Research Opportunities

The scholar will work on a project integrating neuroimaging and genetics across the entire lifespan with the goal of gaining a more fine-grained understanding of the biological mechanisms driving brain morphological changes across the lifespan in health and disease.

Cell-to-cell communication by extracellular vesicles (EVs) is a growing field of investigation in basic cell biology research, biomarker discovery and therapeutic drug delivery. Our lab is investigating how different cargoes are loaded into EVs and the pathways that regulate EV biogenesis, release and uptake.  There are various chaperone proteins within the cell that aid the sorting of cargoes into EVs.  We are particularly interested in the aggregate-prone proteins that are associated with different neurodegenerative diseases (e.g. alpha-synuclein, SOD1, TDP-43, tau and huntingtin) and have shown that these proteins can be loaded into EVs and secreted from cells. We have recently identified that members of the small heat shock protein (sHSP) family can interact with various aggregate-prone proteins to facilitate their loading into EVs and their intercellular spreading.  In particular, we have demonstrated that one of the sHSP family, HSPB1, can interact with the autophagy cargo receptor p62/SQSTM1 to modulate its unconventional secretion by EVs. In cells expressing mutant huntingtin (the aggregate-prone protein associated with Huntington’s disease), these HSPB1-loaded EVs are capable of inducing the spreading of mutant huntingtin to non-expressing cells. Importantly, these findings reveal a novel mechanism for the spreading and seeding of protein aggregates, which may have wider implications for and impact the pathobiological mechanisms underlying other neurodegenerative disorders. In addition, we have identified several signalling pathways and regulatory proteins that are essential for the formation of mutant huntingtin-carrying EVs. 

This project will use a range of cell-based and in vivo assays to investigate how such signalling proteins regulate the interplay between autophagy and unconventional secretion and determine how this affects the accumulation and spreading of neurodegenerative disease-causing proteins. The first part of the project will involve over-expression and knockdown of these signalling proteins in vitro (in cell-based assays), where a range of biochemical and microscopy techniques will be deployed to look at protein interactions, localisation and spreading of these proteins. These findings will be then validated in vivo using a combination of zebrafish fluorescent reporter lines and neurodegenerative disease models. Finally, by using genetic and pharmacological activation and inhibition of signalling pathways, we will monitor EVs in vivo and characterise how perturbation of unconventional secretion can impact the disease progression. 

Our progress in understanding and engineering living systems, and developing therapies, is severely limited by inability to build predictive, data driven models of cellular processes. Much of current cellular biology research, including work with human stem cells, microbes, and cell lines, proceeds by optically labelling cellular components such as proteins and by measuring and manipulating physiological signals optically. Microscope imaging is then used to track and quantify the interactions of these signals and components in living cells, including cells that have been genetically engineered or exposed to pharmacological agents. Quantities of interest, such as where proteins aggregate, or how rapidly cells grow are then extracted from images or movies and then quantified. This is challenging, slow and error prone because the experiments are often done piecemeal, often by hand, and focus on a handful of types of molecules or cellular interactions that are inferred from a condensed snapshot of the data, such as an average protein density.  

This project leverages recent advances in AI to analyse image data gathered from microbial populations (E coli). Our goal is to build predictive models of processes such as cell division and virus infection using high throughput microscope data. We approach this using a fusion of simulated and real data, with model-based predictions tested in automated, high throughput experiments. We wish to scale this up to cover other types of cells, including human stem cells and microbiota through collaboration with suitable groups at NIH.  This project would suit trainees with strong quantitative skills, a first degree in a STEM discipline and proficiency in coding in more than one language.

A major goal in systems neuroscience is to connect the activity of populations of neurons to specific behaviors. However, large scale recordings of neural activity during the execution of learned tasks and during the experience of familiar stimuli have revealed that neural activity patterns continually change over extended periods. This so-called Representational Drift is not accompanied by obvious alterations in behavior, learning or systemic physiology, which raises profound questions about its origin and its implications for learning and memory. For example, textbook theories of learning and memory assert that stable memories require stable relationships between neural activity and learned associations. Representational drift brings these theories into question, while raising practical problems for understanding neural data, designing experiments and developing technology such as brain-machine interfaces.  

This project uses a mix of data science, computational modelling and theory, and collaboration with experimentalists to understand the causes and implications of Representational Drift. We use a variety of statistical methods as well as modelling and analysis of artificial neural networks to generate and test hypotheses. We work closely with experimentalists in Harvard Medical School and UCL, and wish to find experimental partners in the NIH to further this research.  Key skills include proficiency in numerical methods, simulation, strong coding skills and a working knowledge of advanced statistical methods, including generalized linear models and Bayesian inference.  

