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1.
Proc Natl Acad Sci U S A ; 118(43)2021 10 26.
Artículo en Inglés | MEDLINE | ID: mdl-34686609

RESUMEN

The Epstein-Barr virus (EBV) transforms resting B cells and is involved in the development of B cell lymphomas. We report here that the viral noncoding RNA EBER2 accelerates B cell growth by potentiating expression of the UCHL1 deubiquitinase that itself increased expression of the Aurora kinases and of cyclin B1. Importantly, this effect was also visible in Burkitt's lymphoma cells that express none of the virus's known oncogenes. Mechanistically, EBER2 bound the UCHL1 messenger RNA (mRNA), thereby bringing a protein complex that includes PU.1, a UCHL1 transactivator, to the vicinity of its promoter. Although the EBV oncogene LMP1 has been suggested to induce UCHL1, we show here that EBER2 plays a much more important role to reach significant levels of the deubiquitinase in infected cells. However, some viruses that carried a polymorphic LMP1 had an increased ability to achieve full UCHL1 expression. This work identifies a direct cellular target of a viral noncoding RNA that is likely to be central to EBV's oncogenic properties.


Asunto(s)
Proliferación Celular/fisiología , Enzimas Desubicuitinizantes/genética , Herpesvirus Humano 4/fisiología , ARN Viral/fisiología , Activación Transcripcional/fisiología , Linfocitos B/citología , Humanos
2.
J Virol ; 94(10)2020 05 04.
Artículo en Inglés | MEDLINE | ID: mdl-32102884

RESUMEN

The Epstein-Barr virus (EBV) causes human cancers, and epidemiological studies have shown that lytic replication is a risk factor for some of these tumors. This fits with the observation that EBV M81, which was isolated from a Chinese patient with nasopharyngeal carcinoma, induces potent virus production and increases the risk of genetic instability in infected B cells. To find out whether this property extends to viruses found in other parts of the world, we investigated 22 viruses isolated from Western patients. While one-third of the viruses hardly replicated, the remaining viruses showed variable levels of replication, with three isolates replicating at levels close to that of M81 in B cells. We cloned one strongly replicating virus into a bacterial artificial chromosome (BAC); the resulting recombinant virus (MSHJ) retained the properties of its nonrecombinant counterpart and showed similarities to M81, undergoing lytic replication in vitro and in vivo after 3 weeks of latency. In contrast, B cells infected with the nonreplicating Western B95-8 virus showed early but abortive replication accompanied by cytoplasmic BZLF1 expression. Sequencing confirmed that rMSHJ is a Western virus, being genetically much closer to B95-8 than to M81. Spontaneous replication in rM81- and rMSHJ-infected B cells was dependent on phosphorylated Btk and was inhibited by exposure to ibrutinib, opening the way to clinical intervention in patients with abnormal EBV replication. As rMSHJ contains the complete EBV genome and induces lytic replication in infected B cells, it is ideal to perform genetic analyses of all viral functions in Western strains and their associated diseases.IMPORTANCE The Epstein-Barr virus (EBV) infects the majority of the world population but causes different diseases in different countries. Evidence that lytic replication, the process that leads to new virus progeny, is linked to cancer development is accumulating. Indeed, viruses such as M81 that were isolated from Far Eastern nasopharyngeal carcinomas replicate strongly in B cells. We show here that some viruses isolated from Western patients, including the MSHJ strain, share this property. Moreover, replication of both M81 and of MSHJ was sensitive to ibrutinib, a commonly used drug, thereby opening an opportunity for therapeutic intervention. Sequencing of MSHJ showed that this virus is quite distant from M81 and is much closer to nonreplicating Western viruses. We conclude that Western EBV strains are heterogeneous, with some viruses being able to replicate more strongly and therefore being potentially more pathogenic than others, and that the virus sequence information alone cannot predict this property.


