Filamin B restricts vaccinia virus spread and is targeted by vaccinia virus protein C4.

Vaccinia virus (VACV) is a large DNA virus that encodes scores of proteins that modulate the host immune response. VACV protein C4 is one such immunomodulator known to inhibit the activation of both the NF-κB signaling cascade and the DNA-PK-mediated DNA sensing pathway. Here, we show that the N-terminal region of C4, which neither inhibits NF-κB nor mediates interaction with DNA-PK, still contributes to virus virulence. Furthermore, this domain interacts directly and with high affinity to the C-terminal domain of filamin B (FLNB). FLNB is a large actin-binding protein that stabilizes the F-actin network and is implicated in other cellular processes. Deletion of FLNB from cells results in larger VACV plaques and increased infectious viral yield, indicating that FLNB restricts VACV spread. These data demonstrate that C4 has a new function that contributes to virulence and engages the cytoskeleton. Furthermore, we show that the cytoskeleton performs further previously uncharacterized functions during VACV infection. IMPORTANCE: Vaccinia virus (VACV), the vaccine against smallpox and monkeypox, encodes many proteins to counteract the host immune response. Investigating these proteins provides insights into viral immune evasion mechanisms and thereby indicates how to engineer safer and more immunogenic VACV-based vaccines. Here, we report that the N-terminal domain of VACV protein C4 interacts directly with the cytoskeletal protein filamin B (FLNB), and this domain of C4 contributes to virus virulence. Furthermore, VACV replicates and spreads better in cells lacking FLNB, thus demonstrating that FLNB has antiviral activity. VACV utilizes the cytoskeleton for movement within and between cells; however, previous studies show no involvement of C4 in VACV replication or spread. Thus, C4 associates with FLNB for a different reason, suggesting that the cytoskeleton has further uncharacterized roles during virus infection.

Distributed Dynamic Event-Triggered Control to Leader-Following Consensus of Nonlinear Multi-Agent Systems with Directed Graphs.

This paper investigates achieving leader-following consensus in a class of multi-agent systems with nonlinear dynamics. Initially, it introduces a dynamic event-triggered strategy designed to effectively alleviate the strain on the system's communication resources. Subsequently, a distributed control strategy is proposed and implemented in the nonlinear leader-follower system using the dynamic event-triggered mechanism, aiming to ensure synchronization across all nodes at an exponential convergence speed. Thirdly, the research shows that under the dynamic event-triggered strategy the minimum event interval of any two consecutive triggers guarantees the elimination of Zeno behavior. Lastly, the validity of the calculation results is verified by a simulation example.

Preparation of carbon quantum dots from ionic liquid modified biomass for the detection of Fe3+ and Pd2+ in environmental water.

A new type of green carbon quantum dots (ILB-CQDs) was prepared by hydrothermal method using ionic liquid as a modifier and grape skin as carbon source, and was obtained from hydrogen-bonded lattice structure ionic liquid preparation, which makes the CQDs in a ring-like stable structure with a stability period of more than 90 day. There is also the catalytic effect of the ionic liquid on cellulose, which makes the prepared CQDs show good advantages, such as uniform particle size, high quantum yield (26.7%), and very good fluorescence performance. This is a smart material for the selective detection of Fe3+ and Pd2+. It has a detection limit of 0.001 nM for Fe3+ and 0.23 µM for Pd2+ in pure water. It has a detection limit of 3.2 nmol/L for Fe3+ and 0.36 µmol/L for Pd2+ in actual water, both of which meet the requirements of WHO drinking water standards. And there is to achieve more than 90% of water restoration effect.

Vaccinia virus BTB-Kelch proteins C2 and F3 inhibit NF-κB activation.

