- Total grants
- Total funders
- Total recipients
- Earliest award date
- 17 Jan 2014
- Latest award date
- 30 Sep 2018
- Total GBP grants
- Total GBP awarded
- Largest GBP award
- Smallest GBP award
- Total Non-GBP grants
Genetic association studies focusing on common variation have uncovered only a fraction of proposed trait heritability. Some of this so-called missing heritability will be found within rare variation in the population. This hypothesis is supported by the facts that recent explosive population growth has increased the population burden of rare variants and deleterious variants are kept at low allele frequencies. All genetic susceptibility to disease is caused by alterations to the genes or their expression and for this reason it seems fruitful to focus an association study on the genes themselves. Any associations found are then directly informative about the molecular basis of disease without the need for fine mapping. The proposed project aims to develop a statistical method to find genes associated with disease by analysing the rare variation present in a case-control cohort. We aim to extend existing methods by including a previously unconsidered parameter; the position of the variants in a gene. In scenarios where differences in clustering or distribution of variants are observed between cases and controls, this method will have a substantial increase in power. This technique will be useful for elucidating the molecular mechanisms causing the disease and thus discovering new therapeutic targets.
Design and evaluation of a modified vaccinia Ankara vector therapeutic vaccine for hepatitis B immunotherapy 30 Sep 2018
Hepatitis B virus (HBV) is a serious global health problem, with approximately 240 million people chronically infected. Long-term infection can lead liver failure, cancer and death. Current therapy controls but does not eradicate the infection. T cells are a type of immune cell necessary to fight HBV. During chronic hepatitis B these cells become less active. Checkpoint inhibitors are a form of immunotherapy that enables T cells to function again. In a study of woodchucks infected with a similar virus to HBV, treatment with vaccine and checkpoint inhibitor lead to better control of the virus. This project aims to use this combination of vaccine and checkpoint inhibitor, to treat patients with chronic HBV. A vaccine using a virus to carry the HBV proteins has been developed and shown to generate good immune responses in mice. We plan to develop a second vaccine to boost this response and test the vaccines together with checkpoint inhibitors in mice infected with the HBV virus. This will allow us to assess how effective this is at eradicating HBV. If the results from this study are promising, this could pave the way for clinical trials in humans with chronic HBV.
The ATP-sensitive potassium (KATP) channel is a plasma membrane protein present in beta cells of the pacreas which plays a key role in insulin secretion. KATP acts as a metabolic sensor, alerting the beta cells when blood glucose raises too high and stimulating them to release insulin. In diabetes, normal KATP function is disrupted and beta cells no longer secrete insulin properly in response to blood glucose levels. The molecular structure of the channel is closely linked to its function; there have been several genetic studies linking various mutations (which often only affect one molecule in the channel!) to neonatal diabetes or increased propensity to type II diabetes. Our research aims to identify precisely how these small mutations can have such drastic changes in the activity of the channel by using a combination of fluorescent labels and channel current measurements to watch the KATP channel move in real time. We can then try to construct a model of how the channel converts different stimuli into movements, and how this is affected in mutations linked to diabetes.
There is an urgent need to develop new antibiotics against multidrug resistant Gram-negative bacteria such as Pseudomonas aeruginosa and Klebsiella pneumoniae. These organisms are major causes of pneumonia and sepsis, with recent reports identifying hospital isolates of each resistant to all known antibiotics. The present research focuses on the mode of action of a family of antibiotic proteins known as nuclease bacteriocins that have not been developed as antimicrobials, but show promise in animal models of infection. Nuclease bacteriocins are species-specific toxins that are used by bacteria to compete with their neighbours. Although folded proteins these molecules are capable of penetrating the defences of Gram-negative bacteria to deliver an enzyme to the organism’s cytoplasm to degrade essential nucleic acids by an unknown mechanism. Two types of nuclease bacteriocin will be investigated, pyocin AP41 which targets Pseudomonas aeruginosa, and klebicin G which targets Klebsiella pneumoniae. Preliminary computational and experimental work on pyocin AP41 has identified potential candidate proteins involved in its import. This will be followed up with structure and function studies of AP41, a dissection of its import mechanism and new studies on klebicin G, a nuclease bacteriocin that has only recently been identified.
