B-Cell-Epitope-Based Fluorescent Quantum Dot Biosensors for SARS-CoV-2 Enable Highly Sensitive COVID-19 Antibody Detection.

A new antibody diagnostic assay with more rapid and robust properties is demanded to quantitatively evaluate anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) immunity in a large population. Here, we developed a nanometer-scale fluorescent biosensor system consisting of CdSe-ZnS quantum dots (QDs) coupled with the highly sensitive B-cell epitopes of SARS-CoV-2 that could remarkably identify the corresponding antibody with a detection limit of 100 pM. Intriguingly, we found that fluorescence quenching of QDs was stimulated more obviously when coupled with peptides than the corresponding proteins, indicating that the energy transfer between QDs and peptides was more effective. Compared to the traditional enzyme-linked immunosorbent assay (ELISA), the B-cell-epitope-based QD-biosensor could robustly distinguish coronavirus disease 2019 (COVID-19) antibody-positive patients from uninfected individuals with a higher sensitivity (92.3-98.1% positive rates by QD-biosensor vs. 78.3-83.1% positive rates by ELISAs in 207 COVID-19 patients' sera) in a more rapid (5 min) and labor-saving manner. Taken together, the 'QD-peptides' biosensor provided a novel real-time, quantitative, and high-throughput method for clinical diagnosis and home-use tests.

Structural and Functional Insights into a Nonheme Iron- and α-Ketoglutarate-Dependent Halogenase That Catalyzes Chlorination of Nucleotide Substrates.

Nonheme iron- and α-ketoglutarate (αKG)-dependent halogenases (NHFeHals), which catalyze the regio- and stereoselective halogenation of the unactivated C(sp3)-H bonds, exhibit tremendous potential in the challenging asymmetric halogenation. AdeV from Actinomadura sp. ATCC 39365 is the first identified carrier protein-free NHFeHal that catalyzes the chlorination of nucleotide 2'-deoxyadenosine-5'-monophosphate (2'-dAMP) to afford 2'-chloro-2'-deoxyadenosine-5'-monophosphate. Here, we determined the complex crystal structures of AdeV/FeII/Cl and AdeV/FeII/Cl/αKG at resolutions of 1.76 and 1.74 Å, respectively. AdeV possesses a typical ß-sandwich topology with H194, H252, αKG, chloride, and one water molecule coordinating FeII in the active site. Molecular docking, mutagenesis, and biochemical analyses reveal that the hydrophobic interactions and hydrogen bond network between the substrate-binding pocket and the adenine, deoxyribose, and phosphate moieties of 2'-dAMP are essential for substrate recognition. Residues H111, R177, and H192 might play important roles in the second-sphere interactions that control reaction partitioning. This study provides valuable insights into the catalytic selectivity of AdeV and will facilitate the rational engineering of AdeV and other NHFeHals for synthesis of halogenated nucleotides. IMPORTANCE Halogenated nucleotides are a group of important antibiotics and are clinically used as antiviral and anticancer drugs. AdeV is the first carrier protein-independent nonheme iron- and α-ketoglutarate (αKG)-dependent halogenase (NHFeHal) that can selectively halogenate nucleotides and exhibits restricted substrate specificity toward several 2'-dAMP analogues. Here, we determined the complex crystal structures of AdeV/FeII/Cl and AdeV/FeII/Cl/αKG. Molecular docking, mutagenesis, and biochemical analyses provide important insights into the catalytic selectivity of AdeV. This study will facilitate the rational engineering of AdeV and other carrier protein-independent NHFeHals for synthesis of halogenated nucleotides.

[Corrigendum] Nogo­B promotes the epithelial­mesenchymal transition in HeLa cervical cancer cells via Fibulin­5.

