The structure and regulation of the actin cytoskeleton during B cell antigen internalisation

Abstract

B cells are responsible for producing high affinity antibodies, which facilitate the elimination of a variety of infections. In order to achieve this, B cells must extract cognate antigen from antigen presenting cells (APCs) via their B cell receptor (BCR). Antigen is then internalised, processed in endosomal compartments and presented in the context of MHC proteins on the B cell surface. This enables B cells to obtain the T cell help required for proliferation and differentiation into high affinity plasma and memory cells. The formation of an immune synapse between B cells and APCs is necessary for efficient antigen internalisation. Mechanical forces generated by actin and myosin IIA have been implicated in this process. However, how the B cell cytoskeleton is organised and regulated to generate the forces required for antigen extraction is not well understood.

Using STORM (Stochastic optical reconstruction microscopy) and live cell imaging we have visualised the three-dimensional nanoscale organisation of actin filaments and their dynamics during immune synapse formation. As described previously, actin formed lamellipodial structures in the synapse periphery. However, we also revealed an interspersed network of dynamic ARP2/3-generated foci of branched actin and formin-generated linear filaments throughout the B cell synapse. This network formed independently of antigen-mediated BCR stimulation but was regulated by BCR signalling. Functional studies showed that the actin foci associated with antigen clusters during antigen extraction and demonstrated that ARP2/3 activity was crucial for substrate-bound antigen internalisation. Formin activity also contributed to efficient antigen internalisation, likely by regulating actin foci dynamics. Thus, we conclude that ARP2/3 and formin activities are essential for maintaining the actin architecture and dynamics of the B cell immune synapse. We propose that ARP2/3- mediated actin polymerisation within foci is responsible for extraction of antigen clusters from APCs. Understanding how the cytoskeleton facilitates antigen internalisation has implications for the development of novel vaccines and immunotherapies in the future.

Date of Award	1 May 2019
Original language	English
Awarding Institution	King's College London
Supervisor	Susan Cox (Supervisor)