Inferring the DNA replication origin density landscape in human cells from replication timing and fork directionality data

2021 ◽  
Author(s):  
Jean-Michel Arbona ◽  
Benjamin Audit ◽  
Hadi Kabalane ◽  
Olivier Hyrien ◽  
Arach Goldar
Keyword(s):  
Replication Fork ◽  
A Priori ◽  
Analytical Formula ◽  
Replication Timing ◽  
Simulation Framework ◽  
Spatial Frequencies ◽  
Genome Wide ◽  
Dna Replication Origin ◽  
Actual Initiation ◽  
Multiple Replication

The determinants of the locations and firing times of the multiple replication origins are still elusive in human and other metazoan organisms. Experiments can independently profile mean replication timing (MRT) and replication fork directionality (RFD) genome-wide. In the hypothesis of a constant replication fork speed, MRT and RFD are related to each other by an analytical formula so are a priori equivalent. However, we show here that experimental noises result in MRT and RFD profiles containing information at different spatial frequencies. We further demonstrate that one can compute an origin density landscape that, when inserted in an appropriate simulation framework, jointly predicts experimental MRT and RFD profiles with an unprecedented precision. We also extract an analytical formula linking intrinsic origin efficiency with observed origin efficiency and MRT. We then compare the computed origin density landscape with experimental distributions of potential origins (ORC, MCM) or actual initiation events (Bubble-seq, SNS-seq, OK-seq). The results indicate that MRT and RFD data are highly consistent with each other, that our simple model suffices to capture the replication dynamics during S phase given an appropriate initiation probability landscape, but that the density of potential origins is not the sole determinant of this landscape.

scholarly journals A flexible ChIP-sequencing simulation toolkit

2021 ◽  
Vol 22 (1) ◽  
Author(s):  
An Zheng ◽  
Michael Lamkin ◽  
Yutong Qiu ◽  
Kevin Ren ◽  
Alon Goren ◽  
...  
Keyword(s):  
Ground Truth ◽  
Peak Calling ◽  
Simulation Framework ◽  
Experimental Conditions ◽  
Ground Truth Data ◽  
Chip Sequencing ◽  
Genome Wide ◽  
Experimental Parameters ◽  
Differential Binding ◽  
The Impact

Abstract Background A major challenge in evaluating quantitative ChIP-seq analyses, such as peak calling and differential binding, is a lack of reliable ground truth data. Accurate simulation of ChIP-seq data can mitigate this challenge, but existing frameworks are either too cumbersome to apply genome-wide or unable to model a number of important experimental conditions in ChIP-seq. Results We present ChIPs, a toolkit for rapidly simulating ChIP-seq data using statistical models of key experimental steps. We demonstrate how ChIPs can be used for a range of applications, including benchmarking analysis tools and evaluating the impact of various experimental parameters. ChIPs is implemented as a standalone command-line program written in C++ and is available from https://github.com/gymreklab/chips. Conclusions ChIPs is an efficient ChIP-seq simulation framework that generates realistic datasets over a flexible range of experimental conditions. It can serve as an important component in various ChIP-seq analyses where ground truth data are needed.

scholarly journals Repli-seq: genome-wide analysis of replication timing by next-generation sequencing

2017 ◽  
Author(s):  
Claire Marchal ◽  
Takayo Sasaki ◽  
Daniel Vera ◽  
Korey Wilson ◽  
Jiao Sima ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Replication Timing ◽  
Nucleotide Polymorphisms ◽  
Robust Methods ◽  
Next Generation ◽  
Single Nucleotide ◽  
Cellular Processes ◽  
Genome Wide ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

ABSTRACTCycling cells duplicate their DNA content during S phase, following a defined program called replication timing (RT). Early and late replicating regions differ in terms of mutation rates, transcriptional activity, chromatin marks and sub-nuclear position. Moreover, RT is regulated during development and is altered in disease. Exploring mechanisms linking RT to other cellular processes in normal and diseased cells will be facilitated by rapid and robust methods with which to measure RT genome wide. Here, we describe a rapid, robust and relatively inexpensive protocol to analyze genome-wide RT by next-generation sequencing (NGS). This protocol yields highly reproducible results across laboratories and platforms. We also provide computational pipelines for analysis, parsing phased genomes using single nucleotide polymorphisms (SNP) for analyzing RT allelic asynchrony, and for direct comparison to Repli-chip data obtained by analyzing nascent DNA by microarrays.

