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A Dual Laser Scanning Confocal and Transmission Electron Microscopy Analysis of the Intracellular Localization, Aggregation and Particle Formation of African Horse Sickness Virus Major Core Protein VP7

Published online by Cambridge University Press:  23 January 2017

Gayle V. Wall Open the ORCID record for Gayle V. Wall [Opens in a new window] ,
Daria A. Rutkowska ,
Eshchar Mizrachi ,
Henk Huismans  and
Vida van Staden
Gayle V. Wall
Affiliation:
Department of Genetics, University of Pretoria, Pretoria, 0002, South Africa
Daria A. Rutkowska
Affiliation:
Department of Genetics, University of Pretoria, Pretoria, 0002, South Africa
Eshchar Mizrachi
Affiliation:
Department of Genetics, University of Pretoria, Pretoria, 0002, South Africa
Henk Huismans
Affiliation:
Department of Genetics, University of Pretoria, Pretoria, 0002, South Africa
Vida van Staden*
Affiliation:
Department of Genetics, University of Pretoria, Pretoria, 0002, South Africa
*
*Corresponding author. vida.vanstaden@up.ac.za
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Abstract

The bulk of the major core protein VP7 in African horse sickness virus (AHSV) self-assembles into flat, hexagonal crystalline particles in a process appearing unrelated to viral replication. Why this unique characteristic of AHSV VP7 is genetically conserved, and whether VP7 aggregation and particle formation have an effect on cellular biology or the viral life cycle, is unknown. Here we investigated how different small peptide and enhanced green fluorescent protein (eGFP) insertions into the VP7 top domain affected VP7 localization, aggregation, and particle formation. This was done using a dual laser scanning confocal and transmission electron microscopy approach in conjunction with analyses of the solubility, aggregation, and fluorescence profiles of the proteins. VP7 top domain modifications did not prevent trimerization, or intracellular trafficking, to one or two discrete sites in the cell. However, modifications that resulted in a misfolded and insoluble VP7-eGFP component blocked trafficking, and precluded protein accumulation at a single cellular site, perhaps by interfering with normal trimer–trimer interactions. Furthermore, the modifications disrupted the stable layering of the trimers into characteristic AHSV VP7 crystalline particles. It was concluded that VP7 trafficking is driven by a balance between VP7 solubility, trimer forming ability, and trimer–trimer interactions.

Keywords

AHSV VP7protein aggregationparticle formationVP7 localizationprotein solubility
Type
Biological Applications
Information
Microscopy and Microanalysis , Volume 23 , Issue 1 , February 2017 , pp. 56 - 68
Copyright
© Microscopy Society of America 2017 

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Footnotes

†

Current address: Council for Scientific and Industrial Research (CSIR), Biosciences, Pretoria, 0001, South Africa.

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