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)

cAMP is an ubiquitous compound which is involved in the regulation of many biological processes. In bacteria such as E. coli, cAMP mediates the activation of catabolic operons via the CAP protein. The CAP-cAMP complex, whose tridimensional structure has recently been established, binds to the promoter regions of catabolic operons at a specific site, and activates their transcription by inducing RNA polymerase to bind and initiate transcription at the correct site. Various phenomenons including protein-protein interactions or CAP-induced DNA bending or kinking could be involved in the process of forming the open transcription complex. In eukaryotes, cAMP activates cAMP dependent protein kinases which covalently modify proteins by phosphorylation on serine or threonine residues. The catalytically inactive holoenzyme is generally a tetramer containing two regulatory subunits, each capable of binding two molecules of cAMP, and two catalytic subunits. In mammalian cells, two types of cAMP dependent protein kinases (I and II) can be distinguished on the basis of their regulatory subunits; their relative proportion varies from tissue to tissue. Binding of cAMP to the regulatory subunits induces the dissociation of the holoenzyme and releases the free and active catalytic subunits. Phosphorylation of proteins occurs at sequences containing two basic residues in the vicinity of the phosphorylated serine or threonine. A heat-stable protein, present in most eukaryotic cells, specifically interacts with the catalytic subunit and inhibits its activity. The amino-acid sequence of cAMP dependent protein kinases has recently been determined. It is interesting to note that the domains responsible for cAMP binding by the regulatory subunits of mammalian cAMP dependent protein kinases and CAP share important sequence homologies. The same phenomenon is observed concerning the domain responsible for ATP binding to the catalytic subunit of cAMP dependent protein kinases and that of tyrosine-specific protein kinases from oncoviruses. Other eukaryotic proteins such as S-adenosyl-L-homocysteine (SAH) hydrolase are also capable of binding cAMP. The latter is involved in the regulation of S-adenosyl-L-methionine dependent methylations, and its activity could be affected by cAMP. Besides its role as an effector of enzymatic activity via phosphorylation, such as in the regulation of glycogen metabolism, cAMP has recently been shown to activate the transcription of a number of eukaryotic genes. This process probably also involves protein phosphorylation, but its precise mechanism remains to be understood.
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PMID:[Mode of action of cyclic amp in prokaryotes and eukaryotes, CAP and cAMP-dependent protein kinases]. 241 6

In the past, simian virus 40 (SV40) has been used as a cloning vehicle to clone foreign genes by substituting portions of the viral genome vital for viral replication. Propagation of these defective viruses required a helper virus and the recombinant viruses obtained could be grown only as a mixture. In this study, we describe a novel nondefective SV40 vector to clone small RNA polymerase III genes. Two small RNA polymerase III genes, an amber suppressor human serine tRNA gene and the adenovirus (Ad) VAI RNA gene, were cloned in the intron region of the large-T antigen gene of SV40 after deleting DNA sequences coding for the small-t polypeptide. The recombinant viruses grew to wild type levels and showed no growth defects. When CV-1p cells were infected with these viruses, the cloned RNA polymerase III genes were expressed at high levels at late times. Interestingly, large amounts VAI RNA in CV-1p cells infected with SV40-VA recombinant virus, did not enhance translation of viral mRNAs significantly but did lead to a 3 to 4 fold increase in the steady state levels of large-T mRNA suggesting a novel function for VAI RNA in SV40 infected monkey cells. Furthermore, VAI mutants which fail to function in Ad infected human cells also failed to enhance the levels of large-T mRNAs in monkey cells infected with SV40. The simple SV40 vector described here may be useful to study the structure and function of small RNA polymerase III genes in the context of a eucaryotic chromosome. In addition, the nondefective recombinant SV40 which expresses the suppressor tRNA gene at high levels may provide a useful helper system to propagate animal viruses with amber mutations in essential genes.
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PMID:Efficient expression of small RNA polymerase III genes from a novel simian virus 40 vector and their effect on viral gene expression. 246 35

