Proteome and Glycoproteome Analyses Reveal Regulation of Protein Glycosylation Site-Specific Occupancy and Lysosomal Hydrolase Maturation by <i>N</i>-Glycan-Dependent ER-Quality Control

<i>N</i>-Glycan-dependent endoplasmic reticulum quality control (ERQC) primarily mediates protein folding, which determines the fate of the polypeptide. Monoglucose residues on <i>N</i>-glycans determine whether the nascent <i>N</i>-glycosylated proteins enter into and escape from the calnexin (CANX)/calreticulin (CALR) cycle, which is a central system of the ERQC. To reveal the impact of ERQC on glycosylation and protein fate, we performed comprehensive quantitative proteomic and glycoproteomic analyses using cells defective in <i>N</i>-glycan-dependent ERQC. Deficiency of MOGS encoding the ER α-glucosidase I, CANX, or/and CALR broadly affected protein expression and glycosylation. Among the altered glycoproteins, the occupancy of oligomannosidic <i>N</i>-glycans was significantly affected. Besides the expected ER stress, proteins and glycoproteins involved in pathways for lysosome and viral infection are differentially changed in those deficient cells. We demonstrated that lysosomal hydrolases were not correctly modified with mannose-6-phosphates on the <i>N</i>-glycans and were directly secreted to the culture medium in <i>N</i>-glycan-dependent ERQC mutant cells. Overall, the CANX/CALR cycle promotes the correct folding of glycosylated peptides and influences the transport of lysosomal hydrolases.