The Human Cytomegalovirus 86-Kilodalton Major Immediate-Early Protein Interacts Physically and Functionally with Histone Acetyltransferase P/CAF

ABSTRACT The major immediate-early proteins of human cytomegalovirus (HCMV) play a pivotal role in controlling viral and cellular gene expression during productive infection. As well as negatively autoregulating its own promoter, the HCMV 86-kDa major immediate early protein (IE86) activates viral early gene expression and is known to be a promiscuous transcriptional regulator of cellular genes. IE86 appears to act as a multimodal transcription factor. It is able to bind directly to target promoters to activate transcription but is also able to bridge between upstream binding factors such as CREB/ATF and the basal transcription complex as well as interacting directly with general transcription factors such as TATA-binding protein and TFIIB. We now show that IE86 is also able to interact directly with histone acetyltransferases during infection. At least one of these factors is the histone acetyltransferase CBP-associated factor (P/CAF). Furthermore, we show that this interaction results in synergistic transactivation by IE86 of IE86-responsive promoters. Recruitment of such chromatin-remodeling factors to target promoters by IE86 may help explain the ability of this viral protein to act as a promiscuous transactivator of cellular genes.

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Human cytomegalovirus US3 and UL36-38 immediate-early proteins regulate gene expression.

Journal of Virology ◽

10.1128/jvi.66.1.95-105.1992 ◽

1992 ◽

Vol 66 (1) ◽

pp. 95-105 ◽

Cited By ~ 61

Author(s):

A M Colberg-Poley ◽

L D Santomenna ◽

P P Harlow ◽

P A Benfield ◽

D J Tenney

Keyword(s):

Gene Expression ◽

Human Cytomegalovirus ◽

Immediate Early ◽

Regulate Gene Expression ◽

Early Proteins ◽

Immediate Early Proteins ◽

Regulate Gene

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Infection of Cells with Human Cytomegalovirus during S Phase Results in a Blockade to Immediate-Early Gene Expression That Can Be Overcome by Inhibition of the Proteasome

Journal of Virology ◽

10.1128/jvi.76.11.5369-5379.2002 ◽

2002 ◽

Vol 76 (11) ◽

pp. 5369-5379 ◽

Cited By ~ 54

Author(s):

Elizabeth A. Fortunato ◽

Veronica Sanchez ◽

Judy Y. Yen ◽

Deborah H. Spector

Keyword(s):

Gene Expression ◽

Dna Replication ◽

Human Cytomegalovirus ◽

Dna Content ◽

Laser Scanning ◽

Small Population ◽

S Phase ◽

Early Gene ◽

Immediate Early ◽

Laser Scanning Cytometry

ABSTRACT Cells infected with human cytomegalovirus (HCMV) after commencing DNA replication do not initiate viral immediate-early (IE) gene expression and divide before arresting. To determine the nature of this blockade, we examined cells that were infected 24 h after release from G0 using immunofluorescence, laser scanning cytometry, and fluorescence-activated cell sorting (FACS) analysis. Approximately 40 to 50% of the cells had 2N DNA content, became IE+ in the first 12 h, and arrested. Most but not all of the cells with >2N DNA content did not express IE antigens until after mitosis. To define the small population of IE+ cells that gradually accumulated within the S and G2/M compartments, cells were pulsed with bromodeoxyuridine (BrdU) just prior to S-phase infection and analyzed at 12 h postinfection for IE gene expression, BrdU positivity, and cell cycle position. Most of the BrdU+ cells were IE− and had progressed into G2/M or back to G1. The majority of the IE+ cells in S and G2/M were BrdU−. Only a few cells were IE+ BrdU+, and they resided in G2/M. Multipoint BrdU pulse-labeling revealed that, compared to cells actively synthesizing DNA at the beginning of the infection, a greater percentage of the cells that initiated DNA replication 4 h later could express IE antigens and proceed into S. Synchronization of the cells with aphidicolin also indicated that the blockade to the activation of IE gene expression was established in cells soon after initiation of DNA replication. It appears that a short-lived protein in S-phase cells may be required for IE gene expression, as it is partially restored by treatment with the proteasome inhibitor MG132.

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Inhibition of human cytomegalovirus immediate-early gene expression by an antisense oligonucleotide complementary to immediate-early RNA.

