scholarly journals Cell Cycle-Independent Expression of Immediate-Early Gene 3 Results in G1 and G2 Arrest in Murine Cytomegalovirus-Infected Cells

2008 ◽  
Vol 82 (20) ◽  
pp. 10188-10198 ◽  
Author(s):  
Lüder Wiebusch ◽  
Anke Neuwirth ◽  
Linus Grabenhenrich ◽  
Sebastian Voigt ◽  
Christian Hagemeier
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Viral Gene Expression ◽  
Immediate Early Gene ◽  
Early Gene ◽  
Immediate Early ◽  
Viral Gene ◽  
Cycle Arrest ◽  
Infected Cells

ABSTRACT The infectious cycle of human cytomegalovirus (HCMV) is intricately linked to the host's cell cycle. Viral gene expression can be initiated only in G0/G1 phase. Once expressed, the immediate-early gene product IE2 prevents cellular DNA synthesis, arresting infected cells with a G1 DNA content. This function is required for efficient viral replication in vitro. A prerequisite for addressing its in vivo relevance is the characterization of cell cycle-regulatory activities of CMV species for which animal models have been established. Here, we show that murine CMV (MCMV), like HCMV, has a strong antiproliferative capacity and arrests cells in G1. Unexpectedly, and in contrast to HCMV, MCMV can also block cells that have passed through S phase by arresting them in G2. Moreover, MCMV can also replicate in G2 cells. This is made possible by the cell cycle-independent expression of MCMV immediate-early genes. Transfection experiments show that of several MCMV candidate genes, only immediate-early gene 3 (ie3), the homologue of HCMV IE2, exhibits cell cycle arrest activity. Accordingly, an MCMV ie3 deletion mutant has lost the ability to arrest cells in either G1 or G2. Thus, despite interspecies variations in the cell cycle dependence of viral gene expression, the central theme of HCMV IE2-induced cell cycle arrest is conserved in the murine counterpart, raising the possibility of studying its physiological relevance at the level of the whole organism.

scholarly journals HIV-1 Vpr induces cell cycle arrest and enhances viral gene expression by depleting CCDC137

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Fengwen Zhang ◽  
Paul D Bieniasz
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Biological Effects ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Dependent Manner ◽  
M Cell ◽  
Cycle Arrest ◽  
Hiv 1

The HIV-1 Vpr accessory protein induces ubiquitin/proteasome-dependent degradation of many cellular proteins by recruiting them to a cullin4A-DDB1-DCAF1 complex. In so doing, Vpr enhances HIV-1 gene expression and induces (G2/M) cell cycle arrest. However, the identities of Vpr target proteins through which these biological effects are exerted are unknown. We show that a chromosome periphery protein, CCDC137/cPERP-B, is targeted for depletion by HIV-1 Vpr, in a cullin4A-DDB1-DCAF1 dependent manner. CCDC137 depletion caused G2/M cellcycle arrest, while Vpr-resistant CCDC137 mutants conferred resistance to Vpr-induced G2/M arrest. CCDC137 depletion also recapitulated the ability of Vpr to enhance HIV-1 gene expression, particularly in macrophages. Our findings indicate that Vpr promotes cell-cycle arrest and HIV-1 gene expression through depletion of CCDC137.

scholarly journals HIV-1 Vpr induces cell cycle arrest and enhances viral gene expression by depleting CCDC137

2019 ◽  
Author(s):  
Fengwen Zhang ◽  
Paul D. Bieniasz
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Cell Cycle Arrest ◽  
Biological Effects ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Dependent Manner ◽  
M Cell ◽  
Cycle Arrest ◽  
Hiv 1