The Escherichia coli lacZ gene contains a series of latent transcriptional terminators that are responsible for the polar effects of certain mutations. We demonstrate, using gel electrophoretic size analyses and nuclease S1 mapping procedures, that RNA polymerase terminates RNA synthesis in the vicinity of five positions 180, 220, 379, 421 and 463 base-pairs downstream from the start point during transcription of lacZ DNA in vitro in the presence of rho factor. Termination at all but the 421 position depends on rho factor. In the in vitro assays with 0.05 M-KCl and excess rho (36 nM), the terminators are moderately effective, having efficiencies that range from about 8% at the 180 base-pair site to 56% at the 463 base-pair site. These termination stop points correspond to five of the 11 transcriptional pause sites between 180 and 463 base-pairs. Several stop points also correspond to 3' end points of lacZ mRNA isolated from cells containing the strongly polar lacZ-U118 mutation and from cells starved for serine, thus confirming that these latent terminators are responsible for the polar effect and demonstrating that they also function under a condition of physiological stress that prevents the transcription from being translated properly. Two other potential termination factors, NusA protein and cyclic AMP receptor protein have no effect in vitro on the efficiency of termination at the five lacZ sites.
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PMID:Identification and characterization of transcription termination sites in the Escherichia coli lacZ gene. 247 37

Cathepsin G is a 26,000-Da serine protease that is found in the azurophil granules of neutrophils and monocytes. The cathepsin G gene is expressed at high levels in U937 promonocytic cells, but is down-regulated with phorbol-induced differentiation. To characterize the genomic sequences responsible for the regulated expression of this gene, we screened a human genomic fibroblast library using cathepsin G cDNA, and obtained two lambda clones that contained the cathepsin G locus. The cathepsin G gene spans 2.7 kilobase pairs of genomic DNA and consists of 5 exons and 4 introns. The genomic organization of cathepsin G is similar to that of human neutrophil elastase, rat mast cell protease II, murine adipsin, and murine cytotoxic T-cell serine proteases, with protease catalytic residues located near the borders of exons 2, 3, and 5. Using in situ hybridization techniques, we localized cathepsin G to chromosome 14q11.2, a site that is near the alpha/delta T-cell receptor complex. Cathepsin G transcription is abolished in U937 nuclei with 2 micrograms/ml alpha-amanitin, indicating that this gene is probably transcribed by RNA polymerase II. The 5' end of the cathepsin G gene was defined by primer extension and S1 nuclease protection assays. A TATA box is found at position -29, and a CAAT box is found at -69 with respect to the transcription initiation site. Having defined the genomic structure and chromosomal location of cathepsin G, we are now attempting to identify the DNA elements in or near this gene that mediate its tissue and development-specific pattern of expression.
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PMID:Genomic organization and chromosomal localization of the human cathepsin G gene. 256 62

Bovine mitochondrial serine tRNA(AGY) gene transcript was synthesized in vitro with T7 RNA polymerase, and it was capable of being aminoacylated with mitochondrial serine tRNA synthetase. The melting profiles of the transcript was similar to those of native serine tRNA(AGY), suggesting that the higher-order structure of the transcript does not much differ from that of native serine tRNA(AGY). Several transcripts with base-substitution were also constructed and their aminoacylation capacity was investigated.
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PMID:Some properties of bovine mitochondrial serine tRNA gene transcript synthesized with T7 RNA polymerase. 260 55

The unique C-terminal repeat domain (CTD) of the largest subunit (IIa) of eukaryotic RNA polymerase II consists of multiple repeats of the heptapeptide consensus sequence Tyr-Ser-Pro-Thr-Ser-Pro-Ser. The number of repeats ranges from 26 in yeast to 42 in Drosophila to 52 in mouse. The CTD is essential in vivo, but its structure and function are not yet understood. The CTD can be phosphorylated at multiple serine and threonine residues, generating a form of the largest subunit (II0) with markedly reduced mobility in NaDodSO4/polyacrylamide gels. To investigate this extensive phosphorylation, which presumably modulates functional properties of RNA polymerase II, we began efforts to purify a specific CTD kinase. Using CTD-containing fusion proteins as substrates, we have purified a CTD kinase from the yeast Saccharomyces cerevisiae. The enzyme extensively phosphorylates the CTD portion of both the fusion proteins and intact subunit IIa, producing products with reduced electrophoretic mobilities. The properties of the CTD kinase suggest that it is distinct from previously described protein kinases. Analogous activities were also detected in Drosophila and HeLa cell extracts.
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PMID:A protein kinase that phosphorylates the C-terminal repeat domain of the largest subunit of RNA polymerase II. 265 24

Actively transcribing eukaryotic RNA polymerase II is highly phosphorylated on its repetitive carboxyl-terminal domain. We have isolated a protein kinase that phosphorylates serine residues in this repetitive domain. A component of this kinase is cdc2, the product of a cell-cycle control gene previously shown to be a component of M-phase-promoting factor and M-phase-specific histone H1 kinase. This observation suggests a role for the cdc2 protein kinase in transcriptional regulation.
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PMID:Phosphorylation of RNA polymerase by the murine homologue of the cell-cycle control protein cdc2. 266 13

