Research Opportunities

Mitochondrial DNA (mtDNA) mutations have emerged a major cause of neurological disease and may also contribute to the ageing process – but their origin is not well understood. Remarkably, we have shown that most humans harbour a mixture of mutant and wild-type mtDNA (heteroplasmy) at very low levels. Our aims is to understand how mtDNA mutations arise, how they are inherited, and how they accumulate in specific tissues, particularly in the nervous system. Harnessing this knowledge, we will develop new treatments targeting the mitochondrion.

Invent and implement new radioactive probes for imaging specific molecular targets in animal and human brain with positron emission tomography

The cerebellum is a fascinating brain structure. While it has traditionally been regarded solely as a regulator of motor function, recent studies have demonstrated additional roles for the cerebellum in higher-order cognitive functions such as language, emotion, reward, social behaviour and working memory. Accordingly, cerebellar dysfunction is linked to motor diseases such as ataxia, dystonia and tremor, as well as cognitive affective disorders such as autism spectrum disorders and language disorders.

We understand surprisingly little about the molecular processes that underlie the formation of the cerebellum and that, when disrupted, lead to disease. The goal of our research is to provide fundamental insights into the genetic, molecular and cellular mechanisms that govern the development and different diseases of the cerebellum with the  ultimate desire to develop novel treatment options for these disorders.

The project will focus on the functional characterization of novel gene mutations causing cerebellar disorders, with particular emphasis on the effects of disease genes on the dendritic arborisation of developing Purkinje cells in the cerebellum. The approach will be multi-disciplinary and employs a variety of methods including functional experiments in cell lines and primary neurons, as well as modelling of identified patient mutations and their effects using stem cells combined with genome engineering. The project is supervised by two experienced investigators with complementary expertise. Research in the Hammer group at NIH will focus on introducing mutations of interest into primary Purkinje cells and to investigate dendritic phenotypes. Research in the Becker group at Oxford will include further functional analyses in Purkinje cells from mutant mouse models, as well as in human induced pluripotent stem cells.

Becker Group website:
https://www.ndcn.ox.ac.uk/research/cerebellar-disease-group

Hammer Group website:
https://irp.nih.gov/pi/john-hammer

The inflammasome consists of a cytosolic NOD-like receptor, an adaptor molecule (ASC) and an effector molecules caspase 1.  Once activated the inflammasome processes inflammatory cytokines such as interleukin 1 beta (IL1B) and IL18 as well as driving an aggressive form of cell death (pyroptosis).  Inflammasomme protein complexes are central to sustaining inflammation in acute diseases (like COVID-19 associated ARDS) or chronic conditions (such as Alzheimer’s Disease, Parkinson’s, diabetes, arthritis).  Patients with rare autoactivating mutations in the NLR proteins have basally active inflammasomes leading to severe autoinflammatory syndromes. How inflammasome complexes form within the cell, particularly in patients with autoactivating mutations in NLRs are poorly understood.  

The aims of this project are as follows:
1.      Identify the molecular mechanisms by which the gain of function mutations causes constitutive activation of the NLRs
2.      Determine why gain of function mutations in different NLRs (NLRP3 and NLRC4) result in differences in inflammasome cytokine production with NLRP3 biased towards IL1B and NLRC4 towards IL18
3.      Visualise how gain of function mutations alter inflammasome formation by visualising the protein complexes at super resolution and atomic resolution

This project will study how the inflammasome forms using state of the art microscopy techniques including live super resolution imaging and cyroelectron microscopy tomography.  The consequences of the gain of function mutations on inflammasome formation will be studied using these techniques in cell lines where the key proteins are tagged and the gain of function mutations introduced by CRISPR/Cas9 (many of which are already available within the laboratory).  This work will be extended to consider cells from patients with these diseases to map back the biology and the imaging onto the cell line models.   

A balance between neuronal excitation and inhibition is crucial for normal brain physiology; upsetting this balance underlies various brain pathologies. To shed light on the molecular underpinnings of this regulation at the synapse level, this project will investigate the dynamics of glutamate- and GABA-A synapses and receptors in CA1 hippocampus under baseline conditions and in response to synapse potentiation. Specifically, using structural, functional and imaging approaches we will study both, spiny glutamatergic and aspiny GABA-ergic CA1 synapses and associated receptor complexes (AMPA-type glutamate and GABA-A) and how these change at the synapse- and receptor levels in response to LTP (long-term potentiation) induction. Our aim will be to monitor changes of glutamatergic and GABAergic synapses and receptors at pyramidal neurons (glutamate) and/or parvalbumin-positive (PV+) interneurons at various points after LTP induction. We will monitor changes in synapse size and receptor composition using advanced imaging and electrophysiological approaches.

