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Journal of Virology, January 2009, p. 408-419, Vol. 83, No. 1
0022-538X/09/$08.00+0     doi:10.1128/JVI.01568-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Exploring the Nuclear Envelope of Herpes Simplex Virus 1-Infected Cells by High-Resolution Microscopy {triangledown}

Peter Wild,1* Claudia Senn,2 Céline L. Manera,1 Esther Sutter,1 Elisabeth M. Schraner,1 Kurt Tobler,2 Mathias Ackermann,2 Urs Ziegler,3 Miriam S. Lucas,4 and Andres Kaech4,{dagger}

Electron Microscopy, Institute of Veterinary Anatomy,1 Institute of Virology,2 Center for Microscopy and Image Analysis, University of Zürich,3 Electron Microscopy ETH Zürich, Swiss Federal Institute of Technology, Zürich, Switzerland4

Received 24 July 2008/ Accepted 6 October 2008

Herpesviruses are composed of capsid, tegument, and envelope. Capsids assemble in the nucleus and exit the nucleus by budding at the inner nuclear membrane, acquiring tegument and the envelope. This study focuses on the changes of the nuclear envelope during herpes simplex virus 1 (HSV-1) infection in HeLa and Vero cells by employing preparation techniques at ambient and low temperatures for high-resolution scanning and transmission electron microscopy and confocal laser scanning microscopy. Cryo-field emission scanning electron microscopy of freeze-fractured cells showed for the first time budding of capsids at the nuclear envelope at the third dimension with high activity at 10 h and low activity at 15 h of incubation. The mean number of pores was significantly lower, and the mean interpore distance and the mean interpore area were significantly larger than those for mock-infected cells 15 h after inoculation. Forty-five percent of nuclear pores in HSV-1-infected cells were dilated to more than 140 nm. Nuclear material containing capsids protrude through them into the cytoplasm. Examination of in situ preparations after dry fracturing revealed significant enlargements of the nuclear pore diameter and of the nuclear pore central channel in HSV-1-infected cells compared to mock-infected cells. The demonstration of nucleoporins by confocal microscopy also revealed fewer pores but focal enhancement of fluorescence signals in HSV-1-infected cells, whereas Western blots showed no loss of nucleoporins from cells. The data suggest that infection with HSV-1 alters the number, size, and architecture of nuclear pores without a loss of nucleoporins from altered nuclear pore complexes.

* Corresponding author. Mailing address: Electron Microscopy, Institute of Virology, Winterthurerstrasse 266a, CH-8057 Zürich, Switzerland. Phone: 41 1 635 87 84. Fax: 41 1 635 89 11. E-mail: pewild{at}vetanat.uzh.ch

{triangledown} Published ahead of print on 15 October 2008.

{dagger} Present address: Center for Microscopy and Image Analysis, University of Zürich, Zürich, Switzerland.

Journal of Virology, January 2009, p. 408-419, Vol. 83, No. 1
0022-538X/09/$08.00+0     doi:10.1128/JVI.01568-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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