Research Opportunities

This collaboration between the Altan-Bonnet (NCI), Buckley and Coles (Oxford) labs addresses how organs generate distinct inflammatory responses despite sharing common components like immune cells, fibroblasts, and the extracellular matrix. In collaboration with additional teams from the Netherlands, and Canada, we aim to uncover the molecular, cellular, and tissue-level rules governing organ-specific inflammation.

We hypothesize that (1) organ context and cellular experience shape the perception of inflammatory signals, and (2) organ-specific hierarchies integrate responses into coordinated outcomes. Using a data-driven approach, we will combine ex vivo and in silico models of mouse and human tissues to explore these mechanisms. High-throughput robotics will generate diverse tissue models with varying sensitivities to infection or immunopathology. Multimodal datasets from these models will be analyzed using machine learning to build computational models, to guide iterative cycles of discovery.

This project will revolutionize tissue biology by creating a unified framework for understanding tissue-specific inflammation, paving the way for new treatments. We are seeking researchers with expertise in bioengineering, computer science, or immunology to join this interdisciplinary effort.

Project keywords: immunology, systems biology, biomedical engineering. 

Asthma is the world’s most common chronic lung disease. It is increasing rapidly in incidence, but it is not known why. It arises from the complex interplay of genetic predispositions and the influence of environmental factors early in life, particularly early life infections with bacteria like haemophilus influenzae and with viruses like respiratory syncytial virus (RSV), and this points strongly to epigenetic changes being induced in the airway epithelium. 

It has been previously hard to decipher these mechanisms, but that is now becoming possible due to the ability to obtain direct airway samples at bronchoscopy and via nasal brushings and importantly the advent of technologies allowing analysis of epigenetics on small tissue samples and even at a single cell level.

We have generated large epigenetic and immunological datasets of airway epithelium and bronchial biopsies at single cell resolution. We also have DNA methylation data from a large paediatric cohort with early life RSV. We have developed a murine model of long-term infection with haemophilus influenzae and so, for the first time, can model the interactions and consequences of early life bacterial / viral coinfection.

We aim to exploit these datasets and models to understand in detail the specific immunoepigenetic changes in asthma and early life infection in mice and humans with a view to developing targeted epigenetic therapies. 

Our lab is interested in unconventional and innate-like cells (such as MAIT cells) and what their role is in immune protection and pathology.Tmic cells are one such cell - a recently described subset of T helper cells which are Microbially reactive, Innnate-like and Class II restricted. These cells are abundant in the human colon and marked out by high expression of CD161 – a feature normally associated with unconventional T cells – along with other evolving markers. However, as well as behaving like innate-like cells they bear conventional TCRs and are restricted by MHC Class II. They are also found in mice where they adopt a double negative phenotype over time in response to gut commensals. We think these cells are important in gut homeostasis (also other organs potentially) and we have shown they are involved in inflammation, but there are many questions as to their origin and overall functionality to be answered.

This project would explore the development of these cells using CBIR mice which over-express a commensal-reactive TCR, first by scRNASeq and scATACseq to define the steps along the pathway from conventional tissue memory to Tmic phenotype. Secondly using spatial transcriptomic methods to define their colocalization in the steady state and after challenge. Finally we will use a new in vivo CRISPR screen method (CHIME) to define the critical steps in development of Tmics and explore their functions in vivo. We will aim to compare mouse and human Tmic populations to define this novel conserved cell population and explore its role in health and disease.

Reference: https://www.nature.com/articles/s41467-022-35126-3

It is clear that stress responses and immunity are closely entwined, and epidemiological research shows that their complex interplay is key to the development of both psychiatric and autoimmune disorders (Teicher 2013 PMID:23982148, Dube 2009 PMID: 19188532). For example, a major component of the stress response is cortisol release, and depression is associated with hypercortisolaemia and glucocorticoid (GC) resistance. Abnormal responses to GCs likely contribute to the chronic inflammation observed in many patients with depression, as GCs can prime inflammatory responses, and GC-resistant immune cells produce increased levels of pro-inflammatory cytokines. Cytokines can act on the brain to produce the sickness-like behaviours characteristic of depression, and other aspects of GC-induced immune dysregulation (e.g. effects on neutrophils) may also play a role.  Epidemiological studies show that psychological stress interacts with genetic risk to lead to depression and psychosis (e.g. Wang 2023 PMID:36717542), but the risk variants involved and the cellular mechanisms of this effect are unknown. We hypothesize that some risk variants for psychiatric disorders act through glucocorticoid responsive regulatory elements in specific immune cell subsets to lead to symptoms. We further hypothesize that by dissecting the cell subset- and context-specific effects of glucocorticoids in health and in patients, we can develop a better biomarker of impaired neuroendocine signalling in psychiatric disorders, opening the door to biomarker development and more personalised approaches to treatment in stress-responsive autoimmune and psychiatric disorders.   

