Genome wide DNA methylation of Epstein-Barr virus infected immortalized normal oral keratinocytes
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE59842 | GEO | 2014/08/01
SECONDARY ACCESSION(S): PRJNA256343
REPOSITORIES: GEO
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Project description:The oral cavity is the persistent reservoir for EBV with lifelong infection of resident epithelial and B cells. Infection of these cell types results in distinct EBV gene expression patterns that are regulated by epigenetic modifications involving DNA methylation and chromatin structure. Such regulation of EBV gene expression relies on viral manipulation of the host epigenetic machinery that may inadvertently result in long-lasting, oncogenic host epigenetic reprogramming. To test this hypothesis in the context of EBV infection of epithelial cells, we established a transient infection model to identify the epigenetic consequences after EBV infection of immortalized normal oral keratinocytes and subsequent viral loss. NOK cells were single cell cloned (designated as cl1) and EBV infected by co-culture with a recombinant Akata EBV strain carrying neomcycin resistance and GFP expression cassette in place of EBV BXLF1. Cells were selected with 350 ug/ml G418. As a selection control, cells were transfected a plasmid carrying a neomycin resistance cassette. Cells were passaged 10 times with G418, followed by removal of selection pressure. After an additional 10 passages, cells were single cell cloned and the presence of EBV was determined by various methods. Three clones were identified as being EBV negative and said to be transiently infected. Uninfected parental and vector transfected cells were passaged in the same manner and also single cell cloned. The DNA methylation profiles were examined using reduced representation bisulfite sequencing.
2014-08-01 | E-GEOD-59842 | biostudies-arrayexpress
Project description:The oral cavity is the persistent reservoir for EBV with lifelong infection of resident epithelial and B cells. Infection of these cell types results in distinct EBV gene expression patterns that are regulated by epigenetic modifications involving DNA methylation and chromatin structure. Such regulation of EBV gene expression relies on viral manipulation of the host epigenetic machinery that may inadvertently result in long-lasting, oncogenic host epigenetic reprogramming. To test this hypothesis in the context of EBV infection of epithelial cells, we established a transient infection model to identify the epigenetic consequences after EBV infection of immortalized normal oral keratinocytes and subsequent viral loss. With mounting evidence that EBV can induce epigenetic alterations, we developed a transient infection model where a clonal derivative (designated cl1) from a human telomerase immortalized normal oral keratinocyte (NOK) cell line was infected with a recombinant Akata EBV carrying neomycin resistance and GfP cassette in place of the BXLF1 open reading frame. Infected cells were passaged ten times, and then selection pressure was removed for an additional ten passages to allow for viral loss. Three transiently-infected EBV-negative clones were identified by single cell cloning (designated cl1, cl3, cl4). Uninfected parental clone and cells transfected with PTRUF5 plasmid were passaged alongside the transiently-infected clones. The transcription profiles were analyzed using Affymetrix microarray U133Plus2.0 arrays in duplicate.
2014-08-01 | GSE58914 | GEO
Project description:The oral cavity is the persistent reservoir for EBV with lifelong infection of resident epithelial and B cells. Infection of these cell types results in distinct EBV gene expression patterns that are regulated by epigenetic modifications involving DNA methylation and chromatin structure. Such regulation of EBV gene expression relies on viral manipulation of the host epigenetic machinery that may inadvertently result in long-lasting, oncogenic host epigenetic reprogramming. To test this hypothesis in the context of EBV infection of epithelial cells, we established a transient infection model to identify the epigenetic consequences after EBV infection of immortalized normal oral keratinocytes and subsequent viral loss. With mounting evidence that EBV can induce epigenetic alterations, we developed a transient infection model where a clonal derivative (designated cl1) from a human telomerase immortalized normal oral keratinocyte (NOK) cell line was infected with a recombinant Akata EBV carrying neomycin resistance and GfP cassette in place of the BXLF1 open reading frame. Infected cells were passaged ten times, and then selection pressure was removed for an additional ten passages to allow for viral loss. Three transiently-infected EBV-negative clones were identified by single cell cloning (designated cl1, cl3, cl4). Uninfected parental clone and cells transfected with PTRUF5 plasmid were passaged alongside the transiently-infected clones. The transcription profiles were analyzed using Affymetrix microarray U133Plus2.0 arrays in duplicate. NOK cells were single cell cloned (designated as cl1) and EBV infected by co-culture with a recombinant Akata EBV strain carrying neomcycin resistance and GFP expression cassette in place of EBV BXLF1. Cells were selected with 350 ug/ml G418. As a selection control, cells were transfected a plasmid carrying a neomycin resistance cassette. Cells were passaged 10 times with G418, followed by removal of selection pressure. After an additional 10 passages, cells were single cell cloned and the presence of EBV was determined by various methods. Three clones were identified as being EBV negative and said to be transiently infected. Uninfected parental and vector transfected cells were passaged in the same manner and also single cell cloned.
