Oligosaccharide Modification in the Early Secretory Pathway Directs the Selection of a Misfolded Glycoprotein for Degradation by the Proteasome*
- From the Departments of ‡Pathology and§Cell Biology, ¶Cell and Molecular Biology Program, Baylor College of Medicine, Houston, Texas 77030
Abstract
The role of conformation-based quality control in the early secretory pathway is to eliminate misfolded polypeptides and unassembled multimeric protein complexes from the endoplasmic reticulum, ensuring the deployment of only functional molecules to distal sites. The intracellular fate of terminally misfolded human α1-antitrypsin was examined in hepatoma cells to identify the functional role of asparagine-linked oligosaccharide modification in the selection of glycoproteins for degradation by the cytosolic proteasome. Proteasomal degradation required physical interaction with the molecular chaperone calnexin. Altered sedimentation of intracellular complexes following treatment with the specific proteasome inhibitor lactacystin, and in combination with mannosidase inhibition, revealed that the removal of mannose from attached oligosaccharides abrogates the release of misfolded α1-antitrypsin from calnexin prior to proteasomal degradation. Intracellular turnover was arrested with kifunensine, implicating the participation of endoplasmic reticulum mannosidase I in the disposal process. Accelerated degradation occurred in a mannosidase-independent manner and was arrested by lactacystin, in response to the posttranslational inhibition of glucosidase II, demonstrating that the attenuated removal of glucose from attached oligosaccharides functions as the underlying rate-limiting step in the proteasome-mediated pathway. A model is proposed in which the removal of mannose from multiple attached oligosaccharides directs calnexin in the selection of misfolded α1-antitrypsin for degradation by the proteasome.
Footnotes
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↵* This work was supported in part by an American Lung Association research career investigator award, by an American Heart Association grant-in-aid, by the Moran Foundation (to R. N. S), and by an American Lung Association research training fellowship (to P. C.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‖ To whom correspondence should be addressed: Dept. of Pathology, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030. Tel.: 713-798-3169; Fax: 713-798-5838; E-mail:rsifers{at}bcm.tmc.edu.
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↵2 P. Choudhury, unpublished observations.
- Abbreviations:
- ER
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endoplasmic reticulum
- AAT
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α1-antitrypsin
- PAGE
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polyacrylamide gel electrophoresis
- UGTR
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UDP-glucose:glycoprotein glucosyltransferase
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- Received October 13, 1998.
- Revision received December 8, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
