-
Nature Communications Oct 2022The pathways involved in suppressing DNA replication stress and the associated DNA damage are critical to maintaining genome integrity. The Mre11 complex is unique among...
The pathways involved in suppressing DNA replication stress and the associated DNA damage are critical to maintaining genome integrity. The Mre11 complex is unique among double strand break (DSB) repair proteins for its association with the DNA replication fork. Here we show that Mre11 complex inactivation causes DNA replication stress and changes in the abundance of proteins associated with nascent DNA. One of the most highly enriched proteins at the DNA replication fork upon Mre11 complex inactivation was the ubiquitin like protein ISG15. Mre11 complex deficiency and drug induced replication stress both led to the accumulation of cytoplasmic DNA and the subsequent activation of innate immune signaling via cGAS-STING-Tbk1. This led to ISG15 induction and protein ISGylation, including constituents of the replication fork. ISG15 plays a direct role in preventing replication stress. Deletion of ISG15 was associated with replication fork stalling, tonic ATR activation, genomic aberrations, and sensitivity to aphidicolin. These data reveal a previously unrecognized role for ISG15 in mitigating DNA replication stress and promoting genomic stability.
Topics: Aphidicolin; DNA; DNA Damage; DNA Repair; DNA Replication; Nucleotidyltransferases; Ubiquitins
PubMed: 36216822
DOI: 10.1038/s41467-022-33535-y -
Nature Communications Jun 2021Safeguards against excess DNA replication are often dysregulated in cancer, and driving cancer cells towards over-replication is a promising therapeutic strategy. We...
Safeguards against excess DNA replication are often dysregulated in cancer, and driving cancer cells towards over-replication is a promising therapeutic strategy. We determined DNA synthesis patterns in cancer cells undergoing partial genome re-replication due to perturbed regulatory interactions (re-replicating cells). These cells exhibited slow replication, increased frequency of replication initiation events, and a skewed initiation pattern that preferentially reactivated early-replicating origins. Unlike in cells exposed to replication stress, which activated a novel group of hitherto unutilized (dormant) replication origins, the preferred re-replicating origins arose from the same pool of potential origins as those activated during normal growth. Mechanistically, the skewed initiation pattern reflected a disproportionate distribution of pre-replication complexes on distinct regions of licensed chromatin prior to replication. This distinct pattern suggests that circumventing the strong inhibitory interactions that normally prevent excess DNA synthesis can occur via at least two pathways, each activating a distinct set of replication origins.
Topics: Cell Cycle Proteins; Cell Line, Tumor; Cyclopentanes; DNA Replication; Genome, Human; Humans; Mitosis; Models, Biological; Pyrimidines; Replication Origin
PubMed: 34103496
DOI: 10.1038/s41467-021-23835-0 -
ELife Jul 2022Replisomes are multi-protein complexes that replicate genomes with remarkable speed and accuracy. Despite their importance, their dynamics is poorly characterized,...
Replisomes are multi-protein complexes that replicate genomes with remarkable speed and accuracy. Despite their importance, their dynamics is poorly characterized, especially in vivo. In this paper, we present an approach to infer the replisome dynamics from the DNA abundance distribution measured in a growing bacterial population. Our method is sensitive enough to detect subtle variations of the replisome speed along the genome. As an application, we experimentally measured the DNA abundance distribution in populations growing at different temperatures using deep sequencing. We find that the average replisome speed increases nearly fivefold between 17 °C and 37 °C. Further, we observe wave-like variations of the replisome speed along the genome. These variations correlate with previously observed variations of the mutation rate, suggesting a common dynamical origin. Our approach has the potential to elucidate replication dynamics in mutants and in other bacterial species.
