title: Three-Dimensional Structural Characterization of HIV-1 Tethered to Human Cells.
abstract: Tetherin (BST2, CD317, or HM1.24) is a host cellular restriction factor that prevents the release of enveloped viruses by mechanically linking virions to the plasma membrane. The precise arrangement of tetherin molecules at the plasma membrane site of HIV-1 assembly, budding, and restriction is not well understood. To gain insight into the biophysical mechanism underlying tetherin-mediated restriction of HIV-1, we utilized cryo-electron tomography (cryo-ET) to directly visualize HIV-1 virus-like particles (VLPs) and virions tethered to human cells in three dimensions (3D). Rod-like densities that we refer to as tethers were seen connecting HIV-1 virions to each other and to the plasma membrane. Native immunogold labeling showed tetherin molecules located on HIV-1 VLPs and virions in positions similar to those of the densities observed by cryo-ET. The location of the tethers with respect to the ordered immature Gag lattice or mature conical core was random. However, tethers were not uniformly distributed on the viral membrane but rather formed clusters at sites of contact with the cell or other virions. Chains of tethered HIV-1 virions often were arranged in a linear fashion, primarily as single chains and, to a lesser degree, as branched chains. Distance measurements support the extended tetherin model, in which the coiled-coil ectodomains are oriented perpendicular with respect to the viral and plasma membranes.
  
  
  OBJECTIVE
  Tetherin is a cellular factor that restricts HIV-1 release by directly cross-linking the virus to the host cell plasma membrane. We used cryo-electron tomography to visualize HIV-1 tethered to human cells in 3D. We determined that tetherin-restricted HIV-1 virions were physically connected to each other or to the plasma membrane by filamentous tethers that resembled rods ∼15 nm in length, which is consistent with the extended tetherin model. In addition, we found the position of the tethers to be arbitrary relative to the ordered immature Gag lattice or the mature conical cores. However, when present as multiple copies, the tethers clustered at the interface between virions. Tethered HIV-1 virions were arranged in a linear fashion, with the majority as single chains. This study advances our understanding of tetherin-mediated HIV-1 restriction by defining the spatial arrangement and orientation of tetherin molecules at sites of HIV-1 restriction.
authors.affiliation: Department of Pediatrics, Emory University School of Medicine, and Children's Healthcare of Atlanta, Atlanta, Georgia, USA.
authors.forename: Joshua D
authors.initials: JD
authors.lastname: Strauss
authors.affiliation: Department of Pediatrics, Emory University School of Medicine, and Children's Healthcare of Atlanta, Atlanta, Georgia, USA.
authors.forename: Jason E
authors.initials: JE
authors.lastname: Hammonds
authors.affiliation: Robert P. Apkarian Integrated Electron Microscopy Core, Emory University, Atlanta, Georgia, USA.
authors.forename: Hong
authors.initials: H
authors.lastname: Yi
authors.affiliation: Department of Pediatrics, Emory University School of Medicine, and Children's Healthcare of Atlanta, Atlanta, Georgia, USA.
authors.forename: Lingmei
authors.initials: L
authors.lastname: Ding
authors.affiliation: Department of Pediatrics, Emory University School of Medicine, and Children's Healthcare of Atlanta, Atlanta, Georgia, USA paul.spearman@emory.edu erwrigh@emory.edu.
authors.forename: Paul
authors.initials: P
authors.lastname: Spearman
authors.affiliation: Department of Pediatrics, Emory University School of Medicine, and Children's Healthcare of Atlanta, Atlanta, Georgia, USA Robert P. Apkarian Integrated Electron Microscopy Core, Emory University, Atlanta, Georgia, USA paul.spearman@emory.edu erwrigh@emory.edu.
authors.forename: Elizabeth R
authors.initials: ER
authors.lastname: Wright
mesh_terms: D015703:Antigens, CD / Q000378:metabolism*; D055592:Biophysical Phenomena; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure*; D020285:Cryoelectron Microscopy; D055032:Electron Microscope Tomography; D058851:GPI-Linked Proteins / Q000378:metabolism; D015497:HIV-1 / Q000276:immunology / Q000502:physiology* / Q000648:ultrastructure*; D006801:Humans; D014771:Virion / Q000378:metabolism / Q000648:ultrastructure*; D053585:Virus Attachment; D057074:Virus Release; D000074010:Bone Marrow Stromal Antigen 2
chemical_list: D015703:Antigens, CD; D058851:GPI-Linked Proteins; C094572:BST2 protein, human; D000074010:Bone Marrow Stromal Antigen 2