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

PLoS Pathogens
Chad E MireM A Whitt

Abstract

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 whic...Continue Reading

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Citations

Nov 3, 2011·PLoS Pathogens·Eric M FeeleyAbraham L Brass
Apr 8, 2016·Molecular Biology of the Cell·John V CoxMichael A Whitt
Sep 16, 2015·PLoS Pathogens·Qian ShuCarolyn B Coyne
Feb 16, 2007·Virology·Philippe PlattetDaniel Kolakofsky
May 4, 2011·Chembiochem : a European Journal of Chemical Biology·Adam Pomorski, Artur Krężel
Feb 13, 2016·Journal of Virology·Katie M Stiles, Margaret Kielian
Oct 16, 2015·Journal of Virology·James A SimmonsJudith M White
Jun 8, 2012·Journal of Virology·Bernardo A Mainou, Terence S Dermody
Jul 13, 2013·Pathogens and Disease·Sreejith RajasekharanSanjay Gupta
Feb 23, 2012·Viruses·Aurélie A V AlbertiniYves Gaudin
Jan 29, 2021·Journal of Biomolecular Structure & Dynamics·Ritudhwaj TiwariDebasis Nayak

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Methods Mentioned

BETA
confocal microscopy
GTPases
Fluorescence
protein assay

Software Mentioned

Canvas
LSM
Photoshop
Axiovision
Image J
LSM Physiology
Zeiss LSM Physiology
Adobe Photoshop
AIM
Zeiss LSM510

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