scholarly journals DNA- and DNA-Protein-Crosslink Repair in Plants

2019 ◽  
Vol 20 (17) ◽  
pp. 4304 ◽  
Author(s):  
Janina Enderle ◽  
Annika Dorn ◽  
Holger Puchta
Keyword(s):  
S Phase ◽  
Dna Lesions ◽  
Replication Forks ◽  
Genomic Integrity ◽  
Major Threat ◽  
Dna Crosslinks ◽  
Hereditary Disorders ◽  
Toxic Potential ◽  
Protein Crosslinks ◽  
Crosslink Repair

DNA-crosslinks are one of the most severe types of DNA lesions. Crosslinks (CLs) can be subdivided into DNA-intrastrand CLs, DNA-interstrand CLs (ICLs) and DNA-protein crosslinks (DPCs), and arise by various exogenous and endogenous sources. If left unrepaired before the cell enters S-phase, ICLs and DPCs pose a major threat to genomic integrity by blocking replication. In order to prevent the collapse of replication forks and impairment of cell division, complex repair pathways have emerged. In mammals, ICLs are repaired by the so-called Fanconi anemia (FA) pathway, which includes 22 different FANC genes, while in plants only a few of these genes are conserved. In this context, two pathways of ICL repair have been defined, each requiring the interaction of a helicase (FANCJB/RTEL1) and a nuclease (FAN1/MUS81). Moreover, homologous recombination (HR) as well as postreplicative repair factors are also involved. Although DPCs possess a comparable toxic potential to cells, it has only recently been shown that at least three parallel pathways for DPC repair exist in plants, defined by the protease WSS1A, the endonuclease MUS81 and tyrosyl-DNA phosphodiesterase 1 (TDP1). The importance of crosslink repair processes are highlighted by the fact that deficiencies in the respective pathways are associated with diverse hereditary disorders.

scholarly journals Mechanism of replication-coupled DNA-protein crosslink proteolysis by SPRTN and the proteasome

2018 ◽  
Author(s):  
Alan Gao ◽  
Nicolai B. Larsen ◽  
Justin L. Sparks ◽  
Irene Gallina ◽  
Matthias Mann ◽  
...  
Keyword(s):  
Dna Replication ◽  
S Phase ◽  
Dna Lesions ◽  
List Type ◽  
Genomic Integrity ◽  
Dna Metabolism ◽  
Single Stranded Dna ◽  
Egg Extracts ◽  
Protein Crosslinks ◽  
Bulky Dna Lesions

SummaryDNA-protein crosslinks (DPCs) are bulky DNA lesions that interfere with DNA metabolism and therefore threaten genomic integrity. Recent studies implicate the metalloprotease SPRTN in S-phase removal of DPCs, but how SPRTN activity is coupled to DNA replication is unknown. Using Xenopus egg extracts that recapitulate replication-coupled DPC proteolysis, we show that DPCs can be degraded by SPRTN or the proteasome, which act as independent DPC proteases. Proteasome recruitment requires DPC polyubiquitylation, which is triggered by single-stranded DNA, a byproduct of DNA replication. In contrast, SPRTN-mediated DPC degradation is independent of DPC polyubiquitylation but requires polymerase extension of a nascent strand to the lesion. Thus, SPRTN and proteasome activities are coupled to DNA replication by distinct mechanisms and together promote replication across immovable protein barriers.HighlightsThe proteasome, in addition to SPRTN, degrades DPCs during DNA replicationProteasome-dependent DPC degradation requires DPC ubiquitylationDPC ubiquitylation is triggered by ssDNA and does not require the replisomeSPRTN-dependent DPC degradation is a post-replicative process

scholarly journals Multi-BRCT Domain Protein Brc1 Links Rhp18/Rad18 and γH2A To Maintain Genome Stability during S Phase

2017 ◽  
Vol 37 (22) ◽  
Author(s):  
Michael C. Reubens ◽  
Sophie Rozenzhak ◽  
Paul Russell
Keyword(s):  
Genome Stability ◽  
Genome Instability ◽  
Replication Stress ◽  
S Phase ◽  
Dna Lesions ◽  
Brct Domain ◽  
Replication Forks ◽  
Content Type ◽  
Domain Protein ◽  
Chromosome Integrity

