Early Epstein-Barr Virus Research Articles

Susceptibility to primary Epstein-Barr virus infection is associated with interleukin-10 gene promoter polymorphism.

In total, 116 children were investigated to determine whether the interleukin (IL)-10 polymorphism influences the age at primary Epstein-Barr virus (EBV) infection. The promoter of IL-10 is polymorphic, with 3 known single base substitutions (G/A at -1082, C/T at -819, and C/A at -592), which form 3 haplotypes: GCC, ACC, and ATA. This study found that carriage of the ATA haplotype protects against early EBV infection. The presence of the ATA haplotype was associated with EBV seronegativity (odds ratio, 2.6; 95% confidence interval, 1.04-6.7; P=.04), when controlled by age. To examine the effect of haplotypes on IL-10 production, IL-10 plasma levels were measured in 50 newborns and 400 adults and were correlated with the IL-10 haplotype. The IL-10 levels were significantly higher in the ATA carriers than in the noncarriers. These data suggest that the IL-10 ATA haplotype confers protection against primary EBV infection and that the effect is mediated by high IL-10 levels.

Epstein-Barr virus immediate-early protein BRLF1 interacts with CBP, promoting enhanced BRLF1 transactivation.

The Epstein-Barr virus (EBV) immediate-early protein BRLF1 is a transcriptional activator that mediates the switch from latent to lytic viral replication. Many transcriptional activators function, in part, due to an interaction with histone acetylases, such as CREB-binding protein (CBP). Here we demonstrate that BRLF1 interacts with the amino and carboxy termini of CBP and that multiple domains of the BRLF1 protein are necessary for this interaction. Furthermore, we show that the interaction between BRLF1 and CBP is important for BRLF1-induced activation of the early lytic EBV gene SM in Raji cells.

The PKC targeting protein RACK1 interacts with the Epstein-Barr virus activator protein BZLF1.

Phorbol esters reactivate Epstein-Barr virus (EBV) from latently infected cells via transcriptional activation of the viral immediate-early gene BZLF1. BZLF1 is a member of the extended AP-1 family of transcription factors that binds to specific BZLF1-binding motifs within early EBV promoters and to consensus AP-1 sites. Regulation of BZLF1's activity is achieved at the transcriptional level as well as through post-translational modifications. Recently, we reported that the transcriptional activity of BZLF1 is augmented by TPA [Baumann, M., Mischak, H., Dammeier, S., Kolch, W., Gires, O., Pich, D., Zeidler, R., Delecluse, H. J. & Hammerschmidt, W., (1998) J. Virol. 72, 8105-8114]. The increase of BZLF1's activity depends on a single serine residue (S186) that is phosphorylated by protein kinase C (PKC) in vitro and in vivo after stimulation with 12-O-tetradecanoylphorbol-13-acetate (TPA). Here, we identified RACK1 as a binding partner of BZLF1 in a yeast interaction trap assay. RACK stands for receptor of activated C-kinase and is involved in targeting activated PKCs and other signaling proteins. In vivo, RACK1 binds directly to the transactivation domain of BZLF1. Although a functional relationship between BZLF1 and PKC could be mediated by RACKs, RACK1 did not have a detectable effect on the phosphorylation status of BZLF1 in in vitro or in vivo phosphorylation assays. We suggest that RACK1 may act as a scaffolding protein on BZLF1 independently of activated PKCs.

Regulation of Epstein-Barr virus promoters in oral epithelial cells and lymphocytes.