scholarly journals Delineation of the Ancestral Tus-Dependent Replication Fork Trap

2021 ◽  
Author(s):  
Casey Toft ◽  
Morgane Moreau ◽  
Jiri Perutka ◽  
Savitri Mandapati ◽  
Peter Enyeart ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Replication ◽  
Replication Fork ◽  
Wide Distribution ◽  
Replication Forks ◽  
Genome Wide ◽  
Replication Fork Arrest

In Escherichia coli, DNA replication termination is orchestrated by two clusters of Ter sites forming a DNA replication fork trap when bound by Tus proteins. The formation of a ‘locked’ Tus-Ter complex is essential for halting incoming DNA replication forks. However, the absence of replication fork arrest at some Ter sites raised questions about their significance. In this study, we examined the genome-wide distribution of Tus and found that only the six innermost Ter sites (TerA-E and G) were significantly bound by Tus. We also found that a single ectopic insertion of TerB in its non-permissive orientation could not be achieved, advocating against a need for ‘back-up’ Ter sites. Finally, examination of the genomes of a variety of Enterobacterales revealed a new replication fork trap architecture mostly found outside the Enterobacteriaceae family. Taken together, our data enabled the delineation of a narrow ancestral Tus-dependent DNA replication fork trap consisting of only two Ter sites.

scholarly journals Nascent chromatin occupancy profiling reveals locus and factor specific chromatin maturation dynamics behind the DNA replication fork

2018 ◽  
Author(s):  
Mónica P. Gutiérrez ◽  
Heather K. MacAlpine ◽  
David M. MacAlpine
Keyword(s):  
Dna Replication ◽  
Replication Fork ◽  
Histone Variants ◽  
Replication Forks ◽  
Nucleosome Assembly ◽  
Genome Wide ◽  
Dna Binding Factors ◽  
Nucleotide Resolution ◽  
Kinetics Of ◽  
Regulatory Landscape

AbstractProper regulation and maintenance of the epigenome is necessary to preserve genome function. However, in every cell division, the epigenetic state is disassembled and then re-assembled in the wake of the DNA replication fork. Chromatin restoration on nascent DNA is a complex and regulated process that includes nucleosome assembly and remodeling, deposition of histone variants, and the re-establishment of transcription factor binding. To study the genome-wide dynamics of chromatin restoration behind the DNA replication fork, we developed Nascent Chromatin Occupancy Profiles (NCOPs) to comprehensively profile nascent and mature chromatin at nucleotide resolution. While nascent chromatin is inherently less organized than mature chromatin, we identified locus specific differences in the kinetics of chromatin maturation that were predicted by the epigenetic landscape, including the histone variant H2A.Z which marked loci with rapid maturation kinetics. The chromatin maturation at origins of DNA replication was dependent on whether the origin underwent initiation or was passively replicated from distal-originating replication forks suggesting distinct chromatin assembly mechanisms between activated and disassembled pre-replicative complexes. Finally, we identified sites that were only occupied transiently by DNA-binding factors following passage of the replication fork which may provide a mechanism for perturbations of the DNA replication program to shape the regulatory landscape of the genome.

scholarly journals Genome-Wide Analysis of the Arabidopsis Replication Timing Program

2018 ◽  
Vol 176 (3) ◽  
pp. 2166-2185 ◽  
Author(s):  
Lorenzo Concia ◽  
Ashley M. Brooks ◽  
Emily Wheeler ◽  
Gregory J. Zynda ◽  
Emily E. Wear ◽  
...  
Keyword(s):  
Replication Timing ◽  
Genome Wide Analysis ◽  
Genome Wide

