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Recipients:
University of Cambridge
Amounts:
£500 - £1,000

Results

Clinical Characterisation of a Broad Spectrum of Genetic Variation in the General Population 30 Sep 2018

Inborn errors of metabolism (IEM) are severe and extreme changes in metabolism caused by mutations in a single gene. Recent large-scale human studies have shown that genes causal for IEM are associated with nutrients, or ‘metabolites’, in the blood. However, whether these associations cause disease or adverse health outcomes is unknown. In this project, I will use IEM genes identified in these studies to link genetic variation to clinical features in a large human population. To do this, I will assemble a list of IEM genes of interest that were identified in the literature and in large population datasets. I will then test for associations between the variants I find in these genes and a wide range of clinical features found in open-access population datasets. As the IEM genes used in this study have been associated with blood metabolites previously, linking variants in these genes to clinical features will shed light on the molecular mechanisms underlying genes and disease in the general population. Understanding how genetic variation affects disease will help identify novel therapeutic targets and enable health professionals to better manage disease risk.

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

Investigating non-canonical programmed ribosomal frameshifting in porcine reproductive and respiratory syndrome virus 30 Sep 2018

RNA viruses are under selection pressure to maintain a small genome, however they still need to produce a variety of proteins. To overcome these conflicting pressures, many viruses use non-canonical methods of translation control. One example of this is programmed ribosomal frameshifting (PRF), in which a percentage of ribosomes, while translating a ‘slippery sequence’ slip one or two nucleotides out of frame, consequently translating the remainder of the mRNA in a different reading frame and allowing expression of more than one protein from a single gene. This is normally stimulated by a downstream secondary RNA structure, however there are two known examples of a trans-acting viral protein being used as the stimulatory element. One example is found in the genome of porcine reproductive and respiratory syndrome virus (PRRSV): an Arterivirus that infects pigs, causing an estimated annual cost to the US swine industry of $664m. I will use structural techniques such as X-ray crystallography to derive information about the RNA/protein complex, and will investigate the efficiency and mechanism of this non-canonical PRF using ribosomal profiling in parallel with RNASeq. The latter will also allow me to analyse host and viral gene expression, to examine host-virus interactions in this important pathogen.

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

Epigenetic inheritance: the influence of variable silencing of the repeat genome. 10 Apr 2018

Evidence from epidemiological studies and experiments in animal models suggests that effects of environment and lifestyle can be transmitted across generations via non-genetic mechanisms. Such mechanisms are challenging to unravel in mouse and man. In mammals, non-genetic inheritance is best exemplified by the Agouti viable yellow (Avy) mouse where phenotypic differences in genetically identical animals are caused by insertion of a retrotransposon - an endogenous retrovirus (ERV) that provides a cryptic promoter driving ectopic expression of agouti. This ERV is variably DNA methylated in different individuals causing inter-individual variation in coat colour – a non-genetic influence on phenotype. Remarkably, a memory of parental coat colour is transmitted to subsequent generations. Variable expressivity can be modulated by in utero environmental exposures. ERVs represent ~12% of the mouse genome. Inspired by Avy, we propose a research programme, supported by preliminary data, to address the following questions: Aim 1 – To what extent do mammalian ERVs exhibit variable epigenetic silencing and what is the mechanism? Aim 2 – Is this transmitted as non-genetic memory across generations? Aim 3 – Are they sensors of environmental compromise? Aim 4 – Are there implications for phenotype? Aim 5 – Does a related phenomenon occur in humans?

Amount: £2,929,851
Funder: The Wellcome Trust
Recipient: University of Cambridge

Gene Expression Heterogeneity in the Maintenance and Coordinated Differentiation of Neuromesodermal Progenitors in vivo 08 Aug 2018

Modern imaging data in biology is essentially multi-scalar in that raw image data undergoes a series of processing steps until it is at a manageable size to perform quantification and analysis. While this processing pipeline might be beneficial to one set of scientific questions, it may be inappropriate to others. Computational biologists may be interested in improving the pipeline itself, while other researchers may be interested in accessing the already processed data. Thus, a central road-block in the open sharing of large-scale imaging data is the fact that there is no one size fits all solution. This project aims to generate a web-based database that stores experimental and data descriptors together with links to the raw and processed data files. This will greatly enhance our ability to upload this data to repositories that are based placed to share the datasets in question, from the raw unprocessed data files down to feature extracted and processed data. Upon submission, the website will link to the deposited data and thereby act as an integrated platform for other researchers to access and explore the data that is available to them. Thus, researchers will be able to access our data at all levels. In built in the project is a second evaluation phase, whereby will we reach out to collaborating laboratories to assess the effectiveness of our open research platform. These will include computational biologists interested in accessing raw data files and processing pipelines, and other developmental biologists who will interact with processed datasets.

