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 binding of core histones (H2A, H2B, H3, H4) to a circular plasmid DNA and to a circular DNA-RNA hybrid molecule of similar size has been compared. Circular hybrid molecules were formed from single stranded fd DNA by synthesis of the complimentary strand with ribonucleotides using wheat germ RNA polymerase II. Upon reconstitution of plasmid DNA circles with histone, the sedimentation profiles of the DNA remained sharp by increased several fold in rate. Material from the peak fractions of these sedimentations appeared to be condensed circular loops of nucleosomes when examined by electron microscopy (EM), and the mass ratio of DNA to histone (at the histone concentrations which produced the fastest sedimentations) was typical of native chromatin. In contrast, the sedimentation behavior of DNA-RNA hybrid circles after addition of histone remained unchanged except for a minor fraction which exhibited a broad and faster sedimentation rate. Examination by EM revealed that most of the molecules appeared identical to protein free hybrid circles while the minor, faster sedimenting fraction appeared to be two or more circles bound together by protein aggregates. Finally, a linear molecule consisting of about 3000 base pairs of duplex DNA covalently joined on both ends to 1500 base pairs of RNA-DNA hybrid helix was constructed. Reconstitution of this molecule with core histone showed nucleosome formation only on the central DNA duplex region. Isopycnic banding of fixed hybrid-histone mixtures showed that little or no histone had bound to the bulk of the full hybrid molecules. We suggest that the presence of RNA in a nucleic acid duplex inhibits the condensation of the duplex into a nucleosomal structure by histone.
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PMID:The presence of RNA in a double helix inhibits its interaction with histone protein. 616 Apr 77

The regulatory effects of host cellular histones on the transcription of simian virus 40 (SV40) DNA were investigated by using reconstituted and native SV40 nucleoprotein complexes (NPCs). Reconstituted NPCs were prepared from SV40 DNA and the combination fraction of five histones, H1, H2A, H2B, H3, and H4, isolated from the nuclei of permissive (CV-1) or nonpermissive (BALB/c 3T3, rat liver, and calf thymus) cells. Native NPCs were prepared by alkali disruption of purified SV40 virions. Nuclease digestion of these NPCs gave regular patterns of bands similar to those of SV40 NPCs from SV40-infected CV-1 cells, suggesting the presence of a nucleosomal structure. Transcription of NPCs was analyzed in vitro by using Escherichia coli DNA-dependent RNA polymerase. Both histone H1 and the fraction consisting of all five histones inhibited transcription of SV40 DNA by about 90 to 95%. The fraction consisting of four histones lacking H1 reduced the transcription by 30 to 35%, to a level similar to that of transcription with native NPCs. Transcription was inhibited regardless of whether the origin of histones was permissive or nonpermissive cells. Gel electrophoretic patterns of RNA products transcribed from SV40 DNA and reconstituted and native NPCs showed several identical peaks between 13S and 28S. The patterns were identical whether NPCs reconstituted with H1 alone, all five histones, or four histones lacking H1 were used.
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PMID:Transcription of reconstituted simian virus 40 nucleoprotein complexes. 624 6

A 2.56-kbp fragment containing genes coding for histones H2A and H3 that forms a portion of the 10.2-kbp cluster containing all five histone genes isolated from a lambda-Charon 4A library of rainbow trout genomic DNA has been characterized in detail and its complete nucleotide sequence determined. The genes are arranged in tandem, being encoded on the same DNA strand. They are separated by 380 bp of intergenic spacer DNA that contains an alternating purine-pyrimidine stretch of 20 bp and a 46-bp stretch that has the potential of forming a triple cruciform structure. The histone genes contain no introns, have the RNA polymerase II promoter-associated signals known as CAAT and TATA boxes in their 5' flanking regions and contain a conserved inverted repeat sequence, similar to that found in histone genes of other species, capable of forming a hairpin structure at the 3' end of the transcription unit.
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PMID:Organization and nucleotide sequence of rainbow trout histone H2A and H3 genes. 643 79

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

Purified RNA polymerase II from calf thymus can bind to about 15% of the nucleosome core particles prepared from mouse myeloma cells, forming a discrete complex having a sedimentation coefficient of 18S. These bound nucleosome cores are heavily enriched in transcribed DNA sequences, are deficient in histones H2A and H2B, and undergo a reversible change in structure on RNA polymerase II binding.
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PMID:Eukaryotic RNA polymerase II binds to nucleosome cores from transcribed genes. 682 27

