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Recipients:
University of Oxford
Award Year:
2017

Results

Molecular mechanisms of HIV-1 restriction by capsid-sensing host cell proteins 05 Apr 2017

Infections by retroviruses, such as HIV-1, critically depend on the viral capsid. Many host cell defence proteins, including restriction factors Trim5alpha, TrimCyp and MxB, target the viral capsid at the early stages of infection and potently inhibit virus replication. These restriction factors appear to function through a remarkable capsid pattern sensing ability that specifically recognizes the assembled capsid, but not the individual capsid protein. Using an integrative and multidisciplinary approach, I aim to determine the molecular interactions between the viral capsid and host restriction factors, TrimCyp and MxB, that underpin their capsid pattern-sensing capability and ability to inhibit HIV-1 replication. Specifically, I will combine cryoEM and cryoET with all-atom molecular-dynamics simulations to obtain high-resolution structures and atomic models of the capsid and host protein complexes (in vitro), together with mutational and functional analysis as well as correlative light and cryoET imaging of viral infection process (in vivo and in situ), to reveal the essential interfaces in their 3D organization for HIV-1 capsid recognition and inhibition of HIV-1 infection. Information derived from our studies will allow to design more robust therapeutic agents to block HIV-1 replication by strengthening the pattern recognition feature.

Amount: £1,476,229
Funder: The Wellcome Trust
Recipient: University of Oxford

Genomic medicine and statistics 30 Sep 2017

Not available

Amount: £2,580,000
Funder: The Wellcome Trust
Recipient: University of Oxford

Structural studies of bacterial nucleic acid transport proteins 31 Jan 2017

Horizontal gene transfer contributes to genetic plasticity in bacteria and is of great clinical relevance as it contributes to the spread of antibiotic resistance genes. One mechanism of horizontal gene transfer in bacteria is transformation. While the phenomenon of transformation has been known for many decades, little is known about the mechanistic steps of exogenous DNA uptake into bacterial cells. The most obvious problem is how the DNA gets past the cell envelopes. ComEC is believed to be the protein that forms an aqueous pore that allows transport of DNA into the cytoplasm through the bacterial plasma membrane. The protein represents a novel transport protein, and no structural and very little functional information is available. The aim of the project is to structurally and functionally characterize ComEC proteins using modern protein expression and screening techniques, advanced structural approaches (X-ray crystallography, cryo-electron microscopy) and functional studies (fluorescence microscopy, biophysics), in order to build a model for DNA transport across the plasma membrane into the cytoplasm.

Amount: £0
Funder: The Wellcome Trust
Recipient: University of Oxford

KDEL Receptor Stuctural Determination & Biochemical Characterisation 31 Jan 2017

This project aims to characterise the KDEL receptor (KDELR) structurally, biochemically and biophysically. The KDELR is a membrane protein resident in the cis-golgi, where it binds the K-D-E-L amino-acid motif present on resident ER proteins, which have been transported to the Golgi via bulk flow. Once KDELR binds cargo, it initiates transport back to the ER via COPI mediated vesicles, where it releases its cargo, ostensibly because of the differing pH. The molecular mechanisms concerning the actions of the KDELR are largely elusive and would be greatly aided by the structural determination of the KDELR, as well as the structures and characterisations of its interactions with native cargos. Furthermore, KDELR has been predicted to be a GPCR, but does not appear to share homology with proteins in the family. However, distant homology to the SWEET family of sugar transporters has been found, suggesting these ‘receptors’ are in fact transporter like proteins. Furthermore, this project has the potential to involve live cell imaging experiments (in collaboration with Prof. Francis Barr) to test hypothesis borne from the structural and biochemical data.

