation of IL-1RI.

The proinflammatory cytokine IL-1ß plays critical roles in inflammatory and autoimmune diseases. IL-1ß signaling is tightly regulated to avoid excessive inflammatory response. In this study, we identified the E3 ubiquitin ligase membrane-associated RING-CH-type finger 3 (MARCH3) as a critical negative regulator of IL-1ß-triggered signaling. Overexpression of MARCH3 inhibited IL-1ß-triggered activation of NF-κB as well as expression of inflammatory genes, whereas MARCH3 deficiency had the opposite effects. MARCH3-deficient mice produced higher levels of serum inflammatory cytokines and were more sensitive to inflammatory death upon IL-1ß injection or Listeria monocytogenes infection. Mechanistically, MARCH3 was associated with IL-1 receptor I (IL-1RI) and mediated its K48-linked polyubiquitination at K409 and lysosomal-dependent degradation. Furthermore, IL-1ß stimulation triggered dephosphorylation of MARCH3 by CDC25A and activation of its E3 ligase activity. Our findings suggest that MARCH3-mediated IL-1RI degradation is an important mechanism for attenuating IL-1ß-triggered inflammatory response.

CSK promotes innate immune response to DNA virus by phosphorylating MITA.

Upon detection of viral DNA, the cytoplasmic DNA sensor cyclic GMP-AMP (cGAMP) synthase (cGAS) utilizes GTP and ATP as substrates to synthesize the second messenger molecule 2'3'cyclic GMP-AMP (cGAMP), which binds to the ER-associated adaptor protein MITA/STING to signal innate antiviral response to DNA virus. How the cGAS-MITA pathways are post-translationally regulated is not fully understood. In this study, we identified the tyrosine kinase CSK as a positive regulator of cGAS-MITA mediated innate antiviral response. CSK-deficiency inhibits DNA virus-triggered induction of downstream antiviral effector genes. Following DNA virus infection, CSK phosphorylates MITA at Y240 and Y245, which is important for its activation. These results suggest that CSK plays a role in modulating innate immune response to DNA virus.

SNX8 modulates innate immune response to DNA virus by mediating trafficking and activation of MITA.

MITA (also called STING) is a central adaptor protein in innate immune response to cytosolic DNA. Cellular trafficking of MITA from the ER to perinuclear microsomes after DNA virus infection is critical for MITA activation and onset of innate antiviral response. Here we found that SNX8 is a component of DNA-triggered induction of downstream effector genes and innate immune response. Snx8-/- mice infected with the DNA virus HSV-1 exhibited lower serum cytokine levels and higher viral titers in the brains, resulting in higher lethality. Mechanistically, SNX8 recruited the class III phosphatylinositol 3-kinase VPS34 to MITA, which is required for trafficking of MITA from the ER to perinuclear microsomes. Our findings suggest that SNX8 is a critical component in innate immune response to cytosolic DNA and DNA virus.

[Psychological care combined with enhanced recovery after surgery management in perioperative nursing care of andrological patients: A randomized controlled study].

OBJECTIVE: To evaluate the validity of psychological care combined with enhanced recovery after surgery (PC+ERAS) management in perioperative nursing care of andrological patients. METHODS: A total of 300 male patients undergoing andrological surgery were included in this study, 150 given PC+ERAS and the other 150 receiving routine nursing care as controls. We evaluated anxiety and depression of all the patients on admission and discharge using Self-Rating Anxiety Scale (SAS) and Self-Rating Depression Scale (SDS), and compared post-operative hospital days, off-bed time, first passage of flatus, Visual Analog Scale (VAS) score and satisfaction with nursing care between the two groups of patients. RESULTS: On discharge, significant improvement was observed in SAS and SDS scores in the PC+ERAS group compared with the baseline, even more significant than in the control group (P < 0.01), but no obvious improvement was seen in the controls (P > 0.05). The patients in the PC+ERAS group also achieved a significantly shorter post-operative hospital stay, earlier post-operative off-bed time and passage of flatus, lower VAS score, and higher satisfaction with nursing care than those in the control group (P < 0.05). CONCLUSIONS: Psychological care combined with ERAS management deserves wide application in the perioperative nursing care of andrological patients, which can significantly improve the patients' anxiety and depression, shorten post-operative hospital stay, reduce VAS score, and increase their satisfaction with nursing care.

PKACs attenuate innate antiviral response by phosphorylating VISA and priming it for MARCH5-mediated degradation.

