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Recipients:
Broadfield Primary School
University of Cambridge
Amounts:
£0 - £500
£500 - £1,000
Award Year:
2017

Results

The role of aberrant RNA processing in the pathogenesis of Multiple Myeloma. 30 Sep 2017

A role for RNA binding and processing proteins in the control of eukaryotic cellular processes and in disease, including cancer, is emerging . I led the initial sequencing of the myeloma genome at the Broad Institute of MIT and Harvard. A key finding was mutations in RNA processing genes, DIS3 or FAM46C in 25% of cases. These findings have been independently corroborated, establishing these mutations as genuine drivers of the disease. DIS3 is the catalytic component of the exosome, an essenti al RNA processing complex. FAM46C is poorly characterized, but available evidence suggests it has roles in RNA processing in a lineage-dependent manner. This proposal seeks to better characterize these genes and mutations. Characterization of FAM46C mutations will be performed by knock-out of the gene from the DT40 cell line, determination of phenotype and rescue experiments. Lineage-dependent transcriptional pathways affected by altered transcript stability will be identified by RNA sequenci ng and confirmed in primary myeloma samples. Known aberrant RNA processing phenotypes associated with DIS3 loss/mutation will be sought by RNA sequencing in primary myeloma samples. The pathways affected by mutant DIS3 in myeloma will be identified using yeast genetic screens and classical yeast complementation experiments.

Amount: £250,346
Funder: The Wellcome Trust
Recipient: University of Cambridge

Mathematical genomics and medicine 30 Sep 2017

Not available

Amount: £2,490,000
Funder: The Wellcome Trust
Recipient: University of Cambridge

Metabolic and Cardiovascular Disease. 30 Sep 2017

Not available

Amount: £2,352,000
Funder: The Wellcome Trust
Recipient: University of Cambridge

Novel regulators of the HUSH epigenetic repressor complex 31 Jan 2017

<p>Epigenetic regulation of chromatin structure is fundamental to eukaryotic gene expression. The critical role of histone modifications in regulating chromatin has been established in genetic screens in <em>Drosophila. </em>Less information is available in more diverse organisms due, at least in part, to their genetic intractability. Using a gene-trap mutagenesis screen, the Lehner group recently discovered a novel gene silencing complex, the Human Silencing Hub (HUSH), which acts as a repressive H3K9me3 reader-writer complex through recruitment of the SETDB1 methyltransferase. However, regulation of the HUSH complex remains unexplored, and no associated demethylase has been identified.</p> <p>Here, I propose two complementary approaches to identifying novel regulatory components of the HUSH pathway: (A) A forward genetic approach, which will use a genome-wide CRISPR activation library. In a HUSH responsive GFP<sup>dim</sup> reporter cell line, activation of a HUSH regulatory gene will reverse reporter gene silencing. Following enrichment for rare cells reverting to a GFP<sup>bright</sup> reporter phenotype, candidate regulatory genes can be identified by next-generation sequencing. (B) A proteomic approach will use proximity-dependent biotinylation to mark HUSH interaction partners for identification by mass spectrometry. In both approaches, candidate genes will be validated and characterised biochemically to elucidate their mechanistic role in the regulation of heterochromatin.<em></em></p>

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

Analysing the functions of human cytomegalovirus US28 during latent infection 31 Jan 2017

<p>Human cytomegalovirus&nbsp;(HCMV) is a betaherpesvirus which causes lifelong subclinical infection in healthy adults, but significant mortality and morbidity in neonates and immunocompromised individuals. Like all herpesviruses, it establishes latent infection; HCMV undergoes latency in early myeloid lineage cells. Reactivation occurs when these cells differentiate into macrophages and dendritic cells, and reactivation events in immunocompromised individuals often fail to be controlled by the host immune system causing life-threatening disease. The HCMV genome encodes a G protein coupled receptor, US28, which is expressed during both lytic and latent infection, and is essential for establishing latency. I wish to understand how US28 functions in early myeloid lineage cells to establish latency. I will be using existing proteomic analyses as a basis for investigating a number of host proteins as potential mediators and effectors of latency; a number of host proteins are modulated by US28, some of which may be implicated in immune evasion by the virus. I also aim to understand the role of G proteins in establishing latency, given the varying functions of US28 in lytic and latent infection.&nbsp;I hope to establish a picture of the mechanisms by which US28 manipulates the host cell to facilitate latent infection.</p>