Amount: £0
Funder: The Wellcome Trust
Recipient: University of Oxford

Investigating the role of Kv1.6 in pain pathways 31 Jan 2017

Kv1.6 is a member of the Shaker-like Kv1 potassium channel protein family. Widely expressed in the nervous system, these channels have delayed outward rectifier properties and evidence indicates that they act to suppress action potential firing. In the field of neuropathic pain, of which neuronal hyperexcitabilty is a common hallmark, these channels are of interest as their malfunction or downregulation may contribute to the disease pathophysiology. While less is known of Kv1.6 than other subunits of the same family, it has recently been reported that this channel is upregulated following nerve injury, signifying some role for Kv1.6 during the time after injury. Pharmacological inhibition at this stage indicates a functional role for Kv1.6 in restoring hypersensitive pain thresholds somewhat towards more normal values. Having already conducted some preliminary research on the Kv1.6 knock-out mouse, I will employ various in vivo, ex vivo and in vitro techniques from behavioural chronic pain models to electrophysiology, calcium signalling and gene/protein expression analysis in order to further probe the importance of this channel in health and disease, and to determine its sites of action amongst the various neuronal populations along somatosensory/pain pathways in the peripheral and/or central nervous system.

Amount: £29,760
Funder: The Wellcome Trust
Recipient: University of Oxford

Pancreatic Alpha Cells and Glucagon Secretion in Mouse Models of Diabetes 31 Jan 2017

Type 2 diabetes (T2DM) is characterised by insufficient insulin secretion, insulin resistance and dysregulated glucagon secretion. The processes underlying alpha-cell dysfunction in T2DM, leading to elevated glucagon secretion and exacerbated hyperglycaemia, remain poorly understood. With evidence indicating that electrical activity, Ca2+ signalling and hormone secretion occur somewhat independently of each other (at different times, under different conditions), the nature of alpha-cell signal transduction is unclear. We will generate mice expressing the Ca2+ sensitive fluorescent indicator (GCaMP) under the glucagon promoter and will measure Ca2+ oscillations in isolated islets alongside electrical activity. Following fluorescence activated cell sorting (FACS) of whole islets, electrical activity and Ca2+ signalling of isolated alpha-cells will be performed to assess the impact of loss of paracrine effects on alpha-cell function. mRNA and protein expression will also be measured in isolated cells to verify the specificity of marker expression. Also all experiments will be conducted in diabetic mice and littermate controls to assess the effect of hyperglycaemia and beta-cell dysfunction on alpha-cell physiology. These studies, conclusively characterising the nature of stimulus secretion coupling in pancreatic alpha-cells, and its disintegration in T2D, will provide valuable insight into the process underlying pancreatic dysfunction and possible opportunities to reclaim glycaemic control.

Amount: £39,978
Funder: The Wellcome Trust
Recipient: University of Oxford

Understanding the cellular and molecular basis of epithelial migration using the Angiomotin mutant 31 Jan 2017

Epithelial morphogenesis is a complex process involving tissue-level integrity and dynamically coordinated morphogenetic change. The visceral endoderm (VE) in the mouse embryo is one such tissue that undergoes morphogenesis required for embryonic development. In angiomotin (amot) mutants, there is a characteristic abnormal furrowing in the anterior VE accompanied by aberrant apical build up of actin. This leads to reduced displacement of anterior VE cells and embryonic lethality. We do not understand the cellular and molecular basis for abnormal cell migration in this mutant, nor what proteins AMOT interacts with in this context. To address these questions, I will initially make detailed quantitative observations of cell behaviour by analysing light sheet and confocal microscopy data, using automated cell segmentation and tracking approaches I am developing. I will explore actin dynamics using FRAP and laser ablation on mutant and wild-type embryos. To understand the molecular pathway through which AMOT regulates epithelial cell behaviour, I will use a phosphoproteomic approach to identify interacting proteins using wild-type and amot mutant ES cell-derived embryoid bodies. Select interacting proteins will be tested for their role using small molecule inhibitors where available, or CRISPR mediated mutagenesis in ES cells and mice.

