Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.7.7 (DNA polymerase)
17,007 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The gene (pol) encoding the Epstein-Barr virus (EBV) DNA polymerase is a member of the "early" class of viral genes which are expressed shortly after activation of latent virus infection. First, mRNA from the EBV-producing cell line, B95-8, treated with 12-O-tetradecanoylphorbol-13-acetate and sodium butyrate to induce lytic replication and expression of this gene was analyzed. Northern (RNA) analysis revealed a message of 3.7 kb found only in induced cells. 5' mapping of pol mRNA by S1 nuclease and primer extension analyses indicates that transcription initiates at tightly clustered sites within a G + C-rich region 126 bp upstream of the open reading frame. The same initiation region was identified in two other EBV-infected cell lines, P3HR1 and Raji, after induction. Second, a 1.29-kb genomic fragment containing this region, when cloned upstream of the chloramphenicol acetyltransferase reporter gene, demonstrated promoter activity in lymphoid cells cotransfected with pEBV-RZ, a genomic expression construct that includes genes for the EBV immediate-early transactivator proteins, BZLF-1 and BRLF-1. Within the upstream 1.29-kb sequence, two regions of 140 bp and 101 bp appear to be needed for promoter activity. These results demonstrate that unlike most EBV genes studied thus far, the pol gene contains multiple transcriptional start sites. The upstream regulatory region of the promoter for the pol gene does not contain canonical promoter elements such as TATA and CAAT boxes and, furthermore, is not constitutively active but requires transactivation by two or more viral proteins.
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PMID:Regulation of the Epstein-Barr virus DNA polymerase gene. 131 4

The outcome of virus-host interaction after peripheral inoculation of herpes simplex virus (HSV) depends on virus replication at the portal of entry, the ability of the virus to invade nerve endings and capillary endothelium cells and, the rate of virus replication in neurons and nonneural cells of the nervous system. Functions involved are the activity of viral thymidine kinase, DNA polymerase, immediate early transactivator proteins, the transcription initiation protein, the envelope protein(s) governing virus penetration, syncytium formation and natural killing of infected cells as well as some other regulatory DNA sequences.
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PMID:DNA regions and genes determining the virulence of herpes simplex virus. 135 74

The Epstein-Barr virus early antigen diffuse component (EA-D) is essential for Epstein-Barr virus DNA polymerase activity, and its activity is suppressed during latent infection. We investigated the regulation of the promoter (BMRF1) for this early gene by studying its responsiveness in vitro to two immediate-early viral transactivators, BZLF1 (Z) and BRLF1 (R), focusing on the differences in response in lymphoid cells and epithelial cells. In lymphoid cells, Z or R alone produced only small increases in EA-D promoter activity, whereas both transactivators together produced a large stimulatory effect. In epithelial cells, the Z transactivator alone produced maximal stimulation of the EA-D promoter; the effect of R and Z together was no greater than that of Z alone. Deletional analysis and site-directed mutagenesis of the EA-D promoter demonstrated that in epithelial cells the potential AP-1 binding site plays an essential role in Z responsiveness, although sequences further upstream are also important. In lymphoid cells, only the upstream sequences are required for transactivation by the Z/R combination, and the AP-1 site is dispensable. These data suggest that EA-D (BMRF1) promoter regulation by Z and R is cell type specific and appears to involve different mechanisms in each cell type.
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PMID:The Epstein-Barr virus (EBV) BMRF1 promoter for early antigen (EA-D) is regulated by the EBV transactivators, BRLF1 and BZLF1, in a cell-specific manner. 216 95