Vaccinia virus (VACV) encodes scores of proteins that suppress host innate immunity and many of these target intracellular signalling pathways leading to activation of inflammation. The transcription factor NF-κB plays a critical role in the host response to infection and is targeted by many viruses, including VACV that encodes 12 NF-κB inhibitors that interfere at different stages in this signalling pathway. Here we report that VACV proteins C2 and F3 are additional inhibitors of this pathway. C2 and F3 are BTB-Kelch proteins that are expressed early during infection, are non-essential for virus replication, but affect the outcome of infection in vivo. Using reporter gene assays, RT-qPCR analyses of endogenous gene expression, and ELISA, these BTB-Kelch proteins are shown here to diminish NF-κB activation by reducing translocation of p65 into the nucleus. C2 and F3 are the 13th and 14th NF-κB inhibitors encoded by VACV. Remarkably, in every case tested, these individual proteins affect virulence in vivo and therefore have non-redundant functions. Lastly, immunisation with a VACV strain lacking C2 induced a stronger CD8+ T cell response and better protection against virus challenge.

Development and validation of a clinical prediction model to estimate the risk of critical patients with COVID-19.

The outbreak of coronavirus disease 2019 (COVID-19) has globally strained medical resources and caused significant mortality. This study was aimed to develop and validate a prediction model based on clinical features to estimate the risk of patients with COVID-19 at admission progressing to critical patients. Patients admitted to the hospital between January 16, 2020, and March 10, 2020, were retrospectively enrolled, and they were observed for at least 14 days after admission to determine whether they developed into severe pneumonia. According to the clinical symptoms, all patients were divided into four groups: mild, normal, severe, and critical. A total of 390 patients with COVID-19 pneumonia were identified, including 212 severe patients and 178 nonsevere patients. The least absolute shrinkage and selection operator (LASSO) regression reduced the variables in the model to 6, which are age, number of comorbidities, computed tomography severity score, lymphocyte count, aspartate aminotransferase, and albumin. The area under curve of the model in the training set is 0.898, and the specificity and sensitivity were 89.7% and 75.5%. The prediction model, nomogram might be useful to access the onset of severe and critical illness among COVID-19 patients at admission, which is instructive for clinical diagnosis.

NF-κB activation is a turn on for vaccinia virus phosphoprotein A49 to turn off NF-κB activation.

Vaccinia virus protein A49 inhibits NF-κB activation by molecular mimicry and has a motif near the N terminus that is conserved in IκBα, ß-catenin, HIV Vpu, and some other proteins. This motif contains two serines, and for IκBα and ß-catenin, phosphorylation of these serines enables recognition by the E3 ubiquitin ligase ß-TrCP. Binding of IκBα and ß-catenin by ß-TrCP causes their ubiquitylation and thereafter proteasome-mediated degradation. In contrast, HIV Vpu and VACV A49 are not degraded. This paper shows that A49 is phosphorylated at serine 7 but not serine 12 and that this is necessary and sufficient for binding ß-TrCP and antagonism of NF-κB. Phosphorylation of A49 S7 occurs when NF-κB signaling is activated by addition of IL-1ß or overexpression of TRAF6 or IKKß, the kinase needed for IκBα phosphorylation. Thus, A49 shows beautiful biological regulation, for it becomes an NF-κB antagonist upon activation of NF-κB signaling. The virulence of viruses expressing mutant A49 proteins or lacking A49 (vΔA49) was tested. vΔA49 was attenuated compared with WT, but viruses expressing A49 that cannot bind ß-TrCP or bind ß-TrCP constitutively had intermediate virulence. So A49 promotes virulence by inhibiting NF-κB activation and by another mechanism independent of S7 phosphorylation and NF-κB antagonism. Last, a virus lacking A49 was more immunogenic than the WT virus.

Study on the correlations of different clinical types with imaging findings at initial diagnosis and clinical laboratory indexes in COVID-19 patients.