Sea urchin (psammechinus miliaris) H2A histone genes shown to be promoter mutants from oocyte injection experiments were tested for their ability to initiate transcription in vitro. Circular templates were transcribed with HeLa cell extracts, and the transcripts were assayed by mung bean or S1 nuclease mapping of the 5' ends. The transcripts of the H2A mutants produced in vitro were qualitatively similar and, in most cases, identical to those seen in oocyte injection experiments, but quite large quantitative differences were observed for some H2A mutant genes. Both the T-A-T-A box and far upstream sequences residing in the modulator segment E [Grosschedl, R. & Birnstiel, M. L. (1980) Proc. Natl. Acad. Sci. USA 77, 7102--7106] were found to be essential for maximal transcription in vitro. Deletion of either of these sequence elements reduced transcription to 20%. A similar reduction in the amount of H2a transcripts was found when a T-A-T-A-to-T-A-G-A point mutant was tested in vitro. Essential far upstream sequences were mapped between nucleotides -139 and -111, 5' to the initiation site of transcription. In the standard run-off transcription test using restriction fragments, the effects of these sequences could be mimicked by free DNA ends, suggesting that the function of this in vitro upstream sequence might be to provide an entry side for RNA polymerase II.
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PMID:Delimitation of far upstream sequences required for maximal in vitro transcription of an H2A histone gene. 695 85

A histone-like protein (H) from Escherichia coli has been purified to more than 98% homogeneity by using its capacity to inhibit DNA functions. H protein behaves as a dimer of 28,000-dalton subunits. The histone H2A-like properties of H protein are: (i) binding to DNA at a stoichiometry of 1 H protein dimer per 75 bases; (ii) abundance of about 30,000 molecules per cell, sufficient to bind about 20% of the chromosome; (iii) limiting digestion of double-stranded DNA by micrococcal nuclease; (iv) reannealing of complementary single-stranded DNA; (v) amino acid composition resembling that of eukaryotic histone H2A; (vi) neutralization of H protein by antibody specific for H2A; (vii) heat stability; and (viii) acid solubility. The capacity of H protein to bind DNA prevents its template or substrate functions n several reactions in vitro: DNA synthesis by several polymerases; transcription by RNA polymerase; DNA topoisomerase activity; and DNA-dependent ATP hydrolysis by rep protein, dnaB protein, or protein n'. Together with other histone-like proteins of E. coli, H protein may organize the E. coli chromosome into nucleosomes, such as in eukaryotic chromatin.
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PMID:Novel histone H2A-like protein of escherichia coli. 700 71

Direct chemical acetylation of an oligonucleosomal template for bacteriophage T7 RNA polymerase is accompanied by a substantial increase in its capability to support RNA synthesis. The template was assembled from a plasmid, containing a promoter and a terminator for T7 RNA polymerase, plus one (H3-H4)2 tetramer and two H2A.H2B dimers for each 200 base pairs of DNA. Under the employed conditions, acetylation modifies in a preferential way the lysine residues located in the amino-terminal domains of core histones. When the template is assembled with acetylated tetramers and untreated dimers, its efficiency in promoting RNA synthesis is also largely increased. Since a previous work reported transcriptional stimulation upon acetylation of H2A.H2B dimers [Puerta et al. (1995) Biochem. Biophys. Res. Commun. 210, 409], the transcriptional repression brought about by core histone octamers seems to require that the amino-terminal domains of both (H3.H4)2 tetramers and H2A.H2B dimers are not acetylated.
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PMID:Transcriptional properties of oligonucleosomal templates containing acetylated (H3-H4)2 tetramers. 763 40

Histone-DNA templates for bacteriophage T7 RNA polymerase were assembled from a plasmid containing a promoter and a terminator for T7 RNA polymerase, intact (H3.H4)2 tetramers, and either untreated or chemically acetylated H2A.H2B dimers. The nucleosomal particles containing acetylated H2A.H2B dimers protect 145 base pairs of DNA against micrococcal nuclease digestion and prevent the reaction with psoralen of 80 to 145 DNA base pairs. The inhibition of transcriptional initiation caused by the association of DNA with intact core histone octamers decreases significantly when the histone octamers contain acetylated H2A.H2B dimers. These results suggest a role for H2A.H2B dimers in the control of transcription, which might be mediated through acetylation and deacetylation of their lysine residues.
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PMID:Acetylation of histone H2A.H2B dimers facilitates transcription. 775 16

Histone octamers or histone H3/H4 tetramers were reconstituted onto either closed circular plasmids containing a single Xenopus 5S rRNA gene or a reiterated array of Lytechinus 5S rRNA genes. All "reconstitutes" were found to undergo both Na(+)-dependent and Mg(2+)-dependent compaction. However, in each case, the compaction of nucleosomal templates containing H2A/H2B was much more extensive than compaction of templates containing only H3/H4 tetramers. Inclusion of 5 mM MgCl2 in the transcription buffer increased the level of compaction of nucleosomal templates and led to a marked inhibition of both transcription initiation and elongation by RNA polymerase III. The inhibitory effect of Mg2+ was reduced significantly when DNA templates contained only H3/H4 tetramers, consistent with their lesser extent of Mg(2+)-dependent compaction. Thus, the removal of histones H2A/H2B from nucleosomal arrays enhances gene activity, in part because of decreased levels of chromatin folding.
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PMID:A role for histones H2A/H2B in chromatin folding and transcriptional repression. 813 97


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