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

Regulation of the cellular trafficking of TAPBPR and its implication on MHC class I peptide presentation 31 Jan 2017

<p>The process of antigen presentation by major histocompatibility complex (MHC) class I molecules is a crucial process in protecting the human body against intracellular infections and cancer. These molecules present the cellular proteome, in form of short peptides, at the cell surface, for the inspection by cytotoxic T lymphocytes. The MHC class I molecules acquire their peptides in the ER, a process facilitated by the peptide loading complex (PLC). As part of the PLC, tapasin was until recently believed to be the only MHC class I specific chaperone directly involved in peptide selection. However, that has changed with the Boyle laboratory's discovery that a protein called TAPBPR is a second MHC class I dedicated chaperone, intimately involved in peptide selection. It is now essential to understand the distinct roles of the two MHC class I dedicated chaperones and how they work together in determining the final peptide repertoire displayed to the immune system.&nbsp;My key research goals are to investigate how the cellular trafficking of TAPBPR influences its effects on class I antigen presentation and what are the cellular factors that control its trafficking.</p>

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

The molecular basis of KAP1-dependent transcriptional silencing 31 Jan 2017

<p>The transcriptional regulator KRAB-associated protein 1 (KAP1) is crucial for preserving genome integrity by repressing potentially harmful retroelements. Following its recruitment to retrotransposons by KRAB domain containing zinc finger proteins (KRAB-ZFPs), KAP1 coordinates the assembly of a repressor complex containing the histone methyltransferase SETDB1 and histone deacetylase 1 (HDAC1) to induce epigenetic silencing of these elements.</p> <p>In addition to its role in repressing transposons, KAP1 also regulates the expression levels of numerous other genes. Interestingly, the transcriptional activities of KAP1 can be strongly influenced by posttranslational modifications (PTMs). Most notably, phosphorylation at S473 and phosphorylation at S824 both significantly alleviate the repression mediated by KAP1. The molecular mechanisms underlying these processes, however, remain poorly understood.</p> <p>To elucidate the structural basis of KAP1-dependent transcriptional repression, we will reconstitute a complex consisting of KAP1, SETDB1, the KRAB-ZFP ZNF93 and a DNA fragment containing the recognition sequence for ZNF93. Subsequently, we use single-particle cryo-EM to determine the structure of this complex.<br /> In addition, we will investigate the mechanism by which PTMs regulate the transcriptional activities of KAP1. Our central objective in this context will be determining how phosphorylation at S473 or S824 results in transcriptional activation.&nbsp;</p>

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

Functional proteomic analysis of novel antiviral restriction factors in primary leukocytes 31 Jan 2017

<p>This project aims to identify and characterise novel antiviral restriction factors (ARFs) that play key roles in preventing infection of primary leukocytes. ARFs may function by preventing viral entry or exit at the cell surface, or replication at various intracellular stages.&nbsp; I will focus on the subset of plasma membrane (PM) ARFs, which will be identified by two properties: interferon (IFN) induction and virally-induced downregulation.&nbsp; For this I will employ tandem mass tag-based MS3 mass spectrometry, enabling quantitation of PM proteins in primary leukocytes.</p> <p>&nbsp;</p> <p>Key Goals:</p> <p><strong>1.&nbsp;&nbsp;&nbsp;&nbsp; </strong><strong>Use IFNs and infection with two important human pathogens, human cytomegalovirus and HIV as a functional screen to identify novel cell surface ARFs</strong></p> <p><strong>2.&nbsp;&nbsp;&nbsp;&nbsp; </strong><strong>Investigate how these ARFs inhibit viral infection, and how are they targeted for destruction by viruses.</strong></p> <p>&nbsp;</p> <p>The use of IFN as part of the functional screen will additionally enable exploration of the difference in effects between IFN&alpha;, &beta;&nbsp;and &lambda; at the PM, a subject which is currently surprisingly poorly understood.&nbsp; This will provide important insights into human immunity in its own right.&nbsp; Understanding how viruses interacts with and targets ARFs for destruction will have important implications for therapy.</p>

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

Understanding the Pathogenesis of Inflammatory Bowel Disease via Whole-genome Sequencing 31 Jan 2017

