scholarly journals DNA damage and oxidizing conditions activate p53 through differential upstream signaling pathways

2020 ◽  
Author(s):  
Tao Shi ◽  
Paulien E. Polderman ◽  
Boudewijn M.T. Burgering ◽  
Tobias B. Dansen
Keyword(s):  
Reactive Oxygen Species ◽  
Dna Damage ◽  
Double Strand Breaks ◽  
Oxygen Species ◽  
Strand Breaks ◽  
Dna Damage Signaling ◽  
Cycle Arrest ◽  
Dna Damaging Agents ◽  
Tumor Tissues ◽  
Cellular Stresses

AbstractStabilization and activation of the p53 tumour suppressor are triggered in response to various cellular stresses, including DNA damaging agents and elevated Reactive Oxygen Species (ROS) like H2O2. When cells are exposed to exogenously added H2O2, ATR/CHK1 and ATM/CHK2 dependent DNA damage signaling is switched on, suggesting that H2O2 induces both single and double strand breaks. These collective observations have resulted in the widely accepted model that oxidizing conditions lead to DNA damage that subsequently mediates a p53-dependent response like cell cycle arrest and apoptosis. However, H2O2 induces signaling through stress-activated kinases (SAPK, e.g., JNK and p38MAPK) that can activate p53. Here we dissect to what extent these pathways contribute to functional activation of p53 in response to oxidizing conditions. Collectively, our data suggest that p53 can be activated both by SAPK signaling and the DDR independently of each other, and which of these pathways is activated depends on the type of oxidant used. This implies that it could in principle be possible to modulate redox signaling to stimulate p53 without inducing collateral DNA damage, thereby limiting mutation accumulation in both healthy and tumor tissues.

BCR/ABL Oncogenic Kinase Promotes Unfaithful Repair of the Reactive Oxygen Species - Dependent DNA Double-Strand Breaks.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 712-712 ◽  
Author(s):  
Tomasz Skorski ◽  
Michal O. Nowicki ◽  
Rafal Falinski ◽  
Mateusz Koptyra ◽  
Artur Slupianek ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Cell Cycle ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Double Strand Breaks ◽  
Transformed Cells ◽  
Oxygen Species ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Abl Kinase

Abstract The oncogenic BCR/ABL tyrosine kinase induces constitutive DNA damage in Philadelphia chromosome (Ph1)-positive leukemia cells. We find that BCR/ABL kinase - induced reactive oxygen species (ROS) cause chronic oxidative DNA damage as indicated by an enzymatic assay detecting oxidized bases. These DNA lesions result in DNA double-strand breaks (DSBs) detected by comet assay, immunofluorescent gamma-H2AX nuclear foci and linker-ligation PCR (LL-PCR). Combined analysis of the length of LL-PCR products and the sequences of two reference genes DR-GFP and Na+/K+ ATPase revealed that ROS dependent DSBs occurred in the regions containing multiple, 5–9bp long stretches of G/C, in concordance with the notion that oxidative DNA damage is predominantly detected in G/C-rich sequences. Elevated numbers of DSBs were detected in BCR/ABL cell lines, murine bone marrow cells transformed with BCR/ABL and in CML patient samples, in comparison to normal counterparts. Inhibition of the BCR/ABL kinase by STI571 and diminishion of ROS activity by the ROS scavenger PDTC reduced DSBs formation. Cell cycle analysis revealed that most of these DSBs occur during S and G2/M phases, and are probably associated with the stalled replication forks. Homologous recombination repair (HRR) and non-homologous end-joining (NHEJ) represent two major mechanisms of DSBs repair in S and G2/M cell cycle phase. Using the in vivo recombination assay consisting of the DSB-dependent reconstitution of the green fluorescent protein (GFP) gene we found that HRR is stimulated in BCR/ABL-positive cells. In addition, in vitro assay measuring the activity of NHEJ revealed that this repair process is also activated by the BCR/ABL kinase. RAD51 and Ku70 play a key role in HRR and NHEJ, respectively. The reaction sites of HRR and NHEJ in the nuclei could be visualized by double-immunofluorescence detecting co-localization of gamma-H2AX foci (DSBs sites) with RAD51 (HRR sites) or Ku70 (NHEJ sites). Equal co-localization frequency of gamma-H2AX foci with RAD51 and Ku70 was detected, suggesting that both HRR and NHEJ play an important role in reparation of ROS-dependent DSBs in BCR/ABL-transformed cells. Analysis of the DSBs repair products in the reporter DR-GFP gene in BCR/ABL cells identified ~40% of HRR and ~60% of NHEJ events. Sequencing revealed point-mutations in HRR products and large deletions in NHEJ products in BCR/ABL-positive cells, but not in non-transformed cells. We propose that the following series of events may contribute to genomic instability of Ph1-positive leukemias: BCR/ABL → ROS → oxidative DNA damage → DSBs in proliferating cells → unfaithful HRR and NHEJ repair. Since BCR/ABL share many similarities with other members of the fusion tyrosine kinases (FTKs) family, these events may contribute to genomic instability of hematological malignancies caused by FTKs.

