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:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

An RNA polymerase II activator often contains several regions that contribute to its potency, an organization ostensibly analogous to the modular architecture of promoters and enhancers. The regulatory significance of this parallel organization has not been systematically explored. We considered this problem by examining the activation domain of the Epstein-Barr virus transactivator ZEBRA. We performed our experiments in vitro so that the activator concentrations, stabilities, and affinities for DNA could be monitored. ZEBRA and various amino-terminal deletion derivatives, expressed in and purified from Escherichia coli, were assayed in a HeLa cell nuclear extract for the ability to activate model reporter templates bearing one, three, five, and seven upstream ZEBRA binding sites. Our data show that ZEBRA contains four modules that contribute to its potency in vitro. The modules operate interchangeably with promoter sites to determine the transcriptional response such that the loss of modules can be compensated for by increasing promoter sites. Potassium permanganate footprinting was used to show that transcriptional stimulation is a consequence of the activator's ability to promote preinitiation complex assembly. Kinetic measurements of transcription complex assembly in a reconstituted system indicate that ZEBRA promotes formation of a subcomplex requiring the TFIIA and TFIID fractions, where TFIIA acts as an antirepressor. We propose a model in which the concentration of DNA-bound activation modules in the vicinity of the gene initiates synergistic transcription complex assembly.
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PMID:The ZEBRA activation domain: modular organization and mechanism of action. 841 94

We performed high-sensitivity flow cytometry and Western blotting to study the expression of the low-affinity NGF receptor (p75NGFR) and of the transmembrane tyrosine kinase (Trk) family of high-affinity receptors for the different neurotrophic factors on Epstein-Barr virus (EBV)-transformed human B lymphocytes. Reverse transcriptase (RT) polymerase chain reaction (PCR) with single and multiple sets of primers (multiplex RT-PCR) was used to survey the repertoire of neurotrophin receptor transcripts in this cell line. We demonstrated that transformed B cells express detectable levels of Trk b and its mRNA. Conversely, negative results were obtained for p75NGFR, Trk a, and Trk c. Exposure of EBV-transformed B lymphocytes to brain-derived neurotrophic factor (BDNF) triggered the phosphorylation of Trk b, as demonstrated by Western blots of cell lysates probed with monoclonal antibody against phosphotyrosine.
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PMID:Expression of high- and low-affinity neurotrophin receptors on human transformed B lymphocytes. 863 56

Epstein-Barr nuclear antigen 2 (EBNA2), one of the six viral nuclear proteins expressed in latently infected B lymphocytes, is essential to the immortalization of B cells by Epstein-Barr virus (EBV). EBNA2 promotes transcriptional transactivation of viral and cellular genes by acting as an adapter molecule that binds to cellular sequence-specific DNA-binding proteins, JK recombination signal-binding protein (RBP-JK), and PU.1 and engages multiple members of the RNA polymerase II transcription complex. In the present study, we show that EBNA2 also interacts with hSNF5/Ini1, the human homolog of the yeast transcription factor SNF5. Gel filtration fractionation of partially purified EBV-positive lymphocyte nuclear extracts shows that a fraction of EBNA2 coelutes with both hSNF5/Ini1 and BRG1, a human homolog of SWI/SNF2, in the high-molecular-mass region (1.5 to 2.0 MDa) of a Superose 6 chromatogram. An affinity-purified rabbit antibody directed against hSNF5/Ini1 coimmunoprecipitates EBNA2 from this high-molecular-mass nuclear protein fraction, demonstrating that EBNA2 and hSNF5/Ini1 interact in vivo. This interaction is restricted to a subpopulation of phosphorylated viral EBNA2. Deletion mutation analysis of EBNA2 shows that the proline-rich aminoterminal end and a domain within the divergent region of EBNA2 mediate EBNA2-hSNF5/Ini1 interaction. Since the SNF-SWI complex participates in gene regulation through the alteration of nucleosome configuration and may be a component of the RNA polymerase II holoenzyme, the EBNA2-hSNF5/Ini1 interaction supports the hypothesis that EBNA2 facilitates transcriptional transactivation by acting as a transcription adapter molecule. We postulate that EBNA2 engages the hSNF-SWI complex to generate an open chromatin conformation at the EBNA2-responsive target genes, thereby potentiating the function of the RBP-JK-EBNA2-polymerase II transcription complex.
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PMID:Epstein-Barr virus nuclear protein 2 (EBNA2) binds to a component of the human SNF-SWI complex, hSNF5/Ini1. 870 24