Asunto(s)
Linfocitos B/virología , Infecciones por Virus de Epstein-Barr/virología , Herpesvirus Humano 4/genética , Replicación Viral/fisiología , Animales , Linfocitos B/patología , Línea Celular , Clonación Molecular , ADN Viral , Modelos Animales de Enfermedad , Genoma Viral , Células HEK293 , Herpesvirus Humano 4/crecimiento & desarrollo , Herpesvirus Humano 4/aislamiento & purificación , Humanos , Neoplasias Nasofaríngeas/virología , Transactivadores/genética
3.
PLoS Pathog ; 14(12): e1007464, 2018 12.
Artículo en Inglés | MEDLINE | ID: mdl-30521644

RESUMEN

The ubiquitous Epstein-Barr virus (EBV) is the primary cause of infectious mononucleosis and is etiologically linked to the development of several malignancies and autoimmune diseases. EBV has a multifaceted life cycle that comprises virus lytic replication and latency programs. Considering EBV infection holistically, we rationalized that prophylactic EBV vaccines should ideally prime the immune system against lytic and latent proteins. To this end, we generated highly immunogenic particles that contain antigens from both these cycles. In addition to stimulating EBV-specific T cells that recognize lytic or latent proteins, we show that the immunogenic particles enable the ex vivo expansion of cytolytic EBV-specific T cells that efficiently control EBV-infected B cells, preventing their outgrowth. Lastly, we show that immunogenic particles containing the latent protein EBNA1 afford significant protection against wild-type EBV in a humanized mouse model. Vaccines that include antigens which predominate throughout the EBV life cycle are likely to enhance their ability to protect against EBV infection.


Asunto(s)
Antígenos Virales/inmunología , Infecciones por Virus de Epstein-Barr/inmunología , Herpesvirus Humano 4/inmunología , Vacunas contra Herpesvirus/inmunología , Linfocitos T Citotóxicos/inmunología , Animales , Ratones , Latencia del Virus
4.
Nucleic Acids Res ; 46(6): 2802-2819, 2018 04 06.
Artículo en Inglés | MEDLINE | ID: mdl-29529302

RESUMEN

The Epstein-Barr virus (EBV) genome encodes several hundred transcripts. We have used ribosome profiling to characterize viral translation in infected cells and map new translation initiation sites. We show here that EBV transcripts are translated with highly variable efficiency, owing to variable transcription and translation rates, variable ribosome recruitment to the leader region and coverage by monosomes versus polysomes. Some transcripts were hardly translated, others mainly carried monosomes, showed ribosome accumulation in leader regions and most likely represent non-coding RNAs. A similar process was visible for a subset of lytic genes including the key transactivators BZLF1 and BRLF1 in cells infected with weakly replicating EBV strains. This suggests that ribosome trapping, particularly in the leader region, represents a new checkpoint for the repression of lytic replication. We could identify 25 upstream open reading frames (uORFs) located upstream of coding transcripts that displayed 5' leader ribosome trapping, six of which were located in the leader region shared by many latent transcripts. These uORFs repressed viral translation and are likely to play an important role in the regulation of EBV translation.


Asunto(s)
Linfocitos B/metabolismo , Herpesvirus Humano 4/genética , Biosíntesis de Proteínas , Ribosomas/metabolismo , Linfocitos B/citología , Linfocitos B/virología , Células Cultivadas , Regulación Viral de la Expresión Génica , Genoma Viral/genética , Herpesvirus Humano 4/fisiología , Humanos , Proteínas Inmediatas-Precoces/genética , Proteínas Inmediatas-Precoces/metabolismo , Mutación , Sistemas de Lectura Abierta/genética , Ribosomas/genética
5.
PLoS Pathog ; 12(1): e1005405, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-26800049