Hairy leukoplakia (HL) is a proliferative lesion of the tongue that supports abundant Epstein-Barr virus (EBV) replication. Previous work showed high-level expression of the EBV BMRF2 gene in HL. To characterize the regulation of BMRF2 expression in HL, we mapped the 5' ends of the BMRF1 and BMRF2 transcripts and showed that BMRF2 is expressed from a novel internal promoter within the BMRF1 coding region. Mechanisms of BMRF2 regulation were compared in oral epithelial cells and B lymphocytes, as were those of BMRF1 and BDLF3, early and late EBV transcripts, respectively, that are also known to be expressed in HL. Basal activity of the putative BMRF2 promoter was 10-fold higher in HSC-3 epithelial cells than in B lymphocytes. The BMRF2 and the BDLF3 promoters were responsive to induction by phorbol ester, but unlike the BMRF1 promoter, they were not responsive to BZLF1 transactivation. By mutational analysis, the major activity of the BMRF2 promoter mapped to a 50-bp region, which includes a TATA-like element and a GC box. The BMRF2 promoter may be regulated differentially from the BMRF1 promoter and more closely resembles that of BDLF3. This novel BMRF2 promoter likely belongs to a class of viral promoters that is more responsive to mechanisms known to induce epithelial cell differentiation, consistent with its high level of expression in HL.

Natural history of primary Epstein-Barr virus infection in children of mothers infected with human immunodeficiency virus type 1.

The natural history of Epstein-Barr virus (EBV) infection in 556 infants born to 517 human immunodeficiency virus (HIV) type 1-infected mothers was studied in a prospective, multicenter, cohort study. HIV-1-infected children had a cumulative EBV infection rate similar to HIV-1-uninfected children at age 3 years (77.8% vs. 84. 9%) but had more frequent oropharyngeal EBV shedding (50.4% vs. 28. 2%; P<.001). The probability of shedding decreased with longer time from EBV seroconversion and was similar to that of HIV-1-uninfected children 3 years after seroconversion. HIV-1-infected children identified as rapid progressors shed EBV more frequently than nonrapid progressors (69.4% vs.41.0%; P=.01). HIV-1-infected children with EBV infection had higher mean CD8 cell counts. EBV infection did not have an independent effect on mean CD4 cell counts, percent CD4, IgG levels, HIV-1 RNA levels, lymphadenopathy, hepatomegaly, or splenomegaly. Early EBV infection is common in children born to HIV-1-infected mothers. Children with rapidly progressive HIV-1 disease have more frequent EBV shedding.

The Epstein-Barr virus (EBV) SM protein enhances pre-mRNA processing of the EBV DNA polymerase transcript.

The Epstein-Barr virus (EBV) DNA polymerase (pol) mRNA, which contains a noncanonical polyadenylation signal, UAUAAA, is cleaved and polyadenylated inefficiently (S. C. S. Key and J. S. Pagano, Virology 234:147-159, 1997). We postulated that the EBV early proteins SM and M, which appear to act posttranscriptionally and are homologs of herpes simplex virus (HSV) ICP27, might compensate for the inefficient processing of pol pre-mRNA. Here we show that the SM and M proteins interact with each other in vitro. In addition, glutathione S-transferase-SM/M fusion proteins precipitate the heterogeneous ribonucleoprotein (hnRNP) C1 splicing protein. Further, the SM protein is coimmunoprecipitated from SM-expressing cell extracts with an antibody to the hnRNP A1/A2 proteins, which are splicing and nuclear shuttling proteins. Finally, the amount of processed EBV DNA polymerase mRNA was increased three- to fourfold in a HeLa cell line expressing SM; this increase was not due to enhanced transcription. Thus, inefficient processing of EBV pol RNA by cellular cleavage and polyadenylation factors appears to be compensated for and may be regulated by the early EBV protein, SM, perhaps via RNA 3'-end formation.