scholarly journals Through Its Nonstructural Protein NS1, Parvovirus H-1 Induces Apoptosis via Accumulation of Reactive Oxygen Species

2010 ◽  
Vol 84 (12) ◽  
pp. 5909-5922 ◽  
Author(s):  
Georgi Hristov ◽  
Melanie Krämer ◽  
Junwei Li ◽  
Nazim El-Andaloussi ◽  
Rodrigo Mora ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Nonstructural Protein ◽  
Reactive Oxygen ◽  
Double Strand Breaks ◽  
Oxygen Species ◽  
Ns1 Protein ◽  
Dna Double Strand Breaks ◽  
Caspase 9 ◽  
Strand Breaks ◽  
Cycle Arrest

ABSTRACT The rat parvovirus H-1 (H-1PV) attracts high attention as an anticancer agent, because it is not pathogenic for humans and has oncotropic and oncosuppressive properties. The viral nonstructural NS1 protein is thought to mediate H-1PV cytotoxicity, but its exact contribution to this process remains undefined. In this study, we analyzed the effects of the H-1PV NS1 protein on human cell proliferation and cell viability. We show that NS1 expression is sufficient to induce the accumulation of cells in G2 phase, apoptosis via caspase 9 and 3 activation, and cell lysis. Similarly, cells infected with wild-type H-1PV arrest in G2 phase and undergo apoptosis. Furthermore, we also show that both expression of NS1 and H-1PV infection lead to higher levels of intracellular reactive oxygen species (ROS), associated with DNA double-strand breaks. Antioxidant treatment reduces ROS levels and strongly decreases NS1- and virus-induced DNA damage, cell cycle arrest, and apoptosis, indicating that NS1-induced ROS are important mediators of H-1PV cytotoxicity.

BCR/ABL oncogenic kinase promotes unfaithful repair of the reactive oxygen species–dependent DNA double-strand breaks

Blood ◽  
2004 ◽  
Vol 104 (12) ◽  
pp. 3746-3753 ◽  
Author(s):  
Michal O. Nowicki ◽  
Rafal Falinski ◽  
Mateusz Koptyra ◽  
Artur Slupianek ◽  
Tomasz Stoklosa ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Dna Damage ◽  
Oxidative Dna Damage ◽  
Philadelphia Chromosome ◽  
Reactive Oxygen ◽  
Double Strand Breaks ◽  
Oxygen Species ◽  
M Cell ◽  
Proliferating Cells ◽  
Strand Breaks

The oncogenic BCR/ABL tyrosine kinase induces constitutive DNA damage in Philadelphia chromosome (Ph)-positive leukemia cells. We find that BCR/ABL-induced reactive oxygen species (ROSs) cause chronic oxidative DNA damage resulting in double-strand breaks (DSBs) in S and G2/M cell cycle phases. These lesions are repaired by BCR/ABL-stimulated homologous recombination repair (HRR) and nonhomologous end-joining (NHEJ) mechanisms. A high mutation rate is detected in HRR products in BCR/ABL-positive cells, but not in the normal counterparts. In addition, large deletions are found in NHEJ products exclusively in BCR/ABL cells. We propose that the following series of events may contribute to genomic instability of Ph-positive leukemias: BCR/ABL → ROSs → oxidative DNA damage → DSBs in proliferating cells → unfaithful HRR and NHEJ repair.

scholarly journals RexAB promotes the survival of Staphylococcus aureus exposed to multiple classes of antibiotics

2021 ◽  
Author(s):  
Rebecca S. Clarke ◽  
Kam Pou Ha ◽  
Andrew M. Edwards
Keyword(s):  
Reactive Oxygen Species ◽  
Staphylococcus Aureus ◽  
Dna Damage ◽  
Sos Response ◽  
Double Strand Breaks ◽  
Oxygen Species ◽  
Bacterial Survival ◽  
Wild Type ◽  
Dna Double Strand Breaks ◽  
Strand Breaks

Antibiotics inhibit essential bacterial processes, resulting in arrest of growth and in some cases cell death. Many antibiotics are also reported to trigger endogenous production of reactive oxygen species (ROS), which damage DNA, leading to induction of the mutagenic SOS response associated with the emergence of drug resistance. However, the type of DNA damage that arises and how this triggers the SOS response is largely unclear. We found that several different classes of antibiotic triggered dose-dependent induction of the SOS response in Staphylococcus aureus , indicative of DNA damage, including some bacteriostatic drugs. The SOS response was heterogenous and varied in magnitude between strains and antibiotics. However, in many cases, full induction of the SOS response was dependent upon the RexAB helicase/nuclease complex, which processes DNA double strand breaks to produce single-stranded DNA and facilitate RecA nucleoprotein filament formation. The importance of RexAB in repair of DNA was confirmed by measuring bacterial survival during antibiotic exposure, with most drugs having significantly greater bactericidal activity against rexB mutants relative to wild type strains. For some, but not all antibiotics there was no difference in bactericidal activity between wild type and rexB mutant under anaerobic conditions, indicative of a role for reactive oxygen species in mediating DNA damage. Taken together, this work confirms previous observations that several classes of antibiotics cause DNA damage in S. aureus and extends them by showing that processing of DNA double strand breaks by RexAB is a major trigger of the mutagenic SOS response and promotes bacterial survival.

scholarly journals Alterations in hormonal signals spatially coordinate distinct responses to DNA double-strand breaks in Arabidopsis roots

2021 ◽  
Vol 7 (25) ◽  
pp. eabg0993
Author(s):  
Naoki Takahashi ◽  
Soichi Inagaki ◽  
Kohei Nishimura ◽  
Hitoshi Sakakibara ◽  
Ioanna Antoniadi ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Root Apex ◽  
Double Strand Breaks ◽  
Root Tip ◽  
Dna Double Strand Breaks ◽  
Cytokinin Signaling ◽  
Strand Breaks ◽  
Dna Damage Signaling ◽  
Cycle Arrest

Plants have a high ability to cope with changing environments and grow continuously throughout life. However, the mechanisms by which plants strike a balance between stress response and organ growth remain elusive. Here, we found that DNA double-strand breaks enhance the accumulation of cytokinin hormones through the DNA damage signaling pathway in the Arabidopsis root tip. Our data showed that activation of cytokinin signaling suppresses the expression of some of the PIN-FORMED genes that encode efflux carriers of another hormone, auxin, thereby decreasing the auxin signals in the root tip and causing cell cycle arrest at G2 phase and stem cell death. Elevated cytokinin signaling also promotes an early transition from cell division to endoreplication in the basal part of the root apex. We propose that plant hormones spatially coordinate differential DNA damage responses, thereby maintaining genome integrity and minimizing cell death to ensure continuous root growth.

scholarly journals DNA double-strand breaks induced by reactive oxygen species promote DNA polymerase IV activity inEscherichia coli

2019 ◽  
Author(s):  
Sarah S. Henrikus ◽  
Camille Henry ◽  
John P. McDonald ◽  
Yvonne Hellmich ◽  
Elizabeth A. Wood ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Dna Damage ◽  
Dna Polymerase ◽  
Reactive Oxygen ◽  
Double Strand Breaks ◽  
Oxygen Species ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Pol Iv ◽  
Dna Polymerase Iv

Under many conditions the killing of bacterial cells by antibiotics is potentiated by DNA damage induced by reactive oxygen species (ROS)1–3. A primary cause of ROS-induced cell death is the accumulation of DNA double-strand breaks (DSBs)1,4–6. DNA polymerase IV (pol IV), an error-prone DNA polymerase produced at elevated levels in cells experiencing DNA damage, has been implicated both in ROS-dependent killing and in DSBR7–15. Here, we show using single-molecule fluorescence microscopy that ROS-induced DSBs promote pol IV activity in two ways. First, exposure to the antibiotics ciprofloxacin and trimethoprim triggers an SOS-mediated increase in intracellular pol IV concentrations that is strongly dependent on both ROS and DSBR. Second, in cells that constitutively express pol IV, treatment with an ROS scavenger dramatically reduces the number of pol IV foci formed upon exposure to antibiotics, indicating a role for pol IV in the repair of ROS-induced DSBs.

