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Series GSE211037 Query DataSets for GSE211037
Status Public on Apr 19, 2024
Title Acceleration of genome replication uncovered by single-cell nascent DNA sequencing
Organism Homo sapiens
Experiment type Other
Summary Faithful genome replication is a prerequisite for cell division. In a human cell thousands of replication forks simultaneously coordinate the duplication of the entire genome. Single-molecule approaches have been invaluable to monitor this process by incorporating nucleotide analogs during replication and subsequent detection by microscopy3 or long-read sequencing. However, these methods randomly sample molecules from a large population of cells and are therefore insensitive to heterogeneity in replication dynamics between single cells. To overcome this challenge, we developed a sequencing method to detect nascent DNA allowing the detection of replication forks and their respective speeds in single cells. By ordering cells according to overlapping replicated genome regions, we can accurately position individual cells along S-phase. We observe a pronounced variability in replication speeds among cells. Most of this variability can be explained by the position of a cell along S-phase revealing an acceleration of replication as S-phase progresses. Transcribed regions in the genome display particularly lower replication speeds suggesting a potential role for transcription. Indeed, inhibiting transcription increased replication speeds at the start of S-phase. Additionally, we find that the effect of transcription is mediated through DNA damage signaling. We suggest that the lower replication speeds at the beginning of the S phase are caused by the generation of single-strand breaks in the genome. Indeed, cells for which the single-strand break repair pathway is inactive, display lower DNA replication speeds compared to wild-type cells, whereas replication speeds increase when cells are unable to detect single-strand breaks. This novel technology opens up new avenues to quantitatively explore the relationship between replication dynamics and replication stress with single-cell resolution.
Overall design Single cell analysis of Replication fork speeds over different treatment and knockouts.
Contributor(s) van den Berg J, van Batenburg V, van Oudenaarden A
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Submission date Aug 11, 2022
Last update date Apr 20, 2024
Contact name Jeroen van den Berg
Phone +31610597913
Organization name Hubrecht Institute
Lab van Oudenaarden lab
Street address Uppsalalaan 8
City Utrecht
State/province Utrecht
ZIP/Postal code 3584 CT
Country Netherlands
Platforms (2)
GPL18573 Illumina NextSeq 500 (Homo sapiens)
GPL30173 NextSeq 2000 (Homo sapiens)
Samples (50)
GSM6447737 RPE-1 hTERT FUCCI double pulse dT45min_1
GSM6447738 RPE-1 hTERT FUCCI double pulse dT45min_2
GSM6447739 RPE-1 hTERT FUCCI double pulse dT45min_3
BioProject PRJNA868727

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Supplementary file Size Download File type/resource
GSE211037_RAW.tar 2.3 Gb (http)(custom) TAR (of TSV)
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Raw data are available in SRA
Processed data provided as supplementary file

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