You would work with Dr Luis M. Franco and Dr Mary-Ellen Lynall to investigate these hypotheses using immunogenetic and functional genomic techniques, gaining training in cutting edge bioinformatics, statistical genetics, immunology, clinical phenotyping, and (if desired) wet-lab experimental approaches.   You would integrate emerging genetic association results in autoimmune and psychiatric disorders with (a) in-house glucocorticoid-response datasets (see https://www.niams.nih.gov/labs/franco-lab) (b) healthy and patient bulk and single cell datasets from our laboratories.   

Dr Franco's group in the Functional Immunogenomics Section at the NIAMS focuses on the immunobiology of glucocorticoid responses (e.g. Franco 2019 PMID:30674564). 

Dr Lynall's group in the Dept of Psychiatry at Cambridge focuses on immunogenetic analyses and immunophenotyping in psychiatric patients and population cohorts (e.g. Lynall 2022 PMID:36243721).

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   Understanding how ROS is tightly regulated is 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 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.

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 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.

Innate immune responses are the first line of defense against viral infection, but their inappropriate activation can cause autoinflammatory disease. The Best and Modis groups study how the host senses viruses, mounts sufficiently sensitive yet specific responses, and how this balance can be perturbed for example by disease mutations. Our work focuses largely on the roles of two key families of viral restriction factors, the TRIMs and RIG-I-like receptors (RLRs). We integrate an exceptionally broad spectrum of approaches, from in vivo work in high and maximum contain laboratories to state-of-the-art electron microscopy. This uniquely positions us to obtain a more complete understanding of virus-host interactions under physiological conditions with mechanistic insights in atomic-level detail.  

This PhD project will focus on unravelling important insights into how RNA viruses are detected and targeted by their hosts. The aims of this project will apply our full complement of approaches, including electron microscopy, biochemical and cell-based assays, and in vivo work as needed to obtain a detailed mechanistic understanding of the contributions of virus-host interactions to antiviral innate immunity and virus pathogenesis. Our long-term goal is to use contribute to the design of novel therapeutics, such as antivirals, vaccine adjuvants, or immunomodulatory therapeutics, with potential applications in the treatment of infection, autoinflammatory disorders and cancer.

Our teams at the NIAID Rocky Mountain Laboratories and University of Cambridge are strongly committed to fostering a supportive and inclusive work environment in which trainees can thrive and experience the thrill of scientific discovery.

References:
Chiramel AI, Meyerson NR, McNally KL, Broeckel RM, Montoya VR, Méndez-Solís O, Robertson SJ, Sturdevant GL, Lubick KJ, Nair V, Youseff BH, Ireland RM, Bosio CM, Kim K, Luban J, Hirsch VM, Taylor RT, Bouamr F, Sawyer SL, Best SM (2019) TRIM5α Restricts Flavivirus Replication by Targeting the Viral Protease for Proteasomal Degradation. Cell Rep 27:3269  PMC8666140

Yu Q, Herrero del Valle A, Singh R, Modis Y (2021) MDA5 autoimmune disease variant M854K prevents ATP-dependent structural discrimination of viral and cellular RNA. Nat Commun 12:6668  PMCID: PMC8602431

Stoll GA, Pandiloski N, Douse CH, Modis Y (2022) Structure and functional mapping of the KRAB-KAP1 repressor complex. EMBO J 41:e111179  PMC9753469

Shannon JG, Sturdevant GL, Rosenke R, Anzick SL, Forte E, Preuss C, Baker CN, Harder JM, Brunton C, Munger S, Bruno DP, Lack JB, Leung JM, Shamsaddini A, Gardina P, Sturdevant DE, Sun J, Martens C, Holland SM, Rosenthal NA, Best SM (2023) Genetically diverse mouse models of SARS-CoV-2 infection reproduce clinical variation in type I interferon and cytokine responses in COVID-19. Nat Commun 14:4481  PMC10368652

Regulatory T cells (Tregs) have the ability to suppress allogeneic immune responses and have therefore become interesting as a possible cellular therapy.  Our group is leading a phase II randomised clinical trial utilising Tregs as a cellular therapy for kidney transplant recipients.