I plan to explore the effect of nicardipine, a brain-penetrant calcium channel antagonist, on mood instability and its cognitive/neural correlates. This research (carried out within the Collaborative Oxford Network for Bipolar Research to Improve Outcomes [CONBRIO]) has a number of goals. In addition to studying effects of L-type calcium channel (LTCC) antagonism on mood instability, I will investigate effects on sleep/cognition, and on brain activity measured by functional imaging. Rationale is provided by (a) considerable evidence for calcium signalling abnormalities in bipolar, (b) current mood stabilisers correct some of these abnormalities, (c) calcium channel genes contribute to the basis of bipolar as well as to memory/sleep. Moreover, LTCC antagonists are used for heart disease and available for experimental studies; there have also been early studies in bipolar disorder but no robust results. I will study volunteers screened for high mood instability and risk CACNA1C genotype, and assess mood, sleep, cognition, and neural activity, before and after randomisation to nicardipine or placebo. Results will inform whether trials of LTCC antagonists for bipolar disorder are indicated, and their likely efficacy/tolerability. I will learn to conduct a randomised trial, principles of experimental medicine, and aspects of cognition, circadian-biology, and functional brain imaging.
The Effect of Priorizing Information in Working Memory on Later Behavioural Interference 31 May 2018
This experiment will investigate how prioritised information is represented in working memory (WM) through looking at the serial dependence effect. Myers and colleagues (2017) have suggested that items which are prioritised in WM are transformed into action-ready representations. Therefore, the theory predicts that the difference between prioritised and non-prioritised representations in WM will be reflected in behavioural findings. The serial dependence effect occurs when visual information from the recent past biases perception and behaviour at the present moment (Fischer & Whitney, 2014). If prioritised WM items were stored in an action-oriented format, we predict it will show these interference effects in behaviour more than non-prioritised information. By using an orientation adjustment paradigm, we will measure the serial dependence effect for prioritised WM items (which have been retro-cued) versus non-prioritised WM items. In addition, we will vary the type of testing (forced choice versus free recall), predicting that more interference will occur when the tests are the same than when different, due to the action-based nature of the WM representation. Initially we will use behavioural measures (reaction times) to measure the interference effects, extending to EEG to measure neural evidence for the carry-over effects.
Delineating meiotic gene expression of male mice 31 Jan 2017
The mechanisms of meiosis have important consequences for the evolution, fertility, and speciation of sexually reproducing organisms. However, the full gene expression programme of meiosis is not currently known, partly due to heterogeneity of the cell population which is averaged by bulk tissue RNA sequencing or micro-array analysis. It has recently become possible to transcriptionally profile thousands of single cells using RNA sequencing, raising the possibility of identifying the gene expression profile associated with different stages of meiosis and hence delineating the full gene expression programme. The identification of genes expressed in meiosis and its sub-stages may aid in the discovery and understanding of meiotic mechanisms. I will use pseudotime ordering and latent factor analysis type methods to analyse this data and comprehensively define the transcriptional profile of male meiosis.
This PhD focuses on developing statistical methods to discover gene – environment (G-E) interactions. To date there has been some interest in testing for G-E interactions in animal models, but limited success in uncovering examples of G-E interactions in humans. This is in part due to the problem of exposure assessment, or rather, because representative data on the environment of a number of individuals over a lifetime has been hard to acquire. However the data recently made available by the UK BioBank, on over 500000 individuals and a wide array of environmental covariates, may now make it possible to detect these interactions. We aim to use a Bayesian methodology to first test a number of known models, such as a random effects model, against the dataset. We will then attempt to use a Gaussian Process Regression model to identify covariates involved in G-E interactions. This approach is advantageous as GPR is non-parametric, thus avoiding the curse of dimensionality, and places no assumptions on the order of interactions. However as this method currently scales in a cubic manner following the number of samples, significant computational challenges remain.
Z-DNA is an alternative conformation of the DNA double helix. The existence of proteins containing Z-DNA binding domains (ZBDs) suggests that nucleic acids in this conformation have important but so far uncharacterised biological functions. One such protein is the innate immune protein Z-DNA binding protein-1 (ZBP1/DAI/DLM1). We have evidence that ZBP1 mediates virus-induced necroptosis in a manner dependent on its ZBD. This function of ZBP1 is sensitive to RNase but not DNase treatment, suggesting that the trigger is Z-RNA. We therefore hypothesise that Z-RNA may represent a novel pathogen-associated molecular pattern sensed by ZBP1. The aim of my PhD will be to further characterise this role of ZBP1 and to investigate the source and nature of the Z-RNA being sensed. In particular, we will ask if this Z-RNA is of viral and/or cellular origin, and whether specific sequence motifs are recognised by ZBP1. Another question pertains to the relevance of ZBP1 in diverse human virus infections. In parallel to these in vitro studies, we are developing a knock-in mouse in which the ZBD of ZBP1 is mutated. I predict that these animals will be less able to control viral infection due to a failure of ZBP1-mediated virus-induced necroptosis.
