Reglucosylation of N-linked glycans is critical for calnexin assembly with T cell receptor (TCR) alpha proteins but not TCRbeta proteins.

Association of calnexin with newly synthesized glycoproteins involves recognition of monoglucosylated glycans, generated in the endoplasmic reticulum via initial removal of two glucose (Glc) residues from immature glycan chains by glucosidase enzymes (Glc trimming), or addition of a single Glc residue to fully trimmed glycans by glucosyltransferase enzymes (reglucosylation). While it has been established that creation of monoglucosylated glycans is important for chaperone binding, it is unknown if most proteins require both deglucosylation and reglucosylation for calnexin assembly or if initial Glc trimming is sufficient. Here, we studied the deglucosylation and reglucosylation of two related glycoproteins, the alpha and beta subunits of the T cell receptor (TCR) complex, and their assembly with calnexin in BW thymoma cells. Our data demonstrate that TCRalpha/beta glycoproteins undergo multiple cycles of Glc removal and addition within the endoplasmic reticulum and that numerous reglucosylated proteins assemble with calnexin, including TCRalpha/beta glycoproteins. Importantly, the current study shows that TCRbeta proteins, but not TCRalpha proteins, effectively associate with calnexin under conditions of functional Glc trimming but impaired reglucosylation. These data demonstrate that reglucosylated proteins associate with lectin-like chaperones in vivo and provide evidence that reglucosylation is of differential importance for the association of individual, indeed similar, glycoproteins with calnexin.