Minichromosome of simian virus 40: presence of histone HI

Varshavsky, Alexander; Bakayev, Valery V.; Chumackov, P.M.; Georgiev, G.P.

doi:10.1093/nar/3.8.2101

Cited by 116 publications

(74 citation statements)

References 18 publications

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“…After incubation at the nonpermissive temperature a primary virus stock was obtained consisting of over 70% recombinant virions. SV40 minichromosomes containing the various genes were isolated from infected cells by a Triton X-100-EDTA extraction procedure (16), yielding 75S particles; no nucleosomefree DNA that sediments at 21S was observed in these preparations. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that numerous proteins in addition to cellular histones cosediment with the minichromosomes (data not shown).…”

Section: Resultsmentioning

confidence: 99%

“…During infection, SV40 DNA is packaged by the host cell histones into a chromatin structure termed the minichromosome (16). In light of the stable transcription complexes observed on class III genes in vitro (2,9,13) and the association of factor with cellular chromatin (11, 17), we decided to investigate whether the transcription factors that interact with these genes remain bound to the template in a stable complex during the isolation of recombinant minichromosomes containing such genes.…”

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confidence: 99%

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Stable transcription complex on a class III gene in a minichromosome.

Lassar¹,

Hamer²,

Roeder³

1985

Mol. Cell. Biol.

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We have constructed recombinant simian virus 40 molecules containing Xenopus 5S RNA and tRNA genes. Recombinant minichromosomes containing these genes were isolated to study the interaction of RNA polymerase III transcription factors with these model chromatin templates. Minichromosomes containing a tRNAMet gene can be isolated in a stable complex with transcription factors (IIIB and IIIC) and are active in vitro templates for purified RNA polymerase III. In contrast, minichromosomes containing a 5S RNA gene are refractory to transcription by purified RNA polymerase III in either the absence or the presence of other factors.Fractionation of soluble transcription systems has indicated that multiple cellular factors in addition to RNA polymerase III are necessary to transcribe class III genes (14,15). Two factors (designated IIIB and IIIC) are necessary for transcription of the tRNA and the adenovirus VA RNA genes in vitro, whereas the 5S genes require these same factors plus a third gene-specific factor (IIIA) (14, 15). Formation of a stable transcription complex in vitro, containing one or more of these factors, is a general feature of these genes (2, 9, 13). The finding that isolated cellular chromatin is associated with all the factors necessary to promote transcription of the endogenous 5S and tRNA genes by purified RNA polymerase III suggests that such complexes may exist in vivo (11,17). To establish whether these chromatin-associated factors are specifically bound in a stable complex on class III genes, we have studied their interaction with a model chromatin template-the simian virus 40 (SV40) minichromosome.Numerous class III genes cloned into SV40 have been found to be transcriptionally active in the infected cell (8,18,19). During infection, SV40 DNA is packaged by the host cell histones into a chromatin structure termed the minichromosome (16). In light of the stable transcription complexes observed on class III genes in vitro (2, 9, 13) and the association of factor with cellular chromatin (11, 17), we decided to investigate whether the transcription factors that interact with these genes remain bound to the template in a stable complex during the isolation of recombinant minichromosomes containing such genes. Studies were conducted with a Xenopus laevis tRNAMet gene and a Xenopus borealis 5S RNA gene cloned into the late region of SV40. Previous work has shown that formation of a stable complex in vitro requires factors B and C for the tRNAMet gene and factors A and C for the 5S gene (9). Here we show that the tRNAMet gene in the minichromosome can be isolated in a stable complex with factors B and C and is an active in vitro template for purified RNA transcription by purified RNA polymerase III in either the absence or the presence of other factors. MATERIALS AND METHODSVirus construction. The tRNA SV40 fragments, orientations 1 and 2, were constructed as follows. An EcoRI-linkered, 3,055-base-pair fragment of wild-type SV40, extending from the BamHI site to the HpaII site, was cloned into the Ec...

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Stable transcription complex on a class III gene in a minichromosome.

Lassar¹,

Hamer²,

Roeder³

1985

Mol. Cell. Biol.

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“…4) The present study demonstrates that the SV40 viral transcription complex (VTC) can synthesize long RNA transcripts under conditions in which protein does not dissociate from the transcriptionally active chromatin. Electron microscopic and biochemical studies have revealed a close similarity between the structure of SV40 and cellular chromatin (4)(5)(6)(7)(8)(9)(10). On the basis of sedimentation rate (23) and density in CsCl (Fig.…”

Section: H-rna ( ); 14c-vtc (---); 14c-dna (---)mentioning

confidence: 91%

“…The viral complex has been shown by electron microscopic (4)(5)(6) and biochemical (5,(7)(8)(9)(10) analyses to possess a structure which closely resembles that of cellular chromatin. There is an average of 21 beads (nucleosomes) per SV40 genome, each having a diameter of ca.…”

