scholarly journals Bone Marrow Derived SH-SY5Y Neuroblastoma Cells Infected by Kaposi’s Sarcoma Herpes Virus (KSHV) Display Unique Infection Phenotypes and Growth Properties

2021 ◽  
Author(s):  
Xiaohong Kong ◽  
Dongmei Li ◽  
Amirsalar Mansouri ◽  
Guobin Kang ◽  
Khalid Sayood ◽  
...  
Keyword(s):  
Growth Rate ◽  
Cell Adhesion ◽  
Cell Growth ◽  
Neuronal Cells ◽  
Kshv Infection ◽  
Cellular Genes ◽  
Viral Genes ◽  
Latently Infected ◽  
Infected Cells ◽  
Lytic Genes

The Kaposi’s sarcoma-associated herpesvirus (KSHV) is an important oncogenic virus previously shown to be neurotropic, but studies on neuronal cells infection and pathogenesis are still very limited. Here we characterized the effects of KSHV infection on neuronal SH-SY5Y cells by the recombinant virus rKSHV219, which expresses both GFP and RFP to reflect latent and lytic phases of infection. We demonstrated that infected cells have a faster growth rate and the KSHV infection can be sustained. Interestingly, the infected cells can transition spontaneously back and forth between lytic and latent phases of infections, producing progeny viruses but without any adverse effects on cell growth. In addition, transcriptome analysis of viral and cellular genes in latent and lytic cells showed that unlike other infected cell lines, the latently infected cells expressed both latent and most, but not all the lytic genes required for infectious virion production. The uniquely expressed viral genes by the lytic cells were mainly involved in the early steps of virus binding. Some of the cellular genes that were deregulated in both latent and lytically infected cells are involved in cell adhesion, in cell signal pathways and tumorigenesis. The downregulated cellular CCDN1, PAX5, NFASC and upregulated CTGF, BMP4, YAP1, LEF1 and HLA-DRB1 genes were found to be associated with the cell adhesion molecules (CAM), hippo signaling and cancer. These deregulated genes may be involved in creating an environment that are unique in the neuronal cells to sustain cell growth upon KSHV infection not observed in other infected cell types. IMPORTANCE Our study has provided evidence that the neuronal SH-SY5Y cells displayed unique cellular responses upon KSHV infection. Unlike other infected cells, this neuronal cell displayed a more rapid growth rate upon infection and can spontaneously transition back and forth between latent and lytic phases of infection. Unlike other latently infected cells, a number of lytic genes were also expressed in the latent phase of infection in addition to the established latent viral genes. They may play a role in deregulating a number of host genes that are involved in cell signaling and tumorigenesis in order to sustain the infection and growth advantages for the cells. Our study has provided novel insights on KSHV infection of neuronal cells and a potential new model for further studies to explore the underlying mechanism in viral and host interaction for neuronal cells, and the association of KSHV with neuronal diseases.

scholarly journals Nascent Transcriptomics Reveal Cellular Prolytic Factors Upregulated Upstream of the Latent-to-Lytic Switch Protein of Epstein-Barr Virus

2020 ◽  
Vol 94 (7) ◽  
Author(s):  
Tiffany R. Frey ◽  
Jozan Brathwaite ◽  
Xiaofan Li ◽  
Sandeepta Burgula ◽  
Ibukun A. Akinyemi ◽  
...  
Keyword(s):  
Epstein Barr Virus ◽  
Human Herpesvirus ◽  
Lytic Cycle ◽  
Barr Virus ◽  
Viral Spread ◽  
Cellular Genes ◽  
Latently Infected ◽  
Infected Cells ◽  
Host Shutoff ◽  
Epstein Barr

