Similar Datasets
Project description:Faithful DNA replication is essential for genome integrity. Under-replicated DNA leads to chromosome segregation defects, which are reportedly common during embryogenesis. However, DNA replication regulation remains poorly understood in early mammalian embryos. Here, we constructed a single-cell genome-wide DNA replication atlas of pre-implantation mouse embryos and discovered an abrupt replication program switch accompanied by a transient period of genomic instability. In 1- and 2-cell embryos, we observed the complete absence of a replication timing (RT) program, and the entire genome replicated gradually and uniformly using extremely slow-moving forks. In 4-cell embryos, a somatic-cell-like RT program commenced abruptly. However, the fork speed was still slow, S-phase was extended, and SLX4 DNA repair foci increased during G2/M, which was followed by a transient increase in chromosome segregation errors. Importantly, live imaging captured longer S-phase of error cells, and the breakpoints identified by single-cell genome sequencing were enriched in late-replicating regions. By the 8-cell stage, forks gained speed, S-phase was no longer extended, and chromosome aberrations disappeared. Thus, a transient period of genomic instability exists during normal mouse development, which is preceded by a fragile S-phase lacking the coordination between replisome-level regulation and megabase-scale RT regulation, implicating the importance of their coordination for genome integrity.
2024-06-24 | GSE226309 | GEO
Project description:Embryonic genome instability upon DNA replication timing program emergence
| PRJNA1075049 | ENA
Project description:Embryonic genome instability upon DNA replication timing program emergence
| PRJNA939700 | ENA
Project description:Mammalian DNA replication is a highly organized and regulated process. Large, Mb-sized regions are replicated at defined times along S-phase. Replication Timing (RT) is thought to play a role in shaping the mammalian genome by affecting mutation rates. Previous analyses relied on somatic RT profiles. However, only germline mutations are passed on to offspring and affect genomic composition. Therefore, germ cell RT information is necessary to evaluate the influences of RT on the mammalian genome. We adapted the RT mapping technique for limited amounts of cells, and measured RT from two stages in the mouse germline - primordial germ cells (PGCs) and spermatogonial stem cells (SSCs). RT in germline cells exhibited stronger correlations to both mutation rate and recombination hotspots density than those of RT in somatic tissues, emphasizing the importance of using correct tissues-of-origin for RT profiling. Germline RT maps exhibited stronger correlations to additional genetic features including GC-content, transposable elements (SINEs and LINEs), and gene density. GC content stratification and multiple regression analysis revealed independent contributions of RT to SINE, gene, mutation, and recombination hotspot densities. Together, our results establish a central role for RT in shaping multiple levels of mammalian genome composition.
| S-EPMC6144785 | biostudies-literature
Project description:The organization of mammalian DNA replication is poorly understood. We have produced high-resolution dynamic maps of the timing of replication in human erythroid, mesenchymal, and embryonic stem (ES) cells using TimEX, a method that relies on gaussian convolution of massive, highly redundant determinations of DNA copy-number variations during S phase to produce replication timing profiles. We first obtained timing maps of 3% of the genome using high-density oligonucleotide tiling arrays and then extended the TimEX method genome-wide using massively parallel sequencing. We show that in untransformed human cells, timing of replication is highly regulated and highly synchronous, and that many genomic segments are replicated in temporal transition regions devoid of initiation, where replication forks progress unidirectionally from origins that can be hundreds of kilobases away. Absence of initiation in one transition region is shown at the molecular level by single molecule analysis of replicated DNA (SMARD). Comparison of ES and erythroid cells replication patterns revealed that these cells replicate about 20% of their genome in different quarters of S phase. Importantly, we detected a strong inverse relationship between timing of replication and distance to the closest expressed gene. This relationship can be used to predict tissue-specific timing of replication profiles from expression data and genomic annotations. We also provide evidence that early origins of replication are preferentially located near highly expressed genes, that mid-firing origins are located near moderately expressed genes, and that late-firing origins are located far from genes.