RESUMEN

The Epstein-Barr virus (EBV) infects and transforms B-lymphocytes with high efficiency. This process requires expression of the viral latent proteins and of the 3 miR-BHRF1 microRNAs. Here we show that B-cells infected by a virus that lacks these non-coding RNAs (Δ123) grew more slowly between day 5 and day 20, relative to wild type controls. This effect could be ascribed to a reduced S phase entry combined with a moderately increased apoptosis rate. Whilst the first phenotypic trait was consistent with an enhanced PTEN expression in B-cells infected with Δ123, the second could be explained by very low BHRF1 protein and RNA levels in the same cells. Indeed, B-cells infected either by a recombinant virus that lacks the BHRF1 protein, a viral bcl-2 homolog, or by Δ123 underwent a similar degree of apoptosis, whereas knockouts of both BHRF1 microRNAs and protein proved transformation-incompetent. We find that that the miR-BHRF1-3 seed regions, and to a lesser extent those of miR-BHRF1-2 mediate these stimulatory effects. After this critical period, B-cells infected with the Δ123 mutant recovered a normal growth rate and became more resistant to provoked apoptosis. This resulted from an enhanced BHRF1 protein expression relative to cells infected with wild type viruses and correlated with decreased p27 expression, two pro-oncogenic events. The upregulation of BHRF1 can be explained by the observation that large BHRF1 mRNAs are the source of BHRF1 protein but are destroyed following BHRF1 microRNA processing, in particular of miR-BHRF1-2. The BHRF1 microRNAs are unlikely to directly target p27 but their absence may facilitate the selection of B-cells that express low levels of this protein. Thus, the BHRF1 microRNAs allowed a time-restricted expression of the BHRF1 protein to innocuously expand the virus B-cell reservoir during the first weeks post-infection without increasing long-term immune pressure.


Asunto(s)
Linfocitos B/virología , Transformación Celular Viral/genética , Regulación Viral de la Expresión Génica/genética , Herpesvirus Humano 4/genética , MicroARNs/genética , Proteínas Virales/genética , Apoptosis/genética , Proliferación Celular , Infecciones por Virus de Epstein-Barr/genética , Técnicas de Inactivación de Genes , Humanos , Immunoblotting , Etiquetado Corte-Fin in Situ , Fosfohidrolasa PTEN/biosíntesis , Fosfohidrolasa PTEN/genética , Antígeno Nuclear de Célula en Proliferación/biosíntesis , Antígeno Nuclear de Célula en Proliferación/genética , ARN Viral/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa
6.
PLoS Pathog ; 11(12): e1005344, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26694854

RESUMEN

The Epstein-Barr virus (EBV) is a B lymphotropic virus that infects the majority of the human population. All EBV strains transform B lymphocytes, but some strains, such as M81, also induce spontaneous virus replication. EBV encodes 22 microRNAs (miRNAs) that form a cluster within the BART region of the virus and have been previously been found to stimulate tumor cell growth. Here we describe their functions in B cells infected by M81. We found that the BART miRNAs are downregulated in replicating cells, and that exposure of B cells in vitro or in vivo in humanized mice to a BART miRNA knockout virus resulted in an increased proportion of spontaneously replicating cells, relative to wild type virus. The BART miRNAs subcluster 1, and to a lesser extent subcluster 2, prevented expression of BZLF1, the key protein for initiation of lytic replication. Thus, multiple BART miRNAs cooperate to repress lytic replication. The BART miRNAs also downregulated pro- and anti-apoptotic mediators such as caspase 3 and LMP1, and their deletion did not sensitize B-cells to apoptosis. To the contrary, the majority of humanized mice infected with the BART miRNA knockout mutant developed tumors more rapidly, probably due to enhanced LMP1 expression, although deletion of the BART miRNAs did not modify the virus transforming abilities in vitro. This ability to slow cell growth could be confirmed in non-humanized immunocompromized mice. Injection of resting B cells exposed to a virus that lacks the BART miRNAs resulted in accelerated tumor growth, relative to wild type controls. Therefore, we found that the M81 BART miRNAs do not enhance B-cell tumorigenesis but rather repress it. The repressive effects of the BART miRNAs on potentially pathogenic viral functions in infected B cells are likely to facilitate long-term persistence of the virus in the infected host.


