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Molecular and Cellular Biology, July 2000, p. 4922-4931, Vol. 20, No. 13
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Herpes Simplex Virus Type 1 Entry into Host Cells: Reconstitution of Capsid Binding and Uncoating at the Nuclear Pore Complex In Vitro

Päivi M. Ojala,1,* Beate Sodeik,1,dagger Melanie W. Ebersold,1 Ulrike Kutay,2 and Ari Helenius1,Dagger

Department of Cell Biology, Yale University, New Haven, Connecticut,1 and Institute of Biochemistry, ETH-Zürich, Zürich, Switzerland2

Received 1 November 1999/Returned for modification 8 December 1999/Accepted 4 April 2000

During entry, herpes simplex virus type 1 (HSV-1) releases its capsid and the tegument proteins into the cytosol of a host cell by fusing with the plasma membrane. The capsid is then transported to the nucleus, where it docks at the nuclear pore complexes (NPCs), and the viral genome is rapidly released into the nucleoplasm. In this study, capsid association with NPCs and uncoating of the viral DNA were reconstituted in vitro. Isolated capsids prepared from virus were incubated with cytosol and purified nuclei. They were found to bind to the nuclear pores. Binding could be inhibited by pretreating the nuclei with wheat germ agglutinin, anti-NPC antibodies, or antibodies against importin beta . Furthermore, in the absence of cytosol, purified importin beta  was both sufficient and necessary to support efficient capsid binding to nuclei. Up to 60 to 70% of capsids interacting with rat liver nuclei in vitro released their DNA if cytosol and metabolic energy were supplied. Interaction of the capsid with the nuclear pore thus seemed to trigger the release of the viral genome, implying that components of the NPC play an active role in the nuclear events during HSV-1 entry into host cells.

* Corresponding author. Mailing address: Haartman Institute, P. O. Box 21, FIN-00014 University of Helsinki, Finland. Phone: 358-9-191 26439. Fax: 358-9-191 26700. E-mail: Paivi.Ojala{at}helsinki.fi.

dagger Present address: Zentrum Biochemie, Medizinische Hochschule, Hannover, Germany.

Dagger Present address: Institute of Biochemistry, ETH-Zürich, Zürich, Switzerland.

Molecular and Cellular Biology, July 2000, p. 4922-4931, Vol. 20, No. 13
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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