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Trends in Genetics : TIG Feb 2022Cells activate distinctive regulatory pathways that prevent excessive initiation of DNA replication to achieve timely and accurate genome duplication. Excess DNA... (Review)
Review
Cells activate distinctive regulatory pathways that prevent excessive initiation of DNA replication to achieve timely and accurate genome duplication. Excess DNA synthesis is constrained by protein-DNA interactions that inhibit initiation at dormant origins. In parallel, specific modifications of pre-replication complexes prohibit post-replicative origin relicensing. Replication stress ensues when the controls that prevent excess replication are missing in cancer cells, which often harbor extrachromosomal DNA that can be further amplified by recombination-mediated processes to generate chromosomal translocations. The genomic instability that accompanies excess replication origin activation can provide a promising target for therapeutic intervention. Here we review molecular pathways that modulate replication origin dormancy, prevent excess origin activation, and detect, encapsulate, and eliminate persistent excess DNA.
Topics: DNA; DNA Damage; DNA Replication; Genomic Instability; Humans; Replication Origin
PubMed: 34625299
DOI: 10.1016/j.tig.2021.09.008 -
Biochimica Et Biophysica Acta. Reviews... Aug 2017Cell division is a tightly-regulated process that involves the contribution of a large number of proteins. Before they are able to undergo mitosis, cells must first... (Review)
Review
Cell division is a tightly-regulated process that involves the contribution of a large number of proteins. Before they are able to undergo mitosis, cells must first synthesize new DNA, effectively and accurately duplicating their genome. This occurs during what is called the S-phase and requires a fine control in order to avoid replication errors. The synthesis of new DNA takes place in the origin, specific locations in the genome where the double strands of DNA are unwound and separated, allowing for the binding of proteins and complexes that will build new strands of the genomic material, using the existing ones as molds, in what is referred to as semi-conservative process. While the overall flow of the DNA synthesis process has been elucidated, its regulation and the exact role of its contributors are not yet entirely understood. It is believed that the Minichromosome Maintenance (MCM) proteins occupy a central role in DNA synthesis. Given their contribution to a central aspect in the conservation of life, further studies have been launched to understand how the MCM proteins may affect or be affected by pathologies involving cell division, such as neoplasia. In this review, we aim to give an overview on the members of the MCM family, what their functions are in a healthy environment and how they are altered in cancer.
Topics: Animals; Cell Cycle Proteins; DNA; DNA Replication; DNA-Binding Proteins; Humans; Minichromosome Maintenance Proteins; S Phase
PubMed: 28579200
DOI: 10.1016/j.bbcan.2017.06.001 -
Advances in Experimental Medicine and... 2017Successful DNA replication requires intimate coordination with cell-cycle progression. Prior to DNA replication initiation in S phase, a series of essential preparatory... (Review)
Review
Successful DNA replication requires intimate coordination with cell-cycle progression. Prior to DNA replication initiation in S phase, a series of essential preparatory events in G phase ensures timely, complete, and precise genome duplication. Among the essential molecular processes are regulated transcriptional upregulation of genes that encode replication proteins, appropriate post-transcriptional control of replication factor abundance and activity, and assembly of DNA-loaded protein complexes to license replication origins. In this chapter we describe these critical G events necessary for DNA replication and their regulation in the context of both cell-cycle entry and cell-cycle progression.
Topics: Animals; Cell Cycle; Cell Cycle Proteins; Cyclin-Dependent Kinases; DNA Replication; G1 Phase; Humans; Replication Origin; Retinoblastoma Protein; S Phase
PubMed: 29357066
DOI: 10.1007/978-981-10-6955-0_16 -
Current Opinion in Genetics &... Apr 2023Decades of work on the spatiotemporal organization of mammalian DNA replication timing (RT) continues to unveil novel correlations with aspects of transcription and... (Review)
Review
Decades of work on the spatiotemporal organization of mammalian DNA replication timing (RT) continues to unveil novel correlations with aspects of transcription and chromatin organization but, until recently, mechanisms regulating RT and the biological significance of the RT program had been indistinct. We now know that the RT program is both influenced by and necessary to maintain chromatin structure, forming an epigenetic positive feedback loop. Moreover, the discovery of specific cis-acting elements regulating mammalian RT at both the domain and the whole-chromosome level has revealed multiple cell-type-specific and developmentally regulated mechanisms of RT control. We review recent evidence for diverse mechanisms employed by different cell types to regulate their RT programs and the biological significance of RT regulation during development.
