The threshold of B cell activation

Project Overview

Background

The threshold for activation of B cell receptor determines B cell development, the differentiation of subsets, the response to antigen and immune tolerance.  We are beginning to appreciate that the threshold varies through the development of B cells and probably through the lifetime of an individual; but we know little about its molecular basis or how it might be manipulated therapeutically.  These are some of the most fundamental questions in immunology with broad implications. 

Project

The project is to investigate how the activation threshold of the B cell receptor is set in different B cell subsets and during the immune response to antigens.  We will use cellular immunology, gene targeting, and immunoglobulin transgenic models to study these questions, and use ovalbumin (OVA) and hen egg lysozyme (HEL) as model antigens. In recent projects have used similar approaches to study the role of the guanine nucleotide exchange factor DOCK8 and the adaptor protein THEMIS2 in B cells. These studies have highlighted the importance of an antigen’s form and its display on cells in B cell activation, so this is something we would like to explore systematically in more detail. Most studies of B cell activation have used soluble antigens or anti-immunoglobulins to induce B cell activation, whereas most naturally occurring antigens are displayed on membranes.

The first year will be spent developing tools to study the B cell receptor and building models. We will use anti-HEL immunoglobulin transgenic mice to study the response to different forms of HEL or HEL-OVA, and gene targeted cell lines and mice to explore the role of downstream pathways to membrane-bound antigens, starting with the role of Grb2.

Our laboratory is within the MRC Human Immunology Unit and we are well supported with access to cutting edge reagents and technology. There will be strong support for this work in the laboratory, and from the wider community in the WIMM and in Oxford.

The student will study the affected cells at a molecular level, using biochemistry, and genetics to build and address hypotheses. The results will generate new insight into immune mechanisms and the work will provide a broad training in immunology.

Training Opportunities

This programme is suitable for any student with a background in biochemistry, immunology, medicine or a similar degree who is interested in using cellular, genetic and biochemical approaches to studying human disease. It might also be appropriate for a student in chemistry who wants to learn immunology and apply basic principles to the study of cells with imaging etc. A wide variety of lab techniques will be used and you will be become expert in cell biology and immunology. Experience of immunology is not a requirement, since teaching is available in the university and lab.

Students will be enrolled on the MRC WIMM DPhil Course, which takes place in the autumn of their first year. Running over several days, this course helps students to develop basic research and presentation skills, as well as introducing them to a wide-range of scientific techniques and principles, ensuring that students have the opportunity to build a broad-based understanding of differing research methodologies.

Generic skills training is offered through the Medical Sciences Division's Skills Training Programme. This programme offers a comprehensive range of courses covering many important areas of researcher development: knowledge and intellectual abilities, personal effectiveness, research governance and organisation, and engagement, influence and impact. Students are actively encouraged to take advantage of the training opportunities available to them.

As well as the specific training detailed above, students will have access to a wide-range of seminars and training opportunities through the many research institutes and centres based in Oxford.

Theme

Immunology & Infectious Disease and Physiology, Cellular & Molecular Biology

Admissions

Project reference number: 1008

Funding and admissions information

Supervisors

Name Department Institution Country Email
Professor Richard J Cornall FMedSci FRCP Centre for Cellular and Molecular Physiology Oxford University, Henry Wellcome Building for Molecular Physiology GBR richard.cornall@ndm.ox.ac.uk
Dr Mukta Deobagkar-Lele Nuffield Department of Clinical Medicine University of Oxford GBR mukta@well.ox.ac.uk

Ghezraoui H, Oliveira C, Becker JR, Bilham K, Moralli D, Anzilotti C, Fischer R, Deobagkar-Lele M, Sanchiz-Calvo M, Fueyo-Marcos E, Bonham S, Kessler BM, Rottenberg S, Cornall RJ, Green CM, Chapman JR. 2018. 53BP1 cooperation with the REV7-shieldin complex underpins DNA structure-specific NHEJ. Nature, 560 (7716), pp. 122-127. Read abstract | Read more

53BP1 governs a specialized, context-specific branch of the classical non-homologous end joining DNA double-strand break repair pathway. Mice lacking 53bp1 (also known as Trp53bp1) are immunodeficient owing to a complete loss of immunoglobulin class-switch recombination, and reduced fidelity of long-range V(D)J recombination. The 53BP1-dependent pathway is also responsible for pathological joining events at dysfunctional telomeres, and its unrestricted activity in Brca1-deficient cellular and tumour models causes genomic instability and oncogenesis. Cells that lack core non-homologous end joining proteins are profoundly radiosensitive, unlike 53BP1-deficient cells, which suggests that 53BP1 and its co-factors act on specific DNA substrates. Here we show that 53BP1 cooperates with its downstream effector protein REV7 to promote non-homologous end joining during class-switch recombination, but REV7 is not required for 53BP1-dependent V(D)J recombination. We identify shieldin-a four-subunit putative single-stranded DNA-binding complex comprising REV7, c20orf196 (SHLD1), FAM35A (SHLD2) and FLJ26957 (SHLD3)-as the factor that explains this specificity. Shieldin is essential for REV7-dependent DNA end-protection and non-homologous end joining during class-switch recombination, and supports toxic non-homologous end joining in Brca1-deficient cells, yet is dispensable for REV7-dependent interstrand cross-link repair. The 53BP1 pathway therefore comprises distinct double-strand break repair activities within chromatin and single-stranded DNA compartments, which explains both the immunological differences between 53bp1- and Rev7- deficient mice and the context specificity of the pathway. Hide abstract