Sensing of viral RNA by RIG-I-like receptors initiates innate antiviral response, which is mediated by the central adaptor VISA. How the RIG-I-VISA-mediated antiviral response is terminated at the late phase of infection is enigmatic. Here we identified the protein kinase A catalytic (PKAC) subunits α and ß as negative regulators of RNA virus-triggered signaling in a redundant manner. Viral infection up-regulated cellular cAMP levels and activated PKACs, which then phosphorylated VISA at T54. This phosphorylation abrogated virus-induced aggregation of VISA and primed it for K48-linked polyubiquitination and degradation by the E3 ligase MARCH5, leading to attenuation of virus-triggered induction of downstream antiviral genes. PKACs-deficiency or inactivation by the inhibitor H89 potentiated innate immunity to RNA viruses in cells and mice. Our findings reveal a critical mechanism of attenuating innate immune response to avoid host damage at the late phase of viral infection by the house-keeping PKA kinase.

TRIM32-TAX1BP1-dependent selective autophagic degradation of TRIF negatively regulates TLR3/4-mediated innate immune responses.

Toll-like receptor (TLR)-mediated signaling are critical for host defense against pathogen invasion. However, excessive responses would cause harmful damages to the host. Here we show that deficiency of the E3 ubiquitin ligase TRIM32 increases poly(I:C)- and LPS-induced transcription of downstream genes such as type I interferons (IFNs) and proinflammatory cytokines in both primary mouse immune cells and in mice. Trim32-/- mice produced higher levels of serum inflammatory cytokines and were more sensitive to loss of body weight and inflammatory death upon Salmonella typhimurium infection. TRIM32 interacts with and mediates the degradation of TRIF, a critical adaptor protein for TLR3/4, in an E3 activity-independent manner. TRIM32-mediated as well as poly(I:C)- and LPS-induced degradation of TRIF is inhibited by deficiency of TAX1BP1, a receptor for selective autophagy. Furthermore, TRIM32 links TRIF and TAX1BP1 through distinct domains. These findings suggest that TRIM32 negatively regulates TLR3/4-mediated immune responses by targeting TRIF to TAX1BP1-mediated selective autophagic degradation.

TRIM8 Negatively Regulates TLR3/4-Mediated Innate Immune Response by Blocking TRIF-TBK1 Interaction.

TLR-mediated signaling pathways play critical roles in host defense against microbials. However, dysregulation of innate immune and inflammatory responses triggered by TLRs would result in harmful damage to the host. Using a Trim8 gene-knockout mouse model, we show that tripartite motif (TRIM) 8 negatively regulates TLR3- and TLR4-mediated innate immune and inflammatory responses. TRIM8 deficiency leads to increased polyinosinic-polycytidylic acid- and LPS-triggered induction of downstream anti-microbial genes including TNF, Il6, Rantes, and Ifnb, evaluated serum cytokine levels, and increased susceptibility of mice to polyinosinic-polycytidylic acid- and LPS-induced inflammatory death as well as Salmonella typhimurium infection-induced loss of body weight and septic shock. TRIM8 interacted with Toll/IL-1 receptor domain-containing adapter-inducing IFN-ß and mediated its K6- and K33-linked polyubiquitination, leading to disruption of the Toll/IL-1 receptor domain-containing adapter-inducing IFN-ß-TANK-binding kinase-1 association. Our findings uncover an additional mechanism on the termination of TLR3/4-mediated inflammatory and innate immune responses.

Quantitative Proteomics Identified TTC4 as a TBK1 Interactor and a Positive Regulator of SeV-Induced Innate Immunity.

TBK1, STING, and MDA5 are important players within the antiviral innate immune response network. We mapped the interactome of endogenous TBK1, STING, and MDA5 by affinity enrichment MS in virally infected or uninfected THP-1 cells. Based on quantitative data of more than 2000 proteins and stringent statistical analysis, 58 proteins were identified as high-confidence interactors for at least one of three bait proteins. Our data indicated that TBK1 and MDA5 mostly interacted within preexisting protein networks, while STING interacted with different proteins with different viral infections. Functional analysis was performed on 17 interactors, and six were found to have functions in innate immune responses. We identified TTC4 as a TBK1 interactor and positive regulator of sendai virus-induced innate immunity.

MSX1 Modulates RLR-Mediated Innate Antiviral Signaling by Facilitating Assembly of TBK1-Associated Complexes.

Recognition of viral dsRNA by the retinoic acid-inducible gene-1-like receptors (RLRs) triggers signaling cascades that lead to activation of the TBK1 kinase and transcription factor IFN regulatory factor 3, induction of downstream antiviral genes, and innate antiviral responses. In this study, we identified muscle segment homeobox1 (MSX1) as an important modulator of RLR-mediated signaling pathways. Knockdown or knockout of MSX1 significantly impaired Sendai virus-triggered activation of TBK1 and IFN regulatory factor 3, induction of downstream antiviral genes, and cellular antiviral responses. Interestingly, MSX1 was translocated from the nucleus to cytoplasm, particularly mitochondria upon infection of Sendai virus. Biochemcially, MSX1 was important for assembly of TBK1/IKK-related kinase-associated protein 1/TNFR-associated factor-associated NF-κB activator complexes. Our results suggest that MSX1 is an important component of RLR-mediated signaling and reveal mechanisms on innate immune responses against RNA viruses.

