UGGT1/2-mediated reglucosylation of N-glycan competes with ER-associated degradation of unstable and misfolded glycoproteins

Here we investigated how the fate (folding versus degradation) of glycoproteins is determined in the endoplasmic reticulum (ER). Monoglucosylated glycoproteins are recognized by lectin chaperones to facilitate their folding, whereas glycoproteins with well-trimmed mannoses are subjected to glycoprotein ER-associated degradation (gpERAD). Previously we elucidated how mannoses are trimmed by EDEM family members ([George et al., 2020][1], [2021][2] eLife). Though reglucosylation by UGGTs (UGGT1 and UGGT2) was reported to have no effect on substrate degradation, here, we directly test this using genetically disrupted UGGTs. Strikingly, the results showed that UGGTs (mainly UGGT1) delayed the degradation of misfolded substrates and unstable glycoproteins including ATF6α. An experiment with a point mutant of UGGT1 indicated that the glucosylation activity of UGGT was required for the inhibition of early glycoprotein degradation. We further demonstrated the physiological importance of UGGT, since ATF6 cannot function properly without UGGTs. The fate of glycoproteins is determined by a tug-of-war between structure formation by UGGTs and degradation by EDEMs. Thus, our work strongly suggests that UGGTs are central factors in ER protein quality control via regulation of both glycoprotein folding and degradation. ### Competing Interest Statement The authors have declared no competing interest. * UGGT1 : UDP-glucose glycoprotein glucosyltransferase 1 ER : endoplasmic reticulum CNX : Calnexin CRT : Calreticulin gpERAD : glycoprotein ER-associated degradation non-gpERAD : non-glycoprotein ER-associated degradation EDEM : ER degradation enhancing αmannosidase-like protein ATF6 : Activating transcription factor 6 ATF4 : Activating transcription factor 4 PDI : Protein disulfide isomerase GlcNAc : N -acetylglucosamines TXNDC11 : Thioredoxin domain-containing protein 11 WT : wild type DKO : double knockout TKO : triple knockout Puro : puromycin Hygro : hygromycin BiP : immunoglobulin heavy chain binding protein EMC1 : ER membrane protein complex subunit 1 EndoH : endoglycosidase H DNJ : 1-deoxynojirimycin CHX : cycloheximide ERSE : ER stress responsive element UPRE : unfolded protein response element CRISPR : Clustered regularly interspaced short palindromic repeats TALEN : Transcription activator-like effector nuclease BLAST : Basic Local Alignment Search Tool PQQ : pyrrolo-quinoline quinone Tg : thapsigargin XBP1 : X-box binding protein 1 PERK : Protein kinase R (PKR)-like endoplasmic reticulum kinase IRE1 : Inositol-requiring enzyme 1 GAPDH : Glyceraldehyde-3-phosphate dehydrogenase HA : Hemagglutinin A1AT : Alpha-1 antitrypsin NHK : null hong kong EPO : Erythropoietin SDS : sodium dodecyl sulfate [1]: #ref-9 [2]: #ref-8