Nucleosome core particles suppress the thermal untwisting of core DNA and adjacent linker DNA.

Morse, Randall H.; Cantor, Charles R.

doi:10.1073/pnas.82.14.4653

Cited by 50 publications

(12 citation statements)

References 35 publications

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“…Formation of a cruciform by (AT)24 requires a superhelix density (number of supercoils per base pair) of -0.033 within the AT tract (40) in addition to a twist change (number of supercoils introduced) of -4 (45). Furthermore, it is likely that the supercoiling detected by cruciform extrusion must be transmitted through one or more nucleosomes, and it is not yet clear from available evidence to what extent such propogation can occur (21,49,50). An attractive idea is that a site-specific DNA gyrase-like enzyme continually generates the supercoiling observed, perhaps as a consequence of template attachment to the nuclear matrix (37,51,59), which contains topoisomerase II as a major component (14,15,22).…”

Section: Discussionmentioning

confidence: 99%

Evidence for torsional stress in transcriptionally activated chromatin.

Leonard

Patient

1991

Mol. Cell. Biol.

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The existence of torsional stress in eukaryotic chromatin has been controversial. To determine whether it could be detected, we probed the structure of an alternating AT tract. These sequences adopt cruciform geometry when the DNA helix is torsionally strained by negative supercoiling. The single-strand-specific nuclease P1 was used to determine the structure of an alternating AT sequence upstream of the Xenopus P-globin gene when assembled into chromatin in microinjected Xenopus oocytes. The pattern of cleavage by P1 nuclease strongly suggests that the DNA in this chromatin template is under torsional stress. The cruciform was detected specifically in the most fully reconstituted templates at later stages of chromatin assembly, suggesting that negative supercoiling is associated with chromatin maturation. Furthermore, the number of torsionally strained templates increased dramatically at the time when transcription of assembled chromatin templates began. Transcription itself has been shown to induce supercoiling, but the requisite negative supercoiling for cruciform extrusion by (AT). in oocytes was not generated in this way since the characteristic P1 cutting pattern was retained even when RNA polymerase elongation was blocked with a-amanitin. Thus, torsional stress is associated with transcriptional activation of chromatin templates in the absence of ongoing transcription.A role for DNA supercoiling in gene activation is suggested by the observations that initiation of transcription is accompanied by local unwinding of DNA (71) and that such strand separation is facilitated by negative DNA supercoiling (reviewed in reference 39). Furthermore, both the formation of stable initiation complexes and the number of initiated transcripts have been shown to be greater on negatively supercoiled than on relaxed templates in vitro (64). However, although superhelicity has been shown to influence transcription of a number of prokaryotic genes in vivo (62), the situation in eukaryotes is much less clear.Both prokaryotic (88) and eukaryotic (59) chromosomes appear to be organized as a series of independently supercoiled loops or domains. In prokaryotes, the supercoiling is only partially constrained by association with protein (11), so that partitioning between the writhing and twisting of the DNA helix can still take place. Two sources of supercoiling in bacteria are DNA gyrase, which generates negative supercoils (23), and the process of transcription, which introduces both negative and positive supercoils (90). The overall level of unconstrained supercoiling appears to be the result of the combined effects of transcription, DNA gyrase, and the relaxing enzyme DNA topoisomerase I (63,65,78 into chromatin in a site-specific manner. However, there are a number of other potential sources of local supercoiling in eukaryotic chromatin, including the breakdown of higherorder structure (57), the loss of nucleosomes from nucleasehypersensitive sites (32, 69), unfolding of the nucleosome itself (13, 61), histone acetylation (54),...

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Section: Discussionmentioning

confidence: 99%

Evidence for torsional stress in transcriptionally activated chromatin.

Leonard

Patient

1991

Mol. Cell. Biol.

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“…The degree of constraint imposed by chromatin upon DNA has been studied previously by analysis of temperatureinduced topological changes in the DNA of both native (5,6,11,12) and reconstituted (24,25) chromatin. This study carries out such an analysis on transcriptionally active chromatin-i.e., chromatin containing in vivo-initiated RNA polymerases that are capable of transcription run-on in vitro.…”

Section: Discussionmentioning

confidence: 99%

Thermal unwinding of simian virus 40 transcription complex DNA.

Lutter

1989

Proc. Natl. Acad. Sci. U.S.A.

