Endoplasmic reticulum (ER)-associated degradation of misfolded N-linked glycoproteins is suppressed upon inhibition of ER mannosidase I.
To examine the role of early carbohydrate recognition/trimming reactions in targeting endoplasmic reticulum (ER)-retained, misfolded glycoproteins for ER-associated degradation (ERAD), we have stably expressed the cog thyroglobulin (Tg) mutant cDNA in Chinese hamster ovary cells. We found that inhibitors of ER mannosidase I (but not other glycosidases) acutely suppressed Cog Tg degradation and also perturbed the ERAD process for Tg reduced with dithiothreitol as well as for gamma-carboxylation-deficient protein C expressed in warfarin-treated baby hamster kidney cells. Kifunensine inhibition of ER mannosidase I also suppressed ERAD in castanospermine-treated cells; thus, suppression of ERAD does not require lectin-like binding of ER chaperones calnexin and calreticulin to monoglucosylated oligosaccharides. Notably, the undegraded protein fraction remained completely microsome-associated. In pulse-chase studies, kifunensine-sensitive degradation was still inhibitable even 1 h after Tg synthesis. Intriguingly, chronic treatment with kifunensine caused a 3-fold accumulation of Cog Tg in Chinese hamster ovary cells and did not lead to significant induction of the ER unfolded protein response. We hypothesize that, in a manner not requiring lectin-like activity of calnexin/calreticulin, the recognition or processing of a specific branched N-linked mannose structure enhances the efficiency of glycoprotein retrotranslocation from the ER lumen.
Tokunaga
F
F
Department of Life Science, Himeji Institute of Technology, Harima Science Garden City, Hyogo 678-1277, Japan.
Brostrom
C
C
Koide
T
T
Arvan
P
P
D000470:Alkaloids / Q000494:pharmacology*; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D008361:Mannosidases / Q000037:antagonists & inhibitors / Q000502:physiology*; D011486:Protein C / Q000378:metabolism; D017510:Protein Folding; D013954:Thyroglobulin / Q000378:metabolism*
D000470:Alkaloids; D004791:Enzyme Inhibitors; D006023:Glycoproteins; D011486:Protein C; C065629:kifunensine; D013954:Thyroglobulin; D008361:Mannosidases; C046090:mannosyl-oligosaccharide 1,2-alpha-mannosidase