Role of clathrin-mediated endocytosis during vesicular stomatitis virus entry into host cells

Virology. 2005 Jul 20;338(1):53-60. doi: 10.1016/j.virol.2005.05.006.

Abstract

Vesicular stomatitis virus (VSV) is well established to enter cells by pH-dependent endocytosis, but mechanistic aspects of its internalization have remained unclear. Here, we examined the functional role of clathrin in VSV entry by expression of a dominant-negative mutant of Eps15 (GFP-Eps15Delta95/295), a protein essential for clathrin-mediated endocytosis. Whereas expression of GFP alone had no effect on VSV infection, expression of GFP-Eps15Delta95/295 severely limited infection. As independent ways to examine clathrin function, we also examined cells that had been treated with chlorpromazine and utilized small interfering RNA (siRNA) techniques. Inhibition of clathrin-mediated endocytosis by chlorpromazine treatment, as well as clathrin knock-down using siRNA duplexes directed against the clathrin heavy chain, also prevented VSV infection. In combination with previous morphological approaches, these experiments establish clathrin as an essential component needed for endocytosis of VSV.

Publication types

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

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Animals
  • Base Sequence
  • Calcium-Binding Proteins / genetics
  • Calcium-Binding Proteins / physiology
  • Cell Line
  • Chlorpromazine / pharmacology
  • Clathrin / antagonists & inhibitors
  • Clathrin / genetics
  • Clathrin / physiology*
  • Clathrin Heavy Chains / genetics
  • Cricetinae
  • Endocytosis / physiology*
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • HeLa Cells
  • Humans
  • Intracellular Signaling Peptides and Proteins
  • Phosphoproteins / genetics
  • Phosphoproteins / physiology
  • RNA, Small Interfering / genetics
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Transfection
  • Transferrin / metabolism
  • Vesicular stomatitis Indiana virus / pathogenicity*
  • Vesicular stomatitis Indiana virus / physiology

Substances

  • Adaptor Proteins, Signal Transducing
  • Calcium-Binding Proteins
  • Clathrin
  • EPS15 protein, human
  • Intracellular Signaling Peptides and Proteins
  • Phosphoproteins
  • RNA, Small Interfering
  • Recombinant Fusion Proteins
  • Transferrin
  • Clathrin Heavy Chains
  • Green Fluorescent Proteins
  • Chlorpromazine