The genetic architecture of DNA replication timing in human pluripotent stem cells

AbstractDNA replication follows a strict spatiotemporal program that intersects with chromatin structure but has a poorly understood genetic basis. To systematically identify genetic regulators of replication timing, we exploited inter-individual variation in human pluripotent stem cells from 349 individuals. We show that the human genome’s replication program is broadly encoded in DNA and identify 1,617 cis-acting replication timing quantitative trait loci (rtQTLs) – sequence determinants of replication initiation. rtQTLs function individually, or in combinations of proximal and distal regulators, and are enriched at sites of histone H3 trimethylation of lysines 4, 9, and 36 together with histone hyperacetylation. H3 trimethylation marks are individually repressive yet synergistically associate with early replication. We identify pluripotency-related transcription factors and boundary elements as positive and negative regulators of replication timing, respectively. Taken together, human replication timing is controlled by a multi-layered mechanism with dozens of effectors working combinatorially and following principles analogous to transcription regulation.

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The Genetic Architecture of DNA Replication Timing in Human Pluripotent Stem Cells

10.1101/2020.05.08.085324 ◽

2020 ◽

Cited By ~ 1

Author(s):

Qiliang Ding ◽

Matthew M. Edwards ◽

Michelle L. Hulke ◽

Alexa N. Bracci ◽

Ya Hu ◽

...

Keyword(s):

Dna Replication ◽

Dna Sequences ◽

Timing Analysis ◽

Replication Timing ◽

Histone Code ◽

Replication Initiation ◽

Histone Hyperacetylation ◽

Stem Cell Lines ◽

Early Replication ◽

Dna Replication Timing

AbstractDNA replication follows a strict spatiotemporal program that intersects with chromatin structure and gene regulation. However, the genetic basis of the mammalian DNA replication timing program is poorly understood1–3. To systematically identify genetic regulators of DNA replication timing, we exploited inter-individual variation in 457 human pluripotent stem cell lines from 349 individuals. We show that the human genome’s replication program is broadly encoded in DNA and identify 1,617 cis-acting replication timing quantitative trait loci (rtQTLs4) – base-pair-resolution sequence determinants of replication initiation. rtQTLs function individually, or in combinations of proximal and distal regulators, to affect replication timing. Analysis of rtQTL locations reveals a histone code for replication initiation, composed of bivalent histone H3 trimethylation marks on a background of histone hyperacetylation. The H3 trimethylation marks are individually repressive yet synergize to promote early replication. We further identify novel positive and negative regulators of DNA replication timing, the former comprised of pluripotency-related transcription factors while the latter involve boundary elements. Human replication timing is controlled by a multi-layered mechanism that operates on target DNA sequences, is composed of dozens of effectors working combinatorially, and follows principles analogous to transcription regulation: a histone code, activators and repressors, and a promoter-enhancer logic.

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Camptothecin-Induced Replication Stress Affects DNA Replication Profiling by E/L Repli-Seq

Cytogenetic and Genome Research ◽

10.1159/000518263 ◽

2021 ◽

pp. 1-8

Author(s):

Takuya Hayakawa ◽

Rino Suzuki ◽

Kazuhiro Kagotani ◽

Katsuzumi Okumura ◽

Shin-ichiro Takebayashi

Keyword(s):

Dna Replication ◽

Mammalian Cells ◽

Topoisomerase I ◽

Pigment Epithelium ◽

Retinal Pigment ◽

Replication Timing ◽

Replication Stress ◽

Replication Initiation ◽

Domain Structures ◽

Early Replication

E/L Repli-seq is a powerful tool for detecting cell type-specific replication landscapes in mammalian cells, but its potential to monitor DNA replication under replication stress awaits better understanding. Here, we used E/L Repli-seq to examine the temporal order of DNA replication in human retinal pigment epithelium cells treated with the topoisomerase I inhibitor camptothecin. We found that the replication profiles by E/L Repli-seq exhibit characteristic patterns after replication-stress induction, including the loss of specific initiation zones within individual early replication timing domains. We also observed global disappearance of the replication timing domain structures in the profiles, which can be explained by checkpoint-dependent suppression of replication initiation. Thus, our results demonstrate the effectiveness of E/L Repli-seq at identifying cells with replication-stress-induced altered DNA replication programs.

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DNA Fiber Assay for the Analysis of DNA Replication Progression in Human Pluripotent Stem Cells

Current Protocols in Stem Cell Biology ◽

10.1002/cpsc.115 ◽

2020 ◽

Vol 54 (1) ◽

Author(s):

Jason A. Halliwell ◽

Polly Gravells ◽

Helen E. Bryant

Keyword(s):

Stem Cells ◽

Dna Replication ◽

Pluripotent Stem Cells ◽

Human Pluripotent Stem Cells

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Fused in sarcoma regulates DNA replication timing and progression

10.1101/2020.04.22.055343 ◽

2020 ◽

Author(s):

Weiyan Jia ◽

Mark A. Scalf ◽

Peter Tonzi ◽

Robert J. Millikin ◽

Sang Hwa Kim ◽

...

