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Trivedi P Spinsanti P Cuomo L Volpe M Takada K Frati L Faggioni A 《Journal of virology》2001,75(10):4929-4935
Epstein-Barr virus (EBV)-negative Burkitt lymphomas (BLs) can be infected in vitro with prototype EBV strains to study how the virus may affect the phenotype of tumor cells. Studies thus far have concentrated on the use of transforming B95-8 and nontransforming P3HR1 strains. Immunological and phenotypic differences between the sublines infected with these two strains were reported. The majority of these differences, if not all, can be attributed to the lack of EBNA-2 coding sequences in the P3HR1 strain. The recent development of a selectable Akata strain has opened up new possibilities for infecting epithelial and T cells as well. We infected five EBV-negative BL lines with the recombinant Akata virus. Our results indicate that the infected cell lines BL28, Ramos, and DG75 express EBNA-1, EBNA-2, and LMP1, the viral proteins associated with type III latency, and use both YUK and QUK splices. In contrast, two EBV-negative variants of Akata and Mutu when reinfected displayed restricted type I latency and expressed only EBNA-1. All clones of infected Mutu cells used the QUK splice exclusively. The usage of Qp was observed in a majority of Akata clones. Some Akata clones, however, were found to have double promoter usage (Qp and C/Wp) but at 4 months after infection did not express EBNA-2. The results demonstrate differential regulation of EBV latency in BLs with the same recombinant viral strain and suggest that the choice of latency type may be cell dependent. The restricted latency observed for infected Akata and Mutu cells indicates that a BL may opt for type I latency in the absence of immune pressure as well. 相似文献
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CD4+ T-cell responses to Epstein-Barr virus (EBV) latent-cycle antigens and the recognition of EBV-transformed lymphoblastoid cell lines 总被引:2,自引:0,他引:2
Long HM Haigh TA Gudgeon NH Leen AM Tsang CW Brooks J Landais E Houssaint E Lee SP Rickinson AB Taylor GS 《Journal of virology》2005,79(8):4896-4907
There is considerable interest in the potential of Epstein-Barr virus (EBV) latent antigen-specific CD4+ T cells to act as direct effectors controlling EBV-induced B lymphoproliferations. Such activity would require direct CD4+ T-cell recognition of latently infected cells through epitopes derived from endogenously expressed viral proteins and presented on the target cell surface in association with HLA class II molecules. It is therefore important to know how often these conditions are met. Here we provide CD4+ epitope maps for four EBV nuclear antigens, EBNA1, -2, -3A, and -3C, and establish CD4+ T-cell clones against 12 representative epitopes. For each epitope we identify the relevant HLA class II restricting allele and determine the efficiency with which epitope-specific effectors recognize the autologous EBV-transformed B-lymphoblastoid cell line (LCL). The level of recognition measured by gamma interferon release was consistent among clones to the same epitope but varied between epitopes, with values ranging from 0 to 35% of the maximum seen against the epitope peptide-loaded LCL. These epitope-specific differences, also apparent in short-term cytotoxicity and longer-term outgrowth assays on LCL targets, did not relate to the identity of the source antigen and could not be explained by the different functional avidities of the CD4+ clones; rather, they appeared to reflect different levels of epitope display at the LCL surface. Thus, while CD4+ T-cell responses are detectable against many epitopes in EBV latent proteins, only a minority of these responses are likely to have therapeutic potential as effectors directly recognizing latently infected target cells. 相似文献
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E Tatsumi 《Human cell》1992,5(1):79-86
Two facts need to be pointed out to help explain why the history of Epstein-Barr virus (EBV) research has been inseparable from that of the studies with human hematopoietic cell lines of neoplastic and non-neoplastic origin. One is that Burkitt lymphoma (BL) cell lines, EBV-positive or-negative, can be established in culture quite easily. Thus, the BL cell lines which Epstein established were indeed some of the first hematopoietic as well as virus-carrying cell lines of human neoplastic origin. The other is that EBV-positive B-cell lymphoblastoid cell lines (B-LCL) of normal origin can be grown from samples of sero-positive individuals. B-LCL were often mistakenly regarded as being of neoplastic origin, but are almost always of normal cell origin. Very rarely, however, B-LCL with the same clonal markers as those of neoplastic cells have also been obtained. While the development of B-LCL has been referred to as the in vitro viral immortalization of human B cells and as a phenomenon representing the potential oncogenicity of EBV, the phenotypic and genotypic differences between B-LCL and EBV-carrying BL cells are obvious, indicating that the development of B-LCL per se does not prove the oncogenic activity of EBV. Two EBV-derived antigens, EBNA2 and latent-infection membrane protein (LMP), which are strongly expressed by B-LCL but not by BL cells, have recently been detected in EBV-positive proliferative B cells in patients with organ transplants, suggesting that the proliferating of B-LCL-like cells may take place as an initial step of the multi-step in vivo oncogenesis of EBV.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