ABSTRACT Lytic activation from latency is a key transition point in the life cycle of herpesviruses. Epstein-Barr virus (EBV) is a human herpesvirus that can cause lymphomas, epithelial cancers, and other diseases, most of which require the lytic cycle. While the lytic cycle of EBV can be triggered by chemicals and immunologic ligands, the lytic cascade is activated only when expression of the EBV latent-to-lytic switch protein ZEBRA is turned on. ZEBRA then transcriptionally activates other EBV genes and, together with some of those gene products, ensures completion of the lytic cycle. However, not every latently infected cell exposed to a lytic trigger turns on the expression of ZEBRA, resulting in responsive and refractory subpopulations. What governs this dichotomy? By examining the nascent transcriptome following exposure to a lytic trigger, we find that several cellular genes are transcriptionally upregulated temporally upstream of ZEBRA. These genes regulate lytic susceptibility to various degrees in latently infected cells that respond to mechanistically distinct lytic triggers. While increased expression of these cellular genes defines a prolytic state, such upregulation also runs counter to the well-known mechanism of viral-nuclease-mediated host shutoff that is activated downstream of ZEBRA. Furthermore, a subset of upregulated cellular genes is transcriptionally repressed temporally downstream of ZEBRA, indicating an additional mode of virus-mediated host shutoff through transcriptional repression. Thus, increased transcription of a set of host genes contributes to a prolytic state that allows a subpopulation of cells to support the EBV lytic cycle. IMPORTANCE Transition from latency to the lytic phase is necessary for herpesvirus-mediated pathology as well as viral spread and persistence in the population at large. Yet, viral genomes in only some cells in a population of latently infected cells respond to lytic triggers, resulting in subpopulations of responsive/lytic and refractory cells. Our investigations into this partially permissive phenotype of the herpesvirus Epstein-Barr virus (EBV) indicate that upon exposure to lytic triggers, certain cellular genes are transcriptionally upregulated, while viral latency genes are downregulated ahead of expression of the viral latent-to-lytic switch protein. These cellular genes contribute to lytic susceptibility to various degrees. Apart from indicating that there may be a cellular “prolytic” state, our findings indicate that (i) early transcriptional upregulation of cellular genes counters the well-known viral-nuclease-mediated host shutoff and (ii) subsequent transcriptional downregulation of a subset of early upregulated cellular genes is a previously undescribed mode of host shutoff.

scholarly journals Host Cell Gene Expression during Human Immunodeficiency Virus Type 1 Latency and Reactivation and Effects of Targeting Genes That Are Differentially Expressed in Viral Latency

2004 ◽  
Vol 78 (17) ◽  
pp. 9458-9473 ◽  
Author(s):  
Vyjayanthi Krishnan ◽  
Steven L. Zeichner
Keyword(s):  
Gene Expression ◽  
Host Cell ◽  
Cell Lines ◽  
Lytic Replication ◽  
Replication Cycle ◽  
Differentially Expressed ◽  
Cellular Genes ◽  
Latently Infected ◽  
Cell Gene Expression ◽  
Infected Cells

ABSTRACT The existence of reservoirs of cells latently infected with human immunodeficiency virus (HIV) is a major obstacle to the elimination of HIV infection. We studied the changes in cellular gene expression that accompany the reactivation and completion of the lytic viral cycle in cell lines chronically infected with HIV-1. We found that several genes exhibited altered expression in the chronically infected cells compared to the uninfected parental cells prior to induction into lytic replication. A number of gene classes showed increased expression in the chronically infected cells, notably including genes encoding proteasomes, histone deacetylases, and many transcription factors. Following induction of the lytic replication cycle, we observed ordered, time-dependent changes in the cellular gene expression pattern. Approximately 1,740 genes, many of which fall into 385 known pathways, were differentially expressed (P < 0.001), indicating that completion of the HIV replication cycle is associated with distinct, temporally ordered changes in host cell gene expression. Maximum changes were observed in the early and intermediate phases of the lytic replication cycle. Since the changes in gene expression in chronically infected cells suggested that cells latently infected with HIV have a different gene expression profile than corresponding uninfected cells, we studied the expression profiles of three different chronically infected cell lines to determine whether they showed similar changes in common cellular genes and pathways. Thirty-two genes showed significant differential expression in all cell lines studied compared to their uninfected parental cell lines. Notable among them were cdc42 and lyn, which were downregulated and are required for HIV Nef binding and viral replication. Other genes previously unrelated to HIV latency or pathogenesis were also differentially expressed. To determine the effects of targeting products of the genes that were differentially expressed in latently infected cells, we treated the latently infected cells with a proteasome inhibitor, clastolactacystin-beta-lactone (CLBL), and an Egr1 activator, resveratrol. We found that treatment with CLBL and resveratrol stimulated lytic viral replication, suggesting that treatment of cells with agents that target cellular genes differentially expressed in latently infected cells can stimulate lytic replication. These findings may offer new insights into the interaction of the latently infected host cell and HIV and suggest therapeutic approaches for inhibiting HIV infection and for manipulating cells latently infected with HIV so as to trigger lytic replication.

scholarly journals Human splice factors contribute to latent HIV infection in primary cell models and blood CD4+ T cells from ART-treated individuals