Amount: £31,026
Funder: The Wellcome Trust
Recipient: University of Oxford

Early lineage decisions during mouse gastrulation 31 Jan 2017

Diversification of mesoderm and endoderm subtypes occurs at the outset of mouse gastrulation as epiblast cells migrate through the primitive streak (PS). The underlying inductive signals, gene-regulatory networks, and epigenetic modifications that direct lineage diversification at this early stage remains ill-defined. The aim of this project is to dissect the molecular mechanisms that underpin cell fate diversification, as cells egress the PS, by investigating the function of T-box transcription factors (TF), Eomesodermin (Eomes) and Brachyury (T). Eomes and T are expressed in the PS and Eomes is required for specification of cardiac mesoderm (CM) and definitive endoderm (DE). Single cell lineage tracing and RNA-seq experiments will be completed to define the potency and heterogeneity of Eomes and T expressing progenitors. The functional role Eomes plays in haematopoiesis will also be investigated using multiple gain and loss of function experiments. Finally, we will investigate context dependent Eomes binding sites and interacting partners. Eomes tagged mouse embryonic stem cells (mESC) will be differentiated into CM or DE progenitors and with them we will perform ChIP-Seq, RNA-Seq and immunoprecipitation-mass spectrometry(IP-MS). The experiments proposed will help resolve the functional and molecular roles T and Eomes play during early stages of lineage diversification.

Amount: £45,812
Funder: The Wellcome Trust
Recipient: University of Oxford

Functional dissection of a single enhancer sequence at the mouse a-globin locus 31 Jan 2017

Enhancers are cis-regulatory DNA elements that bind transcription factors and chromatin remodelers and drive expression of target genes in a spatiotemporal context. Despite their importance in cell fate specification in development and differentiation, up to now enhancers have largely been studied in assays that are poor predictors of their in vivo requirement and function. In this project, I will probe enhancer sequence requirements for gene regulation in situ, using the mouse a-globin locus as a model. In order to directly dissect the critical sequences, I will first produce a mouse where a-globin expression is driven by a single element (R2) by deleting the other four endogenous enhancers (R1, R3, R4 and Rm). After validating this mouse for its chromatin environment and transcriptional output, I will perform a high-throughput screen of modified R2 enhancer sequences for transcriptional output using an ES cell in vitro haematopoietic differentiation system. From these results I will select three mutant sequences for molecular characterisation in vivo. Resulting data from primary mouse tissues will provide insight into transcription factor binding interdependence, recognition site ‘grammar’ and how binding affects the enhancer-promoter interactions critical for transcription.

Amount: £37,583
Funder: The Wellcome Trust
Recipient: University of Oxford

Dissecting Polo kinase recruitment to the centrosome 31 Jan 2017

Polo kinase is an important cell cycle regulator and it is essential for the correct assembly of centrosomes, major cell organisers. Centrosomes are formed by a pair of cylindrical centrioles surrounded by pericentriolar material (PCM). Polo controls PCM assembly (at least in part through Cnn phosphorylation) and also centriole disengagement and assembly. How Polo is recruited to centrioles and centrosomes is mysterious. During my rotation I have obtained evidence that the PCM protein Spd-2 is necessary for Polo recruitment to centrosomes. During my project I aim to characterise if Polo binding to Spd-2 is necessary for Cnn phosphorylation and correct PCM organisation, what happens when Spd-2 cannot bind Polo and what upstream regulators facilitate this interaction. Furthermore, I aim to identify the other centriole/centrosome proteins involved in Polo recruitment. To do this, I will make use of biochemical assays and advanced microscopy techniques, coupled with fly genetics and a powerful mRNA injection assay to rapidly test the effects of different mutants in fly embryos. Ultimately, I hope to be able to describe in molecular detail which proteins are phosphorylated by which kinases to allow Polo to be recruited to fulfil its many functions at the centrioles and centrosomes.

