scholarly journals Coordinating DNA Replication and Mitosis through Ubiquitin/SUMO and CDK1

2021 ◽  
Vol 22 (16) ◽  
pp. 8796
Author(s):  
Antonio Galarreta ◽  
Pablo Valledor ◽  
Oscar Fernandez-Capetillo ◽  
Emilio Lecona
Keyword(s):  
Dna Replication ◽  
Genomic Stability ◽  
S Phase ◽  
Replication Initiation ◽  
Cancer Therapies ◽  
Replication Machinery ◽  
Post Translational Modification ◽  
Aaa Atpase ◽  
Dna Replication Initiation ◽  
Set Up

Post-translational modification of the DNA replication machinery by ubiquitin and SUMO plays key roles in the faithful duplication of the genetic information. Among other functions, ubiquitination and SUMOylation serve as signals for the extraction of factors from chromatin by the AAA ATPase VCP. In addition to the regulation of DNA replication initiation and elongation, we now know that ubiquitination mediates the disassembly of the replisome after DNA replication termination, a process that is essential to preserve genomic stability. Here, we review the recent evidence showing how active DNA replication restricts replisome ubiquitination to prevent the premature disassembly of the DNA replication machinery. Ubiquitination also mediates the removal of the replisome to allow DNA repair. Further, we discuss the interplay between ubiquitin-mediated replisome disassembly and the activation of CDK1 that is required to set up the transition from the S phase to mitosis. We propose the existence of a ubiquitin–CDK1 relay, where the disassembly of terminated replisomes increases CDK1 activity that, in turn, favors the ubiquitination and disassembly of more replisomes. This model has important implications for the mechanism of action of cancer therapies that induce the untimely activation of CDK1, thereby triggering premature replisome disassembly and DNA damage.

scholarly journals Activation of the DNA Damage Checkpoint in Mutants Defective in DNA Replication Initiation

2008 ◽  
Vol 19 (10) ◽  
pp. 4374-4382 ◽  
Author(s):  
Ling Yin ◽  
Alexandra Monica Locovei ◽  
Gennaro D'Urso
Keyword(s):  
Dna Damage ◽  
Dna Replication ◽  
Replication Fork ◽  
S Phase ◽  
Replication Initiation ◽  
Dna Damage Checkpoint ◽  
Origin Firing ◽  
Dna Replication Initiation ◽  
Fork Stalling ◽  
S Phase Checkpoint

In the fission yeast, Schizosaccharomyces pombe, blocks to DNA replication elongation trigger the intra-S phase checkpoint that leads to the activation of the Cds1 kinase. Cds1 is required to both prevent premature entry into mitosis and to stabilize paused replication forks. Interestingly, although Cds1 is essential to maintain the viability of mutants defective in DNA replication elongation, mutants defective in DNA replication initiation require the Chk1 kinase. This suggests that defects in DNA replication initiation can lead to activation of the DNA damage checkpoint independent of the intra-S phase checkpoint. This might result from reduced origin firing that leads to an increase in replication fork stalling or replication fork collapse that activates the G2 DNA damage checkpoint. We refer to the Chk1-dependent, Cds1-independent phenotype as the rid phenotype (for replication initiation defective). Chk1 is active in rid mutants, and rid mutant viability is dependent on the DNA damage checkpoint, and surprisingly Mrc1, a protein required for activation of Cds1. Mutations in Mrc1 that prevent activation of Cds1 have no effect on its ability to support rid mutant viability, suggesting that Mrc1 has a checkpoint-independent role in maintaining the viability of mutants defective in DNA replication initiation.

scholarly journals The Intra-S-Phase Checkpoint Affects both DNA Replication Initiation and Elongation: Single-Cell and -DNA Fiber Analyses

2007 ◽  
Vol 27 (16) ◽  
pp. 5806-5818 ◽  
Author(s):  
Jennifer A. Seiler ◽  
Chiara Conti ◽  
Ali Syed ◽  
Mirit I. Aladjem ◽  
Yves Pommier
Keyword(s):  
Dna Replication ◽  
Topoisomerase I ◽  
Human Colon ◽  
S Phase ◽  
Replication Initiation ◽  
Dna Double Strand Breaks ◽  
Dna Replication Initiation ◽  
Nucleotide Incorporation ◽  
Replication Foci ◽  
S Phase Checkpoint

ABSTRACT To investigate the contribution of DNA replication initiation and elongation to the intra-S-phase checkpoint, we examined cells treated with the specific topoisomerase I inhibitor camptothecin. Camptothecin is a potent anticancer agent producing well-characterized replication-mediated DNA double-strand breaks through the collision of replication forks with topoisomerase I cleavage complexes. After a short dose of camptothecin in human colon carcinoma HT29 cells, DNA replication was inhibited rapidly and did not recover for several hours following drug removal. That inhibition occurred preferentially in late-S-phase, compared to early-S-phase, cells and was due to both an inhibition of initiation and elongation, as determined by pulse-labeling nucleotide incorporation in replication foci and DNA fibers. DNA replication was actively inhibited by checkpoint activation since 7-hydroxystaurosporine (UCN-01), the specific Chk1 inhibitor CHIR-124, or transfection with small interfering RNA targeting Chk1 restored both initiation and elongation. Abrogation of the checkpoint markedly enhanced camptothecin-induced DNA damage at replication sites where histone γ-H2AX colocalized with replication foci. Together, our study demonstrates that the intra-S-phase checkpoint is exerted by Chk1 not only upon replication initiation but also upon DNA elongation.

scholarly journals Genome duplication in Leishmania major relies on persistent subtelomeric DNA replication

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Jeziel Dener Damasceno ◽  
Catarina A Marques ◽  
Dario Beraldi ◽  
Kathryn Crouch ◽  
Craig Lapsley ◽  
...  
Keyword(s):  
Cell Division ◽  
Dna Replication ◽  
Leishmania Major ◽  
Genome Duplication ◽  
Histone H3 ◽  
S Phase ◽  
Replication Initiation ◽  
G1 Phase ◽  
Dna Replication Initiation ◽  
Single Region

DNA replication is needed to duplicate a cell’s genome in S phase and segregate it during cell division. Previous work in Leishmania detected DNA replication initiation at just a single region in each chromosome, an organisation predicted to be insufficient for complete genome duplication within S phase. Here, we show that acetylated histone H3 (AcH3), base J and a kinetochore factor co-localise in each chromosome at only a single locus, which corresponds with previously mapped DNA replication initiation regions and is demarcated by localised G/T skew and G4 patterns. In addition, we describe previously undetected subtelomeric DNA replication in G2/M and G1-phase-enriched cells. Finally, we show that subtelomeric DNA replication, unlike chromosome-internal DNA replication, is sensitive to hydroxyurea and dependent on 9-1-1 activity. These findings indicate that Leishmania’s genome duplication programme employs subtelomeric DNA replication initiation, possibly extending beyond S phase, to support predominantly chromosome-internal DNA replication initiation within S phase.

Efficiency and equity in origin licensing to ensure complete DNA replication

2021 ◽  
Author(s):  
Liu Mei ◽  
Jeanette Gowen Cook
Keyword(s):  
Cell Cycle ◽  
Cell Division ◽  
Dna Replication ◽  
Genome Stability ◽  
S Phase ◽  
Replication Initiation ◽  
G1 Phase ◽  
Dna Replication Initiation ◽  
Origin Licensing ◽  
Dna Replication Origins

The cell division cycle must be strictly regulated during both development and adult maintenance, and efficient and well-controlled DNA replication is a key event in the cell cycle. DNA replication origins are prepared in G1 phase of the cell cycle in a process known as origin licensing which is essential for DNA replication initiation in the subsequent S phase. Appropriate origin licensing includes: (1) Licensing enough origins at adequate origin licensing speed to complete licensing before G1 phase ends; (2) Licensing origins such that they are well-distributed on all chromosomes. Both aspects of licensing are critical for replication efficiency and accuracy. In this minireview, we will discuss recent advances in defining how origin licensing speed and distribution are critical to ensure DNA replication completion and genome stability.

scholarly journals Schizosacchromyces pombeDpb2 Binds to Origin DNA Early in S Phase and Is Required for Chromosomal DNA Replication