Antibody-free profiling of transcription factor occupancy during early embryogenesis by FitCUT&RUN

2021 ◽  
pp. gr.275837.121
Author(s):  
Xiangxiu Wang ◽  
Wen Wang ◽  
Yiman Wang ◽  
Jia Chen ◽  
Guifen Liu ◽  
...  
Keyword(s):  
Significant Proportion ◽  
K562 Cells ◽  
Replication Timing ◽  
Early Embryogenesis ◽  
Clear Trend ◽  
Genome Wide ◽  
Sequential Binding ◽  
Reliability And Robustness ◽  
Genome Activation ◽  
Transcription Factor Occupancy

Key transcription factors (TFs) play critical roles in zygotic genome activation (ZGA) during early embryogenesis, while genome-wide occupancies of only a few factors have been profiled during ZGA due to the limitation of cell numbers or the lack of high-quality antibodies. Here, we present FitCUT&RUN, a modified CUT&RUN method, in which an Fc fragment of immunoglobulin G is used for tagging, to profile TF occupancy in an antibody-free manner and demonstrate its reliability and robustness using as few as five thousand K562 cells. We applied FitCUT&RUN to zebrafish undergoing embryogenesis to generate reliable occupancy profiles of three known activators of zebrafish ZGA: Nanog, Pou5f3 and Sox19b. By profiling the time-series occupancy of Nanog during zebrafish ZGA, we observed a clear trend toward a gradual increase in Nanog occupancy and found that Nanog occupancy prior to the major phase of ZGA is critical for the activation of a significant proportion of early transcribed genes. Our results further suggested that the sequential binding of Nanog may be controlled by replication timing and the presence of Nanog motifs.

Mapping replication timing domains genome wide in single mammalian cells with single-cell DNA replication sequencing

2020 ◽  
Vol 15 (12) ◽  
pp. 4058-4100
Author(s):  
Hisashi Miura ◽  
Saori Takahashi ◽  
Takahiro Shibata ◽  
Koji Nagao ◽  
Chikashi Obuse ◽  
...  
Keyword(s):  
Dna Replication ◽  
Single Cell ◽  
Mammalian Cells ◽  
Replication Timing ◽  
Genome Wide

scholarly journals Multiple replication origins with diverse control mechanisms in Haloarcula hispanica

2013 ◽  
Vol 42 (4) ◽  
pp. 2282-2294 ◽  
Author(s):  
Zhenfang Wu ◽  
Jingfang Liu ◽  
Haibo Yang ◽  
Hailong Liu ◽  
Hua Xiang
Keyword(s):  
Replication Initiation ◽  
Control Mechanisms ◽  
Replication Origins ◽  
Dna Mutation ◽  
Genome Wide ◽  
Marker Frequency ◽  
Haloarcula Hispanica ◽  
Specific Control ◽  
Main Chromosome ◽  
Multiple Replication

Abstract The use of multiple replication origins in archaea is not well understood. In particular, little is known about their specific control mechanisms. Here, we investigated the active replication origins in the three replicons of a halophilic archaeon, Haloarcula hispanica, by extensive gene deletion, DNA mutation and genome-wide marker frequency analyses. We revealed that individual origins are specifically dependent on their co-located cdc6 genes, and a single active origin/cdc6 pairing is essential and sufficient for each replicon. Notably, we demonstrated that the activities of oriC1 and oriC2, the two origins on the main chromosome, are differently controlled. A G-rich inverted repeat located in the internal region between the two inverted origin recognition boxes (ORBs) plays as an enhancer for oriC1, whereas the replication initiation at oriC2 is negatively regulated by an ORB-rich region located downstream of oriC2-cdc6E, likely via Cdc6E-titrating. The oriC2 placed on a plasmid is incompatible with the wild-type (but not the ΔoriC2) host strain, further indicating that strict control of the oriC2 activity is important for the cell. This is the first report revealing diverse control mechanisms of origins in haloarchaea, which has provided novel insights into the use and coordination of multiple replication origins in the domain of Archaea.

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