<p>We will use a new whole-genome deep-coverage IBD dataset (15x+ coverage, 20 000 cases, 50 000 controls) to conduct genetic association studies. Several analyses are currently planned.</p> <p>&nbsp;</p> <p>The first study will use the data from &gt;1000 IBD patients, who are part of a deep clinical phenotyping experiment, on their response to treatment with anti-TNF medication. We are hoping to determine specific genetic variants associated with successful treatment, non-response, loss of response, and unfavourable drug reactions.</p> <p>&nbsp;</p> <p>Once more samples are sequenced, we will attempt to discover novel low-frequency, rare, and very rare genetic variants associated with IBD. A recent low-coverage sequencing study has identified a rare missense variant in ADCY7 that doubles the risk of ulcerative colitis. In addition, a burden of very rare, damaging missense variants in genes associated with Crohn's disease was detected. The increased coverage and the size of the dataset may confirm the significance of such variants. Discovery of novel rare variants brings important insights into IBD biology, and improves the overall understanding of the genetic landscape of complex diseases.</p>

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

Integrating genomic, transcriptomic, metabolomic and behavioural data from 12 strains of C. elegans to understand gene/environment interactions under different dietary regimens 31 Jan 2017

<p><em>C. elegans</em> can grow on a range of different bacterial diets. It has already been shown that it changes its behaviour depending on the available food. Its lifespan also depends on its diet. Autophagy has been shown to mediate the increase in lifespan when the nematode is grown on certain foods. However, the mechanisms by which the environment leads to a change in beheviour and life history traits remain largely unknown. With this project, we would like to use high-throughput sequencing and metabolomics to build a quantitative and comprehensive map of the underlying molecular networks activated in a specific dietary environment. Furthermore, we would like to harness the genetic diversity of <em>C. elegans</em> to study the genetic basis of its phenotype as well as the interaction of its genome and environment in the determination of its behaviour and lifespan. For this purpose, we plan to extend the latest statistical techniques to integrate all layers of genomic and phenotypic data. Many of the genes involved in metabolism are conserved between humans and nematodes. Therefore, we expect that the findings will be relevant to human physiology.</p>

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

Systematic Identification of Lineage Specification in Murine Gastrulation 31 Jan 2017

<p>Single-cell genomics is a fantastic tool for studying developmental biology: it allows unbiased and large-scale study of gene expression at the correct resolution for cell fate decision making. New fluidics systems provide the capability to study tens of thousands of cells simultaneously - as many as there are in the young embryo.</p> <p>For my PhD, I will analyse scRNA-seq data generated on this platform, studying mouse gastrulation between E6.5 and E8. I will be able to study this process at both an exceptional cell-level resolution (thanks to the fluidics) and at an unprecedented time resolution, at 0.1 day intervals.</p> <p>My focus will be on identification of lineage specification, and how cells make their fate choices. I will need to develop new methods to account for the large numbers of cells assayed, the numerous lineage decisions made, and heterogeneity of speeds of development across and between embryos. I hope to produce a map of lineage specification from epiblast (E6.5) cells through to every cell type present at E8. This work will provide a developmental atlas through gastrulation, and general inferences on cell fate decisions may provide insight for cellular reprogramming and regenerative medicine.&nbsp;</p>

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

New approaches to big data in genome sequencing 31 Jan 2017

<p>The ever increasing throughput and reducing cost of DNA sequencing continue to challenge computational data analysis associated with genome sequencing, requiring novel algorithmic and software solutions that take advantage of structure in the data.&nbsp; We will develop new efficient computational solutions to handle and analyse very large genome sequence data sets.&nbsp; Specifically, we plan to develop software to address the following topics, in the context of the human Haplotype Reference Consortium with tens of thousands of whole human genome sequences, and also large non-human vertebrate reference genome sequencing projects.</p> <ol> <li>We will develop compressed data structures to store deep population resequencing data, while supporting fast genetic analyses that work directly from the compressed data. We will extend these methods to new reference structures based on variation graphs.</li> <li>We will develop new approaches to long range genome sequence assembly, using long reads to scaffold deep short read assemblies, and using linkage disequilibrium structure in population data to scaffold assemblies, incorporating methods from topic (1).</li> <li>We will use machine learning approaches to address statistical problems such as indel and structural variant calling that are hard to model exactly, using large simulated and real data sets and methods from (1) and (2).</li> </ol>

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

Machine Learning methods for microscopy image data in high-content screening 31 Jan 2017