Amount: £48,563
Funder: The Wellcome Trust
Recipient: University of Cambridge

Lineage tracing and early embryonic dynamics using mosaic somatic mutations 30 Sep 2018

Every cell in the human body descends from the fertilised egg cell, and during the journey from zygote to adult cell, many mutations are acquired. The mutations that happen early enough during embryonic development will be present in some parts of the body, but absent in others. One of the difficulties of this project is the discovery of these early embryonic mutations, as they are hard to distinguish from mutations that happened later in life, noise from experimental procedures, and mutations that were already present at fertilisation. Here, we cut out small pieces of various tissues from multiple individuals using a laser and sequence its DNA. Mutations shared between different samples from the same individual are candidate embryonic mutations. Each mosaic mutation gives us information on the development of the human body and with enough mutations, we can construct and visualise the human body as a large evolutionary tree, with the fertilised egg cell as the single root, and all the adult cells as its leaves. This can give us many new insights on the developmental relationship between different organs, how many different early embryonic cells give rise to one tissue or organ and developmental lineages of different cell types.

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

The effect of nutrients on maximal fat oxidation rates in adult humans measured using indirect calorimetry. 31 May 2018

I aim to study the effects of nutrient availability and mitochondrial transport capacity on the variability of maximal fat oxidation (MFO) during exercise in healthy adults. Less than half of MFO can be predicted by variables such as gender, VO2max and body composition. There are two possible reasons for this. First, nutrient availability may have a large effect on MFO and current protocols may not adequately control for it. Second, VO2max - which combines two variables with opposite effects on MFO (oxygen uptake and fat mass) - may not be the optimal predictor. Here, I will test whether heart rates at a given power are a better predictor of MFO and whether short-term fluctuations in nutrient availability can explain some of the variability of MFO seen within the general population. Nutrient availability will be altered using a glucose meal and by glycogen depletion. I will also use nitrate supplementation to test whether MFO can be increased by induced-expression of fat oxidation enzymes. The key goals are to determine to what extent short-term changes in nutrients and the expression of fat oxidation enzymes can alter MFO and whether the resultant fat oxidation rates can be predicted using simple heart rate data.

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

Understanding the Initiation of Viral Replication & its Role in Influenza Virus Pathogenicity 31 May 2018

The development of novel strategies against influenza viruses depends on our understanding of influenza virus replication and pathogenicity. Both are directly linked to the activity of the viral RNA polymerase, which copies and transcribes the viral genome, and generate aberrant RNA products that are non-contiguous in the viral genome and strong inducers of the interferon response. Despite recent crystal structures of the RNA polymerase, we only poorly understand how it interacts with and copies the viral genome, or how it generates aberrant RNA products. This project aims to use i) structure-guided mutagenesis, ii) in vitro and in vivo activity assays, iii) cell culture-based interferon production assays, to ask if RNA polymerase residues that bind and guide the viral genome are important for the initiation of viral replication and the formation of aberrant RNA products and thereby the pathology of influenza virus infections. The project will contribute to our understanding of the mechanics of influenza virus replication and the identification of putative targets for the development of new anti-influenza virus drugs.

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

Molecular mechanisms of alternative splicing regulation 28 Nov 2017

Alternative pre-mRNA splicing (AS) is a widespread regulatory mechanism enabling individual genes to generate multiple protein isoforms. We have investigated the mechanisms controlling AS events that are regulated during the transition of smooth muscle cells (SMCs) between contractile and proliferative phenotypes. We have shown how the widely-expressed RNA binding proteins (RBPs) PTBP1 and MBNL1 regulate SMC splicing events. Recently, we identified RBPMS as a potential "master" regulator of SMC AS. RBPMS is sufficient to switch AS events to the SMC pattern and its activity is strongly modulated by its own AS and by phosphorylation. Critically, RBPMS is sufficient to switch AS to the SMC pattern in vitro. This offers a unique opportunity to determine the molecular anatomy of regulated splicing complexes. We will carry out detailed mechanistic analyses of RBPMS-regulated splicing using a combination of biochemical, proteomic, single-molecule, and structural approaches including Cryo-EM. We will identify critical regulatory interactions between regulatory RBPs and core splicing factors, and test their importance by genome editing and mRNA-Seq. In a complementary aim, we will investigate how peptide-ligand interactions equip PTBP1 to regulate AS and a range of other post-transcriptional processes, and whether a family of such peptide-mediated interactions extends to related RBPs.