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.40.9.2004 ◽

1996 ◽

Vol 40 (9) ◽

pp. 2004-2011 ◽

Cited By ~ 97

Author(s):

K P Anderson ◽

M C Fox ◽

V Brown-Driver ◽

M J Martin ◽

R F Azad

Keyword(s):

Gene Expression ◽

Cell Culture ◽

Antiviral Activity ◽

Protein Expression ◽

Human Cytomegalovirus ◽

Host Cells ◽

Early Gene ◽

Immediate Early ◽

Mrna Levels ◽

Virus Adsorption

ISIS 2922 is a phosphorothioate oligonucleotide that is complementary to human cytomegalovirus (CMV) immediate-early (IE) RNA and that exhibits potent and specific antiviral activity against CMV in cell culture assays. Specific assay systems were developed to separately characterize the antisense and nonantisense components of the antiviral activity mediated by ISIS 2922. In U373 cells transformed with cDNA encoding the CMV IE 55-kDa (IE55) protein, expression was inhibited at nanomolar concentrations comparable to effective concentrations in antiviral assays. The specificity of inhibition was demonstrated by using control oligonucleotides incorporating progressive base changes to destabilize oligonucleotide-RNA base pairing and by showing a lack of inhibition of the CMV IE72 product expressed from the same promoter. Inhibition of IE55 protein expression correlated with a reduction in mRNA levels consistent with an RNase H-mediated termination event. Studies with virus-infected cells demonstrated that antisense and nonantisense mechanisms contribute to the antiviral activity of ISIS 2922. Base complementarity to target RNA was important for optimal activity in antiviral assays, but base changes affecting parameters other than hybridization affinity also influenced antiviral activity. Sequence-independent inhibition of virus adsorption to host cells by phosphorothioate oligonucleotides was also observed at high concentrations. Therefore, at least three different mechanisms may contribute to the antiviral activity of ISIS 2922 in cell culture: antisense-mediated inhibition of target gene expression; nonantisense, sequence-dependent inhibition of virus replication; and sequence-independent inhibition of virus adsorption to host cells.

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Repression of human cytomegalovirus major immediate early gene expression in a monocytic cell line

Journal of General Virology ◽

10.1099/0022-1317-73-2-433 ◽

1992 ◽

Vol 73 (2) ◽

pp. 433-435 ◽

Cited By ~ 60

Author(s):

J. H. Sinclair ◽

J. Baillie ◽

L. A. Bryant ◽

J. A. Taylor-Wiedeman ◽

J. G. P. Sissons

Keyword(s):

Gene Expression ◽

Cell Line ◽

Human Cytomegalovirus ◽

Immediate Early Gene ◽

Early Gene ◽

Immediate Early ◽

Monocytic Cell ◽

Monocytic Cell Line ◽

Early Gene Expression

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The human cytomegalovirus US3 immediate-early protein lacking the putative transmembrane domain regulates gene expression

Nucleic Acids Research ◽

10.1093/nar/21.12.2931 ◽

1993 ◽

Vol 21 (12) ◽

pp. 2931-2937 ◽

Cited By ~ 23

Author(s):

Daniel J. Tenney ◽

Linda D. Santomenna ◽

Karyn B. Goudie ◽

Anamaris M. Colberg-Poley

Keyword(s):

Gene Expression ◽

Human Cytomegalovirus ◽

Transmembrane Domain ◽

Immediate Early ◽

Putative Transmembrane Domain ◽

Early Protein

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Daxx-Mediated Accumulation of Human Cytomegalovirus Tegument Protein pp71 at ND10 Facilitates Initiation of Viral Infection at These Nuclear Domains

Journal of Virology ◽

10.1128/jvi.76.15.7705-7712.2002 ◽

2002 ◽

Vol 76 (15) ◽

pp. 7705-7712 ◽

Cited By ~ 86

Author(s):

Alexander M. Ishov ◽

Olga V. Vladimirova ◽

Gerd G. Maul

Keyword(s):

