Enhancement of endoplasmic reticulum (ER) degradation of misfolded null Hong Kong α1-antitrypsin by human ER mannosidase I

Misfolded glycoproteins synthesized in the endoplasmic reticulum (ER) are degraded by cytoplasmic proteasomes, a mechanism known as ERAD (ER-associated degradation). In the present study, we demonstrate that ERAD of the misfolded genetic variant-null Hong Kong alpha(1)-antitrypsin is enhanced by overexpression of the ER processing alpha1,2-mannosidase (ER ManI) in HEK 293 cells, indicating the importance of ER ManI in glycoprotein quality control. We showed previously that EDEM, an enzymatically inactive mannosidase homolog, interacts with misfolded alpha1-antitrypsin and accelerates its degradation (Hosokawa, N., Wada, I., Hasegawa, K., Yorihuzi, T., Tremblay, L. O., Herscovics, A., and Nagata, K. (2001) EMBO Rep. 2, 415-422). Herein we demonstrate a combined effect of ER ManI and EDEM on ERAD of misfolded alpha(1)-antitrypsin. We also show that misfolded alpha(1)-antitrypsin NHK contains labeled Glc(1)Man(9)GlcNAc and Man(5-9)GlcNAc released by endo-beta-N-acetylglucosaminidase H in pulse-chase experiments with [2-H-3] mannose. Overexpression of ER ManI greatly increases the formation of Man(8)GlcNAc, induces the formation of Glc(1)Man(8)GlcNAc and increases trimming to Man(5-7)GlcNAc. We propose a model whereby the misfolded glycoprotein interacts with ER ManI and with EDEM, before being recognized by downstream ERAD components. This detailed characterization of oligosaccharides associated with a misfolded glycoprotein raises the possibility that the carbohydrate recognition determinant triggering ERAD may not be restricted to Man(8)GlcNAc(2) isomer B as previous studies have suggested.