Topics: Chromosomes; DNA; DNA Replication; DNA, Bacterial; Escherichia coli; Escherichia coli Proteins
PubMed: 35877175
DOI: 10.7554/eLife.75884 -
Genes & Development Aug 2016For more than three decades, investigators have sought to identify the precise locations where DNA replication initiates in mammalian genomes. The development of... (Review)
Review
For more than three decades, investigators have sought to identify the precise locations where DNA replication initiates in mammalian genomes. The development of molecular and biochemical approaches to identify start sites of DNA replication (origins) based on the presence of defining and characteristic replication intermediates at specific loci led to the identification of only a handful of mammalian replication origins. The limited number of identified origins prevented a comprehensive and exhaustive search for conserved genomic features that were capable of specifying origins of DNA replication. More recently, the adaptation of origin-mapping assays to genome-wide approaches has led to the identification of tens of thousands of replication origins throughout mammalian genomes, providing an unprecedented opportunity to identify both genetic and epigenetic features that define and regulate their distribution and utilization. Here we summarize recent advances in our understanding of how primary sequence, chromatin environment, and nuclear architecture contribute to the dynamic selection and activation of replication origins across diverse cell types and developmental stages.
Topics: Animals; DNA Replication; Mammals; Replication Origin
PubMed: 27542827
DOI: 10.1101/gad.285114.116 -
Scientific Reports Jun 2022The spatiotemporal organization of DNA replication produces a highly robust and reproducible replication timing profile. Sequencing-based methods for assaying...
The spatiotemporal organization of DNA replication produces a highly robust and reproducible replication timing profile. Sequencing-based methods for assaying replication timing genome-wide have become commonplace, but regions of high repeat content in the human genome have remained refractory to analysis. Here, we report the first nearly-gapless telomere-to-telomere replication timing profiles in human, using the T2T-CHM13 genome assembly and sequencing data for five cell lines. We find that replication timing can be successfully assayed in centromeres and large blocks of heterochromatin. Centromeric regions replicate in mid-to-late S-phase and contain replication-timing peaks at a similar density to other genomic regions, while distinct families of heterochromatic satellite DNA differ in their bias for replicating in late S-phase. The high degree of consistency in centromeric replication timing across chromosomes within each cell line prompts further investigation into the mechanisms dictating that some cell lines replicate their centromeres earlier than others, and what the consequences of this variation are.
Topics: Centromere; DNA Replication; DNA Replication Timing; Genome, Human; Humans; Telomere
PubMed: 35688856
DOI: 10.1038/s41598-022-13638-8 -
Molecules (Basel, Switzerland) Oct 2019Cells must faithfully duplicate their DNA in the genome to pass their genetic information to the daughter cells. To maintain genomic stability and integrity,... (Review)
Review
Cells must faithfully duplicate their DNA in the genome to pass their genetic information to the daughter cells. To maintain genomic stability and integrity, double-strand DNA has to be replicated in a strictly regulated manner, ensuring the accuracy of its copy number, integrity and epigenetic modifications. However, DNA is constantly under the attack of DNA damage, among which oxidative DNA damage is the one that most frequently occurs, and can alter the accuracy of DNA replication, integrity and epigenetic features, resulting in DNA replication stress and subsequent genome and epigenome instability. In this review, we summarize DNA damage-induced replication stress, the formation of DNA secondary structures, peculiar epigenetic modifications and cellular responses to the stress and their impact on the instability of the genome and epigenome mainly in eukaryotic cells.
Topics: DNA; DNA Damage; DNA Methylation; DNA Replication; Epigenesis, Genetic; Epigenome; Histones; Humans
PubMed: 31717862
DOI: 10.3390/molecules24213870 -
International Journal of Molecular... Dec 2017Within the nucleus, sub-nuclear domains define territories where specific functions occur. Nuclear bodies (NBs) are dynamic structures that concentrate nuclear factors... (Review)
Review
Within the nucleus, sub-nuclear domains define territories where specific functions occur. Nuclear bodies (NBs) are dynamic structures that concentrate nuclear factors and that can be observed microscopically. Recently, NBs containing the p53 binding protein 1 (53BP1), a key component of the DNA damage response, were defined. Interestingly, 53BP1 NBs are visualized during G1 phase, in daughter cells, while DNA damage was generated in mother cells and not properly processed. Unlike most NBs involved in transcriptional processes, replication has proven to be key for 53BP1 NBs, with replication stress leading to the formation of these large chromatin domains in daughter cells. In this review, we expose the composition and organization of 53BP1 NBs and focus on recent findings regarding their regulation and dynamics. We then concentrate on the importance of the replication stress, examine the relation of 53BP1 NBs with DNA damage and discuss their dysfunction.