2020 ◽  
Vol 16 (11) ◽  
pp. e1009060
Author(s):  
Sara Moron-Lopez ◽  
Sushama Telwatte ◽  
Indra Sarabia ◽  
Emilie Battivelli ◽  
Mauricio Montano ◽  
...  
Keyword(s):  
T Cells ◽  
Hiv Infection ◽  
Primary Cell ◽  
Cell Models ◽  
Transcriptional Initiation ◽  
Cellular Genes ◽  
Latently Infected ◽  
Infected Cells ◽  
Latent Hiv

It is unclear what mechanisms govern latent HIV infection in vivo or in primary cell models. To investigate these questions, we compared the HIV and cellular transcription profile in three primary cell models and peripheral CD4+ T cells from HIV-infected ART-suppressed individuals using RT-ddPCR and RNA-seq. All primary cell models recapitulated the block to HIV multiple splicing seen in cells from ART-suppressed individuals, suggesting that this may be a key feature of HIV latency in primary CD4+ T cells. Blocks to HIV transcriptional initiation and elongation were observed more variably among models. A common set of 234 cellular genes, including members of the minor spliceosome pathway, was differentially expressed between unstimulated and activated cells from primary cell models and ART-suppressed individuals, suggesting these genes may play a role in the blocks to HIV transcription and splicing underlying latent infection. These genes may represent new targets for therapies designed to reactivate or silence latently-infected cells.

scholarly journals HACE1, an E3 Ubiquitin Protein Ligase, Mitigates Kaposi’s Sarcoma-Associated Herpesvirus Infection-Induced Oxidative Stress by Promoting Nrf2 Activity

2019 ◽  
Vol 93 (9) ◽  
Author(s):  
Binod Kumar ◽  
Arunava Roy ◽  
Kumari Asha ◽  
Neelam Sharma-Walia ◽  
Mairaj Ahmed Ansari ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Death ◽  
Kaposi's Sarcoma ◽  
Nuclear Translocation ◽  
Kaposi’S Sarcoma ◽  
Induce Cell Death ◽  
Kshv Infection ◽  
Ubiquitin Protein Ligase ◽  
Viral Genes ◽  
Infected Cells

ABSTRACTKaposi’s sarcoma-associated herpesvirus (KSHV)-induced activation of nuclear factor erythroid 2-related factor 2 (Nrf2) is essential for both the expression of viral genes (latency) and modulation of the host antioxidant machinery. Reactive oxygen species (ROS) are also regulated by the ubiquitously expressed HACE1 protein (HECT domain and ankyrin repeat containing E3 ubiquitin protein ligase 1), which targets the Rac1 protein for proteasomal degradation, and this blocks the generation of ROS by Rac1-dependent NADPH oxidases. In this study, we examined the role of HACE1 in KSHV infection. Elevated levels of HACE1 expression were observed inde novoKSHV-infected endothelial cells, KSHV latently infected TIVE-LTC and PEL cells, and Kaposi’s sarcoma skin lesion cells. The increased HACE1 expression in the infected cells was mediated by KSHV latent protein kaposin A. HACE1 knockdown resulted in high Rac1 and Nox 1 (NADPH oxidase 1) activity, increased ROS (oxidative stress), increased cell death, and decreased KSHV gene expression. Loss of HACE1 impaired KSHV infection-induced phosphoinositide 3-kinase (PI3-K), protein kinase C-ζ (PKC-ζ), extracellular signal-regulated kinase 1/2 (ERK1/2), NF-κB, and Nrf2 activation and nuclear translocation of Nrf2, and it reduced the expression of Nrf2 target genes responsible for balancing the oxidative stress. In the absence of HACE1, glutamine uptake increased in the cells to cope with the KSHV-induced oxidative stress. These findings reveal for the first time that HACE1 plays roles during viral infection-induced oxidative stress and demonstrate that HACE1 facilitates resistance to KSHV infection-induced oxidative stress by promoting Nrf2 activity. Our studies suggest that HACE1 could be a potential target to induce cell death in KSHV-infected cells and to manage KSHV infections.IMPORTANCEROS play important roles in several cellular processes, and increased ROS cause several adverse effects. KSHV infection of endothelial cells induces ROS, which facilitate virus entry by amplifying the infection-induced host cell signaling cascade, which, in turn, induces the nuclear translocation of phospho-Nrf2 protein to regulate the expression of antioxidative genes and viral genes. The present study demonstrates that KSHV infection induces the E3 ligase HACE1 protein to regulate KSHV-induced oxidative stress by promoting the activation of Nrf2 and nuclear translocation. Absence of HACE1 results in increased ROS and cellular death and reduced nuclear Nrf2, antioxidant, and viral gene expression. Together, these studies suggest that HACE1 can be a potential target to induce cell death in KSHV-infected cells.