ABSTRACT DNA replication involves the inherent risk of genome instability, since replisomes invariably encounter DNA lesions or other structures that stall or collapse replication forks during the S phase. In the fission yeast Schizosaccharomyces pombe, the multi-BRCT domain protein Brc1, which is related to budding yeast Rtt107 and mammalian PTIP, plays an important role in maintaining genome integrity and cell viability when cells experience replication stress. The C-terminal pair of BRCT domains in Brc1 were previously shown to bind phosphohistone H2A (γH2A) formed by Rad3/ATR checkpoint kinase at DNA lesions; however, the putative scaffold interactions involving the N-terminal BRCT domains 1 to 4 of Brc1 have remained obscure. Here, we show that these domains bind Rhp18/Rad18, which is an E3 ubiquitin protein ligase that has crucial functions in postreplication repair. A missense allele in BRCT domain 4 of Brc1 disrupts binding to Rhp18 and causes sensitivity to replication stress. Brc1 binding to Rhp18 and γH2A are required for the Brc1 overexpression suppression of smc6-74, a mutation that impairs the Smc5/6 structural maintenance of chromosomes complex required for chromosome integrity and repair of collapsed replication forks. From these findings, we propose that Brc1 provides scaffolding functions linking γH2A, Rhp18, and Smc5/6 complex at damaged replication forks.

scholarly journals ATAD5 restricts R-loop formation through PCNA unloading and RNA helicase maintenance at the replication fork

2020 ◽  
Author(s):  
Sangin Kim ◽  
Nalae Kang ◽  
Su Hyung Park ◽  
James Wells ◽  
Taejoo Hwang ◽  
...  
Keyword(s):  
Replication Fork ◽  
Replication Stress ◽  
S Phase ◽  
Replication Forks ◽  
Genomic Integrity ◽  
Loop Formation ◽  
Replication Rate ◽  
Rna Helicases ◽  
Dual Functions

Abstract R-loops are formed when replicative forks collide with the transcriptional machinery and can cause genomic instability. However, it is unclear how R-loops are regulated at transcription-replication conflict (TRC) sites and how replisome proteins are regulated to prevent R-loop formation or mediate R-loop tolerance. Here, we report that ATAD5, a PCNA unloader, plays dual functions to reduce R-loops both under normal and replication stress conditions. ATAD5 interacts with RNA helicases such as DDX1, DDX5, DDX21 and DHX9 and increases the abundance of these helicases at replication forks to facilitate R-loop resolution. Depletion of ATAD5 or ATAD5-interacting RNA helicases consistently increases R-loops during the S phase and reduces the replication rate, both of which are enhanced by replication stress. In addition to R-loop resolution, ATAD5 prevents the generation of new R-loops behind the replication forks by unloading PCNA which, otherwise, accumulates and persists on DNA, causing a collision with the transcription machinery. Depletion of ATAD5 reduces transcription rates due to PCNA accumulation. Consistent with the role of ATAD5 and RNA helicases in maintaining genomic integrity by regulating R-loops, the corresponding genes were mutated or downregulated in several human tumors.

scholarly journals Multi-BRCT domain protein Brc1 links Rhp18/Rad18 and γH2A to maintain genome stability during S-phase

2017 ◽  
Author(s):  
Michael C. Reubens ◽  
Sophie Rozenzhak ◽  
Paul Russell
Keyword(s):  
Genome Stability ◽  
Genome Instability ◽  
Replication Stress ◽  
S Phase ◽  
Dna Lesions ◽  
Brct Domain ◽  
Replication Forks ◽  
Histone H2a ◽  
Domain Protein ◽  
Chromosome Integrity