SummaryThe HIV-1 Vpr accessory protein induces ubiquitin/proteasome-dependent degradation of many cellular proteins by recruiting them to a cullin4A-DDB1-DCAF1 complex. In so doing, Vpr enhances HIV-1 gene expression and induces (G2/M) cell cycle arrest. However, the identities of Vpr target proteins through which these biological effects are exerted are unknown. We show that a chromosome periphery protein, CCDC137/cPERP-B, is targeted for depletion by HIV-1 Vpr, in a cullin4A-DDB1-DCAF1 dependent manner. CCDC137 depletion caused G2/M cell-cycle arrest, while Vpr-resistant CCDC137 mutants conferred resistance to Vpr-induced G2/M arrest. CCDC137 depletion also recapitulated the ability of Vpr to enhance HIV-1 gene expression, particularly in macrophages. Our findings indicate that Vpr promotes cell-cycle arrest and HIV-1 gene expression through depletion of CCDC137.

scholarly journals 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

2000 ◽  
Vol 74 (9) ◽  
pp. 4192-4206 ◽  
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.

scholarly journals Cell-to-cell and genome-to-genome variability of adenovirus transcription tuned by the cell cycle

2020 ◽  
Vol 134 (5) ◽  
pp. jcs252544
Author(s):  
Maarit Suomalainen ◽  
Vibhu Prasad ◽  
Abhilash Kannan ◽  
Urs F. Greber
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Single Molecule ◽  
Click Reaction ◽  
Viral Gene Expression ◽  
Early Gene ◽  
Viral Gene ◽  
Viral Genomes ◽  
E1a Gene ◽  
G1 Cells

ABSTRACTIn clonal cultures, not all cells are equally susceptible to virus infection, and the mechanisms underlying this are poorly understood. Here, we developed image-based single-cell measurements to scrutinize the heterogeneity of adenovirus (AdV) infection. AdV delivers, transcribes and replicates a linear double-stranded DNA genome in the nucleus. We measured the abundance of viral transcripts using single-molecule RNA fluorescence in situ hybridization (FISH) and the incoming 5-ethynyl-2′-deoxycytidine (EdC)-tagged viral genomes using a copper(I)-catalyzed azide–alkyne cycloaddition (click) reaction. Surprisingly, expression of the immediate early gene E1A only moderately correlated with the number of viral genomes in the cell nucleus. Intranuclear genome-to-genome heterogeneity was found at the level of viral transcription and, in accordance, individual genomes exhibited heterogeneous replication activity. By analyzing the cell cycle state, we found that G1 cells exhibited the highest E1A gene expression and displayed increased correlation between E1A gene expression and viral genome copy numbers. The combined image-based single-molecule procedures described here are ideally suited to explore the cell-to-cell variability in viral gene expression in a range of different settings, including the innate immune response.

scholarly journals Small Internal Deletions in the Human Cytomegalovirus IE2 Gene Result in Nonviable Recombinant Viruses with Differential Defects in Viral Gene Expression

2004 ◽  
Vol 78 (4) ◽  
pp. 1817-1830 ◽  
Author(s):  
Elizabeth A. White ◽  
Charles L. Clark ◽  
Veronica Sanchez ◽  
Deborah H. Spector
Keyword(s):  
Gene Expression ◽  
Amino Acids ◽  
Human Cytomegalovirus ◽  
Viral Gene Expression ◽  
Artificial Chromosome ◽  
Early Gene ◽  
Immediate Early ◽  
Viral Gene ◽  
Recombinant Viruses ◽  
Late Genes