Keyword(s):

Dna Replication ◽

Transcriptional Activation ◽

Rna Binding ◽

Soft Tissue Sarcomas ◽

Replication Timing ◽

Strand Break ◽

Replication Initiation ◽

Proliferative Potential ◽

Fused In Sarcoma ◽

Dna Replication Timing

AbstractFused in sarcoma (FUS) encodes a low complexity RNA-binding protein with diverse roles in transcriptional activation and RNA processing. While oncogenic fusions of FUS and transcription factor DNA-binding domains are associated with soft tissue sarcomas, dominant mutations in FUS cause amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). FUS has also been implicated in DNA double-strand break repair (DSBR) and genome maintenance. However, the underlying mechanisms are unknown. Here we employed quantitative proteomics, transcriptomics, and DNA copy number analysis (Sort-Seq), in conjunction with FUS-/- cells to ascertain roles of FUS in genome protection. FUS-/- cells exhibited alterations in the recruitment and retention of DSBR factors BRCA1 and 53BP1 but were not overtly sensitive to genotoxins. By contrast, FUS-deficient cells had reduced proliferative potential that correlated with reduced replication fork speed, diminished loading of pre-replication complexes, and attenuated expression of S-phase associated genes. FUS interacted with lagging strand DNA synthesis factors and other replisome components, but did not translocate with active replication forks. Using a Sort-Seq workflow, we show that FUS contributes to genome-wide control of DNA replication timing and is essential for the early replication of transcriptionally active DNA. These findings illuminate new roles for FUS in DNA replication initiation and timing that may contribute to genome instability and functional defects in cells harboring disease-associated FUS fusions.

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Dynamic changes in replication timing and gene expression during lineage specification of human pluripotent stem cells

Genome Research ◽

10.1101/gr.187989.114 ◽

2015 ◽

Vol 25 (8) ◽

pp. 1091-1103 ◽

Cited By ~ 81

Author(s):

Juan Carlos Rivera-Mulia ◽

Quinton Buckley ◽

Takayo Sasaki ◽

Jared Zimmerman ◽

Ruth A. Didier ◽

...

Keyword(s):

Gene Expression ◽

Stem Cells ◽

Pluripotent Stem Cells ◽

Replication Timing ◽

Human Pluripotent Stem Cells ◽

Lineage Specification ◽

Dynamic Changes

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Continuous chromosomal instability in human pluripotent stem cells – the role of DNA replication

Molecular & Cellular Oncology ◽

10.1080/23723556.2016.1183743 ◽

2016 ◽

Vol 3 (4) ◽

pp. e1183743

Author(s):

Noa Lamm ◽

Batsheva Kerem

Keyword(s):

Stem Cells ◽

Dna Replication ◽

Pluripotent Stem Cells ◽

Chromosomal Instability ◽

Human Pluripotent Stem Cells

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Comprehensive analysis of DNA replication timing in genetic diseases and gene knockouts identifies MCM10 as a novel regulator of the replication program

10.1101/2021.09.08.459433 ◽

2021 ◽

Author(s):

Madison Caballero ◽

Tiffany Ge ◽

Ana Rita Rebelo ◽

Seungmae Seo ◽

Sean Kim ◽

...

Keyword(s):

Dna Replication ◽

Cellular Proliferation ◽

Genetic Diseases ◽

Replication Timing ◽

Cell Types ◽

Replication Initiation ◽

Timing Variability ◽

Gene Knockouts ◽

Dna Replication Timing ◽

Mutant Cells

AbstractCellular proliferation depends on the accurate and timely replication of the genome. Several genetic diseases are caused by mutations in key DNA replication genes; however, it remains unclear whether these genes influence the normal program of DNA replication timing. Similarly, the factors that regulate DNA replication dynamics are poorly understood. To systematically identify trans-acting modulators of replication timing, we profiled replication in 184 cell lines from three cell types, encompassing 60 different gene knockouts or genetic diseases. Through a rigorous approach that considers the background variability of replication timing, we concluded that most samples displayed normal replication timing. However, mutations in two genes showed consistently abnormal replication timing. The first gene was RIF1, a known modulator of replication timing. The second was MCM10, a highly conserved member of the pre-replication complex. MCM10 mutant cells demonstrated replication timing variability comprising 46% of the genome and at different locations than RIF1 knockouts. Replication timing alterations in MCM10-mutant cells was predominantly comprised of replication initiation defects. Taken together, this study demonstrates the remarkable robustness of the human replication timing program and reveals MCM10 as a novel modulator of DNA replication timing.

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