Isolation of single stranded DNA related to the transcriptional activity of animal cells

Tapiero, Haïm; Leibowitch, S.A.; Shaool, D.; Monier, Marie‐Noëlle; Harel, J

doi:10.1093/nar/3.4.953

Cited by 19 publications

(16 citation statements)

References 16 publications

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“…We recently obtained evidence that suggested other mechanisms might be involved. The "major part of ss-DNA could not consist of replication intermediates because its lifespan was similar to that of bulk DNA (10,33). Another possibility that had to be considered is that ss-DNA is related to RNA synthesis.…”

Section: Discussionmentioning

confidence: 99%

“…We have isolated, from the nDNA of various species, a minor fraction of single-stranded DNA (ss-DNA) and demonstrated that about one-third of the ss-DNA from cultured normal embryonic chicken cells (10) or human RD cells (11) Isolation of ss-DNA. The modification of the method for hydroxyapatite chromatography used in these studies has been described (15 of the native nuclear DNA from leukemic chicken cells was isolated as ss-DNA.…”

mentioning

confidence: 99%

“…Similar results, obtained with three different preparations of DNA, indicated that the main part of ss-DNA seems to consist of fragments from only one of the DNA double strands. The complementary sequences of these non-self-reassociable fragments do exist in the cell genome because, like ss-DNA from other cells (10,11), the ss-DNA from AMV-producing cells became hybridized to bulk nuclear DNA with the kinetics of nonrepeated DNA sequences. In addition, up to 35-36% of the ss-DNA from leukemic cells could be annealed with polysomal RNA as compared with 6-7% for bulk DNA [results not shown were similar to those obtained with ss-DNA from other cells (10, 11)].…”

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confidence: 99%

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Single-stranded DNA from oncornavirus-infected cells enriched in virus-specific DNA sequences

Leibovitch¹,

Tapiero²,

Harel³

1977

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

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We previously found that a minor fraction of single-stranded DNA (ss-DNA) isolated from native nuclear DNA of normal chicken embryonic cells and cells of other species hybridized with bulk nuclear DNA or cellular RNA in great excess. At least one-third of ss-DNA belonging to the nonrepetitious part of the cell genome could be hybridized to homologous RNAs. In the present work, similar results were obtained with ss-DNA from cells of chickens infected by avian myeloblastosis virus (AMV). To investigate whether this enrichment of ss-DNA in transcribed DNA sequences involves provirus DNA, radioactive AMV RNA and cDNA copies of AMV RNA were used. Most of the 70S AMV RNA hybridized much faster to ss-DNA from productively infected leukemic cells than to bulk DNA. cDNA, either double-stranded or single-stranded, made in the presence of actinomycin D hybridized to total nuclear DNA with similar kinetics. In contrast, about half of the double-stranded cDNA molecules hybridized 40-50 times faster to ss-DNA than to total DNA, indicating that only one of the provirus DNA strands seems to be present in ss-DNA. This was confirmed by the fact that relatively insignificant amounts of the ss-cDNA molecules made in the presence of actinomycin D could be annealed to ss-DNA as compared with bulk DNA. These results indicate that actively transcribed DNA sequences can be selectively distributed in the ss-DNA fraction, probably because of single strand breaks in the vicinity of transcription sites.The provirus theory of Temin (1), which postulates that the replication of RNA tumor viruses proceeds via DNA copies of viral RNA integrated in the host genome, has been substantiated by the discovery of successful transfection mediated by reverse transcriptase (2, 3), through an intermediate of viral DNA isolated from oncornavirus-transformed cells (4-6) and molecular hybridization between the viral genome and cell nucleic acids (7,8). In productively infected cells, the integrated provirus DNA appears to be more actively transcribed than most of the cellular genes (9).We have isolated, from the nDNA of various species, a minor fraction of single-stranded DNA (ss-DNA) and demonstrated that about one-third of the ss-DNA from cultured normal embryonic chicken cells (10) The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be-hereby marked "advertsement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

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confidence: 99%

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Single-stranded DNA from oncornavirus-infected cells enriched in virus-specific DNA sequences

Leibovitch¹,

Tapiero²,

Harel³

1977

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

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“…DNA, playing critical roles in storage and translation of genetic information [1][2][3][4], is an important target in drug discovery, especially in antiviral, anticancer and antibiotic drugs [5,6]. The study on the ligand-DNA-binding events is of great importance in rational design of novel drugs [7].…”