Section: Introductionmentioning

confidence: 99%

The SV40 transcription complex. II. Non-dissociation of protein from SV40 chromatin during transcription

Green

Brooks

1977

Nucl Acids Res

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ABSTRACT.A small fraction of the SV40 chromatin isolated from infected monkey cell cultures by the Triton method contains active RNA polymerase which had initiated transcription in vivo. This viral transcription complex (VTC) was utilized to answer the question of whether proteins dissociate from chromatin during transcription in vitro. 3H-RNA was synthesized by the VTC under conditions such that over half the label was in transcripts which were longer than half the length of the SV40 genome. Virtually all of the 3H-RNA remained associated with the SV40 chromatin, causing an increase in sedimentation rate from 55S to 78S. The density of the VTC-3H-RNA complex indicated that less than 5% of the original protein dissociated from the SV40 DNA which served as a template for transcription. We conclude that SV40 chromatin can be transcribed while the proteins remain associated with the DNA.INTRODUCTION.

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“…Isolated papovavirus chromatin has been used as a simple model for the study of the fine structure of chromosomes (1)(2)(3)(4)(5). A close similarity between the structural organizations of viral and eukaryotic chromatins was revealed with the main feature being that beaded globular structures termed "nucleosomes" (6), which consist of cellular histones (5,7) are aligned along the DNA filaments on a small number of distinct alternative positions (4).…”

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confidence: 99%

Origin of DNA replication in papovavirus chromatin is recognized by endogenous endonuclease.

Waldeck¹,

Föhring²,

Chowdhury³

et al. 1978

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

116

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Isolated simian virus 40 (SV40) and polyoma nucleoprotein complexes contain endonuclease that, under in vitro conditions, converts part (up to 30%) of the covalently closed superhelical DNA to full-length linear rods. The positions of the cleavage sites within the genomes of SV40 and polyoma were determined by digestion with various single-cut restriction endonucleases and subsequent agarose gel electrophoresis of the cleavage products. Both SV40 and polyoma covalently closed superhelical DNA were cleaved open at their respective origins of DNA replication (±75 base pairs). The full-length linear DNA rods whose ends map adjacent to the origin of DNA replication could also be isolated by sodium dodecyl sulfate/phenol extraction both from SV40-infected permissive cells and from purified SV40 virions. These data reveal the resence of a unique structure of the papovavirus chromatin cose to the initiation site of DNA replication.Isolated papovavirus chromatin has been used as a simple model for the study of the fine structure of chromosomes (1-5). A close similarity between the structural organizations of viral and eukaryotic chromatins was revealed with the main feature being that beaded globular structures termed "nucleosomes" (6), which consist of cellular histones (5, 7) are aligned along the DNA filaments on a small number of distinct alternative positions (4).Viral chromatin also has been successfully used to determine the in vitro factors and conditions that are required for DNA replication. The availability of soluble nucleoprotein complexes derived from the nuclei of simian virus 40 (SV40)-infected cells (8) has fostered such studies (8,9). A number of enzymes such as a and y DNA polymerases (10,11) and RNA polymerase (12) are associated with papovavirus nucleoprotein complexes. As will be shown in this report, there is, in addition, an endonuclease activity present that introduces one double-strand break into the viral DNA. In the case of both SV40 and polyoma DNA, the position that is susceptible to the endonuclease activity within the chromatin was shown to be located at the origin of replication. This finding has revealed a specific structure of the papovavirus chromatin in a biologically important region of the genome. MATERIALS AND METHODSVirus and Cells. SV40 strain Rh 911 was grown on CV-1 cells as described (13). Propagation of polyoma virus and infection of 3T6 cells were carried out as described (14). For isolation of nucleoprotein complexes the cells were infected at a multiplicity of 10 plaque-forming units per cell.Preparation of Nucleoprotein Complexes. Nuclei were isolated from SV40-infected CV-1 cells (107 cells in a 15-cm petri dish, 38 hr after infection) and from polyoma-infected 3T6 cells (2 X 107 cells in a 15-cm petri dish, 26 hr after infection) as described (14). For preparation of nuclear extracts as initially described by Su and De Pamphilis (8) the nuclei were suspended in 2 ml of modified hypotonic extraction buffer (10 mM Hepes, pH 8.0/1 mM MgCl2/0.5 mM CaCl2/1 mM dithio...

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Minichromosome of simian virus 40: presence of histone HI

Cited by 116 publications

References 18 publications

Stable transcription complex on a class III gene in a minichromosome.

Stable transcription complex on a class III gene in a minichromosome.

The SV40 transcription complex. II. Non-dissociation of protein from SV40 chromatin during transcription

Origin of DNA replication in papovavirus chromatin is recognized by endogenous endonuclease.

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