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)

HIV-1 gene expression requires the transactivator Tat, which stimulates viral transcript elongation. Recent results show that two cellular cyclin-dependent kinases, which phosphorylate the carboxy-terminal domain of the RNA polymerase II large subunit, contact Tat and contribute to the control of transcriptional elongation.
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PMID:Transcriptional control: Tat cofactors and transcriptional elongation. 965 70

By binding to the transactivation response element (TAR) RNA, the transcriptional transactivator (Tat) from the human immunodeficiency virus increases rates of elongation rather than initiation of viral transcription. Two cyclin-dependent serine/threonine kinases, CDK7 and CDK9, which phosphorylate the C-terminal domain of RNA polymerase II, have been implicated in Tat transactivation in vivo and in vitro. In this report, we demonstrate that CDK9, which is the kinase component of the positive transcription elongation factor b (P-TEFb) complex, can activate viral transcription when tethered to the heterologous Rev response element RNA via the regulator of expression of virion proteins (Rev). The kinase activity of CDK9 and cyclin T1 is essential for these effects. Moreover, P-TEFb binds to TAR only in the presence of Tat. We conclude that Tat-P-TEFb complexes bind to TAR, where CDK9 modifies RNA polymerase II for the efficient copying of the viral genome.
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PMID:The ability of positive transcription elongation factor B to transactivate human immunodeficiency virus transcription depends on a functional kinase domain, cyclin T1, and Tat. 969 9

Mounting evidence suggests that eukaryotic RNA polymerases preassociate with multiple transcription factors in the absence of DNA, forming RNA polymerase holoenzyme complexes. We have purified an apparent RNA polymerase I (Pol I) holoenzyme from Xenopus laevis cells by sequential chromatography on five columns: DEAE-Sepharose, Biorex 70, Sephacryl S300, Mono Q, and DNA-cellulose. Single fractions from every column programmed accurate promoter-dependent transcription. Upon gel filtration chromatography, the Pol I holoenzyme elutes at a position overlapping the peak of Blue Dextran, suggesting a molecular mass in the range of approximately 2 MDa. Consistent with its large mass, Coomassie blue-stained sodium dodecyl sulfate-polyacrylamide gels reveal approximately 55 proteins in fractions purified to near homogeneity. Western blotting shows that TATA-binding protein precisely copurifies with holoenzyme activity, whereas the abundant Pol I transactivator upstream binding factor does not. Also copurifying with the holoenzyme are casein kinase II and a histone acetyltransferase activity with a substrate preference for histone H3. These results extend to Pol I the suggestion that signal transduction and chromatin-modifying activities are associated with eukaryotic RNA polymerases.
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PMID:Histone acetyltransferase and protein kinase activities copurify with a putative Xenopus RNA polymerase I holoenzyme self-sufficient for promoter-dependent transcription. 985 2

Cyc8(Ssn6)-Tup1, a general co-repressor complex, is recruited to promoter DNA via interactions with DNA-binding regulatory proteins and inhibits the transcription of many different yeast genes. Previous studies have established that repression function of the complex is performed by one subunit of the complex, the Tup1 protein, and requires specific components of the RNA polymerase II holoenzyme such as Sin4 and Rgr1. In this study we test the transcriptional activity of the Cyc8 subunit using a lexA operator-containing reporter. We show that a LexA-Cyc8 hybrid stimulates transcription when expressed in a tup1Delta, a sin4Delta, or a rgr1Delta strain, suggesting that transcriptional activation is an intrinsic property of the Cyc8-Tup1 co-repressor. In support of this notion we demonstrate that Cyc8-Tup1 has a dual function on CIT2, a gene encoding a citrate synthase that is expressed upon mitochondrial dysfunction. First, we show that Cyc8-Tup1 is tethered to CIT2 promoter by interacting with the activation domain of Rtg3, a bHLH/L-Zip DNA-binding transactivator of CIT2. Next we demonstrate that Cyc8-Tup1 activates CIT2 transcription in response to mitochondrial dysfunction, and this stimulatory effect is mediated by Cyc8. In contrast, basal (noninduced) expression of this gene is inhibited by Tup1. These findings establish a positive role for the Cyc8-Tup1 complex in transcription and support a model by which specific metabolic signals may convert the Cyc8-Tup1 transcriptional co-repressor to a co-activator of certain promoters.
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PMID:The Tup1-Cyc8 protein complex can shift from a transcriptional co-repressor to a transcriptional co-activator. 986 31

The potent transactivator Tat recognizes the transactivation response RNA element (TAR) of human immunodeficiency virus type 1 and stimulates the processivity of elongation of RNA polymerase (Pol) II complexes. The cellular proteins Tat-SF1 and human SPT5 (hSPT5) are required for Tat activation as shown by immunodepletion with specific sera and complementation with recombinant proteins. In nuclear extracts, small fractions of both hSPT5 and Pol II are associated with Tat-SF1 protein. Surprisingly, the RAP30 protein of the heterodimeric transcription TFIIF factor is associated with Tat-SF1, while the RAP74 subunit of TFIIF is not coimmunoprecipitated with Tat-SF1. Overexpression of Tat-SF1 and hSPT5 specifically stimulates the transcriptional activity of Tat in vivo. These results suggest that Tat-SF1 and hSPT5 are indispensable cellular factors supporting Tat-specific transcription activation and that they may interact with RAP30 in controlling elongation.
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PMID:Tat-SF1 protein associates with RAP30 and human SPT5 proteins. 1045 43

