Oligosaccharyltransferase Is Involved in Targeting to ER-Associated Degradation

Most membrane and secretory proteins undergo N-glycosylation, catalyzed by oligosaccharyltransferase (OST), a membrane-bound complex in the endoplasmic reticulum (ER). Proteins failing quality control are degraded via ER-associated degradation (ERAD), involving retrotranslocation to cytosolic proteasomes, or relegated to ER subdomains and eliminated via ER-phagy. Using stable isotope labeling by amino acids in cell culture (SILAC) proteomics, we identified OST subunits as differential key interactors with a misfolded ER protein bait upon proteasomal inhibition, suggesting unexpected involvement in ERAD. Previous reports implied additional roles for OST subunits beyond N-glycosylation, such as quality control by ribophorin I. We tested OST engagement in glycoprotein and non-glycosylated protein ERAD; overexpression or partial knockdown of OST subunits interfered with ERAD in conditions that did not affect glycosylation. We studied the effects on model misfolded type I and II membrane-bound proteins, BACE476 and asialoglycoprotein receptor H2a, respectively, and on a soluble luminal misfolded glycoprotein, α1-antitrypsin NHK variant. OST subunits appear to participate in late ERAD stages, interacting with the E3 ligase HRD1 and facilitating retrotranslocation. Molecular dynamics simulations suggest membrane thinning by OST transmembrane domains, possibly assisting retrotranslocation via membrane distortion.