Vaccinia Virus Protein C6 Inhibits Type I IFN Signalling in the Nucleus and Binds to the Transactivation Domain of STAT2.

The type I interferon (IFN) response is a crucial innate immune signalling pathway required for defense against viral infection. Accordingly, the great majority of mammalian viruses possess means to inhibit this important host immune response. Here we show that vaccinia virus (VACV) strain Western Reserve protein C6, is a dual function protein that inhibits the cellular response to type I IFNs in addition to its published function as an inhibitor of IRF-3 activation, thereby restricting type I IFN production from infected cells. Ectopic expression of C6 inhibits the induction of interferon stimulated genes (ISGs) in response to IFNα treatment at both the mRNA and protein level. C6 inhibits the IFNα-induced Janus kinase/signal transducer and activator of transcription (JAK/STAT) signalling pathway at a late stage, downstream of STAT1 and STAT2 phosphorylation, nuclear translocation and binding of the interferon stimulated gene factor 3 (ISGF3) complex to the interferon stimulated response element (ISRE). Mechanistically, C6 associates with the transactivation domain of STAT2 and this might explain how C6 inhibits the type I IFN signalling very late in the pathway. During virus infection C6 reduces ISRE-dependent gene expression despite the presence of the viral protein phosphatase VH1 that dephosphorylates STAT1 and STAT2. The ability of a cytoplasmic replicating virus to dampen the immune response within the nucleus, and the ability of viral immunomodulators such as C6 to inhibit multiple stages of the innate immune response by distinct mechanisms, emphasizes the intricacies of host-pathogen interactions and viral immune evasion.Wellcome-Trust, Lister Institute of Preventive Medicine U

HeapCraft Social Tools: Understanding and Improving Player Collaboration in Minecraft

We introduce a framework to infuence and analyze player collaboration in Minecraft. The framework consists of a telemetry system and several tools to influence player behavior and provide value to server administrators to increase adoption. The data collection includes almost every aspect of gameplay and can be used for analysis beyond player collaboration.1 We started collecting data from several Minecraft servers in March 2015. Most data will be made available to researchers upon request.2 We have also demonstrated the use of our framework to statistically analyze player behavior in Minecraft. More details can be found [1]

HEAPCRAFT: Quantifying and Predicting Collaboration in Minecraft

We present HEAPCRAFT: an open-source suite of tools for monitoring and improving collaboration in Minecraft. At the core of our system is a data collection and analysis framework or recording gameplay. We collected over 3451 playerhours of game behavior from 908 different players, and performed a general study of online collaboration. To make our game analytics easily accessible, we developed interactive information visualization tools and an analysis framework for players, administrators, and researchers to explore graphs, maps and timelines of live server activity. As part of our research, we introduce the collaboration index, a metric which allows server administrators and researchers to quantify, predict, and improve collaboration on Minecraft servers. Our analysis reveals several possible predictors of collaboration which can be used to improve collaboration on Minecraft servers. HEAPCRAFT is designed to be general, and has the potential to be used for other shared online virtual worlds

Life history, patchy distribution, and patchy taxonomy in a shallow-water invertebrate (Mollusca Polyplacophora: Lepidopleurida)

Things without names are difficult to rationalise, and so species that go without names are difficult to conserve or protect. This is a case study in resolving conflicts in historical taxonomy and ‘real’ species (identifiable and evolutionarily relevant groupings) using an approach including population genetics, natural history, and pragmatism. We report the observation that populations of a shallow-water chiton species from Washington and British Columbia demonstrate extremely high site fidelity and patchy distribution. Their limited dispersal potential and isolation could be explained by a brooding life history. This stands in direct contrast with the supposedly wide distribution of this “species”, Leptochiton rugatus (Carpenter in Pilsbry, 1892) sensu lato, from the Sea of Japan to Baja California. But this lineage has previously been suggested to comprise several cryptic species. Indeed, a haplotype network analysis using 61 individual sequences of the cytochrome oxidase c subunit I gene for L. rugatus s.l. revealed four discrete clusters which correspond to different parts of the geographic range. We infer these to represent four distinct species, at least two of which are likely novel. Leptochiton rugatus sensu stricto is herein reinterpreted as restricted to California and Baja California, and the new name L. cascadiensis sp. nov. is established for the lineage with a distribution in the Cascadia coastal bioregion from the panhandle of Alaska to Oregon. There are minor morphological differences among these species in the L. rugatus species complex, but genetic data or morphological observations alone would not have been sufficient to definitively recognise these groups as species-level lineages. The observation that different species within the complex may have different life history strategies provides important support for interpreting different populations as genuinely separate species.</p