Rescue of the Epstein–Barr Virus BZLF1 Mutant, Z(S186A), Early Gene Activation Defect by the BRLF1 Gene Product

Expression of the Epstein–Barr virus (EBV) immediate-early protein, BZLF1 (Z), is sufficient to disrupt viral latency. Z transcriptionally activates the EBV early genes by binding to upstream Z-responsive elements (ZREs). Recently, a serine-to-alanine mutation of Z residue 186 (within the basic DNA binding domain) was shown to inhibit the ability of Z to induce lytic infection in latently infected cells, although the Z(S186A) mutant could still bind several known ZREs and activated an early EBV promoter (BMRF1) in transient reporter gene assays (Francis, A. L., Gradoville, L., and Miller, G. (1997).J. Virol.71, 3054–3061). We now show that a specific deficiency in the ability to bind to ZRE elements in the immediate-early BRLF1 promoter may account for the inability of Z(S186A) to activate BRLF1 expression. Furthermore, we demonstrate that the ability of Z(S186A) to induce early BMRF1 and BHRF1 gene expression is rescued by cotransfection with a BRLF1 expression vector. However, the Z(S186A)/BRLF1 (R) combination cannot induce full lytic replication, suggesting that Z(S186A) may also be deficient in a replication-specific function. These results suggest that in the context of the intact viral genome, both Z and R expression are required for activation of early gene transcription in latently infected cells.

Activation of the Epstein-Barr virus transcription factor BZLF1 by 12-O-tetradecanoylphorbol-13-acetate-induced phosphorylation.

BZLF1 is a member of the extended AP-1 family of transcription factors which binds to specific BZLF1 sequence motifs within early Epstein-Barr virus (EBV) promoters and to closely related AP-1 motifs. BZLF1's activity is regulated at the transcriptional level as well as through protein interactions and posttranslational modifications. Phorbol esters or immunoglobulin cross-linking both reactivate EBV from latently infected B cells via transactivation of BZLF1. We report here that the phorbol ester 12-O-tetradecanoylphorbol-13-acetate (TPA) is capable of inducing BZLF1's activity even further. The induction occurs at the posttranscriptional level and depends on a single serine residue located in the DNA binding domain of BZLF1. This serine residue (S186) is phosphorylated by protein kinase C in vitro and in vivo after stimulation with TPA. Phosphorylation of S186 per se interferes with the DNA binding affinity of BZLF1 in vitro but is mandatory for TPA-induced increase in DNA binding of BZLF1, as shown in gel retardation assays and reconstruction experiments with cellular extracts. In transcriptional reporter assays, S186 is essential for the activation of BZLF1 by TPA. Presumably, a yet-to-be-identified cellular factor restores the DNA binding affinity and enhances the transcriptional activity of S186-phosphorylated BZLF1, which is required to induce the lytic phase of EBV's life cycle.

The Epstein-Barr virus nuclear protein SM is both a post-transcriptional inhibitor and activator of gene expression.

The Epstein-Barr virus (EBV) nuclear protein BS-MLF1 (SM) is expressed early after entry of EBV into the lytic cycle. SM transactivates reporter gene constructs driven by a wide variety of promoters, but the mechanism of SM action is poorly understood. In this study, we demonstrate that the SM protein inhibits expression of intron-containing genes and activates expression of intron-less genes. We demonstrate that SM has the predicted inhibitory effect on expression of a spliced EBV gene but activates an unspliced early EBV gene. SM inhibited gene expression at the post-transcriptional level by preventing the accumulation of nuclear and cytoplasmic RNA transcripts. Conversely, SM led to increased accumulation of nuclear mRNA from intron-less genes without affecting the rate of transcription, indicating that SM enhances nuclear RNA stability. The ratio of cytoplasmic to nuclear polyadenylated mRNA was increased in the presence of SM, suggesting that SM also enhances nucleo-cytoplasmic mRNA transport. The degree of transactivation by SM was dependent on the sequence of the 3'-untranslated region of the target mRNA. Finally, we demonstrate that the amino-terminal portion of SM fused to glutathione-S-transferase binds radioactively labeled RNA in vitro, indicating that SM is a single-stranded RNA binding protein. Importantly, the latent and immediate-early genes of EBV contain introns whereas many early and late genes do not. Thus, SM may down-regulate synthesis of host cell proteins and latent EBV proteins while simultaneously enhancing expression of specific lytic EBV genes by binding to mRNA and modulating its stability and transport.

