A Distinct G
            <sub>1</sub>
            Step Required to Specify the Chinese Hamster DHFR Replication Origin

Wu, Jianpeng; Gilbert, David M.

doi:10.1126/science.271.5253.1270

Cited by 121 publications

(132 citation statements)

References 15 publications

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“…A related model is that prereplication complexes are assembled almost uniformly throughout the genome in early G 1 , but when transcription commences in active genes, are removed from the template, with the result that only intergenic regions usually serve as origins (Gilbert 2001). This would explain why CHO nuclei initiate replication throughout the DHFR gene and intergenic region when they are isolated from early G 1 cells, but initiation is confined to the intergenic region when they are isolated in late G 1 (Wu and Gilbert 1996). Studies in other model systems suggest that more global, highly regulated changes in chromatin environment mediate the interaction between transcription and local origin activity.…”

Section: Genes and Development 405mentioning

confidence: 99%

The promoter of the Chinese hamster ovary dihydrofolate reductase gene regulates the activity of the local origin and helps define its boundaries

Saha¹,

Shan²,

Mesner³

et al. 2004

Genes Dev.

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The dihydrofolate reductase (DHFR) and 2BE2121 genes in the Chinese hamster are convergently transcribed in late G 1 and early S phase, and bracket an early-firing origin of replication that consists of a 55-kb zone of potential initiation sites. To test whether transcription through the DHFR gene is required to activate this origin in early S phase, we examined the two-dimension (2D) gel patterns of replication intermediates from several variants in which parts or all of the DHFR promoter had been deleted. In those variants in which transcription was undetectable, initiation in the intergenic spacer was markedly suppressed (but not eliminated) in early S phase. Furthermore, replication of the locus required virtually the entire S period, as opposed to the usual 3-4 h. However, restoration of transcription with either the wild-type Chinese hamster promoter or a Drosophila-based construct restored origin activity to the wild-type pattern. Surprisingly, 2D gel analysis of promoterless variants revealed that initiation occurs at a low level in early S phase not only in the intergenic region, but also in the body of the DHFR gene. The latter phenomenon has never been observed in the wild-type locus. These studies suggest that transcription through the gene normally increases the efficiency of origin firing in early S phase, but also suppresses initiation in the body of the gene, thus helping to define the boundaries of the downstream origin.

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Section: Genes and Development 405mentioning

confidence: 99%

The promoter of the Chinese hamster ovary dihydrofolate reductase gene regulates the activity of the local origin and helps define its boundaries

Saha¹,

Shan²,

Mesner³

et al. 2004

Genes Dev.

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“…Events that occur during G-S period of the cell cycle, where the distribution of potential origins is defined (Wu and Gilbert 1996), are thus also dependent on previous mitotic events. Although chromosomal loops and loop anchors are still poorly defined biochemically, these data suggest that chromosome architecture plays a predominant role in the regulation of DNA replication origin localization and activation.…”

Section: Higher Levels Of Chromatin Organizationmentioning

confidence: 99%

Programming DNA replication origins and chromosome organization

2010

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During each cell cycle, thousands of DNA replication origins are activated in each cell of a metazoan organism. Although they appear site-specific, their usage and organization are rather plastic. Moreover, no strict sequence specificity has been observed in contrast to bacterial or Saccharomyces cerevisiae DNA replication origins. Epigenetic regulation linked to chromatin structure, chromosome organization, and transcription has been suggested to explain how DNA replication origins are selected and recognized by replication initiation factors. In this paper, we review these epigenetic features and discuss how, during the previous mitosis, chromosomal architecture might prepare DNA replication origins for a new cell cycle.

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“…1) and a widely used chromatin fractionation method [2,29,37]. In order to determine if there were changes occurring at or near the time of the origin decision point (ODP), a G1-phase hallmark at which origins of replication are specified [3,39], we first examined ORC-chromatin association in mitosis and G1-phase using CHO cells synchronized with a brief nocodazole block and release. Consistent with our studies and those of others utilizing the same cell line [3,36,40], we did not detect any significant change in the amounts of ORC1, 2 or 4 in the chromatin-containing fraction during mitosis and throughout G1-phase, although a small amount of ORC1 was consistently found in the soluble fraction during mitosis (Fig.…”

Section: Orc1 2 and 4 Are Stable Proteins That Co-fractionate With Cmentioning

confidence: 99%

Chinese hamster ORC subunits dynamically associate with chromatin throughout the cell-cycle

McNairn

Okuno

Misteli

et al. 2005

Experimental Cell Research

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In yeast, the Origin Recognition Complex (ORC) is bound to replication origins throughout the cellcycle, but in animal cells, there are conflicting data as to whether and when ORC is removed from chromatin. We find ORC1, 2 and ORC4 to be metabolically stable proteins that co-fractionate with chromatin throughout the cell-cycle in Chinese hamster fibroblasts. Since cellular extraction methods cannot directly examine the chromatin binding properties of proteins in vivo, we examined ORC:chromatin interactions in living cells. Fluorescence loss in photobleaching (FLIP) studies revealed ORC1 and ORC4 to be highly dynamic proteins during the cell-cycle with exchange kinetics similar to other regulatory chromatin proteins. In vivo interaction with chromatin was not significantly altered throughout the cell-cycle, including S-phase. These data support a model in which ORC subunits dynamically interact with chromatin throughout the cell-cycle.

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A Distinct G ₁ Step Required to Specify the Chinese Hamster DHFR Replication Origin

Cited by 121 publications

References 15 publications

The promoter of the Chinese hamster ovary dihydrofolate reductase gene regulates the activity of the local origin and helps define its boundaries

The promoter of the Chinese hamster ovary dihydrofolate reductase gene regulates the activity of the local origin and helps define its boundaries

Programming DNA replication origins and chromosome organization

Chinese hamster ORC subunits dynamically associate with chromatin throughout the cell-cycle

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A Distinct G 1 Step Required to Specify the Chinese Hamster DHFR Replication Origin

Cited by 121 publications

References 15 publications

The promoter of the Chinese hamster ovary dihydrofolate reductase gene regulates the activity of the local origin and helps define its boundaries

The promoter of the Chinese hamster ovary dihydrofolate reductase gene regulates the activity of the local origin and helps define its boundaries

Programming DNA replication origins and chromosome organization

Chinese hamster ORC subunits dynamically associate with chromatin throughout the cell-cycle

Contact Info

Product

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A Distinct G ₁ Step Required to Specify the Chinese Hamster DHFR Replication Origin