Asunto(s)
Linfocitos B/virología , Infecciones por Virus de Epstein-Barr/genética , Regulación Viral de la Expresión Génica/genética , Herpesvirus Humano 4/genética , MicroARNs/genética , Replicación Viral/genética , Animales , Western Blotting , Transformación Celular Viral/genética , Genes Virales , Humanos , Inmunohistoquímica , Inmunoprecipitación , Ratones , ARN Viral/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Transfección
7.
J Virol ; 89(23): 12178-88, 2015 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-26401047

RESUMEN

UNLABELLED: Epstein-Barr virus (EBV) is an oncogenic human herpesvirus involved in the pathogenesis of Burkitt's lymphoma (BL) and various other lymphoproliferative disorders. In BL, EBV protein expression is restricted to EBV nuclear antigen 1 (EBNA1), but small noncoding RNAs such as EBV-encoded small RNAs (EBERs) and microRNAs (miRNAs) can also be detected. miRNAs play major roles in crucial processes such as proliferation, differentiation, and cell death. It has recently become clear that alterations in the expression profile of miRNAs contribute to the pathogenesis of a number of malignancies. During latent infection, EBV expresses 25 viral pre-miRNAs and modulates the expression of specific cellular miRNAs, such as miR-155 and miR-146, which potentially play a role in oncogenesis. Here, we established the small-RNA expression profiles of three BL cell lines. Using large-scale sequencing coupled to Northern blotting and real-time reverse transcription-PCR (RT-PCR) analysis validation, we demonstrated the differential expression of some cellular and viral miRNAs. High-level expression of the miR-183-96-182 cluster and EBV miR-BamHI A rightward transcript (miR-BART) cluster was significantly associated with EBV type I latency. This expression was not affected by viral reactivation since transforming growth factor ß1 (TGF-ß1) stimulation did not significantly change the miRNA profiles. However, using several approaches, including de novo infection with a mutant virus, we present evidence that the expression of latent membrane protein 1 (LMP-1) triggered downregulation of the expression of the miR-183-96-182 cluster. We further show that this effect involves the Akt signaling pathway. IMPORTANCE: In addition to expressing their own miRNAs, herpesviruses also impact the expression levels of cellular miRNAs. This regulation can be either positive or negative and usually results in the perturbation of pathways to create a cellular environment that is more "virus-friendly." For example, EBV induces the expression of miR-155, a well-characterized oncomiR, which leads to increased cell proliferation and decreased cell death. Here, we show that EBV-encoded LMP-1 is also involved in the downregulation of a cluster of three miRNAs, miR-183, -96, and -182, which are known to be also repressed in several cancers. We therefore identify yet another potential player in EBV-induced oncogenesis.


Asunto(s)
Regulación de la Expresión Génica/genética , MicroARNs/metabolismo , Familia de Multigenes/genética , Proteínas de la Matriz Viral/genética , Secuencia de Bases , Northern Blotting , Clonación Molecular , Cartilla de ADN/genética , Perfilación de la Expresión Génica , Técnicas de Inactivación de Genes , Humanos , MicroARNs/genética , Datos de Secuencia Molecular , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Análisis de Secuencia de ARN
8.
Curr Top Microbiol Immunol ; 390(Pt 1): 119-48, 2015.
Artículo en Inglés | MEDLINE | ID: mdl-26424646

RESUMEN

The Epstein-Barr virus is etiologically linked with the development of benign and malignant diseases, characterized by their diversity and a heterogeneous geographic distribution across the world. The virus possesses a 170-kb-large genome that encodes for multiple proteins and non-coding RNAs. Early on there have been numerous attempts to link particular diseases with particular EBV strains, or at least with viral genetic polymorphisms. This has given rise to a wealth of information whose value has been difficult to evaluate for at least four reasons. First, most studies have looked only at one particular gene and missed the global picture. Second, they usually have not studied sufficient numbers of diseased and control cases to reach robust statistical significance. Third, the functional significance of most polymorphisms has remained unclear, although there are exceptions such as the 30-bp deletion in LMP1. Fourth, different biological properties of the virus do not necessarily equate with a different pathogenicity. This was best illustrated by the type 1 and type 2 viruses that markedly differ in terms of their transformation abilities, yet do not seem to cause different diseases. Reciprocally, environmental and genetic factors in the host are likely to influence the outcome of infections with the same virus type. However, with recent developments in recombinant virus technology and in the availability of high throughput sequencing, the tide is now turning. The availability of 23 complete or nearly complete genomes has led to the recognition of viral subtypes, some of which possess nearly identical genotypes. Furthermore, there is growing evidence that some genetic polymorphisms among EBV strains markedly influence the biological and clinical behavior of the virus. Some virus strains are endowed with biological properties that explain crucial clinical features of patients with EBV-associated diseases. Although we now have a better overview of the genetic diversity within EBV genomes, it has also become clear that defining phenotypic traits evinced by cells infected by different viruses usually result from the combination of multiple polymorphisms that will be difficult to identify in their entirety. However, the steadily increasing number of sequenced EBV genomes and cloned EBV BACS from diseased and healthy patients will facilitate the identification of the key polymorphisms that condition the biological and clinical behavior of the viruses. This will allow the development of preventative and therapeutic approaches against highly pathogenic viral strains.