Following the publication of the above article, an interested reader drew to the authors' attention that the 'NB­4' and 'NB­2' panels for the invasion and migration assays shown in Fig. 3B and C on p. 113 appeared to contain overlapping data, such that the data may have been derived from the same original source, even though the panels were intending to have shown results obtained under different experimental conditions. The authors have re­examined their raw data and realized that these data were inadvertently mixed up when Fig. 3B and C were assembled. A corrected version of Fig. 3, showing the data as they should have appeared for the 'NB­4' and 'NB­2' invasion and migration assay experiments in Fig. 3B and C, is shown on the next page. The authors sincerely apologize for the errors that were introduced into Fig. 3 of the published article, and thank the Editor of Oncology Reports for allowing them the opportunity to publish a Corrigendum. All the authors agree to the publication of the authors, and they apologize to the readership for any inconvenience caused. [Oncology Reports 29: 109­116, 2013; DOI: 10.3892/or.2012.2069].

Human adenoviruses: A suspect behind the outbreak of acute hepatitis in children amid the COVID-19 pandemic.

As of 10 May 2022, at least 450 cases of pediatric patients with acute hepatitis of unknown cause have been reported worldwide. Human adenoviruses (HAdVs) have been detected in at least 74 cases, including the F type HAdV41 in 18 cases, which indicates that adenoviruses may be associated with this mysterious childhood hepatitis, although other infectious agents or environmental factors cannot be excluded. In this review, we provide a brief introduction of the basic features of HAdVs and describe diseases caused by different HAdVs in humans, aiming to help understand the biology and potential risk of HAdVs and cope with the outbreak of acute child hepatitis.

Etiological, epidemiological, and clinical features of acute diarrhea in China.

National-based prospective surveillance of all-age patients with acute diarrhea was conducted in China between 2009‒2018. Here we report the etiological, epidemiological, and clinical features of the 152,792 eligible patients enrolled in this analysis. Rotavirus A and norovirus are the two leading viral pathogens detected in the patients, followed by adenovirus and astrovirus. Diarrheagenic Escherichia coli and nontyphoidal Salmonella are the two leading bacterial pathogens, followed by Shigella and Vibrio parahaemolyticus. Patients aged <5 years had higher overall positive rate of viral pathogens, while bacterial pathogens were more common in patients aged 18‒45 years. A joinpoint analysis revealed the age-specific positivity rate and how this varied for individual pathogens. Our findings fill crucial gaps of how the distributions of enteropathogens change across China in patients with diarrhea. This allows enhanced identification of the predominant diarrheal pathogen candidates for diagnosis in clinical practice and more targeted application of prevention and control measures.

The TLR3/IRF1/Type III IFN Axis Facilitates Antiviral Responses against Enterovirus Infections in the Intestine.

Enteroviruses infect gastrointestinal epithelium cells, cause multiple human diseases, and present public health risks worldwide. However, the mechanisms underlying host immune responses in intestinal mucosa against the early enterovirus infections remain elusive. Here, we showed that human enteroviruses including enterovirus 71 (EV71), coxsackievirus B3 (CVB3), and poliovirus 1 (PV1) predominantly induce type III interferons (IFN-λ1 and IFN-λ2/3), rather than type I interferons (IFN-α and IFN-ß), in cultured human normal and cancerous intestine epithelial cells (IECs), mouse intestine tissues, and human clinical intestine specimens. Mechanistic studies demonstrated that IFN-λ production is induced upon enterovirus infection through the Toll-like receptor 3/interferon regulatory factor 1 (TLR3/IRF1) signaling pathway in IECs. In turn, the supplementation of IFN-λ subsequently induces intrinsically antiviral responses against enterovirus replication. Notably, intraperitoneal injection in neonatal C57BL/6J mice with mouse recombinant IFN-λ2 protein represses EV71 replication and protects mice from viral lethal effects. Altogether, these results revealed a distinct mechanism by which the host elicited immune responses against enterovirus infections in intestine through activating the TLR3/IRF1/type III IFN axis. The new findings would provide an antiviral strategy for the prevention and treatment of enterovirus infections and associated diseases.IMPORTANCE Enterovirus infections are significant sources of human diseases and public health risks worldwide, but little is known about the mechanism of innate immune response in host intestine epithelial surface during the viral replication. We reported the epithelial immune response in cultured human normal and cancerous cells (IECs), mouse tissues, and human clinical intestine specimens following infection with enterovirus 71. The results mechanistically revealed type III interferons (IFN-λ1 and IFN-λ2/3), rather than type I interferons (IFN-α and IFN-ß), as the dominant production through TLR3/IRF1 signaling upon multiple human enterovirus infection, including enterovirus 71 (EV71), coxsackievirus B3 (CVB3), and poliovirus 1 (PV1). IFN-λ subsequently induced antiviral activity against enterovirus replication in vitro and in vivo. These studies uncovered the role of the novel process of type III IFN production involved in the TLR3/IRF1 pathway in host intestine upon enterovirus infection, which highlighted a regulatory manner of antiviral defense in intestine during enterovirus infection.