Objectives: To investigate the correlations of initial lab and imaging findings in COVID-19 patients of different clinical types. Methods: We retrospective analyzed patients confirmed with COVID-19 in the Fifth Medical Center of the People's Liberation Army (PLA) General Hospital between February to April 2020, selected a total of 58 (N) patients with lab and imaging examinations that met the study criteria, using Artificial intelligence (AI) software to calculate the percentage of COVID-19 lesions in the volume of the whole lung, then the correlations of general information, initial chest CT examination after admission and laboratory examinations were analyzed. Results: The 58 (N) COVID-19 patients were divided into mild group [41(n) cases]: and severe group [17(n) cases]: according to patient's condition. CT findings of the severe group and mild group mainly included single or multiple ground glass opacity (GGO), with lesions mainly distributed in the periphery of lungs or GGO mixed with consolidation, with lesions involved in peripheral and central areas of both lungs, accompanied other signs. A significant difference in CRP, IL-6, D-D, GGT was observed between the two groups (p < 0.05). The ratios regarding lymphocyte abnormality and neutrophil abnormality in the severe group were higher than those in the mild group (p < 0.05). Conclusion: The CT features at initial diagnosis of COVID-19 were mainly characterized by multiple GGO with or without partial consolidation in both lungs, with the lesions mainly distributed at the subpleural regions. Some lab test indexes were correlated with the clinical types of COVID-19.

Recent advances in ionic liquids-based hybrid processes for CO2 capture and utilization.

CO2 capture and utilization (CCU) is an effective strategy to mitigate global warming. Absorption, adsorption and membranes are methods used for CO2 separation and capture, and various catalytic pathways have also been developed for CO2 utilization. Although widely researched and used in industry, these processes are energy-intensive and this challenge needs to be overcome. To realize further optimization, novel materials and processes are continuously being developed. New generation materials such as ionic liquids (ILs) have shown promising potential for cost-effective CO2 capture and utilization. This study reviews the current status of ILs-based solvents, adsorbents, membranes, catalysts and their hybrid processes for CO2 capture and utilization. The special properties of ILs are integrated into new materials through hybridization, which significantly improves the performance in the process of CCU.

Risk Factors Associated With Clinical Outcomes in 323 Coronavirus Disease 2019 (COVID-19) Hospitalized Patients in Wuhan, China.

BACKGROUND: With evidence of sustained transmission in more than 190 countries, coronavirus disease 2019 (COVID-19) has been declared a global pandemic. Data are urgently needed about risk factors associated with clinical outcomes. METHODS: A retrospective review of 323 hospitalized patients with COVID-19 in Wuhan was conducted. Patients were classified into 3 disease severity groups (nonsevere, severe, and critical), based on initial clinical presentation. Clinical outcomes were designated as favorable and unfavorable, based on disease progression and response to treatments. Logistic regression models were performed to identify risk factors associated with clinical outcomes, and log-rank test was conducted for the association with clinical progression. RESULTS: Current standard treatments did not show significant improvement in patient outcomes. By univariate logistic regression analysis, 27 risk factors were significantly associated with clinical outcomes. Multivariate regression indicated age >65 years (P < .001), smoking (P = .001), critical disease status (P = .002), diabetes (P = .025), high hypersensitive troponin I (>0.04 pg/mL, P = .02), leukocytosis (>10 × 109/L, P < .001), and neutrophilia (>75 × 109/L, P < .001) predicted unfavorable clinical outcomes. In contrast, the administration of hypnotics was significantly associated with favorable outcomes (P < .001), which was confirmed by survival analysis. CONCLUSIONS: Hypnotics may be an effective ancillary treatment for COVID-19. We also found novel risk factors, such as higher hypersensitive troponin I, predicted poor clinical outcomes. Overall, our study provides useful data to guide early clinical decision making to reduce mortality and improve clinical outcomes of COVID-19.

Leaky scanning translation generates a second A49 protein that contributes to vaccinia virus virulence.

