Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “DNA Replication Stress”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 469 records · Page 26Linked to original sources
PubMed1998

The mutagenesis proteins UmuD' and UmuC prevent lethal frameshifts while increasing base substitution mutations.

Error-prone DNA repair consists of replicative filling-in of DNA gaps carrying lesions. We have reconstituted E. coli SOS error-prone repair using purified DNA polymerase III holoenzyme, SSB, RecA, UmuD', a UmuC fusion protein, and a gap lesion plasmid. In the absence of UmuDC, or without SOS induction, replication skips over the lesion, forming mostly one-nucleotide deletions. These cause translational frameshifts that usually inactivate genes. UmuD' and UmuC, in the presence of RecA and SSB, stimulate translesion replication and change its mutagenic specificity such that deletions are prevented and base substitutions are increased. This results in mutagenic but nondetrimental gap repair and provides an effective mechanism for generating genetic variation in bacteria adapting to environmental stress.

Bacterial Proteins↗
PubMed2005

A biological network in Saccharomyces cerevisiae prevents the deleterious effects of endogenous oxidative DNA damage.

In this study, we used Saccharomyces cerevisiae to identify a biological network that prevents the deleterious effects of endogenous reactive oxygen species. The absence of Tsa1, a key peroxiredoxin, caused increased rates of mutations, chromosomal rearrangements, and recombination. Defects in recombinational DNA double strand break repair, Rad6-mediated postreplicative repair, and DNA damage and replication checkpoints caused growth defects or lethality in the absence of Tsa1. In addition, the mutator phenotypes caused by a tsa1 mutation were significantly aggravated by defects in Ogg1, mismatch repair, or checkpoints. These results indicate that increased endogenous oxidative stress has broad effects on genome stability and is highly sensitive to the functional state of DNA repair and checkpoints. These findings may provide insight in understanding the consequences of various pathophysiological processes in regard to genomic instability.

DNA Damage↗
PubMed2004

Hallmarks of senescence in carcinogenesis and cancer therapy.

Cellular senescence is a signal transduction program leading to irreversible cell cycle arrest. This growth arrest can be triggered by many different mechanisms including recognition by cellular sensors of DNA double-strand breaks leading to the activation of cell cycle checkpoint responses and recruitment of DNA repair foci. Senescence is initiated by the shortening of telomeres (replicative senescence) or by other endogenous and exogenous acute and chronic stress signals (STASIS: stress or aberrant signaling-induced senescence). The process of carcinogenesis involves a series of changes that allow tumor cells to bypass the senescence program. Nevertheless, tumor cells retain the capacity to undergo senescence. Treatment of tumor cells with many conventional anticancer therapies activates DNA damage signaling pathways, which induce apoptosis in some cells and senescence in others. Overexpression of tumor suppressors or inhibition of oncogenes can also induce rapid senescence in tumor cells. Senescent cells, while not dividing, remain metabolically active and produce many secreted factors, some of which stimulate and others inhibit the growth of tumors. The emerging knowledge about the pathways that lead to senescence and determine the pattern of gene expression in senescent cells may lead to more effective treatments for cancer.

Animals↗
PubMed2006

Hepatocyte nuclear factor (HNF) 1 and HNF4 mediate hepatic multidrug resistance protein 2 up-regulation during hepatitis C virus gene expression.