Amount: £1,503,005
Funder: The Wellcome Trust
Recipient: University of Cambridge

Investigation into the role of RBM8A/Y14 in the development and function of megakaryocytes and platelets using a human pluripotent stem cell model of haematopoiesis 30 Sep 2018

Platelets are small blood cells, which cause blood to clot, preventing bleeding after injury. They are produced by megakaryocytes, large cells in the bone marrow. In people with low platelet counts (thrombocytopenia), life-threatening bleeding occurs spontaneously or after injury. Studying platelet and megakaryocyte development and function is important in understanding a) diseases causing thrombocytopenia, such as genetic disorders and other conditions, particularly cancer (and chemotherapy) and b) strokes and heart attacks, where platelets are excessively activated, forming clots that block vessels. Using stem cells (special cells capable of becoming any cell type) derived from adult skin or blood samples we grow & study megakaryocytes and platelets in the laboratory. We study a rare genetic disease, Thrombocytopenia with Absent Radii (TAR) syndrome, in which babies are born with very few platelets and abnormal bone formation (particularly the radius in the forearm). Our group discovered the cause of TAR, due to abnormalities in a gene called RBM8A, which helps cells control what proteins are produced; however precisely why this causes TAR is unclear. We believe our research will uncover the mechanism of this condition, helping to treat patients with TAR and improve wider understanding of how megakaryocytes & platelets develop and function.

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

Modeling cell differentiation dynamics from single cell genomics data 30 Sep 2018

We aim to understand in detail the dynamics of how white blood cells (specifically T helper cells) react to infections by multiplying rapidly and at the same time adapting their cell state to fight the infection. In particular, we focus on a mouse model of malaria where T helper cells differentiate into two subtypes: Th1 and Tfh. By quantitatively profiling the T helper cell population at different time points during a malaria infection, we expect to improve our understanding of the mechanisms which are responsible for the cell proliferation and specialisation. We study the cell population by detecting RNA expression, surface markers and cell divisions at the single cell level. The RNA expression will provide clues as to genes which are driving this process, and we will test a subset of genes using CRISPR knock outs. In addition to a better knowledge of the immune system, we hope to develop new mathematical and computational methods that will be widely applicable to modeling cell proliferation and differentiation data in diverse biological contexts.

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

Principles of human development and germ cell program 28 Nov 2017

Specification of human primordial germ cells (hPGCs) occurs around gastrulation, a critical juncture when the specification of the primary somatic lineages also occurs. In combination with human preimplantation embryos, in vitro models and hPGCs from aborted fetuses, our objective is to elucidate the origin and properties of the early human germline. For the mechanism of the hPGC fate, we will use experimental models that simulate early human development. We aim to investigate how cells gain competence for germ cell fate, and then respond to combinatorial effects of the critical transcription factors, which induce hPGC specification. Altogether, this study will reveal the organisation of the very early human embryo, and mechanisms of hPGC and somatic outcomes, which is essential for advances in regenerative medicine. Following hPGC specification, epigenetic resetting of the early human germline leads to extensive erasure of DNA methylation and epimutations in response to the critical regulators of chromatin organisation and nuclear architecture towards the epigenetic ground state. Some conserved resistant loci ('escapees') evade reprogramming. We will explore if some escapees have been exapted to function as regulatory elements. If so, this may have a crucial influence on human development, including brain development and neuronal diseases.

Amount: £2,750,000
Funder: The Wellcome Trust
Recipient: University of Cambridge

Metabolic magnetic resonance imaging across the human heart 18 Oct 2017

During my fellowship, I proved the feasibility of measuring cardiac energetics in volunteers and patients using ultra-high field (7T) MRI scanners. The sensitivity and the separation of signals from different metabolites both improved significantly compared to standard research scanners. I recently secured £340k funding to fit a new phosphorus coil on the Oxford 7T scanner, which I am now testing in volunteers. Theory predicts that this coil will have several complementary technical advantages. These will enable mapping of cardiac energy metabolism across the whole heart, with sufficient spatial resolution to distinguish signals from healthy from diseased tissue. It will also enable quantification of cardiac energy metabolism with high precision to study single subjects rather than groups. I request funding to validate these new whole-heart methods, proving their value in three carefully-targeted groups of patients, via an extension of my fellowship. My goals are (A) to study patients in which the metabolic pattern is known by other means; (B) others where the metabolic pattern will reveal previously-inaccessible aspects of disease mechanism; and (C) to prove I can resolve metabolic changes in single patients. Success in each of these studies will give me the pilot data needed for competitive Senior Fellowship applications.