Topics: Animals; DNA Replication Timing; Chromatin; DNA Replication; Mammals
PubMed: 36905782
DOI: 10.1016/j.gde.2023.102031 -
Current Protocols Jan 2022Replication timing (RT) is the temporal order in which genomic DNA is replicated during S phase. Early and late replication correlate with transcriptionally active and...
Replication timing (RT) is the temporal order in which genomic DNA is replicated during S phase. Early and late replication correlate with transcriptionally active and inactive chromatin compartments, but mechanistic links between large-scale chromosome structure, transcription, and replication are still enigmatic. A proper RT program is necessary to maintain the global epigenome that defines cell identity, suggesting that RT is critical for epigenome integrity by facilitating the assembly of different types of chromatin at different times during S phase. RT is regulated during development and has been found to be altered in disease. Thus, RT can identify stable epigenetic differences distinguishing cell types, and can be used to help stratify patient outcomes and identify markers of disease. Most methods to profile RT require thousands of S-phase cells. In cases where cells are rare (e.g., early-stage embryos or rare primary cell types) or consist of a heterogeneous mixture of cell states (e.g., differentiation intermediates), or when the interest is in determining the degree of stable epigenetic heterogeneity within a population of cells, single-cell measurements of RT are necessary. We have previously developed single cell Repli-seq, a method to measure replication timing in single cells using DNA copy number quantification. To date, however, single-cell Repli-seq suffers from relatively low throughput and high costs. Here, we describe an improved single-cell Repli-seq protocol that uses degenerate oligonucleotide-primed PCR (DOP-PCR) for uniform whole-genome amplification and uniquely barcoded primers that permit early pooling of single-cell samples into a single library preparation. We also provide a bioinformatics platform for analysis of the data. The improved throughput and decreased costs of this method relative to previously published single-cell Repli-seq protocols should make it considerably more accessible to a broad range of investigators. © 2022 Wiley Periodicals LLC. Basic Protocol 1: Whole Genome Amplification (WGA) of single cells and sequence library construction. Basic Protocol 2: Deriving and displaying single-cell replication timing data from whole genome sequencing.
Topics: Animals; DNA; DNA Replication; DNA Replication Timing; Humans; S Phase; Sequence Analysis, DNA
PubMed: 34986273
DOI: 10.1002/cpz1.334 -
Advances in Experimental Medicine and... 2017Accurate genome duplication during cell division is essential for life. This process is accomplished by the close collaboration between replication factors and many... (Review)
Review
Accurate genome duplication during cell division is essential for life. This process is accomplished by the close collaboration between replication factors and many additional proteins that provide assistant roles. Replication factors establish the replication machineries capable of copying billions of nucleotides, while regulatory proteins help to achieve accuracy and efficiency of replication. Among regulatory proteins, protein modification enzymes can bestow fast and reversible changes to many targets, leading to coordinated effects on replication. Recent studies have begun to elucidate how one type of protein modification, sumoylation, can modify replication proteins and regulate genome duplication through multiple mechanisms. This chapter summarizes these new findings, and how they can integrate with the known regulatory circuitries of replication. As this area of research is still at its infancy, many outstanding questions remain to be explored, and we discuss these issues in light of the new advances.
Topics: Animals; DNA Replication; Humans; Origin Recognition Complex; Protein Processing, Post-Translational; Replication Origin; SUMO-1 Protein; Sumoylation
PubMed: 29357067
DOI: 10.1007/978-981-10-6955-0_17 -
Genes To Cells : Devoted To Molecular &... Feb 2021Replication initiation, elongation and completion are tightly coordinated to ensure that all sequences replicate precisely once each generation. UV-induced DNA damage...
Replication initiation, elongation and completion are tightly coordinated to ensure that all sequences replicate precisely once each generation. UV-induced DNA damage disrupts replication and delays elongation, which may compromise this coordination leading to genome instability and cell death. Here, we profiled the Escherichia coli genome as it recovers from UV irradiation to determine how these replicational processes respond. We show that oriC initiations continue to occur, leading to copy number enrichments in this region. At late times, the combination of new oriC initiations and delayed elongating forks converging in the terminus appear to stress or impair the completion reaction, leading to a transient over-replication in this region of the chromosome. In mutants impaired for restoring elongation, including recA, recF and uvrA, the genome degrades or remains static, suggesting that cell death occurs early after replication is disrupted, leaving partially duplicated genomes. In mutants impaired for completing replication, including recBC, sbcCD xonA and recG, the recovery of elongation and initiation leads to a bottleneck, where the nonterminus region of the genome is amplified and accumulates, indicating that a delayed cell death occurs in these mutants, likely resulting from mis-segregation of unbalanced or unresolved chromosomes when cells divide.
