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)

The structure of the chicken adult beta-globin gene chromatin in immature and mature erythrocyte nuclei has been analysed using micrococcal nuclease digestion. The resulting DNA fragments were blotted onto DBM-papers and probed with labelled DNA fragments spanning the adult beta-globin gene and its 5'- and 3'-flanking regions. The structure of the nucleosomes within and in the regions flanking the adult beta-globin gene appears to be altered in at least two ways in erythrocyte chromatin, when compared with either bulk or inactive ovalbumin gene chromatin. First, oligomeric DNA fragments containing the beta-globin gene are released faster than those of either bulk or ovalbumin gene chromatin. Second, although the difference in size of the liberated oligomeric DNA fragments is similar to the nucleosomal repeat length of bulk and ovalbumin gene chromatin, the individual oligomers are approximately 100 bp shorter than their bulk or ovalbumin gene counterparts, most noticeably when the nuclease digestion is performed at 37 degrees C. This results in an atypical ladder of approximately 300, 500, 700, 900 bp instead of the canonical chicken erythrocyte ladder which is an integral multiple of 207 bp. The same ladder was obtained from immature erythrocytes, in which the beta-globin gene is actively transcribed, and from mature erythrocytes, in which it is considered to be inactive with RNA polymerase molecules clustered in the 5' moiety of the gene. This indicates that the alteration of the nucleosomal structure is not due to transcription per se.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Digestion of the chicken beta-globin gene chromatin with micrococcal nuclease reveals the presence of an altered nucleosomal array characterized by an atypical ladder of DNA fragments. 301

To investigate the chromatin surrounding an active gene, we have determined the distribution of RNA polymerase molecules, the intactness of nucleosomal structure, and the subnuclear compartmentalization along 15 kilobase pairs (kb) of the mouse kappa immunoglobulin locus of MPC-11 plasmacytoma cells. Hybridization of in vitro nuclear transcripts to probes specific for the template strand reveals that transcription terminates within the region between 1.1 and 2.3 kb downstream from the poly(A) addition site. Ten different short sequences (8-13 base pairs) reside within 460 base pairs of this termination region that exhibit homology with sequences found in the termination regions of mouse beta-globin and chicken ovalbumin genes. Transcription of the nontemplate strand occurs on either side of this termination region. We find that both within the transcription unit and 6.5 kb downstream of the termination region of the kappa gene, the canonical nucleosomal structure is perturbed, the chromatin exhibits pronounced insolubility, and the nucleosomes liberated by micrococcal nuclease appear to lack histone H1. The insolubility is characterized by interactions that are disrupted by 0.3 to 0.6 M NaCl treatment. We conclude that the active chromatin phenotype spreads a considerable distance along the kappa locus, well beyond the region of transcription termination.
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PMID:Transcription termination and chromatin structure of the active immunoglobulin kappa gene locus. 308 10

The N gene protein of bacteriophage lambda prevents termination of transcription by E. coli RNA polymerase. We describe here the conditions of a cell-free reaction system in which pure N stimulates net transcription up to tenfold and therefore nearly stoichiometrically modifies transcribing RNA polymerase molecules. The reaction contains micrococcal nuclease-treated S100 extract derived from E. coli and a plasmid template DNA containing the lambda early promoter PL, the N utilization site nutL, and the Rho-dependent terminator tL1. Stimulation by N in this system is specific and biologically relevant since it is absent with vector pBR322 DNA and with extracts derived from E. coli strains bearing the nusA1 and nusE71 mutations known to block N function in vivo. We use the system to provide further evidence that ribosomes are not necessary for N function and to demonstrate the direct involvement in N function of the NusA protein of E. coli.
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PMID:Efficient modification of E. coli RNA polymerase in vitro by the N gene transcription antitermination protein of bacteriophage lambda. 315 83

