title: A strategy for O-glycoproteomics of enveloped viruses--the O-glycoproteome of herpes simplex virus type 1.
abstract: Glycosylation of viral envelope proteins is important for infectivity and interaction with host immunity, however, our current knowledge of the functions of glycosylation is largely limited to N-glycosylation because it is difficult to predict and identify site-specific O-glycosylation. Here, we present a novel proteome-wide discovery strategy for O-glycosylation sites on viral envelope proteins using herpes simplex virus type 1 (HSV-1) as a model. We identified 74 O-linked glycosylation sites on 8 out of the 12 HSV-1 envelope proteins. Two of the identified glycosites found in glycoprotein B were previously implicated in virus attachment to immune cells. We show that HSV-1 infection distorts the secretory pathway and that infected cells accumulate glycoproteins with truncated O-glycans, nonetheless retaining the ability to elongate most of the surface glycans. With the use of precise gene editing, we further demonstrate that elongated O-glycans are essential for HSV-1 in human HaCaT keratinocytes, where HSV-1 produced markedly lower viral titers in HaCaT with abrogated O-glycans compared to the isogenic counterpart with normal O-glycans. The roles of O-linked glycosylation for viral entry, formation, secretion, and immune recognition are poorly understood, and the O-glycoproteomics strategy presented here now opens for unbiased discovery on all enveloped viruses.
authors.affiliation: Copenhagen Center for Glycomics, Institute of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Ieva
authors.initials: I
authors.lastname: Bagdonaite
authors.affiliation: Department of Clinical Virology, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Rickard
authors.initials: R
authors.lastname: Nordén
authors.affiliation: Copenhagen Center for Glycomics, Institute of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Hiren J
authors.initials: HJ
authors.lastname: Joshi
authors.affiliation: Institute of Odontology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Sally
authors.initials: S
authors.lastname: Dabelsteen
authors.affiliation: Department of Clinical Virology, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Kristina
authors.initials: K
authors.lastname: Nyström
authors.affiliation: Copenhagen Center for Glycomics, Institute of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Sergey Y
authors.initials: SY
authors.lastname: Vakhrushev
authors.affiliation: Department of Clinical Virology, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Sigvard
authors.initials: S
authors.lastname: Olofsson
authors.affiliation: Copenhagen Center for Glycomics, Institute of Cellular and Molecular Medicine, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Hans H
authors.initials: HH
authors.lastname: Wandall
mesh_terms: D000818:Animals; D005434:Flow Cytometry; D054794:Glycomics; D006023:Glycoproteins / Q000378:metabolism; D006031:Glycosylation; D018259:Herpesvirus 1, Human / Q000378:metabolism*; D006801:Humans; D007150:Immunohistochemistry; D013058:Mass Spectrometry / Q000379:methods*; D016133:Polymerase Chain Reaction; D040901:Proteomics / Q000379:methods*; D014759:Viral Envelope Proteins / Q000378:metabolism*
chemical_list: D006023:Glycoproteins; D014759:Viral Envelope Proteins
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