The bipartite POU domain of transcription factor Oct-1 stimulates adenovirus DNA replication through an interaction with the octamer sequence present in the auxiliary origin. Employing an immobilized in vitro DNA replication system, we show that the POU domain enhances the formation of a pre-initiation complex composed of the viral precursor terminal protein-DNA polymerase (pTP-pol) complex and the origin. To investigate the mechanism of stimulation we have explored protein-protein interactions between the POU domain and the pTP-pol complex. Such an interaction could be detected using a GST-POU fusion protein bound to glutathione-agarose beads. Binding was also observed with the POU homeodomain (POUHD), albeit weaker than with the intact POU domain, but not with the POU specific subdomain. Four point mutations localized in the POUHD were analyzed for pTP-pol binding. Two of these, E22A and E30A, bound pTP-pol equally as well as the wild-type, while the other two, Q24A and E29A, were able to bind 2- to 4-fold better. These mutations are localized in the same region where the HSV transactivator VP16 binds, but did not coincide with the VP16 contacts. A direct correlation between pTP-pol binding and stimulation of DNA replication in vitro was observed for all mutants, suggesting that stimulation by the POU domain is caused by an interaction with the viral pTP-pol complex.
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PMID:The Oct-1 POU domain stimulates adenovirus DNA replication by a direct interaction between the viral precursor terminal protein-DNA polymerase complex and the POU homeodomain. 795 6

Late gene expression follows and is dependent upon lytic replication of the viral genome. Although experimental evidence is lacking, lytic viral DNA replication is believed to remove modifications or binding factors from the genome which serve to repress late gene expression during latency or the early lytic cycle. We have developed a reporter assay to begin characterizing the mechanisms that regulate late gene expression in Epstein-Barr virus (EBV). In this model system, the activities of late promoter-reporter fusions are measured following transient transfection into tissue culture cells expressing EBV during different stages of the lytic cycle. This system faithfully recapitulates late expression patterns from the endogenous virus, implicating specific cis-active sequences in the control of late gene expression. In addition, these promoters respond only indirectly to the viral immediate-early transactivator, ZEBRA. This indirect response is mediated by other viral or virally induced activities downstream of ZEBRA in the lytic cascade. In this system, late gene expression is sensitive to inhibitors of the viral DNA polymerase such as phosphonoacetic acid, although the reporters lack a eukaryotic origin of replication and are not replicated under the assay conditions. Thus, replication of the transcriptional template is not a prerequisite for expression with late kinetics, a finding inconsistent with the current models which posit a cis-active relationship between lytic EBV DNA replication and late gene expression. Rather, analysis of this system has revealed a trans relationship between late gene expression and viral DNA replication and highlights the indirect and complex link between these two events.
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PMID:Late gene expression from the Epstein-Barr virus BcLF1 and BFRF3 promoters does not require DNA replication in cis. 934 31

Viral proteins E1 and E2 are essential for transient human papillomavirus (HPV) DNA replication. E1 is a multifunctional protein which can bind DNA and complex with E2, has ATPase and helicase activities, and interacts with DNA polymerase alpha-primase. E2 is a transactivator-repressor protein, playing an important role in replication and transcriptional regulation. A series of deletion mutants of HPV-11 E1 were constructed and tested in functional assays to define those domains of HPV-11 E1 which are important for binding to the origin DNA and E2. The domain of HPV-11 E1 involved in binding to the origin was located between aa 186 and 649, and that for binding to E2 was between aa 346 and 649. Since E1 binds to the origin more efficiently in the presence of E2, we also mapped the DNA binding domain of E1 in the presence of E2, and found that when binding was enhanced, the region of E1 involved in binding was similar to that observed with E1 alone. The same deletion mutation constructs of E1 were subcloned into an expression vector for use in transient replication assays to study the effect of the deletions on the replication of the origin DNA in vivo and the data suggest that the C-terminal domain contains important functions for replication.
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PMID:Active domains of human papillomavirus type 11 E1 protein for origin replication. 968 Jan 27

The Epstein-Barr virus (EBV) BMRF1 gene product is an essential component of the viral DNA polymerase and is absolutely required for lytic virus replication. In addition to its polymerase accessory protein function, we recently demonstrated that BMRF1 is a transactivator, inducing expression of the essential oriLyt promoter, BHLF1. However, the regions of BMRF1 required for transactivation of BHLF1 are unknown. Here we demonstrate that the carboxy-terminal portion of the BMRF1 protein (amino acids 378404), although not required for DNA binding or polymerase processivity function, is required for transactivator function as well as nuclear localization. Site-directed mutagenesis of this region allowed us to separate the transactivator and nuclear localization motifs of BMRF1. The two DNA-binding domains of BMRF1 are also required for efficient transactivation of the BHLF1 promoter.
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PMID:Identification of transactivator and nuclear localization domains in the Epstein-Barr virus DNA polymerase accessory protein, BMRF1. 993 86