Inherited neurological disorders are disabling, progressive, often fatal conditions, representing an enormous unmet medical need with devastating impacts on affected families, the healthcare system, and the economy. There are no cures and the limited therapies available treat symptoms without addressing the underlying disease.

Next-generation sequencing has facilitated a molecular diagnosis for many inherited neurological disorders, such as mitochondrial diseases and other neuromuscular diseases, which are the focus of this research. The development of targeted therapies requires detailed laboratory investigation of molecular and mutational mechanisms, and a systematic evaluation of well-chosen agents as well as gene and transcript directed strategies using standardized experimental systems. Our research is focusing on understanding the molecular pathogenesis of childhood onset inherited neurological diseases, such as mitochondrial disease and other neuromuscular diseases to develop targeted therapies.

Using a translational approach, we aim to
1. understand the clinical course of patients in relation to the underlying disease mechanism
2. delineate the mutational and molecular mechanisms of the molecular defect in the appropriate cell types by developing model systems such as induced neuronal progenitor cells (in vitro) and zebrafish (in vivo)
3. improve the treatment options for patients by developing novel therapies that are directed at these mechanisms, including directly at the genetic mutation or resulting transcript.

We use a combination of exome sequencing, genome sequencing, and other omics technologies to identify novel disease genes and disease mechanisms. By functional evaluation in vitro (induced neuronal progenitor cells) and in vivo (zebrafish) we confirm pathogenicity and uncover molecular mechanisms of disease. To address the mutational mechanisms, we use gene transfer, splice modulation, allele silencing and CRISPR/cas systems.

My group studies the human malaria parasite Plasmodium falciparum.  Collaborative PhD projects can be offered in research areas centered around Plasmodium DNA biology: we are particularly interested in the molecular mechanisms underlying DNA replication and cell cycle control in Plasmodium, which replicates by an unusual method called schizogony.  We are also interested in mechanisms for silencing and promoting the recombination of a family of key virulence genes called var genes - particularly the role that G-quadruplex DNA structures may play in var gene control.  In fact, we have recently discovered that G-quadruplexes and their helicases have more general roles in genome stability and evolution in the malaria parasite as well.

This collaborative project between Dr. Steven Holland’s laboratory at the NIH and Dr. Lalita Ramakrishnan’s lab at the University of Cambridge will seek to understand the mechanistic basis of human susceptibility to environmental mycobacteria that are nonpathogenic to most people but can cause serious disease in individuals with specific immune deficiencies. Dr. Holland runs an international referral service that takes care of a unique cohort of patients with genetic susceptibility to nontuberculous mycobacterial infections. In the lab, they are mapping these susceptibilities and have found then to map to distinct immune genes, e.g., IRF8 and GATA-2, myeloid growth factors, IL-12R, the GTPase Rac2, to name only a few. 

Dr. Ramakrishnan’s group has pioneered the optically transparent and genetically tractable zebrafish as a model for mycobacterial pathogenesis. The use of the zebrafish has enabled discoveries about TB immunopathogenesis and the genetic basis of susceptibility to TB which has led to the discovery of a variety of inexpensive, approved drugs that can be used to treat TB, often in a patient genotype-directed manner. They have also used the zebrafish to understand the mechanism of leprosy neuropathy.

Through this joint project, the two labs will work together to harness the power of the zebrafish to understand the molecular and cellular basis of the human susceptibilities identified by Holland. The student will move between humans and fish (and Bethesda and Cambridge) to uncover fundamental mechanisms of mycobacterial disease pathogenesis while acquiring mastery over the disciplines immunology, infectious diseases, genetics, molecular biology and cell biology.

There are more than 200 million clinical cases of malaria each year, leading to nearly half a million deaths, primarily among children in Africa. The two major tools for malaria control, antimalarial drugs and insecticides, are both seriously threatened by resistance, making the search for a highly effective malaria vaccine more urgent than ever. My lab focuses on the malaria parasite blood stages, during which parasites invade, multiply inside and consume human erythrocytes. The process of erythrocyte invasion represents a brief extracellular window in the parasite life cycle when parasites are exposed to the antibody-mediated immune system, making it a potential vaccine target. A number of vaccine-related projects are available that intersect with the interests of NIH collaborators in the NIAID Malaria Research Program, from systematic screening of new potential vaccine candidates, to deep structural understanding of current high-profile candidates, to understanding natural immunity to malaria by working with partners in endemic countries in order to inform better vaccine design. All could involve a mix of new technologies, including CRISPR/Cas9 engineering of parasite genomes, and represent an opportunity to contribute to the long-term battle against one of humanities oldest and most persistent infectious disease foes.