Hepatitis C virus (HCV) is known to induce hepatic oxidative stress that is implicated in the up-regulation of multidrug resistance proteins (MRPs). The relationship between increased prooxidant production, MRPs, and HCV has not been investigated. Here, we report that a homeodomain-containing transcription factor, hepatocyte nuclear factor (HNF) 1, plays a central role in liver gene regulation during HCV gene expression and/or subgenome replication. MRP2 protein and mRNA expression were increased and MRP2 promoter activity was increased 7-fold. Mutations within the putative HNF1 binding site of the human MRP2 promoter abrogated HCV-induced activation, implicating HNF1 in the induction of MRP2 by HCV. The mechanism by which HNF1-mediated activation occurs seems to be transcriptional, because the regulated expression of HNF4, which is known to control HNF1 expression, was also increased. Consistent with this finding, HNF1 mRNA was increased 10-fold. A promoter-luciferase construct of the human HNF1 gene was activated in an HNF4-dependent manner, and a mutant construct lacking the HNF4 binding site was not activated in HCV-positive cells. Consistent with this hypothesis, HNF4 protein and mRNA levels as well as HNF4 promoter activity and DNA binding activity were increased. The expression of HNF1 seems to play a critical role in the induction of hepatic MRP2 secondary to HCV subgenomic replication. The ability of HCV to induce HNF1 and HNF4 is attributed to 1) increased oxidative stress and 2) direct protein-protein interactions between HCV nonstructural component (NS) 5A and HNF1, leading to enhanced HNF1 DNA binding. In conclusion, we describe a novel mechanism by which HCV gene expression may induce adaptive responses involving MRP2 via HNF1 activation. This may constitute, in part, the cellular detoxification task force during HCV infection.

Cell Line↗
PubMed2006

Involvement of the ATR- and ATM-dependent checkpoint responses in cell cycle arrest evoked by pierisin-1.

Pierisin-1 identified from the cabbage butterfly, Pieris rapae, is a novel mono-ADP-ribosylating toxin that transfers the ADP-ribose moiety of NAD at N(2) of dG in DNA. Resulting mono-ADP-ribosylated DNA adducts cause mutations and the induction of apoptosis. However, little is known about checkpoint responses elicited in mammalian cells by the formation of such bulky DNA adducts. In the present study, it was shown that DNA polymerases were blocked at the specific site of mono-ADP-ribosylated dG, which might lead to the replication stress. Pierisin-1 treatment of HeLa cells was found to induce an intra-S-phase arrest through both ataxia telangiectasia mutated (ATM) and Rad3-related (ATR) and ATM pathways, and ATR pathway also contributes to a G(2)-M-phase delay. In the colony survival assays, Rad17(-/-) DT40 cells showed greater sensitivity to pierisin-1-induced cytotoxicity than wild-type and ATM(-/-) DT40 cells, possibly due to defects of checkpoint responses, such as the Chk1 activation. Furthermore, apoptotic 50-kb DNA fragmentation was observed in the HeLa cells, which was well correlated with occurrence of phosphorylation of Chk2. These results thus suggest that pierisin-1 treatment primarily activates ATR pathway and eventually activates ATM pathway as a result of the induction of apoptosis. From these findings, it is suggested that mono-ADP-ribosylation of DNA causes a specific type of fork blockage that induces checkpoint activation and signaling.

ADP Ribose Transferases↗
PubMed2004

Error-prone replication for better or worse.

Precise genome duplication requires accurate copying by DNA polymerases and the elimination of occasional mistakes by proofreading exonucleases and mismatch repair enzymes. The commonly held belief that 'if something is worth doing, then it's worth doing well' normally applies to DNA replication and repair, however, there are exceptions. This review describes elements that are crucial to cell fitness, evolution and survival in the recently discovered error-prone DNA polymerases. Large numbers of errant DNA polymerases, spanning microorganisms to humans, are used to rescue stalled replication forks by copying damaged DNA and even undamaged DNA to generate 'purposeful' mutations that generate genetic diversity in times of stress. Here we focus on low-fidelity polymerases from bacteria, comparing Escherichia coli, archeabacteria and those most recently discovered in Gram-positive Bacilli, Streptococcus, pathogenic Mycobacterium and intein-containing cyanobacteria.

Adaptation, Biological↗
PubMed1998

The beta subunit sliding DNA clamp is responsible for unassisted mutagenic translesion replication by DNA polymerase III holoenzyme.