Amount: £0
Funder: The Wellcome Trust
Recipient: University of Oxford

Inducing and disrupting white matter plasticity 31 Jan 2017

Recent evidence has shown that dynamical changes in white matter underlie the learning of motor skills. This has been observed both in humans with Diffusion Tensor Imaging (DTI, Schulz et al., 2009) and in rodents with histology (Sampaio-Baptista et al., 2013). These findings raise questions on which conditions are necessary for WM plasticity induction and how the plasticity they induce is modulated by genetic factors. Can an artificial induction of white matter plasticity in humans lead to behavioural changes, even without actual motor experience? To answer this question, the first aim of my DPhil will be to induce white matter plasticity at rest; more specifically, I will attempt to do so through fMRI neurofeedback, and will monitor the outcome with DTI and other myelin-specific MRI modalities. Given that white matter is often implicated in psychiatric disorders, a second question regards whether genes related to mental health conditions (in this case, NR1), can influence how white matter responds to learning, and possibly underlie a deficient or maladaptive response. Therefore, the second aim of my DPhil will be to investigate whether WM plasticity is affected in a mouse model of white matter-specific knockout of NMDA receptors.

Amount: £83,102
Funder: The Wellcome Trust
Recipient: University of Oxford

The role of regulatory T cells in immunity to malaria 31 Jan 2017

In malaria vaccine trials conducted in the target population of semi-immune people from endemic African countries, vaccine immunogenicity is sometimes substantially reduced compared to European malaria-naïve participants. This could result from the suppression of vaccine-induced immune responses by regulatory T cells (Tregs) acquired through prior malaria infections, however there are few studies in man which have previously explored this. Using samples from controlled human malaria infection studies in semi-immune Kenyan individuals, we will investigate how Tregs affect natural immunity to infection and see if increased Treg responses correlate with vaccine efficacy following malaria challenge in participants with varying prior exposures to malaria. We will also directly compare the effect and induction of Tregs in different pre-erythrocytic candidate vaccines and adjuvants to understand how vaccine-specific effects might affect Treg responses. Additionally, we will investigate if malaria-induced Tregs affect responses to other childhood vaccines. Single cell transcriptomic analysis using the Fluidigm platform will be employed to explore the phenotype and functional heterogeneity of Tregs. This will provide insight into the mechanisms by which Tregs are involved in immunity to malaria. This work will have important implications for the design and evaluation of malaria vaccines for use in endemic populations.

Amount: £0
Funder: The Wellcome Trust
Recipient: University of Oxford

Not just a virulence factor: capsule biology across the Neisseria genus 31 Jan 2017

This project seeks to address the relatively unexplored topic of the genetics, function and evolutionary history of the Neisseria polysaccharide capsule, beyond its established role as a virulence factor in Neisseria meningitidis (Nme). This will be achieved by examining capsular types not associated with disease, both from Nme, and capsules recently discovered in the commensal Neisseria species. The first goal is to complete genetic and phenotypic characterisation of the novel commensal capsular types. Once this is established, a key goal is to seek comparisons between these novel capsules and those of Nme within the coding sequences and regulatory regions, and at the structural level. I also plan to address the question of what the role of capsule is in colonisation and transmission, given that it most likely was not selected for its virulence properties. Finally, I seek to build a clearer history of the acquisition and evolution of capsule in Neisseria. This will bring forward new insights into the roles of capsule in normal, healthy colonisation of the nasopharynx, both by Nme and the strictly commensal Neisseria species. This work may also have implications for our interpretation of Nme dynamics and the rare transition to a state of disease.

Amount: £0
Funder: The Wellcome Trust
Recipient: University of Oxford

Understanding mechanisms that drive pain perception in early human development 11 Jul 2017

Pain in infancy has negative long-term consequences and its prevention is a clinical priority, but adequate treatment requires mechanistic understanding of the structural and functional development of human nociceptive circuitry. Recent scientific and technological advances provide insights into how noxious information is transmitted to the infant brain, providing a platform to ask how intrinsic brain network connectivity and the environment affect noxious-evoked brain activity, behaviour and ultimately pain perception in the developing infant nervous system. The fellowship goal is to understand the mechanisms that drive and modulate pain perception in early human development. I will ask whether inherent differences in how the brain behaves at rest influence variability in noxious-evoked activity, and will determine how this relationship is altered by environmental factors and pathology. I will establish how the development of structural and functional network connectivity alters noxious-evoked brain activity, and influences the dynamic relationship between brain activity and behaviour. I will translate this mechanistic understanding into clinical practice by conducting a clinical trial of an analgesic (fentanyl) during a minor surgical procedure, and will establish whether our newly-developed measures of noxious-evoked brain activity are suitable for use in infant analgesic dose-finding studies.

