Characterisation of the mechanism of B-cell receptor triggering

Thesis

B cells are key mediators of humoral immunity and rely on signals initiated by the B cell receptor (BCR). The structure of the BCR and mechanism of ligand binding are well characterised, but what remains to be understood is how ligand binding triggers intracellular signalling. A number of opposing models have been proposed. In this thesis, these models have been tested experimentally using a toolkit of reagents based on the HyHEL10 BCR and its natural ligand Hen-Egg Lysozyme (HEL). Whilst a receptor crosslinking-based model accounts for multivalent ligand presented in solution, the inability of monovalent ligands to trigger robust B cell signalling, except when they are surface-associated, implicates a second triggering mechanism. To narrow down potential theories of BCR triggering, the stoichiometry of the BCR in the resting state was established using multicolour fluorescence microscopy and shown to be predominantly monomeric. Using fluorescence imaging, it was found that BCR triggering by surface associated monovalent ligands correlated with the exclusion of the large phosphatases, CD45 and CD148, from regions of BCR/ligand engagement. In addition, BCR triggering was shown to be dependent on both the size of the BCR/ligand complex and the size of the extracellular domain of CD148. These findings suggest that the size-dependent exclusion of CD45/CD148 functions as a second mechanism of BCR triggering for surface bound ligand, and complements the ability of BCR to be directly triggered by cross- linking multivalent ligands. These observations have important implications in terms of differential signalling, and the positive and negative selection of B cells. The mature B cell immune response to foreign antigen is thought to be predominantly focused on membrane bound antigen, displayed via complement or immunogloblin Fc receptors, whilst soluble antigens may be tolerogenic or lead to T-independent activation, at high avidity. It is also to be expected that differential expression of CD45/CD148 on B cell subsets will tune their response to membrane bound antigens.

Authors: Wilcock, M

• DOI: 10.5287/ora-r5ojqxd8r
• Type of award: DPhil
• Level of award: Doctoral
• Awarding institution: University of Oxford