Article
Global analysis of herpes simplex virus type 1 transcription using an oligonucleotide-based DNA microarray.
Department of Molecular Biology and Biochemistry, University of California, Irvine, California 92697, USA.
Journal of Virology (impact factor:
5.4).
12/2000;
74(21):9916-27.
pp.9916-27
Source: PubMed
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Article: Replication of herpes simplex virus type 1 within trigeminal ganglia is required for high frequency but not high viral genome copy number latency.
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ABSTRACT: The replication properties of a thymidine kinase-negative (TK(-)) mutant of herpes simplex virus type 1 (HSV-1) were exploited to examine the relative contributions of replication at the body surface and within trigeminal ganglia (TG) on the establishment of latent infections. The replication of a TK(-) mutant, 17/tBTK(-), was reduced by approximately 12-fold on the mouse cornea compared to the rescued isolate 17/tBRTK(+), and no replication of 17/tBTK(-) in the TG of these mice was detected. About 1.8% of the TG neurons of mice infected with 17/tBTK(-) harbored the latent viral genome compared to 23% of those infected with 17/tBRTK(+). In addition, the latent sites established by the TK(-) mutant contained fewer copies of the HSV-1 genome (average, 2.3/neuron versus 28/neuron). On the snout, sustained robust replication of 17tBTK(-) in the absence of significant replication within the TG resulted in a modest increase in the number of latent sites. Importantly, these latently infected neurons displayed a wild-type latent-genome copy number profile, with some neurons containing hundreds of copies of the TK(-) mutant genome. As expected, the replication of the TK(-) mutant appeared to be blocked prior to DNA replication in most ganglionic neurons in that (i) virus replication was severely restricted in ganglia, (ii) the number of neurons expressing HSV proteins was reduced 30-fold compared to the rescued isolate, (iii) cell-to-cell spread of virus was not detected within ganglia, and (iv) the proportion of infected neurons expressing late proteins was reduced by 89% compared to the rescued strain. These results demonstrate that the viral TK gene is required for the efficient establishment of latency. This requirement appears to be primarily for efficient replication within the ganglion, which leads to a sixfold increase in the number of latent sites established. Further, latent sites with high genome copy number can be established in the absence of significant virus genome replication in neurons. This suggests that neurons can be infected by many HSV virions and still enter the latent state.Journal of Virology 02/2000; 74(2):965-74. · 5.40 Impact Factor -
Article: Herpes simplex virus alpha protein ICP27 possesses separable positive and negative regulatory activities.
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ABSTRACT: The HSV-1 alpha (immediate-early) protein ICP27 expressed in transfected cells can activate the expression of certain HSV-1 promoters as well as inhibit the transactivated expression of others. We constructed a set of plasmids encoding mutant ICP27 molecules truncated at their carboxyl termini and used transfection assays to determine the functional properties of the mutant proteins. A polypeptide containing the amino-terminal 263 amino acid residues of ICP27 retained partial ability to activate gene expression but was unable to inhibit transactivation. Mutant proteins possessing 406 or 504 amino acids of ICP27 were unable to activate gene expression but retained full ability to inhibit transactivation. These results define two separable regulatory activities of ICP27, one positive and one negative, which can modulate gene expression in transfected cells. Immunoblot and immunofluorescence experiments were used to study the immunological reactivities and intracellular localizations of the mutant proteins. All proteins possessing the amino-terminal 263 amino acids of ICP27 reacted with an ICP27-specific monoclonal antibody and were localized to the cell nucleus. The mutant proteins, however, exhibited a number of phenotypes with regard to intranuclear localization. A mutant possessing 504 residues of ICP27 was similar to the wild-type protein in apparently localizing to all regions of the nucleus. A mutant containing 406 residues of ICP27, on the other hand, was mostly excluded from the nucleolar regions, while a 263-residue mutant was localized predominantly in the nucleoli. Thus, some aspect of ICP27 structure or function can dramatically affect its intranuclear distribution.Journal of Virology 09/1989; 63(8):3399-407. · 5.40 Impact Factor -
Article: Transcription of the derepressed open reading frame P of herpes simplex virus 1 precludes the expression of the antisense gamma(1)34.5 gene and may account for the attenuation of the mutant virus.
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ABSTRACT: Open reading frame P (ORF P), located at the 3' terminus of the 8.5-kb DNA sequence transcribed during latency and almost completely antisense to the gamma(1)34.5 gene, is naturally repressed by infected cell protein 4 (ICP4), the major herpes simplex virus 1 regulatory protein. Earlier studies on cells infected with a mutant in which the expression of ORF P is derepressed have shown that (i) the accumulation of the alpha infected cell proteins 0 (ICP0) and 22 (ICP22), the products of spliced mRNAs, is reduced congruent with the binding of ORF P protein to p32, a component of the ASF/SF2 splicing factors, (ii) ORF P protein colocalizes with spliceosomes, (iii) both gamma(1)34.5 mRNA and protein are virtually undetectable, and (iv) the virus is attenuated on intracerebral inoculation in mice. We report the construction and characterization of two recombinant viruses: R7546, in which ORF P transcription was derepressed and the initiator methionine codon was replaced; and R7547, in which both mutations were repaired to the wild-type genotype. The mutations in R7546 do not alter the amino acid sequence of the gamma(1)34.5 gene. We report that (i) the reduction in the accumulation of gamma(1)34.5 mRNA and protein in cells infected with mutant viruses expressing derepressed ORF P genes reflects the effects of antisense transcription of ORF P rather than a function of ORF P protein, (ii) the attenuated phenotype of the viruses carrying derepressed ORF P genes is due largely to the absence of the gamma(1)34.5 protein, and (iii) the reduction in accumulation of ICP0 and ICP22 requires the expression of ORF P protein.Journal of Virology 11/1997; 71(10):7750-7. · 5.40 Impact Factor
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Keywords
100 transcripts
57 human genes
75-mer oligonucleotides specific
alpha27 gene
cellular gene expression
cellular transcripts
essential immediate-early alpha27
excellent correlation
herpes simplex virus
HSV infection
immediate-early transcripts
individual viral transcripts
latent infections
others function
particular kinetic class
single transcripts
various abundances
various conditions
viral regulatory genes
viral transcripts