2014-08-01 | E-GEOD-58914 | biostudies-arrayexpress
Project description:Epstein Barr virus (EBV) replication contributes to multiple human diseases, including infectious mononucleosis, nasopharyngeal carcinoma, B-cell lymphomas, and oral hairy leukoplakia. We performed systematic quantitative analyses of temporal changes in host and EBV proteins during lytic replication to gain novel insights into virus-host interactions, using conditional Burkitt lymphoma models of type I and II EBV infection. We quantified profiles of >8000 cellular and 69 EBV proteins, including >500 plasma membrane proteins, providing temporal views of the lytic B-cell proteome and EBV virome. Our approach revealed EBV-induced remodelling of cell cycle, innate and adaptive immune pathways, including upregulation of the complement cascade and proteasomal degradation of the B-cell receptor complex, conserved between EBV types I and II. Cross-comparison with proteomic analyses of human cytomegalovirus infection and of a Kaposi sarcoma associated herpesvirus immunoevasin identified host factors targeted by multiple herpesviruses. Our results provide an important resource for studies of EBV replication.
2017-05-19 | PXD006317 | Pride
Project description:Comprehensive Profiling of Epstein-Barr Virus-Encoded miRNAome Associated with Specific Latent Type in Tumor Cells Epstein-Barr virus (EBV) is an etiological cause of many human lymphocytic and epithelial malignancies. EBV expressed different genes associated with three latent types. So far as many as 44 EBV-encoded miRNA species have been found but their comprehensive and comparative profiling is not well documented in three latent infection states linked to various tumor cells. In this study, we utilized the polyA-tailed quantitative real time RT-PCR procedure to measure the relative abundance of viral miRNA species that linked to individual viral genome locations in combination with microarray evaluation in a subset of representative lymphoid and epithelial tumor cells undergoing various types of EBV latent infection. The results showed that miR-BHRF1 family and miR-BART family are expressed differentially in a tissue-dependent and latency-dependent manner. In particular, in NPC tissue and the only EBV consistently harboring cell line C666-1 with latency type II, there were highly abundant miR-BART family but not miR-BHRF1 family members that accounted for more than 10% of the whole known human miRNA library, implicating their important roles in maintaining EBV latent infection and driving NPC tumorigenesis. In addition, EBV miRNAome-based clustering analysis could classify three distinct EBV latency types, meanwhile, for the first time, we found and subsequently evaluated a novel secret latent switch in BL cell line Daudi from type I to III, which was unable to be identified by traditional latent biomarkers. Together, our data provided an in-depth and comparative profiling of EBV miRNA transcriptome in correspondence with three EBV latent infections, suggesting that different viral miRNA species were involved in divergent host cell carcinogenesis. Finally, EBV miRNAome, as a cluster of novel latency biomarkers expressed variedly in tumor cells, greatly complements and improves the classical typing criteria in conjunction with other latently expressed marker genes. 2 NPC tissue samples and 2 NPC cell lines and 5 lymphocytic cell lines
2012-07-06 | E-GEOD-33225 | biostudies-arrayexpress
Project description:Epstein-Barr virus (EBV) causes Burkitt, Hodgkin, and post-transplant B-cell lymphomas. How EBV remodels metabolic pathways to support rapid B-cell outgrowth remains largely unknown. To gain insights, primary human B-cells were profiled by tandem-mass-tag based proteomics at rest and at 9 time points after infection. >8000 host and 29 viral proteins were quantified, revealing mitochondrial remodeling and induction of one-carbon (1C) metabolism. EBV-encoded EBNA2 and its target MYC were required for upregulation of the central mitochondrial 1C enzyme MTHFD2, which played key roles in EBV-driven B-cell growth and survival. MTHFD2 was critical for maintaining elevated NADPH levels in infected cells, and oxidation of mitochondrial NADPH diminished B-cell proliferation. Tracing studies underscored contributions of 1C to nucleotide synthesis, NADPH production and redox defense. EBV upregulated import and synthesis of serine to augment 1C flux. Our results highlight EBV-induced 1C as a potential therapeutic target and provide a new paradigm for viral onco-metabolism.