Light treatment of nuclei of Physarum polycephalum microplasmodia with DNase I, at low MgCl2 concentration (less than or equal to 3% DNA acid solubility, 0.1 mM MgCl2) selectively solubilizes a defined fraction of chromatin, in the form of a macromolecular complex. This fraction (up to 15% of the total chromatin) contains a full complement of the core histones and a reduced amount of histone H1, and is enriched in the high-mobility-group type of proteins. It is preferentially associated with nascent RNA and RNA polymerase B actively engaged in transcription. Digestion of DNAase-I-solubilized chromatin by micrococcal nuclease releases a size-heterogeneous population of cleavage products, indicative of lack of a typical nucleosomal packaging. It is concluded that the procedure used allows the isolation of structurally and functionally distinct regions of Physarum chromatin.
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PMID:Lack of nucleosomal structure in a DNase-I-solubilized transcriptionally active chromatin fraction of Physarum polycephalum. 397 88

The reversible effect of dietary methionine deficiency was studied in young adult rats. The sensitivity of nuclear chromatin to micrococcal nuclease (EC3.1.4.7) digestion and the composition of the chromatin proteins were unaffected by the dietary regimens. The specific chromatin-bound RNA polymerase II activity decreased during methionine deficiency. Refeeding of methionine for 2 days restored the activity in the nuclease-released chromatin. RNA polymerase I plus III activity remained unchanged. Total RNA polymerase activity changed with the liver wet weight which was reduced during methionine deficiency and was not restored to control level after 2 days of methionine refeeding. RNA polymerase activity was altered by methionine deficiency. The recovery was independent of major modifications of the chromatin structure and protein composition.
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PMID:RNA polymerase activities and chromatin protein composition of rat liver during methionine deprivation and refeeding. 400 43

Extremely mild treatment with micrococcal nuclease of isolated nuclei yields subnuclear fractions in which the majority of RNA polymerase II transcriptional complexes formed in vivo are segregated [Tata & Baker (1978) J. Mol. Biol. 118, 249-272]. We now describe different approaches followed to established whether or not the nuclei are thus resolved into transcribed and non-transcribed DNA. First, we have compared the sensitivity to deoxyribonuclease I, which is known to digest preferably expressed genes as present in nuclei or chromatin, of three micrococcal-nuclease-derived fractions from nuclei of different transcriptional activities. In transcriptionally active nuclei (rat liver, hen liver and oviduct, and Xenopus liver), the DNA in a polynucleosomal fraction comprising 6-15% of DNA and the majority of template-engaged RNA polymerase II (fraction P2) was 10-50 times as sensitive to deoxyribonuclease I as the DNA in the other two fractions (fractions P1 and S, comprising 78-88% of total nuclear DNA as large polynucleosomal aggregates and 2-6% of DNA mostly as mononucleosomes, respectively). In transcriptionally inactive nuclei obtained from hen erythrocytes, micrococcal nuclease did not separate DNA into fractions exhibiting such differential sensitivities. Second, we have monitored the partition of an expressed gene. Hybridization of complementary DNA to Xenopus albumin mRNA revealed a 5-10-fold enrichment of the albumin (but not the globin) gene in the P2 fraction of nuclei from Xenopus liver in which this gene is fully expressed. Third, a large part of the nascent rapidly labelled RNA synthesized in vivo in rat liver nuclei was recovered in the micrococcal-nuclease-derived fraction that is more susceptible to digestion with deoxyribonuclease I. It is concluded that mild micrococcal-nuclease treatment of nuclei causes their separation into transcribed and non-transcribed DNA as determined by a number of very different criteria.
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PMID:Subnuclear fractionation by mild micrococcal-nuclease treatment of nuclei of different transcriptional activities causes a partition of expressed and non-expressed genes. 615 73

