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)

The tRNATyr precursor molecule, synthesized from phi 80 psu3+ DNA (containing a single tRNA gene) by DNA-dependent RNA polymerase and q factor, was about 205 nucleotides long. The main product of its digestion with a ribonuclease tii preparation from Escherichia coli showed the same electrophoretic mobility as tRNAtyr precursor isolated in vivo and was found to be identical to it when analysed using fingerprint techniques. This intermediate precursor synthesized in vitro was converted further by processing with ribonuclease P into an RNA identical size to mature tRNATyr. It was concluded that the initiation of transcription of the tRNATyr gene in vitro occurs at the same site as that of transcription in vivo and a termination occurs at about 80 nucleotides beyond the CCA end of tRNATyr.
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PMID:Processing by ribonuclease II of the tRNATyr precursor of Escherichia coli synthesized in vitro. 32 7

The gene for the RNA subunit of ribonuclease P from the extreme thermophilic eubacterium T. thermophilus HB8 was cloned using oligonucleotide probes complementary to conserved regions of RNase P RNA subunits from proteobacteria. The monocistronic gene and its flanking regions were sequenced. The gene is enclosed by a promoter and a rho-independent terminator. Nuclease S1 protection analyses showed that the primary transcript is identical with the mature RNA, i.e. no processing events are involved. The stem and loop structure of the terminator remains part of the mature molecule. In vitro transcription of the cloned gene with purified RNA polymerase from T. thermophilus yields the same RNA product as in vivo, indicating that no other components except RNA polymerase are involved in the synthesis of the RNA. RNase P RNA from T. thermophilus cleaved a pre-tRNA(Tyr) from E. coli with highest efficiency between 55 degrees C and 65 degrees C. The T. thermophilus RNA, which has a G-C content of 86% in helical regions, displays several structural idiosyncrasies, although its secondary structure is similar to that of proteobacteria. Numerous invariable nucleotides in the structural core of eubacterial RNase P RNAs are also conserved in the RNA from the extreme thermophilic eubacterium.
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PMID:Analysis of the gene encoding the RNA subunit of ribonuclease P from T. thermophilus HB8. 171 85

We have constructed a plasmid expressing E. coli M1 RNA, the catalytic RNA subunit of ribonuclease P, under the control of a phage T7 promoter. The active M1 RNA species synthesized in vitro by T7 RNA polymerase from this vector was reacted with the tRNA(Gln) - tRNA(Leu) precursor RNA (Band K) encoded by phage T4. Only the tRNA(Leu) moiety of this dimeric precursor RNA contains the 3' terminal C-C-A sequence common to all tRNAs. We observed that protein-free M1 RNA was capable of processing the precursor RNA at the 5' ends of both tRNA tRNA sequences. The rate of cleavage of the tRNA(Gln) sequence was more strongly dependent on [Mg2+] than that of tRNA(Leu), increasing severalfold between 100 and 500 mM Mg2+, conditions under which the rate of cleavage at the tRNA(Leu) sequence was constant.
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PMID:Dependence of M1 RNA substrate specificity on magnesium ion concentration. 245 26

A transfer RNA complete devoid of modified nucleosides was synthesized by in vitro transcription, and some of its properties in aminoacylation and protein synthesis in vitro were studied. For this purpose, a plasmid was constructed which contained a glycine tRNA gene from Mycoplasma mycoides under the promoter of the T7 RNA polymerase, as well as a BstNI restriction site at the 3'-end of the tRNA gene. Cleavage of plasmid DNA with BstNI followed by T7 RNA polymerase transcription in vitro yielded an RNA which was processed with M1 RNA, the catalytic subunit of ribonuclease P, to give a tRNA of mature length. The tRNA synthesized in this manner can be esterified with glycine in vitro, and the rate of aminoacylation is the same as when using the corresponding fully modified glycine tRNA from M. mycoides. Furthermore, in protein synthesis in vitro, the tRNA lacking modified nucleosides was essentially as efficient as the corresponding normal glycine tRNA. However, the Escherichia coli extract used in our protein-synthesizing system introduced one modification, pseudouridine, into the in vitro-synthesized tRNA, and it cannot be excluded that this modification has an essential role in protein synthesis.
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PMID:Properties of a transfer RNA lacking modified nucleosides. 284 31