Key recent publications include:  
Micou, C., & O'Leary, T. (2023). Representational drift as a window into neural and behavioural plasticity. Current opinion in neurobiology, 81, 102746. https://www.sciencedirect.com/science/article/pii/S0959438823000715  Rule, M. E., & O’Leary, T. (2022). 

Self-healing codes: How stable neural populations can track continually reconfiguring neural representations. Proceedings of the National Academy of Sciences, 119(7), e2106692119. https://www.pnas.org/doi/abs/10.1073/pnas.2106692119

The transmission of viruses between species faces significant barriers due to differences in host immune systems. A virus adapted to an animal host might not be well-equipped to evade the human immune system. However, mutations and other viral adaptations can occasionally overcome these barriers, leading to zoonotic infections. This concept is exemplified by the ongoing avian influenza pandemic which is now spread from birds to mammals, including livestock cow herds. Understanding and strengthening antiviral immunity is therefore crucial in preventing and controlling zoonotic diseases and for improving human and livestock health by, for example, driving next-generation vaccine development.   The molecular and cellular mechanisms by which human cells sense and respond to infection are well characterised and known to be essential for host defence against viruses. Despite their importance as sources of food, their economic importance, and as sources of zoonotic pathogens, for the majority of livestock species these innate immune systems are relatively poorly defined.  

In this project we the student will define the functions of PRRs that sense viral nucleic acids across multiple species, including sheep, cows, chickens, and ducks and compare them to humans. The project will employ loss of function assays, using CRISPR/Cas9, signalling and targeted perturb-seq experiments to understand the functions of these receptors. The project will also include a range of virus infection models, for example influenza viruses and poxviruses, to define how PRRs from these key livestock species impacts antiviral responses in the context of zoonotic infections.

We have established an approach to accelerate vaccine development, through our Plug-and-Protect platform. A limiting factor in vaccine generation is the difficulty of turning a promising target protein into the kind of assembly that would give long-lasting disease protection. We demonstrated potent immunization towards the global health challenge of malaria. This approach is now being used by many groups against cancer and various infectious diseases, e.g. HIV, influenza, coronaviruses and other outbreak pathogens. This project will involve creating new protein antigen and nanoparticle designs to achieve the most effective and broadly protective immune responses. By inducing potent mucosal immunity, the project will contribute to developing a new generation of vaccine systems, towards protection against the most challenging diseases.

The aberrant modification status of proteins is universally recognized as a crucial component of disease. In order to develop therapeutic agents to combat disease, we need to understand the role that posttranslational modifications (PTMs) play within pathological systems. Focusing on infectious diseases using mutant cell lines, mouse models and patient data, we will study the link between PTM status and subcellular location which has been so far poorly captured in the majority of experimental workflows. The knowledge of the PTM affecting relocalization of the protein and, in turn, its function, will be pivotal to the correct drug design. This project combines development of state of the art quantitative proteomics methodologies, computational workflows and whole cell modelling which will be used to decipher the mechanism of immunity to infection and propose new ways of treatment. 

Genetics only explain a small proportion of phenotypic variance, with common diseases typically having 10%-30% heritability (Loh et al. 2017 Nature Genetics). This project aims to explain the remaining 70%-90% of variance using molecular data. Past efforts have attributed genetic variance to expression data (Yao et al. 2020 Nature Genetics) and different tissues (Amariuta et al. 2023 Nature Genetics); yet limited attention is paid to the non-genetic variance.  We aim to develop methods to provide an unbiased estimate of the environment variance in complex traits that are mediated through molecular traits. Specifically, we are interested in the proportion of non-genetic variance that are mediated by gene expression, protein level, and metabolomics. We will utilize large-scale proteomic and metabolomic data that are linked to electronic health records to validate the model and provide the molecular explanation for common complexity traits.

Mitochondrial diseases are caused by defects in genes required for energy production and oxidative phosphorylation (OxPhos). We find it intriguing that some patients with mitochondrial disease present late in life, with very tissue-specific phenotypes. It seems that not all cells and tissues are equally susceptible to mitochondrial disease.  We mainly study how mitochondrial dysfunction and mutations in the mitochondrial genome affect neural stem cell behaviour in Drosophila and mouse. 

The questions we address are:  
(1)    how mitochondrial dysfunction affects normal and pathological cell fate decisions in the developing brain. We previously showed that neural stem cells in the brain rely heavily on mitochondrial energy production and now study how they interact with the glial cells that make up their stem cell niche. 
(2)    how transcription of the nuclear genome is regulated when a cell is confronted with mitochondrial dysfunction. We employ and develop innovative DamID-seq based in vivo chromatin profiling technology to study metabolism of chromatin modification.  
(3)    how mutations in the mitochondrial genome evolve over time, during brain development and aging. We use single-cell forward genetic CRISPR screening to identify novel regulators of mitochondrial genome maintenance.    

In order to study these questions in an in vivo context, in (stem) cells surrounded by their appropriate tissue environment, our primary model system is the fruit fly, Drosophila melanogaster. In addition, we actively translate our findings and the technology we develop into mammalian model systems, in particular the mouse embryonic cortex.   

Key References: 
Viscomi C, van den Ameele J, Meyer KC, Chinnery PF. Opportunities for mitochondrial disease gene therapy. Nat Rev Drug Discov. 2023 Jun;22(6):429-430. 

van den Ameele J, Krautz R, Cheetham SW, et al., Reduced chromatin accessibility correlates with resistance to Notch activation. Nat Commun. 2022;13(1):2210. 

van den Ameele J, Li AYZ, Ma H, Chinnery PF. Mitochondrial heteroplasmy beyond the oocyte bottleneck. Semin Cell Dev Biol. 2020 Jan. 97:156-66.  

van den Ameele J, Brand AH. Neural stem cell temporal patterning and brain tumour growth rely on oxidative phosphorylation. eLife. 2019;8:e47887. 

Adult neurogenesis, the process of generating new neurons in the adult brain, is a rare occurrence in mammals, confined mainly to the olfactory bulb and the hippocampus. Unlike the body's ability to repair most tissues, the limited scope of neurogenesis in the brain restricts our capacity to recover from neurological damage, a limitation that profoundly impacts the treatment of brain disorders.  In the olfactory system, ongoing neurogenesis supports the regeneration of key neuron types, including dopaminergic cells, granule cells, and olfactory sensory neurons, all of which are essential for sensory processing and adaptability. Recent studies have revealed that dopaminergic neurons born during embryonic development differ significantly from those generated postnatally, suggesting that they perform distinct functions rather than acting as simple replacements.  Our project aims to expand on these findings by exploring whether these differences extend to other regenerating neuron populations in the olfactory system. Using a combination of transgenic mouse models, in vivo calcium imaging, immunohistochemistry, electrophysiology, and behavioral testing, we will investigate the specific roles of embryonic versus adult-born dopaminergic neurons in olfactory processing.  By addressing these questions, our research will contribute to a deeper understanding of the unique contributions of adult neurogenesis to brain function, with the potential to inform new approaches for treating neurological disorders.

The ability to sense and respond to environmental cues is vital for the survival of all organisms. This process hinges on the integration of sensory information to generate appropriate behaviors, a capability rooted in neuronal plasticity. Neuronal plasticity encompasses structural, synaptic, and intrinsic modifications within neurons. However, these mechanisms are often studied in isolation, leaving their collective impact on behavior poorly understood.  Our lab aims to bridge this gap by exploring how mice adapt to olfactory stimuli. In this project, we will manipulate the olfactory environment of mice through sensory deprivation (akin to experiencing a mild cold) or olfactory enrichment (comparable to exposure to a perfume shop). Using advanced genetic tools, we will label neurons responsive to specific odors. Our approach integrates immunohistochemistry, in vivo calcium imaging, and patch-clamp electrophysiology to examine how olfactory bulb neurons adjust their synaptic connections, morphology, and intrinsic properties in response to varying durations of sensory alteration.  To link these cellular changes to behavior, we will employ automated behavioral testing to evaluate the mice's ability to detect and differentiate odors. This will allow us to assess how adaptive plasticity influences learning and behavioral flexibility. By combining cellular and behavioral analyses, this interdisciplinary project aims to uncover the complex neural mechanisms underlying behavioral adaptability in response to changing olfactory environments.

La Crosse virus (LACV), a member of the Bunyaviridae family, is a mosquito-borne pathogen that is leading cause of paediatric encephalitis in North America. Most cases of LACV infection are asymptomatic. However, the virus can cause severe disease when it infects the central nervous system: approximately 1% of LACV neuroinvasive disease cases are fatal, and it causes neurological sequelae including epilepsy and cognitive abnormalities in a further 6-15% of cases. LACV neuroinvasive disease is most common in children under the age of 16.

LACV is an enveloped virus with a tripartite, negative-sense RNA genome consisting of small (S), medium (M), and large (L) segments. The M segment encodes the glycoproteins Gn and Gc that are present on the surface of virus particles. These glycoproteins recognise receptors on the surface of suitable host cells and catalyse the membrane fusion that allows entry of the viral genome into the host cell, the essential first step of the infection cycle. The cell surface receptor(s) of LACV have not been characterised, limiting our understanding of which cells can be infected and of why children but not adults suffer severe LACV neuroinvasive disease.

We have performed an unbiased protein-based screen to identify human cell surface proteins that bind the surface glycoproteins of LACV. We identified that LACV binds to receptors of the Notch signalling pathway. This result is highly significant because Notch receptors are regulators of neuronal development, helping determine whether neural progenitor cells differentiate into neurons or glial cells. Biochemical and biophysical experiments using purified components have confirmed that LACV binds these receptors with high affinity, and we have a high-confidence structural prediction of the interaction between LACV and these cell surface receptors.