The project aims to utilise mutations identified in congenital myasthenic syndromes to study the interactions of the muscle acetylcholine receptor (AChR) and its anchoring protein, Rapsyn.Widely quoted publications suggest that Rapsyn interacts with the M3-M4 intracellular loop of the AChR alpha, beta and epsilon subunits, however we have identified several kinships in which mutations in the M3-M4 intracellular loop of the AChR delta subunit underlie a phenotype that mimics myasthenic syndromes caused by mutations in RAPSN. The project will use in vitro mutagenesis and cell culture experiments to investigate how mutations in CHRND impair agrin induced-AChR clustering in C2C12 myotubes. Variants identified in CMS patients, ie. p.(Glu381Lys) or p.(Arg376His) will be incorporated into expression constructs and transfected in chrnd-/- C2C12 cell that have been created using CRIPR/Cas9 techniques. Similar experiments will be performed following in vitro mutatagenesis that is designed to disrupt potential PKA, PKC and tyrosine kinase phosphorylation sites within the M3-M4 intracellular loop of the delta subunit. The effects of the mutations on agrin-induced AChR cluster formation will be assessed by using fluorecent-labelled alpha bungarotoxin and microscopy. This short project will provide novel data on how mutations within the AChR itself can impair the cluster formation.
Many fundamental cell biological processes take place at the juxtaposition of two membranes. Having methods for examining such systems is therefore of considerable interest, but few experimental techniques have the required spatial and temporal resolutions. Molecular dynamics (MD) simulations can theoretically offer insight into the time-dependent behaviour of membrane interfaces at near-atomic detail. In this work, we will develop the methodology for investigating membrane interfaces using multiscale MD simulations. During the first stage of the project, we will use a structurally and experimentally well-characterised system to develop and refine the methodological underpinnings of the simulation work. Specifically, we will use the available data on the myelin-axon interface bridged by myelin-associated glycoprotein (MAG) and its cognate ganglioside. Once the model is established, we will use it to address specific questions about this system, including the energetics of ganglioside binding, and the roles of various MAG post-translational modifications, including an unusual Trp C-mannosylation in the ganglioside-binding domain.
Nuclear Magnetic Resonance has traditionally provided much information about metabolism in living systems ranging from enzymes to humans, owing to its unique chemical specificity. Due to the Boltzmann distribution, conventional magnetic resonance is inherently insensitive and resolving many metabolic reactions is impossible in vivo. Hyperpolarised Magnetic Resonance with Dynamic Nuclear Polarisation (DNP) provides a way to overcome the Boltzmann distribution by using low temperature quantum mechanics. A 13C-labelled metabolite is prepared at 0.8K, rapidly melted, and then injected into a living system in an MRI scanner. The 13C labelled molecule can then be briefly tracked through space, time and biochemistry before relaxing back to thermodynamic equilibrium. Currently, the field quantifies DNP data poorly because it estimates pseudo-first-order metabolic rate constants and neglects physical facts such as membrane transport. My project would propose differential equations that model the underlying biology and physics, solve them using advanced Bayesian methods and produce quantitative estimates of metabolic rate constants with bounded uncertainty. The aim is to quantify unidirectional metabolic fluxes on data acquired in vivo in the human heart, with external validation. The key goals would be the production of mathematical models, comparison of fitting algorithms and in vitro enzymatic comparison.
Social anxiety in adolescents: Testing aspects of the cognitive model and developing an Internet version of Cognitive Therapy. 30 Sep 2018
While it is common for teenagers to report feeling somewhat self-conscious and worried about what others think of them, for some adolescents social anxiety can be overwhelming and markedly interfere with their day-to-day life. Around 4% of young people experience clinical levels of social anxiety by the age of 18. Their success at school and their relationships with family and friends are all seriously affected. It is therefore essential that we have effective treatments for this disorder. Unfortunately, evidence suggests that many young people do not get better with the talking treatments that are available. In contrast we know that Cognitive Therapy (a type of talking treatment) is very effective with socially anxious adults. Cognitive Therapy was developed to target key factors that cause and maintain the illness in adults. I would therefore like to answer two questions in my research. First, are the factors that are important in maintaining social anxiety in adults also important in adolescent social anxiety? Second, can we develop an accessible and effective version of Cognitive Therapy for adolescents online? We will test if Cognitive Therapy works by comparing its effects to online stress management.