Amount: £584,259
Funder: The Wellcome Trust
Recipient: University of Cambridge

Investigating the mechanism of influenza virus translation 30 Sep 2018

When viruses infect a cell, they need to hijack host machinery to produce their own proteins from mRNA, in a process called translation. The host cell requires several factors for translation, including proteins called eukaryotic initiation factors (eIFs). EIF4F plays a central role in this process and is a complex made up of the proteins eIF4A, eIF4G, and eIF4E. Together, these proteins act along with other factors to recruit the cellular machinery required for translation to begin. Influenza can promote the translation of its own proteins whilst host protein synthesis is impaired. As viral and host mRNA are highly similar, influenza virus was thought to only use the same mechanism of translation as the host. However several findings, such as the fact that influenza can translate its proteins without eIF4E, suggest that this is not the case. My hypothesis is that influenza can employ a different mechanism of translation from the host. I will use several RNA/protein analysis approaches to identify the key components required for influenza translation, and attempt to dissect the mechanism(s) of translation used by influenza. Identifying key differences between host and viral processes is important for identifying novel therapeutic targets.

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

INVESTIGATING THE ROLE OF DDX17 IN ANTIVIRAL INNATE IMMUNE SIGNALLING DURING VIRAL INFECTION AND ITS IMPACT ON VIRUS SPREAD AND REPLICATION 31 May 2018

The host DEAD box RNA helicases are master regulators of pathogen RNA and DNA sensing dependent IRF3 signalling and are crucial for host survival and infection outcome in response to a multitude of both viral and bacterial pathogens. Here we report the DEAD box RNA helicase DDX17 as a novel pathogen recognition receptor essential for IRF3 driven IFNbeta expression in response to immunostimulatory DNA and dsRNA. Our current data maps DDX17 to act independent to the canonical IRF3 signalling cascade at the level of gene transcription, independent of IRF3 phosphorylation. We hypothesise that DDX17 may regulate beta-catenin nuclear shuttling, an essential IRF3 transcriptional cofactor. We aim to investigate the impact of DDX17 on beta-catenin activation and phosphorylation status as well as subcellular localisation following stimulation of WT or KO MEFs. Furthermore, we aim to investigate the biological relevance in viral infection following vaccinia virus and herpes simplex virus type 1 infection of WT and KO MEFs through quantification of viral replication and spread as well as IRF3 pathway activation. This project will contribute to understanding the role of DDX17 in innate immunity and host-pathogen interaction with implications in the immunological understanding of viral infection. Key words: DDX17, IRF3, IFNbeta, helicase

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

Epigenetic control of neurodevelopmental gene regulatory networks linked to neurodegeneration 30 Sep 2018

Dementia is predicted to affect 130 million people worldwide by 2050 according to the World Alzheimer 2015 Report30. Some familial forms of dementia inherited in autosomal-dominant fashion are linked to mutations altering gene dosage2,8,14,16,19,23,25. Patients with the mutations display a long pre-symptomatic phase during which cellular changes may take place before the onset of the disease decades later23. The cellular changes are reflected in gene regulatory networks3,4,5,12,28,31. As evidence from other neurodevelopmental conditions suggests10,13,17, changes during early neural development may lead to onset of the disease decades later. In order to study the neurodevelopmental gene regulatory networks and their links to dementia, I would like to focus on two forms of their regulation: small RNAs and demethylation escapees. Demethylation escapees are regions of the genome that escape epigenome resetting during early embryonic development27. Small RNAs have an important role in neural development and gene regulatory networks controlling them1,6,20,24. In order to address the question, I will use RNA and whole genome bisulfite sequencing methods of neurons derived from human stem-cells from familial dementia patients, combined with bioinformatics analyses. Focusing on small RNAs and demethylation escapees, the project might hint at neurodevelopmental gene regulatory pathways dysregulated in autosomal familial dementia.

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

Investigating stiffness alterations in spinal cord injury 30 Sep 2018

Spinal cord injury is a devastating condition that may lead to loss of limb movement, sensation and bladder control. Despite intense research, treatment is still very limited. Most research to date has focused on biochemical signalling. However, some more recent studies have hinted that mechanics might play an important role in spinal cord regeneration. Using atomic force microscopy (AFM), a cutting-edge technique which allows us to very precisely measure stiffness maps of biological tissues, we will investigate the stiffness of spinal cord tissue at various time points after injury and compare this to the stiffness of healthy spinal cord. We will test whether artificially modifying the stiffness of the damaged spinal cord or modifying mechanosensing in spinal cord cells improves regeneration of neurons after spinal cord injury. Our studies will be carried out in a cervical contusion model in rats which closely mimics the pathology seen in the human spinal cord after injury, even though the behavioural impairments the animals show are markedly less grave.