The proposed project aims to in depth investigate the local immune response within the transplanted allograft, specifically focusing on infiltrating regulatory T cells and their local interactions with other infiltrating immune cells, utilising renal biopsy samples from the cell therapy trial.  We hypothesise, that cellular therapy with regulatory T cells will result in trafficking of the Tregs cells to the allograft and creation of a local tolerogenic microenvironment.

We are proposing to utilise our unique collection of Treg treated patients’ biopsy samples employing the cutting-edge technology of spatial transcriptomics and single cell RNA sequencing to determine the cellular and molecular composition of leukocyte infiltrates in allograft biopsies, dissect cellular interactions in situ and determine Treg homing to the allograft.

How are different B cell populations developmentally linked in human health and disease?

We are investigating the generation, function and plasticity of B cell populations in human health. In particular, we are interested in how different lymphocyte subsets are developmentally linked and differences in function, and therefore providing a platform to understand how B cell fate may be different in human disease. We are defining how B cells select a particular developmental pathway, and will use this information to develop methods for modulating B cell function as potential therapeutic approaches.

How can B and T cells may be therapeutically modulated across cancers and autoimmune diseases?

There is accumulating evidence for the role of both T and B cells in modulating immune responses to both solid tumours and haematological malignancies. We are investigating the contributions, function and heterogeneity of B and T cells on the immune responses to tumours and their potential role in cancer detection and treatment. We are determining the nature of B and T cell immuno-surveillance, regulation and activation across cancers and autoimmune diseases, as well as the immunological features associated with better prognosis and immunomodulation. With this, we aim to highlight novel therapeutic avenues. Our lab is affiliated with the Oxford Cancer Centre (https://www.cancer.ox.ac.uk/research/research-themes/developments-in-immuno-oncology) and non-cancer clinicians, with strong clinical links to a wide range of hard-to-treat diseases.

What is the effect of genetic and environmental variation on B and T cell fate?

Immunological health relies on a balance between the ability to mount an immune response against potential pathogens and tolerance to self. B and T cells are key to the immune response by producing antibodies and cytotoxic T cells. B/T cell clones selectively expand following antigen recognition by B and T cell receptors (BCR and TCR) respectively. BCRs are the membrane-form of antibodies and are generated through DNA recombination resulting in the potential to recognise a vast array of pathogens. Defects in the ability to mount effective B cell or T cell responses have been implicated in infectious susceptibility, impaired surveillance of cancer and immunodeficiencies, whereas a breakdown of immunological tolerance has been attributed to autoimmune diseases such as through autoantibody production and reduced numbers of regulatory B/T cells. Through integrating genomics, bulk and single-cell transcriptomics, and metabolomics data, serological, B /T cell repertoire and viromics datasets we will investigate the effect of both genetic variation and environmental factors on B cell fate, regulation, and the relationship to disease susceptibility.

T cells are a major component of adaptive immune response, continuously screening lymphoid tissues for antigen peptides presented by major histocompatibility complex (peptide/MHC or pMHC)3. These antigen peptides are recognized by T-cell receptors (TCRs). Thymocytes with a low-affinity TCRs mature into T cells and enter the lymphoid organs, where they are exposed to foreign antigen peptides by MHC molecules from antigen-presenting cells including macrophages, dendritic cells, and B-cells, during infection. When the T-cell receptors bind to antigenic peptide, T-cells are activated and undergo clonal expansion, resulting in immune response. CD8+ T cells play an important for immune response and viral clearing, but their role in protection and pathogenesis of SARS-CoV-2 remains poorly understood4. In addition to extensively studied spike protein, open reading frame 3a (ORF3a), a highly conserved protein within the Betacoronavirus subgenus, has been considered as a potential target for vaccines or therapeutics, with deletion of ORF3a resulting in decreased viral titer and morbidity. We have identified shared CD8+ T cell clonotypes responding to a ORF3a in COVID-19 infections.  Importantly, shared clonotypes in severe COVID-19 infections provides a target for development of novel antiviral immunotherapies. The aim of this project is to analyse shared TCR clonotypes in ORF3a recognition and provide structural basis for the recognition of ORF3a-pMHC complex by T-cell receptors.