The replication of damaged nucleotides that have escaped DNA repair leads to the formation of mutations caused by misincorporation opposite the lesion. In Escherichia coli, this process is under tight regulation of the SOS stress response and is carried out by DNA polymerase III in a process that involves also the RecA, UmuD' and UmuC proteins. We have shown that DNA polymerase III holoenzyme is able to replicate, unassisted, through a synthetic abasic site in a gapped duplex plasmid. Here, we show that DNA polymerase III*, a subassembly of DNA polymerase III holoenzyme lacking the beta subunit, is blocked very effectively by the synthetic abasic site in the same DNA substrate. Addition of the beta subunit caused a dramatic increase of at least 28-fold in the ability of the polymerase to perform translesion replication, reaching 52% bypass in 5 min. When the ssDNA region in the gapped plasmid was extended from 22 nucleotides to 350 nucleotides, translesion replication still depended on the beta subunit, but it was reduced by 80%. DNA sequence analysis of translesion replication products revealed mostly -1 frameshifts. This mutation type is changed to base substitution by the addition of UmuD', UmuC, and RecA, as demonstrated in a reconstituted SOS translesion replication reaction. These results indicate that the beta subunit sliding DNA clamp is the major determinant in the ability of DNA polymerase III holoenzyme to perform unassisted translesion replication and that this unassisted bypass produces primarily frameshifts.

DNA Damage↗
PubMed2006

Chromosomal telomere attrition as a mechanism for the increased risk of epithelial cancers and senescent phenotypes in type 2 diabetes.

Telomeres are the repeat DNA sequences at the end of chromosomes necessary for successful DNA replication and chromosomal integrity. Telomeres shorten at cell division at a rate determined by oxidative DNA damage, and cells are triggered into replicative senescence once telomeres shorten to a critical length. Telomere-related chromosomal maintenance also has a role in carcinogenesis. Type 2 diabetes is characterised by increased oxidative stress, increased oxidative DNA damage, senescent retinal and renal phenotypes, and an increased risk of epithelial malignancy. We suggest that increased oxidative DNA damage and telomere attrition in type 2 diabetes leads to: (1) carcinogenic telomere-dependent chromosomal non-reciprocal translocations, genomic instability, and the development of epithelial cancers; (2) senescent retinal and renal phenotypes (expressed as diabetic retinopathy and nephropathy); and (3) senescent vascular endothelial, monocyte-macrophage and vascular smooth muscle cells (expressed as endothelial dysfunction and accelerated atherogenesis). An adverse intrauterine environment leads to increased feto-placental oxidative stress and feto-placental oxidative DNA damage. We also suggest that intrauterine oxidative DNA damage and telomere shortening is another point at which increased oxidative stress could contribute to a pre-programmed increased risk of senescent phenotypes in adult offspring, characterised by type 2 diabetes and epithelial malignancy. These suggestions can be used to understand early glucose intolerance in the young children of type 1 diabetes pregnancies, poor cancer outcomes in type 2 diabetes, beta cell fatigue in type 2 diabetes and the absence of increased epithelial cancer risk in type 1 diabetes.

Carcinoma↗
PubMed2005

Cleavage of claspin by caspase-7 during apoptosis inhibits the Chk1 pathway.

Claspin is required for the phosphorylation and activation of the Chk1 protein kinase by ATR during DNA replication and in response to DNA damage. This checkpoint pathway plays a critical role in the resistance of cells to genotoxic stress. Here, we show that human Claspin is cleaved by caspase-7 during the initiation of apoptosis. In cells, induction of DNA damage by etoposide at first produced rapid phosphorylation of Chk1 at a site targeted by ATR. Subsequently, etoposide caused activation of caspase-7, cleavage of Claspin, and dephosphorylation of Chk1. In apoptotic cell extracts, Claspin was cleaved by caspase-7 at a single aspartate residue into a large N-terminal fragment and a smaller C-terminal fragment that contain different functional domains. The large N-terminal fragment was heavily phosphorylated in a human cell-free system in response to double-stranded DNA oligonucleotides, and this fragment retained Chk1 binding activity. In contrast, the smaller C-terminal fragment did not bind Chk1, but did associate with DNA and inhibited the DNA-dependent phosphorylation of Chk1 associated with its activation. These results indicate that cleavage of Claspin by caspase-7 inactivates the Chk1 signaling pathway. This mechanism may regulate the balance between cell cycle arrest and induction of apoptosis during the response to genotoxic stress.