<p>An important area in drug development is understanding low-level molecular processes and pathways that cause diseases. These cellular phenotypes are high-dimensional and are increasingly being captured using single-cell assays and high-content imaging. In understanding natural cell trait variation and engineered variants, we can elucidate the cellular consequences of disease mutations.</p> <p>&nbsp;</p> <p>In my project, I will exploit cellular images in a range of contexts to investigate the link between genetic variation and cell trait variability using both natural genetic variation and engineered variants. To do so, I will develop machine learning methods to extract features from high throughput microscopy data, and to accurately account for genetic, environmental, and experimental sources of variability in them. Furthermore, I will work on integrative approaches using public genomic data to bring in other omics modalities, thereby tackling key challenges in the larger aim of deciphering disease and fostering drug development.</p> <p><br />I will use existing data from the HipSci project, high throughput drug screens from AstraZeneca, and, in addition, will design and oversee the generation of datasets through high-throughput CRISPR knockouts as part of Leopold Parts&rsquo; group at the Wellcome Trust Sanger Institute and Oliver Stegle&rsquo;s group at the European Bioinformatics Institute.</p>

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

Dynamic interplay between the ageing CNS progenitor cells and their surrounding niche 31 Jan 2017

<p>The pathological loss of myelin sheath from axons predisposes neurons to atrophy and subsequent death. In response to brain demyelination, a cell population called oligodendrocyte progenitor cells (OPCs) proliferate and differentiate into oligodendrocytes (OLs), which remyelinate exposed axons. In ageing remyelination often fails due to a reduction in OPC regenerative capacity.&nbsp;The age-related reduction in remyelination coincides with changes in the brain biochemical and mechanical properties. Preliminary data have demonstrated that exposure of aged OPCs to young-like environments improves aged OPC function in terms of proliferation and differentiation into myelinating OLs. This project aims to identify factors from the physical extracellular microenvironment, which negatively influence remyelination in the ageing CNS. In order to elucidate the underlying molecular mechanisms, the composition of the brain tissue changes with age on a biochemical and mechanical level and and its influence on the OPC proliferation and differentiation into OLs will be investigated. Moreover, the negative effects of the ageing microenvironment will be mitigated by mimicking the young brain niche using synthetic scaffolds as well as modified decellularised tissue. Understanding this interplay using a range of <em>in vitro</em> and <em>in vivo</em> methods will therefore aid the development of therapies for enhancing endogenous remyelination.</p>

Amount: £62,117
Funder: The Wellcome Trust
Recipient: University of Cambridge

The role and regulation of UHRF1 during epigenetic reprogramming 31 Jan 2017

<p>During both mouse and human embryonic development there are two waves of global DNA demethylation associated with an increase in developmental potential: firstly during the development of the inner cell mass from the gametes, and secondly during the development of primordial germ cells. Although some of these changes in DNA methylation are correlated with gene expression, it is not understood why this epigenetic reprogramming is consistently so extensive. Until recently it was believed that most of this erasure of DNA methylation occurred actively. However, recent research has revealed that regulation of maintenance methylation accounts for the differing rates of demethylation in different reprogramming contexts. Across the majority of these different contexts, the activity of maintenance methylation is controlled by regulation of UHRF1 protein. This project aims to elucidate the mechanism by which UHRF1 protein is regulated during epigenetic reprogramming, then to use this knowledge to assess the importance of this regulation to the acquisition of pluripotency during mammalian embryonic development. This will improve our understanding of developmental and reproductive health and may even shed light on new or improved methods of reprogramming cells for therapeutic applications.</p>

Amount: £26,820
Funder: The Wellcome Trust
Recipient: University of Cambridge

Investigating the Impact of Extrinsic Factors on Intestinal Stem Cell Dynamics 31 Jan 2017

<p>The rapid turnover of the mammalian intestinal epithelium is fuelled by division of stem cells residing at the bottom of the crypts of Lieberk&uuml;hn. Our understanding of how stem cells populate the intestine in humans lags behind that of the murine system. In the latter, stem cells populate the crypts by a process of neutral drift. Moreover the number of functional stem cells per crypt and their replacement rate has been quantified in the mouse intestinal epithelium. However, it is not known how extrinsic factors, such as regular drug use, influence these dynamics. In humans, randomised clinical trials have linked low-dose aspirin to a reduction in colorectal cancer incidence and mortality. However, the mechanism is unclear. Since tumours originate from stem cells, I plan to conduct an in vivo study to investigate the impact of aspirin on murine intestinal stem cell dynamics.</p> <p>In the human system, functional stem cell numbers and replacement rates are still not well characterised. Somatic mutations can be used as clonal marks to investigate these parameters in patient samples. I thus aim to find and validate novel human somatic clonal marks and collect data for mathematical modelling of human intestinal stem cell dynamics.</p>