ABL-MYC is a recombinant retrovirus that constitutively expresses the v-abl and c-myc oncogenes. When used to infect immunized mice this virus rapidly and efficiently induces plasmacytomas of which an unusually high percentage secrete antigen (Ag)-specific monoclonal antibodies. These findings suggested that ABL-MYC targets Ag-stimulated B cells for transformation and that infection of lymphoid cells in vitro might be a useful, alternative method for generating monoclonal, Ag-specific plasmacytomas (ASPCTs). Therefore, we used helper virus-free ABL-MYC to infect suspensions of cells from spleens and other lymphoid organs from mice that had been immunized with a variety of Ags and transplanted them into naive mice. The results show that ABL-MYC preferentially transforms splenocytes that are Ag-reactive. They also demonstrate that ASPCTs can be produced by in vitro infection of cell suspensions from the spleen, lymph nodes and Peyer's patches of mice that had been immunized intraperitoneally with sheep red blood cells, Escherichia coli core RNA polymerase or Epstein-Barr virus gp340 protein or immunized orally with live Giardia lamblia parasites. The ASPCTs usually consisted of one to three colnes, secreted antibodies that were quantitatively and qualitatively similar to those obtained from hybridomas, and could continue to secrete Ag-reactive antibody over eight transplant generations.
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PMID:The ABL-MYC retrovirus generates antigen-specific plasmacytomas by in vitro infection of activated B lymphocytes from spleen and other murine lymphoid organs. 889 Aug 96

The anion transporter, band 3, is a ubiquitous protein. It is present in brain and all other tissues examined. Not only is band 3 present in cell membranes, but also in nuclear, Golgi and mitochondria membranes. There are four isoforms of band 3, the anion exchanger (AE) proteins, thus far discovered. They are products of different genes. Lymphocytes are reported to contain AE2, but not AE1. We hypothesized that induction or up-regulation of AE1 occurs when lymphocytes are transformed as an initial event in the path to malignancy. We transformed lymphocytes containing a single base mutation with Epstein Barr Virus (EBV). The mutation of band 3, high transport band 3 (HTbd3), exhibits anion transport that is 2-3 times normal in erythrocytes which contain AE1. This facilitated our identification of AE1 since the probability that 2 different gene products would have the same mutation approaches zero. Thus, we have a base mutation in addition to linear sequence to identify AE1. A 133 base pair (bp) fragment including the affected region was amplified from the mRNA of lymphocytes from the HTbd3 mutant. AE1 primers were used to amplify regions of interest. Reverse transcriptase polymerase chain reaction (RT-PCR) was used to generate cDNA which was sequenced. The sequence of the crucial 133 base pair segment from high transport lymphocytes was 100% identical to the sequence published for the red blood cell band 3 of the same mutant. As reported previously for erythrocytes, this mutation is a C-->T base change which changes a proline to leucine in the protein sequence. Restriction enzyme digests of AE1 cDNA from normal and HTBD3 lymphocytes confirmed that the proposita was homozygous for the mutation, and showed the father to be heterozygous. Anion transport was increased in HTbd3 EBV transformed lymphocytes, as was the case with HTbd3 erythrocytes. AE2 was identified in lymphocytes by sequence. Thus, EBV transformed lymphocytes express the erythroid band 3 (AE1) in addition to AE2.
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PMID:Human erythroid band 3 "anion exchanger 1" is expressed in transformed lymphocytes. 896 Jul 72

The expression of herpes simplex virus 1 gamma (late) genes requires functional alpha proteins (gamma1 genes) and the onset of viral DNA synthesis (gamma2 genes). We report that late in infection after the onset of viral DNA synthesis, cell nuclei exhibit defined structures which contain two viral regulatory proteins (infected cell proteins 4 and 22) required for gamma gene expression, RNA polymerase II, a host nucleolar protein (EAP or L22) known to be associated with ribosomes and to bind small RNAs, including the Epstein-Barr virus small nuclear RNAs, and newly synthesized progeny DNA. The formation of these complexes required the onset of viral DNA synthesis. The association of infected cell protein 22, a highly posttranslationally processed protein, with these structures did not occur in cells infected with a viral mutant deleted in the genes U(L)13 and U(S)3, each of which specifies a protein kinase known to phosphorylate the protein.
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PMID:Association of herpes simplex virus regulatory protein ICP22 with transcriptional complexes containing EAP, ICP4, RNA polymerase II, and viral DNA requires posttranslational modification by the U(L)13 proteinkinase. 899 34