scholarly journals Induction of Autophagy to Achieve a Human Immunodeficiency Virus Type 1 Cure

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1798
Author(s):  
Grant R. Campbell ◽  
Stephen A. Spector
Keyword(s):  
Human Immunodeficiency Virus ◽  
Antiretroviral Therapy ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Functional Cure ◽  
Immunodeficiency Virus ◽  
Latently Infected ◽  
Infected Cells ◽  
Hiv 1

Effective antiretroviral therapy has led to significant human immunodeficiency virus type 1 (HIV-1) suppression and improvement in immune function. However, the persistence of integrated proviral DNA in latently infected reservoir cells, which drive viral rebound post-interruption of antiretroviral therapy, remains the major roadblock to a cure. Therefore, the targeted elimination or permanent silencing of this latently infected reservoir is a major focus of HIV-1 research. The most studied approach in the development of a cure is the activation of HIV-1 expression to expose latently infected cells for immune clearance while inducing HIV-1 cytotoxicity—the “kick and kill” approach. However, the complex and highly heterogeneous nature of the latent reservoir, combined with the failure of clinical trials to reduce the reservoir size casts doubt on the feasibility of this approach. This concern that total elimination of HIV-1 from the body may not be possible has led to increased emphasis on a “functional cure” where the virus remains but is unable to reactivate which presents the challenge of permanently silencing transcription of HIV-1 for prolonged drug-free remission—a “block and lock” approach. In this review, we discuss the interaction of HIV-1 and autophagy, and the exploitation of autophagy to kill selectively HIV-1 latently infected cells as part of a cure strategy. The cure strategy proposed has the advantage of significantly decreasing the size of the HIV-1 reservoir that can contribute to a functional cure and when optimised has the potential to eradicate completely HIV-1.

scholarly journals Targeting the latent human cytomegalovirus reservoir for T-cell-mediated killing with virus-specific nanobodies

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Timo W. M. De Groof ◽  
Elizabeth G. Elder ◽  
Eleanor Y. Lim ◽  
Raimond Heukers ◽  
Nick D. Bergkamp ◽  
...  
Keyword(s):  
Gene Expression ◽  
Human Cytomegalovirus ◽  
Early Gene ◽  
Latent Reservoir ◽  
Solid Organ ◽  
Cell Transplant ◽  
Viral Receptor ◽  
Transplant Patients ◽  
Latently Infected ◽  
Infected Cells

AbstractLatent human cytomegalovirus (HCMV) infection is characterized by limited gene expression, making latent HCMV infections refractory to current treatments targeting viral replication. However, reactivation of latent HCMV in immunosuppressed solid organ and stem cell transplant patients often results in morbidity. Here, we report the killing of latently infected cells via a virus-specific nanobody (VUN100bv) that partially inhibits signaling of the viral receptor US28. VUN100bv reactivates immediate early gene expression in latently infected cells without inducing virus production. This allows recognition and killing of latently infected monocytes by autologous cytotoxic T lymphocytes from HCMV-seropositive individuals, which could serve as a therapy to reduce the HCMV latent reservoir of transplant patients.

82 Methyl Donor Supplementation Alters Cytosine Methylation and Biological Processes of Cells Cultured in Divergent Glucose Media Reflecting Improvements in Mitochondrial Respiration and Cell Growth Rate

2021 ◽  
Vol 99 (Supplement_1) ◽  
pp. 109-109
Author(s):  
Matthew S Crouse ◽  
Wellison Jarles Da Silva Diniz ◽  
Joel Caton ◽  
Carl R Dahlen ◽  
Lawrence P Reynolds ◽  
...  
Keyword(s):  
Growth Rate ◽  
Cell Proliferation ◽  
Cell Growth ◽  
Vitamin B12 ◽  
Mitochondrial Respiration ◽  
Cytosine Methylation ◽  
Biological Processes ◽  
Cell Growth Rate ◽  
Metabolic Processes ◽  
Positive Regulation