Amount: £60,000
Funder: The Wellcome Trust
Recipient: University of Oxford

Enhancing the WT MOP Vietnam's Engagement Programme 30 Aug 2017

In 2015 the WT Major Overseas Programme Vietnam was awarded a renewal of its Core funding. The MOP has a history of successful public engagement, funded through International Engagement awards and from industry sponsorship. However, with the introduction of the Provisions for Public Engagement funding scheme, we applied for funding for engagement at an institutional level, enabling us to create a 5-year strategic plan for developing engagement capacity within the MOP and in the region. Now, 20 months into the award, we reflect on activities to date, and plan strategically for the second part of the programme. The 5-year public engagement programme includes a schools engagement programme (SEP) and a capacity building programme (CBP), both of which have proved to be very successful and highly valued by our local government and school partners. The third focus has been to develop researcher capacity for engagement – through small grants and offering training and mentoring. We have had a good uptake of these ‘seed awards’ from MOP researchers and increasing interest in engagement from researchers at local institutes in Vietnam. Schools Engagement: The SEP has been very successful (http://www.mediafire.com/file/td3kaomtu9t7ia7/Application.7z), in particular: afterschool science clubs; weekly science articles in a children’s magazine; science theatre; and lab visits enabling young people to interact with scientists. The SEP has also included ‘I’m a scientist, Get me out of here’ - a competition linking children and scientists, run with Gallomanor UK (https://imascientist.org.uk) (https://www.youtube.com/watch?v=n--SJOtFm1w). Capacity building: The CBP was developed in recognition that much of the ‘front-line’ contact with patients and communities enrolled in clinical trials or cohort studies is from hospital or government study staff. In response we have started a CBP to train and support hospital health care workers (HCMC), community-based data collectors (Nepal) and local vets (in provinces where we conduct research on zoonosis). As the funding for the IAS project and other awards come to an end, we need additional funding to support the current PE team. This application is for additional staff salary costs and to run PE workshops to develop engagement capacity across the region.

Amount: £343,289
Funder: The Wellcome Trust
Recipient: University of Oxford

Understanding the link between CpG islands and gene transcription 28 Nov 2017

CpG islands(CGIs) are epigenetically specified elements that are intimately associated with over two-thirds of human gene promotors, yet whether CGIs regulate gene expression has remained enigmatic. This gap in our understanding of gene promoter function has serious implications for human health given that CGIs are perturbed in cancer and other debilitating human diseases. We have recently discovered that CGIs are recognized by reader proteins which can regulate gene expression. Capitalising on these advances, I will now discover how CGIs and the proteins that read them control the transcriptional machinery at gene promoters. I will achieve this transformative new mechanistic understanding through a multidisciplinary and hypothesis-driven programme of research that builds on a series of exciting new and unpublished observations to discover how CGIs function to activate(Aim1) and maintain(Aim2) transcription, and test whether CGIs create gene expression switches(Aim3). These new discoveries will help to redefine our understanding of how gene promoters function to control gene expression and will provide the basis on which new therapeutic interventions can be developed for diseases where normal CGI biology is perturbed.