Breast milk is not a significant source for early Epstein-Barr virus or human herpesvirus 6 infection in infants: a seroepidemiologic study in 2 endemic areas of human T-cell lymphotropic virus type I in Japan.

In order to evaluate the possibility of Epstein-Barr virus (EBV) and human herpesvirus 6 (HHV-6) transmission via breast milk, a total of 331 serum specimens collected from bottle-fed and breast-fed children and their mothers, in 2 endemic areas of human T-cell lymphotropic virus type I (HTLV-I) in Japan, were assayed for antibodies to EBV and HHV-6. The seroprevalences of EBV and HHV-6 were over 95% both in the mothers of bottle-fed children and in those of breast-fed children. The seroprevalence of EBV at 12-23 months of age was 54.5% (36/66) and 55.8% (24/43) in breast-fed children and bottle-fed children, respectively. The seroprevalence of HHV-6 at 12-23 months of age was 90.9% (60/66) and 93.0% (40/43) in breast-fed children and bottle-fed children, respectively. No difference was observed between the seroprevalences of EBV and HHV-6 in breast-fed and bottle-fed children at 12-23 months of age. Our seroepidemiologic data indicate that breast milk is not a significant source of early EBV or HHV-6 infection in infancy.

Endoscopic and pathologic features of Epstein-Barr virus–associated gastric carcinoma

Background: Although the presence of Epstein-Barr virus has been documented in approximately 7% of patients with gastric carcinoma, the clinical features of Epstein-Barr virus–associated carcinoma have not been well documented. We studied the histologic and endoscopic characteristics of Epstein-Barr virus–associated gastric carcinoma. Methods: We tested 124 gastric carcinomas from 117 patients using in situ hybridization for Epstein-Barr virus encoded small RNA1. The histologic and endoscopic findings in the Epstein-Barr virus–associated groups and the negative control groups were analyzed and compared. Results: Twelve tumors (9.7%) were identified as Epstein-Barr virus associated. These lesions were located mainly in the upper part of the stomach (p < .05) and had a diffuse-type histology (p < .05) compared with those in the control group. Six of seven (85.7%) early Epstein-Barr virus–associated lesions were type 0 IIc (superficial depressed) or a combined type, and 42.9% were accompanied by submucosal nodules of carcinoma with lymphoid stroma. Four of five (80%) advanced Epstein-Barr virus–associated tumors were type 3 (ulcerated without definite limits), thought to be the advanced shape of superficial depressed lesions. Conclusions: Epstein-Barr virus–associated gastric carcinomas often appear as superficial depressed or ulcerated lesions in the upper part of the stomach and have a diffuse-type histology with lymphoid infiltration. (Gastrointest Endosc 1997;45:236-42.)

Rheumatic disease patients, prone to Sjögren's syndrome and/or lymphoma, mount an antibody response to BHRF1, the Epstein-Barr viral homologue of BCL-2.

The IgG response to Epstein-Barr virus (EBV) early antigens [BHRF1 (p 17.1), the viral homologue of bcl-2, and BMRF1 (p50.10), a DNA binding protein] was measured in patients with rheumatic disease to see whether there was any association with lymphoma. Patients with rheumatoid arthritis (RA), systemic lupus erythematosus (SLE), Sjögren's syndrome (SS), rheumatic disease patients with lymphoma, patients with lymphoma who did not have a rheumatic disease and normal individuals were tested for the presence of anti-EA peptide antibodies by ELISA. Whereas antibodies to early EBV peptides were detected only in one normal individual, patients with rheumatic diseases, especially those with either SS and/or lymphoma, had a much higher frequency of antibody detection. Antibodies to BMRF1 p50.10 were found in 7-50% of patients, and to BHRF1 p17.1 in 4-27%, depending on the group studied. Patients with lymphoma lacking a rheumatic disease had a 2-fold lower frequency of anti-BHRF1 antibodies, compared to the lymphoma plus rheumatic disease group. The increased immune response to the EBV EA proteins in the rheumatic diseases probably reflects the presence of reactivated virus, and the BHRF1 protein (the viral homologue to bcl-2) could, via inhibiting apoptosis, contribute to the lymphoproliferative nature of these diseases.