Vaccinia virus (VACV) strain Western Reserve gene A49L encodes a small intracellular protein with a Bcl-2 fold that is expressed early during infection and has multiple functions. A49 co-precipitates with the E3 ubiquitin ligase ß-TrCP and thereby prevents ubiquitylation and proteasomal degradation of IκBα, and consequently blocks activation of NF-κB. In a similar way, A49 stabilizes ß-catenin, leading to activation of the wnt signalling pathway. However, a VACV strain expressing a mutant A49 that neither co-precipitates with ß-TrCP nor inhibits NF-κB activation, is more virulent than a virus lacking A49, indicating that A49 has another function that also contributes to virulence. Here we demonstrate that gene A49L encodes a second, smaller polypeptide that is expressed via leaky scanning translation from methionine 20 and is unable to block NF-κB activation. Viruses engineered to express either only the large protein or only the small A49 protein both have lower virulence than wild-type virus and greater virulence than an A49L deletion mutant. This demonstrates that the small protein contributes to virulence by an unknown mechanism that is independent of NF-κB inhibition. Despite having a large genome with about 200 genes, this study illustrates how VACV makes efficient use of its coding potential and from gene A49L expresses a protein with multiple functions and multiple proteins with different functions.

Vaccinia Virus BBK E3 Ligase Adaptor A55 Targets Importin-Dependent NF-κB Activation and Inhibits CD8+ T-Cell Memory.

Viral infection of cells is sensed by pathogen recognition receptors that trigger an antiviral innate immune response, and consequently viruses have evolved countermeasures. Vaccinia virus (VACV) evades the host immune response by expressing scores of immunomodulatory proteins. One family of VACV proteins are the BTB-BACK (broad-complex, tram-trac, and bric-a-brac [BTB] and C-terminal Kelch [BACK]) domain-containing, Kelch-like (BBK) family of predicted cullin-3 E3 ligase adaptors: A55, C2, and F3. Previous studies demonstrated that gene A55R encodes a protein that is nonessential for VACV replication yet affects viral virulence in vivo Here, we report that A55 is an NF-κB inhibitor acting downstream of IκBα degradation, preventing gene transcription and cytokine secretion in response to cytokine stimulation. A55 targets the host importin α1 (KPNA2), acting to reduce p65 binding and its nuclear translocation. Interestingly, while A55 was confirmed to coprecipitate with cullin-3 in a BTB-dependent manner, its NF-κB inhibitory activity mapped to the Kelch domain, which alone is sufficient to coprecipitate with KPNA2 and inhibit NF-κB signaling. Intradermal infection of mice with a virus lacking A55R (vΔA55) increased VACV-specific CD8+ T-cell proliferation, activation, and cytotoxicity in comparison to levels of the wild-type (WT) virus. Furthermore, immunization with vΔA55 induced increased protection to intranasal VACV challenge compared to the level with control viruses. In summary, this report describes the first target of a poxvirus-encoded BBK protein and a novel mechanism for DNA virus immune evasion, resulting in increased CD8+ T-cell memory and a more immunogenic vaccine.IMPORTANCE NF-κB is a critical transcription factor in the innate immune response to infection and in shaping adaptive immunity. The identification of host and virus proteins that modulate the induction of immunological memory is important for improving virus-based vaccine design and efficacy. In viruses, the expression of BTB-BACK Kelch-like (BBK) proteins is restricted to poxviruses and conserved within them, indicating the importance of these proteins for these medically important viruses. Using vaccinia virus (VACV), the smallpox vaccine, we report that the VACV BBK protein A55 dysregulates NF-κB signaling by disrupting the p65-importin interaction, thus preventing NF-κB translocation and blocking NF-κB-dependent gene transcription. Infection with VACV lacking A55 induces increased VACV-specific CD8+ T-cell memory and better protection against VACV challenge. Studying viral immunomodulators therefore expands not only our understanding of viral pathogenesis and immune evasion strategies but also of the immune signaling cascades controlling antiviral immunity and the development of immune memory.

Astrocytic NDRG2 is critical in the maintenance of neuropathic pain.

