PERSISTENCE OF GLUCOSE RESIDUES ON CORE OLIGOSACCHARIDES PREVENTS ASSOCIATION OF TCR-ALPHA AND TCR-BETA PROTEINS WITH CALNEXIN AND RESULTS SPECIFICALLY IN ACCELERATED DEGRADATION OF NASCENT TCR-ALPHA PROTEINS WITHIN THE ENDOPLASMIC-RETICULUM

The alpha beta T-cell antigen receptor (TCR) is a multisubunit transmembrane complex composed of at least six different proteins (alpha, beta, gamma, delta, epsilon and zeta) that are assembled in the endoplasmic reticulum (ER). In this report we have examined the role of oligosaccharide processing on survival and assembly of nascent TCR proteins within the ER and their associations with molecular chaperone proteins important in TCR assembly. We found that treatment of BW5147 T cells with the glucosidase inhibitor castanospermine resulted in markedly accelerated degradation of nascent TCR alpha proteins with a half-life of similar to 20 min. Accelerated degradation was unique to TCR alpha proteins, as the stability of nascent TCR beta and CD3 gamma,epsilon chains was unaltered. Consistent with a requirement for glucose (Glc) trimming for survival of nascent TCR alpha proteins within the ER, we found that newly synthesized TCR alpha chains were innately unstable in the glucosidase II-deficient BW5147 mutant cell line PHAR2.7. In addition to destabilizing nascent TCR alpha proteins we found that persistence of Glc residues on core oligosaccharides markedly interfered with association of both TCR alpha and TCR beta glycoproteins with the molecular chaperone calnexin. Finally, using 2B4 T hybridoma cells in which TCR complexes are efficiently assembled, we found that rapid degradation of nascent TCR alpha proteins induced by impaired Glc trimming severely limits assembly of TCR alpha proteins with TCR beta proteins. These results demonstrate that the persistence of Glc residues on asparagine-linked (N-linked) oligosaccharide side chains (i) disrupts associations of calnexin with both TCR alpha and TCR beta proteins, (ii) results in destabilization of nascent TCR alpha proteins, and (iii) restricts the ability of TCR alpha and TCR beta proteins to assemble together. Thus, association with calnexin is uniquely important for the stabilization of nascent TCR alpha proteins within the ER.