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Chromosomal localization of the Epstein-Barr virus (EBV) genome in Bloom's syndrome B-lymphoblastoid cell lines transformed with EBV 总被引:3,自引:0,他引:3
The localization of the Epstein-Barr virus (EBV) genome in chromosomes of human B-lymphoblastoid cell lines (LCLs) transformed with EBV, and the effect of EBV DNA on the level of sister chromatid exchange (SCE) in Bloom's syndrome (BS) B-LCLs, were examined with chromosomal in situ hybridization techniques using a 3H-EBV DNA probe. EBV DNA was detected in chromosomes 1–5 and 13–15 at specific G band regions in BS as well as in normal B-LCLs, regardless of SCE. Several chromosomal sites (1p31, 1q31, 4q22–24, 5q21, 13q21, 14q21) carrying EBV DNA seemed to be very characteristic in normal as well as in BS B-LCLs. There was no statistically significant difference in silver grain counts due to EBV DNA and their distribution in different chromosomes or groups among normal and BS B-LCLs with normal and high SCE. These findings strongly indicate that EBV infection did not introduce a correcting factor for BS SCE. 相似文献
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Different activation of Epstein-Barr virus immediate-early and early genes in Burkitt lymphoma cells and lymphoblastoid cell lines. 总被引:6,自引:2,他引:4
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C Bogedain P Alliger F Schwarzmann M Marschall H Wolf W Jilg 《Journal of virology》1994,68(2):1200-1203
Specific expression of the Epstein-Barr virus (EBV) immediate-early and early gene products Zta, Rta, I'ta, and MSta by a recombinant vaccinia virus system allowed us to analyze the first steps in the induction of the lytic cycle in EBV-infected Burkitt lymphoma (BL) cells and lymphoblastoid cell lines (LCLs). Significant differences in the induction of early genes were found between these cell types: whereas in BL cells the trans activator Zta was found to induce key steps of the early lytic cycle, only minor activities of Zta were noted in LCLs. Contrary to Zta, the trans activator Rta was found to be highly effective in LCLs. These observations suggest that Rta may play an important role in the activation of the early lytic cycle in LCLs, although it cannot be activated by Zta. The latter may be a reason for the lower tendency of LCLs to switch into the lytic cycle compared with BL cells or differentiated epithelial cells. 相似文献
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The spliced BZLF1 gene of Epstein-Barr virus (EBV) transactivates an early EBV promoter and induces the virus productive cycle. 总被引:38,自引:26,他引:12
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A spliced cDNA spanning the Epstein-Barr virus BZLF1 gene expresses the BZLF1 protein and is active in inducing the virus productive cycle. A deletion mutant which lacks the N-terminal half of the protein is inactive. Cotransfection experiments in EBV-negative B-lymphocyte cell lines demonstrated that the BZLF1 gene activates the promoter for the BSLF2 + BMLF1 gene in the absence of any other EBV gene product. These results confirmed that the spliced BZLF1 gene is the transactivating gene structure in BamHI-Z. 相似文献
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Epstein-Barr virus (EBV)-negative B-lymphoma cell lines for clonal isolation and replication of EBV recombinants.
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Previous experiments have demonstrated that positive selection markers recombined into the Epstein-Barr virus (EBV) genome enable the isolation of transforming or nontransforming mutant EBV recombinants in EBV-negative B-lymphoma (BL) cell lines (A. Marchini, J. I. Cohen, and E. Kieff, J. Virol. 66:3214-3219, 1992; F. Wang, A. Marchini, and E. Kieff, J. Virol. 65:1701-1709, 1991). However, virus has been recovered from a BL cell clone (BL41) infected with an EBV recombinant in only one instance (Wang et al., J. Virol. 65:1701-1709, 1991). We now compare the utility of four EBV-negative BL lines, BJAB, BL30, BL41, and Loukes, for isolating EBV recombinants and supporting their subsequent replication. Transforming or nontransforming EBV recombinants carrying a simian virus 40 promoter-hygromycin phosphotransferase (HYG) cassette were cloned by selecting newly infected BL cells for HYG expression. Most of the infected BL clones contained EBV episomes, and EBV gene expression was largely restricted to EBNA-1. Although the BJAB cell line was a particularly good host for isolating EBV recombinants (Marchini et al., J. Virol. 66:3214-3219, 1992), it was largely nonpermissive for virus replication, even in response to heterologous expression of the BZLF1 immediate-early transactivator. In contrast, approximately 50% of infected BL41, BL30, or Loukes cell clones responded to lytic cycle induction. Frequently, a substantial fraction of infected cells expressed the late lytic infection viral protein, gp350/220, and released infectious virus. Since BL cells do not depend on EBV for growth, transforming and nontransforming EBV recombinants were isolated and passaged. 相似文献
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Ohashi M Horie K Hoshikawa Y Nagata K Osaki M Ito H Sairenji T 《Microbes and infection / Institut Pasteur》2007,9(2):150-159
