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)

Several DNA-histone complexes were reconstituted in the presence and absence of urea. The fiber size of DNA-histone H1 complex was about 20 A in width with knobs 100 to 250 A in diameter interspersed at an average interval of about 1,100 A. H1-was associated with DNA segments corresponding to a DNA size of fewer than 100 base pairs. DNA-histones H2A, H2B, H3 and H4 complex consisted of globular subunits 100 to 150 A in diameter alternating with thin strands, like beads on a string. DNA-whole histones complex was 200 to 250 A in width and had a condensed configuration. The nuclease digestion pattern of the complexes containing histones H2A, H2B, H3 and H4 was regular, similar to that of chromatin, and was disrupted by urea. The complex containing H1 was inactive for in vitro RNA synthesis by escherichia coli RNA polymerase, whereas the other complexes were active. The complexes reconstituted in the absence of urea had template activities slightly less than in the presence of urea.
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PMID:Finestructure and template activities of DNA-histone complexes reconstituted in the presence and absence of urea. 644 34

The properties of active and repressed 5S rna genes in somatic cell chromatin from Xenopus laevis cultured cells were studied by transcription in vitro. The somatic 5S RNA genes, which are active in vivo, are packaged in chromatin as active stable transcription complexes lacking only RNA polymerase III for transcription activity. The oocyte 5S RNA genes, which are inactive in somatic tissues, do not have transcription factors bound to them in purified chromatin and are prevented from binding these factors by a structure dependent on histone H1. In chromatin active stable transcription complexes protect 5S RNA genes from histone H1-mediated repression and H1 binding prevents the formation of stable active transcription complexes.
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PMID:The transcriptional regulation of Xenopus 5s RNA genes in chromatin: the roles of active stable transcription complexes and histone H1. 654 Jan 47

Inactive Drosophila heat-shock genes of isolated diploid nuclei can be induced to a transcriptionally active state by exposure to cytoplasmic extracts from heat-shocked Drosophila cells. No effect was observed on histone gene transcription, and extracts from non-heat-shocked cells were ineffective. The factor in the cytoplasmic extract has been partially purified and characterized. It is protease-sensitive and heat-labile. A striking change accompanies in vitro activation that permits transcription by E. coli RNA polymerase of the chromatin 5' -distal to the structural genes at the 87A and 87C heat-shock gene loci; we have previously observed a similar change after in vivo heat-shock induction. That this change occurred in the absence of endogenous RNA polymerase II activity suggests that these changes may reflect the earliest event in gene activation. Inasmuch as activation also took place after histone H1 depletion, this histone does not appear to be essential for this step of gene activation.
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PMID:Activation of the major drosophila heat-shock genes in vitro. 679 38

By means of indirect immunofluorescence microscopy, we have studied the distribution of RNA polymerase B, of the nucleosomal histones H2b, H3, and H4 and of histone H1, in nuclei of primary spermatocytes of Drosophila hydei. RNA polymerase B and histones, including H1, are found to be present on the loop structures of the Y chromosome. The nucleolus stains only for the histones, but not for RNA polymerase B. Various mutants deficient for some of the loops or altering their morphology, were used to identify the individual chromosomal segments. In growing spermatocytes of the genetic constitution X/0, autosomes and the chromosome X react strongly with antibodies against RNA polymerase B, but not with antibodies against histones. The results suggest that the autosomes, the chromosome X and the Y chromosomal loop structures, with the exception of the nucleolus, are transcribed mostly by RNA polymerase B.
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PMID:Localization of RNA polymerase B and histones in the nucleus of primary spermatocytes of Drosophila hydei, studied by immunofluorescence microscopy. 722 44

We describe the preparation and use of a nuclear extract derived from Drosophila embryos that is highly active for transcription in vitro by RNA polymerase This extract, which is termed the soluble nuclear fraction (SNF), can support multiple rounds of transcription and generate about 0.45 transcripts per template per 30 min. Furthermore, the SNF is deficient in nonspecific DNA binding factors that inhibit transcription, such as histone H1, and can be used for the analysis of transcriptional regulation by promoter- and enhancer-binding factors with either naked DNA or chromatin templates.
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PMID:The soluble nuclear fraction, a highly efficient transcription extract from Drosophila embryos. 770 54

The effect of histone H1 on transcription by bacteriophage T7 RNA polymerase was examined using reconstituted chromatin templates. A 3.8 kb linear DNA template consisting of a specific transcription promoter for T7 RNA polymerase placed upstream of 18 tandem repeats of a 207 bp nucleosome positioning sequence derived from the 5S rRNA gene of Lytechinus variegatus was used as a template for chromatin reconstitution. Regularly spaced arrays of nucleosome cores were assembled onto this DNA template from donor histone octamers by salt step dialysis. Histone H1 was incorporated onto free DNA or reconstituted chromatin templates and double label transcription assays were performed. The experiments indicated that histone H1 has a strong inhibitory effect on both transcription initiation and elongation. These effects are especially pronounced on chromatin templates, where both transcription initiation and elongation are virtually halted. The inhibition of transcription elongation appears to result from a dramatic increase in premature termination of transcripts. These experiments indicate that assembly of histone H1 into chromatin can result in structures which are completely repressed with respect to transcription.
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PMID:Deposition of histone H1 onto reconstituted nucleosome arrays inhibits both initiation and elongation of transcripts by T7 RNA polymerase. 773 95