2019-06-28 | PXD013034 | Pride
Project description:The prototypic gammaherpesvirus Epstein-Barr virus (EBV) is carried by over 90% of the adult human population. Although infection often proceeds asymptomatically, the virus is causally involved in infectious mononucleosis and various malignancies. Like all herpesviruses, EBV establishes a lifelong latent infection in its human host. To successfully establish a persistent infection and spread to new hosts, EBV needs to avoid clearance by the host immune system. In addition to ~100 proteins, some of which perform immunomodulatory functions, EBV encodes over 40 mature miRNAs. Cellular miRNAs regulate diverse cellular processes, but the functions of the EBV miRNAs remain poorly understood. In this study, we identify EBV-encoded miR-BART16 as a novel viral immune evasion factor that interferes with host type I IFN responses. miR-BART16-5p downregulates the cellular histone acetyltransferase CBP (CREBB-binding protein) by directly targeting the CBP-encoding mRNA. CBP plays an important role in the type I IFN signaling pathway. Inhibiting endogenously expressed miR-BART16 in EBV-transformed B cells and gastric carcinoma cells results in increased CBP levels. Consistent with the important role of CBP in type I IFN signaling, miR-BART16 expression interferes with ISRE-promoter activity and production of interferon-stimulated genes in response to IFN-α treatment. These combined data show that EBV employs miRNAs to modify IFN-induced gene transcription. Given the importance of the host IFN system in the antiviral defence, obstructing this response may provide a way for EBV to favour viral replication and could play an etiological role in EBV-associated malignancies.
2017-04-05 | GSE69697 | GEO
Project description:Epstein-Barr virus (EBV) causes endemic Burkitt lymphoma and immunosuppression-related lymphomas. These B-cell malignancies arise by distinct transformation pathways and utilize divergent viral and host expression programs. To identify host dependency factors elicited by EBV latent-infection states, we performed parallel genome-wide CRISPR/Cas9 screens in Burkitt lymphoma (BL) and lymphoblasotid cell lines (LCL). Our results highlighted 57 BL and 87 LCL genes selectively critical for their growth and survival. LCL hits were enriched for EBV-induced genes, including viral super-enhancer targets and multiple kinases. We uncovered key CD19/CD81 roles in EBV membrane protein-driven PI3K/AKT activation and mechanisms by which EBV evades tumor suppressor responses to its growth program. LMP1-induced cFLIP was critical for LCL defense against TNFa-mediated programmed cell death, while EBV-induced BATF/IRF4 were critical for LCL BIM suppression and MYC induction. EBV super-enhancer targeted IRF2 protected LCLs against BLIMP1 responses. Collectively, our results identify host/pathogen interaction-driven synthetic lethal targets for therapeutic intervention.
2017-05-11 | GSE93681 | GEO
Project description:Epstein-Barr virus (EBV) causes infectious mononucleosis, triggers multiple sclerosis and is associated with 200,000 cancers/year. EBV colonizes the B-cell compartment and periodically reactivates, inducing expression of 80 viral proteins. Yet much remains unknown about how EBV remodels host cells and dismantles key antiviral responses. We therefore created a proteomic map of EBV-host and EBV-EBV interactions in B-cells undergoing EBV replication, uncovering conserved herpesvirus versus EBV-specific host cell targets. The EBV-encoded G-protein coupled receptor BILF1 associated with MAVS and the UFM1 E3 ligase UFL1. Whereas UFMylation of 14-3-3 proteins drives RIG-I/MAVS signaling, BILF1-directed MAVS UFMylation instead triggered MAVS packaging into mitochondrial-derived vesicles and lysosomal proteolysis. In the absence of BILF1, EBV replication activated the NLRP3 inflammasome, which impaired viral replication and triggered pyroptosis. Our results provide a viral protein interaction network resource, reveal a UFM1-dependent pathway for selective degradation of mitochondrial cargo and highlight BILF1 as a novel therapeutic target.
2023-07-20 | PXD041336 | Pride
Project description:Whether the human tumor virus, Epstein-Barr virus (EBV) promotes breast cancers remains controversial and a potential mechanism has remained elusive. Here we show EBV can infect primary mammary epithelial cells (MECs) that express the attachment receptor, CD21. EBV infection leads to the expansion of early MEC progenitor cells with a stem cell phenotype and enforces a differentiation block. When MECs were implanted as xenografts, EBV infection cooperated with activated Ras and accelerated the formation of breast cancer. Infection in EBV-related tumors was of a latency type II pattern, including expression of latent membrane proteins 1 (LMP1) and 2 (LMP2), similar to nasopharyngeal carcinoma (NPC). A human gene expression signature for EBVness was generated based on the RNA expression profile of the EBV infected primary mammary epithelial cells, tumors. This was signature associated with high grade (40 vs 13.5%) estrogen-receptor-negative status (31.8 vs. 10.5%, p53 mutation (37.5 vs 14.5%) and poor survival. In 11/33 (33%) of tumors positive for EBVness EBV-DNA was found in tumor cells by fluorescent in situ hybridization for the viral LMP1 and BXLF2 genes, while only 4/36 (11%) of EBVness-negative tumors tested positive for EBV DNA. An analysis of the TCGA breast cancer data revealed a correlation of EBVness with presence of the APOBEC mutational signatures consistent with past viral infection. We conclude that a contribution of EBV to breast cancer etiology via a ‘hit-and-run’ mechanism is plausible, in which EBV infection predisposes mammary epithelial cells to malignant transformation, but is not required for the maintenance of the malignant phenotype.
2016-05-04 | GSE81068 | GEO