2003 ◽  
Vol 14 (8) ◽  
pp. 3427-3436 ◽  
Author(s):  
Wenyi Feng ◽  
Luis Rodriguez-Menocal ◽  
Gökhan Tolun ◽  
Gennaro D'Urso
Keyword(s):  
Dna Replication ◽  
S Phase ◽  
Replication Initiation ◽  
Temperature Sensitive ◽  
The Novel ◽  
Chromosomal Dna ◽  
Dna Polymerase Epsilon ◽  
Dna Replication Initiation ◽  
Cycle Arrest ◽  
C Terminus

Genetic evidence suggests that DNA polymerase epsilon (Pol ϵ) has a noncatalytic essential role during the early stages of DNA replication initiation. Herein, we report the cloning and characterization of the second largest subunit of Pol ϵ in fission yeast, called Dpb2. We demonstrate that Dpb2 is essential for cell viability and that a temperature-sensitive mutant of dpb2 arrests with a 1C DNA content, suggesting that Dpb2 is required for initiation of DNA replication. Using a chromatin immunoprecipitation assay, we show that Dpb2, binds preferentially to origin DNA at the beginning of S phase. We also show that the C terminus of Pol ϵ associates with origin DNA at the same time as Dpb2. We conclude that Dpb2 is an essential protein required for an early step in DNA replication. We propose that the primary function of Dpb2 is to facilitate assembly of the replicative complex at the start of S phase. These conclusions are based on the novel cell cycle arrest phenotype of the dpb2 mutant, on the previously uncharacterized binding of Dpb2 to replication origins, and on the observation that the essential function of Pol ϵ is not dependent on its DNA synthesis activity.

scholarly journals Unscheduled DNA replication in G1 causes genome instability through head-to-tail replication fork collisions

2021 ◽  
Author(s):  
Karl-Uwe Reusswig ◽  
Julia Bittmann ◽  
Martina Peritore ◽  
Michael Wierer ◽  
Matthias Mann ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Replication Fork ◽  
Genome Instability ◽  
S Phase ◽  
Replication Initiation ◽  
Dna Replication Initiation ◽  
Chromosome Breaks ◽  
Naturally Occurring ◽  
Identical Response

DNA replicates once per cell cycle. Interfering with the regulation of DNA replication initiation generates genome instability through over-replication and has been linked to early stages of cancer development. Here, we engineered genetic systems in budding yeast to induce unscheduled replication in the G1-phase of the cell cycle. Unscheduled G1 replication initiated at canonical S-phase origins across the genome. We quantified differences in replisomes in G1- and S-phase and identified firing factors, polymerase α, and histone supply as factors that limit replication outside S-phase. G1 replication per se did not trigger cellular checkpoints. Subsequent replication during S-phase, however, resulted in over-replication and led to chromosome breaks via head-to-tail replication fork collisions that are marked by chromosome-wide, strand-biased occurrence of RPA-bound single-stranded DNA. Low-level, sporadic induction of G1 replication induced an identical response, indicating findings from synthetic systems are applicable to naturally occurring scenarios of unscheduled replication initiation by G1/S deregulation.

scholarly journals Evidence That the pre-mRNA Splicing Factor Clf1p Plays a Role in DNA Replication in Saccharomyces cerevisiae

Genetics ◽  
2002 ◽  
Vol 160 (4) ◽  
pp. 1319-1333
Author(s):  
Wenge Zhu ◽  
Irene R Rainville ◽  
Min Ding ◽  
Margaret Bolus ◽  
Nicholas H Heintz ◽  
...  
Keyword(s):  
Dna Replication ◽  
S Phase ◽  
Replication Initiation ◽  
Mrna Splicing ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Direct Role ◽  
Dna Replication Initiation ◽  
Single Nucleus ◽  
2C Dna Content

