scholarly journals XRCC1 Is Specifically Associated with Poly(ADP-Ribose) Polymerase and Negatively Regulates Its Activity following DNA Damage

1998 ◽  
Vol 18 (6) ◽  
pp. 3563-3571 ◽  
Author(s):  
Murielle Masson ◽  
Claude Niedergang ◽  
Valérie Schreiber ◽  
Sylviane Muller ◽  
Josiane Menissier-de Murcia ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Cell Cycle Checkpoint ◽  
Dna Breaks ◽  
Protective Function ◽  
C Terminus ◽  
Base Excision

ABSTRACT Poly(ADP-ribose) polymerase (PARP; EC 2.4.2.30 ) is a zinc-finger DNA-binding protein that detects and signals DNA strand breaks generated directly or indirectly by genotoxic agents. In response to these breaks, the immediate poly(ADP-ribosyl)ation of nuclear proteins involved in chromatin architecture and DNA metabolism converts DNA damage into intracellular signals that can activate DNA repair programs or cell death options. To have greater insight into the physiological function of this enzyme, we have used the two-hybrid system to find genes encoding proteins putatively interacting with PARP. We have identified a physical association between PARP and the base excision repair (BER) protein XRCC1 (X-ray repair cross-complementing 1) in theSaccharomyces cerevisiae system, which was further confirmed to exist in mammalian cells. XRCC1 interacts with PARP by its central region (amino acids 301 to 402), which contains a BRCT (BRCA1 C terminus) module, a widespread motif in DNA repair and DNA damage-responsive cell cycle checkpoint proteins. Overexpression of XRCC1 in Cos-7 or HeLa cells dramatically decreases PARP activity in vivo, reinforcing the potential protective function of PARP at DNA breaks. Given that XRCC1 is also associated with DNA ligase III via a second BRCT module and with DNA polymerase β, our results provide strong evidence that PARP is a member of a BER multiprotein complex involved in the detection of DNA interruptions and possibly in the recruitment of XRCC1 and its partners for efficient processing of these breaks in a coordinated manner. The modular organizations of these interactors, associated with small conserved domains, may contribute to increasing the efficiency of the overall pathway.

scholarly journals How are base excision DNA repair pathways deployed in vivo?

F1000Research ◽  
2017 ◽  
Vol 6 ◽  
pp. 279 ◽  
Author(s):  
Upasna Thapar ◽  
Bruce Demple
Keyword(s):  
Dna Repair ◽  
Base Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Biochemical Level ◽  
Repair Pathways ◽  
Base Excision ◽  
Base Excision Dna Repair

Since the discovery of the base excision repair (BER) system for DNA more than 40 years ago, new branches of the pathway have been revealed at the biochemical level by in vitro studies. Largely for technical reasons, however, the confirmation of these subpathways in vivo has been elusive. We review methods that have been used to explore BER in mammalian cells, indicate where there are important knowledge gaps to fill, and suggest a way to address them.

scholarly journals ZGRF1 promotes end resection of DNA homologous recombination via forming complex with BRCA1/EXO1

2021 ◽  
Vol 7 (1) ◽  
Author(s):  
Shuang Yan ◽  
Man Song ◽  
Jie Ping ◽  
Shu-ting Lai ◽  
Xiao-yu Cao ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Dna Damage ◽  
Dna Repair ◽  
Homologous Recombination ◽  
Mammalian Cells ◽  
Cell Cycle Checkpoint ◽  
Dependent Manner ◽  
Dna Breaks ◽  
M Cell ◽  
End Resection

AbstractTo maintain genomic stability, the mammalian cells has evolved a coordinated response to DNA damage, including activation of DNA repair and cell cycle checkpoint processes. Exonuclease 1 (EXO1)-dependent excision of DNA ends is important for the initiation of homologous recombination (HR) repair of DNA breaks, which is thought to play a key role in activating the ATR-CHK1 pathway to induce G2/M cell cycle arrest. But the mechanism is still not fully understood. Here, we report that ZGRF1 forms complexes with EXO1 as well as other repair proteins and promotes DNA repair through HR. ZGRF1 is recruited to DNA damage sites in a MDC1-RNF8-BRCA1 dependent manner. Furthermore, ZGRF1 is important for the recruitment of RPA2 to DNA damage sites and the following ATR-CHK1 mediated G2/M checkpoint in response to irradiation. ZGRF1 null cells show increased sensitivity to many DNA-damaging agents, especially PARPi and irradiation. Collectively,our findings identify ZGRF1 as a novel regulator of DNA end resection and G2/M checkpoint. ZGRF1 is a potential target of radiation and PARPi cancer therapy.

scholarly journals An interaction between the mammalian DNA repair protein XRCC1 and DNA ligase III.

