title: N-glycosylation of asparagine 8 regulates surface expression of major histocompatibility complex class I chain-related protein A (MICA) alleles dependent on threonine 24.
abstract: NKG2D is an activating receptor expressed on several types of human lymphocytes. NKG2D ligands can be induced upon cell stress and are frequently targeted post-translationally in infected or transformed cells to avoid immune recognition. Virus infection and inflammation alter protein N-glycosylation, and we have previously shown that changes in cellular N-glycosylation are involved in regulation of NKG2D ligand surface expression. The specific mode of regulation through N-glycosylation is, however, unknown. Here we investigated whether direct N-glycosylation of the NKG2D ligand MICA itself is critical for cell surface expression and sought to identify the essential residues. We found that a single N-glycosylation site (Asn(8)) was important for MICA018 surface expression. The frequently expressed MICA allele 008, with an altered transmembrane and intracellular domain, was not affected by mutation of this N-glycosylation site. Mutational analysis revealed that a single amino acid (Thr(24)) in the extracellular domain of MICA018 was essential for the N-glycosylation dependence, whereas the intracellular domain was not involved. The HHV7 immunoevasin, U21, was found to inhibit MICA018 surface expression by affecting N-glycosylation, and the retention was rescued by T24A substitution. Our study reveals N-glycosylation as an allele-specific regulatory mechanism important for regulation of surface expression of MICA018, and we pinpoint the residues essential for this N-glycosylation dependence. In addition, we show that this regulatory mechanism of MICA surface expression is likely targeted during different pathological conditions.
authors.affiliation: From the Laboratory of Immunology, Section for Experimental Animal Models, Faculty of Health and Medical Sciences, University of Copenhagen, DK-1870 Frederiksberg, Denmark and.
authors.forename: Maiken
authors.initials: M
authors.lastname: Mellergaard
authors.affiliation: From the Laboratory of Immunology, Section for Experimental Animal Models, Faculty of Health and Medical Sciences, University of Copenhagen, DK-1870 Frederiksberg, Denmark and.
authors.forename: Sarah Line
authors.initials: SL
authors.lastname: Skovbakke
authors.affiliation: the Department of Biology, Carroll University, Waukesha, Wisconsin 53186.
authors.forename: Christine L
authors.initials: CL
authors.lastname: Schneider
authors.affiliation: From the Laboratory of Immunology, Section for Experimental Animal Models, Faculty of Health and Medical Sciences, University of Copenhagen, DK-1870 Frederiksberg, Denmark and.
authors.forename: Felicia
authors.initials: F
authors.lastname: Lauridsen
authors.affiliation: From the Laboratory of Immunology, Section for Experimental Animal Models, Faculty of Health and Medical Sciences, University of Copenhagen, DK-1870 Frederiksberg, Denmark and.
authors.forename: Lars
authors.initials: L
authors.lastname: Andresen
authors.affiliation: From the Laboratory of Immunology, Section for Experimental Animal Models, Faculty of Health and Medical Sciences, University of Copenhagen, DK-1870 Frederiksberg, Denmark and.
authors.forename: Helle
authors.initials: H
authors.lastname: Jensen
authors.affiliation: From the Laboratory of Immunology, Section for Experimental Animal Models, Faculty of Health and Medical Sciences, University of Copenhagen, DK-1870 Frederiksberg, Denmark and sosk@sund.ku.dk.
authors.forename: Søren
authors.initials: S
authors.lastname: Skov
mesh_terms: D000483:Alleles; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001216:Asparagine / Q000737:chemistry; D001665:Binding Sites / Q000235:genetics; D002352:Carrier Proteins / Q000276:immunology / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane / Q000276:immunology / Q000378:metabolism; D006031:Glycosylation; D016199:Herpesvirus 7, Human / Q000276:immunology; D015395:Histocompatibility Antigens Class I / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D055655:NK Cell Lectin-Like Receptor Subfamily K / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013912:Threonine / Q000737:chemistry; D014764:Viral Proteins / Q000276:immunology / Q000378:metabolism
chemical_list: D002352:Carrier Proteins; D015395:Histocompatibility Antigens Class I; C527085:KLRK1 protein, human; D008024:Ligands; C099187:MHC class I-related chain A; D055655:NK Cell Lectin-Like Receptor Subfamily K; D011994:Recombinant Proteins; C488324:U21 glycoprotein, human herpesvirus 7; D014764:Viral Proteins; D013912:Threonine; D001216:Asparagine
keywords: Genetic Polymorphism; Glycosylation; Herpesvirus; Innate Immunity; Melanoma; Natural Killer Cells (NK Cells)