This project will define the molecular interactions between LACV and its cell surface receptors at atomic resolution, using a combination of structural biology, biochemical and biophysical techniques. We will exploit this information to generate structurally informed mutations that disrupt these interactions, allowing us to test the functional consequences of Notch receptor binding for LACV infection and neuropathogenesis. Students undertaking this project will gain experience of biochemistry, biophysics, structural biology and molecular virology, with the opportunity to perform virus infection experiments in a high-containment environment. They will also advance our knowledge of an important paediatric disease, potentially identifying new strategies to prevent LACV infection or minimise the subsequent neurological sequelae.

Generation of reactive oxygen species (ROS) by the phagocyte NADPH oxidase is a critical and highly conserved antimicrobial function of myeloid immune cells such as neutrophils and monocytes. ROS production must be tightly regulated to ensure constant readiness for immune defence, while restraining inappropriate activation. A lack of ROS from this complex results in the devastating inborn error of immunity chronic granulomatous disease (CGD), characterised by recurrent infection but also autoinflammation and autoimmunity. Common hypomorphic variation in the genes encoding components of the phagocyte NADPH oxidase also drives pre-disposition to common autoimmune diseases such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA). Excess ROS production can, however, result in oxidative stress. Understanding how ROS is tightly regulated is thus important for the development of rational therapeutics immune-mediated diseases. 

Despite the elucidation of the NADPH oxidase complex structure and function, upstream regulators of ROS production remain largely undiscovered due to a lack of robust biological model systems. The Thomas Lab characterised EROS (Essential for Reactive Oxygen Species) as an indispensable regulator of ROS generation but we believe that there are many more. Recent developments in CRISPR-Cas9 technology now allows both the introduction of precise edits (homology-directed repair, HDR) and genome-wide forward genetic screening by introducing knockout (CRISPRko) libraries. This may identify therapeutic targets in inflammatory disease.

We will use CRISPR-HDR methods to endogenously tag key components of the NADPH oxidase complex with fluorescent proteins to generate reporter lines for iterative selection by flow cytometry. By screening these at genome-wide scale with CRISPRko libraries and sorting cells based on component expression, followed by functional screens using fluorescent ROS probes, we will elucidate upstream regulators of the complex expression and function. The function of these novel regulators can then be investigated and validated using primary and immortalised cells, structural biology, and selective mutagenesis. Interrogation of publicly available genomic datasets will guide ‘hit’ selection and possible therapeutic relevance.

The placenta is the interface between mother and fetus, integrating signals between the two. The placenta releases factors such as proteins and miRNAs that can impact on maternal and fetal physiology. Some of these will be released from the placenta within extra-cellular vesicles (EVs), but how their content is modulated by an obese diabetic environment and how it impacts on maternal and offspring health is unknown. 

This project will explore how the placental secretome, including EV content, is modulated by obesity and diabetes during pregnancy and define how these changes have short and long-term consequences on maternal and offspring metabolism. The project will involve:
(1) profiling the placental secretome in healthy and obese diabetic pregnancies
(2) determining the protein and miRNA content of placental EVs isolated from lean and obese diabetic murine pregnancies
(3) establishing the effects of secreted placental proteins on maternal and offspring metabolism
(4) a combination of in vitro and in vivo experiments to establish the functional consequences of the changes in placental EV protein and miRNA content 

Lymphocytes respond to infection by rapidly increasing and decreasing the expression of many genes in a highly regulated manner. This regulation requires the integration of transcription, mRNA decay and translation. We are only just beginning to understand how these processes are integrated with each other. The host labs are studying how the multiprotein CCR4-NOT complex and its associated RNA binding proteins control gene expression. By combining structural and molecular biology approaches with cellular immunology and mouse models of immune responses we offer a broad training experience and the opportunity to discover fundamental mechanisms of gene regulation in the immune system. The discoveries have application in modifying the potency and durability of cellular immune therapies such as anti-tumour CART cell responses and the student will have an opportunity to apply fundamental knowledge to these applications.

References
The timing of differentiation and potency of CD8 effector function is set by RNA binding proteins. Nat Commun. 2022 doi: 10.1038/s41467-022-29979-x. PMID: 35477960; PMCID: PMC9046422.

Regulation of the multisubunit CCR4-NOT deadenylase in the initiation of mRNA degradation. Curr Opin Struct Biol. 2022 doi: 10.1016/j.sbi.2022.102460. Epub 2022 Sep 16. PMID: 36116370; PMCID: PMC9771892.

The nexus between RNA-binding proteins and their effectors. Nat Rev Genet. 2023. doi: 10.1038/s41576-022-00550-0. Epub 2022 Nov 23. PMID: 36418462; PMCID: PMC10714665.