Paxillin mediates ATP-induced activation of P2X7 receptor and NLRP3 inflammasome.

BACKGROUND: Extracellular adenosine triphosphate (ATP), a key danger-associated molecular pattern (DAMP) molecule, is released to the extracellular medium during inflammation by injured parenchymal cells, dying leukocytes, and activated platelets. ATP directly activates the plasma membrane channel P2X7 receptor (P2X7R), leading to an intracellular influx of K+, a key trigger inducing NLRP3 inflammasome activation. However, the mechanism underlying P2X7R-mediated activation of NLRP3 inflammasome is poorly understood, and additional molecular mediators have not been identified. Here, we demonstrate that Paxillin is the molecule connecting the P2X7 receptor and NLRP3 inflammasome through protein interactions. RESULTS: We show a distinct mechanism by which Paxillin promotes ATP-induced activation of the P2X7 receptor and NLRP3 inflammasome. Extracellular ATP induces Paxillin phosphorylation and then facilitates Paxillin-NLRP3 interaction. Interestingly, Paxillin enhances NLRP3 deubiquitination and activates NLRP3 inflammasome upon ATP treatment and K+ efflux. Moreover, we demonstrated that USP13 is a key enzyme for Paxillin-mediated NLRP3 deubiquitination upon ATP treatment. Notably, extracellular ATP promotes Paxillin and NLRP3 migration from the cytosol to the plasma membrane and facilitates P2X7R-Paxillin interaction and PaxillinNLRP3 association, resulting in the formation of the P2X7R-Paxillin-NLRP3 complex. Functionally, Paxillin is essential for ATP-induced NLRP3 inflammasome activation in mouse BMDMs and BMDCs as well as in human PBMCs and THP-1-differentiated macrophages. CONCLUSIONS: We have identified paxillin as a mediator of NLRP3 inflammasome activation. Paxillin plays key roles in ATP-induced activation of the P2X7 receptor and NLRP3 inflammasome by facilitating the formation of the P2X7R-Paxillin-NLRP3 complex.

MYSM1 Represses Innate Immunity and Autoimmunity through Suppressing the cGAS-STING Pathway.

The immune system is not only required for preventing threats exerted by pathogens but also essential for developing immune tolerance to avoid tissue damage. This study identifies a distinct mechanism by which MYSM1 suppresses innate immunity and autoimmunity. The expression of MYSM1 is induced upon DNA virus infection and by intracellular DNA stimulation. MYSM1 subsequently interacts with STING and cleaves STING K63-linked ubiquitination to suppress cGAS-STING signaling. Notably, Mysm1-deficient mice exhibit a hyper-inflammatory response, acute tissue damage, and high mortality upon virus infection. Moreover, in the PBMCs of patients with systemic lupus erythematosus (SLE), MYSM1 production decreases, while type I interferons and pro-inflammatory cytokine expressions increase. Importantly, MYSM1 treatment represses the production of IFNs and pro-inflammatory cytokines in the PBMCs of SLE patients. Thus, MYSM1 is a critical repressor of innate immunity and autoimmunity and is thus a potential therapeutic agent for infectious, inflammatory, and autoimmune diseases.

miR-4454 up-regulated by HPV16 E6/E7 promotes invasion and migration by targeting ABHD2/NUDT21 in cervical cancer.