Adaptor Proteins, Signal Transducing↗
PubMed2005

Mitochondrial DNA in somatic cells: a promising target of routine clinical tests.

Alterations of mitochondrial DNA have long been considered only from a point of view of rare genetic disorders causing neuromyopathy. Recently, alterations of mitochondrial DNA have been found in so-called common diseases such as heart failure, diabetes, and cancer; some of these alterations are inherited, and some are generated and/or accumulated in somatic cells with age. Mitochondrial DNA is more vulnerable to alteration than is nuclear DNA. For example, mitochondria produce a large amount of reactive oxygen species as an inevitable byproduct of oxidative phosphorylation. Therefore, mitochondrial DNA is under much stronger oxidative stress than is nuclear DNA. In spite of the importance, it is much less elucidated in the mitochondrial genome than in the nuclear genome how the genome is maintained. In this review, we focus on maintenance of mitochondrial DNA in somatic cells and its clinical importance.

Animals↗
PubMed1995

Metabolic rate and directional nucleotide substitution in animal mitochondrial DNA.

There is marked heterogeneity of nucleotide composition in mitochondrial DNA across divergent animals. Differences in nucleotide composition presumably reflect differences in directional nucleotide substitution for A+T or G+C nucleotides. In mitochondrial DNA, there is A+T directional nucleotide substitution in most (if not all) animals surveyed, and the magnitude of directional A+T nucleotide substitution differs greatly within and among groups. Differences in directional nucleotide substitution among lineages of mammals can be explained by changes in metabolic physiology. This relationship is thought to be mediated by the effect of oxygen radicals because these toxic compounds are by-products of aerobic metabolism and are known mutagens. Association between metabolism and nucleotide composition provides additional evidence in favor of the hypothesis that rates and patterns of nucleotide substitution in mitochondrial DNA can be influenced by factors that impinge on rates of endogenous DNA damage.

Animals↗
PubMed1984

In vitro response of chondrocytes to mechanical loading. The effect of short term mechanical tension.

Chick epiphyseal chondrocytes were grown in high density cultures for 14 days, after which the cell layers were placed in a cyclical stretching apparatus and subjected to a strain of 5.5% at a frequency of 0.2 Hz. There was a 1.4- and 1.7-fold increase in the incorporation of 35SO4 and 14C-glucosamine, respectively, into glycosaminoglycans in cultures subjected to mechanical loading for 24 h. No significant change was observed in the hydrodynamic size of the proteoglycans synthesized by chondrocytes subjected to mechanical loading. In this time period there was no increase in 3H-glycine incorporation into acid-insoluble protein, but there was a 2.4-fold increase of 3H-thymidine into DNA in cultures subjected to tensional strain. Concomitant with these changes, the cellular levels of cyclic AMP increased 2.2 times in the mechanically loaded cultures. This is discussed as a possible mechanism whereby chondrocytes respond to mechanical stimuli.

Animals↗
PubMed2024

Reversion from basal histone H4 hypoacetylation at the replication fork increases DNA damage in FANCA deficient cells.