Eukaryotic tRNA expression initiates with transcription by RNA polymerase III and requires two additional protein factors and two regions within the tRNA gene (the 5'-internal control region (ICR) or A-box and the 3'-ICR or B-box). Using a reconstituted Saccharomyces cerevisiae RNA polymerase III system, the transcription of various 5'-ICR, 3'-ICR, and double mutation alleles of the Schizosaccharomyces pombe sup3-e dimeric tRNA gene were studied. The sup3-e tRNA locus consists of an upstream serine tRNA gene and a downstream initiator methionine tRNA gene which are transcribed as a dimeric precursor and processed to give two tRNAs. Only the ICRs of the tRNA(Ser) gene are active in directing dimeric gene transcription. Mutations in the 3'-ICR of the tRNA(Ser) gene reduce transcription of the dimer more than those in the 5'-ICR. Mutations in the 5'-ICR were found which greatly increased or decreased transcription of the dimer, while base changes in the 3'-ICR were only found to decrease transcription. This suggests a modulatory role for the 5'-ICR in transcription regulation. Mutation of the methionine tRNA gene ICR has little effect on sup3-e transcription, and no detectable transcripts initiate from the methionine tRNA gene when the tRNA(Ser) gene promoter is inactivated by mutation. Comparison with transcription studies of other mutant tRNA genes suggests that nucleotides sites within the ICRs, such as nucleotides 8, 10, 13, 18, and 19 in the 5'-ICR and 48, 53, 56, 57, and 58 in the 3'-ICR, appear to have evolved universal importance for RNA polymerase III transcription in eukaryotes. Thus these ICR sequences may play a critical role in regulation of tRNA expression.
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PMID:Multiple mutations of the first gene of a dimeric tRNA gene abolish in vitro tRNA gene transcription. 267 99

The C-terminal domain of the largest subunit of RNA polymerase II in higher eukaryotes is present in the protozoan parasite Trypanosoma brucei in a strongly modified form. To determine whether this is a general feature of the Kinetoplastida and to determine the role of this domain in RNA polymerase II transcription, we have analysed the C-terminal domain of the distantly related species Crithidia fasciculata. No positional identity of amino acid residues between the C-termini of C. fasciculata and T. brucei can be found. Moreover, both domains lack the heptapeptide repeat structure present in higher eukaryotes. The two domains are, however, very similar in amino acid composition, being rich in acidic residues as well as serine and tryosine. The latter observation is compatible with the concept that in vivo phosphorylation of the C-terminus activates RNA polymerase II.
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PMID:Unusual C-terminal domain of the largest subunit of RNA polymerase II of Crithidia fasciculata. 272 83

Beet necrotic yellow vein virus (BNYVV) has a quadripartite plus-strand RNA genome in which the two smallest genome components, RNA 3 and 4, are not necessary for virus multiplication in leaves. Infectious transcripts of BNYVV RNA 3 and 4 have already been described (V. Ziegler-Graff, S. Bouzoubaa, I. Jupin, H. Guilley, G. Jonard, and K. Richards (1988) J. Gen. Virol. 69, 2347-2357). In this paper we describe synthesis of a full-length RNA-1 transcript by bacteriophage T7 RNA polymerase-directed run-off transcription of cloned viral cDNA. A recombinant plasmid containing a full-length cDNA insert of RNA 2 could not be maintained in Escherichia coli. Therefore full-length transcript of RNA 2 was produced by transcription of cDNA ligation products without amplification in bacteria. When inoculated together to leaves of Chenopodium quinoa or Tetragonia expansa the RNA 1 and 2 transcripts were infectious; they also supported multiplication of the BNYVV RNA 3 and 4 transcripts, providing a totally synthetic inoculum of the virus. In one recombinant clone of RNA 2 a point mutation causing an arginine to serine substitution at position 119 of the viral coat protein was discovered. The mutation was detected because the resulting coat protein had altered electrophoretic mobility. RNA 2 transcripts containing this mutation were infectious but viral RNA was not encapsidated. The mutation also interfered with long distance movement of the virus in spinach, presumably as a consequence of the packaging deficiency.
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PMID:In vitro synthesis of biologically active beet necrotic yellow vein virus RNA. 277 20


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