Cooperativity and Segregation of Function within the Ig-α/β Heterodimer of the B Cell Antigen Receptor Complex

Abstract

A B cell's response to antigen, whether it be proliferation, differentiation, anergy, or deletion, is dependent upon recognition of that antigen by the B cell antigen receptor (BCR) 1 (1-3). The receptor is a multimeric complex consisting of the antigenrecognition substructure, membrane-bound immunoglobulin non-covalently associated with heterodimer(s) of Ig-␣ and Ig-␤ (4 -6). Present evidence indicates that the cytoplasmic tails of Ig-␣ and ␤ (7) translate antigen engagement into cytoplasmic signaling events that initiate cellular responses (8 -12). Most proximally in the signaling cascade, one or more tyrosine kinases, including Syk and members of the Src family, are activated (13-15). These in turn activate a variety of pathways whose constituents include Ras, phosphatidylinositol 3-kinase, and phospholipase C (1). Embedded within the cytoplasmic tails of both Ig-␣ and ␤ is a sequence common to other multichain immune recognition receptor (MIRR) subunits including CD3␦, CD3␥, TCR, Fc␥RIII␥, and Fc⑀RI␥, termed the immunoreceptor tyrosine-based activation motif (ITAM) (16,17). The motif contains two tyrosines, both of which are critical for initiating tyrosine kinase activation (10, 18). Phosphorylation of these tyrosines facilitates the recruitment and activation of tyrosine kinases which contain SH2 domains, such as . Substrates for these kinases may also be recruited (22). The presence of the ITAM in all MIRR chains involved in signal transduction has led some to suggest that apparently heterologous chains such as CD3⑀ and TCR are functionally redundant and the presence of multiple ITAMs within each MIRR serve to increase the strength of signal which can be generated via the receptor. Evidence for this assertion has been obtained in studies of both the B and T cell antigen receptors (8,12,23,24). In contrast, we and others have provided evidence indicating that each heterologous ITAM containing chain has a distinct function (10,11,19,25,26).Many of the above studies utilized chimeras in which irrelevant extracellular and transmembrane domains were fused to the single cytoplasmic domain under study (18,27,28). Although this approach has yielded considerable insight into ITAM-containing chains, it assumes that functions observed in the isolated circumstance of a single chimera are reflective of the function of that cytoplasmic domain within the intact receptor complex. This might not be true since most ITAM-containing chains, such as Ig-␣ and Ig-␤, are expressed on cell surfaces as heterodimers (29 -32). Therefore, we postulated that within the context of a heterodimer, Ig-␣ and Ig-␤ would have new, complementary or even synergistic functions, not predicted from studies of single chain chimeras.As demonstrated in this report, Ig-␣ and ␤ have new and unpredicted functions in the context of a heterodimer. Using a novel chimera system, which allowed us to form either heteroor homodimers of the cytoplasmic domains of Ig-␣ and Ig-␤, we observed that when Ig-␤ is ligated independently it is able to activate tyrosine...

B Cell Antigen Receptor Signaling and Internalization Are Mutually Exclusive Events

“…Surprisingly, there has been no systematic examination of the role of the Igα/Igβ cytosolic tyrosines in BCR internalization. Therefore, we undertook such an analysis with a focus on Igα because it is the main signaling chain of the BCR [ 46 , 47 ], and it has been implicated in constitutive receptor internalization [ 44 ].…”

“…To simplify our initial analysis, we chose to examine the cytosolic tail of Igα in isolation as a chimera with the extracellular and transmembrane domains of human platelet derived growth factor receptor (PDGFR) [ 47 , 48 ]. For these experiments, we derived chimeras containing several Igα cytosolic mutants.…”

Cis and Trans Regulatory Mechanisms Control AP2-Mediated B Cell Receptor Endocytosis via Select Tyrosine-Based Motifs

“…Thus, the mechanism of trans regulation of BCR signaling appears to be completely distinct from that of cis regulation and likely involves crosstalk between the two cytoplasmic domains (Figure 9D). This general idea is not completely new, as it has been suggested that there is crosstalk between the two cytoplasmic domains of the BCR during signaling [29]. However, this is the first time trans regulation has been shown to control receptor internalization.…”

“…However, in vivo analysis of BCR internalization clearly demonstrates a role for the CD79b ITAM tyrosines in both constitutive and inducible BCR endocytosis [28]. Moreover, while CD79b is not internalization competent as a monomeric construct, artificial pairing of two CD79b cytoplasmic domains [24], [29]–[31] or generation of select mutations within CD79b [24] leads to receptor endocytosis with kinetics remarkably similar to wild-type BCR. Finally, a recent study of B cell lymphoma suggests a role for the membrane proximal ITAM tyrosine in CD79b, Y195, in influencing BCR surface expression levels [32].…”

Recruitment of Cbl-b to B Cell Antigen Receptor Couples Antigen Recognition to Toll-Like Receptor 9 Activation in Late Endosomes

“…Lysates or immunoprecipitates were resolved on a 4–15% Mini-Protean TGX gel (Bio-Rad) and transferred onto Immun-Blot PVDF membrane (Bio-Rad). Membranes were probed with antibodies specific for ubiquitin (P4D1, Santa Cruz), Igβ [24] or Cbl-b.…”