The FA/BRCA pathway safeguards DNA replication by repairing interstrand crosslinks (ICL) and maintaining replication fork stability. Chromatin structure, which is in part regulated by histones posttranslational modifications (PTMs), has a role in maintaining genomic integrity through stabilization of the DNA replication fork and promotion of DNA repair. An appropriate balance of PTMs, especially acetylation of histones H4 in nascent chromatin, is required to preserve a stable DNA replication fork. To evaluate the acetylation status of histone H4 at the replication fork of FANCA deficient cells, we compared histone acetylation status at the DNA replication fork of isogenic FANCA deficient and FANCA proficient cell lines by using accelerated native immunoprecipitation of nascent DNA (aniPOND) and in situ protein interactions in the replication fork (SIRF) assays. We found basal hypoacetylation of multiple residues of histone H4 in FA replication forks, together with increased levels of Histone Deacetylase 1 (HDAC1). Interestingly, high-dose short-term treatment with mitomycin C (MMC) had no effect over H4 acetylation abundance at the replication fork. However, chemical inhibition of histone deacetylases (HDAC) with Suberoylanilide hydroxamic acid (SAHA) induced acetylation of the FANCA deficient DNA replication forks to levels comparable to their isogenic control counterparts. This forced permanence of acetylation impacted FA cells homeostasis by inducing DNA damage and promoting G2 cell cycle arrest. Altogether, this caused reduced RAD51 foci formation and increased markers of replication stress, including phospho-RPA-S33. Hypoacetylation of the FANCA deficient replication fork, is part of the cellular phenotype, the perturbation of this feature by agents that prevent deacetylation, such as SAHA, have a deleterious effect over the delicate equilibrium they have reached to perdure despite a defective FA/BRCA pathway.

Histones↗
PubMed2007

Increased common fragile site expression, cell proliferation defects, and apoptosis following conditional inactivation of mouse Hus1 in primary cultured cells.

Targeted disruption of the mouse Hus1 cell cycle checkpoint gene results in embryonic lethality and proliferative arrest in cultured cells. To investigate the essential functions of Hus1, we developed a system for the regulated inactivation of mouse Hus1 in primary fibroblasts. Inactivation of a loxP site-flanked conditional Hus1 allele by using a cre-expressing adenovirus resulted in reduced cell doubling, cell cycle alterations, and increased apoptosis. These phenotypes were associated with a significantly increased frequency of gross chromosomal abnormalities and an S-phase-specific accumulation of phosphorylated histone H2AX, an indicator of double-stranded DNA breaks. To determine whether these chromosomal abnormalities occurred randomly or at specific genomic regions, we assessed the stability of common fragile sites, chromosomal loci that are prone to breakage in cells undergoing replication stress. Hus1 was found to be essential for fragile site stability, because spontaneous chromosomal abnormalities occurred preferentially at common fragile sites upon conditional Hus1 inactivation. Although p53 levels increased after Hus1 loss, deletion of p53 failed to rescue the cell-doubling defect or increased apoptosis in conditional Hus1 knockout cells. In summary, we propose that Hus1 loss leads to chromosomal instability during DNA replication, triggering increased apoptosis and impaired proliferation through p53-independent mechanisms.

Animals↗
PubMed2014

The N-terminus of Mcm10 is important for interaction with the 9-1-1 clamp and in resistance to DNA damage.

Accurate replication of the genome requires the evolutionarily conserved minichromosome maintenance protein, Mcm10. Although the details of the precise role of Mcm10 in DNA replication are still debated, it interacts with the Mcm2-7 core helicase, the lagging strand polymerase, DNA polymerase-α and the replication clamp, proliferating cell nuclear antigen. Loss of these interactions caused by the depletion of Mcm10 leads to chromosome breakage and cell cycle checkpoint activation. However, whether Mcm10 has an active role in DNA damage prevention is unknown. Here, we present data that establish a novel role of the N-terminus of Mcm10 in resisting DNA damage. We show that Mcm10 interacts with the Mec3 subunit of the 9-1-1 clamp in response to replication stress evoked by UV irradiation or nucleotide shortage. We map the interaction domain with Mec3 within the N-terminal region of Mcm10 and demonstrate that its truncation causes UV light sensitivity. This sensitivity is not further enhanced by a deletion of MEC3, arguing that MCM10 and MEC3 operate in the same pathway. Since Rad53 phosphorylation in response to UV light appears to be normal in N-terminally truncated mcm10 mutants, we propose that Mcm10 may have a role in replication fork restart or DNA repair.

