The U28 ORF of human herpesvirus-7 does not encode a functional ribonucleotide reductase R1 subunit.
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Herpesvirus ribonucleotide reductases, essential for the de novo synthesis of viral DNA, are composed of two non-identical subunits, termed R1 and R2. The U28 ORF from human herpesvirus-7 has been classified, by sequence comparisons, as a homologue of the R1 subunit from ribonucleotide reductase but no R2 ORF is present. Detailed analysis of the U28 amino acid sequence indicated that a number of essential R1 catalytic residues are absent. Cloning and expression of the U28 protein in E. coli and its subsequent characterization in subunit interaction and enzyme activity assays confirmed that it is not a functional equivalent of a herpesvirus R1. In the absence of the R2 gene, we propose that the R1 ORF has evolved a distinct, as yet unidentified, function not only in human herpesvirus-7 but also in other human betaherpes-viruses. (+info)
Modulatory effects of human herpes virus-7 on cytokine synthesis and cell proliferation in human peripheral blood mononuclear cell cultures.
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Human herpes virus-7 (HHV-7) infects cells of the immune system and thus may modulate their function. To investigate the potential immunomodulatory effects of this virus, we performed an in vitro study in which we investigated effects of HHV-7 on the synthesis of several key immunomodulatory cytokines, i.e. tumor necrosis factor alpha (TNF-alpha), interleukin-2 (IL-2), interferon-gamma (IFN-gamma), IL-4, IL-6, and transforming growth factor beta (TGF-beta). This was examined after in vitro infection of human peripheral blood mononuclear cells (PBMC) with HHV-7. We found elevated levels of TNF-alpha, TGF-beta, and IFN-gamma in the supernatants of HHV-7-infected cells. By reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, using cytokine-specific primers, we found that levels of TNF-alpha, TGF-beta, and IFN-gamma mRNA were increased in the infected cells. The HHV-7 infection also significantly (P < 0.05) decreased the production of IL-2 from activated, IL-2-producing PBMC. Furthermore, mitogen- and cytokine-induced cellular proliferative responses of human PBMC were found to be significantly (P < 0.05) down-regulated by this virus. On the other hand, HHV-7 did not affect IL-4 and IL-6 synthesis. Overall, our results indicate that HHV-7 infection causes significant immunomodulatory effects with regard to cytokine synthesis in these cells as well as inhibiting lymphocyte proliferation by various stimuli. (+info)
CXC-chemokine receptor 4 is not a coreceptor for human herpesvirus 7 entry into CD4(+) T cells.
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Human herpesvirus 7 (HHV-7) is a T-lymphotropic virus which utilizes the CD4 receptor as its main receptor to enter the target cells. Hence, HHV-7 can interfere with human immunodeficiency virus type 1 (HIV-1) infection in CD4(+) T cells. It was recently suggested that the CXC chemokine receptor 4 (CXCR4), which was found to be a crucial coreceptor for T-tropic HIV-1 strains, may also play a role in the HHV-7 infection process. However, the results presented here demonstrate that CXCR4 is not involved in HHV-7 infection. The natural ligand of CXCR4, SDF-1alpha, was not able to inhibit HHV-7 infection in SupT1 cells or in CD8(+) T-cell-depleted peripheral blood mononuclear cells. Also, AMD3100, a specific CXCR4 antagonist with potent antiviral activity against T-tropic HIV strains (50% inhibitory concentration inverted question markIC(50), 1 to 10 ng/ml), completely failed to inhibit HHV-7 infection (IC(50), >250 microg/ml). Thus, two different agents known to specifically interact with CXCR4 were not able to inhibit HHV-7 infection. Other T-lymphoid cell lines, expressing both CD4 and CXCR4 (e.g., HUT-78 and MT-4) could not be infected by HHV-7. In addition, the CD4-transfected cell lines HOS. CD4 and U87.CD4 and the CD4/CXCR4 double-transfected cell lines HOS. CD4.CXCR4 and U87.CD4.CXCR4 were not infectable with HHV-7. Also, we found no down-regulation of surface-bound or intracellular CXCR4 in HHV-7-infected CD4(+) T cells. As compared to uninfected SupT1 cells, stromal cell-derived factor 1alpha (SDF-1alpha)/CXCR4-mediated intracellular calcium flux was unchanged in SupT1 cells that were acutely or persistently infected with HHV-7. All these data argue against CXCR4 as a receptor involved in the HHV-7 infection process. (+info)
Detection of human herpesviruses 6 and 7 genomic sequences in brain tumours.
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BACKGROUND: Human herpesviruses 6 and 7 (HHV-6, HHV-7) are ubiquitous, with primary infection occurring early in life followed by persistence, which may involve neural tissue. While HHV-6 and HHV-7 are predominantly T lymphotropic, the extent of tissue tropism in persistent infection is not known. AIM: To investigate neuropersistence and the role of HHV-6 and HHV-7 in brain tumorigenesis. METHODS: Nested polymerase chain reaction was used to detect HHV-6 and HHV-7 genomic sequences in preparations of total DNA extracted from 98 formalin fixed, paraffin embedded primary brain tumours. HHV-6 detected was further characterized into variants A and B by restriction fragment length analysis. RESULTS: HHV-6 was detected in 8.2% of cases and HHV-7 in 14.3% (14/98). None of the positive samples contained both viruses. Among the eight HHV-6 positive tumours, three harboured variant A and five variant B. Four of the five ependymomas studied contained viral DNA. Otherwise, both HHV-6 and HHV-7 were present at similar low frequencies in most of the tumour types investigated. CONCLUSIONS: The findings do not support an aetiological role of HHV-6 and HHV-7 in primary brain tumour, but they suggest that HHV-6 and HHV-7 are neurotropic in vivo and that the central nervous system seems to be one of the reservoirs for persistent infection. (+info)
Suppressive effects of human herpesvirus-6 on thrombopoietin-inducible megakaryocytic colony formation in vitro.