Amount: £2,072,282
Funder: The Wellcome Trust
Recipient: University of Oxford

CHIM - Shigella sonnei in Vietnamese adults - Development funds 17 Nov 2017

Diarrhoea remains a major cause of childhood morbidity and mortality globally. The vast majority of the 2 billion annual diarrhoeal infections occur in low and middle-income countries (LMICs). Members of the genus Shigella are key agents of diarrhoea in LMICs, and S. sonnei is replacing S. flexneri as the predominant species globally. There is a necessity to improve our knowledge of S. sonnei infections in LMICs, with a specific requirement to better understand host-pathogen interactions and the natural history of disease in a setting where the organism is well understood, well described, and associated with a significant disease burden. Therefore, we aim to establish a Controlled Human Infection Model (CHIM) of S. sonnei diarrhoea in healthy Vietnamese adults. This is an innovative project will be the first CHIM study conducted at the Vietnam MOP. Therefore, it is imperative that the project is carefully designed in consultation with all relevant stakeholders. In order to ensure that the proposal is developed to the required standard in the timeframe available, I am requesting funds to employ an international postdoctoral assistant, with a background in microbiology and clinical research on a consultancy basis.

Amount: £21,120
Funder: The Wellcome Trust
Recipient: University of Oxford

Control of T cell responses by accessory receptors revealed by phenotypic models 11 Jul 2017

T cells orchestrate immune responses crucial for the elimination of infections and cancers. They do this by initiating a diverse set of effector responses when their T cell surface receptors (TCRs) recognise these threats. It is now appreciated that a large number of other, "accessory", receptors shape these responses. Indeed, the remarkable clinical success of checkpoint inhibitors and chimeric antigen receptors is based on perturbing accessory receptor signalling. Despite extensive research into the underlying biochemistry, we have yet to formulate canonical models of signalling that can predict how accessory receptors shape T cell responses. Here, we propose to use a mathematical method known as adaptive inference to identify signalling models directly from T cell response data, without prior biochemical assumptions. The method produces what we term phenotypic models because it coarse-grains over molecular information. These models provide effective pathway architectures showing how accessory receptors integrate (or not) with TCR signalling to shape response phenotypes. This will move the field beyond the current stimulatory/inhibitory binary paradigm of accessory receptors. The work offers a different way to study receptor regulated signalling pathways and the predictive power of the phenotypic model will be exploited for T cell-based therapies.

Amount: £1,783,754
Funder: The Wellcome Trust
Recipient: University of Oxford

ISA-InterMine: accelerating and rewarding data sharing 06 Jul 2017

There is a growing recognition that research should be carried out in an open fashion, making data available early and in a reusable form to maximise worldwide research output. However, fulfilling this promise requires front-line researchers to comply with current data management standards as required by the data policies of funders and journals. These are additional burdens to research that will give them little immediate return. Thus we propose to create a cloud-based, open-source, extensible data collection and presentation platform that will provide scientists with: (1) immediate reward for their annotation efforts through sharable data visualisation, integration outputs and exploration tools; (2) standardised web services to facilitate script-based data manipulation and analysis; (3) an easy-to-use pipeline for preparing their data for publication; (4) incentives to improve data quality, accessibility, and machine-actionability at the appropriate level of granularity; and (5) allow institutions and other parties to host the platform to ensure its availability and reliability. We will do this by building on the success and complementarity of the ISA tools suite (Oxford) and the InterMine platform (Cambridge) to make it quicker and easier to generate rich integrated dynamic web sites at single paper/lab scale up to consortium scale.

Amount: £504,250
Funder: The Wellcome Trust
Recipient: University of Oxford

Evidence for a pathogenic link between lung inflammation and autoimmune arthritis 27 Apr 2017

Our overall objective is to experimentally identify the early immunological events which trigger a spontaneous breach of tolerance in genetically susceptible mice, and, which can be linked to the development of arthritis. We will analyse the immunological reactivity of the lungs of susceptible mice to environmental stresses such as microbes. Our purpose is to identify early biomarkers of autoimmune initiation, which will be assessed for suitability as treatment targets with translational relevance. We will seek to determine the benefits of administration of anti-microbial agents for amelioration of lung inflammation with a view to prevention of autoimmune disease progression.

Amount: £0
Funder: The Wellcome Trust
Recipient: University of Oxford