Guillain-Barré syndrome: a century of progress

In 1916, Guillain, Barré and Strohl reported on two cases of acute flaccid paralysis with high cerebrospinal fluid protein levels and normal cell counts — novel findings that identified the disease we now know as Guillain–Barré syndrome (GBS). 100 years on, we have made great progress with the clinical and pathological characterization of GBS. Early clinicopathological and animal studies indicated that GBS was an immune-mediated demyelinating disorder, and that severe GBS could result in secondary axonal injury; the current treatments of plasma exchange and intravenous immunoglobulin, which were developed in the 1980s, are based on this premise. Subsequent work has, however, shown that primary axonal injury can be the underlying disease. The association of Campylobacter jejuni strains has led to confirmation that anti-ganglioside antibodies are pathogenic and that axonal GBS involves an antibody and complement-mediated disruption of nodes of Ranvier, neuromuscular junctions and other neuronal and glial membranes. Now, ongoing clinical trials of the complement inhibitor eculizumab are the first targeted immunotherapy in GBS

Phenotypic Variation and Bistable Switching in Bacteria

Microbial research generally focuses on clonal populations. However, bacterial cells with identical genotypes frequently display different phenotypes under identical conditions. This microbial cell individuality is receiving increasing attention in the literature because of its impact on cellular differentiation, survival under selective conditions, and the interaction of pathogens with their hosts. It is becoming clear that stochasticity in gene expression in conjunction with the architecture of the gene network that underlies the cellular processes can generate phenotypic variation. An important regulatory mechanism is the so-called positive feedback, in which a system reinforces its own response, for instance by stimulating the production of an activator. Bistability is an interesting and relevant phenomenon, in which two distinct subpopulations of cells showing discrete levels of gene expression coexist in a single culture. In this chapter, we address techniques and approaches used to establish phenotypic variation, and relate three well-characterized examples of bistability to the molecular mechanisms that govern these processes, with a focus on positive feedback.

Disrupting HIV-1 capsid formation causes cGAS sensing of viral DNA

Detection of viral DNA by cyclic GMP-AMP synthase (cGAS) is a first line of defence leading to the production of type I interferon (IFN). As HIV-1 replication is not a strong inducer of IFN, we hypothesised that an intact capsid physically cloaks viral DNA from cGAS. To test this, we generated defective viral particles by treatment with HIV-1 protease inhibitors or by genetic manipulation of gag. These viruses had defective Gag cleavage, reduced infectivity and diminished capacity to saturate TRIM5α. Importantly, unlike wild-type HIV-1, infection with cleavage defective HIV-1 triggered an IFN response in THP-1 cells that was dependent on viral DNA and cGAS. An IFN response was also observed in primary human macrophages infected with cleavage defective viruses. Infection in the presence of the capsid destabilising small molecule PF-74 also induced a cGAS-dependent IFN response. These data demonstrate a protective role for capsid and suggest that antiviral activity of capsid- and protease-targeting antivirals may benefit from enhanced innate and adaptive immunity in vivo

Lentiviral Vector Production Titer Is Not Limited in HEK293T by Induced Intracellular Innate Immunity

Most gene therapy lentiviral vector (LV) production platforms employ HEK293T cells expressing the oncogenic SV40 large T-antigen (TAg) that is thought to promote plasmid-mediated gene expression. Studies on other viral oncogenes suggest that TAg may also inhibit the intracellular autonomous innate immune system that triggers defensive antiviral responses upon detection of viral components by cytosolic sensors. Here we show that an innate response can be generated after HIV-1-derived LV transfection in HEK293T cells, particularly by the transgene, yet, remarkably, this had no effect on LV titer. Further, overexpression of DNA sensing pathway components led to expression of inflammatory cytokine and interferon (IFN) stimulated genes but did not result in detectable IFN or CXCL10 and had no impact on LV titer. Exogenous IFN-β also did not affect LV production or transduction efficiency in primary T cells. Additionally, manipulation of TAg did not affect innate antiviral responses, but stable expression of TAg boosted vector production in HEK293 cells. Our findings demonstrate a measure of innate immune competence in HEK293T cells but, crucially, show that activation of inflammatory signaling is uncoupled from cytokine secretion in these cells. This provides new mechanistic insight into the unique suitability of HEK293T cells for LV manufacture

A randomised, double-blind, placebo-controlled trial of repeated nebulisation of non-viral cystic fibrosis transmembrane conductance regulator (CFTR) gene therapy in patients with cystic fibrosis