ABSTRACTDNA replication involves the inherent risk of genome instability, as replisomes invariably encounter DNA lesions or other structures that stall or collapse replication forks during S-phase. In the fission yeast Schizosaccharomyces pombe, the multi-BRCT domain protein Brc1, which is related to budding yeast Rtt107 and mammalian PTIP, plays an important role in maintaining genome integrity and cell viability when cells experience replication stress. The C-terminal pair of BRCT domains in Brc1 were previously shown to bind phospho-histone H2A (γH2A) formed by Rad3/ATR checkpoint kinase at DNA lesions; however, the putative scaffold interactions involving the N-terminal BRCT domains 1-4 of Brc1 have remained obscure. Here we show that these domains bind Rhp18/Rad18, which is an E3 ubiquitin protein ligase that has crucial functions in postreplication repair. A missense allele in BRCT domain 4 of Brc1 disrupts binding to Rhp18 and causes sensitivity to replication stress. Brc1 binding to Rhp18 and γH2A are required for the Brc1-overexpression suppression of smc6-74, which impairs the Smc5/6 structural maintenance of chromosomes complex required for chromosome integrity and repair of collapsed replication forks. From these findings we propose that Brc1 provides scaffolding functions linking γH2A, Rhp18, and Smc5/6 complex at damaged replication forks.

scholarly journals Mutations in SID2, a Novel Gene in Saccharomyces cerevisiae, Cause Synthetic Lethality With sic1 Deletion and May Cause a Defect During S Phase

Genetics ◽  
2001 ◽  
Vol 159 (1) ◽  
pp. 17-33
Author(s):  
Matthew D Jacobson ◽  
Claudia X Muñoz ◽  
Kirstin S Knox ◽  
Beth E Williams ◽  
Lenette L Lu ◽  
...  
Keyword(s):  
Dna Damage ◽  
Synthetic Lethality ◽  
S Phase ◽  
Temperature Sensitive ◽  
Replication Forks ◽  
Novel Gene ◽  
Mutant Strains ◽  
Single Nucleus ◽  
2C Dna Content ◽  
Slow Growing

Abstract SIC1 encodes a nonessential B-type cyclin/CDK inhibitor that functions at the G1/S transition and the exit from mitosis. To understand more completely the regulation of these transitions, mutations causing synthetic lethality with sic1Δ were isolated. In this screen, we identified a novel gene, SID2, which encodes an essential protein that appears to be required for DNA replication or repair. sid2-1 sic1Δ strains and sid2-21 temperature-sensitive strains arrest preanaphase as large-budded cells with a single nucleus, a short spindle, and an ~2C DNA content. RAD9, which is necessary for the DNA damage checkpoint, is required for the preanaphase arrest of sid2-1 sic1Δ cells. Analysis of chromosomes in mutant sid2-21 cells by field inversion gel electrophoresis suggests the presence of replication forks and bubbles at the arrest. Deleting the two S phase cyclins, CLB5 and CLB6, substantially suppresses the sid2-1 sic1Δ inviability, while stabilizing Clb5 protein exacerbates the defects of sid2-1 sic1Δ cells. In synchronized sid2-1 mutant strains, the onset of replication appears normal, but completion of DNA synthesis is delayed. sid2-1 mutants are sensitive to hydroxyurea indicating that sid2-1 cells may suffer DNA damage that, when combined with additional insult, leads to a decrease in viability. Consistent with this hypothesis, sid2-1 rad9 cells are dead or very slow growing even when SIC1 is expressed.

scholarly journals The Interactions of DNA Repair, Telomere Homeostasis, and p53 Mutational Status in Solid Cancers: Risk, Prognosis, and Prediction

Cancers ◽  
2021 ◽  
Vol 13 (3) ◽  
pp. 479
Author(s):  
Pavel Vodicka ◽  
Ladislav Andera ◽  
Alena Opattova ◽  
Ludmila Vodickova
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Repair ◽  
Dna Lesions ◽  
Cell Cycle Checkpoint ◽  
Solid Cancer ◽  
Genomic Integrity ◽  
Repair Capacity ◽  
Mutational Status ◽  
Telomere Homeostasis