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Two long-standing questions in the control of eukaryotic gene expression have been how the structure of transcribing chromatin compares with that of nontranscribing chromatin and how chromatin structure differs among various eukaryotic organisms. We have addressed aspects of these two questions by characterizing the rotational flexibility of the DNA of the simian virus 40 (SV40) transcription complex. When transcription complex samples are incubated with topoisomerase at 0C or 37C, the DNA of the 37C sample is unwound by 1.8 turns relative to that of the 0C sample. This amount of unwinding is similar to that observed for bulk, untranscribed SV40 minichromosome DNA, indicating that the chromatin structure of a transcribed gene resembles that of a nontranscribed gene in the degree of constraint that it imposes on its DNA. However, this amount of unwinding differs substantially from the value observed for yeast plasmid chromatin DNA, suggesting that yeast chromatin differs significantly from mammalian chromatin in this fundamental property.Substantial progress has been made in describing structural details of bulk eukaryotic chromatin, but as yet little is known about the structural details of that minor fraction of chromatin that is transcribed (1-3). The work of Weintraub and Groudine (4) demonstrates that some structural difference exists because the enzyme DNase I digests the DNA of transcriptionally active chromatin more rapidly than that of transcriptionally inactive chromatin. This indicates that active chromatin has a more "open" or accessible structure, suggesting that the DNA-histone interactions may perhaps be less stable than those of inactive chromatin.Recent results from studies of yeast plasmid chromatin (5, 6) would seen to support this view. The topology of the DNA of yeast plasmid chromatin was found to change with temperature in an amount that is --70% the value exhibited by bare DNA. This substantial rotational flexibility of the yeast chromatin DNA differs markedly from that of the DNA of the simian virus 40 (SV40) minichromosome (7,8), which has been shown to unwind by only 25% [or =--1.5 turns per 5243 base pairs (bp)] of the value for bare DNA. However, the bulk chromatin of SV40 is predominantly inactive transcriptionally: only ==1% of the SV40 minichromosomes in an infected cell appear to be transcriptionally active (9). Yeast 2-,um plasmid chromatin, on the other hand, is thought for the most part to be transcriptionally active. Noting these functional differences, Saavedra and Huberman (5) have suggested that the relatively flexible structure that they observe in the yeast 2-,um minichromosome may be a fundamental feature of the transcriptionally active subclass of eukaryotic chromatin. By this proposal, the transcribed fraction of mammalian chromatin would be expected to exhibit the enhanced DNA flexibility characteristic of the yeast chromatin.We have carried out experiments here that directly test this proposal by analyzing the topological properties of that fraction of SV40 m...

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“…autoradiographs were measured by densitometric scanning using the Apple Color OneScanner and analyzed with NIH Image. version 1.49. [The center of a given distribution is defined as the midpoint of integrated density along the length of the densitometric scan (see Morse and Cantor. 1985).…”

Section: Plasmid Isolation and Topology Analysismentioning

confidence: 99%

All four core histone N-termini contain sequences required for the repression of basal transcription in yeast.

et al. 1996

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Nucleosomes prevent the recognition of TATA promoter elements by the basal transcriptional machinery in the absence of induction. However, while Saccharomyces cerevisiae histones H3 and H4 contain N‐terminal regions involved in the activation and repression of GAL1 and in the expression of heterochromatin‐like regions, the sequences involved in repressing basal transcription have not yet been identified. Here, we describe the mapping of new N‐terminal domains, in all four core histones (H2A, H2B, H3 and H4), required for the repression of basal, uninduced transcription. Basal transcription was monitored by the use of a GAL1 promoter‐URA3 reporter construct whose uninduced activity can be detected through cellular sensitivity to the drug, 5‐fluoroorotic acid. We have found for each histone that the N‐terminal sequences repressing basal activity are in a short region adjacent to the structured alpha‐helical core. Analysis of minichromosome DNA topology demonstrates that the basal domains are required for the proper folding of DNA around the chromosomal particle. Deletion of the basal domain at each histone significantly decreases plasmid superhelical density, which probably reflects a release of DNA from the constraints of the nucleosome into the linker region. This provides a means by which basal factors may recognize otherwise repressed regulatory elements.

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Nucleosome core particles suppress the thermal untwisting of core DNA and adjacent linker DNA.

Cited by 50 publications

References 35 publications

Evidence for torsional stress in transcriptionally activated chromatin.

Evidence for torsional stress in transcriptionally activated chromatin.

Thermal unwinding of simian virus 40 transcription complex DNA.

All four core histone N-termini contain sequences required for the repression of basal transcription in yeast.

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