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

Regulation of Neural Stem Cells 30 Sep 2018

Of all the tissues and organs in the human body the nervous system is the most intricate and complex, consisting of more than 100 billion neurons. These neurons make precise connections with each other to form functional networks that can transmit information at amazing speed over considerable distances. Neurons are produced by neural stem cells, which renew themselves at each cell division while also giving rise to all of the diverse types of neurons in the brain. The Brand lab is interested in how the environment influences stem cell behaviour, in particular how nutrition regulates neural stem cell proliferation. Uncovering the molecular mechanisms that control whether a stem cell chooses to proliferate or remain dormant is crucial for understanding tissue regeneration under normal and pathological conditions and in response to ageing. It is critical to learn not only how stem cell proliferation is induced but also how stem cells can return to a dormant (‘quiescent’) state, as uncontrolled stem cell division can lead to cancer, including brain tumours like glioma. A thorough appreciation of the signals, both extrinsic and intrinsic, that control stem cell behaviour is necessary to understand how homeostasis is achieved and maintained in the brain.

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

What is the functional significance of the interaction between Eros and OS9? 31 May 2018

Eros is a recently described endoplasmic-reticulum transmembrane protein that controls the phagocyte respiratory burst. Eros is essential for the generation of reactive oxygen species because it is necessary for protein (but not mRNA) expression of the gp91phox-p22phox heterodimer, which is almost absent in Eros-deficient mice. Consequently, Eros-/- animals succumb quickly following infection with Salmonella or Listeria. Eros is highly evolutionarily conserved and has a human orthologue C17ORF62 , which exhibits approximately 90% sequence similarity. Dr Thomas's group have preliminary data that the role of Eros is fully conserved in humans. However, the exact mechanism by which Eros controls gp91phox-p22phox abundance remains unclear. Using a yeast 2 hybrid screen Dr Thomas has shown that Eros's most significant interaction partner was OS9, an ER-resident lectin that regulates the degradation of misfolded transmembrane glycoproteins. Given that gp91phox is a transmembrane glcyoprotein, it is possible that Eros regulates gp91phox through a mechanism that involves OS9. While OS9 is highly expressed in the immune system, its role in the respiratory burst has not been studied. Using OS9 knockout mice and lentiviral over-expression systems, I will determine whether it regulates expression of gp91phox-p22phox or indeed, Eros itself.

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

Analysis of Inflammatory markers in chronic subdural haematoma 31 May 2018

This project plans to measure levels of tissue plasminogen activator (tPA) which is involved in fibrinolysis of blood clots within the CSDH lesions. This bleeding is an essential part of CSDH formation, followed by coagulation and fibrinolysis which is triggered by the cleavage of plasminogen by tPA to generate plasmin. tPA will be measured in these samples using the commercially available ELISA kit. I will determine whether levels of tPA are correlated with levels of other inflammatory markers in CSDH fluid or in blood, and also to examine if increased tPA levels at the site of the haematoma predicts risk of CSDH recurrence. If tPA concentrations in blood or CSDH fluid correlate with clinical outcome, this could be used clinically to decide whether surgical or pharmacological management are most appropriate for individual patients. Finally, the effects of dexamethasone treatment on levels of tPA and other cytokines will also be determined, by comparison of dexamthasone and placebo-treated patients. These patient samples are anonymised and will only be unblinded after measurement of the above analytes has been completed.

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

The role of Factor Inhibiting HIF (FIH) in T lymphocyte fate determination. 30 Sep 2018

Oxygen is essential for almost all animal life on the planet, acting as a key player in the energy production in cells. Cells are able to adapt to low oxygen environments by activating a factor named hypoxia-inducible factor (HIF), which shifts the metabolism of cells away from oxygen-consuming processes while increasing alternative energy-producing pathways. In T cells, an important type of immune cell involved in the body’s defence against infections and tumours, this metabolic shift alters cell function, resulting in a more aggressive immune response. Understanding how this process is regulated may allow us to target and harness immune cells to treat a variety of diseases, such as autoimmune disease or cancer. Our group and others have shown that boosting HIF levels in T cells makes them more effective in clearing tumours and resolving viral infections. We have also studied the effect of factor inhibiting HIF 1-alpha (FIH), a protein that blocks HIF activity. Interestingly, FIH itself also appears to alter the metabolism of cells, although its effect on immune cells is currently unknown. During my PhD I will assess the role of FIH in mouse T cells, focussing on how FIH-driven metabolic changes can augment immune responses.

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