The disruption of genomic integrity due to the accumulation of various kinds of DNA damage, deficient DNA repair capacity, and telomere shortening constitute the hallmarks of malignant diseases. DNA damage response (DDR) is a signaling network to process DNA damage with importance for both cancer development and chemotherapy outcome. DDR represents the complex events that detect DNA lesions and activate signaling networks (cell cycle checkpoint induction, DNA repair, and induction of cell death). TP53, the guardian of the genome, governs the cell response, resulting in cell cycle arrest, DNA damage repair, apoptosis, and senescence. The mutational status of TP53 has an impact on DDR, and somatic mutations in this gene represent one of the critical events in human carcinogenesis. Telomere dysfunction in cells that lack p53-mediated surveillance of genomic integrity along with the involvement of DNA repair in telomeric DNA regions leads to genomic instability. While the role of individual players (DDR, telomere homeostasis, and TP53) in human cancers has attracted attention for some time, there is insufficient understanding of the interactions between these pathways. Since solid cancer is a complex and multifactorial disease with considerable inter- and intra-tumor heterogeneity, we mainly dedicated this review to the interactions of DNA repair, telomere homeostasis, and TP53 mutational status, in relation to (a) cancer risk, (b) cancer progression, and (c) cancer therapy.

scholarly journals DNA–protein crosslink proteases in genome stability

2021 ◽  
Vol 4 (1) ◽  
Author(s):  
Annamaria Ruggiano ◽  
Kristijan Ramadan
Keyword(s):  
Dna Replication ◽  
Cancer Therapy ◽  
Genome Stability ◽  
Dna Lesions ◽  
Protein Component ◽  
Human Diseases ◽  
Direct Link ◽  
The Past ◽  
Protein Crosslinks ◽  
Bulky Dna Lesions

AbstractProteins covalently attached to DNA, also known as DNA–protein crosslinks (DPCs), are common and bulky DNA lesions that interfere with DNA replication, repair, transcription and recombination. Research in the past several years indicates that cells possess dedicated enzymes, known as DPC proteases, which digest the protein component of a DPC. Interestingly, DPC proteases also play a role in proteolysis beside DPC repair, such as in degrading excess histones during DNA replication or controlling DNA replication checkpoints. Here, we discuss the importance of DPC proteases in DNA replication, genome stability and their direct link to human diseases and cancer therapy.

scholarly journals RuvAB and RecG Are Not Essential for the Recovery of DNA Synthesis Following UV-Induced DNA Damage in Escherichia coli

Genetics ◽  
2004 ◽  
Vol 166 (4) ◽  
pp. 1631-1640 ◽  
Author(s):  
Janet R Donaldson ◽  
Charmain T Courcelle ◽  
Justin Courcelle
Keyword(s):  
Dna Damage ◽  
Dna Synthesis ◽  
Dna Lesions ◽  
Replication Forks ◽  
Wild Type ◽  
Replication Machinery ◽  
Dna Junctions ◽  
Fork Regression

Abstract Ultraviolet light induces DNA lesions that block the progression of the replication machinery. Several models speculate that the resumption of replication following disruption by UV-induced DNA damage requires regression of the nascent DNA or migration of the replication machinery away from the blocking lesion to allow repair or bypass of the lesion to occur. Both RuvAB and RecG catalyze branch migration of three- and four-stranded DNA junctions in vitro and are proposed to catalyze fork regression in vivo. To examine this possibility, we characterized the recovery of DNA synthesis in ruvAB and recG mutants. We found that in the absence of either RecG or RuvAB, arrested replication forks are maintained and DNA synthesis is resumed with kinetics that are similar to those in wild-type cells. The data presented here indicate that RecG- or RuvAB-catalyzed fork regression is not essential for DNA synthesis to resume following arrest by UV-induced DNA damage in vivo.

scholarly journals Interactions and Localization of Escherichia coli Error-Prone DNA Polymerase IV after DNA Damage