Section: Introductionmentioning

confidence: 99%

Interaction of bis-aryl functionalized molecules with nucleosides and nucleic acids

et al. 2010

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A series of novel molecules with a cyclen (1,4,7,10-tetraazacyclododecane) moiety appended on and bearing different aromatic fragments in the structures were synthesized and characterized. The binding activities of these compounds towards DNA were systematically studied by spectroscopic, viscometric and gel electrophoresis methods. The results suggest that the stacking interaction plays an important role in improving the DNA binding ability of the compounds. The binding modes of the compounds towards DNA are also affected by the sizes of the aromatic rings. The binding interaction between binaphthyl compound 1b and several nucleosides was studied by fluorescence titration. Stacking interaction and hydrophobic interaction play the key role in such non-selective binding process.stacking and hydrophobic interactions, DNA binding, aromatic compounds, nucleosides, oligonucleotides

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“…The method utilized to prepare RNA from the cytoplasmic phase of cells at 3 days (myoblasts: stage I), and 9 days (myotubes: stage IV) was previously reported [4]. In brief, after two successive centrifugations, 15 min at 5000 rev./min and 30 min at 12000 rev./min in a Sorvall swinging rotor, the final supernatant was made 0.2 M in MgClz and kept overnight at 4 "C. The precipitated RNA was pelleted and solubilized in 0.15 M NaCl, 0.01 M Tris-HC1 pH 7.4, 0.002 M EDTA.…”

Section: Preparation Of Cytoplasmic Rhtasmentioning

confidence: 99%

Relationship between Single‐Stranded DNA Isolated from Cultured Muscular Cells during Differentiation and the Transcription of Messenger RNA

Leibovitch¹,

Leibovitch²,

Kruh³

et al. 1979

European Journal of Biochemistry

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Single-stranded DNA (ssDNA), equivalent to about 2 "/, of the total nuclear DNA, was isolated by an improved method of hydroxyapatite chromatography from native nuclear DNA of rat myoblast cells and myotubes, of the Lg line. Small quantities of '251-labelled ssDNA were annealed with a large excess of unlabelled DNA, cytoplasmic RNA and mRNA from myoblasts or myotubes. The results indicated that ssDNA belongs to the non-repetitious portion of the cell genome and is formed of two distinct molecular fractions. The major ssDNA fractions (75 %) consist of non-selfreassociating DNA sequences and the minor fraction (25 x) consists of self-reassociating DNA sequences. About 30 -32 % and 25 -26 "/, of ssDNA from myoblast represent DNA sequences complementary to total cytplasmic RNAs and polyadenylated RNAs respectively. Hybridizations of ssDNA with an excess of RNA from myoblasts and/or myotubes show differences in the abundance and the diversity of mRNA during muscular differentiation. These differences were confirmed by DNA-driven reactions between 'Z51-labelled polyadenylated RNA and ssDNA in great excess.Various observations favour models of DNA transcription based on local opening of the DNA helix [l -31. Previous studies from our laboratory showed that a minor fraction of single-stranded DNA (ssDNA), usually amounting to 1.5 -2 of the total nuclear DNA, is related to the transcriptional activity, avian myeloblastosis virus [5] and ssDNA from mouse SV-3T3 cells (Shaool, D., Hanania, N., Harel, J . and May, E., unpublished results) non-productively transformed by simian virus 40 was found much enriched for transcribed virus-specific DNA sequences but contained no or very little DNA from the non-transcribed viral (or proviral) sequences. Muscle cells grown in culture provide a model system to study the role of gene expression during cell differentiation. After a defined time in culture, myoblast cells (stage I) from the L g line [7] fuse to form multinucleated myotubes (stage IV). This morphological event is characterized biochemically by the synthesis of musclespecific proteins [8-101 and changes in certain messenger RNAs [11 -131 producing modifications in chromatin protein kinases and phosphoproteins [14]. Our previous studies, based on hybridization of complementary DNAs (cDNAs) synthesized in a cellfree system t o cytoplasmic RNAs, had demonstrated typical changes in both the total number and relative abundance of polyadenylated RNAs during the course of muscular differentiation [14a]. The present work indicates that these changes in the transcriptional activity can be demonstrated at the level of ssDNA.

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Isolation of single stranded DNA related to the transcriptional activity of animal cells

Cited by 19 publications

References 16 publications

Single-stranded DNA from oncornavirus-infected cells enriched in virus-specific DNA sequences

Single-stranded DNA from oncornavirus-infected cells enriched in virus-specific DNA sequences

Interaction of bis-aryl functionalized molecules with nucleosides and nucleic acids

Relationship between Single‐Stranded DNA Isolated from Cultured Muscular Cells during Differentiation and the Transcription of Messenger RNA

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