Amount: £46,765
Funder: The Wellcome Trust
Recipient: University of Cambridge

Development and leptin signalling in human stem cell-derived POMC neurons 31 Jan 2017

<p>Energy homeostasis in mammals is tightly controlled by a distinct neural circuit in the hypothalamus and its dysfunction leads to obesity. Pro-opiomelanocortin (POMC) neurons are a central component of this circuit. While obesity research has been largely limited to studies in rodents in the past decades, recently published protocols now allow the<em> in vitro</em> generation of human POMC neurons from human pluripotent stem cells (hPSCs). This enables us to study disease-associated mechanisms directly in the human cell type relevant to obesity. In my PhD I will contribute to a thorough characterisation and optimisation of this <em>in vitro</em> system. I will study when human POMC neurons are born in culture, assess their responsiveness to metabolic cues that regulate their activity and test different means of enhancing this responsiveness. This <em>in vitro</em> characterisation of POMC neuronal activity will be complemented by transplantation studies, in which I will test whether hPSC-derived POMC neurons possess all the features of functional maturity required to reinstall energy homeostasis in obese mice. I will finally use this novel tool to study the molecular mechanisms of certain signalling pathways in human POMC neurons, which might lead to the identification of potential targets for therapeutic intervention in human obesity.</p>

Amount: £46,168
Funder: The Wellcome Trust
Recipient: University of Cambridge

Quantitative visualization of polarity protein complexes in Drosophila and mouse polarized epithelia 31 Jan 2017

<p>Epithelial cells have to establish and maintain apicobasal polarity to perform their vectorial secretory and absorptive functions. At the cellular level, polarity is defined by apical, lateral and basal domains, each marked by distinct polarity proteins localized at the cell cortex and in the cytoplasm. Based on biochemical and genetic evidence from C. elegans, D. melanogaster and cultured mammalian cells, these proteins are proposed to associate with each other and form spatially-restricted, mutually-antagonistic complexes. This model has proven difficult to test, since some proteins are thought to be involved in multiple complexes. An overarching goal for the field is to untangle the complicated model for polarity determination. I propose to contribute in the following two ways: firstly, to provide a direct evidence of polarity protein interactions and their stoichiometry within distinct complexes in Drosophila and mouse polarized epithelial cells using nanoscopy, and secondly, to investigate the molecular regulation of polarity orientation in intestinal organoids.</p>

Amount: £21,342
Funder: The Wellcome Trust
Recipient: University of Cambridge

Activity and specificity of Notch regulated enhancers 31 Jan 2017

<p>Signalling pathways are required iteratively during development, coordinating many different developmental processes.&nbsp; This diversity relies on them eliciting different qualitative and quantitative transcriptional responses depending on the context.&nbsp; The Notch pathway is one of the small set of core pathways and can promote distinct outcomes through different sets of target genes. How Notch responsive enhancers achieve this level of regulation is not understood. In this project, well characterized Notch responsive enhancers will be used to study how activity and specificity in response to Notch are encoded within the enhancer sequence and how different factors play a role in the control of these properties.</p> <p>1. Activity: Characterize the transcriptional output of a Notch responsive enhancer and analyze how it is governed.</p> <p>2. Specificity: Compare two enhancers that exhibit different response to Notch to determine how their response specificity is achieved.</p> <p>3. Sufficiency: Assess if a defined enhancer is sufficient to confer a specific Notch response when introduced into a heterologous locus.</p>

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

Microtubule organisation in developing neurons 31 Jan 2017

<p>Microtubule (MT) bundles found in neuronal axons are highly organised, as around 90% of MTs point with their growing end (commonly referred to as the +end) away from the cell body. This MT orientation is crucial for correct cargo localisation. Known for more than 30 years, the underlying mechanisms for axonal MT organisation are yet to be discovered. Preliminary experimental data from the Franze group suggest that correctly oriented (or &ldquo;+end out&rdquo;) MTs in axons are selectively stabilised whereas oppositely oriented ones are not. The stabilisation requires both axonal cortex integrity and cytoplasmic dynein function, and MT destabilisation leads to the breakdown of MT organisation. In this project, I will focus on the exploration of the mechanism(s) that lead to the stabilisation of &ldquo;correctly&rdquo; oriented MTs. Furthermore, I will investigate how MT organisation could be related to common pathologies.</p>

Amount: £14,678
Funder: The Wellcome Trust
Recipient: University of Cambridge