The abnormal expression of HPV16 E6/E7 activates oncogenes and/or inactivates tumor suppressor genes, resulting in the selective growth and malignant transformation of cancer cells. miR-4454 was selected by sequencing due to its abnormal high expression in HPV16 E6/E7 positive CaSki cell compared with HPV16 E6/E7 negative C33A cell. Overexpression of miR-4454 enhances cervical cancer cell invasion and migration. ABHD2 and NUDT21 are identified as a target gene of miR-4454.The effects of ABHD2 and NUDT21 on migration and invasion of CaSki and C33A cells were determined. The dual luciferase and RT-qPCR assays confirmed that miR-4454 might regulate its targets ABHD2 and NUDT21 to promote the proliferation, invasion and migration, whereas, inhibit the apoptosis in CaSki and C33A cells.

Enterovirus 71 induces neural cell apoptosis and autophagy through promoting ACOX1 downregulation and ROS generation.

Enterovirus 71 (EV71) infection causes hand, foot, and mouth disease (HFMD), and even fatal neurological complications. However, the mechanisms underlying EV71 neurological pathogeneses are largely unknown. This study reveals a distinct mechanism by which EV71 induces apoptosis and autophagy in neural cells. EV71 non-structure protein 3D (also known as RNA-dependent RNA polymerase, RdRp) interacts with the peroxisomal protein acyl-CoA oxidase 1 (ACOX1), and contributes to ACOX1 downregulation. Further studies demonstrate that EV71 reduces peroxisome numbers. Additionally, knockdown of ACOX1 or peroxin 19 (PEX19) induces apoptosis and autophagy in neural cells including human neuroblastoma (SK-N-SH) cells and human astrocytoma (U251) cells, and EV71 infection induces neural cell death through attenuating ACOX1 production. Moreover, EV71 infection and ACOX1 knockdown facilitate reactive oxygen species (ROS) production and attenuate the cytoprotective protein deglycase (DJ-1)/Nuclear factor erythroid 2-related factor 2 (NRF2)/Heme oxygenase 1 (HO-1) pathway (DJ-1/NRF2/HO-1), which collectively result in ROS accumulation in neural cells. In conclusion, EV71 downregulates ACOX1 protein expression, reduces peroxisome numbers, enhances ROS generation, and attenuates the DJ-1/NRF2/HO-1 pathway, thereby inducing apoptosis and autophagy in neural cells. These findings provide new insights into the mechanism underlying EV71-induced neural pathogenesis, and suggest potential treatments for EV71-associated diseases.

Transcriptomic characteristics of bronchoalveolar lavage fluid and peripheral blood mononuclear cells in COVID-19 patients.

Circulating in China and 158 other countries and areas, the ongoing COVID-19 outbreak has caused devastating mortality and posed a great threat to public health. However, efforts to identify effectively supportive therapeutic drugs and treatments has been hampered by our limited understanding of host immune response for this fatal disease. To characterize the transcriptional signatures of host inflammatory response to SARS-CoV-2 (HCoV-19) infection, we carried out transcriptome sequencing of the RNAs isolated from the bronchoalveolar lavage fluid (BALF) and peripheral blood mononuclear cells (PBMC) specimens of COVID-19 patients. Our results reveal distinct host inflammatory cytokine profiles to SARS-CoV-2 infection in patients, and highlight the association between COVID-19 pathogenesis and excessive cytokine release such as CCL2/MCP-1, CXCL10/IP-10, CCL3/MIP-1A, and CCL4/MIP1B. Furthermore, SARS-CoV-2 induced activation of apoptosis and P53 signalling pathway in lymphocytes may be the cause of patients' lymphopenia. The transcriptome dataset of COVID-19 patients would be a valuable resource for clinical guidance on anti-inflammatory medication and understanding the molecular mechansims of host response.

Osteopetrosis-Associated Transmembrane Protein 1 Recruits RNA Exosome To Restrict Hepatitis B Virus Replication.