| S-EPMC2792175 | biostudies-literature
Project description:We describe the DNA replication timing programs of 14 yeast mutants with an extended S phase identified by a novel genome-wide screen. These mutants are associated with the DNA replication machinery, cell-cycle control, and dNTP synthesis and affect different parts of S phase. In 13 of the mutants, origin activation time scales with the duration of S phase. A limited number of origins become inactive in these strains, with inactive origins characterized by small replicons and distributed throughout S phase. In sharp contrast, cells deleted of MRC1, a gene implicated in replication fork stabilization and in the replication checkpoint pathway, maintained wild-type firing times despite over twofold lengthening of S phase. Numerous dormant origins were activated in this mutant. Our data suggest that most perturbations that lengthen S phase affect the entire program of replication timing, rather than a specific subset of origins, maintaining the relative order of origin firing time and delaying firing with relative proportions. Mrc1 emerges as a regulator of this robustness of the replication program.
| S-EPMC2877575 | biostudies-literature
Project description:Human embryonic stem cells (hESCs), due to their pluripotent nature, represent a particularly relevant model system to study the relationship between the replication program and differentiation state. Here, we define the basic properties of the replication program in hESCs and compare them to the programs of hESC-derived multipotent cells (neural rosette cells) and primary differentiated cells (microvascular endothelial cells [MECs]). We characterized three genomic loci: two pluripotency regulatory genes, POU5F1 (OCT4) and NANOG, and the IGH locus, a locus that is transcriptionally active specifically in B-lineage cells. We applied a high-resolution approach to capture images of individual replicated DNA molecules. We demonstrate that for the loci studied, several basic properties of replication, including the average speed of replication forks and the average density of initiation sites, were conserved among the cells analyzed. We also demonstrate, for the first time, the presence of initiation zones in hESCs. However, significant differences were evident in other aspects of replication for the DNA segment containing the POU5F1 gene. Specifically, the locations of centers of initiation zones and the direction of replication fork progression through the POU5F1 gene were conserved in two independent hESC lines but were different in hESC-derived multipotent cells and MECs. Thus, our data identify features of the replication program characteristic of hESCs and define specific changes in replication during hESC differentiation.
| S-EPMC2937526 | biostudies-literature
Project description:The DNA polymerase zeta (Polζ) plays a critical role in bypassing DNA damage. REV3L, the catalytic subunit of Polζ is also essential in mouse embryonic development and cell proliferation for reasons that remain incompletely understood. In this study, we reveal that REV3L protein interacts with heterochromatin components including repressive histone marks, and localizes in pericentromeric regions through direct interaction with HP1 dimer. We demonstrate that Polζ/REV3L ensures progression of replication forks through difficult-to-replicate pericentromeric heterochromatin, thereby preventing spontaneous chromosome break formation. We also find that Rev3l-deficient cells are compromised in the repair of heterochromatin-associated double-strand breaks, eliciting deletions in late replicating regions. Lack of REV3L leads to further consequences that may be ascribed to heterochromatin replication and repair-associated functions of Polζ, with a disruption of the temporal replication program at specific loci. This is correlated with changes in epigenetic landscape and transcriptional control of developmentally regulated genes. These results reveal a new function of Polζ in preventing chromosome instability during replication of heterochromatic regions.
| S-SCDT-EMBOJ-2020-104543 | biostudies-other
Project description:DNA replication enables genetic inheritance across the kingdoms of life. Replication occurs with a defined temporal order known as the replication timing (RT) programme, leading to organization of the genome into early- or late-replicating regions. RT is cell-type specific, is tightly linked to the three-dimensional nuclear organization of the genome1,2 and is considered an epigenetic fingerprint3. In spite of its importance in maintaining the epigenome4, the developmental regulation of RT in mammals in vivo has not been explored. Here, using single-cell Repli-seq5, we generated genome-wide RT maps of mouse embryos from the zygote to the blastocyst stage. Our data show that RT is initially not well defined but becomes defined progressively from the 4-cell stage, coinciding with strengthening of the A and B compartments. We show that transcription contributes to the precision of the RT programme and that the difference in RT between the A and B compartments depends on RNA polymerase II at zygotic genome activation. Our data indicate that the establishment of nuclear organization precedes the acquisition of defined RT features and primes the partitioning of the genome into early- and late-replicating domains. Our work sheds light on the establishment of the epigenome at the beginning of mammalian development and reveals the organizing principles of genome organization.
| S-EPMC10781638 | biostudies-literature
Project description:This SuperSeries is composed of the SubSeries listed below.
2017-12-19 | GSE102077 | GEO