A stringent quality control process selects misfolded polypeptides generated in the endoplasmic reticulum (ER) for ER-associated degradation (ERAD). Here we assessed the maintenance of efficient glycoprotein folding in cells with defective ERAD caused by lack of adaptation of the intralumenal level of ER degradationenhancing ␣-mannosidase-like protein (EDEM) to an increase in the ER cargo load. When these cells were converted into factories for production of high levels of human -secretase, maturation of this N-glycosylated aspartic protease progressed as in wild-type cells initially to gradually become less efficient. Up-regulation of EDEM to strengthen the ERAD machinery (but not up-regulation of calnexin to reinforce the folding machinery) was instrumental in maintaining folding efficiency and secretory capacity. Our data underscore the important role that the degradation machinery plays in maintaining a functional folding environment in the ER.Only a fraction of the polypeptide chains expressed in the ER 1 is transported to its final destination. A stringent quality control process prevents export of orphan subunits of oligomeric complexes, folding-incompetent products of mutated genes, and by-products of protein biosynthesis that do not acquire the native conformation (1-3). Polypeptides that do not pass quality control are either retained by the ER chaperone network to be used for further folding attempts or are sorted out for cytosolic degradation. The mechanisms regulating ER quality control are better understood for N-glycosylated polypeptides. N-glycans are added to asparagines in nascent chains as pre-assembled tri-antennary oligosaccharides composed of three terminal glucoses, nine mannoses, and two Nacetylglucosamines. The two outermost glucoses are rapidly trimmed by ER glucosidases I and II. Polypeptides exposing mono-glucosylated intermediates of N-glycan trimming associate with the ER lectins calnexin (Cnx) and calreticulin (Crt).The opposing actions of glucosidase II that trim the innermost glucose residue (thereby releasing the newly synthesized polypeptide from Cnx/Crt) and of UDP-glucose-glycoprotein glucosyltransferase that adds back one glucose to non-native conformers sending them back to Cnx/Crt, drive the so-called Cnx cycle (1, 2). For folding-incompetent and terminally misfolded glycopolypeptides, termination of unproductive folding attempts coincides with their definitive release from the Cnx cycle and deviation into the ERAD machinery. Interruption of unproductive folding attempts is a crucial step of ER quality control because it prevents clogging of the ER chaperone system.Emerging evidence underscores the importance of a tightly controlled extraction of folding-incompetent glycopolypeptides from the Cnx cycle to promote their degradation. Extraction is regulated by the ER ␣-mannosidase I (4) and by EDEM (ER degradation enhancing ␣-mannosidase-like protein (5)), an enzymatically inactive mannosidase-like protein. The ER ␣-mannosidase I cleaves one (6) or more (7) mannose residues...