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Single-cell replication profiling reveals stochastic regulation of the mammalian replication-timing program [WGA]

ABSTRACT: This study demonstrates the stochastic nature of DNA replication-timing regulation by measures genome-wide replication timing in single-cells

ORGANISM(S): Mus musculus

PROVIDER: GSE102075 | GEO | 2017/12/19

SECONDARY ACCESSION(S): PRJNA396646

REPOSITORIES: GEO

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Single-cell replication profiling reveals stochastic regulation of the mammalian replication-timing program [population]

Project description:This study demonstrates the stochastic nature of DNA replication-timing regulation by measures genome-wide replication timing in single-cells

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Single-cell replication profiling reveals stochastic regulation of the mammalian replication-timing program [SEQXE]

Project description:This study demonstrates the stochastic nature of DNA replication-timing regulation by measuring genome-wide replication timing in single-cells

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Single-cell replication profiling reveals stochastic regulation of the mammalian replication-timing program

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Project description:We report the establishment of a single-cell DNA replication sequencing method, scRepli-seq, which is a simple genome-wide methodology that measures copy number differences between replicated and unreplicated DNA. Using scRepli-seq, we demonstrate that replication domain organization is conserved among individual mouse embryonic stem cells (mESCs). Differentiated mESCs exhibited distinct replication profiles, which were conserved from cell to cell. Haplotype-resolved scRepli-seq revealed similar replication timing profiles of homologous autosomes, while the inactive X chromosome was clearly replicated later than its active counterpart. However, a small degree of cell-to-cell replication timing heterogeneity was present, and we discovered that developmentally regulated domains are a source of such variability, suggesting a link between cell-to-cell heterogeneity and developmental plasticity. Together, our results form a foundation for single-cell-level understanding of DNA replication regulation and provide insights into 3D genome organization.

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Project description:Deregulated DNA replication is a major contributor to human developmental disorders and cancer, yet our understanding of how replication is coordinated with changes in transcription and chromatin structure is limited. Our lab has employed the zebrafish model to investigate the mechanisms driving changes in the replication timing program during development. Previous studies have identified changes in replication timing patterns from the onset of zygotic transcription through gastrulation in zebrafish embryos. The protein Rif1 is crucial for replication timing in a wide range of eukaryotes, yet its role in establishing the replication timing program and chromatin structure during early vertebrate development is not well understood. Using Rif1 mutant zebrafish and performing RNA sequencing and whole-genome replication timing analysis, we found that Rif1 mutants were viable but had a defect in female sex determination. Interestingly, Rif1 loss primarily affected DNA replication timing after gastrulation, while its impact on transcription was more pronounced during zygotic genome activation. Our results indicate that Rif1 has distinct roles in regulating DNA replication and transcription at different stages of development.

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Replication Timing in Yeast: CGH microarrays: S vs G1 cells in various stress conditions

Project description:Replication timing in Saccharomyces cerevisiae in various replication stress conditions using CGH microarrays of S vs G1 cells. Replication of the eukaryotic genomes follows a characteristic temporal program, with different regions replicating at different times in S phase. We describe a stochastic model of DNA replication that naturally explains the replication profiles of a dozen of S. cerevisiae mutants. Each profile is described by a single parameter that differ between the mutants: the ratio between the fork velocity and the typical initiation rate (the “replicon length”), while all other origin-specific parameters remain unchanged. We devise an analytical method to infer the replicon lengths from the available data, and predicted its value in different mutant backgrounds. Further experiments verified model’s predictions. Our study supports a stochastic model of DNA replication in S. cerevisiae and explains the apparent robustness of this profile to perturbations that extend S phase. DNA from FACS-sorted cells was extracted, labeled with Cy dyes, hybridized to the CGH microarray and scanned. 18 strains were used, with variable biological repeats number.

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Project description:As a part of the project looking into the role of WEE1 in regulating replication initiation, we report the replication timing analysis of the U2OS cells treated with WEE1 kinase inhibitor AZD1775

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