Activation of astrocytes and abnormal synaptic glutamate metabolism are closely associated with the induction and maintenance of neuropathic pain (NP), but the exact mechanism underlying this association remains unclear. N-myc downstream-regulated gene 2 (NDRG2), a novel tumor-suppressor protein and stress-response gene, is involved in the pathogenesis of several neurodegenerative diseases. However, its role in nociceptive transduction has rarely been investigated. Here, we found that NDRG2, which was mainly expressed in the astrocytes in the central nervous system (CNS), was increased in the spinal cord of a spinal nerve ligation (SNL) rat model for NP. Suppression of NDRG2 by intrathecal injection of an NDRG2-RNAi-adenovirus significantly alleviated SNL-induced mechanical and thermal hypersensitivity, as well as elevated astrocytic glutamate transporter 1 (GLT-1) expression and downregulated pro-inflammatory cytokine levels, in the spinal dorsal horn of rats on Day 10 after SNL. Furthermore, in lipopolysaccharide (LPS)-stimulated primary astrocytic cultures derived from neonatal rats, inhibition of NDRG2 significantly reversed both the LPS-induced activation of astrocytes and decreased expression of GLT-1. By contrast, overexpression of NDRG2 by an adenoviral vector carrying NDRG2 resulted in astrocytic activation, aberrant glutamatergic neurotransmission, and spontaneous nociceptive responses in rats. Intrathecal injection of AG490, which is an inhibitor of the Janus tyrosine kinase and signal transducer and activator of the transcription 3 (JAK/STAT3) signaling pathway, significantly attenuated both mechanical and thermal hyperalgesia, as well as inhibited reactive astrocytes and restored normal expression levels of astrocytic GLT-1, in the spinal dorsal horn of NDRG2-overexpression rats. In conclusion, spinal astrocytic NDRG2 is critical in the maintenance of NP. Moreover, NDRG2 modulates astrocytic activation and inflammatory responses via regulating GLT-1 expression through the JAK/STAT3 signaling pathway. Our findings suggested that NDRG2 could be a novel therapeutic target for the treatment of NP.

Aberrant dorsal attention network homogeneity in patients with right temporal lobe epilepsy.

The dorsal attention network (DAN) is involved in the process that causes wide-ranging cognitive damage resulted in right temporal lobe epilepsy (rTLE). Nevertheless, few studies have evaluated the relationship between DAN and rTLE. There has been little research on alterations in the network homogeneity (NH) of the DAN in rTLE. The aim of the present study was to investigate NH changes in DAN in patients with rTLE. We included 85 patients with rTLE and 69 healthy controls in this study, and resting-state functional magnetic resonance imaging (rs-fMRI) data were acquired. The NH method was used for data analysis. All subjects took the attention network test (ANT). Network homogeneity in the right superior parietal lobule (SPL) and right precuneus (PCU) was significantly higher in patients with rTLE than in healthy controls. The reaction time (RT) was significantly longer in patients with rTLE than in controls. Notably, we observed no significant relationship between the clinical variables and the abnormal NH. These results indicated that abnormal alterations in DAN existed in patients with rTLE and highlighted the crucial role of DAN in the pathophysiology of cognitive damage in rTLE. Our findings suggested that the executive function (EF) significantly weakened in patients with rTLE.

Hybrid Sequencing of Full-Length cDNA Transcripts of the Medicinal Plant Scutellaria baicalensis.

