Three-dimensional imaging of the intracellular assembly of a functional viral RNA replicase complex

J Cell Sci. 2017 Jan 1;130(1):260-268. doi: 10.1242/jcs.181586. Epub 2016 Mar 29.

Abstract

Positive-strand RNA viruses, which can be devastating pathogens in humans, animals and plants, replicate their genomes on intracellular membranes. Here, we describe the three-dimensional ultrastructural organization of a tombusvirus replicase in yeast, a valuable model for exploring virus-host interactions. We visualized the intracellular distribution of a viral replicase protein using metal-tagging transmission electron microscopy, a highly sensitive nanotechnology whose full potential remains to be developed. These three-dimensional images show how viral replicase molecules are organized when they are incorporated into the active domains of the intracellular replication compartment. Our approach provides a means to study protein activation mechanisms in cells and to identify targets for new antiviral compounds.

Keywords: 3D electron microscopy; Clonable tag; Metal-tagging transmission electron microscopy; Tomography; Viral replication factory.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Antibodies / metabolism
  • Imaging, Three-Dimensional*
  • Intracellular Space / virology*
  • Metallothionein / metabolism
  • Models, Biological
  • RNA Replicase / metabolism*
  • RNA, Double-Stranded / metabolism
  • RNA, Viral / metabolism*
  • Saccharomyces cerevisiae / ultrastructure
  • Saccharomyces cerevisiae / virology
  • Tombusvirus / physiology*
  • Tombusvirus / ultrastructure
  • Tomography
  • Virus Assembly*
  • Virus Replication

Substances

  • Antibodies
  • RNA, Double-Stranded
  • RNA, Viral
  • Metallothionein
  • RNA Replicase

Associated data

  • figshare/10.6084/m9.figshare.3159094.v1