Ebolavirus Is Internalized Into Host Cells via Macropinocytosis in a Viral Glycoprotein-Dependent Manner

PLoS Pathog. 2010 Sep 23;6(9):e1001121. doi: 10.1371/journal.ppat.1001121.

Abstract

Ebolavirus (EBOV) is an enveloped, single-stranded, negative-sense RNA virus that causes severe hemorrhagic fever with mortality rates of up to 90% in humans and nonhuman primates. Previous studies suggest roles for clathrin- or caveolae-mediated endocytosis in EBOV entry; however, ebolavirus virions are long, filamentous particles that are larger than the plasma membrane invaginations that characterize clathrin- or caveolae-mediated endocytosis. The mechanism of EBOV entry remains, therefore, poorly understood. To better understand Ebolavirus entry, we carried out internalization studies with fluorescently labeled, biologically contained Ebolavirus and Ebolavirus-like particles (Ebola VLPs), both of which resemble authentic Ebolavirus in their morphology. We examined the mechanism of Ebolavirus internalization by real-time analysis of these fluorescently labeled Ebolavirus particles and found that their internalization was independent of clathrin- or caveolae-mediated endocytosis, but that they co-localized with sorting nexin (SNX) 5, a marker of macropinocytosis-specific endosomes (macropinosomes). Moreover, the internalization of Ebolavirus virions accelerated the uptake of a macropinocytosis-specific cargo, was associated with plasma membrane ruffling, and was dependent on cellular GTPases and kinases involved in macropinocytosis. A pseudotyped vesicular stomatitis virus possessing the Ebolavirus glycoprotein (GP) also co-localized with SNX5 and its internalization and infectivity were affected by macropinocytosis inhibitors. Taken together, our data suggest that Ebolavirus is internalized into cells by stimulating macropinocytosis in a GP-dependent manner. These findings provide new insights into the lifecycle of Ebolavirus and may aid in the development of therapeutics for Ebolavirus infection.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Animals
  • Blotting, Western
  • Caveolae / metabolism
  • Caveolae / virology
  • Cells, Cultured
  • Chlorocebus aethiops
  • Clathrin / metabolism
  • Ebolavirus / physiology*
  • Endocytosis / physiology
  • Hemorrhagic Fever, Ebola / metabolism
  • Hemorrhagic Fever, Ebola / virology*
  • Humans
  • Microscopy, Fluorescence
  • Pinocytosis / physiology*
  • RNA, Messenger / genetics
  • Reverse Transcriptase Polymerase Chain Reaction
  • Signal Transduction
  • Sorting Nexins / genetics
  • Sorting Nexins / metabolism*
  • Vero Cells
  • Vesiculovirus
  • Viral Envelope Proteins / genetics
  • Viral Envelope Proteins / metabolism*
  • Virion / genetics
  • Virus Internalization*
  • Virus Replication
  • rab GTP-Binding Proteins / genetics
  • rab GTP-Binding Proteins / metabolism

Substances

  • Clathrin
  • RNA, Messenger
  • Sorting Nexins
  • Viral Envelope Proteins
  • envelope glycoprotein, Ebola virus
  • rab7 protein
  • rab GTP-Binding Proteins