Simian virus 40 origin- and T-antigen-dependent DNA replication with Drosophila factors in vitro.

Kamakaka, Rohinton T.; Kaufman, Paul D.; Stillman, Bruce; Mitsis, Paul G.; Kadonaga, J T

doi:10.1128/mcb.14.8.5114

Cited by 28 publications

(21 citation statements)

References 48 publications

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“…In a previous study, we had observed a CAF-1-like activity in a crude extract derived from Drosophila embryos (14). In that work, it was shown that factors derived from Drosophila embryos were able to mediate SV40 origin-and T-antigen-dependent DNA replication in vitro.…”

Section: Resultsmentioning

confidence: 97%

“…Thus far, CAF-1 has been identified and characterized only in human cells. However, earlier studies that we had performed on SV40 origin-and T-antigen-dependent DNA replication with factors derived from Drosophila embryos had suggested that there is a related CAF-1 activity in Drosophila melanogaster (14). Because of the potentially significant insight to be gained from the study of chromatin assembly in both mammals and D. melanogaster, we felt that it would be important to identify and to characterize the Drosophila CAF-1 activity.…”

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confidence: 99%

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Postreplicative Chromatin Assembly by Drosophila and Human Chromatin Assembly Factor 1

Kamakaka

Bulger

Kaufman

et al. 1996

Molecular and Cellular Biology

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To study the relationship between DNA replication and chromatin assembly, we have purified a factor termed Drosophila chromatin assembly factor 1 (dCAF-1) to approximately 50% homogeneity from a nuclear extract derived from embryos. dCAF-1 appears to consist of four polypeptides with molecular masses of 180, 105, 75, and 55 kDa. dCAF-1 preferentially mediates chromatin assembly of newly replicated DNA relative to unreplicated DNA during T-antigen-dependent simian virus 40 DNA replication in vitro, as seen with human CAF-1. Analysis of the mechanism of DNA replication-coupled chromatin assembly revealed that both dCAF-1 and human CAF-1 mediate chromatin assembly preferentially with previously yet newly replicated DNA relative to unreplicated DNA. Moreover, the preferential assembly of the postreplicative DNA was observed at 30 min after inhibition of DNA replication by aphidicolin, but this effect slowly diminished until it was no longer apparent at 120 min after inhibition of replication. These findings suggest that the coupling between DNA replication and chromatin assembly may not necessarily involve a direct interaction between the replication and assembly factors at a replication fork.Replication of the eukaryotic genome involves both the replication of DNA and the assembly of the newly synthesized DNA into chromatin (for reviews, see references 7, 11, 34, 35, and 36), and it is therefore important to study the relationship between DNA replication and chromatin assembly. The current data suggest that the assembly of chromatin in vivo begins immediately after DNA replication by a mechanism that leads to a random distribution of the newly synthesized and preexisting histones between the daughter DNA strands. In addition, it appears that nucleosome assembly occurs by the initial deposition of histones H3 and H4 followed by the incorporation of histones H2A and H2B. Structures that resemble bulk nucleosomes have been observed at replication forks by elec-

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Section: Resultsmentioning

confidence: 97%

mentioning

confidence: 99%

Postreplicative Chromatin Assembly by Drosophila and Human Chromatin Assembly Factor 1

Kamakaka

Bulger

Kaufman

et al. 1996

Molecular and Cellular Biology

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“…DNA polymerase ␣-primase interacts directly with each of the three other components of the complex, and any of these interactions could, in principle, form the basis of the observed species specificity of DNA replication (4,5,34). However, the functional cooperation of RPA and DNA polymerase ␣-primase from mammalian origin has been shown not to be species-specific (27,29,54). Therefore, we concentrated on the functional interactions between SV40 TAg and DNA polymerase ␣-primase and, more specifically, on the p180 subunit known to control SV40 species specificity (29).…”

Section: Discussionmentioning

confidence: 99%

Species Specificity of Simian Virus 40 DNA Replication in Vitro Requires Multiple Functions of Human DNA Polymerase α