BACKGROUND: Cystic fibrosis (CF) is a chronic, life-limiting disease caused by mutations in the CF transmembrane conductance regulator (CFTR) gene leading to abnormal airway surface ion transport, chronic lung infections, inflammation and eventual respiratory failure. With the exception of the small-molecule potentiator, ivacaftor (Kalydeco®, Vertex Pharmaceuticals, Boston, MA, USA), which is suitable for a small proportion of patients, there are no licensed therapies targeting the basic defect. The UK Cystic Fibrosis Gene Therapy Consortium has taken a cationic lipid-mediated CFTR gene therapy formulation through preclinical and clinical development. OBJECTIVE: To determine clinical efficacy of the formulation delivered to the airways over a period of 1 year in patients with CF. DESIGN: This was a randomised, double-blind, placebo-controlled Phase IIb trial of the CFTR gene–liposome complex pGM169/GL67A. Randomisation was performed via InForm™ version 4.6 (Phase Forward Incorporated, Oracle, CA, USA) and was 1 : 1, except for patients in the mechanistic subgroups (2 : 1). Allocation was blinded by masking nebuliser chambers. SETTINGS: Data were collected in the clinical and scientific sites and entered onto a trial-specific InForm, version 4.6 database. PARTICIPANTS: Patients with CF aged ≥ 12 years with forced expiratory volume in the first second (FEV1) between 50% and 90% predicted and any combination of CFTR mutations. The per-protocol group (≥ 9 doses) consisted of 54 patients receiving placebo (62 randomised) and 62 patients receiving gene therapy (78 randomised). INTERVENTIONS: Subjects received 5 ml of nebulised pGM169/G67A (active) or 0.9% saline (placebo) at 28 (±5)-day intervals over 1 year. MAIN OUTCOME MEASURES: The primary end point was the relative change in percentage predicted FEV1 over the 12-month period. A number of secondary clinical outcomes were assessed alongside safety measures: other spirometric values; lung clearance index (LCI) assessed by multibreath washout; structural disease on computed tomography (CT) scan; the Cystic Fibrosis Questionnaire – Revised (CFQ-R), a validated quality-of-life questionnaire; exercise capacity and monitoring; systemic and sputum inflammatory markers; and adverse events (AEs). A mechanistic study was performed in a subgroup in whom transgene deoxyribonucleic acid (DNA) and messenger ribonucleic acid (mRNA) was measured alongside nasal and lower airway potential difference. RESULTS: There was a significant (p = 0.046) treatment effect (TE) of 3.7% [95% confidence interval (CI) 0.1% to 7.3%] in the primary end point at 12 months and in secondary end points, including forced vital capacity (FVC) (p = 0.031) and CT gas trapping (p = 0.048). Other outcomes, although not reaching statistical significance, favoured active treatment. Effects were noted by 1 month and were irrespective of sex, age or CFTR mutation class. Subjects with a more severe baseline FEV1 had a FEV1 TE of 6.4% (95% CI 0.8% to 12.1%) and greater changes in many other secondary outcomes. However, the more mildly affected group also demonstrated benefits, particularly in small airway disease markers such as LCI. The active group showed a significantly (p = 0.032) greater bronchial chloride secretory response. No difference in treatment-attributable AEs was seen between the placebo and active groups. CONCLUSIONS: Monthly application of the pGM169/GL67A gene therapy formulation was associated with an improvement in lung function, other clinically relevant parameters and bronchial CFTR function, compared with placebo. LIMITATIONS: Although encouraging, the improvement in FEV1 was modest and was not accompanied by detectable improvement in patients’ quality of life. FUTURE WORK: Future work will focus on attempts to increase efficacy by increasing dose or frequency, the coadministration of a CFTR potentiator, or the use of modified viral vectors capable of repeated administration. TRIAL REGISTRATION: ClinicalTrials.gov NCT01621867

Development and initial psychometric properties of the Warwick–Edinburgh Mental Wellbeing Scale-Intellectual Disability version

Background: The Warwick–Edinburgh Mental Wellbeing Scale (WEMWBS; Tennant et al., 2007) is yet to be validated in the intellectual disability (ID) population. The aim of this study was to report the development process and assess the psychometric properties of a newly adapted version of the WEMWBS and the Short WEMWBS for individuals with mild to moderate IDs (WEMWBS-ID/SWEMWBS-ID). / Method: The WEMWBS item wordings and response options were revised by clinicians and researchers expert in the field of ID, and a visual aid was added to the scale. The adapted version was reviewed by 10 individuals with IDs. The measure was administered by researchers online using screenshare, to individuals aged 16+ years with mild to moderate IDs. Data from three UK samples were collated to evaluate the WEMWBS-ID (n = 96). A subsample (n = 22) completed the measure again 1 to 2 weeks later to assess test–retest reliability, and 95 participants additionally completed an adapted version of the adapted Rosenberg Self-Esteem Scale to examine convergent validity. Additional data from a Canadian sample (n = 27) were used to evaluate the SWEMWBS-ID (n = 123). / Results: The WEMWBS-ID demonstrated good internal consistency (ω = 0.77–0.87), excellent test–retest reliability [intraclass correlation coefficient (ICC) =.88] and good convergent validity with the self-esteem scale (r =.48–.60) across samples. A confirmatory factor analysis for a single factor model demonstrated an adequate fit. The SWEMWBS-ID showed poor to good internal consistency (ω = 0.36–0.74), moderate test–retest reliability (ICC =.67) and good convergent validity (r =.48–.60) across samples, and a confirmatory factor analysis indicated good model fit for a single factor structure. / Conclusions: The WEMWBS-ID and short version demonstrated promising psychometric properties, when administered virtually by a researcher. Further exploration of the scales with larger, representative samples is warranted