title: A glycosylated type I membrane protein becomes cytosolic when peptide: N-glycanase is compromised.
abstract: The human cytomegalovirus-encoded glycoprotein US2 catalyzes proteasomal degradation of Class I major histocompatibility complex (MHC) heavy chains (HCs) through dislocation of the latter from the endoplasmic reticulum (ER) to the cytosol. During this process, the Class I MHC HCs are deglycosylated by an N-glycanase-type activity. siRNA molecules designed to inhibit the expression of the light chain, beta(2)-microglobulin, block the dislocation of Class I MHC molecules, which implies that US2-dependent dislocation utilizes correctly folded Class I MHC molecules as a substrate. Here we demonstrate it is peptide: N-glycanase (PNGase or PNG1) that deglycosylates dislocated Class I MHC HCs. Reduction of PNGase activity by siRNA expression in US2-expressing cells inhibits deglycosylation of Class I MHC HC molecules. In PNGase siRNA-treated cells, glycosylated HCs appear in the cytosol, providing the first evidence for the presence of an intact N-linked type I membrane glycoprotein in the cytosol. N-glycanase activity is therefore not required for dislocation of glycosylated Class I MHC molecules from the ER.
authors.affiliation: Department of Pathology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Daniël
authors.initials: D
authors.lastname: Blom
authors.forename: Christian
authors.initials: C
authors.lastname: Hirsch
authors.forename: Patrick
authors.initials: P
authors.lastname: Stern
authors.forename: Domenico
authors.initials: D
authors.lastname: Tortorella
authors.forename: Hidde L
authors.initials: HL
authors.lastname: Ploegh
mesh_terms: D045744:Cell Line, Tumor; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006031:Glycosylation; D015395:Histocompatibility Antigens Class I / Q000378:metabolism*; D006801:Humans; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000378:metabolism*; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D014759:Viral Envelope Proteins / Q000378:metabolism*; D001613:beta 2-Microglobulin / Q000235:genetics / Q000378:metabolism*
chemical_list: D015395:Histocompatibility Antigens Class I; D034741:RNA, Small Interfering; C118670:US2 protein, Varicellovirus; D014759:Viral Envelope Proteins; D001613:beta 2-Microglobulin; D046988:Proteasome Endopeptidase Complex; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase