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)

Anti-La antibodies are frequently found in patients with autoimmune diseases; the antigen was reported to be a 50,000-Da protein (Rinke, J., and Steitz, J. A. (1982) Cell 29, 149-159). Because this protein was associated with many nascent RNA polymerase III transcripts, it was suggested to be an RNA polymerase III transcription factor. The present study was designed to analyze 4.5 I ribonucleoprotein, an RNA polymerase III transcript which contains the La antigen. It was found that the 3'-end 20-30-nucleotide portion was the most protected portion of 4.5 I RNA when 4.5 I ribonucleoprotein was digested with T1 RNase. When U2 RNA (an RNA polymerase II transcript) and 4.5 I RNA were incubated with the S-100 fraction of Novikoff hepatoma cells, the 4.5 I RNA bound La antigen but the U2 RNA did not. When partial and complete T1 RNase digestion fragments of 4.5 I RNA were incubated with the S-100 fraction, the 3'-end fragments bound preferentially to the La antigen. However, the fragments of 4.5 I RNA bound less efficiently to La antigen than whole 4.5 I RNA. These results indicate that the 3'-end of 4.5 I RNA is the La antigen binding site in this molecule and suggest that the overall conformation of RNA aids in the binding of La antigen.
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PMID:Identification of a La protein binding site in a RNA polymerase III transcript (4.5 I RNA). 686 91

Total HeLa cell extract was separated into multiple components using first DEAE-Sephadex and then phosphocellulose column chromatography. Four major fractions, DE175, DE500, P100, and P1000, from HeLa cells are found to be essential for accurate and efficient transcription of cloned ovalbumin DNA fragments in the reconstituted system. DE175 serves as the source for RNA polymerase II and DE500 minimizes the synthesis of small molecular size RNA. P100 enhances the levels of specific transcription, while P1000 is absolutely required for correct initiation. Chick oviduct crude extract was also fractionated into multiple components according to the same procedure. Similar efficiency of specific initiation could be obtained when an individual fraction (DE175, DE500, P100, or P1000) from oviduct cells was exchanged for a fraction from HeLa cells. These results indicate that chick oviduct tissue also contains general transcription factors like that of HeLa cells and these factors can be fractionated according to the same procedure. In this fractionation scheme, we were able to separate the bulk of RNase activity into the P350 fraction which was not required for initiation of ovalbumin RNA transcription. Thus, this reconstituted system is suitable for studying in vitro transcription in a homologous system derived from tissue extracts, even though a substantial amount of cellular ribonuclease may be associated with it.
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PMID:Transcription factors from oviduct and HeLa cells are similar. 689 17

We have examined the effects of steroid hormones in the chromatin sensitivity of the ovalbumin gene to micrococcal nuclease and have attempted to define the importance of the nucleosome core, higher order chromatin folding, and transcription in the maintenance of the nuclease-sensitive conformation of the ovalbumin chromatin. Solution hybridization studies demonstrated that the sensitivity of the ovalbumin gene in oviduct nuclei to micrococcal nuclease paralleled the hormone-dependent transcription of the ovalbumin gene in the immature chick. Blot hybridization analysis also revealed a hormone-dependent change in this chromatin region since ovalbumin DNA fragments from nuclease-treated hen and estrogen-stimulated chick oviduct nuclei exhibited nucleosomal repeat patterns that were less discrete than those observed for the ovalbumin specific fragments from liver and hormone-withdrawn oviducts. This transcription-related conformation was not the result of enhanced sensitivity of the ovalbumin-containing nucleosomal cores since the bulk of the nucleosomes associated with the ovalbumin chromatin were not preferentially cleaved internally by micrococcal nuclease. Rather, an analysis of the fragmentation of the ovalbumin chromatin as a function of digestion extent suggested a mechanism in which the heightened sensitivity resulted from the collective expansion of the nuclease cutting sites in the linker regions of the ovalbumin chromatin because the gene was in an unfolded conformation. The transcription-specific conformation was not merely a consequence of RNA synthesis per se since the selective sensitivity of the gene was unaffected by treatment of oviduct nuclei with alpha-amanitin, actinomycin D, or RNase. In addition, the presence of the transcriptional complex on the ovalbumin chromatin was presumably not required for selective nuclease recognition since preferential cleavage was observed under conditions expected to deplete oviduct nuclei of template-bound RNA polymerase and nascent RNA chains. These results are consistent with a model in which the expressed ovalbumin gene is in an unfolded polynucleosomal structure whose formation is related to transcriptional activity but not dependent on the transcriptional process.
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PMID:Hormonal regulation of the conformation of the ovalbumin gene in chick oviduct chromatin. 713 Jan 93

To study the mechanism of the synthesis of Semliki Forest virus (SFV) 26-S RNA, we have isolated the 5'-terminal 'cap'-containing RNase-T1-resistant oligonucleotide (T1 cap) from the genomic 42-S RNA and from the subgenomic 26-S RNA and determined their nucleotide sequences. The T1 caps were purified from 32P-labelled RNAs on two successive two-dimensional fractionation systems: (a) electrophoresis on cellulose acetate paper followed by homochromatography and (b) two-dimensional polyacrylamide gel electrophoresis. The T1 caps derived from the two RNAs had different mobilities in both systems. Their nucleotide sequence was found to be: m7G(5')ppp-(5')ApUpGp- for the 42-S RNA and m7G(5')ppp(5')ApUpUpGp- for the 26-S RNA, respectively. Thus, it appears that the 26-S RNA is not formed by initiation of the RNA polymerase at the 3' end of the negative-strand template followed by 'cleavage and splicing' or as the result of a 'polymerase jump'. Our results, instead, favour the model of internal initiation of the polymerase on the 42-S negative-strand RNA template.
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PMID:The nucleotide sequences of the 5'-terminal T1 oligonucleotides of Semliki-Forest-virus 42-S and 26-S RNAs are different. 737 41

Transgenic mice expressing rat parvalbumin under the control of the human metallothionein IIA (MTII A), SV-40 early, and neuron-specific enolase (NSE) promoters were produced. Ectopic expression was analyzed by RNA polymerase chain reaction and RNase protection in combination with immunohistochemistry. From a total of 25 transgenic lines 18 were found to express the transgene. Expression strength and tissue specificity were dependent upon the promoter used and varied considerably among animal lines produced with the same construct. Highest constitutive MT IIA-driven expression was found in lung, liver, heart, and kidney, as well as in brain, and lower amounts of transgene expression were found in spleen, testis, and muscle. Immunohistochemistry of tissue sections of metallothionein-parvalbumin transgenic strain 29 in the non-induced state revealed that ectopic PV mRNA is translated into protein. Short-term induction of the MT IIA promoter by CdSO4 or CdCl2 leads to a shift in tissue specificity and does not increase ectopic expression in tissues where the transgene is active in the noninduced state. As expected the NSE promoter showed highest activity in brain. However, NSE-driven expression could also be detected to various degrees in all investigated tissues. SV-40-dependent PV expression showed no tissue preference and varied considerably among different strains. Except for the observation that the SV-40-PV construct showed lower yields in transgenic production and reduced numbers of positive offspring no obvious impairment of growth or behavior as a consequence of transgenic PV expression could be detected.
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PMID:Production and analysis of transgenic mice with ectopic expression of parvalbumin. 753 34

We have investigated the expression of diazepam binding inhibitor (DBI) (also called acyl-CoA-binding protein or endozepine) transcripts in different human tissues and tissue culture cell lines by reverse-transcriptase assisted PCR and RNase protection assay. Two different DBI transcripts capable of encoding polypeptides of 86 and 104 amino acids were detected in all the human tissues and cell lines studied. The transcript coding for the 86 amino acid DBI polypeptide was found to represent the majority of the total DBI transcript pool.
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PMID:The characterization of two diazepam binding inhibitor (DBI) transcripts in humans. 753 63

We have investigated the subcellular organization of the four human Y RNAs. These RNAs, which are transcribed by RNA polymerase III, are usually found complexed with the Ro autoantigen, a 60-kD protein. We designed 2'-OMe oligoribonucleotides that were complementary to accessible single-stranded regions of Y RNAs within Ro RNPs and used them in fluorescence in situ hybridization. Although all four Y RNAs were primarily cytoplasmic, oligonucleotides directed against three of the RNAs hybridized to discrete structures near the nucleolar rim. We have termed these structures "perinucleolar compartments" (PNCs). Double labeling experiments with appropriate antisera revealed that PNCs are distinct from coiled bodies and fibrillar centers. Co-hybridization with a genomic DNA clone spanning the human Y1 and Y3 genes showed that PNCs are not stably associated with the transcription site for these Y RNAs. Although 5S rDNA was often located near the nucleolar periphery, PNCs are not associated with 5S gene loci. Two additional pol III transcripts, the RNA components of RNase P and RNase MRP, did colocalize within PNCs. Most interestingly, the polypyrimidine tract-binding protein hnRNP I/PTB was also concentrated in this compartment. Possible roles for this novel nuclear subdomain in macromolecular assembly and/or nucleocytoplasmic shuttling of these five pol III transcripts, along with hnRNP I/PTB, are discussed.
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PMID:A perinucleolar compartment contains several RNA polymerase III transcripts as well as the polypyrimidine tract-binding protein, hnRNP I. 753 9

The transcripts of the citrate synthase-encoding gene (gltA) in Rickettsia prowazekii (Rp), an obligate intracellular parasitic bacterium, were analyzed by RNase protection (RP), primer extension (PE) and in vitro transcription assays. Analysis of the 5' end of the gltA mRNA by RP and PE assays revealed that there were two gltA mRNAs with the 5' ends located at 16 bp and 307 bp upstream from the gltA coding region. Since these two mRNAs might represent two species of mRNA transcribed from two different promoters or a single transcript that was processed to give two mRNAs, an in vitro transcription analysis with purified Rp RNA polymerase (RNAP) was performed to distinguish these two possibilities. Purified Rp RNAP catalyzed the formation of two transcripts initiated from the same nucleotides indicated by RP and PE. Sequence analysis identified Escherichia coli (Ec) promoter-like sequences immediately upstream from both transcription start points (tsp). The first promoter (promoter P1) had the core sequence TTCTAA-N17-TATACT, was 6 bp upstream from the tsp (base A) and was centered at 37 bp upstream from the coding region. The second promoter (promoter P2) had the core sequence ATGAAA-N17-TAAAGT, was 7 bp upstream from the tsp (base T) and was centered at 329 bp upstream from the coding region. This is the first demonstration of multiple promoters in this obligate intracellular parasite which has implications concerning transcriptional regulation.
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PMID:The citrate synthase-encoding gene of Rickettsia prowazekii is controlled by two promoters. 755 59

In the Solanaceae, self-incompatibility is controlled by a single, multi-allelic ('S') locus. One product of this locus is a ribonuclease, the S-RNase, which is expressed predominantly in mature pistils and has recently been shown to cause allele-specific pollen rejection in transgenic plants. Hybrid Nicotiana plumbaginifolia x N. alata plants were used to test the effects of antisense suppression of the SA2-RNase from N. alata using three different gene constructs: two driven by RNA polymerase II-transcribed promoters, and the third, containing a truncated soybean tRNA (met-i) gene, transcribed by RNA polymerase III. All three constructs caused suppression of S-RNase activity in the transgenic plants. Unexpectedly, the CaMV 35S promoter was more effective for antisense suppression than the tissue specific tomato ChiP promoter. Antisense suppression of S-RNase correlated with low sense SA2 transcript levels and high antisense SA2 transcript levels. Untransformed hybrids that contained the N. alata SA2 allele were incompatible with N. alata SA2 pollen, while transgenic plants with suppressed SA2 gene expression accepted the pollen. The utility of this hybrid plant system for studying some aspects of antisense gene suppression is discussed.
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PMID:Antisense suppression of S-RNase expression in Nicotiana using RNA polymerase II- and III-transcribed gene constructs. 757 73

Four variant AE1 anion exchangers with predicted molecular masses of approximately 99, approximately 102, approximately 104, and approximately 108 kDa are expressed in chicken erythroid cells. These variant polypeptides differ in sequence only at the N terminus of their cytoplasmic domains. Molecular analyses have shown that transcripts derived from both of the erythroid-specific promoters, P1 and P2, encode all four of these AE1 anion exchanger variants. However, quantitative RNase protection analyses have shown that the transcripts derived from the P1 promoter are much more prevalent than those derived from the P2 promoter. Reverse transcriptase polymerase chain reaction studies have indicated that the extensive diversity in the transcripts derived from the AE1 gene occurs both in primitive and definitive lineage erythroid cells. Transient transfection analyses using human erythroleukemia cells have investigated the functional significance of the alternative sequences at the N terminus of these variant exchangers. These studies have shown that the erythroid AE1 variants are sorted to different membrane compartments in these cells. The approximately 99- and approximately 102-kDa variants are primarily sorted to the plasma membrane, whereas the approximately 108-kDa variant is retained in a perinuclear compartment. These results suggest that the alternative N-terminal cytoplasmic sequences of these polypeptides may serve as signals to direct these variant transporters to different membrane compartments within cells.
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PMID:Four variant chicken erythroid AE1 anion exchangers. Role of the alternative N-terminal sequences in intracellular targeting in transfected human erythroleukemia cells. 764 85


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