Hepatitis B virus (HBV) chronically infects approximately 350 million people worldwide, and 600,000 deaths are caused by HBV-related hepatic failure, liver cirrhosis, and hepatocellular carcinoma annually. It is important to reveal the mechanism underlying the regulation of HBV replication. This study demonstrated that osteopetrosis-associated transmembrane protein 1 (Ostm1) plays an inhibitory role in HBV replication. Ostm1 represses the levels of HBeAg and HBsAg proteins, HBV 3.5-kb and 2.4/2.1-kb RNAs, and core-associated DNA in HepG2, Huh7, and NTCP-HepG2 cells. Notably, Ostm1 has no direct effect on the activity of HBV promoters or the transcription of HBV RNAs; instead, Ostm1 binds to HBV RNA to facilitate RNA decay. Detailed studies further demonstrated that Ostm1 binds to and recruits the RNA exosome complex to promote the degradation of HBV RNAs, and knockdown of the RNA exosome component exonuclease 3 (Exosc3) leads to the elimination of Ostm1-mediated repression of HBV replication. Mutant analyses revealed that the N-terminal domain, the transmembrane domain, and the C-terminal domain are responsible for the repression of HBV replication, and the C-terminal domain is required for interaction with the RNA exosome complex. Moreover, Ostm1 production is not regulated by interferon-α (IFN-α) or IFN-γ, and the expression of IFN signaling components is not affected by Ostm1, suggesting that Ostm1 anti-HBV activity is independent of the IFN signaling pathway. In conclusion, this study revealed a distinct mechanism underlying the repression of HBV replication, in which Ostm1 binds to HBV RNA and recruits RNA exosomes to degrade viral RNA, thereby restricting HBV replication.IMPORTANCE Hepatitis B virus (HBV) is a human pathogen infecting the liver to cause a variety of diseases ranging from acute hepatitis to advanced liver diseases, fulminate hepatitis, liver cirrhosis, and hepatocellular carcinoma, thereby causing a major health problem worldwide. In this study, we demonstrated that Ostm1 plays an inhibitory role in HBV protein production, RNA expression, and DNA replication. However, Ostm1 has no effect on the activities of the four HBV promoters; instead, it binds to HBV RNA and recruits RNA exosomes to promote HBV RNA degradation. We further demonstrated that the anti-HBV activity of Ostm1 is independent of the interferon signaling pathway. In conclusion, this study reveals a distinct mechanism underlying the repression of HBV replication and suggests that Ostm1 is a potential therapeutic agent for HBV infection.

SOX2 Represses Hepatitis B Virus Replication by Binding to the Viral EnhII/Cp and Inhibiting the Promoter Activation.

Hepatitis B virus (HBV) replication is controlled by four promoters (preS1, preS2, Cp, and Xp) and two enhancers (EnhI and EnhII). EnhII stimulates Cp activity to regulate the transcriptions of precore, core, polymerase, and pregenomic RNAs, and therefore, EnhII/Cp is essential for the regulation of HBV replication. This study revealed a distinct mechanism underlying the suppression of EnhII/Cp activation and HBV replication. On the one hand, the sex determining region Y box2 (SOX2), a transcription factor, is induced by HBV. On the other hand, SOX2, in turn, represses the expression levels of HBV RNAs, HBV core-associated DNA, hepatitis B surface antigen (HBsAg), and hepatitis B e antigen (HBeAg), thereby playing an inhibitory role during HBV replication. Further studies indicated that SOX2 bound to the EnhII/Cp DNA and repressed the promoter activation. With the deletion of the high mobility group (HMG) domain, SOX2 loses the ability to repress EnhII/Cp activation, viral RNA transcription, HBV core-associated DNA replication, HBsAg and HBeAg production, as well as fails to enter the nucleus, demonstrating that the HMG domain is required for the SOX2-mediated repression of HBV replication. Moreover, SOX2 represses HBsAg and HBeAg secretion in BALB/c mice sera, and attenuates HBV 3.5kb RNA transcription and hepatitis B virus core protein (HBc) production in the liver tissues, demonstrating that SOX2 suppresses HBV replication in mice. Furthermore, the results revealed that the HMG domain was required for SOX2-mediated repression of HBV replication in the mice. Taken together, the above facts indicate that SOX2 acts as a new host restriction factor to repress HBV replication by binding to the viral EnhII/Cp and inhibiting the promoter activation through the HMG domain.