scholarly journals Staphylococcal DNA Repair Is Required for Infection

mBio ◽  
2020 ◽  
Vol 11 (6) ◽  
Author(s):  
Kam Pou Ha ◽  
Rebecca S. Clarke ◽  
Gyu-Lee Kim ◽  
Jane L. Brittan ◽  
Jessica E. Rowley ◽  
...  
Keyword(s):  
Reactive Oxygen Species ◽  
Staphylococcus Aureus ◽  
Human Blood ◽  
Immune Cells ◽  
Double Strand Breaks ◽  
Oxygen Species ◽  
Dna Double Strand Breaks ◽  
Gram Positive ◽  
Content Type ◽  
Strand Breaks

ABSTRACT To cause infection, Staphylococcus aureus must withstand damage caused by host immune defenses. However, the mechanisms by which staphylococcal DNA is damaged and repaired during infection are poorly understood. Using a panel of transposon mutants, we identified the rexBA operon as being important for the survival of Staphylococcus aureus in whole human blood. Mutants lacking rexB were also attenuated for virulence in murine models of both systemic and skin infections. We then demonstrated that RexAB is a member of the AddAB family of helicase/nuclease complexes responsible for initiating the repair of DNA double-strand breaks. Using a fluorescent reporter system, we were able to show that neutrophils cause staphylococcal DNA double-strand breaks through reactive oxygen species (ROS) generated by the respiratory burst, which are repaired by RexAB, leading to the induction of the mutagenic SOS response. We found that RexAB homologues in Enterococcus faecalis and Streptococcus gordonii also promoted the survival of these pathogens in human blood, suggesting that DNA double-strand break repair is required for Gram-positive bacteria to survive in host tissues. Together, these data demonstrate that DNA is a target of host immune cells, leading to double-strand breaks, and that the repair of this damage by an AddAB-family enzyme enables the survival of Gram-positive pathogens during infection. IMPORTANCE To cause infection, bacteria must survive attack by the host immune system. For many bacteria, including the major human pathogen Staphylococcus aureus, the greatest threat is posed by neutrophils. These immune cells ingest the invading organisms and try to kill them with a cocktail of chemicals that includes reactive oxygen species (ROS). The ability of S. aureus to survive this attack is crucial for the progression of infection. However, it was not clear how the ROS damaged S. aureus and how the bacterium repaired this damage. In this work, we show that ROS cause breaks in the staphylococcal DNA, which must be repaired by a two-protein complex known as RexAB; otherwise, the bacterium is killed, and it cannot sustain infection. This provides information on the type of damage that neutrophils cause S. aureus and the mechanism by which this damage is repaired, enabling infection.

scholarly journals RAD51 localization and activation following DNA damage

2004 ◽  
Vol 359 (1441) ◽  
pp. 87-93 ◽  
Author(s):  
Madalena Tarsounas ◽  
Adelina A. Davies ◽  
Stephen C. West
Keyword(s):  
Dna Damage ◽  
Genome Stability ◽  
Critical Role ◽  
S Phase ◽  
Double Strand Breaks ◽  
Focus Formation ◽  
Rad51 Protein ◽  
Strand Breaks ◽  
Dna Damaging Agents ◽  
Binding Level

The efficient repair of double–strand breaks in DNA is critical for the maintenance of genome stability. In response to ionizing radiation and other DNA–damaging agents, the RAD51 protein, which is essential for homologous recombination, relocalizes within the nucleus to form distinct foci that can be visualized by microscopy and are thought to represent sites where repair reactions take place. The formation of RAD51 foci in response to DNA damage is dependent upon BRCA2 and a series of proteins known as the RAD51 paralogues (RAD51B, RAD51C, RAD51D, XRCC2 and XRCC3), indicating that the components present within foci assemble in a carefully orchestrated and ordered manner. By contrast, RAD51 foci that form spontaneously as cells undergo DNA replication at S phase occur without the need for BRCA2 or the RAD51 paralogues. It is known that BRCA2 interacts directly with RAD51 through a series of degenerative motifs known as the BRC repeats. These interactions modulate the ability of RAD51 to bind DNA. Taken together, these observations indicate that BRCA2 plays a critical role in controlling the actions of RAD51 at both the microscopic (focus formation) and molecular (DNA binding) level.