Abstract We hypothesized that supplementation of one-carbon metabolites (OCM: methionine, folate, choline, and vitamin B12) to bovine embryonic tracheal fibroblasts in divergent glucose media would alter cytosine methylation, and alterations in cytosine methylation will reflect biological processes matching previously improved mitochondrial respiration, cell proliferation, and cell growth rate data. Cells were cultured with 1g/L glucose (Low) or 4.5g/L glucose (High). Control medium (CON) contained basal concentrations of folate (0.001g/L), choline (0.001g/L), vitamin B12 (4µg/L), and methionine (0.015g/L). The OCM were supplemented at 2.5 and 5 times (2.5X and 5X, respectively) the CON media, except methionine was limited to 2X across all supplemented treatments. Cells were passaged three times in their treatment media before DNA extraction. Reduced representation bisulfite sequencing was adopted to analyze and compare the genomic methylation patterns within and across treatments using edgeR. Biological processes (BP) were retrieved based on the nearest genes of differentially methylated cytosines (P &lt; 0.01) for each comparison between treatments. In both Low and High treatments, greater OCM increased the proportion of hypomethylated vs. hypermethylated cytosines. Functional analyses pointed out positive regulation of BP related to energy metabolism, except for the contrasts within the High group. Among the BP, we can highlight positive regulation of: GTPase activity, catalytic activity, molecular function, protein modification processes, phosphorylation, protein phosphorylation, cellular protein metabolic processes, MAPK cascade, and metabolic processes. These data support previously reported results from this experiment that showed increased mitochondrial respiration, cell proliferation, and growth rates with increasing OCM levels. We interpret these data to imply that when energy and OCM requirements are met for growth and basal methylation levels, DNA methylation levels decrease which may allow for greater transcription. Thus, OCM can be utilized for other functions such as polyamine synthesis, nucleotide synthesis, energetic metabolites, and phosphatidylcholine synthesis. USDA is an equal opportunity provider and employer.

Galectin-3 is involved in HIV-1 expression through NF-κB activation and associated with Tat in latently infected cells

2019 ◽  
Vol 260 ◽  
pp. 86-93 ◽  
Author(s):  
Mika Okamoto ◽  
Akemi Hidaka ◽  
Masaaki Toyama ◽  
Masanori Baba
Keyword(s):  
Latently Infected ◽  
Galectin 3 ◽  
Infected Cells ◽  
Hiv 1

scholarly journals The Early Gene hhi1 Reactivates Heliothis zea Nudivirus 1 in Latently Infected Cells

2009 ◽  
Vol 84 (2) ◽  
pp. 1057-1065 ◽  
Author(s):  
Yueh-Lung Wu ◽  
Carol P. Wu ◽  
Song-Tay Lee ◽  
Han Tang ◽  
Chi-Hua Chang ◽  
...  
Keyword(s):  
Viral Infection ◽  
Critical Role ◽  
Heliothis Zea ◽  
Virus Titer ◽  
Viral Reactivation ◽  
Early Gene ◽  
Mechanistic Study ◽  
Early Genes ◽  
Latently Infected ◽  
Infected Cells

ABSTRACT Heliothis zea nudivirus 1 (HzNV-1), previously known as Hz-1 virus, is an insect virus able to establish both productive and latent infections in several lepidopteran insect cells. Here, we have cloned and characterized one of the HzNV-1 early genes, hhi1, which maps to the HindIII-I fragment of the viral genome. During the productive viral infection, a 6.2-kb hhi1 transcript was detectable as early as 0.5 h postinfection (hpi). The level of transcript reached a maximum at 2 hpi and gradually decreased after 4 hpi. The transcript was not detectable during the latent phase of viral infection. Upon cycloheximide treatment, much higher levels of hhi1 transcript were detected throughout the productive viral infection cycle, suggesting that newly synthesized proteins are not needed for the expression of hhi1. Nevertheless, viral coinfection can further stimulate the expression of transfected hhi1 promoter in a plasmid. Transient hhi1 expression in latently infected cells resulted in a significant increase in virus titer and viral DNA propagation, suggesting that hhi1 plays a critical role in viral reactivation. Additional experiments showed that six early genes, which possibly function in transcription or DNA replication, were activated in the latent cells upon hhi1 transfection. Among these six genes, orf90 and orf121 expression could be induced by hhi1 alone without the need for other viral genes. Our discovery should be useful for future mechanistic study of the switches of latent/productive HzNV-1 viral infections.

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