Functional and physical interactions between the Epstein-Barr virus (EBV) proteins BZLF1 and BMRF1: Effects on EBV transcription and lytic replication.

The Epstein-Barr virus (EBV) proteins BZLF1 and BMRF1 are both essential for lytic EBV replication. BZLF1 is a transcriptional activator which binds directly to the lytic origin of replication (oriLyt) and plays a critical role in the disruption of viral latency. The BMRF1 protein is required for viral polymerase processivity. Here we demonstrate that the BMRF1 gene product functions as a transcriptional activator and has direct (as well as indirect) interactions with the BZLF1 gene product. The BMRF1 gene product activates an essential oriLyt promoter, BHLF1, but does not activate two other early EBV promoters (BMRF1 and BHRF1). Direct interaction between the BMRF1 and BZLF1 gene products requires the first 45 amino acids of BMRF1 and the bZip domain of BZLF1. The effect of the BZLF1-BMRF1 interaction on early EBV transcription is complex and is promoter specific. The oriLyt BHLF1 promoter is activated by either the BZLF1 or BMRF1 gene product alone and is further activated by the combination of the BZLF1 and BMRF1 gene products. Enhanced activation of BHLF1 transcription by the BMRF1-BZLF1 combination does not require direct interaction between these proteins. In contrast, BZLF1-induced activation of the BMRF1 promoter is inhibited in the presence of the BMRF1 gene product. A point mutation in the BZLF1 protein (amino acid 200), which prevents in vitro interaction with the BMRF1 protein but which does not reduce BZLF1 transactivator function, allows the BZLF1 protein to activate the BMRF1 promoter equally well in the presence or absence of the BMRF1 gene product. Therefore, direct interaction between the BZLF1 and BMRF1 proteins may inhibit BZLF1-induced transcription of the BMRF1 promoter. BZLF1 mutated at amino acid 200 is as efficient as wild-type BZLF1 in promoting replication of an oriLyt plasmid. However, this mutation reduces the ability of BZLF1 to induce lytic replication of the endogenous viral genome in D98/HE-R-1 cells. Our results indicate that functional and physical interactions between the BMRF1 and BZLF1 proteins may modulate the efficiency of lytic EBV infection. The BMRF1 gene product clearly has a transcriptional, as well as replicative, role during lytic EBV infection.

Nasal T‐cell lymphoma causally associated with Epstein‐Barr virus: Clinicopathologic, phenotypic, and genotypic studies

BACKGROUND The authors have previously demonstrated nasal T-cell lymphoma (NTL) associated with Epstein–Barr virus (EBV). The detailed clinical, phenotypic, and genotypic features and the role of EBV in lymphomagenesis remain to be clarified. METHODS The study group consisted of 18 patients with NTL. The phenotype was determined by immunoperoxidase staining with various monoclonal antibodies. Genotypic study was done using Southern blot hybridization. The presence of EBV-encoded small nuclear early region (EBER) RNA and EBV DNA were determined by in situ hybridization. The expression of EBV-encoded nuclear antigen (EBNA) and latent membrane protein (LMP1) were identified by immunohistologic methods. Clonotypic analysis of EBV genomes was performed by Southern blot hybridization with EBV termini fragment probe. RESULTS The clinical features of NTL were characterized as prolonged fever (16 patients), widespread dissemination into distant sites (13 patients), and poor prognosis with a median survival of only 6 months. EBER transcripts were identified in 16 of 18 patients. Monoclonal EBV genomes EBNA1 and LMP1 were also detected in all EBER-positive cases tested. All 18 patients expressed pan-T antigens such as MT1, CD45RO, and/or CD2. The rearrangements of T-cell receptor (TCR)-β, -γ, and/or -δ genes were shown in all 11 patients tested. The natural killer (NK) cell phenotype CD56 was expressed in all EBV-positive cases tested, and was not detected in EBV-negative cases. Seven EBV-positive cases expressed a TCR-δ chain with rearranged TCR-γ or -δ genes whereas both EBV-negative cases corresponded to αβT-cell lymphoma, which expressed a TCR-β chain with a rearranged TCR-β gene. CONCLUSIONS These data suggest that EBV-positive NTL may be derived from the lineage of NK-like T-cells or γδT-cells, and that EBV may play a role in lymphomagenesis. Therefore, we propose that NTL which has peculiar clinical and histologic features, could be classified as a new lymphoma entity. Cancer 1996;77:2137-49.

