scholarly journals EGR1 transcriptional control of human cytomegalovirus latency

2019 ◽  
Author(s):  
Jason Buehler ◽  
Ethan Carpenter ◽  
Sebastian Zeltzer ◽  
Suzu Igarashi ◽  
Michael Rak ◽  
...  
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Human Cytomegalovirus ◽  
Transcriptional Control ◽  
Viral Reactivation ◽  
Erk Signaling ◽  
Viral Gene ◽  
Egfr Signaling ◽  
Down Regulation ◽  
Cmv Reactivation

ABSTRACTSustained phosphotinositide3-kinase (PI3K) signaling is critical to the maintenance of herpesvirus latency. We have previously shown that the beta-herpesvirus, human cytomegalovirus (CMV), regulates epidermal growth factor receptor (EGFR), upstream of PI3K, to control states of latency and reactivation. Inhibition of EGFR signaling enhances CMV reactivation from latency and increases viral replication, but the mechanisms by which EGFR impacts replication and latency is not known. We demonstrate that HCMV downregulates MEK/ERK and AKT phosphorylation, but not STAT3 or PLCγ for productive replication. Similarly, inhibition of either MEK/ERK or PI3K/AKT, but not STAT or PLCγ, pathways increases viral reactivation from latently infected CD34+hematopoietic progenitor cells (HPCs), defining a role for these pathways in latency. We hypothesized that CMV modulation of EGFR signaling might impact viral transcription. Indeed, EGF-stimulation increased expression of theUL138latency gene, but not immediate early or early viral genes, suggesting that EGFR signaling promotes latent gene expression. The early growth response-1 (EGR1) transcription factor is induced downstream of EGFR signaling through both PI3K/AKT and MEK/ERK pathways. EGR1 expression is important for the maintenance of HPC stemness and its downregulation drives HPC differentiation and mobilization. We demonstrate that EGR1 binds upstream ofUL138and is sufficient to promoteUL138expression. Further, disruption of EGR1 binding upstream ofUL138prevented CMV from establishing a latent infection in CD34+HPCs. Our results indicate a model whereby UL138 modulation of EGFR signaling feeds back to promote UL138 expression and suppression of replication to establish or maintain viral quiescence.AUTHOR SUMMARYCMV regulates EGFR signaling to balance states of viral latency and replication. CMV blocks downstream PI3K/AKT and MEK/ERK signaling pathways through down-regulation of EGFR at the plasma membrane. PI3K/AKT and MEK/ERK signaling increases expression of the EGR1 transcription factor that is necessary for the maintenance of stem cell stemness. A decrease in EGR1 expression promotes HPC mobilization to the periphery and differentiation, a known stimulus for CMV reactivation. We identified functional EGR1 binding sites upstream of theUL138gene and EGR-1 binding stimulatesUL138expression. Additionally, down-regulation of EGR1 by CMV miR-US22 decreasesUL138expression emphasizing the importance of this transcription factor in expression of this latency gene. Lastly, we demonstrate that a CMV mutant virus lacking an upstream EGR1 binding site is unable to establish latency in CD34+HPCs. This study defines one mechanism by which EGFR signaling impacts viral gene expression to promote CMV latency.

scholarly journals Human cytomegalovirus transcriptome activity differs during replication in human fibroblast, epithelial and astrocyte cell lines

2012 ◽  
Vol 93 (5) ◽  
pp. 1046-1058 ◽  
Author(s):  
James C. Towler ◽  
Bahram Ebrahimi ◽  
Brian Lane ◽  
Andrew J. Davison ◽  
Derrick J. Dargan
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Growth Kinetics ◽  
Viral Gene Expression ◽  
Cell Types ◽  
Viral Gene ◽  
Genome Replication ◽  
Cell Type ◽  
Low Passage ◽  
Temporal Expression Pattern

