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Cold Spring Harbor Perspectives in... Mar 2013One of the fundamental challenges facing the cell is to accurately copy its genetic material to daughter cells. When this process goes awry, genomic instability ensues... (Review)
Review
One of the fundamental challenges facing the cell is to accurately copy its genetic material to daughter cells. When this process goes awry, genomic instability ensues in which genetic alterations ranging from nucleotide changes to chromosomal translocations and aneuploidy occur. Organisms have developed multiple mechanisms that can be classified into two major classes to ensure the fidelity of DNA replication. The first class includes mechanisms that prevent premature initiation of DNA replication and ensure that the genome is fully replicated once and only once during each division cycle. These include cyclin-dependent kinase (CDK)-dependent mechanisms and CDK-independent mechanisms. Although CDK-dependent mechanisms are largely conserved in eukaryotes, higher eukaryotes have evolved additional mechanisms that seem to play a larger role in preventing aberrant DNA replication and genome instability. The second class ensures that cells are able to respond to various cues that continuously threaten the integrity of the genome by initiating DNA-damage-dependent "checkpoints" and coordinating DNA damage repair mechanisms. Defects in the ability to safeguard against aberrant DNA replication and to respond to DNA damage contribute to genomic instability and the development of human malignancy. In this article, we summarize our current knowledge of how genomic instability arises, with a particular emphasis on how the DNA replication process can give rise to such instability.
Topics: Cell Cycle Checkpoints; Cell Cycle Proteins; DNA Damage; DNA Replication; Geminin; Genomic Instability; Humans; Minichromosome Maintenance Complex Component 2; Models, Biological; Neoplasms; Nuclear Proteins; Origin Recognition Complex; Ubiquitin-Protein Ligases
PubMed: 23335075
DOI: 10.1101/cshperspect.a012914 -
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 -
Microbiology Spectrum Feb 2015Plasmids are autonomously replicating pieces of DNA. This article discusses theta plasmid replication, which is a class of circular plasmid replication that includes... (Review)
Review
Plasmids are autonomously replicating pieces of DNA. This article discusses theta plasmid replication, which is a class of circular plasmid replication that includes ColE1-like origins of replication popular with expression vectors. All modalities of theta plasmid replication initiate synthesis with the leading strand at a predetermined site and complete replication through recruitment of the host's replisome, which extends the leading strand continuously while synthesizing the lagging strand discontinuously. There are clear differences between different modalities of theta plasmid replication in mechanisms of DNA duplex melting and in priming of leading- and lagging-strand synthesis. In some replicons duplex melting depends on transcription, while other replicons rely on plasmid-encoded trans-acting proteins (Reps); primers for leading-strand synthesis can be generated through processing of a transcript or in other replicons by the action of host- or plasmid-encoded primases. None of these processes require DNA breaks. The frequency of replication initiation is tightly regulated to facilitate establishment in permissive hosts and to achieve a steady state. The last section of the article reviews how plasmid copy number is sensed and how this feedback modulates the frequency of replication.
Topics: Bacteria; DNA Replication; DNA, Circular; Plasmids
PubMed: 26104556
DOI: 10.1128/microbiolspec.PLAS-0029-2014 -
Nature Structural & Molecular Biology Jan 2019Although DNA replication is a fundamental aspect of biology, it is not known what determines where DNA replication starts and stops in the human genome. We directly...
Although DNA replication is a fundamental aspect of biology, it is not known what determines where DNA replication starts and stops in the human genome. We directly identified and quantitatively compared sites of replication initiation and termination in untransformed human cells. We found that replication preferentially initiates at the transcription start site of genes occupied by high levels of RNA polymerase II, and terminates at their polyadenylation sites, thereby ensuring global co-directionality of transcription and replication, particularly at gene 5' ends. During replication stress, replication initiation is stimulated downstream of genes and termination is redistributed to gene bodies; this globally reorients replication relative to transcription around gene 3' ends. These data suggest that replication initiation and termination are coupled to transcription in human cells, and propose a model for the impact of replication stress on genome integrity.
Topics: DNA Replication; Humans; Polyadenylation; RNA Polymerase II; Replication Origin; Transcription Initiation Site; Transcription, Genetic
PubMed: 30598550
DOI: 10.1038/s41594-018-0171-0 -
Robust linear DNA degradation supports replication-initiation-defective mutants in Escherichia coli.G3 (Bethesda, Md.) Nov 2022RecBCD helicase/nuclease supports replication fork progress via recombinational repair or linear DNA degradation, explaining recBC mutant synthetic lethality with...