Asunto(s)
Infecciones por Virus de Epstein-Barr/virología , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/aislamiento & purificación , Polimorfismo Genético , Secuencia de Aminoácidos , Genotipo , Herpesvirus Humano 4/química , Herpesvirus Humano 4/clasificación , Humanos , Datos de Secuencia Molecular , Filogenia , Alineación de Secuencia , Proteínas Virales/química , Proteínas Virales/genética , Proteínas Virales/metabolismo
9.
mBio ; 14(5): e0178423, 2023 Oct 31.
Artículo en Inglés | MEDLINE | ID: mdl-37830871

RESUMEN

IMPORTANCE: The Epstein-Barr virus efficiently infects and transforms B lymphocytes. During this process, infectious viral particles transport the viral genome to the nucleus of target cells. We show here that these complex viral structures serve additional crucial roles by activating transcription of the transforming genes encoded by the virus. We show that components of the infectious particle sequentially activate proinflammatory B lymphocyte signaling pathways that, in turn, activate viral gene expression but also cause cytokine release. However, virus infection activates expression of ZFP36L1, an RNA-binding stress protein that limits the length and the intensity of the cytokine response. Thus, the infectious particles can activate viral gene expression and initiate cellular transformation at the price of a limited immune response.


Asunto(s)
Infecciones por Virus de Epstein-Barr , Humanos , Herpesvirus Humano 4/fisiología , Citocinas/metabolismo , Linfocitos B , Transducción de Señal , Factor 1 de Respuesta al Butirato/metabolismo
10.
Front Immunol ; 12: 796379, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-34975903

RESUMEN

Whole genome sequencing of Epstein-Barr virus (EBV) isolates from around the world has uncovered pervasive strain heterogeneity, but the forces driving strain diversification and the impact on immune recognition remained largely unknown. Using a data mining approach, we analyzed more than 300 T-cell epitopes in 168 published EBV strains. Polymorphisms were detected in approximately 65% of all CD8+ and 80% of all CD4+ T-cell epitopes and these numbers further increased when epitope flanking regions were included. Polymorphisms in CD8+ T-cell epitopes often involved MHC anchor residues and resulted in changes of the amino acid subgroup, suggesting that only a limited number of conserved T-cell epitopes may represent generic target antigens against different viral strains. Although considered the prototypic EBV strain, the rather low degree of overlap with most other viral strains implied that B95.8 may not represent the ideal reference strain for T-cell epitopes. Instead, a combinatorial library of consensus epitopes may provide better targets for diagnostic and therapeutic purposes when the infecting strain is unknown. Polymorphisms were significantly enriched in epitope versus non-epitope protein sequences, implicating immune selection in driving strain diversification. Remarkably, CD4+ T-cell epitopes in EBNA2, EBNA-LP, and the EBNA3 family appeared to be under negative selection pressure, hinting towards a beneficial role of immune responses against these latency type III antigens in virus biology. These findings validate this immunoinformatics approach for providing novel insight into immune targets and the intricate relationship of host defense and virus evolution that may also pertain to other pathogens.


