Projects funded by the National Network but that sit outside of individual clusters whilst contributing to a number of our key goals
The Network is also supporting exciting research projects that are of interest across the Clusters. These have the potential to help establish future Clusters and support technology development that will benefit the entire scientific community.
Improving experimental reproducibility is a major goal in mouse genetics research. It is well established that microbiomes vary between mouse facilities and in consequence the microbiome represents a major source of uncontrolled variation, with the potential to significantly impact key experimental outcomes.
A highly collaborative network like the National Mouse Genetics Network provides the perfect opportunity to systematically assess the extent of laboratory mouse microbiome variation across UK. We propose to do this by non-invasively sampling faecal microbiomes from C57BL/6 breeding colonies at participating facilities.
The information gathered will ultimately allow us to predict the extent to which gut microbiome variation leads to functional changes in the microbiome that may directly impact different disease models being studied across the Network. Data generated will also contribute to efforts to integrate data resources across the Network, allowing researchers to better consider the effect of the microbiome on their own experiments.
Excess cholesterol is a major contributor to many human disorders, especially cardiac and neurodegenerative diseases, yet relatively few studies examine cholesterol’s behaviour in biological membranes in vivo.
Recently, we identified an unexpected link between mitochondria and cholesterol homeostasis in a genetic form of mitochondrial disorder. Taking a probe that binds cholesterol in the membrane of culture cells (D4-mKate) we adapted it for expression in fly, and found that the reporter is well tolerated in the animal. Strikingly, the probe revealed cholesterol aggregates in the disease model that are absent from controls. As well as yielding new readouts of disease mechanisms, the new model will be used to screen for treatments. With the project funded by the Netwrork, we aim to create an equivalent mouse model for the study of the full panoply of diseases characterized by aberrant cholesterol metabolism.
Leads: Antonella Spinazzola (UCL) and Ian Holt (UCL)
Although the mouse remains the major in vivo model system used in both fundamental and applied cancer research, there are major differences between mice and man in the metabolism of drugs, and these differences are a significant contributing factor why data obtained in mice does not extrapolate to the clinic. To circumvent these issues, particularly in drug development, we have created a unique mouse model (8HUM) which has been extensively humanised for the major genes involved in drug metabolism in man (Henderson et al 2019, Drug Metab Dispos. 47(6):601), to improve the translation of efficacy and safety assessments and to test and optimise therapies involving drug combinations. In this project we will use this 8HUM syngeneic model in proof-of-concept studies using clinically relevant mouse models of colon and melanoma cancers, available through the Cancer Cluster, to establish their utility in anti-cancer drug development.
Specific aims of this project will allow us to:
Collaborators: Karen Blyth, Kevin Read, Roland Wolf and the Mary Lyon Centre
Although mutations and variation that contribute to human disease can be modelled in mouse, as we learn more about disease mechanisms, it is becoming increasingly clear that simple genetically altered mouse models may be of limited use for investigating human pathologies.
We know that genetic variation and mutation that is associated with disease risk often lies within regions of the genome which do not encode proteins. Yet there are limited technologies which allow these non-coding variations to be modelled in vivo.
Similarly, we are learning that the structure of genes plays an important role in their regulation and are identifying human diseases in which structural changes play an important role. Understanding gene regulation at scale necessitates large-scale gene engineering, technologies for which are frequently not routinely available within core facilities.
Establishing simple protocols to allow large regions of the mouse genome to be manipulated would unlock the potential of the mouse for modelling and interrogating all human genetic variation.
Working mainly in mouse embryonic stem cells, we will test, compare and optimize technologies for large scale engineering of genetic loci. Technologies which will be investigated include
In addition, under the remit of horizon scanning, we aim to implement and test newly described technologies to establish whether they provide a robust and reproducible methodology for disease modelling.
Once established and optimized, we hope to deliver a dedicated tool kit of selection cassettes, vector backbones and defined genetic strategies to help disseminate the streamlined protocols to the community.
Working with the network’s research theme clusters, we hope to develop proof-of-concept models which use the developed technology to model complex human disease in the mouse.
The Degron Tagging Cluster is developing tools to evaluate different approaches to degrade target proteins in human disease models. In most cases, these approaches make use of small molecules that induce physical proximity between tagged target proteins and E3 ubiquitin ligases, which results in ubiquitination of target proteins followed by rapid degradation via the proteasome. While this strategy is proving effective for many proteins of interest, there are specific protein classes for which targeted degradation is likely to require a different approach. For example, proteins that localise within mitochondria, or on the cell surface, are not exposed to E3 ubiquitin ligase activities that can be easily recruited to promote turnover. Other proteins are already rapidly turned over by the ubiquitin proteasome system, and so induced proximity with E3 ligases might do little to accelerate this process.
Using this award from the NMGN Director’s Fund, we will investigate an alternative approach for inducible degradation of these protein classes that works via a fundamentally different mechanism. The Small Molecule Assisted Shut-off system (SMASh) makes use of tags encoding an unstructured protease from the Hepatitis C virus together with a destabilising hydrophobic domain (Chung et al, Nature Chem. Biol. 2015). Immediately following translation, the protease cleaves the tag from the nascent protein-of-interest (POI) leaving a short peptide scar of 6 amino acids which typically does not perturb normal protein function. Cleavage can be inhibited using clinically approved protease inhibitors, resulting in retention of the destabilising tag and turnover following induction. We will evaluate the utility of the SMASh system against other approaches for inducible protein degradation, focusing on challenging proteins-of-interest to our partners in the NMGN disease clusters.
This multi-institute collaborative approach aims to:
The UK PDX biobank is led by Dr. Sara Wells (Mary Lyon Centre at MRC Harwell), Prof. Dominique Bonnet (The Francis Crick Institute) and Dr. William Grey (University of York). Projects undertaken will range from studying multiple blood cancers and testing drug efficacy but also provide humanised PDX to study and test potential immunotherapies responses to solid tumours and provide a deep phenotypic analysis of murine xenograft behaviour.
The mouse AA467197 gene (orthologue of the human C15orf48 gene), is highly conserved throughout evolution, but despite this the precise function of the gene is not understood. Our group and others have recently shown that this gene encodes a small mitochondrial protein that localises to complex IV of the electron transport chain (the enzyme cytochrome c oxidase). The protein shows tissue- and condition-specific expression, is strongly induced during inflammation and infection, and is dysregulated in various types of cancer.
In collaboration with the Network’s Mitochondrial Cluster and experts at the Mary Lyon Centre, this project aims to understand the role of the AA467197 gene using a novel knock-out mouse line. Detailed phenotyping of mice under resting and immune-challenged conditions will give important insight into the function of this poorly studied gene during physiological development and the immune response.
Figure: The human orthologue of AA467197 (C15orf48) is expressed in regions of inflammation in the joints of rheumatoid arthritis patients. Brown = macrophage marker CD68; Pink = C15orf48 transcript; Purple = nuclei. (Clayton et al. 2021 Sci. Adv. 7,eabl5182)
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