Broad cell tropism contributes to the pathogenesis of human cytomegalovirus (HCMV), but the extent to which cell type influences HCMV gene expression is unclear. A bespoke HCMV DNA microarray was used to monitor the transcriptome activity of the low passage Merlin strain of HCMV at 12, 24, 48 and 72 h post-infection, during a single round of replication in human fetal foreskin fibroblast cells (HFFF-2s), human retinal pigmented epithelial cells (RPE-1s) and human astrocytoma cells (U373MGs). In order to correlate transcriptome activity with concurrent biological responses, viral cytopathic effect, growth kinetics and genomic loads were examined in the three cell types. The temporal expression pattern of viral genes was broadly similar in HFFF-2s and RPE-1s, but dramatically different in U373MGs. Of the 165 known HCMV protein-coding genes, 41 and 48 were differentially regulated in RPE-1s and U373MGs, respectively, compared with HFFF-2s, and 22 of these were differentially regulated in both RPE-1s and U373MGs. In RPE-1s, all differentially regulated genes were downregulated, but, in U373MGs, some were down- and others upregulated. Differentially regulated genes were identified among the immediate-early, early, early late and true-late viral gene classes. Grouping of downregulated genes according to function at landmark stages of the replication cycle led to the identification of potential bottleneck stages (genome replication, virion assembly, and virion maturation and release) that may account for cell type-dependent viral growth kinetics. The possibility that cell type-specific differences in expressed cellular factors are responsible for modulation of viral gene expression is discussed.

scholarly journals The AP-1 Transcription Factor Is a Key Determinant of Human Cytomegalovirus Latency and Reactivation

Proceedings ◽  
2020 ◽  
Vol 50 (1) ◽  
pp. 127
Author(s):  
Benjamin A. Krishna ◽  
Amanda B. Wass ◽  
Christine M. O’Connor
Keyword(s):  
Transcription Factor ◽  
Human Cytomegalovirus ◽  
Infected Individual ◽  
Virus Production ◽  
Mapk Signaling ◽  
Viral Reactivation ◽  
Immediate Early ◽  
Activator Protein 1 ◽  
Latently Infected ◽  
Signaling Axis

Human cytomegalovirus (HCMV) is a ubiquitous pathogen that latently infects hematopoietic progenitor cells (HPCs). Individuals with a competent immune system are, for the most part, asymptomatic for the disease. However when a latently infected individual becomes immunosuppressed, HCMV can reactivate, causing severe morbidity and mortality. While much of the viral genome is transcriptionally silenced during latency, some genes are expressed, including the HCMV-encoded G-protein coupled receptor US28. We showed that US28 expression is required for latency, as it suppressed the activator protein-1 (AP-1) transcription factor by attenuating the AP-1 subunit, fos. In turn, this prevents AP-1 from binding and activating the major immediate early promoter (MIEP), the key promoter regulating the latent-to-lytic transcriptional “switch”. Our new data suggest that US28-mediated signaling during latency attenuates the Src-MAPK signaling axis to regulate AP-1. We find that US28 expression suppresses Src, MEK, and ERK, as well as fos phosphorylation and AP-1 binding to the MIEP. Conversely, the pharmacological inhibition of Src, MEK, or ERK in US28Δ-latently infected HPCs suppresses infectious virus production, demonstrating the important role for this signaling axis during latency. Our recent data also reveal that regulating AP-1 is a key determinant in balancing HCMV latency and reactivation. Infection with a virus in which we disrupted the proximal AP-1 binding site in the MIEP (AP-1Δp) leads to reduced AP-1 binding and inefficient viral reactivation compared to wild type. Furthermore, AP-1 is critical for the de-repression of MIEP-driven transcripts and downstream early and late genes, while other immediate early genes remain unaffected. Collectively, these data suggest that AP-1 binding to the MIEP is suppressed during latency, but is required for the efficient transactivation of the MIEP during reactivation. We are currently elucidating US28’s involvement in recruiting AP-1 to the MIEP during reactivation.

scholarly journals Interaction between STAT-3 and HNF-3 Leads to the Activation of Liver-Specific Hepatitis B Virus Enhancer 1 Function

2002 ◽  
Vol 76 (6) ◽  
pp. 2721-2729 ◽  
Author(s):  
Gulam Waris ◽  
Aleem Siddiqui
Keyword(s):  
Gene Expression ◽  
Hepatitis B Virus ◽  
Hepatitis B ◽  
Transcriptional Control ◽  
Viral Gene Expression ◽  
Cooperative Interaction ◽  
Viral Gene ◽  
Mobility Shift ◽  
B Virus ◽  
Stimulation Of