Asunto(s)
Variación Antigénica , Antígenos Virales/genética , Epítopos de Linfocito T/genética , Heterogeneidad Genética , Herpesvirus Humano 4/genética , Polimorfismo Genético , Algoritmos , Antígenos Virales/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD4-Positivos/virología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/virología , Minería de Datos , Bases de Datos Genéticas , Epítopos de Linfocito T/inmunología , Herpesvirus Humano 4/inmunología
11.
Nat Microbiol ; 4(12): 2475-2486, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31501540

RESUMEN

The Epstein-Barr virus M81 strain, isolated from a nasopharyngeal carcinoma, induces potent spontaneous virus production in infected B cells. We found that the M81 non-coding Epstein-Barr-encoded RNA EBER2, which carries polymorphisms that are mainly restricted to viruses found in endemic nasopharyngeal carcinomas, markedly stimulated this process. M81 EBER2 increased CXCL8 expression, and this chemokine enhanced spontaneous lytic replication levels in M81-infected B cells. Both events resulted from the endocytosis of extracellular vesicles containing EBER2 that were generated by neighbouring M81-infected B cells, thereby generating a paracrine loop. These effects were strictly dependent on a functional Toll-like receptor 7 (TLR7), a sensor of single-stranded RNA located in the endosome of these cells. These unique properties of M81 EBER2 could be ascribed to its unusually high expression level and to the ability of its single-stranded region to activate TLR7; both of these properties were dependent on M81-specific polymorphisms. Thus, M81 induced chronic inflammation in its target cells and this resulted in increased virus production. These observations provide a mechanistic molecular link between M81 virus replication-a central viral function and a cancer risk factor-and the production of a chemokine involved in inflammation and carcinogenesis.


Asunto(s)
Infecciones por Virus de Epstein-Barr/virología , Herpesvirus Humano 4/crecimiento & desarrollo , Herpesvirus Humano 4/genética , Carcinoma Nasofaríngeo/metabolismo , Carcinoma Nasofaríngeo/virología , Neoplasias Nasofaríngeas/virología , ARN no Traducido/genética , Replicación Viral , Linfocitos B/virología , Quimiocinas/metabolismo , Infecciones por Virus de Epstein-Barr/inmunología , Células HEK293 , Interacciones Huésped-Patógeno , Humanos , Interleucina-8/metabolismo , Carcinoma Nasofaríngeo/genética , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/metabolismo , Virus Oncogénicos , ARN Viral , Receptor Toll-Like 7/metabolismo , Cultivo de Virus
12.
PLoS One ; 14(9): e0222847, 2019.
Artículo en Inglés | MEDLINE | ID: mdl-31568538

RESUMEN

The Epstein-Barr virus (EBV) induces B-cell proliferation with high efficiency through expression of latent proteins and microRNAs. This process takes place in vivo soon after infection, presumably to expand the virus reservoir, but can also induce pathologies, e.g. an infectious mononucleosis (IM) syndrome after primary infection or a B-cell lymphoproliferation in immunosuppressed individuals. In this paper, we investigated the growth characteristics of EBV-infected B-cells isolated from transplant recipients or patients with IM. We found that these cells grew and withstood apoptosis at highly variable rates, suggesting that the expansion rate of the infected B-cells widely varies between individuals, thereby influencing the size of the B-cell reservoir and the ability to form tumors in infected individuals. All viruses investigated were type 1 and genetically close to western strains. EBV-infected B-cells expressed the transforming EBV latent genes and microRNAs (miRNAs) at variable levels. We found that the B-cell growth rates positively correlated with the BHRF1 miRNA levels. Comparative studies showed that infected B-cells derived from transplant recipients with iEBVL on average expressed higher levels of EBV miR-BHRF1 miRNAs and grew more rapidly than B-cells from IM patients, suggesting infection by more transforming viruses. Altogether, these findings suggest that EBV infection has a highly variable impact on the B-cell compartment that probably reflects the genetic diversity of both the virus and the host. It also demonstrates the unexpected finding that B-cells from different individuals can grow at different speed under the influence of the same virus infection.


