CD23 Sheddase A Disintegrin and Metalloproteinase 10 (ADAM10) Is Also Required for CD23 Sorting into B Cell-derived Exosomes

Abstract

The low affinity receptor for IgE, CD23, is the natural regulator of IgE synthesis, and understanding both the synthesis and the catabolism of CD23 are, thus, important issues. Membrane CD23 is cleaved by a disintegrin and metalloproteinase 10 (ADAM10) and this cleavage influences the ability of CD23 to regulate IgE. In contrast to the belief that cleavage is a cell surface event, endosomal neutralization with NH 4 Cl was found to dramatically reduce CD23 cleavage, suggesting that the majority of CD23 cleavage occurred subsequent to internalization in the endosomal pathway and not at the cell surface. In line with this, full-length CD23 was shown to be sorted in an ADAM10-dependent manner into exosomes. Greatly increased ADAM10-mediated CD23 cleavage was seen at endosomal pH. Additionally, the stalk region of CD23 was found to interact with ADAM10 and ADAM10 binding of CD23 was found to be protease independent. SPR analysis of the interaction indicated about a 10-fold increase in the R max at endosomal pH (pH 5.8) compared with pH 7.4, whereas the affinity of the interaction was not significantly changed. The R max change, combined with the increased cleavage at endosomal pH, indicates greater accessibility of the CD23 stalk region for ADAM10 at the lower pH. These results indicate a model where CD23 internalization results in ADAM10-dependent incorporation into exosomes, followed by partial cleavage of CD23 by ADAM10 prior to being released from the cell. The increased cleavage at endosomal pH also has implications for other ADAM10 substrates.The cleavage of plasma membrane-bound proteins and their subsequent release from the cell is described as ectodomain shedding. Historically this was believed to be a cell surface process. However, recently a second mechanism was discovered, involving the release of proteins after their internalization and subsequent trafficking through the endosomal pathway to multivesicular bodies (MVB) 2 (1). In the MVB, the proteins can interact with their sheddase and be cleaved and released or be sorted into exosomes and released as full-length proteins. Exosomes are small, 30 -100 nm, membrane containing vesicles that are released by a wide variety of cells. As they have high expression of major histocompatibility complexes (MHC) molecules they have been investigated as cell-free transporters of cancer antigens in cancer immunotherapy treatments (2). Exosome released from B cells have also been extensively studied. B cell-derived exosomes express integrins (3), which allow them to bind, and potentially present antigen to T cells. They also transport antigens, including peptides derived from allergen between antigen presenting cells (reviewed in Ref. 4). B cell exosome secretion is also increased upon activation (5) and interaction with T cells (6). However, the secretion of exosomes by cells as well as the mechanism that controls the sorting of proteins into exosomes are still poorly understood. The low-affinity receptor of immunoglobulin E (CD23) is an important regulator of IgE pro...

Immune Cell-derived Vesicles: Modulators and Mediators of Inflammation

“…The low levels of lysosomal proteases present in B cells may assure that the antigen is poorly degraded during the sorting process into secreted vesicles. Second, the low affinity receptor for IgE (CD23) found in B cell-derived vesicles [65] could mediate IgE-immune complex binding for shuttling antigen to other APC.…”

Pericellular proteolysis in cancer

“…Khokha and colleagues (Shimoda et al 2014) recently demonstrated that fibroblasts engineered to be deficient for all four Timp genes functionally recapitulate cancer-associated fibroblasts (CAFs) and activate oncogenic signaling in cancer cells through ADAM10-rich exosomes, which lead to increased cancer cell motility. Finally, proteolysis has also been shown to occur within exosomes in the case of ADAM10, which rapidly cleaves the immunoglobulin E (IgE) receptor CD23 prior to its release from B cells (Mathews et al 2010).…”

ADAM10 is required for SCF-induced mast cell migration

“…ADAM10, effective in site 2 cleavage in Notch1 and 2 signaling, and is important for T cell and marginal zone B cell development [15; 19; 24]. Several papers have shown ADAM10 to regulate B cell Ig production, including its role as the primary sheddase of CD23, a negative regulator of IgE production [17; 19; 23; 26]. Important to our work, Mathews et al showed that B cell-restricted ADAM10 deficiency decreased antigen-specific IgE, eosinophila and IL-5 production in an IgE/mast cell-dependent airway hyperresponsiveness (AHR) model [22].…”

“…However, ADAM10 cleaves many substrates involved in immunity, including Notch 1, Notch 2, DL1, CD23, TNF, and IL-6R [14; 16; 17; 18; 19; 20; 21; 22; 23; 24; 25; 26]. ADAM10-dependent Notch signaling has been demonstrated in recent years to be crucially important for the proper differentiation and distribution of T cells and B cells [17; 19; 24].…”