Topics: Animals; Cell Nucleus; Chromatin; DNA Damage; DNA Replication; Humans; Tumor Suppressor p53-Binding Protein 1
PubMed: 29206178
DOI: 10.3390/ijms18122611 -
Open Biology Aug 2023Metabolism and DNA replication are the two most fundamental biological functions in life. The catabolic branch of metabolism breaks down nutrients to produce energy and... (Review)
Review
Metabolism and DNA replication are the two most fundamental biological functions in life. The catabolic branch of metabolism breaks down nutrients to produce energy and precursors used by the anabolic branch of metabolism to synthesize macromolecules. DNA replication consumes energy and precursors for faithfully copying genomes, propagating the genetic material from generation to generation. We have exquisite understanding of the mechanisms that underpin and regulate these two biological functions. However, the molecular mechanism coordinating replication to metabolism and its biological function remains mostly unknown. Understanding how and why living organisms respond to fluctuating nutritional stimuli through cell-cycle dynamic changes and reproducibly and distinctly temporalize DNA synthesis in a wide-range of growth conditions is important, with wider implications across all domains of life. After summarizing the seminal studies that founded the concept of the metabolic control of replication, we review data linking metabolism to replication from bacteria to humans. Molecular insights underpinning these links are then presented to propose that the metabolic control of replication uses signalling systems gearing metabolome homeostasis to orchestrate replication temporalization. The remarkable replication phenotypes found in mutants of this control highlight its importance in replication regulation and potentially genetic stability and tumorigenesis.
Topics: Humans; DNA Replication; Cell Cycle; Cell Division
PubMed: 37582405
DOI: 10.1098/rsob.230220 -
Chromosome Research : An International... Mar 2020Genomic DNA replicates according to a defined temporal program in which early-replicating loci are associated with open chromatin, higher gene density, and increased... (Review)
Review
Genomic DNA replicates according to a defined temporal program in which early-replicating loci are associated with open chromatin, higher gene density, and increased gene expression levels, while late-replicating loci tend to be heterochromatic and show higher rates of genomic instability. The ability to measure DNA replication dynamics at genome scale has proven crucial for understanding the mechanisms and cellular consequences of DNA replication timing. Several methods, such as quantification of nucleotide analog incorporation and DNA copy number analyses, can accurately reconstruct the genomic replication timing profiles of various species and cell types. More recent developments have expanded the DNA replication genomic toolkit to assays that directly measure the activity of replication origins, while single-cell replication timing assays are beginning to reveal a new level of replication timing regulation. The combination of these methods, applied on a genomic scale and in multiple biological systems, promises to resolve many open questions and lead to a holistic understanding of how eukaryotic cells replicate their genomes accurately and efficiently.
Topics: Animals; Chromosome Mapping; DNA Replication; DNA Replication Timing; Eukaryotic Cells; Genome; Genome-Wide Association Study; Genomics; Humans; Replication Origin; Single-Cell Analysis
PubMed: 31848781
DOI: 10.1007/s10577-019-09624-y -
Trends in Biochemical Sciences Jan 2024DNA single-strand breaks (SSBs) are among the most common lesions arising in human cells, with tens to hundreds of thousands arising in each cell, each day. Cells have... (Review)
Review
DNA single-strand breaks (SSBs) are among the most common lesions arising in human cells, with tens to hundreds of thousands arising in each cell, each day. Cells have efficient mechanisms for the sensing and repair of these ubiquitous DNA lesions, but the failure of these processes to rapidly remove SSBs can lead to a variety of pathogenic outcomes. The threat posed by unrepaired SSBs is illustrated by the existence of at least six genetic diseases in which SSB repair (SSBR) is defective, all of which are characterised by neurodevelopmental and/or neurodegenerative pathology. Here, I review current understanding of how SSBs arise and impact on critical molecular processes, such as DNA replication and gene transcription, and their links to human disease.
Topics: Humans; DNA Repair; DNA Breaks, Single-Stranded; DNA Damage; DNA Replication; DNA
PubMed: 38040599
DOI: 10.1016/j.tibs.2023.11.001