Scutellaria baicalensis is a well-known medicinal plant that produces biologically active flavonoids, such as baicalin, baicalein, and wogonin. Pharmacological studies have shown that these compounds have anti-inflammatory, anti-bacterial, and anti-cancer activities. Therefore, it is of great significance to investigate the genetic information of S. baicalensis, particularly the genes related to the biosynthetic pathways of these compounds. Here, we constructed the full-length transcriptome of S. baicalensis using a hybrid sequencing strategy and acquired 338,136 full-length sequences, accounting for 93.3% of the total reads. After the removal of redundancy and correction with Illumina short reads, 75,785 nonredundant transcripts were generated, among which approximately 98% were annotated with significant hits in the protein databases, and 11,135 sequences were classified as lncRNAs. Differentially expressed gene (DEG) analysis showed that most of the genes related to flavonoid biosynthesis were highly expressed in the roots, consistent with previous reports that the flavonoids were mainly synthesized and accumulated in the roots of S. baicalensis. By constructing unique transcription models, a total of 44,071 alternative splicing (AS) events were identified, with intron retention (IR) accounting for the highest proportion (44.5%). A total of 94 AS events were present in five key genes related to flavonoid biosynthesis, suggesting that AS may play important roles in the regulation of flavonoid biosynthesis in S. baicalensis. This study provided a large number of highly accurate full-length transcripts, which represents a valuable genetic resource for further research of the molecular biology of S. baicalensis, such as the development, breeding, and biosynthesis of active ingredients.

Neuroprotective Effects of Ginsenosides against Cerebral Ischemia.

Ginseng has been used worldwide as traditional medicine for thousands of years, and ginsenosides have been proved to be the main active components for their various pharmacological activities. Based on their structures, ginsenosides can be divided into ginseng diol-type A and ginseng triol-type B with different pharmacological effects. In this study, six ginsenosides, namely ginsenoside Rb1, Rh2, Rg3, Rg5 as diol-type ginseng saponins, and Rg1 and Re as triol-type ginseng saponins, which were reported to be effective for ischemia-reperfusion (I/R) treatment, were chosen to compare their protective effects on cerebral I/R injury, and their mechanisms were studied by in vitro and in vivo experiments. It was found that all ginsenosides could reduce reactive oxygen species (ROS), inhibit apoptosis and increase mitochondrial membrane potential in cobalt chloride-induced (CoCl2-induced) PC12 cells injury model, and they could reduce cerebral infarction volume, brain neurological dysfunction of I/R rats in vivo. The results of immunohistochemistry and western blot showed that the expression of Toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88), silencing information regulator (SIRT1) and nuclear transcription factor P65 (NF-κB) in hippocampal CA1 region of some ginsenoside groups were also reduced. In general, the effect on cerebral ischemia of Rb1 and Rg3 was significantly improved compared with the control group, and was the strongest among all the ginsenosides. The effect on SIRT1 activation of ginsenoside Rb1 and the inhibition effect of TLR4/MyD88 protein expression of ginsenoside Rb1 and Rg3 were significantly stronger than that of other groups. The results indicated that ginsenoside Rg1, Rb1, Rh2, Rg3, Rg5 and Re were effective in protecting the brain against ischemic injury, and ginsenoside Rb1 and Rg3 have the strongest therapeutic activities in all the tested ginsenosides. Their neuroprotective mechanism is associated with TLR4/MyD88 and SIRT1 activation signaling pathways, and they can reduce cerebral ischemic injury by inhibiting NF-κB transcriptional activity and the expression of proinflammatory cytokines, including interleukin-1ß (IL-1ß), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6).

Nuclear magnetic resonance evaluation of inflammatory activity from chronic viral Hepatitis B.

OBJECTIVES: To discuss the value of applying magnetic resonance diffusion-weighted imaging (DWI) to evaluate inflammatory activity from chronic viral hepatitis B. METHODS: One hundred forty-two patients with chronic viral hepatitis B who received treatment at The Fifth Medical Center of Chinese PLA General Hospital from January 2014 to December 2015 and 20 healthy persons in the control group who were scheduled to undergo nuclear magnetic resonance scanning and DWI examinations (b value = 0, 800 s/mm2), and the apparent diffusion coefficients (ADCs) were measured and compared with the biopsy results of hepatic tissue. RESULTS: The ADC value of the group with hepatitis B was lower than that of the healthy group (P<0.05), and the ADC value of the group with mild inflammation (G1) significantly differed from that of the group with moderate inflammation (G2) and that of the group with severe inflammation (G3-G4) (P<0.05). CONCLUSIONS: Magnetic resonance diffusion-weighted imaging technology has high clinical value for evaluating the inflammatory activity from chronic hepatitis B, and the measured ADC value corresponds to the pathological grade well, so this method is worth clinical promotion and application.