ABSTRACT The signal transducer and activator of transcription 3 (STAT-3), a member of the STAT family of proteins, binds to a large number of transcriptional control elements and regulates gene expression in response to cytokines. While it binds to its cognate nucleotide sequences, it has been recently shown to directly interact with other transcriptional factors in the absence of DNA. We report here one such novel interaction between STAT-3 and hepatocyte nuclear factor 3 (HNF-3) in the absence of DNA. We have identified a STAT-3 binding site within the core domain of hepatitis B virus (HBV) enhancer 1. The HBV enhancer 1 DNA-STAT-3 protein interaction is shown to be stimulated by interleukin-6 (IL-6) and epidermal growth factor, which leads to an overall stimulation of HBV enhancer 1 function and viral gene expression. Using mobility shift assays and transient transfection schemes, we demonstrate a cooperative interaction between HNF-3 and STAT-3 in mediating the cytokine-mediated HBV enhancer function. Cytokine stimulation of HBV gene expression represents an important regulatory scheme of direct relevance to liver disease pathogenesis associated with HBV infection.

scholarly journals Lytic infection of permissive cells with human cytomegalovirus is regulated by an intrinsic ‘pre-immediate-early’ repression of viral gene expression mediated by histone post-translational modification

2009 ◽  
Vol 90 (10) ◽  
pp. 2364-2374 ◽  
Author(s):  
Ian J. Groves ◽  
Matthew B. Reeves ◽  
John H. Sinclair
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Chromatin Structure ◽  
Viral Gene Expression ◽  
Lytic Infection ◽  
Immediate Early ◽  
Late Gene ◽  
Viral Gene ◽  
Gene Promoters ◽  
Late Gene Expression

Human cytomegalovirus (HCMV) lytic gene expression occurs in a regulated cascade, initiated by expression of the viral major immediate-early (IE) proteins. Transcribed from the major IE promoter (MIEP), the major IE genes regulate viral early and late gene expression. This study found that a substantial proportion of infecting viral genomes became associated with histones immediately upon infection of permissive fibroblasts at low m.o.i. and these histones bore markers of repressed chromatin. As infection progressed, however, the viral MIEP became associated with histone marks, which correlate with the known transcriptional activity of the MIEP at IE time points. Interestingly, this chromatin-mediated repression of the MIEP at ‘pre-IE’ times of infection could be overcome by inhibition of histone deacetylases, as well as by infection at high m.o.i., and resulted in a temporal advance of the infection cycle by inducing premature viral early and late gene expression and DNA replication. As well as the MIEP, and consistent with previous observations, the viral early and late promoters were also initially associated with repressive chromatin. However, changes in histone modifications around these promoters also occurred as infection progressed, and this correlated with the known temporal regulation of the viral early and late gene expression cascade. These data argue that the chromatin structure of all classes of viral genes are initially repressed on infection of permissive cells and that the chromatin structure of HCMV gene promoters plays an important role in regulating the time course of viral gene expression during lytic infection.

scholarly journals Transcription factor IIIA gene expression in Xenopus oocytes utilizes a transcription factor similar to the major late transcription factor.

1989 ◽  
Vol 9 (11) ◽  
pp. 5003-5011 ◽  
Author(s):  
R K Hall ◽  
W L Taylor
Keyword(s):  
Gene Expression ◽  
Transcription Factor ◽  
Transcriptional Control ◽  
Somatic Cells ◽  
State Level ◽  
Control Element ◽  
Transcription Factor Iiia ◽  
Shift Analysis ◽  
Late Transcription ◽  
Upstream Element

Xenopus transcription factor IIIA (TFIIIA) gene expression is stringently regulated during development. The steady-state level of TFIIIA mRNA in a somatic cell is approximately 10(6) times less than in an immature oocyte. We have undertaken studies designed to identify differences in how the TFIIIA gene is transcribed in oocytes and somatic cells. In this regard, we have localized an upstream transcriptional control element in the TFIIIA promoter that stimulates transcription from the TFIIIA promoter approximately threefold in microinjected oocytes. The upstream element, in cis. does not stimulate transcription from the TFIIIA promoter in somatic cells. Thus, the element appears to be oocyte specific in the context of the TFIIIA promoter. However, both oocytes and somatic cells contain a protein (or a related protein) that binds the upstream element. We have termed this protein from oocytes the TFIIIA distal element factor. The sequence of the upstream element is similar to the sequence of the upstream element found in the adenovirus major late promoter that is a binding site for the major late transcription factor. By gel shift analysis, chemical footprinting, methylation intereference, and point mutation analysis, we demonstrate that the TFIIIA distal element factor and major late transcription factor have similar DNA-binding properties.

