Prolonged activation of NF-kappaB by human cytomegalovirus promotes efficient viral replication and late gene expression.
(2006)
Journal - Virology (United States )
Abstract :
Infection of fibroblasts by human cytomegalovirus (HCMV) rapidly activates the NF-kappaB signaling pathway, which we documented promotes efficient transactivation of the major immediate-early promoter (DeMeritt, I.B., Milford, L.E., Yurochko, A.D. (2004). Activation of the NF-kappaB pathway in human cytomegalovirus-infected cells is necessary for efficient transactivation of the major immediate-early promoter. J. Virol. 78, 4498-4507). Because a second, sustained increase in NF-kappaB activity following the initial phase of NF-kappaB activation was also observed, we investigated the role that this prolonged NF-kappaB activation played in viral replication and late gene expression. We first investigated HCMV replication in cells in which NF-kappaB activation was blocked by pretreatment with NF-kappaB inhibitors: HCMV replication was significantly decreased in these cultures. A decrease in replication was also observed when NF-kappaB was inhibited up to 48 h post-infection, suggesting a previously unidentified role for NF-kappaB in the regulation of the later class of viral genes.
| ISSN : | 0042-6822 |
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| Mesh Heading : | Aspirin Cells, Cultured Cytomegalovirus Fibroblasts Humans Leupeptins NF-kappa B Time Factors Viral Proteins pharmacology metabolism pathogenicity pharmacology antagonists & inhibitors genetics |
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| Mesh Heading Relevant : | Gene Expression Regulation, Viral Virus Replication physiology metabolism metabolism |
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Activation of the NF-kappaB pathway in human cytomegalovirus-infected cells is necessary for efficient transactivation of the major immediate-early promoter.
(2004)
Journal - Journal of virology (United States )
Abstract :
We previously demonstrated that human cytomegalovirus (HCMV) infection induced the activation of the cellular transcription factor NF-kappaB. Here, we investigate the mechanism for the HCMV-induced NF-kappaB activation and the role that the induced NF-kappaB plays in transactivation of the major immediate-early promoter (MIEP) and production of immediate-early (IE) proteins. Using a dominant-negative inhibitor of NF-kappaB, the IkappaB-superrepressor, we demonstrated that active NF-kappaB is critical for transactivation of the HCMV MIEP. Investigation of the mechanisms of NF-kappaB activation following HCMV infection showed a rapid and sustained decrease in the inhibitors of NF-kappaB, IkappaBalpha and IkappaBbeta. Because the IkappaB kinases (IKKs) regulate the degradation of the IkappaBs, virus-mediated changes in the IKKs were examined next. Using dominant-negative forms of the IKKs, we showed significant decreases in transactivation of the MIEP in the presence of these mutants. In addition, protein levels of members of the IKK complex and IKK kinase activity were upregulated throughout the time course of infection. Lastly, the role NF-kappaB plays in HCMV IE mRNA and protein production during infection was examined. Using aspirin and MG-132, we demonstrated that production of IE protein and mRNA was significantly decreased and delayed in infected cells treated with these drugs. Together, the results of these studies suggest that virus-mediated NF-kappaB activation, through the dysregulation of the IKK complex, plays a primary role in the initiation of the HCMV gene cascade in fibroblasts and may provide new targets for therapeutic intervention.
| ISSN : | 0022-538X |
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| Mesh Heading : | Cells, Cultured Cytomegalovirus Cytomegalovirus Infections Down-Regulation Fibroblasts Gene Expression Regulation, Viral Humans I-kappa B Kinase I-kappa B Proteins Immediate-Early Proteins NF-kappa B Promoter Regions, Genetic Protein-Serine-Threonine Kinases genetics virology metabolism genetics metabolism |
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| Mesh Heading Relevant : | Transcriptional Activation pathogenicity virology metabolism metabolism physiology |
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The Human Cytomegalovirus Virion Possesses an Activated Casein Kinase II That Allows for the Rapid Phosphorylation of the Inhibitor of NF-?B, I?Ba
(2007)
Journal - Journal of Virology
Abstract :
We documented that the NF-?B signaling pathway was rapidly induced following human cytomegalovirus (HCMV) infection of human fibroblasts and that this induced NF-?B activity promoted efficient transactivation of the major immediate-early promoter (MIEP). Previously, we showed that the major HCMV envelope glycoproteins, gB and gH, initiated this NF-?B signaling event. However, we also hypothesized that there were additional mechanisms utilized by the virus to rapidly upregulate NF-?B. In this light, we specifically hypothesized that the HCMV virion contained I?Ba kinase activity, allowing for direct phosphorylation of I?Ba following virion entry into infected cells. In vitro kinase assays performed on purified HCMV virion extract identified bona fide I?Ba kinase activity in the virion. The enzyme responsible for this kinase activity was identified as casein kinase II (CKII), a cellular serine-threonine protein kinase. CKII activity was necessary for efficient transactivation of the MIEP and IE gene expression. CKII is generally considered to be a constitutively active kinase. We suggest that this molecular characteristic of CKII represents the biologic rationale for the viral capture and utilization of this kinase early after infection. The packaging of CKII into the HCMV virion identifies that diverse molecular mechanisms are utilized by HCMV for rapid NF-?B activation. We propose that HCMV possesses multiple pathways to increase NF-?B activity to ensure that the correct temporal regulation of NF-?B occurs following infection and that sufficient threshold levels of NF-?B are reached in the diverse array of cells, including monocytes and endothelial cells, infected in vivo.
Activation of the NF-?B Pathway in Human Cytomegalovirus-Infected Cells Is Necessary for Efficient Transactivation of the Major Immediate-Early Promoter
(2004)
Journal - Journal of Virology
Abstract :
We previously demonstrated that human cytomegalovirus (HCMV) infection induced the activation of the cellular transcription factor NF-?B. Here, we investigate the mechanism for the HCMV-induced NF-?B activation and the role that the induced NF-?B plays in transactivation of the major immediate-early promoter (MIEP) and production of immediate-early (IE) proteins. Using a dominant-negative inhibitor of NF-?B, the I?B-superrepressor, we demonstrated that active NF-?B is critical for transactivation of the HCMV MIEP. Investigation of the mechanisms of NF-?B activation following HCMV infection showed a rapid and sustained decrease in the inhibitors of NF-?B, I?Ba and I?Bß. Because the I?B kinases (IKKs) regulate the degradation of the I?Bs, virus-mediated changes in the IKKs were examined next. Using dominant-negative forms of the IKKs, we showed significant decreases in transactivation of the MIEP in the presence of these mutants. In addition, protein levels of members of the IKK complex and IKK kinase activity were upregulated throughout the time course of infection. Lastly, the role NF-?B plays in HCMV IE mRNA and protein production during infection was examined. Using aspirin and MG-132, we demonstrated that production of IE protein and mRNA was significantly decreased and delayed in infected cells treated with these drugs. Together, the results of these studies suggest that virus-mediated NF-?B activation, through the dysregulation of the IKK complex, plays a primary role in the initiation of the HCMV gene cascade in fibroblasts and may provide new targets for therapeutic intervention.