Nasal T-cell lymphoma causally associated with Epstein-Barr virus: clinicopathologic, phenotypic, and genotypic studies.

The authors have previously demonstrated nasal T-cell lymphoma (NTL) associated with Epstein-Barr virus (EBV). The detailed clinical, phenotypic, and genotypic features and the role of EBV in lymphomagenesis remain to be clarified. The study group consisted of 18 patients with NTL. The phenotype was determined by immunoperoxidase staining with various monoclonal antibodies. Genotypic study was done using Southern blot hybridization. The presence of EBV-encoded small nuclear early region (EBER) RNA and EBV DNA were determined by in situ hybridization. The expression of EBV-encoded nuclear antigen (EBNA) and latent membrane protein (LMP1) were identified by immunohistologic methods. Clonotypic analysis of EBV genomes was performed by Southern blot hybridization with EBV termini fragment probe. The clinical features of NTL were characterized as prolonged fever (16 patients), widespread dissemination into distant sites (13 patients), and poor prognosis with a median survival of only 6 months. EBER transcripts were identified in 16 of 18 patients. Monoclonal EBV genomes EBNA1 and LMP1 were also detected in all EBER-positive cases tested. All 18 patients expressed pan-T antigens such as MT1, CD45RO, and/or CD2. The rearrangements of T-cell receptor (TCR)-beta, -gamma, and/or -delta genes were shown in all 11 patients tested. The natural killer (NK) cell phenotype CD56 was expressed in all EBV-positive cases tested, and was not detected in EBV-negative cases. Seven EBV-positive cases expressed a TCR-delta chain with rearranged TCR-gamma or -delta genes whereas both EBV-negative cases corresponded to alpha beta T-cell lymphoma, which expressed a TCR-beta chain with a rearranged TCR-beta gene. These data suggest that EBV-positive NTL may be derived from the lineage of NK-like T-cells or gamma delta T-cells, and that EBV may play a role in lymphomagenesis. Therefore, we propose that NTL which has peculiar clinical and histologic features could be classified as a new lymphoma entity.

Activation of the Epstein-Barr virus DNA polymerase promoter by the BRLF1 immediate-early protein is mediated through USF and E2F

The Epstein-Barr virus (EBV) DNA polymerase (pol) is essential for the replication of viral genomes during productive EBV infection. We have previously reported that the EBV DNA pol promoter, which is TATA-less and constitutively inactive, is activated by a genomic clone expressing both immediate-early viral transactivators, BZLF1Z and BRLF1 (R), in EBV-infected lymphoid cells. Here we demonstrate that R alone is sufficient to activate the pol promoter in EBV-negative B cells. Unlike other early promoters to which the R protein binds directly, its effect on the pol promoter does not appear to involve a direct DNA-binding mechanism. Instead, we found that two cellular transcription factors, an upstream stimulatory factor USF, and a member of the E2F family of proteins, bind directly to the pol promoter at positions -795 to -786 and -186 to -170, respectively, regions previously identified as important for activation of the pol promoter. These two sites contribute to or are essential for transactivation of the pol promoter by R in EBV-noninfected B cells. These data suggest that the R immediate-early protein may activate a key early EBV promoter (pol) through both USF and E2F.