ABSTRACT The human cytomegalovirus (HCMV) IE2 86-kDa protein is a key viral transactivator and an important regulator of HCMV infections. We used the HCMV genome cloned as a bacterial artificial chromosome (BAC) to construct four HCMV mutants with disruptions in regions of IE2 86 that are predicted to be important for its transactivation and autoregulatory functions. Three of these mutants have mutations that remove amino acids 356 to 359, 427 to 435, and 505 to 511, which disrupts a region of IE2 86 implicated in the activation of HCMV early promoters, a predicted zinc finger domain, and a putative helix-loop-helix motif, respectively, while the fourth carries three arginine-to-alanine substitution mutations in the region of amino acids 356 to 359. The resulting recombinant viruses are not viable, and by using quantitative real-time reverse transcription-PCR and immunofluorescence we have determined the location of the block in their replicative cycles. The IE2 86Δ356-359 mutant is able to support early gene expression, as indicated by the presence of UL112-113 transcripts and UL112-113 and UL44 proteins in cells transfected with the mutant BAC. This mutant does not express late genes and behaves nearly indistinguishably from the IE2 86R356/7/9A substitution mutant. Both exhibit detectable upregulation of major immediate-early transcripts at early times. The IE2 86Δ427-435 and IE2 86Δ505-511 recombinant viruses do not activate the early genes examined and are defective in repression of the major immediate-early promoter. These two mutants also induce the expression of selected delayed early (UL89) and late genes at early times in the infection. We conclude that these three regions of IE2 86 are necessary for productive infections and for differential control of downstream viral gene expression.

scholarly journals Inactivating a Cellular Intrinsic Immune Defense Mediated by Daxx Is the Mechanism through Which the Human Cytomegalovirus pp71 Protein Stimulates Viral Immediate-Early Gene Expression

2006 ◽  
Vol 80 (8) ◽  
pp. 3863-3871 ◽  
Author(s):  
Ryan T. Saffert ◽  
Robert F. Kalejta
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Immediate Early Gene ◽  
Nuclear Body ◽  
Immune Defense ◽  
Early Gene ◽  
Immediate Early ◽  
Innate Immune Responses ◽  
Early Gene Expression ◽  
Infected Cells

ABSTRACT Human cytomegalovirus (HCMV) masterfully evades adaptive and innate immune responses, allowing infection to be maintained and periodically reactivated for the life of the host. Here we show that cells also possess an intrinsic immune defense against HCMV that is disarmed by the virus. In HCMV-infected cells, the promyelocytic leukemia nuclear body (PML-NB) protein Daxx silences viral immediate-early gene expression through the action of a histone deacetylase. However, this antiviral tactic is efficiently neutralized by the viral pp71 protein, which is incorporated into virions, delivered to cells upon infection, and mediates the proteasomal degradation of Daxx. This work demonstrates the mechanism through which pp71 activates viral immediate-early gene expression in HCMV-infected cells. Furthermore, it provides insight into how a PML-NB protein institutes an intrinsic immune defense against a DNA virus and how HCMV pp71 inactivates this defense.

scholarly journals Human cytomegalovirus immediate-early gene expression is restricted by the nuclear domain 10 component Sp100

2011 ◽  
Vol 92 (7) ◽  
pp. 1532-1538 ◽  
Author(s):  
Martina Adler ◽  
Nina Tavalai ◽  
Regina Müller ◽  
Thomas Stamminger
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Viral Gene Expression ◽  
Early Gene ◽  
Immediate Early ◽  
Viral Gene ◽  
Major Protein ◽  
Hcmv Replication ◽  
Protein Components ◽  
Nuclear Domains

Nuclear domains 10 (ND10s) are discrete subnuclear structures that contain the three major protein components promyelocytic leukaemia protein (PML), hDaxx and Sp100. Previous studies identified the ND10-components PML and hDaxx as cellular restriction factors that independently counteract human cytomegalovirus (HCMV) infection via the repression of viral immediate-early (IE) gene expression. Consequently, we asked whether Sp100 is likewise involved in this repressive activity. Infection of Sp100 knockdown (kd) cells with HCMV resulted in a significantly increased plaque-forming ability. In addition, ablation of Sp100 led to a considerable increase in the number of IE1-expressing cells, indicating that Sp100 suppresses the initiation of viral gene expression. Next, double-kd cells, lacking either Sp100/hDaxx or Sp100/PML, were generated. Here, infection resulted in an additional enhancement in HCMV replication efficacy compared with the single-kd cells. Thus, our results further strengthen the concept that the three major ND10-components independently contribute to the cellular restriction of HCMV replication.

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