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Two clinical observations, the association of human herpesvirus-6 (HHV-6) with delayed engraftment after stem cell transplantation and thrombocytopenia concomitant with exanthema subitum, prompted us to evaluate the suppressive effects of HHV-6 on thrombopoiesis in vitro. Different culture conditions for thrombopoietin (TPO)-inducible colonies in semi-solid matrices were examined. Using cord blood mononuclear cells as the source of haematopoietic progenitors, two types of colonies, megakaryocyte colony-forming units (CFU-Meg) and non-CFU-Meg colonies, were established. The former colonies were identified by the presence of cells with translucent cytoplasm and highly refractile cell membrane, most of which were positive for the CD41 antigen. Although the plating efficiency of both types was much higher under serum-containing conditions than under serum-free conditions, the proportion of CFU-Meg to non-CFU-Meg colonies was consistently higher under serum-free conditions. The plating efficiency of CFU-Meg colonies was doubled by adding stem cell factor to the serum-free matrix. The effects of two variants of HHV-6 (HHV-6A and 6B) and human herpesvirus-7 (HHV-7) on TPO-inducible colonies were then compared. HHV-6B inhibited both CFU-Meg and non-CFU-Meg colony formation under serum-free and serum-containing conditions. HHV-6A had similar inhibitory effects. In contrast, HHV-7 had no effect on TPO-inducible colony formation. Heat-inactivation and ultra-filtration of the virus sample completely abolished the suppressive effect. After infection of CD34(+) cells with HHV-6, the viral genome was consistently detected by in situ hybridization. These data suggest that the direct effect of HHV-6 on haematopoietic progenitors is one of the major causes of the suppression of thrombopoiesis. (+info)
Identification and analysis of a novel heparin-binding glycoprotein encoded by human herpesvirus 7.
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Human herpesvirus 6 (HHV-6) and HHV-7 are closely related betaherpesviruses that encode a number of genes with no known counterparts in other herpesviruses. The product of one such gene is the HHV-6 glycoprotein gp82-105, which is a major virion component and a target for neutralizing antibodies. A 1.7-kb cDNA clone from HHV-7 was identified which contains a large open reading frame capable of encoding a predicted primary translational product of 468 amino acids (54 kDa) with 13 cysteine residues and 9 potential N-linked glycosylation sites. This putative protein, which we have termed gp65, was homologous to HHV-6 gp105 (30% identity) and contained a single potential membrane-spanning domain located near its amino terminus. Comparison of the cDNA sequence with that of the viral genome revealed that the gene encoding gp65 contains eight exons, spanning almost 6 kb of the viral genome at the right (3') end of the HHV-7 genome. Northern (RNA) blot analysis with poly(A)(+) RNA from HHV-7-infected cells revealed that the cDNA insert hybridized to a single major RNA species of 1.7 kb. Antiserum raised against a purified, recombinant form of gp65 recognized a protein of roughly 65 kDa in sucrose density gradient-purified HHV-7 preparations; treatment with PNGase F reduced this glycoprotein to a putative precursor of approximately 50 kDa. Gp65-specific antiserum also neutralized the infectivity of HHV-7, while matched preimmune serum did not do so. Finally, analysis of the biochemical properties of recombinant gp65 revealed a specific interaction with heparin and heparan sulfate proteoglycans and not with closely related molecules such as N-acetylheparin and de-N-sulfated heparin. At least two domains of the protein were found to contribute to heparin binding. Taken together, these findings suggest that HHV-7 gp65 may contribute to viral attachment to cell surface proteoglycans. (+info)
Sensitive method for detection of human herpesviruses 6 and 7 in saliva collected in field studies.
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To facilitate studies of the epidemiology and natural history of human herpesviruses 6 and 7 in infants, a practical method for collecting and quantifying the DNA of these viruses was developed. Saliva was collected using small strips of filter paper, and virus was detected using a real-time quantitative fluorescent-probe PCR assay. The sensitivity and specificity of this method even after prolonged drying of the specimens compared favorably to those of our traditional method of collecting and assaying saliva. (+info)
The prevalence of human herpesvirus-7 in renal transplant recipients is unaffected by oral or intravenous ganciclovir.
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The purpose of this study was to compare the prevalence of human herpesvirus (HHV)-7 and cytomegalovirus (CMV) viremia and the effects of oral and intravenous (iv) ganciclovir in renal transplant recipients at risk for CMV. Stored lysates from peripheral blood leukocytes from 92 patients, who had been previously analyzed for CMV viremia by polymerase chain reaction (PCR) for 12 weeks after transplantation, were analyzed for HHV-7 viremia. Baseline and peak prevalences of HHV-7 viremia were 22% and 54%, respectively (P<. 0001). Eighty-two (89%) of 92 patients had at least 1 positive PCR for HHV-7. Oral ganciclovir and treatment with iv ganciclovir had no effect on the prevalence of HHV-7 viremia. In contrast, CMV was almost completely suppressed in patients who received oral ganciclovir, and when present, CMV responded to iv therapy. These results indicate that HHV-7 is resistant to ganciclovir at levels that were effective for prevention and treatment of CMV. (+info)