We examined the expression of messenger RNA (mRNA) of the human inducible nitric oxide synthase (hiNOS) gene in a panel of human T-cell lines. Reverse transcriptase-polymerase chain reaction showed that human T-cell leukemia virus type-I (HTLV-I)-infected T-cell lines (MT-1, SLB-1, and C5/MJ) expressed mRNA for the hiNOS, but TL-Om1 or uninfected Jurkat, H9, and CCRF-CEM did not. The MT-1, SLB-1, and C5/MJ cell lines are infected with HTLV-I and express the viral transactivator Tax, whereas TL-Om1 cells, although derived from adult T-cell leukemia (ATL) leukemic cells, do not express Tax. There was, thus, a correlation between Tax and hiNOS mRNA expression. The transcriptional regulatory region of the hiNOS gene was activated by Tax in Jurkat, in which endogenous hiNOS is induced by Tax. Deletion analysis showed that the region of hiNOS encompassing nucleotides -159 to -111 contained the minimum Tax-responsive elements. Mutations in the NF-kappaB element at position -115 and -106 bp in the hiNOS promoter were still activated by Tax, and a Tax mutant defective for activation of the NF-kappaB pathway retained the ability to activate the hiNOS promoter. In addition, overexpression of the dominant-negative mutants of IkappaBalpha and I kappaBbeta failed to reduce Tax-induced activation of hiNOS gene. Furthermore, hiNOS mRNA was detected in leukemic cells from ATL patients. Our results show that the hiNOS promoter contains a minimum Tax-responsive element located between nucleotides -159 and -111, and imply that the expression of the hiNOS gene is involved in the pathogenesis of HTLV-I-associated diseases.
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PMID:Expression of human inducible nitric oxide synthase gene in T-cell lines infected with human T-cell leukemia virus type-I and primary adult T-cell leukemia cells. 1051 90

HIV-1 gene expression relies upon a complex machinery that is primarily controlled by two viral regulatory proteins, Tat and Rev. Rev is involved in regulating post-transcriptional events of HIV-1 gene expression. The Tat protein transactivates transcription from the HIV-1 5' long terminal repeat (LTR) and acts in synergy with specific cellular factors. Recently, it has been shown that one set of these cellular factors is a protein kinase activity termed TAK (Tat-associated kinase), which activates transcription by hyperphosphorylation of the carboxyl-terminal domain (CTD) of the large subunit of RNA polymerase II. TAK also enhances transcription of HIV-2, together with the retroviral transactivator, Tat-2. The TAK activity appears to be related to the CTD kinase P-TEFb, which stabilizes transcription elongation of many genes and was originally isolated from Drosophila extracts. Both TAK and P-TEFb contain at least two subunits: the cyclin-dependent kinase, CDK9 (PITALRE), the catalytic subunit, and the regulatory subunit, cyclin T1. CDK9 and cyclin T1 are ubiquitous factors that affects many cellular processes, including cell differentiation and apoptosis. The involvement of TAK in HIV-1 and HIV-2 gene expression is an important aspect in the biology of these two retroviruses, and may lead to the development of novel antiretroviral drugs and/or gene therapy approaches for the treatment of patients with AIDS.
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PMID:Regulatory functions of Cdk9 and of cyclin T1 in HIV tat transactivation pathway gene expression. 1053 59

Transcription from the HIV-1 long terminal repeat (LTR) is regulated by the viral transactivator Tat, which increases RNA polymerase II (RNAP II) processivity. Previous reports have demonstrated that phosphorylation of the RNAP II carboxy-terminal domain by TFIIH and P-TEFb is important for Tat transactivation. Our present results demonstrate that phosphorylation of the RAP74 subunit of TFIIF is also an important step in Tat transactivation. Interestingly, while the general transcription factor TFIIF is required for both basal and Tat-activated transcription, phosphorylation of the RAP74 subunit occurs in the presence of Tat and correlates with a high level of transcription activity. Using a biotinylated DNA template transcription assay, we provide evidence that RAP74 is phosphorylated by TAF(II)250 during Tat-activated transcription. Depletion of RAP74 from the HeLa nuclear extract inhibited HIV-1 LTR-driven basal transcription and Tat transactivation. The addition of TFIIF, reconstituted from recombinant RAP30 and RAP74, to the depleted HeLa nuclear extract resulted in restoration of Tat transactivation. Of importance, the exogenous RAP74 was rapidly phosphorylated in the presence of Tat. These results suggest that RAP74 phosphorylation is one important step, of several, in the Tat transactivation cascade.
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PMID:Phosphorylation of the RAP74 subunit of TFIIF correlates with Tat-activated transcription of the HIV-1 long terminal repeat. 1070 53

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

The activation of the HIV-1 long terminal repeat (LTR) by the viral transcriptional transactivator Tat is an essential step in the viral replication cycle. To increase the processivity of RNA polymerase II, Tat interacts with the positive transcription elongation factor b (P-TEFb) and cyclin-dependent kinase (CDK)-activating kinase (CAK). In this study, we demonstrate that a pseudo-substrate peptide for CDK7, mC2p, inhibits HIV-1 replication as well as Tat transactivation. Specifically, mC2p blocks only the activity of CAK and not that of P-TEFb. Moreover, mC2p inhibits Tat transactivation and HIV replication. Therefore, the activation of CDK7 by Tat is considered a critical step of Tat transactivation and mC2p and related compounds represent potential candidates for novel anti-HIV therapeutics.
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PMID:HIV-1 replication is inhibited by a pseudo-substrate peptide that blocks Tat transactivation. 1079 93


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