scholarly journals HMCES Maintains Replication Fork Progression and Prevents Double-Strand Breaks in Response to APOBEC Deamination and Abasic Site Formation

Cell Reports ◽  
2020 ◽  
Vol 31 (9) ◽  
pp. 107705 ◽  
Author(s):  
Kavi P.M. Mehta ◽  
Courtney A. Lovejoy ◽  
Runxiang Zhao ◽  
Darren R. Heintzman ◽  
David Cortez
Keyword(s):  
Replication Fork ◽  
Double Strand Breaks ◽  
Site Formation ◽  
Abasic Site ◽  
Strand Breaks ◽  
Replication Fork Progression

Cobalt and nickel impair DNA metabolism by the oxidative stress independent pathway

Metallomics ◽  
2017 ◽  
Vol 9 (11) ◽  
pp. 1596-1609 ◽  
Author(s):  
Vineet Kumar ◽  
Rajesh Kumar Mishra ◽  
Gursharan Kaur ◽  
Dipak Dutta
Keyword(s):  
Oxidative Stress ◽  
Metal Ions ◽  
Replication Fork ◽  
Double Strand Breaks ◽  
Dna Metabolism ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Replication Fork Progression ◽  
Nickel Exposure ◽  
Cobalt And Nickel

Cobalt and nickel exposure leads to DNA double-strand breaks, decelerating replication fork progression. In parallel, the metal ions inhibit RecBCD function to block SOS-mediated repair of the damaged DNA.

scholarly journals Defective Ribonucleoside Diphosphate Reductase Impairs Replication Fork Progression in Escherichia coli

2007 ◽  
Vol 189 (9) ◽  
pp. 3496-3501 ◽  
Author(s):  
Estrella Guarino ◽  
Alfonso Jiménez-Sánchez ◽  
Elena C. Guzmán
Keyword(s):  
Replication Fork ◽  
Double Strand Breaks ◽  
Permissive Temperature ◽  
Replication Forks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Replication Fork Progression ◽  
Holliday Junction Resolvase ◽  
Ribonucleoside Diphosphate Reductase ◽  
Stalled Replication Forks

ABSTRACT The observed lengthening of the C period in the presence of a defective ribonucleoside diphosphate reductase has been assumed to be due solely to the low deoxyribonucleotide supply in the nrdA101 mutant strain. We show here that the nrdA101 mutation induces DNA double-strand breaks at the permissive temperature in a recB-deficient background, suggesting an increase in the number of stalled replication forks that could account for the slowing of replication fork progression observed in the nrdA101 strain in a Rec+ context. These DNA double-strand breaks require the presence of the Holliday junction resolvase RuvABC, indicating that they have been generated from stalled replication forks that were processed by the specific reaction named “replication fork reversal.” Viability results supported the occurrence of this process, as specific lethality was observed in the nrdA101 recB double mutant and was suppressed by the additional inactivation of ruvABC. None of these effects seem to be due to the limitation of the deoxyribonucleotide supply in the nrdA101 strain even at the permissive temperature, as we found the same level of DNA double-strand breaks in the nrdA + strain growing under limited (2-μg/ml) or under optimal (5-μg/ml) thymidine concentrations. We propose that the presence of an altered NDP reductase, as a component of the replication machinery, impairs the progression of the replication fork, contributing to the lengthening of the C period in the nrdA101 mutant at the permissive temperature.

scholarly journals Replication of the Mammalian Genome by Replisomes Specific for Euchromatin and Heterochromatin

2021 ◽  
Vol 9 ◽  
Author(s):  
Jing Zhang ◽  
Marina A. Bellani ◽  
Jing Huang ◽  
Ryan C. James ◽  
Durga Pokharel ◽  
...  
Keyword(s):  
Replication Fork ◽  
Single Species ◽  
S Phase ◽  
Double Strand Breaks ◽  
Replication Forks ◽  
Strand Breaks ◽  
Interstrand Crosslink ◽  
Genomic Location ◽  
Replication Restart ◽  
Atr Kinase