Immunity and immune-tolerance at mucosal and other barrier surfaces is vital for host survival and homeostasis with antibodies or immunoglobulins (Ig) playing a key role. However, the specific roles of B cell sub-types, particularly, the innate-like B-1 cell subset is poorly understood. The surgeon James Rutherford Morrison (1906) called the omentum in the peritoneal cavity the "abdominal policeman" and it promotes gut IgA production by peritoneal B-1 cells. IgA is the most abundantly produced antibody isotype and is known to be important for mucosal immunity. It is estimated that ~50% of IgA is derived from B-1 cells. In addition, B-1 cells are thought to be important because they make T cell-independent “natural” IgM circulating in our blood, and they can rapidly respond to mucosal perturbations such as an infection. By contrast, it will take conventional B-2 cells weeks to mount a germinal center (GC) reaction and generate antibodies that have undergone T cell-dependent affinity maturation and isotype switching. Textbooks currently do not entertain the possibility that B-1 cells can also participate in GC reactions. This project aims to challenge such an assumption. After all, B-1 cells in the gut mucosa and probably other mucosal tissues can undergo class switch recombination to IgA. However, the differentiation program that leads to this distinct pathway of IgA production is not well understood: for example, it is unknown if this process occurs outside or within GCs in mucosa-associated lymphoid tissues (MALT). Expertise on B-1 cells in the Muljo lab and the GC reaction in the Turner lab will be combined to explore this potentially paradigm-shifting research. Both wet-bench and bioinformatic research opportunities are available.

Students will learn about the fundamentals of transcriptional; epigenetic and post-transcriptional regulation; immunometabolism; in vivo CRISPR screening; CRISPR editing in primary B cells; and systems immunology. The combination of classical immunological techniques and cutting-edge, multi-disciplinary approaches will enable important discoveries to define the in vivo biology of B-1 cells. Ultimately, we seek novel insights that can be translated to inform vaccine design targeted to activate B-1 cells and/or therapeutics to inhibit their activity when necessary.
 

Recent publications:

Turner, D. J., Saveliev, A., Salerno, F., Matheson, L. S., Screen, M., Lawson, H., Wotherspoon, D., Kranc, K. R., and Turner, M. (2022). A functional screen of RNA binding proteins identifies genes that promote or limit the accumulation of CD138+ plasma cells. eLife, 11, e72313. PMID: 35451955; DOI: 10.7554/eLife.72313.

Osma-Garcia, I.C., Capitan-Sobrino, D., Mouysset, M., Bell, S.E., Lebeurrier, M., Turner, M. and Diaz-Muñoz, M.D. (2021). The RNA-binding protein HuR is required for maintenance of the germinal centre response. Nature Communications, 12(1):6556. PMID: 34772950; DOI: 10.1038/s41467-021-26908-2.

Wang, S., Chim, B., Su, Y., Khil, P., Wong, M., Wang, X., Foroushani, A., Smith, P. T., Liu, X., Li, R., Ganesan, S., Kanellopoulou, C., Hafner, M. and S. A. Muljo. (2109). Enhancement of LIN28B-induced hematopoietic reprogramming by IGF2BP3. Genes & Development, 33: 1048-1068. PMID: 31221665; DOI: 10.1101/gad.325100.119.

We have several lines of research that accommodate excellent PhD candidates. These revolve around the theme of RNA viral pathogens, antibodies/B-cell responses and immunodeficiencies.

The first involves understanding Immune Correlates of protective immunity, specifically which types of B-cell response and their fine specificities are important for protection against specific RNA viral pathogens (RNA viruses from HIV, HCV to Ebola) how B-cell responses to correlate with protection by vaccines to specific pathogens. The 2nd project involves using broadly neutralizing monoclonal antibodies to develop improved and novel vaccines against notoriously variable viruses. The 3rd project involves understanding how the resident virome in primary, acquired or induced immunodeficiencies leads to chronic immune activation and poor prognosis, with an emphasis on mucosal immunity.

We have the largest world-wide collection of patients suffering from rare-inherited immunodeficiency that have been whole-genome sequenced (1500+ cases). Using established analytical expertise the candidate will use novel methods to interrogate and filter potential genetic mutations, we will identify novel candidate genetic loci in patients grouped by disease phenotype or familial relationship. Candidate genetic loci will be investigated using CRISPR-editing of patient derived material (lymphoblastoid, fibroblast and iPS cell lines). Confirmatory studies at mRNA, protein and functional level will be carried out to validate the link between variant and disease.