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Journal of Virology, February 2001, p. 1450-1458, Vol. 75, No. 3
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.3.1450-1458.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Genetic Evidence of an Essential Role for
Cytomegalovirus Small Capsid Protein in Viral Growth
Eva-Maria
Borst,
Sibylle
Mathys,
Markus
Wagner,
Walter
Muranyi, and
Martin
Messerle*
Max von Pettenkofer-Institut für
Hygiene und Medizinische Mikrobiologie, Lehrstuhl Virologie,
Genzentrum, Ludwig-Maximilians-Universität
München, D-81377 Munich, Germany
Received 21 August 2000/Accepted 6 November 2000
Many steps in the replication cycle of cytomegalovirus (CMV), like
cell entry, capsid assembly, and egress of newly synthesized virions,
have not been completely analyzed yet. In order to facilitate these
studies, we decided to construct a recombinant CMV that incorporates
the green fluorescent protein (GFP) into the nucleocapsid. A comparable
herpes simplex virus type 1 (HSV-1) mutant has recently been generated
by fusion of the GFP open reading frame (ORF) with the HSV-1 ORF
encoding small capsid protein (SCP) VP26 (P. Desai and S. Person,
J. Virol. 72:7563-7568, 1998). Recombinant CMV genomes expressing
a fusion protein consisting of GFP and the SCP were constructed by the
recently established bacterial artificial chromosome mutagenesis
procedure. In transfected cells, the SCP-GFP fusion protein localized
to distinct foci in the nucleus that may represent sites for capsid
assembly (assemblons). However, no viable progeny was reconstituted
from these mutant CMV genomes. CMV genomes with deletion of the SCP ORF
also did not give rise to infectious virus. Rescue of the mutation by
insertion of the SCP gene at an ectopic position in an SCP knockout
genome indicates that, in contrast to the HSV-1 SCP, the CMV SCP is
essential for viral growth. Expression of the SCP-GFP fusion protein
together with the authentic SCP blocked the CMV infection cycle,
suggesting that the SCP-GFP fusion protein exerts a dominant-negative
effect on the assembly of new virions. The results of this study are discussed with regard to recently published data about the structure of
the CMV virion and its differences from the HSV-1 virion.
*
Corresponding author. Mailing address: Max von
Pettenkofer-Institut, Genzentrum, Feodor-Lynen-Strasse 25, D-81377
Munich, Germany. Phone: 49 89 2180 6850. Fax: 49 89 2180 6898. E-mail: messerle{at}lmb.uni-muenchen.de.
Journal of Virology, February 2001, p. 1450-1458, Vol. 75, No. 3
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.3.1450-1458.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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