Reovirus FAST Proteins Drive Pore Formation and Syncytiogenesis Using a Novel Helix-Loop-Helix Fusion-Inducing Lipid Packing Sensor

PLoS Pathog. 2015 Jun 10;11(6):e1004962. doi: 10.1371/journal.ppat.1004962. eCollection 2015 Jun.

Abstract

Pore formation is the most energy-demanding step during virus-induced membrane fusion, where high curvature of the fusion pore rim increases the spacing between lipid headgroups, exposing the hydrophobic interior of the membrane to water. How protein fusogens breach this thermodynamic barrier to pore formation is unclear. We identified a novel fusion-inducing lipid packing sensor (FLiPS) in the cytosolic endodomain of the baboon reovirus p15 fusion-associated small transmembrane (FAST) protein that is essential for pore formation during cell-cell fusion and syncytiogenesis. NMR spectroscopy and mutational studies indicate the dependence of this FLiPS on a hydrophobic helix-loop-helix structure. Biochemical and biophysical assays reveal the p15 FLiPS preferentially partitions into membranes with high positive curvature, and this partitioning is impeded by bis-ANS, a small molecule that inserts into hydrophobic defects in membranes. Most notably, the p15 FLiPS can be functionally replaced by heterologous amphipathic lipid packing sensors (ALPS) but not by other membrane-interactive amphipathic helices. Furthermore, a previously unrecognized amphipathic helix in the cytosolic domain of the reptilian reovirus p14 FAST protein can functionally replace the p15 FLiPS, and is itself replaceable by a heterologous ALPS motif. Anchored near the cytoplasmic leaflet by the FAST protein transmembrane domain, the FLiPS is perfectly positioned to insert into hydrophobic defects that begin to appear in the highly curved rim of nascent fusion pores, thereby lowering the energy barrier to stable pore formation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Chlorocebus aethiops
  • Chromatography, High Pressure Liquid
  • Circular Dichroism
  • Magnetic Resonance Spectroscopy
  • Membrane Fusion / physiology*
  • Protein Conformation
  • Reoviridae / pathogenicity
  • Reoviridae / physiology*
  • Transfection
  • Vero Cells
  • Viral Fusion Proteins / chemistry
  • Viral Fusion Proteins / metabolism*

Substances

  • Viral Fusion Proteins

Associated data

  • PDB/2MNS