The Epstein-Barr virus (EBV) open reading frame BGLF4 was identified as a potential Ser/Thr protein kinase gene through the recognition of amino acid sequence motifs characteristic of conserved regions within the catalytic domains of protein kinases. In order to investigate this potential kinase activity, BGLF4 was expressed in Escherichia coli and the purified protein was used to generate a specific antiserum. Recombinant vaccinia virus vTF7-3, which expresses the T7 RNA polymerase, was used to infect 293 and 293T cells after transient transfection with a plasmid containing BGLF4 under the control of the T7 promoter. Autophosphorylation of the BGLF4 protein was demonstrated using the specific antiserum in an immune complex kinase assay. In addition, EBNA-1-tagged BGLF4 and EBNA-1 monoclonal antibody 5C11 were used to demonstrate the specificity of the kinase activity and to locate BGLF4 in the cytoplasm of transfected cells. Manganese ions were found to be essential for autophosphorylation of BGLF4, and magnesium can stimulate the activity. BGLF4 can utilize GTP, in addition to ATP, as a phosphate donor in this assay. BGLF4 can phosphorylate histone and casein in vitro. Among the potential viral protein substrates we examined, the EBV early antigen (EA-D, BMRF1), a DNA polymerase accessory factor and an important transactivator during lytic infection, was found to be phosphorylated by BGLF4 in vitro. Amino acids 1 to 26 of BGLF4, but not the predicted conserved catalytic domain, were found to be essential for autophosphorylation of BGLF4.
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PMID:A protein kinase activity associated with Epstein-Barr virus BGLF4 phosphorylates the viral early antigen EA-D in vitro. 1070 24

Antisense RNA complementary to the Epstein-Barr virus (EBV) Zta gene, an immediate-early gene encoding a transactivator, was applied to inhibit EBV protein synthesis during its lytic cycle. A DNA fragment containing the Zta gene sequence was inserted into an expression vector, pMAMneo, in a sense and antisense direction under a dexamethasone-inducible murine mammary tumor virus LTR promoter, resulting in the construction of plasmids pZ(+) and pZ(-), respectively. Synthesis of Zta protein was reduced in pZ(-)-transfected cells upon dexamethasone induction. Because D-form early antigen and DNA polymerase are essential for viral DNA replication, the contents of these two viral proteins were examined. Amounts of the two lytic proteins were observed to be significantly repressed in pZ(-)-transfected cells. In contrast, both proteins were normally expressed in the sense plasmid pZ(+) or cells transfected with vector alone. Above results demonstrate that Zta antisense RNA can reduce the production of Zta protein and the other lytic proteins, possibly resulting in the inhibition of EBV replication. Copyright 1997 S. Karger AG, Basel
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PMID:Inhibition of the Synthesis of Proteins Needed for Epstein-Barr Virus Replication by Antisense RNA against the Zta Gene. 1172 46

Human herpesvirus 6 (HHV-6), which is present in more than 90% of the human, is known to cause infectious diseases in immuno-compromised patients, e.g., transplant patients. To clarify the possible role of the pattern of expression of HHV-6 genes in various types of HHV-6B infection, we sought to determine whether or not viral DNA microarray could be used for detailed characterization of viral transcription using a HHV-6B DNA microarray that contains 97 known open reading frames of HHV-6B. A subset of genes are preferentially expressed in persistent infection: U16 (IE-B, transactivator, US22 gene family), U18 (IE-B, homolog to HCMV IE glycoprotein), U20 (glycoprotein), U27 (DNA polymerase processivity transactivator), U82 (gL, gH accessory protein), U83 (chemokine), U85 (OX-2 homology, glycoprotein), U90 (IE-A), and U94 (transactivator), respectively. Although the function of each HHV-6B is not fully understood, our study suggests that comprehensive analysis of HHV-6B transcription is useful not only to clarify the pathogenesis of the virus but also to develop new strategies for anti-viral drugs.
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PMID:Transcriptional profiling of human herpesvirus type B (HHV-6B) in an adult T cell leukemia cell line as in vitro model for persistent infection. 1572 Dec 66


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