| The B cell antigen receptor (BCR) relays extracellular signals and delivers antigen to the endocytic compartment for processing and presentation. Concurrently, BCR ligation destabilizes Ig-alpha/Ig-beta from mu-heavy chain (mum). Here, we describe that receptor destabilization represents a physical separation of mum from Ig-alpha/Ig-beta. Sucrose gradient fractionation localized Ig-alpha/Ig-beta to GM1-containing lipid microdomains in the absence of mum. Confocal and transmission electron microscopy studies revealed colocalization of unsheathed mum with clathrin-coated vesicles (CCVs). Further, mum failed to associate with CCVs when receptor destabilization was inhibited, suggesting that unsheathing of mum is required for clathrin-mediated endocytosis. In summary, we found that antigen stimulation physically separates Ig-alpha/Ig-beta from mum, facilitating concomitant signal transduction and antigen delivery to the endocytic compartment. In addition, we demonstrate the molecular basis and the role of BCR destabilization in antigen processing. Dissociation of Ig-aalpha/Ig-beta from mum requires tyrosine-587 (Y587) of the mum transmembrane domain. Receptors expressing an Y587F mutation transduced signals that were comparable to wild type receptors, yet failed to dissociate mum from Ig-alpha/Ig-beta. Further, receptors harboring the Y587F mutation failed to associate with CCVs, resulting in diminished antigen in the lysosome-associated membrane protein-1 positive (LAMP-1 +) compartments and impaired antigen presentation. Thus, the transmembrane tyrosine of mum mediates destabilization of the BCR complex and the association of antigen with CCVs. Our data also indicate that association of receptors with CCVs is required for antigen presentation. Together, we demonstrate that BCR destabilization reveals a novel and prerequisite step prior to the targeting of antigen into the endocytic pathway. |