The repertoire of 4,431 open reading frames (ORFs), eight rRNA operons and 98 tRNA genes of Chromobacterium violaceum must be expressed in a regulated manner for successful adaptation to a wide variety of environmental conditions. To accomplish this feat, the organism relies on protein machineries involved in transcription, RNA processing and translation. Analysis of the C. violaceum genome showed that transcription initiation, elongation and termination are performed by the five well-known RNA polymerase subunits, five categories of sigma 70 factors, one sigma 54 factor, as well as six auxiliary elongation and termination factors. RNA processing is performed by a variety of endonucleases and exonucleases, such as ribonuclease H, ribonuclease E, ribonuclease P, and ribonuclease III, in addition to poly(A) polymerase and specific methyltransferases and pseudouridine synthases. ORFs for all ribosomal proteins, except S22, were found. Only 19 aminoacyl-tRNA synthetases were found, in addition to three aminoacyl-tRNA synthetase-related proteins. Asparaginyl-tRNA (Asn) is probably obtained by enzymatic modification of a mischarged aminoacyl-tRNA. The translation factors IF-1, IF-2, IF-3, EF-Ts, EF-Tu, EF-G, RF-1, RF-2 and RF-3 are all present in the C. violaceum genome, although the absence of selB suggests that C. violaceum does not synthesize selenoproteins. The components of trans-translation, tmRNA and associated proteins, are present in the C. violaceum genome. Finally, a large number of ORFs related to regulation of gene expression were also found, which was expected, considering the apparent adaptability of this bacterium.
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PMID:Gene expression in Chromobacterium violaceum. 1510 Sep 88

RNA polymerase III promoters of human ribonuclease P RNA component H1, human U6, and mouse U6 small nuclear RNA genes are commonly used in short hairpin RNA (shRNA) expression vectors due their precise initiation and termination sites. During transient transfection of shRNA vectors, we observed that H1 or U6 promoters also express longer transcripts enough to express several reporter genes including firefly luciferase, green fluorescent protein EGFP, and red fluorescent protein JRed. Expression of such longer transcripts was augmented by upstream RNA polymerase II enhancers and completely inhibited by downstream polyA signal sequences. Moreover, the transcription of firefly luciferase from human H1 promoter was sensitive to RNA polymerase II inhibitor alpha-amanitin. Our findings suggest that commonly used polymerase III promoters in shRNA vectors are also prone to RNA polymerase II mediated transcription, which may have negative impacts on their targeted use.
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PMID:RNA polymerase II mediated transcription from the polymerase III promoters in short hairpin RNA expression vector. 1630 Jul 30

The physical and functional links between transcription and processing machines of tRNA in the cell remain essentially unknown. We show here that whole HeLa extracts depleted of ribonuclease P (RNase P), a tRNA-processing ribonucleoprotein, exhibit a severe deficiency in RNA polymerase (Pol) III transcription of tRNA and other small, noncoding RNA genes. However, transcription can be restored by the addition of a purified holoenzyme. Targeted cleavage of the H1 RNA moiety of RNase P alters enzyme specificity and diminishes Pol III transcription. Moreover, inactivation of RNase P by targeting its protein subunits for destruction using small interfering RNAs inhibits Pol III function and Pol III-directed promoter activity in the cell. RNase P exerts its role in transcription through association with Pol III and chromatin of active tRNA and 5S rRNA genes. The results demonstrate a role for RNase P in Pol III transcription and suggest that transcription and early processing of tRNA may be coordinated.
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PMID:A role for the catalytic ribonucleoprotein RNase P in RNA polymerase III transcription. 1677 78

Influenza A virus (IAV) is a major human pathogen whose genotypic diversity results in unpredictable pandemics and epidemics. Interaction with the cell nucleus is essential to IAV infection, allowing recruitment of cellular components to facilitate virus replication. Viral proteins are also targeted to the nucleolus, a subnuclear structure involved in ribosomal biogenesis, RNA maturation, stress response, and control of cell growth, but the functional consequences of this are unclear. We took an unbiased approach to studying IAV-nucleolar interactions by using stable isotope labeling with amino acids in cell culture (SILAC) in conjunction with LC-MS/MS to quantify changes in the nucleolar proteome following infection with A/PR/8/34 (H1N1) and A/Udorn/72 (H3N2) strains of the virus. Only a minority of nucleolar proteins showed significant changes in abundance after infection; these alterations were mostly different between the two strains but could be validated by confocal microscopy of infected cells. Many of the affected proteins comprised functional groupings, including components of ribonuclease P, RNA polymerase I, the MLL1 histone methyltransferase complex, as well as nuclear paraspeckles and the RNA editing apparatus. This, as well as comparison with other viruses that cause changes in the nucleolar proteome, suggests that IAV targets specific nucleolar pathways.
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PMID:Quantitative proteomics using SILAC coupled to LC-MS/MS reveals changes in the nucleolar proteome in influenza A virus-infected cells. 2070 60