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Tagging of the vaccinia virus protein F13 with mCherry causes aberrant virion morphogenesis

Published version
Peer-reviewed

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Type

Article

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Authors

Hollinshead, MS 
Ewles, HA 
Lee, S-A 
Smith, GL 

Abstract

Vaccinia virus produces two distinct infectious virions; the single-enveloped intracellular mature virus (IMV), which remains in the cell until cell lysis, and the double-enveloped extracellular enveloped virus (EEV), which mediates virus spread. The latter is derived from a triple-enveloped intracellular enveloped virus (IEV) precursor, which is transported to the cell periphery by the kinesin-1 motor complex. This transport involves the viral protein A36 as well as F12 and E2. A36 is an integral membrane protein associated with the outer virus envelope and is the only known direct link between virion and kinesin-1 complex. Yet in the absence of A36 virion egress still occurs on microtubules, albeit at reduced efficiency. In this paper double-fluorescent labelling of the capsid protein A5 and outer-envelope protein F13 was exploited to visualize IEV transport by live-cell imaging in the absence of either A36 or F12. During the generation of recombinant viruses expressing both A5-GFP and F13-mCherry a plaque size defect was identified that was particularly severe in viruses lacking A36. Electron microscopy showed that this phenotype was caused by abnormal wrapping of IMV to form IEV, and this resulted in reduced virus egress to the cell surface. The aberrant wrapping phenotype suggests that the fluorescent fusion protein interferes with an interaction of F13 with the IMV surface that is required for tight association between IMVs and wrapping membranes. The severity of this defect suggests that these viruses are imperfect tools for characterizing virus egress.

Description

Keywords

Poxviridae, Vaccinia, F13, A36, F12, virus morphogenesis, virus wrapping, IEV, electron microscopy, virus egress, fluorescent protein tagging

Journal Title

Journal of General Virology

Conference Name

Journal ISSN

0022-1317
1465-2099

Volume Title

Publisher

Society for General Microbiology
Sponsorship
Medical Research Council (G1000207)
Wellcome Trust (090315/Z/09/Z)
This work was supported by grants from the Medical Research Council (UK) G1000207 and the Wellcome Trust (UK) grant 090315.