DNA replication timing during development anticipates transcriptional programs and parallels enhancer activation

Siefert, Joseph C.; Georgescu, Constantin; Wren, Jonathan D.; Koren, Amnon; Sansam, Christopher L.

doi:10.1101/gr.218602.116

Cited by 63 publications

(84 citation statements)

References 64 publications

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“…Differences in replication timing can have significant consequences, as late replication is associated with higher frequencies of mutation and genome rearrangement (Chen et al, 2010;Koren et al, 2012;Lang & Murray, 2011;Stamatoyannopoulos et al, 2009;Weber, Pink, & Hurst, 2012;Yehuda et al, 2018). The importance of replication timing is underscored by the observation that it is sometimes modulated according to the utility of the region being replicated: For example, some chromosomal regions containing developmentally regulated genes replicate early only during those developmental phases during which they are activated (Hiratani et al, 2010;Rivera-Mulia et al, 2015;Siefert, Georgescu, Wren, Koren, & Sansam, 2017). An even more striking example is the X chromosome in female mammals, where the active X chromosome (Xa) replicates early while the inactive X (Xi) replicates late (Gilbert, Muldal, Lajtha, & Rowley, 1962;Hansen, Canfield, Fjeld, & Gartler, 1996;Heard & Disteche, 2006).…”

Section: Introductionmentioning

confidence: 99%

Chromosomal Mcm2-7 distribution is the primary driver of the genome replication program in species from yeast to humans

Foss

Sripathy

Gatbonton-Schwager

et al. 2019

Preprint

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The spatio-temporal program of genome replication across eukaryotes is thought to be driven both by the uneven loading of pre-replication complexes (pre-RCs) across the genome at the onset of S-phase, and by differences in the timing of activation of these complexes during Sphase. To determine the degree to which distribution of pre-RC loading alone could account for chromosomal replication patterns, we identified the binding sites of the Mcm2-7 helicase complex, a key component of the pre-RC that is required for initiation of DNA replication, in budding yeast, fission yeast and mouse. In budding yeast, we detected Mcm2 binding in sharply focused peaks, generally with a single double hexamer bound at known origins of replication. In fission yeast, Mcm2 binding, while still concentrated at known origins, was more diffuse, often with 6 to 8 helicase complexes distributed along 0.5-1.5 kb sized origins, and with significantly more binding between origins. Finally, in mouse, we found even more diffuse Mcm2-7 distribution, with the density of Mcm2-7 binding in G1 recapitulating to a remarkable degree the replication program implemented in S-phase. Computer simulations that assign each licensed origin an equal probability of firing show that the observed Mcm2-7 density distribution in G1 across all three species largely recapitulated the DNA replication program. We conclude that the pattern of origin licensing from yeast to mammals is sufficient to explain most differences in replication timing without invoking an overarching temporal program of origin firing. ResultsInitiation of DNA replication requires activation of the MCM2-7 helicase complex (Mcm2-7) at sites at which this complex was loaded during the preceding G2/M and G1 phases [1][2][3][4].However, the order in which different regions of the genome complete replication is widely assumed to reflect not only the locations of Mcm2-7 loading but also a program of control above the level of loading, in which certain complexes are activated before others [5,6]. The distribution of Mcm2-7 is therefore expected to constrain but not fully determine the program of genome replication. Mcm2-7 encircles approximately 60 base pairs (bp) of DNA as a double hexamer whose monomer components are juxtaposed head-to-head at their N-termini with C-

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Section: Introductionmentioning

confidence: 99%

Chromosomal Mcm2-7 distribution is the primary driver of the genome replication program in species from yeast to humans

Foss

Sripathy

Gatbonton-Schwager

et al. 2019

Preprint

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“…However, it is not late replicating until the end of gastrulation or bud stage (10 hpf) (Siefert et al, 2017). Previous work in zebrafish has found ZnF proteins on chromosome 4 to undergo robust expression from the initiation of zygotic transcription until mid-gastrula stage (White et al, 2017).…”

