Cryo-electron Microscopy Structure of the Native Prototype Foamy Virus Glycoprotein and Virus Architecture

PLoS Pathog. 2016 Jul 11;12(7):e1005721. doi: 10.1371/journal.ppat.1005721. eCollection 2016 Jul.

Abstract

Foamy viruses (FV) belong to the genus Spumavirus, which forms a distinct lineage in the Retroviridae family. Although the infection in natural hosts and zoonotic transmission to humans is asymptomatic, FVs can replicate well in human cells making it an attractive gene therapy vector candidate. Here we present cryo-electron microscopy and (cryo-)electron tomography ultrastructural data on purified prototype FV (PFV) and PFV infected cells. Mature PFV particles have a distinct morphology with a capsid of constant dimension as well as a less ordered shell of density between the capsid and the membrane likely formed by the Gag N-terminal domain and the cytoplasmic part of the Env leader peptide gp18LP. The viral membrane contains trimeric Env glycoproteins partly arranged in interlocked hexagonal assemblies. In situ 3D reconstruction by subtomogram averaging of wild type Env and of a Env gp48TM- gp80SU cleavage site mutant showed a similar spike architecture as well as stabilization of the hexagonal lattice by clear connections between lower densities of neighboring trimers. Cryo-EM was employed to obtain a 9 Å resolution map of the glycoprotein in its pre-fusion state, which revealed extensive trimer interactions by the receptor binding subunit gp80SU at the top of the spike and three central helices derived from the fusion protein subunit gp48TM. The lower part of Env, presumably composed of interlaced parts of gp48TM, gp80SU and gp18LP anchors the spike at the membrane. We propose that the gp48TM density continues into three central transmembrane helices, which interact with three outer transmembrane helices derived from gp18LP. Our ultrastructural data and 9 Å resolution glycoprotein structure provide important new insights into the molecular architecture of PFV and its distinct evolutionary relationship with other members of the Retroviridae.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Blotting, Western
  • Cell Line
  • Cryoelectron Microscopy
  • Gene Products, env / ultrastructure*
  • Glycoproteins / ultrastructure*
  • Humans
  • Image Processing, Computer-Assisted
  • Protein Conformation
  • Spumavirus / chemistry
  • Spumavirus / ultrastructure*
  • Transfection

Substances

  • Gene Products, env
  • Glycoproteins

Grant support

This work is supported by Agence Nationale de la Recherche (ANR-14-CE09-0003-01 to WW and GS) and Deutsche Forschungsgemeinschaft (LI 621/3-3, LI 621/10-1 to DL). GE was supported by a post-doctoral fellowship from the ANRS at early stages of the project. WW acknowledges support from the Institut Universitaire de France. This work used the platforms of the Grenoble Instruct Center (ISBG: UMS 3518 CNRS-CEA-UJF-EMBL) with support from FRISBI (ANR-10-INSB-05-02) and GRAL (ANR-10-LABX-49-01) within the Grenoble Partnership for Structural Biology (PSB). The electron microscopy facility is supported by the Rhône-Alpes Region and by the Fondation pour la Recherche Medicale. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.