N-linked oligosaccharide processing, but not association with calnexin/calreticulin is highly correlated with endoplasmic reticulum-associated degradation of antithrombin Glu313-deleted mutant.
Previously we showed that two antithrombin mutants were degraded through an endoplasmic reticulum (ER)-associated degradation (ERAD) pathway [F. Tokunaga et al., FEBS Lett. 412 (1997) 65]. Here, we examined the combined effects of inhibitors of glycosidases, protein synthesis, proteasome, and tyrosine phosphatase on ERAD of a Glu313-deleted (DeltaGlu) mutant of antithrombin. We found that kifunensine, an ER mannosidase I inhibitor, suppressed ERAD, indicating that specific mannose trimming plays a critical role. Cycloheximide and puromycin, inhibitors of protein synthesis, also suppressed ERAD, the effects being cancelled by pretreatment with castanospermine. In contrast, kifunensine suppressed ERAD even in castanospermine-treated cells, suggesting that suppression of ERAD does not always require the binding of lectin-like ER chaperones-like calnexin and/or calreticulin. These results indicate that, besides proteasome inhibitors, inhibitors of ER mannosidase I and protein synthesis suppress ERAD of the antithrombin deltaGlu mutant at different stages, and processing of N-linked oligosaccharides highly correlated with the efficiency of ERAD.
Tokunaga
Fuminori
F
Department of Life Science, Graduate School of Science, Himeji Institute of Technology, Harima Science Garden City, Hyogo 678-1297, Japan.
Hara
Kazuya
K
Koide
Takehiko
T
D017485:1-Deoxynojirimycin / Q000494:pharmacology; D000470:Alkaloids / Q000494:pharmacology; D000818:Animals; D000991:Antithrombins / Q000235:genetics / Q000378:metabolism*; D037281:Calnexin / Q000378:metabolism*; D037282:Calreticulin / Q000378:metabolism*; D002236:Carbohydrate Conformation; D002478:Cells, Cultured; D006224:Cricetinae; D003513:Cycloheximide / Q000494:pharmacology; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005959:Glucosidases / Q000037:antagonists & inhibitors; D007212:Indolizines / Q000494:pharmacology; D008358:Mannose / Q000378:metabolism; D008361:Mannosidases / Q000037:antagonists & inhibitors; D009154:Mutation; D009844:Oligosaccharides / Q000378:metabolism*; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011691:Puromycin / Q000494:pharmacology; D054836:Indolizidines
D017485:1-Deoxynojirimycin; D000470:Alkaloids; D000991:Antithrombins; D037281:Calnexin; D037282:Calreticulin; D003513:Cycloheximide; D004791:Enzyme Inhibitors; D005959:Glucosidases; D007212:Indolizines; D008358:Mannose; D008361:Mannosidases; D009844:Oligosaccharides; D011500:Protein Synthesis Inhibitors; D017027:Protein Tyrosine Phosphatases; D011691:Puromycin; C065629:kifunensine; C046090:mannosyl-oligosaccharide 1,2-alpha-mannosidase; C037806:castanospermine; D054836:Indolizidines