Cell-cycle-dependent localization of human cytomegalovirus UL83 phosphoprotein in the nucleolus and modulation of viral gene expression in human embryo fibroblasts in vitro

2011 ◽  
Vol 112 (1) ◽  
pp. 307-317 ◽  
Author(s):  
Maria-Cristina Arcangeletti ◽  
Isabella Rodighiero ◽  
Prisco Mirandola ◽  
Flora De Conto ◽  
Silvia Covan ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Human Cytomegalovirus ◽  
Human Embryo ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Embryo Fibroblasts ◽  
Cycle Dependent

scholarly journals MicroRNA miR-21 Attenuates Human Cytomegalovirus Replication in Neural Cells by Targeting Cdc25a

2014 ◽  
Vol 89 (2) ◽  
pp. 1070-1082 ◽  
Author(s):  
Ya-Ru Fu ◽  
Xi-Juan Liu ◽  
Xiao-Jun Li ◽  
Zhang-zhou Shen ◽  
Bo Yang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cell Cycle ◽  
Stem Cells ◽  
Birth Defects ◽  
Human Cytomegalovirus ◽  
Hcmv Infection ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Gene Products ◽  
Hcmv Replication

ABSTRACTCongenital human cytomegalovirus (HCMV) infection is a leading cause of birth defects, primarily manifesting as neurological disorders. HCMV infection alters expression of cellular microRNAs (miRs) and induces cell cycle arrest, which in turn modifies the cellular environment to favor virus replication. Previous observations found that HCMV infection reduces miR-21 expression in neural progenitor/stem cells (NPCs). Here, we show that infection of NPCs and U-251MG cells represses miR-21 while increasing the levels of Cdc25a, a cell cycle regulator and known target of miR-21. These opposing responses to infection prompted an investigation of the relationship between miR-21, Cdc25a, and viral replication. Overexpression of miR-21 in NPCs and U-251MG cells inhibited viral gene expression, genome replication, and production of infectious progeny, while shRNA-knockdown of miR-21 in U-251MG cells increased viral gene expression. In contrast, overexpression of Cdc25a in U-251MG cells increased viral gene expression and production of infectious progeny and overcame the inhibitory effects of miR-21 overexpression. Three viral gene products—IE1, pp71, and UL26—were shown to inhibit miR-21 expression at the transcriptional level. These results suggest that Cdc25a promotes HCMV replication and elevation of Cdc25a levels after HCMV infection are due in part to HCMV-mediated repression of miR-21. Thus, miR-21 is an intrinsic antiviral factor that is modulated by HCMV infection. This suggests a role for miR-21 downregulation in the neuropathogenesis of HCMV infection of the developing CNS.IMPORTANCEHuman cytomegalovirus (HCMV) is a ubiquitous pathogen and has very high prevalence among population, especially in China, and congenital HCMV infection is a major cause for birth defects. Elucidating virus-host interactions that govern HCMV replication in neuronal cells is critical to understanding the neuropathogenesis of birth defects resulting from congenital infection. In this study, we confirm that HCMV infection downregulates miR-21 but upregulates Cdc25a. Further determined the negative effects of cellular miRNA miR-21 on HCMV replication in neural progenitor/stem cells and U-251MG glioblastoma/astrocytoma cells. More importantly, our results provide the first evidence that miR-21 negatively regulates HCMV replication by targeting Cdc25a, a vital cell cycle regulator. We further found that viral gene products of IE1, pp71, and UL26 play roles in inhibiting miR-21 expression, which in turn causes increases in Cdc25a and benefits HCMV replication. Thus, miR-21 appears to be an intrinsic antiviral factor that represents a potential target for therapeutic intervention.

scholarly journals DNA Microarrays of the Complex Human Cytomegalovirus Genome: Profiling Kinetic Class with Drug Sensitivity of Viral Gene Expression