Impact of accessory receptors on close-contact formation and ligand discrimination by T cells 30 Sep 2018
The immune system of vertebrates has evolved a remarkable ability to adapt to new pathogens. The ability of T cells to discriminate between self and foreign antigen is crucial for adaptive immune responses. However, we do not fully understand the mechanism behind T cell activation. Interactions between TCR and other surface molecules are responsible for activation at the interface between T cell and antigen-presenting cell (APC). Unlike previously thought, this interface is not just two flat membrane surfaces, but the T cell engages the APC with numerous small membrane protrusions, that form microdomains called close contacts. We hypothesise that the temporal and spatial properties of those microdomains are important for antigen discrimination. We will measure the effect of accessory receptors on the proximal signalling indirectly by stimulation assays in combination with mathematical modelling. Furthermore, we investigate close contacts directly my microscopy, measuring properties like size, distribution and movement. We believe our results will improve the comprehension of T cell activation, a process central for the defence against invading pathogens and cancer and the prevention of autoimmunity.
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.
I will study the mechanism of protein export in the injectisome by elucidating the structure of one of its central components, the export gate. The injectisome is a large, molecular nanomachine which injects effector proteins into cells of the hosts of many bacterial pathogens. The export gate is the protein channel at the heart of this syringe-like type three secretion system (T3SS), and the structure of this channel will provide important insight into how it may function: For example, the complex may be actively involved in guiding substrates across the inner membrane or merely provide a passive conduit for unfolded proteins. In addition, there is the question of how such a complex may be "plugged" during periods of inactivity. I have recombinantly produced the export gate complex from the injectisome and also from the flagellum which possesses a related T3SS (rotation reports), and the tools of structural biology will continue to be used on them. Furthermore, I will purify larger complexes, consisting of the export gate and other T3SS proteins. This will enable me to study the structure and interactions between the gate and subunits such as the substrate specificity switch.
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.
Genome-wide association studies (GWAS) have found over 100 loci associated with type 2 diabetes (T2D), but the causal mechanisms of the disease remain unclear. Pancreatic islets play a role in the development of T2D, and although the contribution of developmental processes to disease susceptibility has not yet been established, several of the GWAS loci localise near known genes for islet development. I will investigate the role of islet development in T2D susceptibility through analysis of iPSC differentiation models of human endocrine pancreas cells (concretely, beta-cells). I will use transcriptomic and regulatory information to identify genes involved in their development, finding the most influential genes in each stage of differentiation, and characterizing temporal patterns and networks of gene expression. I will investigate specially the presence and abundance of genes located in T2D or fasting glucose -associated loci. The bioinformatics analyses will generate a list of important genes for endocrine pancreas development. I will then evaluate experimentally the function of some of these genes in development and T2D risk. Using CRISPR to modulate the expression of candidate targets in specific stages of development, either activating or interfering gene expression, I will measure how this alters the transcriptome and beta-cell differentiation.
The recent evolution of artemisin-resistant strains of P.falciparum in Southeast Asia have highlighted the need for a deeper understanding of the genetic epidemiology of malaria and for more strategic implementation of malaria control resources. Assessment of the efficacy of these control efforts requires accurate and standardised estimates of epidemological parameters, such as transmission intensity. As of now, these estimates are not standarised across regions, are often labor-intensive, and work poorly in low-transmission areas. Genomics, via the sequencing of geolocated patient blood-derived parasites, has the power to provide key insights into the migration and evolution of malaria. Moreover, the rich data provided by genome sequencing represents an attractive alternative from which epidemological parameters can be calculated. To this end, my DPhil will involve the development and benchmarking of genetic epidemological models of malaria built from thousands of spatially-referenced malaria genomes collected from across Africa and Asia. These models will analysze the structure of regional genetic diversity to make estimates of parasite migration, population size and transmission intensity. Estimates will be systematically compared to field gold-standards and against simulated environments of malaria infection. The overarching goal is to guide malaria eradication policy through improved disease survelliance and control effort assessment.
With the rise in multidrug resistance as a result of broad spectrum antibiotic use, bacteriocins have received attention as potential new antibiotics. Bacteriocins are bacterial toxins that kill closely related strains and species. Pyocin S5, a bacteriocin targeting Pseudomonas aeruginosa, a leading cause of multidrug resistant nosocomial infections, was shown to be effective against pneumonia in mice. Pyocin S5 depends on the iron receptor FptA and kills cells by depolarizing the inner membrane. Little else is known about its mode of action. Pyocin S5-producing cells harbour a gene encoding a small immunity protein, predicted to localize to the inner membrane, which protects from the action of this pyocin. In this PhD project, I aim to investigate the molecular mechanism of pyocin S5 binding to P. aeruginosa cells, its mechanism of translocation to the periplasm and the immunity protein’s mode of action. These aims will be approached using a combination of structural and biophysical methods as well as functional assays and a variety of fluorescence microscopy techniques.