Asunto(s)
Linfocitos B/virología , Regulación Viral de la Expresión Génica , Genes Virales , Herpesvirus Humano 4/genética , Huésped Inmunocomprometido , Mononucleosis Infecciosa/virología , MicroARNs/genética , Adulto , Anciano , Linfocitos B/inmunología , Linfocitos B/patología , Línea Celular Transformada , Proliferación Celular , Femenino , Trasplante de Células Madre Hematopoyéticas , Herpesvirus Humano 4/crecimiento & desarrollo , Herpesvirus Humano 4/metabolismo , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/inmunología , Humanos , Mononucleosis Infecciosa/inmunología , Mononucleosis Infecciosa/patología , Trasplante de Riñón , Masculino , MicroARNs/inmunología , Persona de Mediana Edad , Cultivo Primario de Células , Proteínas Virales/genética , Proteínas Virales/inmunología
13.
Oncotarget ; 8(6): 10238-10254, 2017 Feb 07.
Artículo en Inglés | MEDLINE | ID: mdl-28052012

RESUMEN

The Epstein-Barr virus (EBV) is etiologically associated with the development of multiple types of tumors, but it is unclear whether this diversity is due to infection with different EBV strains. We report a comparative characterization of SNU719, GP202, and YCCEL1, three EBV strains that were isolated from gastric carcinomas, M81, a virus isolated in a nasopharyngeal carcinoma and several well-characterized laboratory type A strains. We found that B95-8, Akata and GP202 induced cell growth more efficiently than YCCEL1, SNU719 and M81 and this correlated positively with the expression levels of the viral BHRF1 miRNAs. In infected B cells, all strains except Akata and B95-8 induced lytic replication, a risk factor for carcinoma development, although less efficiently than M81. The panel of viruses induced tumors in immunocompromised mice with variable speed and efficacy that did not strictly mirror their in vitro characteristics, suggesting that additional parameters play an important role. We found that YCCEL1 and M81 infected primary epithelial cells, gastric carcinoma cells and gastric spheroids more efficiently than Akata or B95-8. Reciprocally, Akata and B95-8 had a stronger tropism for B cells than YCCEL1 or M81. These data suggest that different EBV strains will induce the development of lymphoid tumors with variable efficacy in immunocompromised patients and that there is a parallel between the cell tropism of the viral strains and the lineage of the tumors they induce. Thus, EBV strains can be endowed with properties that will influence their transforming abilities and the type of tumor they induce.


Asunto(s)
Carcinoma/virología , Transformación Celular Viral , Infecciones por Virus de Epstein-Barr/virología , Herpesvirus Humano 4/patogenicidad , Neoplasias Nasofaríngeas/virología , Neoplasias Gástricas/virología , Animales , Linfocitos B/metabolismo , Linfocitos B/patología , Linfocitos B/virología , Células CACO-2 , Carcinoma/genética , Carcinoma/metabolismo , Carcinoma/patología , Proliferación Celular , Técnicas de Cocultivo , Células Epiteliales/metabolismo , Células Epiteliales/patología , Células Epiteliales/virología , Infecciones por Virus de Epstein-Barr/genética , Infecciones por Virus de Epstein-Barr/metabolismo , Infecciones por Virus de Epstein-Barr/patología , Células HEK293 , Herpesvirus Humano 4/clasificación , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/aislamiento & purificación , Interacciones Huésped-Patógeno , Humanos , Ratones SCID , MicroARNs/genética , MicroARNs/metabolismo , Carcinoma Nasofaríngeo , Neoplasias Nasofaríngeas/genética , Neoplasias Nasofaríngeas/metabolismo , Neoplasias Nasofaríngeas/patología , ARN Viral/genética , ARN Viral/metabolismo , Neoplasias Gástricas/genética , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patología , Factores de Tiempo , Proteínas Virales/genética , Proteínas Virales/metabolismo , Tropismo Viral , Internalización del Virus , Replicación Viral
14.
Environ Sci Pollut Res Int ; 9(2): 122-9, 2002.
Artículo en Inglés | MEDLINE | ID: mdl-12008292

RESUMEN

In declining forests of the Vosges mountains (northeast of France), we previously observed that the yellowing of spruce (Picea abies L. cv. Karsten) needles was associated with impairment of the free radical scavenging capacity of the cells and coincided with chronic exposure to ozone. Chloroplasts of yellow needles were characterized by an abnormal accumulation of photosystem II (PSII) D1-protein in the thylakoids. Further experiments carried out on declining and decline-resistant individual spruce trees characterized in previous studies showed that needle yellowing was associated with impairment of the overall anti-oxidative defense in both the cytosol and the chloroplasts. Both enzymic (peroxidases) and non-enzymic (carotenoids) oxidant scavengers were shown to be affected in the declining spruce. PSII D1-protein accumulation seemed to result from a stabilization of the polypeptide, which led us to hypothesize that oxidative processes might interfere with the specific degradation of this protein in declining spruce, with destructive consequences for the photosystems.