Antibody responses to Epstein-Barr virus, human herpesvirus 6 and human herpesvirus 7 in patients with chronic fatigue syndrome.

To test for an association between chronic fatigue syndrome (CFS) and infections with Epstein-Barr virus (EBV), human herpesvirus 6 (HHV-6) and human herpesvirus 7 (HHV-7), antibodies to these viruses were tested in the serum from three groups of individuals: (1) 10 CFS patients with chronic fatigue beginning with a clinical pattern of acute infectious mononucleosis [IM; true chronic IM (CIM)]; (2) 10 CFS patients whose illness did not start with acute IM (non-CIM), and (3) healthy controls. High EBV antibody titers were demonstrated in most patients. Antibodies to ZEBRA, a product of the immediate early EBV gene BZLF1, were detected in the serum of CFS patients at a higher frequency than in healthy controls. Antibody titers to HHV-6 and HHV-7 were also higher in the patients with CFS than in the controls. These results are consistent with the view that CFS patients may have reactivations of EBV, HHV-6 and HHV-7.

Epstein-Barr Virus Shedding in Breast Milk

One hundred healthy women already donating to the Children's Hospital Breast Milk bank consented to provide a sample of breast milk for this study. Using a DNA-DNA hybridization dot-blot assay Epstein-Barr virus (EBV) genome (Bam HIW region) was detected in cells shed into breast milk of 46 out of 100 women studied and in 60 out of 132 (46%) of samples donated overall. The prevalence of EBV shedding increased postnatally to a peak of 74% (26/35 positive samples) between 3 and 12 weeks postdelivery. Women delivering prematurely had an initially lower prevalence of shedding with only six out of 30 (20%) positive samples in the first week after delivery, compared to 16 out of 35 (46%) for women delivering at term. Of the 18 women donating more than one sample, 13 showed consistently positive (n = 8) or negative (n = 5) results, and the remaining five had intermittent shedding detected. Only seven out of 42 (17%) breast milk samples studied were EBV-IgG antibody positive, and none showed IgM or IgA-EBV antibodies. Further studies and prospective followup of infants are needed to confirm that breast milk is a significant source for early EBV infection of infants, as indicated by serologic studies.

The spliced BZLF1 gene of Epstein-Barr virus (EBV) transactivates an early EBV promoter and induces the virus productive cycle

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.

Expression of normal and translocated c-myc alleles in Burkitt's lymphoma cells: evidence for different regulation.

In Burkitt's lymphoma (BL) cells the normal c-myc allele is usually silent or expressed at very low levels. Here we demonstrate that the normal c-myc allele can be induced in BL cells by 12-O-tetradecanoylphorbol-13-acetate (TPA). TPA did activate the normal c-myc alleles in Raji(P207), BL36, P3HR1, Jijoye and LY91 cells, but not in Raji(DE88), BL41, BL67, LY47 and KK124 cells. C-myc RNA derived from the normal allele appeared 6 h after treatment with TPA and showed the characteristic preferential usage of the second promoter. This induction could not be inhibited by cycloheximide. Despite the differences in c-myc induction in Raji(P207) and Raji(DE88) cells, c-fos and the early Epstein-Barr virus gene DR were induced to a similar extent and with similar kinetics by TPA. Nuclear run-on experiments suggest that the normal c-myc allele in Raji cells is activated at least in part by releasing a block to RNA elongation at the end of c-myc exon 1. Expression of the translocated c-myc alleles was also affected by TPA; however, only if cycloheximide was simultaneously present. TPA plus cycloheximide induced a rapid decrease of c-myc RNA derived from the translocated allele within 6 h, whereas cycloheximide alone led to abolition of c-myc RNA after 16-24 h. This rapid decline of c-myc RNA was observed in Raji and BL41 cells, but not in three cell lines with variant t(2;8) and t(8;22) translocations.