Cell Cycle Proteins↗
PubMed2000

Heat stress reduces poly(ADPR)polymerase expression in rat testis.

Poly(ADPR)polymerase (PARP) is a chromatin-associated enzyme with a presumptive role in DNA repair during replication and recovery from strand breaks caused by genotoxic agents. It catalyses the attachment and elongation of ADPribose polymers (pADPR) to a variety of acceptor proteins (including PARP itself, and histones) and is particularly active in the testis where its expression varies according to the stage of germ cell differentiation. PARP degradation is also one of the classic indicators of apoptosis. In this investigation we have examined the effects of heat stress on the adult rat testis with respect to the concentration and activity of PARP, the nature of the pADRP nuclear acceptor proteins, the length of ADPR polymers and the activity of the ADPR depolymerizing enzyme, poly(ADPR)glycohydrolase (PARG). Our results show a significant reduction in the concentration and activity of PARP 4 and 8 days after artificial cryptorchidism, but no significant changes were observed in PARG activity or in pADPR length. Unexpectedly, the apoptotic degradation of PARP was not detected following heat stress. These results confirm that PARP gene expression is developmentally regulated during spermatogenesis and indicate that it is suppressed coincidentally with the loss of meiotic spermatocytes during artificial cryptorchidism.

Animals↗
PubMed2005

Replication checkpoint kinase Cds1 regulates Mus81 to preserve genome integrity during replication stress.

The replication checkpoint kinase Cds1 preserves genome integrity by stabilizing stalled replication forks. Cds1 targets substrates through its FHA domain. The Cds1 FHA domain interacts with Mus81, a subunit of the Mus81-Eme1 structure-specific endonuclease. We report here that Mus81 and Rhp51 are required for generating deletion mutations in fission yeast replication mutants that experience replication stress. A mutation in the Mus81 FHA-binding motif eliminates its Cds1-binding and Cds1-dependent phosphorylation. Furthermore, this mutation exacerbates the deletion mutator phenotype of a replication mutant, and induces a hyper-recombination phenotype in hydroxyurea-treated cells. In unperturbed cells, Mus81 associates with chromatin throughout S phase. In replication mutants grown at semipermissive temperature, Mus81 undergoes minor Cds1-dependent phosphorylation, remains chromatin-associated, generates deletion mutations, and maintains cell growth. Upon S-phase arrest by acute hydroxyurea treatment, Mus81 is not required for cell viability but is essential for recovery from replication fork collapse. Moreover, Mus81 undergoes extensive Cds1-dependent phosphorylation and dissociates from chromatin in hydroxyurea-arrested cells, thereby preventing it from cleaving stalled replication forks that could lead to fork breakage and chromosomal rearrangement. These results provide novel insights into how Cds1 regulates Mus81 accordingly when cells experience different replication stress to preserve genome integrity.

Checkpoint Kinase 2↗
PubMed2006

Evidence that the ATR/Chk1 pathway maintains normal replication fork progression during unperturbed S phase.

If cells are treated with DNA damaging agents or inhibitors that interfere with ongoing DNA replication, the intra-S and S/M checkpoints delay progression through S phase and mitotic entry, respectively, to allow time for DNA repair and replication restart. In vertebrates, these checkpoint responses to replication blocks are largely mediated by the sensor kinase ATR and its major downstream effector kinase Chk1. Increasing evidence suggests that the ATR pathway is also vital in the absence of exogenous stresses, i.e., during "unperturbed" replication. Both ATR and Chk1 are essential proteins in vertebrates, and lack of components of the ATR/Chk1 pathway can result in impaired replication and spontaneous DNA damage. Here we give an overview of how the ATR/Chk1 pathway responds to exogenously blocked replication and then describe evidence for roles of this pathway during replication in an unperturbed S phase.

Animals↗