title: Endoplasmic reticulum retention, degradation, and aggregation of olfactory G-protein coupled receptors.
abstract: The mammalian olfactory G-protein coupled receptor family is comprised of hundreds of proteins that mediate odorant binding and initiate signal transduction cascades leading to the sensation of smell. However, efforts to functionally express olfactory receptors and identify specific odorant ligand-olfactory receptor interactions have been severely impeded by poor olfactory receptor surface expression in heterologous systems. Therefore, experiments were performed to elucidate the cellular mechanism(s) responsible for inefficient olfactory receptor cell surface expression. We determined that the mouse odorant receptors mI7 and mOREG are not selected for export from the ER and therefore are not detectable at the Golgi apparatus or plasma membrane. Specifically, olfactory receptors interact with the ER chaperone calnexin, are excluded from ER export sites, do not accumulate in ER-Golgi transport intermediates at 15 degrees C, and contain endoglycosidase H-sensitive oligosaccharides, consistent with olfactory receptor exclusion from post-ER compartments. A labile pool of ER-retained olfactory receptors are post-translationally modified by polyubiquitination and targeted for degradation by the proteasome. In addition, olfactory receptors are sequestered into ER aggregates that are degraded by autophagy. Collectively, these data demonstrate that poor surface expression of olfactory receptors in heterologous cells is attributable to a combination of ER retention due to inefficient folding and poor coupling to ER export machinery, aggregation, and degradation via both proteasomal and autophagic pathways.
authors.affiliation: Senomyx, Inc., 11099 North Torrey Pines Road, La Jolla, CA 92037, USA.
authors.forename: Min
authors.initials: M
authors.lastname: Lu
authors.forename: Fernando
authors.initials: F
authors.lastname: Echeverri
authors.forename: Bryan D
authors.initials: BD
authors.lastname: Moyer
mesh_terms: D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases / Q000378:metabolism; D017931:DNA Primers; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D044385:GTP-Binding Protein alpha Subunits; D006056:Golgi Apparatus / Q000378:metabolism; D020962:Heterotrimeric GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007668:Kidney / Q000502:physiology; D051379:Mice; D008954:Models, Biological; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex; D021381:Protein Transport; D043562:Receptors, G-Protein-Coupled / Q000737:chemistry* / Q000378:metabolism*; D018035:Receptors, Odorant / Q000737:chemistry* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D017931:DNA Primers; D044385:GTP-Binding Protein alpha Subunits; D009097:Multienzyme Complexes; D043562:Receptors, G-Protein-Coupled; D018035:Receptors, Odorant; D011994:Recombinant Proteins; C084618:olfactory G protein subunit alpha olf; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex; D020962:Heterotrimeric GTP-Binding Proteins