-
Cell Cycle (Georgetown, Tex.) 2016Heat stress is one of the best-studied exogenous stress factors; however little is known about its delayed effects. Recently, we have shown that heat stress induces... (Review)
Review
Heat stress is one of the best-studied exogenous stress factors; however little is known about its delayed effects. Recently, we have shown that heat stress induces cellular senescence-like G2 arrest exclusively in early S-phase cells. The mechanism of this arrest includes the generation of heat stress-induced single-stranded DNA breaks, the collision of replication forks with these breaks and the formation of difficult-to-repair double-stranded DNA breaks. However, the early S phase-specific effects of heat stress are not limited to the induction of single-stranded DNA breaks. Here, we report that HS induces partial DNA re-replication and centrosome amplification. We suggest that HS-induced alterations in the expression levels of the genes encoding the replication licensing factors are the primary source of such perturbations. Notably, these processes do not contribute to acquisition of a senescence-like phenotype, although they do elicit postponed effects. Specifically, we found that the HeLa cells can escape from the heat stress-induced cellular senescence-like G2 arrest, and the mitosis they enter is multipolar due to the amplified centrosomes.
Topics: Cellular Senescence; Centrosome; DNA Damage; DNA Replication; Hot Temperature; Humans; S Phase; Tubulin
PubMed: 26689112
DOI: 10.1080/15384101.2015.1127477 -
Current Opinion in Structural Biology Dec 2023In eukaryotic cells, genome duplication is temporally organised according to a program referred to as the replication-timing (RT) program. The RT of individual genomic... (Review)
Review
In eukaryotic cells, genome duplication is temporally organised according to a program referred to as the replication-timing (RT) program. The RT of individual genomic domains strikingly parallels the three-dimensional architecture of their chromatin contacts and subnuclear distribution. However, it is unclear whether this correspondence is coincidental or whether it indicates a causal and regulatory relationship. In either case, the nature of the molecular mechanisms ensuring this spatio-temporal coordination is still unknown. Here, we review recent evidence that begins to uncover the existence of a shared molecular machinery at the core of the spatio-temporal co-regulation of DNA replication and genome architecture. Finally, we discuss the outstanding, key question of the biological role of their coordination.
Topics: DNA Replication Timing; Chromatin; DNA Replication; Eukaryotic Cells; Genome
PubMed: 37741142
DOI: 10.1016/j.sbi.2023.102704 -
Proceedings of the National Academy of... May 2020
Topics: DNA; DNA Replication; DNA-Directed DNA Polymerase; Mutation; Saccharomyces cerevisiae
PubMed: 32350136
DOI: 10.1073/pnas.2005160117 -
Chemical Reviews Dec 2023The paradigm of cellular systems as deterministic machines has long guided our understanding of biology. Advancements in technology and methodology, however, have... (Review)
Review
The paradigm of cellular systems as deterministic machines has long guided our understanding of biology. Advancements in technology and methodology, however, have revealed a world of stochasticity, challenging the notion of determinism. Here, we explore the stochastic behavior of multi-protein complexes, using the DNA replication system (replisome) as a prime example. The faithful and timely copying of DNA depends on the simultaneous action of a large set of enzymes and scaffolding factors. This fundamental cellular process is underpinned by dynamic protein-nucleic acid assemblies that must transition between distinct conformations and compositional states. Traditionally viewed as a well-orchestrated molecular machine, recent experimental evidence has unveiled significant variability and heterogeneity in the replication process. In this review, we discuss recent advances in single-molecule approaches and single-particle cryo-EM, which have provided insights into the dynamic processes of DNA replication. We comment on the new challenges faced by structural biologists and biophysicists as they attempt to describe the dynamic cascade of events leading to replisome assembly, activation, and progression. The fundamental principles uncovered and yet to be discovered through the study of DNA replication will inform on similar operating principles for other multi-protein complexes.
Topics: DNA Replication; DNA; Molecular Conformation
PubMed: 37971892
DOI: 10.1021/acs.chemrev.3c00436 -
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 -
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 -
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 -
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 -
Open Biology Dec 2020The tree of life shows the relationship between all organisms based on their common ancestry. Until 1977, it comprised two major branches: prokaryotes and eukaryotes.... (Review)
Review
The tree of life shows the relationship between all organisms based on their common ancestry. Until 1977, it comprised two major branches: prokaryotes and eukaryotes. Work by Carl Woese and other microbiologists led to the recategorization of prokaryotes and the proposal of three primary domains: Eukarya, Bacteria and Archaea. Microbiological, genetic and biochemical techniques were then needed to study the third domain of life. , a halophilic species belonging to the phylum Euryarchaeota, has provided many useful tools to study Archaea, including easy culturing methods, genetic manipulation and phenotypic screening. This review will focus on DNA replication and DNA repair pathways in , how this work has advanced our knowledge of archaeal cellular biology, and how it may deepen our understanding of bacterial and eukaryotic processes.
Topics: Biomarkers; DNA Repair; DNA Replication; DNA, Archaeal; Gene Expression Regulation, Archaeal; Haloferax volcanii; Recombination, Genetic
PubMed: 33259746
DOI: 10.1098/rsob.200293 -
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