Asunto(s)
Cloroplastos/metabolismo , Proteínas del Complejo del Centro de Reacción Fotosintética/metabolismo , Árboles/metabolismo , Northern Blotting , Carotenoides/metabolismo , Clorofila/metabolismo , Depuradores de Radicales Libres/metabolismo , Humanos , Complejos de Proteína Captadores de Luz , Peroxidasas/metabolismo , Complejo de Proteína del Fotosistema II , Hojas de la Planta , Tilacoides/metabolismo
15.
J Virol ; 79(4): 2274-86, 2005 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-15681429

RESUMEN

Autonomous parvoviruses are characterized by their stringent dependency on host cell S phase and their cytopathic effects on neoplastic cells. To better understand the interactions between the virus and its host cell, we used oligonucleotide arrays that carry more than 19,000 unique human gene sequences to profile the gene expression of the human hepatocellular carcinoma cell line QGY-7703 at two time points after parvovirus H-1 infection. At the 6-h time point, a single gene was differentially expressed with a >2.5-fold change. At 12 h, 105 distinct genes were differentially expressed in virus-infected cells compared to mock-treated cells, with 93% of these genes being down-regulated. These repressed genes clustered mainly into classes involved in transcriptional regulation, signal transduction, immune and stress response, and apoptosis, as exemplified by genes encoding the transcription factors Myc, Jun, Fos, Ids, and CEBPs. Quantitative real-time reverse transcription-PCR analysis on selected genes validated the array data and allowed the changes in cellular gene expression to be correlated with the accumulation of viral transcripts and NS1 protein. Western blot analysis of several cellular proteins supported the array results and substantiated the evidence given by these and other data to suggest that the H-1 virus kills QGY-7703 cells by a nonapoptotic process. The promoter regions of most of the differentially expressed genes analyzed fail to harbor any motif for sequence-specific binding of NS1, suggesting that direct binding of NS1 to cellular promoters may not participate in the modulation of cellular gene expression in H-1 virus-infected cells.


Asunto(s)
Apoptosis , Línea Celular Tumoral/virología , Perfilación de la Expresión Génica , Parvovirus/fisiología , Carcinoma Hepatocelular/patología , Proliferación Celular , Regulación de la Expresión Génica , Humanos , ARN Mensajero/metabolismo , ARN Viral/genética , ARN Viral/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Factores de Tiempo
16.
Microbiology (Reading) ; 148(Pt 7): 2111-2123, 2002 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-12101299

RESUMEN

The DUP240 gene family of Saccharomyces cerevisiae is composed of 10 members. They encode proteins of about 240 amino acids which contain two predicted transmembrane domains. Database searches identified only one homologue in the closely related species Saccharomyces bayanus, indicating that the DUP240 genes encode proteins specific to Saccharomyces sensu stricto. The short-flanking homology PCR gene-replacement strategy with a variety of selective markers for replacements, and classical genetic methods, were used to generate strains deleted for all 10 DUP240 genes. All of the knock-out strains were viable and had similar growth kinetics to the wild-type. Two-hybrid screens, hSos1p fusions and GFP fusions were carried out; the results indicated that the Dup240 proteins are membrane associated, and that some of them are concentrated around the plasma membrane.


Asunto(s)
Proteínas de la Membrana/metabolismo , Familia de Multigenes , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/genética , Secuencia de Aminoácidos , Membrana Celular/metabolismo , Eliminación de Gen , Genes Esenciales , Proteínas Fluorescentes Verdes , Proteínas Luminiscentes/genética , Proteínas Luminiscentes/metabolismo , Proteínas de la Membrana/genética , Proteínas Recombinantes de Fusión , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Análisis de Secuencia de ADN , Fracciones Subcelulares/metabolismo , Secuencias Repetidas en Tándem/genética , Transformación Genética , Técnicas del Sistema de Dos Híbridos
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