Smith¹,

Steffen²,

Grosse³

et al. 2002

Journal of Biological Chemistry

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Human cell extracts support the replication of SV40 DNA, whereas mouse cell extracts do not. Species specificity is determined at the level of initiation of DNA replication, and it was previously found that this requires the large subunit, p180, of DNA polymerase ␣-primase to be of human origin. Furthermore, a functional interaction between SV40 large T antigen (TAg) and p180 is essential for viral DNA replication. In this study we determined that the N-terminal regions of human p180, which contain the TAg-binding sites, can be replaced with those of murine origin without losing the ability to support SV40 DNA replication in vitro. The same substitutions do not prevent SV40 TAg from stimulating the activity of DNA polymerase ␣-primase on single-stranded DNA in the presence of replication protein A. Furthermore, biophysical studies show that the interactions of human and murine DNA polymerase ␣-primase with SV40 TAg are of a similar magnitude. These studies strongly suggest that requirement of SV40 DNA replication for human DNA polymerase ␣ depends neither on the TAg-binding site being of human origin nor on the strength of the binary interaction between SV40 TAg and DNA polymerase ␣-primase but rather on sequences in the C-terminal region of human p180.Polyomavirus DNA replication has been studied extensively because of the ease with which viruses of this family, which include SV40 and mouse polyoma virus (PyV), 1 can be grown in cell culture and moreover because of the availability of an in vitro replication system, which allows detailed investigation of the individual factors that play a role in the replication process (1, 2). Polyomavirus DNA replication has been found to be largely dependent upon factors involved in replication of the host DNA but does contribute one essential trans-acting factor known as large T antigen (TAg) to the replication complex. TAg is responsible for recognition of the viral origin of replication, at which it forms a double hexamer (3). In the presence of the single-stranded DNA-binding protein, RPA (replication protein A), and topoisomerase I, TAg proceeds to unwind the origin DNA and recruits the DNA polymerase ␣-primase heterotetramer, which then initiates bidirectional DNA synthesis (4 -11). DNA polymerase ␣-primase initiates DNA replication through the action of its smallest subunit, p48, which synthesizes RNA primers that are subsequently elongated by the large subunit, p180 (12-15). The bulk of DNA synthesis is, however, carried out by a more processive enzyme complex consisting of DNA polymerase ␦ and its processivity factor, PCNA (16 -18). The transition between DNA synthesis by DNA polymerases ␣ and ␦ is mediated by the PCNA loading factor replication factor C (16, 19). Throughout the viral replication cycle, TAg double hexamers are thought to act as the replicative helicase (20). For a more extensive account of the DNA replication process and its participating factors see Refs. 1, 2, and 21-23. SV40 and PyV are very similar with regard to DNA replication; their respectiv...

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“…32 P]dATP were performed and analyzed as described previously (38,42), with the specific modifications reported by Kamakaka et al (17). CAF-1-dependent assembly of nucleosomes during the replication reaction was observed by the incorporation of negative supercoils into the newly replicated DNA, as detected by autoradiography.…”

Section: Methodsmentioning

confidence: 99%

The p55 Subunit of Drosophila Chromatin Assembly Factor 1 Is Homologous to a Histone Deacetylase-Associated Protein

Tyler

Bulger

Kamakaka

et al. 1996

Molecular and Cellular Biology

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124

116

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To gain a better understanding of DNA replication-coupled chromatin assembly, we have isolated the cDNA encoding the smallest (apparent molecular mass, 55 kDa; termed p55) subunit of Drosophila melanogaster chromatin assembly factor 1 (dCAF-1), a multisubunit protein that is required for the assembly of nucleosomes onto newly replicated DNA in vitro. The p55 polypeptide comprises seven WD repeat motifs and is homologous to the mammalian RbAp48 protein, which is associated with the HD1 histone deacetylase. dCAF-1 was immunopurified by using affinity-purified antibodies against p55; the resulting dCAF-1 preparation possessed the four putative subunits of dCAF-1 (p180, p105, p75, and p55) and was active for DNA replication-coupled chromatin assembly. Moreover, dCAF-1 activity was specifically depleted with antibodies against p55. Thus, p55 is an integral component of dCAF-1. p55 is localized to the nucleus and is present throughout Drosophila development. Consistent with the homology between p55 and the HD1-associated RbAp48 protein, histone deacetylase activity was observed to coimmunoprecipitate specifically with p55 from a Drosophila nuclear extract. Furthermore, a fraction of the p55 protein becomes associated with the newly assembled chromatin following DNA replication. These findings collectively suggest that p55 may function as a link between DNA replication-coupled chromatin assembly and histone modification.The assembly of newly replicated DNA into chromatin is a fundamental biological process that affects the structure and function of the genome (for reviews, see references 10, 18, 19, 22, 43, 45, and 49). In the analysis of chromatin assembly, it is important to consider not only the factors that mediate the formation of nucleosomes but also the nature of the components-the histones and DNA-from which chromatin is derived. With respect to the DNA, chromatin assembly appears to commence immediately after DNA replication, and nucleosomes are assembled onto the newly synthesized DNA. With regard to the histones, newly synthesized histones are modified (such as by acetylation and/or phosphorylation) and then deposited onto newly synthesized DNA, after which further modifications (such as deacetylation) can occur. In addition, roughly one-half of the histones in newly assembled chromatin are derived from the parental preexisting nucleosomes that are disassembled and reassembled during passage of the replication fork. Thus, in this manner, the process of nucleosome assembly is an important step in the creation of daughter chromosomes from the parental chromosome.The biochemical analysis of DNA replication and chromatin assembly has led to the identification and characterization of a protein termed chromatin assembly factor 1 (CAF-1) from human cells (20,37,38,42) and Drosophila melanogaster embryos (16). CAF-1 was identified by its ability to facilitate the assembly of newly replicated DNA into chromatin during Tantigen-mediated simian virus 40 (SV40) DNA replication. In these experiments, DNA replication-chr...

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Simian virus 40 origin- and T-antigen-dependent DNA replication with Drosophila factors in vitro.

Cited by 28 publications

References 48 publications

Postreplicative Chromatin Assembly by Drosophila and Human Chromatin Assembly Factor 1

Postreplicative Chromatin Assembly by Drosophila and Human Chromatin Assembly Factor 1

Species Specificity of Simian Virus 40 DNA Replication in Vitro Requires Multiple Functions of Human DNA Polymerase α

The p55 Subunit of Drosophila Chromatin Assembly Factor 1 Is Homologous to a Histone Deacetylase-Associated Protein

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