1999 ◽  
Vol 73 (7) ◽  
pp. 5757-5766 ◽  
Author(s):  
James Chambers ◽  
Ana Angulo ◽  
Dhammika Amaratunga ◽  
Hongqing Guo ◽  
Ying Jiang ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Dna Sequences ◽  
De Novo ◽  
Expression Profiles ◽  
Viral Gene Expression ◽  
Viral Gene ◽  
Viral Dna ◽  
Viral Protein Synthesis ◽  
Entire Genome

ABSTRACT We describe, for the first time, the generation of a viral DNA chip for simultaneous expression measurements of nearly all known open reading frames (ORFs) in the largest member of the herpesvirus family, human cytomegalovirus (HCMV). In this study, an HCMV chip was fabricated and used to characterize the temporal class of viral gene expression. The viral chip is composed of microarrays of viral DNA prepared by robotic deposition of oligonucleotides on glass for ORFs in the HCMV genome. Viral gene expression was monitored by hybridization to the oligonucleotide microarrays with fluorescently labelled cDNAs prepared from mock-infected or infected human foreskin fibroblast cells. By using cycloheximide and ganciclovir to block de novo viral protein synthesis and viral DNA replication, respectively, the kinetic classes of array elements were classified. The expression profiles of known ORFs and many previously uncharacterized ORFs provided a temporal map of immediate-early (α), early (β), early-late (γ1), and late (γ2) genes in the entire genome of HCMV. Sequence compositional analysis of the 5′ noncoding DNA sequences of the temporal classes, performed by using algorithms that automatically search for defined and recurring motifs in unaligned sequences, indicated the presence of potential regulatory motifs for β, γ1, and γ2 genes. In summary, these fabricated microarrays of viral DNA allow rapid and parallel analysis of gene expression at the whole viral genome level. The viral chip approach coupled with global biochemical and genetic strategies should greatly speed the functional analysis of established as well as newly discovered large viral genomes.

scholarly journals The KSHV ORF20 Protein Interacts With The Viral Processivity Factor ORF59 And Promote Viral Reactivation

2020 ◽  
Author(s):  
D. Hoffman ◽  
W. Rodriguez ◽  
D. Macveigh-Fierro ◽  
J. Miles ◽  
M. Muller
Keyword(s):  
Gene Expression ◽  
Viral Replication ◽  
Viral Proteins ◽  
Lytic Replication ◽  
Viral Reactivation ◽  
Wild Type Virus ◽  
Viral Gene ◽  
Late Gene Expression ◽  
Lytic Gene ◽  
Processivity Factor

AbstractUpon KSHV lytic reactivation, rapid and widespread amplification of viral DNA (vDNA) triggers significant nuclear reorganization. As part of this striking shift in nuclear architecture, viral replication compartments are formed as sites of lytic vDNA production along with remarkable spatial remodeling and relocalization of cellular and viral proteins. These viral replication compartments house several lytic gene products that coordinate viral gene expression, vDNA replication, and nucleocapsid assembly. The viral proteins and mechanisms that regulate this overhaul of the nuclear landscape during KSHV replication remain largely unknown. KSHV’s ORF20 is a widely conserved lytic gene among all herpesviruses suggesting it may have a fundamental contribution to the progression of herpesviral infection. Here, we utilized a promiscuous biotin ligase proximity labeling method to identify the proximal interactome of ORF20, which includes several replication-associated viral proteins, one of which is ORF59, the KSHV DNA processivity factor. Using co-immunoprecipitation and immunofluorescence assays, we confirmed the interaction between ORF20 and ORF59 and tracked the localization of both proteins to KSHV replication compartments. To further characterize the function of ORF20, we generated an ORF20-deficient KSHV and compared its replicative fitness relative to wild type virus. Virion production was significantly diminished in the ORF20-deficient virus as observed by supernatant transfer assays. Additionally, we tied this defect in viable virion formation to a reduction in viral late gene expression. Lastly, we observed an overall reduction in vDNA replication in the ORF20-deficient virus implying a key role for ORF20 in the regulation of lytic replication. Taken together, these results capture the essential role of KSHV ORF20 in progressing viral lytic infection by regulating vDNA replication alongside other crucial lytic proteins within KSHV replication compartments.

Sign in / Sign up

Export Citation Format

Share Document