title: The RING-CH ligase K5 antagonizes restriction of KSHV and HIV-1 particle release by mediating ubiquitin-dependent endosomal degradation of tetherin.
abstract: Tetherin (CD317/BST2) is an interferon-induced membrane protein that inhibits the release of diverse enveloped viral particles. Several mammalian viruses have evolved countermeasures that inactivate tetherin, with the prototype being the HIV-1 Vpu protein. Here we show that the human herpesvirus Kaposi's sarcoma-associated herpesvirus (KSHV) is sensitive to tetherin restriction and its activity is counteracted by the KSHV encoded RING-CH E3 ubiquitin ligase K5. Tetherin expression in KSHV-infected cells inhibits viral particle release, as does depletion of K5 protein using RNA interference. K5 induces a species-specific downregulation of human tetherin from the cell surface followed by its endosomal degradation. We show that K5 targets a single lysine (K18) in the cytoplasmic tail of tetherin for ubiquitination, leading to relocalization of tetherin to CD63-positive endosomal compartments. Tetherin degradation is dependent on ESCRT-mediated endosomal sorting, but does not require a tyrosine-based sorting signal in the tetherin cytoplasmic tail. Importantly, we also show that the ability of K5 to substitute for Vpu in HIV-1 release is entirely dependent on K18 and the RING-CH domain of K5. By contrast, while Vpu induces ubiquitination of tetherin cytoplasmic tail lysine residues, mutation of these positions has no effect on its antagonism of tetherin function, and residual tetherin is associated with the trans-Golgi network (TGN) in Vpu-expressing cells. Taken together our results demonstrate that K5 is a mechanistically distinct viral countermeasure to tetherin-mediated restriction, and that herpesvirus particle release is sensitive to this mode of antiviral inhibition.
authors.affiliation: MRC Centre for Medical Molecular Virology, University College London, London, United Kingdom.
authors.forename: Claire
authors.initials: C
authors.lastname: Pardieu
authors.forename: Raphaël
authors.initials: R
authors.lastname: Vigan
authors.forename: Sam J
authors.initials: SJ
authors.lastname: Wilson
authors.forename: Alessandra
authors.initials: A
authors.lastname: Calvi
authors.forename: Trinity
authors.initials: T
authors.lastname: Zang
authors.forename: Paul
authors.initials: P
authors.lastname: Bieniasz
authors.forename: Paul
authors.initials: P
authors.lastname: Kellam
authors.forename: Greg J
authors.initials: GJ
authors.lastname: Towers
authors.forename: Stuart J D
authors.initials: SJ
authors.lastname: Neil
mesh_terms: D015703:Antigens, CD / Q000378:metabolism*; D002469:Cell Separation; D011992:Endosomes / Q000378:metabolism; D005434:Flow Cytometry; D058851:GPI-Linked Proteins; D015658:HIV Infections / Q000378:metabolism*; D015497:HIV-1 / Q000378:metabolism / Q000472:pathogenicity; D006367:HeLa Cells; D006566:Herpesviridae Infections / Q000378:metabolism*; D019288:Herpesvirus 8, Human / Q000378:metabolism / Q000472:pathogenicity; D054298:Human Immunodeficiency Virus Proteins; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism*; D018613:Microscopy, Confocal; D020133:Reverse Transcriptase Polymerase Chain Reaction; D025801:Ubiquitin / Q000378:metabolism; D054875:Ubiquitination; D054334:Viral Regulatory and Accessory Proteins; D014771:Virion / Q000378:metabolism; D057074:Virus Release / Q000502:physiology*; D000074010:Bone Marrow Stromal Antigen 2; D000086082:Viroporin Proteins
chemical_list: D015703:Antigens, CD; D058851:GPI-Linked Proteins; D054298:Human Immunodeficiency Virus Proteins; D017874:Immediate-Early Proteins; D008562:Membrane Glycoproteins; D025801:Ubiquitin; D054334:Viral Regulatory and Accessory Proteins; C094572:BST2 protein, human; D000074010:Bone Marrow Stromal Antigen 2; C405867:K5 protein, Human herpesvirus 8; C514859:vpu protein, Human immunodeficiency virus 1; D000086082:Viroporin Proteins