Replisomes follow a schedule in which replication of DNA in euchromatin is early in S phase while sequences in heterochromatin replicate late. Impediments to DNA replication, referred to as replication stress, can stall replication forks triggering activation of the ATR kinase and downstream pathways. While there is substantial literature on the local consequences of replisome stalling–double strand breaks, reversed forks, or genomic rearrangements–there is limited understanding of the determinants of replisome stalling vs. continued progression. Although many proteins are recruited to stalled replisomes, current models assume a single species of “stressed” replisome, independent of genomic location. Here we describe our approach to visualizing replication fork encounters with the potent block imposed by a DNA interstrand crosslink (ICL) and our discovery of an unexpected pathway of replication restart (traverse) past an intact ICL. Additionally, we found two biochemically distinct replisomes distinguished by activity in different stages of S phase and chromatin environment. Each contains different proteins that contribute to ICL traverse.

scholarly journals RecO impedes RecG-SSB binding to impair the strand annealing recombination pathway in E.coli

2019 ◽  
Author(s):  
Xuefeng Pan ◽  
Li Yang ◽  
Nan Jiang ◽  
Xifang Chen ◽  
Bo Li ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Dna Repair ◽  
Dna Replication ◽  
Replication Fork ◽  
Double Strand Breaks ◽  
Recombinational Repair ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Homologous Recombinational Repair ◽  
Differential Binding

AbstractFaithful duplication of genomic DNA relies not only on the fidelity of DNA replication itself, but also on fully functional DNA repair and homologous recombination machinery. We report a molecular mechanism responsible for deciding homologous recombinational repair pathways during replication dictated by binding of RecO and RecG to SSB in E.coli. Using a RecG-yfp fusion protein, we found that RecG-yfp foci appeared only in the ΔrecG, ΔrecO and ΔrecA, ΔrecO double mutants. Surprisingly, foci were not observed in wild-type ΔrecG, or double mutants where recG and either recF or, separately recR were deleted. In addition, formation of RecG-yfp foci in the ΔrecO::kanR required wildtype ssb, as ssb-113 could not substitute. This suggests that RecG and RecO binding to SSB is competitive. We also found that the UV resistance of recO alone mutant increased to certain extent by supplementing RecG. In an ssb-113 mutant, RecO and RecG worked following a different pattern. Both RecO and RecG were able to participate in repairing UV damages when grown at permissive temperature, while they could also be involved in making DNA double strand breaks when grown at nonpermissive temperature. So, our results suggested that differential binding of RecG and RecO to SSB in a DNA replication fork in Escherichia coli.may be involved in determining whether the SDSA or DSBR pathway of homologous recombinational repair is used.Author summarySingle strand DNA binding proteins (SSB) stabilize DNA holoenzyme and prevent single strand DNA from folding into non-B DNA structures in a DNA replication fork. It has also been revealed that SSB can also act as a platform for some proteins working in DNA repair and recombination to access DNA molecules when DNA replication fork needs to be reestablished. In Escherichia coli, several proteins working primarily in DNA repair and recombination were found to participate in DNA replication fork resumption by physically interacting with SSB, including RecO and RecG etc. However the hierarchy of these proteins interacting with SSB in Escherichia coli has not been well defined. In this study, we demonstrated a differential binding of RecO and RecG to SSB in DNA replication was used to establish a RecO-dependent pathway of replication fork repair by abolishing a RecG-dependent replication fork repair. We also show that, RecG and RecO could randomly participate in DNA replication repair in the absence of a functional SSB, which may be responsible for the generation of DNA double strand breaks in an ssb-113 mutant in Escherichia coli.