Human Cytomegalovirus ◽

Coiled Coil ◽

Immediate Early ◽

Productive Infection ◽

Tegument Protein ◽

Knockout Mutant ◽

Nuclear Site ◽

Immediate Early Proteins ◽

Nuclear Domains

ABSTRACT Human cytomegalovirus (HCMV) starts immediate-early transcription at nuclear domains 10 (ND10), forming a highly dynamic immediate transcript environment at this nuclear site. The reason for this spatial correlation remains enigmatic, and the mechanism for induction of transcription at ND10 is unknown. We investigated whether tegument-based transactivators are involved in the specific intranuclear location of HCMV. Here, we demonstrate that the HCMV transactivator tegument protein pp71 accumulates at ND10 before the production of immediate-early proteins. Intracellular trafficking of pp71 is facilitated through binding to a coiled-coil region of Daxx. The C-terminal domain of Daxx then interacts with SUMO-modified PML, resulting in the deposition of pp71 at ND10. In Daxx-deficient cells, pp71 does not accumulate at ND10, proving in vivo the necessity of Daxx for pp71 deposition. Also, HCMV forms immediate transcript environments at sites other than ND10 in Daxx-deficient cells, and so does the HCMV pp71 knockout mutant UL82−/− in normal cells. This result strongly suggests that pp71 and Daxx are essential for HCMV transcription at ND10. Lack of Daxx had the effect of reducing the infection rate. We conclude that the tegument transactivator pp71 facilitates viral genome deposition and transcription at ND10, possibly priming HCMV for more efficient productive infection.

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The Immediate Early Gene 1 Product of Human Cytomegalovirus Is Sufficient for Up-Regulation of Interleukin-8 Gene Expression

Biochemical and Biophysical Research Communications ◽

10.1006/bbrc.2000.3923 ◽

2000 ◽

Vol 279 (1) ◽

pp. 298-304 ◽

Cited By ~ 37

Author(s):

Tsugiya Murayama ◽

Naofumi Mukaida ◽

Hidetaka Sadanari ◽

Nobuo Yamaguchi ◽

Khalid S.A. Khabar ◽

...

Keyword(s):

Gene Expression ◽

Human Cytomegalovirus ◽

Immediate Early Gene ◽

Interleukin 8 ◽

Early Gene ◽

Immediate Early

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Dysregulation of Cyclin E Gene Expression in Human Cytomegalovirus-Infected Cells Requires Viral Early Gene Expression and Is Associated with Changes in the Rb-Related Protein p130

Journal of Virology ◽

10.1128/jvi.74.9.4192-4206.2000 ◽

2000 ◽

Vol 74 (9) ◽

pp. 4192-4206 ◽

Cited By ~ 33

Author(s):

Anita K. McElroy ◽

Roopashree S. Dwarakanath ◽

Deborah H. Spector

Keyword(s):

Gene Expression ◽

Cell Cycle ◽

Human Cytomegalovirus ◽

Cyclin E ◽

Early Gene ◽

Immediate Early ◽

Mrna Levels ◽

Early Gene Expression ◽

Cell Extracts ◽

Infected Cells

ABSTRACT We have previously shown that many cell cycle regulatory gene products are markedly affected by infection of primary fibroblasts with human cytomegalovirus (HCMV) (F. M. Jault, J. M. Jault, F. Ruchti, E. A. Fortunato, C. Clark, J. Corbeil, D. D. Richman, and D. H. Spector, J. Virol. 69:6697–6704, 1995). One of these proteins, cyclin E, is a key determinant of cell cycle progression during G1, and its mRNA levels are significantly increased in HCMV-infected fibroblasts (B. S. Salvant, E. A. Fortunato, and D. H. Spector, J. Virol. 72:3729–3741, 1998). To determine the molecular basis of this effect, we have examined the events that occur at the endogenous cyclin E promoter during the course of infection. In vivo dimethyl sulfate footprinting of the cyclin E promoter revealed several regions of protection and hypersensitivity that were unique to infected cells. In accord with this observation, we find that the virus-induced cyclin E transcripts initiate downstream of the start site identified in mock-infected cells, in regions where these newly appearing protected and hypersensitive sites occur. Viral gene expression is required for this induction. However, the viral immediate-early proteins IE1-72 and IE2-86, either alone or in combination, cannot induce expression of the endogenous cyclin E. The virus must progress past the immediate-early phase and express an early gene product(s) for activation of cyclin E expression. Moreover, IE1-72 does not appear to be required, as infection of cells with an HCMV mutant containing a deletion in the IE1-72 gene leads to full upregulation of cyclin E expression. Using electrophoretic mobility shift assays with infected cell extracts and a region of the cyclin E promoter that includes two previously defined E2F sites as the probe, we detected the appearance of an infection-specific banding pattern. One of the infection-specific bands contained the proteins E2F-4, DP-1, and p130, which were maintained in the infected cells as uniquely phosphorylated species. These results suggest that an altered E2F-4–DP-1–p130 complex along with viral early gene expression may play a role in the transcriptional regulation of cyclin E mRNA during HCMV infection.

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