NS5 Conservative Site Is Required for Zika Virus to Restrict the RIG-I Signaling.

During host-virus co-evolution, cells develop innate immune systems to inhibit virus invasion, while viruses employ strategies to suppress immune responses and maintain infection. Here, we reveal that Zika virus (ZIKV), a re-emerging arbovirus causing public concerns and devastating complications, restricts host immune responses through a distinct mechanism. ZIKV nonstructural protein 5 (NS5) interacts with the host retinoic acid-inducible gene I (RIG-I), an essential signaling molecule for defending pathogen infections. NS5 subsequently represses K63-linked polyubiquitination of RIG-I, attenuates the phosphorylation and nuclear translocation of interferon regulatory factor 3 (IRF3), and inhibits the expression and production of interferon-ß (IFN-ß), thereby restricting the RIG-I signaling pathway. Interestingly, we demonstrate that the methyltransferase (MTase) domain of NS5 is required for the repression of RIG-I ubiquitination, IRF3 activation, and IFN-ß production. Detailed studies further reveal that the conservative active site D146 of NS5 is critical for the suppression of the RIG-I signaling. Therefore, we uncover an essential role of NS5 conservative site D146 in ZIKV-mediated repression of innate immune system, illustrate a distinct mechanism by which ZIKV evades host immune responses, and discover a potential target for anti-viral infection.

SOX9 represses hepatitis B virus replication through binding to HBV EnhII/Cp and inhibiting the promoter activity.

Hepatitis B virus (HBV) infection affects 364 million people worldwide and causes a serious global public health problem. The SRY-related high mobility group-box 9 (SOX9) is a risk of developing cirrhosis in patients with chronic hepatitis B and a cancer stem cell marker. However, the role of SOX9 in HBV replication has not been reported. This study revealed a distinct mechanism underling the regulation of HBV replication mediated by SOX9. HBV induces SOX9 mRNA and protein expression in human hepatoma cells, including HepG2.2.15, HepG2, Huh7, and HepG2-NTCP cells. Further study demonstrated that HBV activates SOX9 expression at the transcriptional level through inducing SOX9 promoter activity and HBc could induce the activity of SOX9 promoter. Interestingly, SOX9 in turn represses HBV replication in human hepatoma cells. More importantly, SOX9 inhibits HBV infection in HepG2-NTCP cells and C57/BL6 mice. Detailed study revealed that SOX9 suppresses HBV replication through directly binding to HBV EnhII/Cp (HBV 1667-1672 nt) to inhibit EnhII/Cp activation. Results from deletion mutant analysis, ChIP assay, nuclear and cytoplasmic extraction analysis, and immunofluorescence demonstrated that SOX9 high mobility group (HMG) domain is required for SOX9 anti-HBV activity. Moreover, we demonstrated that SOX9 and hepatocyte nuclear factor 4 alpha (HNF4α) can bind to HBV EnhII/Cp (HBV 1667-1672 nt) individually and simultaneously to regulate the promoter activity. Collectively, the results revealed a distinct negative feedback mechanism underlying HBV replication and SOX9 expression, and identified SOX9 as a new host restriction factor in HBV replication and infection. IMPORTANCE: HBV infection is a global public health problem by causing serious liver diseases, but the mechanisms underlying HBV pathogenesis remain largely unknown. SOX9 is a risk of developing cirrhosis and a cancer stem cell marker, however, the role of SOX9 in HBV infection has not been reported. The authors revealed a distinct mechanism underling the regulation of HBV replication and SOX9 expression. On the one hand, HBV induces SOX9 expression in human hepatoma cells through activating SOX9 promoter. On the other hand, SOX9 in turn represses HBV replication in human hepatoma cells by binding to and inhibiting HBV EnhII/Cp through its HMG domain. More importantly, SOX9 inhibits HBV infection in HepG2-NTCP cells and C57/BL6 mice. Therefore, this study identifies SOX9 as a novel and potential therapeutic reagent for the prevention and treatment of HBV-associated diseases.