scholarly journals STK38 promotes ATM activation by acting as a reader of histone H4 ufmylation

2020 ◽  
Vol 6 (23) ◽  
pp. eaax8214 ◽  
Author(s):  
Bo Qin ◽  
Jia Yu ◽  
Somaira Nowsheen ◽  
Fei Zhao ◽  
Liewei Wang ◽  
...  
Keyword(s):  
Dna Damage ◽  
Hippo Pathway ◽  
Double Strand Breaks ◽  
Binding Motif ◽  
Histone H4 ◽  
Ataxia Telangiectasia Mutated ◽  
Independent Manner ◽  
Strand Breaks ◽  
Dna Damage Signaling ◽  
The Hippo Pathway

The ATM (ataxia-telangiectasia mutated) kinase is rapidly activated following DNA damage and phosphorylates its downstream targets to launch DDR signaling. Recently, we and others showed that UFM1 signaling promotes ATM activation. We further discovered that monoufmylation of histone H4 at Lys31 by UFM1-specific ligase 1 (UFL1) is an important step in the amplification of ATM activation. However, how monoufmylated H4 enhances ATM activation is still unknown. Here, we report STK38, a kinase in the Hippo pathway, serves as a reader for histone H4 ufmylation to promote ATM activation in a kinase-independent manner. STK38 contains a potential UFM1 binding motif which recognizes ufmylated H4 and recruits the SUV39H1 to the double-strand breaks, resulting in H3K9 trimethylation and Tip60 activation to promote ATM activation. Together, STK38 is a previously unknown player in DNA damage signaling and functions as a reader of monoufmylated H4 at Lys31 to promote ATM activation.

scholarly journals DNA double-strand breaks in the Toxoplasma gondii-infected cells by the action of reactive oxygen species

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Haohan Zhuang ◽  
Chaoqun Yao ◽  
Xianfeng Zhao ◽  
Xueqiu Chen ◽  
Yimin Yang ◽  
...  
Keyword(s):  
Dna Damage ◽  
Toxoplasma Gondii ◽  
Double Strand Breaks ◽  
Host Cells ◽  
Oxygen Species ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Damage Response ◽  
Infected Cells

Abstract Background Toxoplasma gondii is an obligate parasite of all warm-blooded animals around the globe. Once infecting a cell, it manipulates the host’s DNA damage response that is yet to be elucidated. The objectives of the present study were three-fold: (i) to assess DNA damages in T. gondii-infected cells in vitro; (ii) to ascertain causes of DNA damage in T. gondii-infected cells; and (iii) to investigate activation of DNA damage responses during T. gondii infection. Methods HeLa, Vero and HEK293 cells were infected with T. gondii at a multiplicity of infection (MOI) of 10:1. Infected cells were analyzed for a biomarker of DNA double-strand breaks (DSBs) γH2AX at 10 h, 20 h or 30 h post-infection using both western blot and immunofluorescence assay. Reactive oxygen species (ROS) levels were measured using 2′,7′-dichlorodihydrofluorescein diacetate (H2DCFDA), and ROS-induced DNA damage was inhibited by a ROS inhibitor N-acetylcysteine (NAC). Lastly, DNA damage responses were evaluated by detecting the active form of ataxia telangiectasia mutated/checkpoint kinase 2 (ATM/CHK2) by western blot. Results γH2AX levels in the infected HeLa cells were significantly increased over time during T. gondii infection compared to uninfected cells. NAC treatment greatly reduced ROS and concomitantly diminished γH2AX in host cells. The phosphorylated ATM/CHK2 were elevated in T. gondii-infected cells. Conclusions Toxoplasma gondii infection triggered DNA DSBs with ROS as a major player in host cells in vitro. It also activated DNA damage response pathway ATM/CHK2. Toxoplasma gondii manages to keep a balance between survival and apoptosis of its host cells for the benefit of its own survival.

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