Research Opportunities

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.

The Phosphoinositide 3-kinases (PI3Ks) are a family of lipid kinases that control diverse signalling pathways affecting gene-transcription, cellular adhesion and trafficking, autophagy and metabolism via the generation of PIP3. While some of these readouts are controlled by the evolutionarily conserved PI3K-AKT-FOXO, PI3K-AKT-mTOR axes, there is a diverse network of PI3K effectors that are less well studied, especially in lymphocytes, but which nonetheless can have profound effects on lymphocyte biology. We have recently used CRISPR/Cas9 to perform a targeted screen of PI3K effectors by generating a library that specifically targets PIP3-binding proteins. Screening for genes that affect T cell adhesion, we identified RASA3 as a key protein linking PI3K to the activation of the integrin LFA-1 and found that RASA3 is critical for T cell migration, homeostasis and responses to immunization (Johansen et al Sci Signaling 2022; Trend Immunol 2023; Front Immunol. 2021). We have now generated extended CRISPR/Cas9 libraries that target the entire PI3K-ome (including the kinases, phosphatases and all known effector proteins). Potential projects include designing and implementing new screens for downstream readouts of PI function, including autophagy, endocytosis, regulation of humoral immunity in vivo or other readouts, and/or understanding how RASA3 and Kindlin3 regulate T cell function and the signaling pathways. Interestingly, while RASA3 and Kindlin3 are both regulated by PIP3, they have opposite effects on LFA1 activity.  We will use advanced imaging technologies to determine the differential effects of PI3K signalling on these two proteins.

Antiviral restriction factors (ARF) are a critical element of cellular innate immunity, representing the first barrier to viral infection that can determine outcome. We aim to identify and characterise novel ARF and their viral antagonists, since therapeutic interruption of viral antagonism can enable restoration of endogenous antiviral activity.

We employ a number of human pathogens, in particular Human Cytomegalovirus (HCMV), Monkeypox virus (MPXV) and its vaccine, Modified Vaccinia Ankara (MVA). Our systematic proteomic analyses determine which cellular factors each pathogen targets for destruction, since we have shown these to be enriched in novel ARFs. For example, we recently developed a multiplexed proteomic technique that identified proteins degraded in the proteasome or lysosome very early during HCMV infection (Nightingale et al, Cell Host & Microbe 2018). A shortlist of 35 proteins were degraded with high confidence, and we have since shown that several are novel ARF, with characterisation of these factors forming ongoing projects. Application to MVA infection indicated further candidates, and identified novel mechanisms of vaccine action (Albarnaz et al, Nature Communications 2023); a study on MPXV will shortly be submitted. Furthermore, interactome screens can identify the viral factor(s) responsible for targeting each ARF, and indicate mechanism (Nobre et al eLife 2019).

This project will now identify and characterise critical pan-viral ARF, which can restrict diverse viruses. For the most potent, we will determine both the mechanism of restriction and the mechanism of virally mediated protein degradation. In order to prioritise the most important factors, there will also be the opportunity to use novel multiplexed proteomic screens.

This is an opportunity to work with Prof Fitzgerald and her group who are internationally renowned for their work on Barrett’s oesophagus including the development of a novel, non-endoscopic capsule sponge test, and Dr. Katki’s group leaders in developing quantitative methods for risk-based approaches to cancer screening.

The incidence of oesophageal cancer has increased rapidly in the past 30 years. Oesophageal adenocarcinoma (OAC), the most common form of oesophageal cancer in the US and UK, has <20% 5-year survival, which improves with early detection. Individuals known to have Barrett’s oesophagus, an asymptomatic precursor condition to oesophageal adenocarcinoma, undergo surveillance with the goal of treating advanced (dysplastic) Barrett’s oesophagus before it develops into cancer, thereby preventing cancer. Even if a cancer is not prevented, surveillance may detect it an earlier, asymptomatic, stage, where survival is better. However, currently the vast majority of Barrett’s oesophagus cases are undetected. The recent development of the capsule-sponge (Cytosponge) by Prof Fitzgerald’s lab has made screening for Barrett’s oesophagus more accessible, since it is cheaper than endoscopy, the prior screening method, and can be performed at a GP practice. We therefore expect an increase in the number of people with detected Barrett’s oesophagus, and therefore the number of people undergoing surveillance.  

Current surveillance guidelines depend only on characteristics of the Barrett’s oesophagus, and have not been updated to reflect the use of capsule-sponge as a surveillance tool. We will model the time to progression among patients with Barrett’s oesophagus, to inform surveillance intervals. Since Barrett’s oesophagus is asymptomatic, it can only be detected at times when the oesophagus is evaluated; special statistical methods are therefore required to model this, such as prevalence-incidence models, developed by Dr Katki’s group. The estimates will focus on absolute risk estimates, since ideally individuals would have a surveillance visit when their absolute risk exceeded a threshold, following the concept of ‘equal management of equal risk’. 

The projects will use clinical data from large cohorts of individuals undergoing endoscopy surveillance in England, Scotland and Northern Ireland. We will use both capsule-sponge and endoscopy data to inform surveillance intervals, including how surveillance intervals could vary based on the number of previous surveillance visits.