Measurement of Chromosomal DNA Single‐Strand Breaks and Replication Fork Progression Rates

2006 ◽  
pp. 410-425 ◽  
Author(s):  
Claire Breslin ◽  
Paula M. Clements ◽  
Sherif F. El‐Khamisy ◽  
Eva Petermann ◽  
Natasha Iles ◽  
...  
Keyword(s):  
Replication Fork ◽  
Single Strand ◽  
Chromosomal Dna ◽  
Strand Breaks ◽  
Replication Fork Progression ◽  
Single Strand Breaks

scholarly journals The non-homologous end joining factor Ku orchestrates replication fork resection and fine-tunes Rad51-mediated fork restart

2017 ◽  
Author(s):  
Ana Teixeira-Silva ◽  
Anissia Ait Saada ◽  
Ismail Iraqui ◽  
Marina Charlotte Nocente ◽  
Karine Fréon ◽  
...  
Keyword(s):  
Replication Fork ◽  
Double Strand Breaks ◽  
Fine Tuning ◽  
List Type ◽  
End Joining ◽  
Strand Breaks ◽  
Replication Restart ◽  
Dna End Resection ◽  
Non Homologous End Joining ◽  
End Resection

AbstractReplication requires Homologous Recombination (HR) to stabilize and restart terminally-arrested forks. HR-mediated fork processing requires single stranded DNA (ssDNA) gaps and not necessarily Double Strand Breaks. We used genetic and molecular assays to investigate fork-resection and restart at dysfunctional, unbroken forks in Schizosaccharomyces pombe. We found that fork-resection is a two-step process coordinated by the non-homologous end joining factor Ku. An initial resection mediated by MRN/Ctp1 removes Ku from terminally-arrested forks, generating ~ 110 bp sized gaps obligatory for subsequent Exo1-mediated long-range resection and replication restart. The lack of Ku results in slower fork restart, excessive resection, and impaired RPA recruitment. We propose that terminally-arrested forks undergo fork reversal, providing a single DNA end for Ku binding which primes RPA-coated ssDNA. We uncover an unprecedented role for Ku in orchestrating resection of unbroken forks and in fine-tuning HR-mediated replication restart.Ku orchestrates a two-steps DNA end-resection of terminally-arrested and unbroken forksMRN/Ctp1 removes Ku from terminally-arrested forks to initiate fork-resectiona ~110 bp sized ssDNA gap is sufficient and necessary to promote fork restart.The lack of Ku decreases ssDNA RPA-coating, and slows down replication fork restart.

scholarly journals DNA Ligase IV Prevents Replication Fork Stalling and Promotes Cellular Proliferation in Triple Negative Breast Cancer

2019 ◽  
Vol 2019 ◽  
pp. 1-11 ◽  
Author(s):  
Rashmi R. Joshi ◽  
Sk Imran Ali ◽  
Amanda K. Ashley
Keyword(s):  
Breast Cancer ◽  
Dna Damage ◽  
Replication Fork ◽  
Triple Negative ◽  
Double Strand Breaks ◽  
Dna Ligase ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Ligase Iv ◽  
Fork Stalling

DNA damage is a hallmark of cancer, and mutation and misregulation of proteins that maintain genomic fidelity are associated with the development of multiple cancers. DNA double strand breaks are arguably considered the most deleterious type of DNA damage. The nonhomologous end-joining (NHEJ) pathway is one mechanism to repair DNA double strand breaks, and proteins involved in NHEJ may also regulate DNA replication. We previously established that DNA-PKcs, a NHEJ protein, promotes genomic stability and cell viability following cellular exposure to replication stress; we wanted to discern whether another NHEJ protein, DNA ligase IV (Lig4), shares this phenotype. Our investigations focused on triple negative breast cancer cells, as, compared to nonbasal breast cancer, LIG4 is frequently amplified, and an increased gene dose is associated with higher Lig4 expression. We depleted Lig4 using siRNA and confirmed our knockdown by qPCR and western blotting. Cell survival diminished with Lig4 depletion alone, and this was associated with increased replication fork stalling. Checkpoint protein Chk1 activation and dephosphorylation were unchanged in Lig4-depleted cells. Lig4 depletion resulted in sustained DNA-PKcs phosphorylation following hydroxyurea exposure. Understanding the effect of Lig4 on genomic replication and the replication stress response will clarify the biological ramifications of inhibiting Lig4 activity. In addition, Lig4 is an attractive clinical target for directing CRISPR/Cas9-mediated repair towards homology-directed repair and away from NHEJ, thus understanding of how diminishing Lig4 impacts cell biology is critical.