Section: Discussionmentioning

confidence: 99%

Gene-environment interactions characterized by single embryo transcriptomics

Sidik¹,

Dixon²,

Kirby³

et al. 2019

Preprint

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Fetal Alcohol Spectrum Disorders (FASD) describes the full range of defects that result from prenatal alcohol exposure. The clinical diagnosis of Fetal Alcohol Syndrome (FAS) is the most severe outcome, characterized by facial dysmorphism and neurological deficits. Gene-ethanol interactions underlie susceptibility to FASD but we lack a clear understanding of the genetic risk factors. Here, we leverage the genetic tractability of zebrafish to address this problem. We first show that vangl2, a member of the Wnt/planar cell polarity (Wnt/PCP) pathway that mediates convergent extension movements, strongly interacts with ethanol during late blastula and early gastrula stages. Embryos mutant or heterozygous for vangl2 are sensitized to ethanol-induced midfacial hypoplasia. However, we have no understanding of the potential mechanism of this sensitization. We performed single-embryo RNA-Seq during early embryonic stages, to assess individual variation to the transcriptional response to ethanol and determine the mechanism of the vangl2-ethanol interaction. Transcriptional changes due to ethanol are indicative of increased oxidative stress and ion transport as well as reduced DNA replication and cell division. To identify the pathway(s) that are disrupted by ethanol we used these global changes in gene expression to identify small molecules, often pathway inhibitors, that mimic the effects of ethanol via the Library of Integrated Network-based Cellular Signatures (LINCS L1000) dataset. This dataset predicted that the Sonic Hedgehog (Shh) pathway inhibitor, cyclopamine, would mimic the effects of ethanol, despite ethanol not altering the expression levels of direct targets of Shh signaling. Indeed, we found that ethanol and cyclopamine strongly interact to disrupt midfacial development. Collectively, these results suggest that the midfacial defects in ethanol-exposed vangl2 mutants are due to an indirect interaction between ethanol and the Shh pathway. Vangl2 functions as part of a signaling pathway that regulates coordinated cell movements during midfacial development. 3Consistent with an indirect model, a critical source of Shh signaling that separates the developing eye field into bilateral eyes, allowing the expansion of the midface, becomes mispositioned in ethanol-exposed vangl2 mutants.

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“…We wanted to know if START-R suite can run the correct analyses with this type of data and also with other organisms than mouse and human. We performed exactly the same pipeline used for early-late Repli-seq data described above for Drosophila, zebrafish and human S/G1 data (Armstrong et al, 2018;Siefert et al, 2017;Massey et al, 2019), in order to be sure that the integration into the START-R pipeline was correct. Then and as expected,…”

Section: Validation Of Start-r With Early-late Repli-seq Data From Mousementioning

confidence: 99%

“…Different laboratories analyze variations of DNA copy number between G1 and S phase cells (S/G1 ratio) to study the replication timing program with Repli-seq. We used data obtained from different organisms such as Drosophila, zebrafish and human (Armstrong et al, 2018;Siefert et al, 2017;Massey et al, 2019), to validate the START-R suite (GEO accession numbers: GSM3154888 and GSM3154890 for HWT Drosophila melanogaster female larvae wing disc cells in S and G1 phase, respectively; GSM2282090 for 28hpf Danio rerio embryos; SRX3413939-40 for HEK293T human cells in S and G1 phase, respectively). As previously, reads from G1 and S fractions are mapped with Bowtie2, then PCR duplicates are removed by RmDUP tool, and Bamcoverage is used to obtain the coverage with RPKM.…”

Section: Validation Of Start-r Suite Using S/g1 Data From Multiple Spmentioning

confidence: 99%

New, easy, quick and efficient DNA replication timing analysis by high-throughput approaches

Hadjadj

Denecker

et al. 2019

Preprint

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DNA replication must be faithful and follow a well-defined spatio-temporal program closely linked to transcriptional activity, epigenomic marks, intra-nuclear structures, mutation rate and cell fate determination. Among the readouts of the DNA replication spatio-temporal program, replication timing (RT) analyses require complex, precise and time-consuming experimental procedures, and the study of large-size computer files. We improved the RT protocol to speed it up and increase its quality and reproducibility. Also, we partly automated the RT protocol and developed a user-friendly software: the START-R suite (Simple Tool for the Analysis of the Replication Timing based on R). START-R suite is an open source web application using an R script and an HTML interface to analyze DNA replication timing in a given cell line with microarray or deep-sequencing results. This novel approach can be used by every biologist without requiring specific knowledge in bioinformatics. It also reduces the time required for generating and analyzing simultaneously data from several samples. START-R suite detects constant timing regions (CTR) but also, and this is a novelty, it identifies temporal transition regions (TTR) and detects significant differences between two experimental conditions. The informatic global analysis requires less than 10 minutes.

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DNA replication timing during development anticipates transcriptional programs and parallels enhancer activation

Cited by 63 publications

References 64 publications

Chromosomal Mcm2-7 distribution is the primary driver of the genome replication program in species from yeast to humans

Chromosomal Mcm2-7 distribution is the primary driver of the genome replication program in species from yeast to humans

Gene-environment interactions characterized by single embryo transcriptomics

New, easy, quick and efficient DNA replication timing analysis by high-throughput approaches

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