SUMO1 SUMOylates and SENP3 deSUMOylates NLRP3 to orchestrate the inflammasome activation.

The NLRP3 inflammasome regulates innate immune and inflammatory responses by promoting caspase1-dependent induction of pro-inflammatory cytokines. However, aberrant inflammasome activation causes diverse diseases, and thus inflammasome activity must be tightly controlled. Here, we reveal a molecular mechanism underlying the regulation of NLRP3 inflammasome. NLRP3 interacts with SUMO-conjugating enzyme (UBC9), which subsequently promotes small ubiquitin-like modifier 1 (SUMO1) to catalyze NLRP3 SUMOylation at residue Lys204. SUMO1-catalyzed SUMOylation of NLRP3 facilitates ASC oligomerization, inflammasome activation, and interleukin-1ß secretion. Moreover, this study also reveals that SUMO-specific protease 3 (SENP3) is required for the deSUMOylation of NLRP3. Interestingly, SENP3 deSUMOylates NLRP3 to attenuate ASC recruitment and speck formation, the NLRP3 inflammasome activation, as well as IL-1ß cleavage and secretion. In conclusion, we reveal that SUMO1-catalyzed SUMOylation and SENP3-mediated deSUMOylation of NLRP3 orchestrate the inflammasome activation.

Nickel-Catalyzed Reductive Coupling for Transforming Unactivated Aryl Electrophiles into ß-Fluoroethylarenes.

We report herein a facile synthetic method for converting unactivated (hetero)aryl electrophiles into ß-fluoroethylated (hetero)arenes via nickel-catalyzed reductive cross-couplings. This coupling reaction features the involvement of FCH2 CH2 radical intermediate rather than ß-fluoroethyl manganese species which provides effective solutions to the problematic ß-fluoride side eliminations. The practical value of this protocol is further demonstrated by the late-stage modification of several complex ArCl or ArOH-derived bioactive molecules.

Enterovirus 71 Represses Interleukin Enhancer-Binding Factor 2 Production and Nucleus Translocation to Antagonize ILF2 Antiviral Effects.

Enterovirus 71 (EV71) infection causes hand-foot-mouth disease (HFMD), meningoencephalitis, neonatal sepsis, and even fatal encephalitis in children, thereby presenting a serious risk to public health. It is important to determine the mechanisms underlying the regulation of EV71 infection. In this study, we initially show that the interleukin enhancer-binding factor 2 (ILF2) reduces EV71 50% tissue culture infective dose (TCID50) and attenuates EV71 plaque-formation unit (PFU), thereby repressing EV71 infection. Microarray data analyses show that ILF2 mRNA is reduced upon EV71 infection. Cellular studies indicate that EV71 infection represses ILF2 mRNA expression and protein production in human leukemic monocytes (THP-1) -differentiated macrophages and human rhabdomyosarcoma (RD) cells. In addition, EV71 nonstructural protein 2B interacts with ILF2 in human embryonic kidney (HEK293T) cells. Interestingly, in the presence of EV71 2B, ILF2 is translocated from the nucleus to the cytoplasm, and it colocalizes with 2B in the cytoplasm. Therefore, we present a distinct mechanism by which EV71 antagonizes ILF2-mediated antiviral effects by inhibiting ILF2 expression and promoting ILF2 translocation from the nucleus to the cytoplasm through its 2B protein.

EV71 infection induces neurodegeneration via activating TLR7 signaling and IL-6 production.