Abstract Clf1p is an essential, highly conserved protein in S. cerevisiae that has been implicated in pre-mRNA splicing. Clf1p's ortholog in Drosophila, Crn, is required for normal cell proliferation. Cells depleted of Clf1p arrest primarily with large buds, a single nucleus, a 2C DNA content, and a short, intact mitotic spindle. We isolated temperature-sensitive clf1 mutants that exhibit similar mitotic defects when released to the restrictive temperature from an early S-phase block. While these mutants also accumulate unspliced pre-mRNA at the restrictive temperature, the mitotic arrest does not appear to result from a failure to splice tubulin pre-mRNA. Moreover, the same mutants exhibit a delayed entry into S phase when released to the restrictive temperature from a G1 phase block. This delay could not be suppressed by disruption of the S-phase CDK inhibitor SIC1, suggesting that Clf1p is involved in DNA replication. Consistent with this possibility, we find that Clf1p (but not the mutant clf1p) interacts with the DNA replication initiation protein Orc2p in two-hybrid and co-immunoprecipitation assays, that Clf1p preferentially associates with origins of DNA replication, and that this association is Orc2p dependent. These observations suggest that Clf1p plays a direct role in the initiation of DNA replication.

scholarly journals Genome duplication in Leishmania major relies on DNA replication outside S phase

2019 ◽  
Author(s):  
Jeziel D. Damasceno ◽  
Catarina A. Marques ◽  
Dario Beraldi ◽  
Kathryn Crouch ◽  
Craig Lapsley ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Replication ◽  
Genome Stability ◽  
Leishmania Major ◽  
Genome Duplication ◽  
S Phase ◽  
Replication Initiation ◽  
Dna Replication Initiation ◽  
Synthesis Activity ◽  
Important Parasite

AbstractOnce every cell cycle, DNA replication takes place to allow cells to duplicate their genome and segregate the two resulting copies into offspring cells. In eukaryotes, the number of DNA replication initiation loci, termed origins, is proportional to chromosome size. However, previous studies have suggested that in Leishmania, a group of single-celled eukaryotic parasites, DNA replication starts from just a single origin per chromosome, which is predicted to be insufficient to secure complete genome duplication within S phase. Here, we show that the paucity of origins activated in early S phase is balanced by DNA synthesis activity outside S phase. Simultaneous recruitment of acetylated histone H3 (AcH3), modified base J and the kinetochore factor KKT1 is exclusively found at the origins used in early S phase, while subtelomeric DNA replication can only be linked to AcH3 and displays persistent activity through the cell cycle, including in G2/M and G1 phases. We also show that subtelomeric DNA replication, unlike replication from the previously mapped origins, is sensitive to hydroxyurea and dependent on subunits of the 9-1-1 complex. Our work indicates that Leishmania genome transmission relies on an unconventional DNA replication programme, which may have implications for genome stability in this important parasite.

scholarly journals How MCM loading and spreading specify eukaryotic DNA replication initiation sites

F1000Research ◽  
2016 ◽  
Vol 5 ◽  
pp. 2063 ◽  
Author(s):  
Olivier Hyrien
Keyword(s):  
Dna Replication ◽  
Mammalian Cells ◽  
Origin Recognition Complex ◽  
Cell Types ◽  
S Phase ◽  
Replication Initiation ◽  
Replication Process ◽  
Dna Replication Initiation ◽  
Eukaryotic Dna ◽  
Transcription Complexes

DNA replication origins strikingly differ between eukaryotic species and cell types. Origins are localized and can be highly efficient in budding yeast, are randomly located in early fly and frog embryos, which do not transcribe their genomes, and are clustered in broad (10-100 kb) non-transcribed zones, frequently abutting transcribed genes, in mammalian cells. Nonetheless, in all cases, origins are established during the G1-phase of the cell cycle by the loading of double hexamers of the Mcm 2-7 proteins (MCM DHs), the core of the replicative helicase. MCM DH activation in S-phase leads to origin unwinding, polymerase recruitment, and initiation of bidirectional DNA synthesis. Although MCM DHs are initially loaded at sites defined by the binding of the origin recognition complex (ORC), they ultimately bind chromatin in much greater numbers than ORC and only a fraction are activated in any one S-phase. Data suggest that the multiplicity and functional redundancy of MCM DHs provide robustness to the replication process and affect replication time and that MCM DHs can slide along the DNA and spread over large distances around the ORC. Recent studies further show that MCM DHs are displaced along the DNA by collision with transcription complexes but remain functional for initiation after displacement. Therefore, eukaryotic DNA replication relies on intrinsically mobile and flexible origins, a strategy fundamentally different from bacteria but conserved from yeast to human. These properties of MCM DHs likely contribute to the establishment of broad, intergenic replication initiation zones in higher eukaryotes.

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