RecBCD helicase/nuclease supports replication fork progress via recombinational repair or linear DNA degradation, explaining recBC mutant synthetic lethality with replication elongation defects. Since replication initiation defects leave chromosomes without replication forks, these should be insensitive to the recBCD status. Surprisingly, we found that both Escherichia coli dnaA46(Ts) and dnaC2(Ts) initiation mutants at semi-permissive temperatures are also recBC-colethal. Interestingly, dnaA46 recBC lethality suppressors suggest underinitiation as the problem, while dnaC2 recBC suppressors signal overintiation. Using genetic and physical approaches, we studied the dnaA46 recBC synthetic lethality, for the possibility that RecBCD participates in replication initiation. Overproduced DnaA46 mutant protein interferes with growth of dnaA+ cells, while the residual viability of the dnaA46 recBC mutant depends on the auxiliary replicative helicase Rep, suggesting replication fork inhibition by the DnaA46 mutant protein. The dnaA46 mutant depends on linear DNA degradation by RecBCD, rather than on recombinational repair. At the same time, the dnaA46 defect also interacts with Holliday junction-moving defects, suggesting reversal of inhibited forks. However, in contrast to all known recBC-colethals, which fragment their chromosomes, the dnaA46 recBC mutant develops no chromosome fragmentation, indicating that its inhibited replication forks are stable. Physical measurements confirm replication inhibition in the dnaA46 mutant shifted to semi-permissive temperatures, both at the level of elongation and initiation, while RecBCD gradually restores elongation and then initiation. We propose that RecBCD-catalyzed resetting of inhibited replication forks allows replication to displace the "sticky" DnaA46(Ts) protein from the chromosomal DNA, mustering enough DnaA for new initiations.
Topics: Escherichia coli; Escherichia coli Proteins; DNA Replication; DNA Helicases; DNA; Mutant Proteins; DNA, Bacterial; Mutation; Bacterial Proteins; Chromosomes, Bacterial
PubMed: 36165702
DOI: 10.1093/g3journal/jkac228 -
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 -
The Biochemical Journal Sep 2020Eukaryotic DNA replication is a highly dynamic and tightly regulated process. Replication involves several dozens of replication proteins, including the initiators ORC...
Eukaryotic DNA replication is a highly dynamic and tightly regulated process. Replication involves several dozens of replication proteins, including the initiators ORC and Cdc6, replicative CMG helicase, DNA polymerase α-primase, leading-strand DNA polymerase ε, and lagging-strand DNA polymerase δ. These proteins work together in a spatially and temporally controlled manner to synthesize new DNA from the parental DNA templates. During DNA replication, epigenetic information imprinted on DNA and histone proteins is also copied to the daughter DNA to maintain the chromatin status. DNA methyltransferase 1 is primarily responsible for copying the parental DNA methylation pattern into the nascent DNA. Epigenetic information encoded in histones is transferred via a more complex and less well-understood process termed replication-couple nucleosome assembly. Here, we summarize the most recent structural and biochemical insights into DNA replication initiation, replication fork elongation, chromatin assembly and maintenance, and related regulatory mechanisms.
Topics: Animals; DNA; DNA Polymerase II; DNA Polymerase III; DNA Replication; Epigenesis, Genetic; Eukaryotic Cells; Histones; Humans; Nucleosomes; Poly-ADP-Ribose Binding Proteins
PubMed: 32970141
DOI: 10.1042/BCJ20200065 -
Bioscience Reports Jul 2023To pass on genetic information to the next generation, cells must faithfully replicate their genomes to provide copies for each daughter cell. To synthesise these... (Review)
Review
To pass on genetic information to the next generation, cells must faithfully replicate their genomes to provide copies for each daughter cell. To synthesise these duplicates, cells employ specialised enzymes called DNA polymerases, which rapidly and accurately replicate nucleic acid polymers. However, most polymerases lack the ability to directly initiate DNA synthesis and required specialised replicases called primases to make short polynucleotide primers, from which they then extend. Replicative primases (eukaryotes and archaea) belong to a functionally diverse enzyme superfamily known as Primase-Polymerases (Prim-Pols), with orthologues present throughout all domains of life. Characterised by a conserved catalytic Prim-Pol domain, these enzymes have evolved various roles in DNA metabolism, including DNA replication, repair, and damage tolerance. Many of these biological roles are fundamentally underpinned by the ability of Prim-Pols to generate primers de novo. This review examines our current understanding of the catalytic mechanisms utilised by Prim-Pols to initiate primer synthesis.
Topics: DNA Primase; DNA-Directed DNA Polymerase; DNA Replication; Catalytic Domain; DNA
PubMed: 37358261
DOI: 10.1042/BSR20221986 -
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 -
Molecules (Basel, Switzerland) Sep 2019G-quadruplexes are four-stranded guanine-rich structures that have been demonstrated to occur across the genome in humans and other organisms. They provide regulatory... (Review)
Review
G-quadruplexes are four-stranded guanine-rich structures that have been demonstrated to occur across the genome in humans and other organisms. They provide regulatory functions during transcription, translation and immunoglobulin gene rearrangement, but there is also a large amount of evidence that they can present a potent barrier to the DNA replication machinery. This mini-review will summarize recent advances in understanding the many strategies nature has evolved to overcome G-quadruplex-mediated replication blockage, including removal of the structure by helicases or nucleases, or circumventing the deleterious effects on the genome through homologous recombination, alternative end-joining or synthesis re-priming. Paradoxically, G-quadruplexes have also recently been demonstrated to provide a positive role in stimulating the initiation of DNA replication. These recent studies have not only illuminated the many roles and consequences of G-quadruplexes, but have also provided fundamental insights into the general mechanisms of DNA replication and its links with genetic and epigenetic stability.
Topics: DNA Helicases; DNA Replication; Deoxyribonucleases; G-Quadruplexes; Humans
PubMed: 31546714
DOI: 10.3390/molecules24193439