Glycoprotein-dependent acidification of vesicular stomatitis virus enhances release of matrix protein.
To study vesicular stomatitis virus (VSV) entry and uncoating, we generated a recombinant VSV encoding a matrix (M) protein containing a C-terminal tetracysteine Lumio tag (rVSV-ML) that could be fluorescently labeled using biarsenical compounds. Quantitative confocal microscopy showed that there is a transient loss of fluorescence at early times after the initiation of endocytosis of rVSV-ML-Green (rVSV-MLG) virions, which did not occur when cells were treated with bafilomycin A1. The reduction in fluorescence occurred 5 to 10 min postentry, followed by a steady increase in fluorescence intensity from 15 to 60 min postentry. A similar loss of fluorescence was observed in vitro when virions were exposed to acidic pH. The reduction in fluorescence required G protein since "bald" DeltaG-MLG particles did not show a similar loss of fluorescence at low pH. Based on the pH-dependent fluorescence properties of Lumio Green, we hypothesize that the loss of fluorescence of rVSV-MLG virions during virus entry is due to a G ectodomain-dependent acidification of the virion interior. Biochemical analysis indicated that low pH also resulted in an enhancement of M protein dissociation from partially permeabilized, but otherwise intact, wild-type virions. From these data we propose that low-pH conformational changes in G protein promote acidification of the virus interior, which facilitates the release of M from ribonucleoprotein particles during uncoating.
Mire
Chad E
CE
Department of Molecular Sciences, University of Tennessee Health Science Center, Memphis, Tennessee 38163, USA.
Dube
Derek
D
Delos
Sue E
SE
White
Judith M
JM
Whitt
Michael A
MA
D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D006023:Glycoproteins / Q000378:metabolism*; D018613:Microscopy, Confocal; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013194:Staining and Labeling; D013997:Time Factors; D018116:Vesiculovirus / Q000502:physiology*; D014763:Viral Matrix Proteins / Q000235:genetics / Q000378:metabolism*; D014764:Viral Proteins / Q000378:metabolism*; D053586:Virus Internalization
D006023:Glycoproteins; D011994:Recombinant Proteins; D014763:Viral Matrix Proteins; D014764:Viral Proteins