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Journal of Virology, May 2001, p. 4048-4055, Vol. 75, No. 9
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.9.4048-4055.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.

Regulation of Viral Intermediate Gene Expression by the Vaccinia Virus B1 Protein Kinase†

Gerald R. Kovacs,1,* Nikos Vasilakis,1 and Bernard Moss2

Department of Viral Vaccine Research, Wyeth-Lederle Vaccines, Pearl River, New York 10965,1 and Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 208922

Received 2 November 2000/Accepted 2 February 2001

The B1 gene of vaccinia virus encodes a serine/threonine protein kinase that is expressed early after infection. Under nonpermissive conditions, temperature-sensitive mutants (ts2 and ts25) that map to B1 fail to efficiently replicate viral DNA. Our goal was to extend studies on the function of B1 by determining if the kinase is required for intermediate or late gene expression, two events that ordinarily depend on viral DNA replication. First, we established that early viral gene expression occurred at the nonpermissive temperature. By using a transfection procedure that circumvents the viral DNA replication requirement, we found that reporter genes regulated by an intermediate promoter were transcribed only under conditions permissive for expression of active B1. To assay late gene expression, the T7 RNA polymerase gene was inserted into the genome of ts25 to form ts25/T7. A DNA replication-independent late gene transcription system was established by cotransfecting plasmids containing T7 promoter-driven late gene transcription factors and a late promoter reporter gene into ts25/T7-infected cells. Late genes, unlike intermediate genes, were expressed at the nonpermissive temperature. Last, we showed that overexpression of B1 stimulated intermediate but inhibited late gene expression in cells infected with wild-type virus.

* Corresponding author. Mailing address: Wyeth-Lederle Vaccines, Department of Viral Vaccine Research, Pearl River, NY 10965. Phone: (845) 732-5974. Fax: (845) 732-4941. E-mail: KOVACSG{at}WAR.WYETH.COM.

dagger This work is dedicated to the memory of Roskey Jennings, whose character was an inspiration to the many people who passed through the Laboratory of Viral Diseases.

Journal of Virology, May 2001, p. 4048-4055, Vol. 75, No. 9
0022-538X/01/$04.00+0   DOI: 10.1128/JVI.75.9.4048-4055.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.


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