1994 ◽  
Vol 14 (1) ◽  
pp. 68-76 ◽  
Author(s):  
K W Caldecott ◽  
C K McKeown ◽  
J D Tucker ◽  
S Ljungquist ◽  
L H Thompson
Keyword(s):  
Dna Repair ◽  
Affinity Chromatography ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Affinity Purification ◽  
Alkylating Agents ◽  
Chinese Hamster ◽  
Dna Ligase ◽  
Base Excision ◽  
Dna Ligase Iii

XRCC1, the human gene that fully corrects the Chinese hamster ovary DNA repair mutant EM9, encodes a protein involved in the rejoining of DNA single-strand breaks that arise following treatment with alkylating agents or ionizing radiation. In this study, a cDNA minigene encoding oligohistidine-tagged XRCC1 was constructed to facilitate affinity purification of the recombinant protein. This construct, designated pcD2EHX, fully corrected the EM9 phenotype of high sister chromatid exchange, indicating that the histidine tag was not detrimental to XRCC1 activity. Affinity chromatography of extract from EM9 cells transfected with pcD2EHX resulted in the copurification of histidine-tagged XRCC1 and DNA ligase III activity. Neither XRCC1 or DNA ligase III activity was purified during affinity chromatography of extract from EM9 cells transfected with pcD2EX, a cDNA minigene that encodes untagged XRCC1, or extract from wild-type AA8 or untransfected EM9 cells. The copurification of DNA ligase III activity with histidine-tagged XRCC1 suggests that the two proteins are present in the cell as a complex. Furthermore, DNA ligase III activity was present at lower levels in EM9 cells than in AA8 cells and was returned to normal levels in EM9 cells transfected with pcD2EHX or pcD2EX. These findings indicate that XRCC1 is required for normal levels of DNA ligase III activity, and they implicate a major role for this DNA ligase in DNA base excision repair in mammalian cells.

scholarly journals Dynamic Compartmentalization of Base Excision Repair Proteins in Response to Nuclear and Mitochondrial Oxidative Stress

2008 ◽  
Vol 29 (3) ◽  
pp. 794-807 ◽  
Author(s):  
Lyra M. Griffiths ◽  
Dan Swartzlander ◽  
Kellen L. Meadows ◽  
Keith D. Wilkinson ◽  
Anita H. Corbett ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Dna Damage ◽  
Dna Repair ◽  
Base Excision Repair ◽  
Oxidative Dna Damage ◽  
Excision Repair ◽  
Intracellular Localization ◽  
Genotoxic Stress ◽  
Mitochondrial Oxidative Stress ◽  
Base Excision

ABSTRACT DNAs harbored in both nuclei and mitochondria of eukaryotic cells are subject to continuous oxidative damage resulting from normal metabolic activities or environmental insults. Oxidative DNA damage is primarily reversed by the base excision repair (BER) pathway, initiated by N-glycosylase apurinic/apyrimidinic (AP) lyase proteins. To execute an appropriate repair response, BER components must be distributed to accommodate levels of genotoxic stress that may vary considerably between nuclei and mitochondria, depending on the growth state and stress environment of the cell. Numerous examples exist where cells respond to signals, resulting in relocalization of proteins involved in key biological transactions. To address whether such dynamic localization contributes to efficient organelle-specific DNA repair, we determined the intracellular localization of the Saccharomyces cerevisiae N-glycosylase/AP lyases, Ntg1 and Ntg2, in response to nuclear and mitochondrial oxidative stress. Fluorescence microscopy revealed that Ntg1 is differentially localized to nuclei and mitochondria, likely in response to the oxidative DNA damage status of the organelle. Sumoylation is associated with targeting of Ntg1 to nuclei containing oxidative DNA damage. These studies demonstrate that trafficking of DNA repair proteins to organelles containing high levels of oxidative DNA damage may be a central point for regulating BER in response to oxidative stress.

scholarly journals Databases and Bioinformatics Tools for the Study of DNA Repair

2011 ◽  
Vol 2011 ◽  
pp. 1-9 ◽  
Author(s):  
Kaja Milanowska ◽  
Kristian Rother ◽  
Janusz M. Bujnicki
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Excision Repair ◽  
Uv Light ◽  
Dna Lesions ◽  
Environmental Chemicals ◽  
Nonhomologous End Joining ◽  
End Joining ◽  
Repair Mechanisms ◽  
Base Excision