[Unusual Isolated Sellar Spindle Cell Carcinoma:Report of One Case].

Sellar malignant tumors are uncommon and usually reported as metastatic diseases from breast or lung cancers. Spindle cell carcinoma (SCC) is a rare malignancy and has been found in breast,oral cavity,lungs,kidneys,and hepatobiliary pancreatic system but not in sellar region. We report here the first case of isolated sellar SCC with aggressive features in Peking Union Medical College Hospital. This patient was referred to our hospital on September 9,2015 and discharged on October 16,2015. We described the clinical manifestations,imaging findings,and pathological features of this rare disease.

Monitoring and Control of an Adsorption System Using Electrical Properties of the Adsorbent for Organic Compound Abatement.

Adsorption systems typically need gas and temperature sensors to monitor their adsorption/regeneration cycles to separate gases from gas streams. Activated carbon fiber cloth (ACFC)-electrothermal swing adsorption (ESA) is an adsorption system that has the potential to be controlled with the electrical properties of the adsorbent and is studied here to monitor and control the adsorption/regeneration cycles without the use of gas and temperature sensors and to predict breakthrough before it occurs. The ACFC's electrical resistance was characterized on the basis of the amount of adsorbed organic gas/vapor and the adsorbent temperature. These relationships were then used to develop control logic to monitor and control ESA cycles on the basis of measured resistance and applied power values. Continuous sets of adsorption and regeneration cycles were performed sequentially entirely on the basis of remote electrical measurements and achieved ≥95% capture efficiency at inlet concentrations of 2000 and 4000 ppmv for isobutane, acetone, and toluene in dry and elevated relative humidity gas streams, demonstrating a novel cyclic ESA system that does not require gas or temperature sensors. This contribution is important because it reduces the cost and simplifies the system, predicts breakthrough before its occurrence, and reduces emissions to the atmosphere.

TRIM38 Negatively Regulates TLR3/4-Mediated Innate Immune and Inflammatory Responses by Two Sequential and Distinct Mechanisms.

Tripartite motif (TRIM)38 is an E3 ubiquitin ligase that was reported to regulate signaling in innate immune and inflammatory responses in certain cell lines. In this study, we show that Trim38 deficiency markedly increased TLR3- and TLR4-mediated induction of type I IFNs and proinflammatory cytokines, such as TNF-α, IL-1ß, and IL-6, in immune cells and in vivo. Trim38 deficiency also caused the mice to be more susceptible to death triggered by polyinosinic-polycytidylic acid, LPS, and Salmonella typhimurium. Mechanistically, TRIM38 catalyzed K48-linked polyubiquitination of the TLR3/4 adapter protein TIR domain-containing adapter-inducing IFN-ß at K228 and promoted its proteasomal degradation in immune cells. Moreover, Trim38 was highly induced by type I IFNs, which then negatively regulated TNF-α/IL-1ß signaling in IFN-ß-primed immune cells, but not unprimed immune cells, by mediating degradation of Tab2 in a lysosomal-dependent process. These results suggest that Trim38 negatively regulates TLR3/4-mediated innate immune and inflammatory responses by two sequential and distinct mechanisms. This study increases our understanding of how the innate immune response is initiated during the early phase of infection to defend against microbial invasion and is efficiently terminated during the late phase to prevent excessive and harmful inflammatory responses.

TRIM38 inhibits TNFα- and IL-1ß-triggered NF-κB activation by mediating lysosome-dependent degradation of TAB2/3.

TNFα and IL-1ß are two proinflammatory cytokines that play critical roles in many diseases, including rheumatoid arthritis and infectious diseases. How TNFα- and IL-1ß-mediated signaling is finely tuned is not fully elucidated. Here, we identify tripartite-motif protein 38 (TRIM38) as a critical negative regulator of TNFα- and IL-1ß-triggered signaling. Overexpression of TRIM38 inhibited activation of NF-κB and induction of downstream cytokines following TNFα and IL-1ß stimulation, whereas knockdown or knockout of TRIM38 had the opposite effects. TRIM38 constitutively interacted with critical components TGF-ß-activated kinase 1 (TAK1)-binding protein 2/3 (TAB2/3) and promoted lysosome-dependent degradation of TAB2/3 independent of its E3 ubiquitin ligase activity. Consistently, deficiency of TRIM38 resulted in abolished translocation of TAB2 to the lysosome, increased level of TAB2 in cells, and enhanced activation of TAK1 after TNFα and IL-1ß stimulation. We conclude that TRIM38 negatively regulates TNFα- and IL-1ß-induced signaling by mediating lysosome-dependent degradation of TAB2/3, two critical components in TNFα- and IL-1ß-induced signaling pathways. Our findings reveal a previously undiscovered mechanism by which cells keep the inflammatory response in check to avoid excessive harmful immune response triggered by TNFα and IL-1ß.