As a neurotropic virus, human Enterovirus 71 (EV71) infection causes hand-foot-and-mouth disease (HFMD) and may develop severe neurological disorders in infants. Toll-like receptor 7 (TLR7) acts as an innate immune receptor and is also a death receptor in the central nervous system (CNS). However, the mechanisms underlying the regulation of TLR7-mediated brain pathogenesis upon EV71 infection remain largely elusive. Here we reveal a novel mechanism by which EV71 infects astrocytes in the brain and induces neural pathogenesis via TLR7 and interleukin-6 (IL-6) in C57BL/6 mice and in human astroglioma U251 cells. Upon EV71 infection, wild-type (WT) mice displayed more significant body weight loss, higher clinical scores, and lower survival rates as compared with TLR7-/- mice. In the cerebral cortex of EV71-infected mice, neurofilament integrity was disrupted, and inflammatory cell infiltration and neurodegeneration were induced in WT mice, whereas these were largely absent in TLR7-/- mice. Similarly, IL-6 production, Caspase-3 cleavage, and cell apoptosis were significantly higher in EV71-infected WT mice as compared with TLR7-/- mice. Moreover, EV71 preferentially infected and induced IL-6 in astrocytes of mice brain. In U251 cells, EV71-induced IL-6 production and cell apoptosis were suppressed by shRNA-mediated knockdown of TLR7 (shTLR7). Moreover, in the cerebral cortex of EV71-infected mice, the blockade of IL-6 with anti-IL-6 antibody (IL-6-Ab) restored the body weight loss, attenuated clinical scores, improved survival rates, reduced the disruption of neurofilament integrity, decreased cell apoptotic induction, and lowered levels of Caspase-3 cleavage. Similarly, in EV71-infected U251 cells, IL-6-Ab blocked EV71-induced IL-6 production and cell apoptosis in response to viral infection. Collectively, it's exhibited TLR7 upregulation, IL-6 induction and astrocytic cell apoptosis in EV71-infected human brain. Taken together, we propose that EV71 infects astrocytes of the cerebral cortex in mice and human and triggers TLR7 signaling and IL-6 release, subsequently inducing neural pathogenesis in the brain.

Dengue Virus M Protein Promotes NLRP3 Inflammasome Activation To Induce Vascular Leakage in Mice.

Dengue virus (DENV) infection causes serious clinical symptoms, including dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Vascular permeability change is the main feature of the diseases, and the abnormal expression of proinflammatory cytokines is the important cause of vascular permeability change. However, the mechanism underlying vascular permeability induced by DENV has not been fully elucidated. Here, we reveal a distinct mechanism by which DENV infection promotes NLRP3 inflammasome activation and interleukin-1 beta (IL-1ß) release to induce endothelial permeability and vascular leakage in mice. DENV M protein interacts with NLRP3 to facilitate NLRP3 inflammasome assembly and activation, which induce proinflammatory cytokine IL-1ß activation and release. Notably, M can induce vascular leakage in mouse tissues by activating the NLRP3 inflammasome and IL-1ß. More importantly, inflammatory cell infiltration and tissue injuries are induced by M in wild-type (WT) mouse tissues, but they are not affected by M in NLRP3 knockout (NLRP3-/-) mouse tissues. Evans blue intensities in WT mouse tissues are significantly higher than in NLRP3-/- mouse tissues, demonstrating an essential role of NLRP3 in M-induced vascular leakages in mice. Therefore, we propose that upon DENV infection, M interacts with NLRP3 to facilitate inflammasome activation and IL-1ß secretion, which lead to the induction of endothelial permeability and vascular leakage in mouse tissues. The important role of the DENV-M-NLRP3-IL-1ß axis in the induction of vascular leakage provides new insights into the mechanisms underlying DENV pathogenesis and DENV-associated DHF and DSS development.IMPORTANCE Dengue virus (DENV) is a mosquito-borne pathogen, and infections by this virus are prevalent in over 100 tropical and subtropical countries or regions, with approximately 2.5 billion people at risk. DENV infection induces a spectrum of clinical symptoms, ranging from classical dengue fever (DF) to severe dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). Therefore, it is important to understand the mechanisms underlying DENV pathogenesis. In this study, we reveal that the DENV membrane protein (M) interacts with the host NLRP3 protein to promote NLRP3 inflammasome activation, which leads to the activation and release of a proinflammatory cytokine, interleukin-1 beta (IL-1ß). More importantly, we demonstrate that M protein can induce vascular permeability and vascular leakage and that NLRP3 is required for M-induced vascular leakage in mouse tissues. Collectively, this study reveals a distinct mechanism underlying DENV pathogeneses and provides new insights into the development of therapeutic agents for DENV-associated diseases.