A signalling pathway linking nutrient availability to changes in gene expression that hinges on the phosphorylation of translation initiation 2 (eIF2) has long been known to exist. Recognized initially as the yeast General Control Response, recent convergent lines of research have implicated its metazoan counterpart, the Integrated Stress Response, in diverse physiological processes ranging from immunity to memory formation.

This PhD programme will exploit our emerging detailed understanding of translation initiation and termination to shed light on unanticipated mechanistic aspects of the ISR. An understanding of these details may inform efforts to target the ISR to therapeutic ends.

Apicomplexan pathogens are highly-adapted intracellular parasites of humans causing disease including malaria, toxoplasmosis and cryptosporidiosis. These parasites actively confront, subvert and defend themselves against host immune attack using a complex suite of parasite surface and secreted proteins that hijack immune signalling pathways. Moreover, transmission and generation of genetic novelty occurs in definitive hosts where differentiation into sexual parasite forms occurs. Relatively little is known, however, of the molecules and processes that drive these events, particularly during the sexual stages of parasite development. This project will use new methods in in vitro culture of sexual development in Toxoplasma, advanced methods for global spatial characterisation of parasite cell proteomes in order to identify specific proteins thought to be implicated in these interactions, and then utilise CRISPR/cas9 mutagenesis tools to engineer pools of strains deficient in these specific proteins. By assaying mutant pools both in vitro, and through the definitive host we will identify proteins and processes required for sexual stage conversion.

Primary progressive multiple sclerosis (PPMS) is a chronic demyelinating disease of the central nervous system, which currently lacks restorative therapies. Transplantation of neural stem cells (NSCs) has been shown to promote healing of the injured CNS, but previous work has demonstrated that NSCs from patients with PPMS are prematurely senescent. Cellular senescence causes a pro-inflammatory cellular phenotype that impairs tissue regeneration. Senescence in PPMS NSCs was found to be associated with increased secretion of HMGB1, a pro-inflammatory alarmin found to inhibit oligodendrocyte differentiation, and also found increased within white matter lesions of PPMS autopsy tissue. This project aims to understand the role of HMGB1 in PPMS NSC senescence using techniques such as CRISPR-Cas9, RNA sequencing, and functional NSC assays. The longterm goal of this project will be to determine the cause of senescence in NSCs from patients with PPMS and if these cells are suitable for therapeutic use.

Andrew Holmes’ Lab (NIH/NIAAA) and Armin Lak’s Lab, Oxford University

Several decades of research has shown that medial frontal cortical neurons, as well as the neuromodulatory system that innervate the medial frontal cortex, notably the dopamine system, are important in reward valuation and decision making. However, it is not known whether different regions of medial frontal cortex play distinct roles in guiding decisions. Moreover, the role of frontal dopamine signals in shaping and regulating decisions has yet to be established. To address these questions, this project will use a combination of large-scale Neuropixels recording across the medial frontal cortex, as well as optical measurement of dopamine release in the medial frontal cortex during decision making in mice. At Oxford, the project will use Neuropixels probes to record the activity of many neurons across different regions of medial frontal cortex while mice perform a task that systematically manipulates the value of choice options. This data will allow us to investigate the relation between frontal neuronal activity and decision-making variables, and characterize distinct roles of different medial frontal regions in choice behavior. At NIH, the project will take advantage of optical and genetic methods to measure the dynamics of dopamine release in frontal cortical regions identified in the electrophysiological recordings. These experiments will reveal the roles that frontal dopamine play during decision making. In analyzing the electrophysiological and optical data, we will use computational models of learning and decision making to relate neuronal signals with trial-by-trial model-driven estimates of decision variables. The project is primarily experimental in nature but will provide an opportunity to develop computational skills. Together, the project will provide fundamental insights into behaviorally-relevant computations that neurons across the medial frontal cortex perform during decision making, and will reveal the roles of frontal dopamine signals in shaping choice behavior. For more information please visit: https://www.niaaa.nih.gov/laboratory-behavioral-and-genomic-neuroscience and https://www.laklab.org.

*This project is available for the 2021 Oxford-NIH Pilot Programme*