A spatio-temporal analysis of matrix protein and nucleocapsid trafficking during vesicular stomatitis virus uncoating

To study VSV entry and the fate of incoming matrix (M) protein during virus uncoating we used recombinant viruses encoding M proteins with a C-terminal tetracysteine tag that could be fluorescently labeled using biarsenical (Lumio) compounds. We found that uncoating occurs early in the endocytic pat...

Full description

Saved in:
Bibliographic Details
Published in:PLoS pathogens 2010-07, Vol.6 (7), p.e1000994-e1000994
Main Authors: Mire, Chad E, White, Judith M, Whitt, Michael A
Format: Article
Language:English
Subjects:
Care and treatment
Cell Biology/Membranes and Sorting
Cellular biology
Cytoplasm
Endosomes
Extracellular matrix
Genetic aspects
Genomes
Glycoproteins - metabolism
Infections
Influenza
Membrane Fusion
Membrane proteins
Membranes
Molecular Probes
Nucleocapsid - metabolism
Polyclonal antibodies
Properties
Protein Transport
Proteins
Ribonucleoproteins
RNA polymerase
Stomatitis
Vesicular stomatitis virus
Vesiculovirus - physiology
Viral envelopes
Viral Matrix Proteins - metabolism
Virology/Host Invasion and Cell Entry
Virus Internalization
Virus Physiological Phenomena
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:To study VSV entry and the fate of incoming matrix (M) protein during virus uncoating we used recombinant viruses encoding M proteins with a C-terminal tetracysteine tag that could be fluorescently labeled using biarsenical (Lumio) compounds. We found that uncoating occurs early in the endocytic pathway and is inhibited by expression of dominant-negative (DN) Rab5, but is not inhibited by DN-Rab7 or DN-Rab11. Uncoating, as defined by the separation of nucleocapsids from M protein, occurred between 15 and 20 minutes post-entry and did not require microtubules or an intact actin cytoskeleton. Unexpectedly, the bulk of M protein remained associated with endosomal membranes after uncoating and was eventually trafficked to recycling endosomes. Another small, but significant fraction of M distributed to nuclear pore complexes, which was also not dependent on microtubules or polymerized actin. Quantification of fluorescence from high-resolution confocal micrographs indicated that after membrane fusion, M protein diffuses across the endosomal membrane with a concomitant increase in fluorescence from the Lumio label which occurred soon after the release of RNPs into the cytoplasm. These data support a new model for VSV uncoating in which RNPs are released from M which remains bound to the endosomal membrane rather than the dissociation of M protein from RNPs after release of the complex into the cytoplasm following membrane fusion.
ISSN:1553-7374
1553-7366
1553-7374
DOI:10.1371/journal.ppat.1000994