Santos AM, Ponjavic A, Fritzsche M, Fernandes RA, de la Serna JB, Wilcock MJ, Schneider F, Urbančič I, McColl J, Anzilotti C, Ganzinger KA, Aßmann M, Depoil D, Cornall RJ, Dustin ML, Klenerman D, Davis SJ, Eggeling C, Lee SF. 2018. Capturing resting T cells: the perils of PLL. Nat. Immunol., 19 (3), pp. 203-205. | Read more

Cheng D, Deobagkar-Lele M, Zvezdova E, Choi S, Uehara S, Baup D, Bennett SC, Bull KR, Crockford TL, Ferry H, Warzecha C, Marcellin M, de Peredo AG, Lesourne R, Anzilotti C, Love PE, Cornall RJ. 2017. Themis2 lowers the threshold for B cell activation during positive selection. Nat. Immunol., 18 (2), pp. 205-213. Read abstract | Read more

The positive and negative selection of lymphocytes by antigen is central to adaptive immunity and self-tolerance, yet how this is determined by different antigens is not completely understood. We found that thymocyte-selection-associated family member 2 (Themis2) increased the positive selection of B1 cells and germinal center B cells by self and foreign antigens. Themis2 lowered the threshold for B-cell activation by low-avidity, but not high-avidity, antigens. Themis2 constitutively bound the adaptor protein Grb2, src-kinase Lyn and signal transducer phospholipase γ2 (PLC-γ2), and increased activation of PLC-γ2 and its downstream pathways following B cell receptor stimulation. Our findings identify a unique function for Themis2 in differential signaling and provide insight into how B cells discriminate between antigens of different quantity and quality. Hide abstract

Randall KL, Lambe T, Johnson AL, Treanor B, Kucharska E, Domaschenz H, Whittle B, Tze LE, Enders A, Crockford TL, Bouriez-Jones T, Alston D, Cyster JG, Lenardo MJ, Mackay F, Deenick EK, Tangye SG, Chan TD, Camidge T, Brink R, Vinuesa CG, Batista FD, Cornall RJ, Goodnow CC. 2009. Dock8 mutations cripple B cell immunological synapses, germinal centers and long-lived antibody production. Nat. Immunol., 10 (12), pp. 1283-91. Read abstract | Read more

To identify genes and mechanisms involved in humoral immunity, we did a mouse genetic screen for mutations that do not affect the first wave of antibody to immunization but disrupt response maturation and persistence. The first two mutants identified had loss-of-function mutations in the gene encoding a previously obscure member of a family of Rho-Rac GTP-exchange factors, DOCK8. DOCK8-mutant B cells were unable to form marginal zone B cells or to persist in germinal centers and undergo affinity maturation. Dock8 mutations disrupted accumulation of the integrin ligand ICAM-1 in the B cell immunological synapse but did not alter other aspects of B cell antigen receptor signaling. Humoral immunodeficiency due to Dock8 mutation provides evidence that organization of the immunological synapse is critical for signaling the survival of B cell subsets required for long-lasting immunity. Hide abstract

Johnson AL, Aravind L, Shulzhenko N, Morgun A, Choi SY, Crockford TL, Lambe T, Domaschenz H, Kucharska EM, Zheng L, Vinuesa CG, Lenardo MJ, Goodnow CC, Cornall RJ, Schwartz RH. 2009. Themis is a member of a new metazoan gene family and is required for the completion of thymocyte positive selection. Nat. Immunol., 10 (8), pp. 831-9. Read abstract | Read more

T cell antigen receptor (TCR) signaling in CD4(+)CD8(+) double-positive thymocytes determines cell survival and lineage commitment, but the genetic and molecular basis of this process is poorly defined. To address this issue, we used ethylnitrosourea mutagenesis to identify a previously unknown T lineage-specific gene, Themis, which is critical for the completion of positive selection. Themis contains a tandem repeat of a unique globular domain (called 'CABIT' here) that includes a cysteine motif that defines a family of five uncharacterized vertebrate proteins with orthologs in most animal species. Themis-deficient thymocytes showed no substantial impairment in early TCR signaling but did show altered expression of genes involved in the cell cycle and survival before and during positive selection. Our data suggest a unique function for Themis in sustaining positive selection. Hide abstract