Inhibition of Translation Initiation by Protein 169: A Vaccinia Virus Strategy to Suppress Innate and Adaptive Immunity and Alter Virus Virulence.

Vaccinia virus (VACV) is the prototypic orthopoxvirus and the vaccine used to eradicate smallpox. Here we show that VACV strain Western Reserve protein 169 is a cytoplasmic polypeptide expressed early during infection that is excluded from virus factories and inhibits the initiation of cap-dependent and cap-independent translation. Ectopic expression of protein 169 causes the accumulation of 80S ribosomes, a reduction of polysomes, and inhibition of protein expression deriving from activation of multiple innate immune signaling pathways. A virus lacking 169 (vΔ169) replicates and spreads normally in cell culture but is more virulent than parental and revertant control viruses in intranasal and intradermal murine models of infection. Intranasal infection by vΔ169 caused increased pro-inflammatory cytokines and chemokines, infiltration of pulmonary leukocytes, and lung weight. These alterations in innate immunity resulted in a stronger CD8+ T-cell memory response and better protection against virus challenge. This work illustrates how inhibition of host protein synthesis can be a strategy for virus suppression of innate and adaptive immunity.

Enhancement of CD8(+) T-cell memory by removal of a vaccinia virus nuclear factor-κB inhibitor.

Factors influencing T-cell responses are important for vaccine development but are incompletely understood. Here, vaccinia virus (VACV) protein N1 is shown to impair the development of both effector and memory CD8(+) T cells and this correlates with its inhibition of nuclear factor-κB (NF-κB) activation. Infection with VACVs that either have the N1L gene deleted (vΔN1) or contain a I6E mutation (vN1.I6E) that abrogates its inhibition of NF-κB resulted in increased central and memory CD8(+) T-cell populations, increased CD8(+) T-cell cytotoxicity and lower virus titres after challenge. Furthermore, CD8(+) memory T-cell function was increased following infection with vN1.I6E, with more interferon-γ production and greater protection against VACV infection following passive transfer to naive mice, compared with CD8(+) T cells from mice infected with wild-type virus (vN1.WT). This demonstrates the importance of NF-κB activation within infected cells for long-term CD8(+) T-cell memory and vaccine efficacy. Further, it provides a rationale for deleting N1 from VACV vectors to enhance CD8(+) T-cell immunogenicity, while simultaneously reducing virulence to improve vaccine safety.

Poxvirus targeting of E3 ligase ß-TrCP by molecular mimicry: a mechanism to inhibit NF-κB activation and promote immune evasion and virulence.

The transcription factor NF-κB is essential for immune responses against pathogens and its activation requires the phosphorylation, ubiquitination and proteasomal degradation of IκBα. Here we describe an inhibitor of NF-κB from vaccinia virus that has a closely related counterpart in variola virus, the cause of smallpox, and mechanistic similarity with the HIV protein Vpu. Protein A49 blocks NF-κB activation by molecular mimicry and contains a motif conserved in IκBα which, in IκBα, is phosphorylated by IKKß causing ubiquitination and degradation. Like IκBα, A49 binds the E3 ligase ß-TrCP, thereby preventing ubiquitination and degradation of IκBα. Consequently, A49 stabilised phosphorylated IκBα (p-IκBα) and its interaction with p65, so preventing p65 nuclear translocation. Serine-to-alanine mutagenesis within the IκBα-like motif of A49 abolished ß-TrCP binding, stabilisation of p-IκBα and inhibition of NF-κB activation. Remarkably, despite encoding nine other inhibitors of NF-κB, a VACV lacking A49 showed reduced virulence in vivo.