Oligosaccharyltransferase is involved in targeting to ER-associated degradation

Abstract

AbstractThe majority of membrane and secretory proteins undergo N-glycosylation, a process catalyzed by oligosaccharyltransferase (OST), a membrane-bound protein complex that associates with the translocation channels within the endoplasmic reticulum (ER). Proteins failing quality control undergo ER-associated degradation (ERAD) by retrotranslocation to cytosolic proteasomes. Using a proteomics approach, we unexpectedly identified several OST subunits as significant interactors of a misfolded ER protein bait. Previous reports have suggested other roles for OST in addition to N-glycosylation, such as participation of the OST subunit ribophorin I in quality control. Our findings demonstrate OST engagement in ERAD of glycoproteins and non-glycosylated proteins, both affected by OST subunit overexpression and partial knockdown, which 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 appears to be involved in late ERAD stages, interacting with the E3 ligase HRD1 and impacting retrotranslocation. We discuss the possibility that OST, harboring multiple transmembrane domains, might assist retrotranslocation by contributing to membrane distortion for protein dislocation.Significance statementAbout thirty percent of cellular proteins undergo processing through the endoplasmic reticulum (ER), with most being glycosylated on asparagine residues in a process called N-glycosylation. In this process, oligosaccharyltransferase (OST), an ER membrane-bound protein complex, transfers sugar chains to the protein. Here we elucidate an additional role of OST, in targeting misfolded proteins to ER-associated degradation (ERAD), a process that requires their transport from the ER to the cytosol. Employing a mass spectrometry proteomics approach, we observed differential binding of OST subunits to accumulated misfolded protein molecules following inhibition of their degradation. Interference with OST activity in conditions where its function in N-glycosylation was not affected, resulted in ERAD inhibition.ClassificationBiological Sciences, Cell Biology