Transcription of 5S rRNA and tRNA genes by RNA polymerase III (pol III) in cytosolic extracts of unfertilized Xenopus eggs and in a reconstituted system derived from Xenopus oocytes is repressed by the action of one or more mitotic protein kinases. Repression is due to the phosphorylation of a component of the pol III transcription apparatus. We find that the maturation/mitosis-promoting factor kinase (MPF, p34cdc2-cyclin B) can directly mediate this repression in vitro. Affinity-purified MPF and immune complexes formed with antibodies to the protein subunits of MPF (p34cdc2 and cyclin B) retain both histone H1 kinase activity and the capacity to repress transcription in the reconstituted transcription system. Transcription complexes of oocyte-type 5S RNA genes and tRNA genes are quantitatively more sensitive to MPF repression than the corresponding transcription complexes of the somatic-type 5S RNA gene. The differential transcription of oocyte- and somatic-type genes observed during early Xenopus embryogenesis has been reproduced with the reconstituted transcription system and affinity-purified MPF. This differential transcription may be due to the instability of transcription complexes on the oocyte-type genes and the heightened sensitivity of soluble transcription factors to inactivation by mitotic phosphorylation. Our results suggest that MPF may play a role in vivo in the establishment of the embryonic pattern of pol III gene expression.
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PMID:Role of maturation-promoting factor (p34cdc2-cyclin B) in differential expression of the Xenopus oocyte and somatic-type 5S RNA genes. 800 72

We have isolated from a crude Hela cell cofactor fraction (USA) a novel positive cofactor that cooperates with the general transcription machinery to effect efficient stimulation of transcription by GAL4-AH, a derivative of the Saccharomyces cerevisiae regulatory factor GAL4. PC2 was shown to be a 500-kDa protein complex and to be functionally and biochemically distinct from native TFIID and previously identified cofactors. In the presence of native TFIID and other general factors, PC2 was necessary and sufficient for activation by GAL4-AH. Cofactor function was specific for transcriptional activation domains of GAL4-AH. The repressor histone H1 further potentiated but was not required for activation of transcription by GAL4-AH. On the basis of the observation that PC2 exerts entirely positive effects on transcription, we propose a model in which PC2 increases the activity of the preinitiation complex in the presence of an activator, thereby establishing a specific pathway during activation of RNA polymerase II.
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PMID:RNA polymerase II cofactor PC2 facilitates activation of transcription by GAL4-AH in vitro. 819 33

Chromatin reconstituted in an extract from preblastoderm Drosophila embryos represses transcription by RNA polymerase II. We have assembled regularly spaced nucleosomes on DNA attached to paramagnetic beads enabling the efficient purification of chromatin templates for transcription studies. We have used diagnostic salt extractions to establish that transcriptional repression of immobilized chromatin was largely due to nucleosome cores. When purified H1 was incorporated into chromatin, resulting in increased repeat lengths to 200-220 bp, the contribution of H1 to transcriptional repression was negligible. If more H1 was added no regularly spaced chromatin was obtained and only under these conditions was transcriptional inhibition by H1 apparent. We conclude that efficient repression of transcription by polymerase II in this system does not require the presence of histone H1.
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PMID:Transcriptional repression by nucleosomes but not H1 in reconstituted preblastoderm Drosophila chromatin. 831 82

An experimental assay was developed to search for proteins capable of antagonizing histone H1-mediated general repression of transcription. T7 RNA polymerase templates containing an upstream scaffold-associated region (SAR) were highly selectively repressed by H1 relative to non-SAR control templates. This is due to the nucleation of H1 assembly into flanking DNA brought about by the numerous A-tracts (AT-rich sequences containing short homopolymeric runs of dA.dT base pairs) of the SAR. Partial, selective titration of these A-tracts by the high mobility group (HMG) protein HMG-I/Y led to the complete derepression of transcription from the SAR template by inducing the redistribution of H1 on to non-SAR templates. SARs are associated with many highly transcribed regulated genes where they may serve to facilitate the HMG-I/Y-mediated displacement of histone H1 in chromatin. Indeed, HMG-I/Y was found to be strongly enriched in the H1-depleted subfraction which can be isolated from chromatin.
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PMID:SAR-dependent mobilization of histone H1 by HMG-I/Y in vitro: HMG-I/Y is enriched in H1-depleted chromatin. 834 61


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