As a new model to elucidate molecular mechanisms in Epstein-Barr virus (EBV) activation, we tested the tetracycline-inducible (Tet-On)/BZLF1-oriP plasmid system in Raji cells. Cells transfected with this Tet-On plasmid did not activate EBV by doxycycline and surprisingly EBV latency was disrupted with large amounts of BMRF1 protein (EA-D) being accumulated in the cells. Brilliant EA-D fluorescence was markedly condensed in small sized cells, intra-cellular vesicles, and extra-cellular particles. Scanning electron microscopy demonstrated the extra-cellular particles to be covered with a membrane. EA-D molecules of 58, 50, 48, and 44kDa were expressed in the cells. The high (58 and 50kDa) and low (48 and 44kDa) EA-D molecules appeared in the early and late stages, respectively. Low EA-D molecules were detected mostly in EA-D positive cells separated into the heaviest density layer of a discontinuous Percoll gradient. Such molecules could be created from high EA-D molecules by protein phosphatase treatment. The EA-D molecules that appeared similar were detected in EBV-activated P3HR-1 and Akata cells. Several hypotheses concerning the accumulation of EA-D molecules of various polymorphic forms and their phosphorylation/dephosphorylation in this model system are presented, with possible biological and clinical relevance. 相似文献
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trans activation of the latent Epstein-Barr virus (EBV) genome after transfection of the EBV DNA fragment. 总被引:56,自引:37,他引:19
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Transfection of Epstein-Barr virus (EBV)-nonproducer Raji cells with the BamHI Z fragment of EBV DNA induced antigens that were detected with human antiserum against EBV-specific early antigens. Northern blot analysis of transfected cells revealed that one intense RNA band hybridized with the BamHI H and F fragments but not with the BamHI Z fragment. Cooperation between the BamHI H, F, and BamHI Z regions was also confirmed in baby hamster kidney cells that were cotransfected with both fragments. These results indicate that the transfected BamHI Z fragment of EBV DNA induces a trans-acting factor which activates the gene expression of the BamHI H and F region and that the BamHI Z region possibly plays an important role in the latency of EBV. 相似文献
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Isolation of Epstein-Barr virus (EBV)-negative cell clones from the EBV-positive Burkitt's lymphoma (BL) line Akata: malignant phenotypes of BL cells are dependent on EBV. 总被引:20,自引:9,他引:11
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During cultivation of the Epstein-Barr virus (EBV)-positive Burkitt's lymphoma (BL) line Akata, it was noted that EBV DNA is lost from some of the cells. Isolation of EBV-positive and EBV-negative clones with the same origin made it possible to examine the effects of EBV in BL cells. The results indicate that malignant phenotypes of BL, such as growth in low serum, anchorage-independent growth in soft agar, and tumorigenicity in nude mice, are dependent on the presence of EBV genomes and underline the oncogenic function of EBV in human cancer. 相似文献
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Increased hexose uptake is a marker for viral transformation, as has been shown in non-human fibroblasts transformed by oncogenic viruses. If this phenomenon is a general expression of viral induced transformation it should also apply on different oncogenic virus-cell systems. Recently two human EBV-negative lymphoma lines were converted to a stable EBV-positive state by infection with EBV. According to their biochemical and biological properties they enable us to study events associated with EBV-transformation. We analysed the uptake of (3H) glucosamine and (3H) 2-deoxy-D-glucose into BJAB and Ramos and their EBV-converted sublines and found a clear increase of the rate of uptake of both sugars in the EBV-positive sublines. Control experiments confirmed that the increased uptake was due to alterations on the level of the hexose membrane carriers and not due to increased metabolism. The observation of increased hexose uptake in the only presented available virus transformed human cell system is a strong argument for the general importance of this transformation-associated membrane change. 相似文献
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Spontaneous reduction in Epstein-Barr virus (EBV) DNA copy number in EBV-infected epithelial cell lines. 总被引:1,自引:0,他引:1
M Kanamori M Murakami T Takahashi N Kamada M Tajima K Okinaga Y Miyazawa T Kurata T Sairenji 《Microbes and infection / Institut Pasteur》2001,3(13):1085-1091
We found that spontaneous and 12-0-tetradecanoylphorbol-13-acetate-induced Epstein-Barr virus (EBV) reactivation occurred in short-term (ST)-cultured EBV-infected epithelial cell lines GT38 and GT39 after their establishment; however, it diminished in the long-term (LT)-cultured cells passaged for more than 2 years from ST-cultured cells. We hypothesized that the EBV reactivation may be related to the EBV DNA copy number in the cells. A higher level of EBV DNA content was detected in ST-cultured cells than in LT-cultured cells by Southern hybridization using an EBV DNA XhoI probe. Fluorescence in situ hybridization using EBV DNA BamHI W fragments showed that ST-cultured cells contained a higher EBV DNA copy number than that of LT-cultured cells. EBV DNA-negative cells were detected in small proportions in LT-cultured cells, but were undetected in ST-cultured cells. These results demonstrate that EBV genomes are not maintained stably in the cell lines, and some of them are lost in continuous passages of the cells. We discuss the mechanisms of reduction of EBV reactivation and EBV DNA in the cell lines. 相似文献