Abstract
Engagement of the B cell receptor (BCR) with surface-tethered antigens leads to the formation of an immune synapse (IS), where cell signaling and antigen uptake are tightly coordinated. Centrosome re-orientation to the immune synapse has emerged as a critical regulatory step to guide the local recruitment and secretion of lysosomes, which can facilitate the extraction of immobilized antigens. This process is coupled to actin remodeling at the centrosome and at the immune synapse, which is crucial to promote cell polarity. How B cells balance both pools of actin cytoskeleton to achieve a polarized phenotype during the formation of an immune synapse is not fully understood. Here, we reveal that B cells rely on proteasome activity to achieve this task. The proteasome is a multi-catalytic protease that degrades cytosolic and nuclear proteins and its dysfunction is associated with diseases, such as cancer and autoimmunity. Our results show that resting B cells contain an active proteasome pool at the centrosome, which is required for efficient actin clearance at this level. As a result of proteasome inhibition, activated B cells do not deplete actin at the centrosome and are unable to separate the centrosome from the nucleus and thus display impaired polarity. Consequently, lysosome recruitment to the immune synapse, antigen extraction and presentation are severely compromised in B cells with diminished proteasome activity. Additionally, we found that proteasome inhibition leads to impaired actin remodeling at the immune synapse, where B cells display defective spreading responses and distribution of key signaling molecules at the synaptic membrane. Overall, our results reveal a new role for the proteasome in regulating the immune synapse of B cells, where the intracellular compartmentalization of proteasome activity controls cytoskeleton remodeling between the centrosome and synapse, with functional repercussions in antigen extraction and presentation.
Highlights
B Lymphocytes mediate humoral responses by the recognition of antigens tethered at the surface of presenting cells such as follicular dendritic cells or macrophages, forming a domain known as an Immune Synapse (IS) [1,2,3]
These results indicate that inhibition of proteasome activity in B cells does not affect cell surface levels of MHC-II molecules and does not influence B-T cell interactions per se, but could affect critical steps upstream of antigen presentation, such as antigen extraction and processing
To discard that our observations were due to off target effects elicited by MG-132, we evaluated the antigen extraction capacity of B cells pretreated with another proteasome inhibitor, Epoxomicin
Summary
B Lymphocytes mediate humoral responses by the recognition of antigens tethered at the surface of presenting cells such as follicular dendritic cells or macrophages, forming a domain known as an Immune Synapse (IS) [1,2,3]. The spreading reaction exerted by B cells is tightly coupled to their signaling capacity, as cells that recruit fewer signaling molecules to micro-clusters show deficient spreading responses to membrane-bound antigens [4] This is followed by a contraction phase where BCR-Ag complexes are concentrated into a central cluster by the concerted action of the microtubulebased motor protein, dynein, and actin rearrangements [4]. The extraction of antigens immobilized on stiffer substrates can be facilitated by proteases, which originate from the local secretion of MHC-II+ lysosomes at the synaptic membrane This process is controlled by the cooperative action of polarity complex proteins, such as Cdc, Par, and aPKC, which promote the recruitment of the centrosome and associated lysosomes toward the immune synapse [6,7,8]. The peptides generated from uptaken antigens are further mounted onto MHC-II molecules and presented to T cells in order to trigger B-T cooperation, thereby promoting the maturation and differentiation of B cells to memory B cells or plasma cells [9, 10]
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