Nuclei isolated from uninfected HEp-2 cells synthesized RNA for 60 to 90 min. The individual RNA polymerase activities were determined by alpha-amanitin differential inhibition and the RNA products characterized by electron microscope (EM) autoradiography and sucrose gradient centrifugation. In nuclei prepared from poliovirus-infected cells, the capacity to synthesize RNA in vitro decreased with time after infection. RNA polymerase II activity (hnRNA synthesis) was preferentially inhibited more than was the polymerase I activity (rRNA synthesis). Poliovirus-infected cytoplasm (S-30) inhibited in vitro RNA synthesis in uninfected nuclei by selectively affecting the polymerase II activity. Selective inhibition of hnRNA synthesis by the crude extracts could be monitored by EM autoradiography directly. Determinations of individual RNA polymerase activities by differential alpha-amanitin inhibition were done only after treatment of the infected cytoplasm with micrococcal nuclease to abolish virus RNA replication. Selective inhibition of hnRNA synthesis depended on preincubation of the nuclei together with the infected cytoplasm, indicating that inhibitory substances from the infected cytoplasm entered the nuclei. Isolated nuclei therefore provide a useful system for studying the nature of the inhibitor(s) and of host RNA synthesis inhibition by picornaviruses.
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PMID:Poliovirus-induced inhibition of host RNA synthesis studied in isolated HEp-2 cell nuclei. 618 46

The effect of dimethylnitrosamine on functional activities of liver chromatin was studied in mice. After a single dose of dimethylnitrosamine injected i.v. (25 mg/kg body wt, 45 min before sacrifice) liver nuclei were isolated and incubated with micrococcal nuclease (EC 3.1.4.7) to an acid-solubility of 2.5% of total DNA. Chromatin was fractionated into a 1,200 g pellet P1, 102,000 g pellet P2 and supernatant fraction S2. Chromatin-bound RNA polymerase I plus III activity decreased 15% in the P1 and 25% in the P2 fraction. No changes in activity were observed in the S2 fraction. Chromatin-bound RNA polymerase II activity decreased 19% in the P1, 49% in the P2 and 32% in the S2 fraction. Heparin stimulated RNA polymerase II activity decreased 10% in the P1 and 44% in the P2 fraction. Formation of initiation in nuclear lysates with RNA polymerase from Escherichia coli increased after administration of dimethylnitrosamine suggesting an increase in the number of sites available for the start of new RNA chains. The results show that limited digestion of nuclei with endonuclease cleaves chromatin regions which are more affected by dimethylnitrosamine than the total chromatin suggesting a non-random effect of the hepatotoxin on chromatin. Modifications of the DNA template by dimethylnitrosamine is indicated by the change in number of initiation complexes.
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PMID:Non-random effect on RNA synthesis in liver chromatin by administration of dimethylnitrosamine to mice. 619 51

The administration of thyroxine to neonatal rats stimulates RNA synthesis by neuronal nuclei isolated from the developing rat brain cortex. Glial nuclei are relatively resistant to thyroxine treatment. The activity of neuronal RNA polymerase II is particularly stimulated by the hormone. Thyroxine also affects neuronal chromatin structure as shown by changes in the relative proportion of different subnuclear fractions obtained by gentle micrococcal nuclease digestion of nuclei from hormone-treated rats.
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PMID:Thyroxine preferentially stimulates transcription by isolated neuronal nuclei in the developing rat brain cortex. 620 58

Electron microscope spread preparations of nuclear chromatin derived from early S-phase of Physarum reveal 'beads on a string' for nonreplicated and a portion of newly replicated chromatin. Many of the early replicons contain transcription units as visualized by nascent transcripts. They are, in most cases, arranged in continuous length gradients on both newly replicated strands of chromatin, the putative origin of replication being within the transcription unit. Preferential release of DNA as acid precipitable material by DNAse I and of RNA polymerase B (estimated as release of labeled alpha-amanitin bound to isolated nuclei) is observed in early S-phase, but only if DNA synthesis is not inhibited. Also, generation of a small particle (peak A) by staphylococcal nuclease, characteristic of transcriptionally active chromatin, depends on concomitant replication of early replicons. It is concluded that DNA replication is a prerequisite for its transcription by RNA polymerase B. Thus, the sequential replication of the genome of Physarum dictates the order of transcription during S-phase which may in part control the proliferative mitotic cycle of Physarum.
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PMID:Physical relationship between replicons and transcription units in Physarum polycephalum. 621 73


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