University of Oulu

Lackman, J. J., Markkanen, P. M. H., Hogue, M., Bouvier, M., & Petäjä-Repo, U. E. (2014). N-Glycan-dependent and -independent Quality Control of Human δ Opioid Receptor N-terminal Variants. Journal of Biological Chemistry, 289(25), 17830–17842. https://doi.org/10.1074/jbc.m114.566273

N-glycan-dependent and -independent quality control of human δ opioid receptor N-terminal variants

Saved in:
Author: Lackman, Jarkko J.1; Markkanen, Piia M. H.1; Hogue, Mireille2;
Organizations: 1Department of Anatomy and Cell Biology and the Medical Research Center Oulu, Institute of Biomedicine, University of Oulu, FI-90014 Oulu, Finland
2Department of Biochemistry, Institute for Research in Immunology and Cancer and Groupe de Recherche Universitaire sur le Médicament, Université de Montréal, Montréal, Québec H3C 3J7, Canada
Format: article
Version: published version
Access: open
Online Access: PDF Full Text (PDF, 1.7 MB)
Persistent link: http://urn.fi/urn:nbn:fi-fe2019120545771
Language: English
Published: American Society for Biochemistry and Molecular Biology, 2014
Publish Date: 2019-12-05
Description:

Abstract

Quality control (QC) in the endoplasmic reticulum (ER) scrutinizes newly synthesized proteins and directs them either to ER export or ER-associated degradation (ERAD). Here, we demonstrate that the human δ-opioid receptor (hδOR) is subjected to ERQC in both N-glycan-dependent and -independent manners. This was shown by investigating the biosynthesis and trafficking of wild-type and non-N-glycosylated F27C variants in metabolic pulse-chase assays coupled with flow cytometry and cell surface biotinylation. Both QC mechanisms distinguished the minute one-amino acid difference between the variants, targeting a large fraction of hδOR-Cys²⁷ to ERAD. However, the N-glycan-independent QC was unable to compensate the N-glycan-dependent pathway, and some incompletely folded non-N-glycosylated hδOR-Cys²⁷ reached the cell surface in conformation incompatible with ligand binding. The turnover of receptors associating with the molecular chaperone calnexin (CNX) was significantly slower for the hδOR-Cys²⁷, pointing to an important role of CNX in the hδOR N-glycan-dependent QC. This was further supported by the fact that inhibiting the co-translational interaction of hδOR-Cys²⁷ precursors with CNX led to their ERAD. Opioid receptor pharmacological chaperones released the CNX-bound receptors to ER export and, furthermore, were able to rescue the Cys²⁷ variant from polyubiquitination and retrotranslocation to the cytosol whether carrying N-glycans or not. Taken together, the hδOR appears to rely primarily on the CNX-mediated N-glycan-dependent QC that has the capacity to assist in folding, whereas the N-glycan-independent mechanism constitutes an alternative, although less accurate, system for directing misfolded/incompletely folded receptors to ERAD, possibly in altered cellular conditions.

see all

Series: Journal of biological chemistry
ISSN: 0021-9258
ISSN-E: 1083-351X
ISSN-L: 0021-9258
Volume: 289
Issue: 25
Pages: 17830 - 17842
DOI: 10.1074/jbc.M114.566273
OADOI: https://oadoi.org/10.1074/jbc.M114.566273
Type of Publication: A1 Journal article – refereed
Field of Science: 1182 Biochemistry, cell and molecular biology
Subjects:
Funding: This work was supported by the Sigrid Jusélius Foundation, the Medical Research Center Oulu and Grant 127199 from the Academy of Finland (to U. E. P.-R.). Jarkko J. Lackman was supported by the Finnish Cultural Foundation.
Academy of Finland Grant Number: 127199
Detailed Information: 127199 (Academy of Finland Funding decision)
Copyright information: This research was originally published in the Journal of Biological Chemistry. Lackman, J. J., Markkanen, P. M. H., Hogue, M., Bouvier, M., & Petäjä-Repo, U. E.. N-Glycan-dependent and -independent Quality Control of Human δ Opioid Receptor N-terminal Variants. J. Biol. Chem. 2012; 289:17830-17842. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.