2015 ◽  
Vol 197 (17) ◽  
pp. 2792-2809 ◽  
Author(s):  
Sarita Mallik ◽  
Ellen M. Popodi ◽  
Andrew J. Hanson ◽  
Patricia L. Foster
Keyword(s):  
Dna Damage ◽  
Dna Polymerase ◽  
Dna Helicase ◽  
Dna Lesions ◽  
Replication Forks ◽  
Content Type ◽  
Pol Iv ◽  
Dna Polymerase Iv ◽  
Stalled Replication Forks

ABSTRACTEscherichia coli's DNA polymerase IV (Pol IV/DinB), a member of the Y family of error-prone polymerases, is induced during the SOS response to DNA damage and is responsible for translesion bypass and adaptive (stress-induced) mutation. In this study, the localization of Pol IV after DNA damage was followed using fluorescent fusions. After exposure ofE. colito DNA-damaging agents, fluorescently tagged Pol IV localized to the nucleoid as foci. Stepwise photobleaching indicated ∼60% of the foci consisted of three Pol IV molecules, while ∼40% consisted of six Pol IV molecules. Fluorescently tagged Rep, a replication accessory DNA helicase, was recruited to the Pol IV foci after DNA damage, suggesting that thein vitrointeraction between Rep and Pol IV reported previously also occursin vivo. Fluorescently tagged RecA also formed foci after DNA damage, and Pol IV localized to them. To investigate if Pol IV localizes to double-strand breaks (DSBs), an I-SceI endonuclease-mediated DSB was introduced close to a fluorescently labeled LacO array on the chromosome. After DSB induction, Pol IV localized to the DSB site in ∼70% of SOS-induced cells. RecA also formed foci at the DSB sites, and Pol IV localized to the RecA foci. These results suggest that Pol IV interacts with RecAin vivoand is recruited to sites of DSBs to aid in the restoration of DNA replication.IMPORTANCEDNA polymerase IV (Pol IV/DinB) is an error-prone DNA polymerase capable of bypassing DNA lesions and aiding in the restart of stalled replication forks. In this work, we demonstratein vivolocalization of fluorescently tagged Pol IV to the nucleoid after DNA damage and to DNA double-strand breaks. We show colocalization of Pol IV with two proteins: Rep DNA helicase, which participates in replication, and RecA, which catalyzes recombinational repair of stalled replication forks. Time course experiments suggest that Pol IV recruits Rep and that RecA recruits Pol IV. These findings providein vivoevidence that Pol IV aids in maintaining genomic stability not only by bypassing DNA lesions but also by participating in the restoration of stalled replication forks.

Replication forks pause at yeast centromeres

1992 ◽  
Vol 12 (9) ◽  
pp. 4056-4066
Author(s):  
S A Greenfeder ◽  
C S Newlon
Keyword(s):  
Gel Electrophoresis ◽  
Replication Fork ◽  
S Phase ◽  
Core Structure ◽  
Replication Forks ◽  
Agarose Gel Electrophoresis ◽  
Unusual Structure ◽  
Dna Complex ◽  
Derivatives Of ◽  
Pause Site

The 120 bp of yeast centromeric DNA is tightly complexed with protein to form a nuclease-resistant core structure 200 to 240 bp in size. We have used two-dimensional agarose gel electrophoresis to analyze the replication of the chromosomal copies of yeast CEN1, CEN3, and CEN4 and determine the fate of replication forks that encounter the protein-DNA complex at the centromere. We have shown that replication fork pause sites are coincident with each of these centromeres and therefore probably with all yeast centromeres. We have analyzed the replication of plasmids containing mutant derivatives of CEN3 to determine whether the replication fork pause site is a result of an unusual structure adopted by centromere DNA or a result of the protein-DNA complex formed at the centromere. The mutant centromere derivatives varied in function as well as the ability to form the nuclease-resistant core structure. The data obtained from analysis of these derivatives indicate that the ability to cause replication forks to pause correlates with the ability to form the nuclease-resistant core structure and not with the presence or absence of a particular DNA sequence. Our findings further suggest that the centromere protein-DNA complex is present during S phase when replication forks encounter the centromere and therefore may be present throughout the cell cycle.

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