scholarly journals Interstitial telomere sequences disrupt break-induced replication and drive formation of ectopic telomeres

2020 ◽  
Vol 48 (22) ◽  
pp. 12697-12710
Author(s):  
Elizabeth A Stivison ◽  
Kati J Young ◽  
Lorraine S Symington
Keyword(s):  
Replication Fork ◽  
Double Strand Breaks ◽  
Replication Forks ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Homology Directed Repair ◽  
Telomere Sequence ◽  
D Loop ◽  
Break Induced Replication ◽  
Telomeric Protein

Abstract Break-induced replication (BIR) is a mechanism used to heal one-ended DNA double-strand breaks, such as those formed at collapsed replication forks or eroded telomeres. Instead of utilizing a canonical replication fork, BIR is driven by a migrating D-loop and is associated with a high frequency of mutagenesis. Here we show that when BIR encounters an interstitial telomere sequence (ITS), the machinery frequently terminates, resulting in the formation of an ectopic telomere. The primary mechanism to convert the ITS to a functional telomere is by telomerase-catalyzed addition of telomeric repeats with homology-directed repair serving as a back-up mechanism. Termination of BIR and creation of an ectopic telomere is promoted by Mph1/FANCM helicase, which has the capacity to disassemble D-loops. Other sequences that have the potential to seed new telomeres but lack the unique features of a natural telomere sequence, do not terminate BIR at a significant frequency in wild-type cells. However, these sequences can form ectopic telomeres if BIR is made less processive. Our results support a model in which features of the ITS itself, such as the propensity to form secondary structures and telomeric protein binding, pose a challenge to BIR and increase the vulnerability of the D-loop to dissociation by helicases, thereby promoting ectopic telomere formation.

scholarly journals Analysis of RuvABC and RecG Involvement in the Escherichia coli Response to the Covalent Topoisomerase-DNA Complex

2010 ◽  
Vol 192 (17) ◽  
pp. 4445-4451 ◽  
Author(s):  
Jeanette H. Sutherland ◽  
Yuk-Ching Tse-Dinh
Keyword(s):  
Escherichia Coli ◽  
Homologous Recombination ◽  
Dna Cleavage ◽  
Topoisomerase I ◽  
Replication Fork ◽  
Double Strand Breaks ◽  
Strand Breaks ◽  
Sos Induction ◽  
Cleavage Complex ◽  
Dna Complex

ABSTRACT Topoisomerases form a covalent enzyme-DNA intermediate after initial DNA cleavage. Trapping of the cleavage complex formed by type IIA topoisomerases initiates the bactericidal action of fluoroquinolones. It should be possible also to identify novel antibacterial lead compounds that act with a similar mechanism on type IA bacterial topoisomerases. The cellular response and repair pathways for trapped topoisomerase complexes remain to be fully elucidated. The RuvAB and RecG proteins could play a role in the conversion of the initial protein-DNA complex to double-strand breaks and also in the resolution of the Holliday junction during homologous recombination. Escherichia coli strains with ruvA and recG mutations are found to have increased sensitivity to low levels of norfloxacin treatment, but the mutations had more pronounced effects on survival following the accumulation of covalent complexes formed by mutant topoisomerase I defective in DNA religation. Covalent topoisomerase I and DNA gyrase complexes are converted into double-strand breaks for SOS induction by the RecBCD pathway. SOS induction following topoisomerase I complex accumulation is significantly lower in the ruvA and recG mutants than in the wild-type background, suggesting that RuvAB and RecG may play a role in converting the initial single-strand DNA-protein cleavage complex into a double-strand break prior to repair by homologous recombination. The use of a ruvB mutant proficient in homologous recombination but not in replication fork reversal demonstrated that the replication fork reversal function of RuvAB is required for SOS induction by the covalent complex formed by topoisomerase I.

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