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Journal of Virology, February 2000, p. 1973-1984, Vol. 74, No. 4
Center for Immunology and Departments of
Pathology and Molecular Microbiology, Washington University School
of Medicine, St. Louis, Missouri 63110
Received 6 July 1999/Accepted 11 November 1999
Murine gammaherpesvirus 68 (
0022-538X/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Disruption of the Murine Gammaherpesvirus 68 M1
Open Reading Frame Leads to Enhanced Reactivation from
Latency
HV68, or MHV-68) is a genetically
tractable, small animal model for the analysis of gammaherpesvirus pathogenesis. The
HV68 genome is colinear with the genomes of other
sequence gammaherpesviruses, containing large blocks of conserved genes
interspersed by a number of putative genes without clear homologs in
the other gammaherpesviruses. One of these putative unique genes, the
M1 open reading frame (ORF), exhibits sequence homology to a poxvirus
serine protease inhibitor, SPI-1, as well as to another
HV68 gene,
M3, which we have recently shown encodes an abundantly secreted
chemokine binding protein. To assess the contribution of the M1 ORF to
HV68 pathogenesis, we have generated a recombinant
HV68 in which
the M1 ORF has been disrupted through targeted insertion of a
lacZ expression cassette (M1.LacZ). Although M1.LacZ
replicated normally in tissue culture, it exhibited decreased splenic
titers at days 4 and 9 postinfection in both immunocompetent and
immunodeficient mice. Despite decreased levels of acute virus replication, M1.LacZ established a latent infection comparable to
wild-type (wt)
HV68, but exhibited an approximately fivefold increase in efficiency of reactivation from latency. M1.LacZ also caused severe vasculitis of the great elastic arteries in gamma interferon receptor (IFN-
R)-deficient mice with a frequency
comparable to wt
HV68, but did not cause the mortality or splenic
pathology observed with wt
HV68 infection of IFN-
R-deficient
mice. Restoration of M1 ORF sequences into M1.LacZ (M1 marker rescue,
or M1.MR) demonstrated that M1.LacZ phenotypic alterations in growth in vivo and latency were not due to the presence of additional mutations located elsewhere in the M1.LacZ genome. Generation of a second M1
mutant virus containing a deletion at the 5' end of the M1 ORF
(M1
511), but lacking the LacZ expression cassette, revealed the same
latency phenotype observed with the M1.LacZ mutant. However, M1
511
was not attenuated for acute virus replication in the spleen. We
conclude that (i) the induction of arteritis in
HV68-infected IFN-
R-deficient mice can occur in the absence of splenic pathology and mortality, (ii) replication during acute infection is not the
primary determinant for the establishment of latent infection, and
(iii) the M1 ORF, or a closely linked gene, encodes a gene product that
functions to suppress virus reactivation.
*
Corresponding author. Mailing address: Department of
Pathology, Box 8118, Washington University School of Medicine, 660 S. Euclid Ave., St. Louis, MO 63110. Fax: (314) 362-4096. H.W.V.: Phone:
(314) 362-9223. E-mail: virgin{at}immunology.wustl.edu.
S.H.S.: Phone: (314) 362-0367. E-mail:
speck{at}pathology.wustl.edu.
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