Topics: Chromosomes, Bacterial; DNA Damage; DNA Repair; DNA Replication; Escherichia coli; Escherichia coli Proteins; Gene Dosage; Genome, Bacterial; Mutation; Ultraviolet Rays
PubMed: 33382157
DOI: 10.1111/gtc.12826 -
Protoplasma May 2017Scientific discoveries and technological advancements are inseparable but not always take place in a coherent chronological manner. In the next, we will provide a... (Review)
Review
Scientific discoveries and technological advancements are inseparable but not always take place in a coherent chronological manner. In the next, we will provide a seemingly unconnected and serendipitous series of scientific facts that, in the whole, converged to unveil DNA and its duplication. We will not cover here the many and fundamental contributions from microbial genetics and in vitro biochemistry. Rather, in this journey, we will emphasize the interplay between microscopy development culminating on super resolution fluorescence microscopy (i.e., nanoscopy) and digital image analysis and its impact on our understanding of DNA duplication. We will interlace the journey with landmark concepts and experiments that have brought the cellular DNA replication field to its present state.
Topics: DNA; DNA Replication; Green Fluorescent Proteins; History, 19th Century; History, 20th Century; History, 21st Century; Humans; Image Processing, Computer-Assisted; Microscopy, Fluorescence
PubMed: 27943022
DOI: 10.1007/s00709-016-1058-8 -
Cold Spring Harbor Perspectives in... Aug 2014A particularly relevant phenomenon in cell physiology and proliferation is the fact that spontaneous mitotic recombination is strongly enhanced by transcription. The... (Review)
Review
A particularly relevant phenomenon in cell physiology and proliferation is the fact that spontaneous mitotic recombination is strongly enhanced by transcription. The most accepted view is that transcription increases the occurrence of double-strand breaks and/or single-stranded DNA gaps that are repaired by recombination. Most breaks would arise as a consequence of the impact that transcription has on replication fork progression, provoking its stalling and/or breakage. Here, we discuss the mechanisms responsible for the cross talk between transcription and recombination, with emphasis on (1) the transcription-replication conflicts as the main source of recombinogenic DNA breaks, and (2) the formation of cotranscriptional R-loops as a major cause of such breaks. The new emerging questions and perspectives are discussed on the basis of the interference between transcription and replication, as well as the way RNA influences genome dynamics.
Topics: DNA; DNA Breaks, Double-Stranded; DNA Replication; Humans; Models, Genetic; RNA; Recombination, Genetic; Transcription, Genetic
PubMed: 25085910
DOI: 10.1101/cshperspect.a016543 -
Critical Reviews in Biochemistry and... Jun 2017Life as we know it, simply would not exist without DNA replication. All living organisms utilize a complex machinery to duplicate their genomes and the central role in... (Review)
Review
Life as we know it, simply would not exist without DNA replication. All living organisms utilize a complex machinery to duplicate their genomes and the central role in this machinery belongs to replicative DNA polymerases, enzymes that are specifically designed to copy DNA. "Hassle-free" DNA duplication exists only in an ideal world, while in real life, it is constantly threatened by a myriad of diverse challenges. Among the most pressing obstacles that replicative polymerases often cannot overcome by themselves are lesions that distort the structure of DNA. Despite elaborate systems that cells utilize to cleanse their genomes of damaged DNA, repair is often incomplete. The persistence of DNA lesions obstructing the cellular replicases can have deleterious consequences. One of the mechanisms allowing cells to complete replication is "Translesion DNA Synthesis (TLS)". TLS is intrinsically error-prone, but apparently, the potential downside of increased mutagenesis is a healthier outcome for the cell than incomplete replication. Although most of the currently identified eukaryotic DNA polymerases have been implicated in TLS, the best characterized are those belonging to the "Y-family" of DNA polymerases (pols η, ι, κ and Rev1), which are thought to play major roles in the TLS of persisting DNA lesions in coordination with the B-family polymerase, pol ζ. In this review, we summarize the unique features of these DNA polymerases by mainly focusing on their biochemical and structural characteristics, as well as potential protein-protein interactions with other critical factors affecting TLS regulation.
Topics: Animals; DNA Damage; DNA Repair; DNA Replication; DNA-Directed DNA Polymerase; Humans
PubMed: 28279077
DOI: 10.1080/10409238.2017.1291576