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Journal of Virology, March 2001, p. 2575-2583, Vol. 75, No. 6
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.6.2575-2583.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Fluorescent Tagging of Herpes Simplex Virus Tegument Protein VP13/14 in Virus Infection

Michelle Donnelly and Gillian Elliott*

Virus Assembly Group, Marie Curie Research Institute, The Chart, Oxted, Surrey RH8 0TL, United Kingdom

Received 12 October 2000/Accepted 19 December 2000

The cellular site of herpesvirus tegument assembly has yet to be defined. We have previously used a recombinant herpes simplex virus type 1 expressing a green fluorescent protein (GFP)-tagged tegument protein, namely VP22, to show that VP22 is localized exclusively to the cytoplasm during infection. Here we have constructed a similar virus expressing another fluorescent tegument protein, YFP-VP13/14, and have visualized the intracellular localization of this second tegument protein in live infected cells. In contrast to VP22, VP13/14 is targeted predominantly to the nuclei of infected cells at both early and late times in infection. More specifically, YFP-13/14 localizes initially to the nuclear replication compartments and then progresses into intense punctate domains that appear at around 12 h postinfection. At even later times this intranuclear punctate fluorescence is gradually replaced by perinuclear micropunctate and membranous fluorescence. While the vast majority of YFP-13/14 seems to be targeted to the nucleus, a minor subpopulation also appears in a vesicular pattern in the cytoplasm that closely resembles the pattern previously observed for GFP-22. Moreover, at late times weak fluorescence appears at the cell periphery and in extracellular virus particles, confirming that YFP-13/14 is assembled into virions. This predominantly nuclear targeting of YFP-13/14 together with the cytoplasmic targeting of VP22 may imply that there are multiple sites of tegument protein incorporation along the virus maturation pathway. Thus, our YFP-13/14-expressing virus has revealed the complexity of the intracellular targeting of VP13/14 and provides a novel insight into the mechanism of tegument, and hence virus, assembly.


* Corresponding author. Mailing address: Virus Assembly Group, Marie Curie Research Institute, The Chart, Oxted, Surrey RH8 0TL, United Kingdom. Phone: 441883 722306. Fax: 441883 714375. E-mail: g.elliott{at}mcri.ac.uk.


Journal of Virology, March 2001, p. 2575-2583, Vol. 75, No. 6
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.6.2575-2583.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.



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