Reverse-transcriptase polymerase chain reaction has been used to analyze the expression of 2 latent genes (EBNA-1 and LMP-1) and one replicative gene (BZLF-1) of Epstein-Barr virus in mononuclear cells from bone marrow and peripheral blood of healthy donors. EBV-gene transcription was detected in 8 out of 15 bone-marrow samples. Among these, 5 allowed the detection of latency-associated transcripts in the absence of BZLF-1 expression. Only one sample showed positivity for expression of both latent and lytic genes. In 2 cases, BZLF-1 was the only transcript detected. In peripheral blood, 4 out of 7 samples showed evidence of EBNA-1 transcription; LMP-1 was expressed in 5 samples, and in 2 cases concomitant expression of EBNA-1 and BZLF-1 was detected. These results provide a direct demonstration by RT-PCR of EBV-gene transcription in bone-marrow-resident viral infected cells and suggest, in contrast to previous studies on peripheral blood, that LMP-1 and BZLF-1 are frequently transcribed also in absence of EBV-related disease. The heterogeneous viral gene expression found makes it difficult to define a pattern of viral latency in vivo which coincides with that described for lymphoblastoid or Burkitt's-lymphoma cell lines at different stages of differentiation.
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PMID:Transcription of latent and replicative Epstein-Barr-virus genes in bone-marrow and peripheral-blood mononuclear cells of healthy donors. 905 50

Epstein-Barr virus (EBV) has been shown to establish latency in resting B lymphocytes of the peripheral blood. This creates a virus reservoir in contrast to lytic virus replication, which is thought to be restricted to differentiated epithelial cells in vivo. So far, the route of transmission between B cells and the production of progeny virus in the epithelial tissue has remained unclear. Reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry analysis of 16 patients with acute infectious mononucleosis (IM) and 25 healthy seropositive donors was performed to detect lytic replication gene products in B lymphocytes of the peripheral blood. Transcriptional activity was found in peripheral blood B lymphocytes (PBLs) for BZLF1 in 88%, BALF2 in 50%, and BcLF1 in 25% of the tested IM patients. All positive results were further confirmed in enriched B-cell populations by antigen determination using immunostaining with the APAAP technique. Furthermore, we detected transcripts for BZLF1 in 72% and for BALF2 in 16% of peripheral B lymphocytes of healthy seropositive donors. In contrast to patients with IM, no signals for BcLF1 were ever found in healthy seropositive donors. In these individuals, lytic replication of EBV is probably restricted by immunologic and gene regulatory mechanisms, whereas in the absence of immunologic control, reflected here by IM patients, the production of infectious virus becomes visible in PBLs.
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PMID:Lytic replication of Epstein-Barr virus in the peripheral blood: analysis of viral gene expression in B lymphocytes during infectious mononucleosis and in the normal carrier state. 929 55

Structures consistent in size, shape and character with various stages of a Lentivirus replicative cycle were observed by electron microscopy in 12-day peripheral-blood lymphocyte cultures from 10 of 17 Chronic Fatigue Syndrome patients and not in controls. Attempts to identify a lymphoid phenotype containing these structures by immunogold labelling failed and the results of reverse-transcriptase assay of culture supernatants were equivocal. The study was blind and case-controlled, patients being paired with age, sex and ethnically matched healthy volunteers. Prescreening of subjects included the common metabolic and immunological disorders, functional conditions and a virus-screen against hepatitis B and C, Epstein-Barr Virus, Cytomegalovirus and Human Immunodeficiency Virus.
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PMID:Electron microscopic immunocytological profiles in chronic fatigue syndrome. 920 53

Recent experiments have strongly suggested that the process of somatic mutation is linked to transcription initiation. It was postulated that a mutator factor loads onto the RNA polymerase and, during elongation, causes transcriptional arrest that activates DNA repair, thus occasionally causing errors in the DNA sequence. We report the analysis of the role of one of the known DNA repair systems, nucleotide excision repair (NER), in somatic mutation. Epstein-Barrvirus-transformed B cells from patients with defects in NER (XP-B, XP-D, XP-V, and CS-A) were studied. Their heavy and light chain genes show a high frequency of point mutations in the variable (V), but not in the constant (C) regions. This suggests that these B cells can undergo somatic hypermutation despite significant defects in NER. Thus, it is doubtful that NER is an essential part of the mechanism of somatic hypermutation of Ig genes. As an aside, NER seems also not involved in Ig gene switch recombination.
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PMID:B lymphocytes of xeroderma pigmentosum or Cockayne syndrome patients with inherited defects in nucleotide excision repair are fully capable of somatic hypermutation of immunoglobulin genes. 923 93


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