DNA is continuously exposed to many different damaging agents such as environmental chemicals, UV light, ionizing radiation, and reactive cellular metabolites. DNA lesions can result in different phenotypical consequences ranging from a number of diseases, including cancer, to cellular malfunction, cell death, or aging. To counteract the deleterious effects of DNA damage, cells have developed various repair systems, including biochemical pathways responsible for the removal of single-strand lesions such as base excision repair (BER) and nucleotide excision repair (NER) or specialized polymerases temporarily taking over lesion-arrested DNA polymerases during the S phase in translesion synthesis (TLS). There are also other mechanisms of DNA repair such as homologous recombination repair (HRR), nonhomologous end-joining repair (NHEJ), or DNA damage response system (DDR). This paper reviews bioinformatics resources specialized in disseminating information about DNA repair pathways, proteins involved in repair mechanisms, damaging agents, and DNA lesions.

scholarly journals The ATPase Domain but Not the Acidic Region of Cockayne Syndrome Group B Gene Product Is Essential for DNA Repair

1999 ◽  
Vol 10 (11) ◽  
pp. 3583-3594 ◽  
Author(s):  
Robert M. Brosh ◽  
Adayabalam S. Balajee ◽  
Rebecca R. Selzer ◽  
Morten Sunesen ◽  
Luca Proietti De Santis ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Excision Repair ◽  
Uv Light ◽  
Genetic Disorder ◽  
Cockayne Syndrome ◽  
Premature Aging ◽  
Atpase Domain ◽  
Acidic Region

Cockayne syndrome (CS) is a human genetic disorder characterized by UV sensitivity, developmental abnormalities, and premature aging. Two of the genes involved, CSA andCSB, are required for transcription-coupled repair (TCR), a subpathway of nucleotide excision repair that removes certain lesions rapidly and efficiently from the transcribed strand of active genes. CS proteins have also been implicated in the recovery of transcription after certain types of DNA damage such as those lesions induced by UV light. In this study, site-directed mutations have been introduced to the human CSB gene to investigate the functional significance of the conserved ATPase domain and of a highly acidic region of the protein. The CSB mutant alleles were tested for genetic complementation of UV-sensitive phenotypes in the human CS-B homologue of hamster UV61. In addition, theCSB mutant alleles were tested for their ability to complement the sensitivity of UV61 cells to the carcinogen 4-nitroquinoline-1-oxide (4-NQO), which introduces bulky DNA adducts repaired by global genome repair. Point mutation of a highly conserved glutamic acid residue in ATPase motif II abolished the ability of CSB protein to complement the UV-sensitive phenotypes of survival, RNA synthesis recovery, and gene-specific repair. These data indicate that the integrity of the ATPase domain is critical for CSB function in vivo. Likewise, the CSB ATPase point mutant failed to confer cellular resistance to 4-NQO, suggesting that ATP hydrolysis is required for CSB function in a TCR-independent pathway. On the contrary, a large deletion of the acidic region of CSB protein did not impair the genetic function in the processing of either UV- or 4-NQO-induced DNA damage. Thus the acidic region of CSB is likely to be dispensable for DNA repair, whereas the ATPase domain is essential for CSB function in both TCR-dependent and -independent pathways.

Potentiation of Melphalan-Induced Cytotoxicity through Targeting of the Base Excision Repair Pathway in Multiple Myeloma.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 4799-4799
Author(s):  
April M. Reed ◽  
Melissa L. Fishel ◽  
Mark R. Kelley ◽  
Rafat Abonour
Keyword(s):  
Multiple Myeloma ◽  
Dna Damage ◽  
Dna Repair ◽  
Side Effects ◽  
Small Molecule ◽  
Base Excision Repair ◽  
Excision Repair ◽  
Chemotherapeutic Agents ◽  
Repair Pathways ◽  
Base Excision

Abstract Melphalan (M) is an active agent against multiple myeloma (MM). One of the obstacles with M treatment is the patient’s ability to tolerate side effects such as mucositis and pancytopenia. This is especially true for those patients >70 years of age. We hypothesize that potentiation of M-induced cytotoxicity is possible in MM with agents that target, and therefore further imbalance, multiple DNA repair pathways. A key protein in the Base Excision Repair (BER) pathway, Apurinic/apyrimidinic endonuclease/ redox factor (APE1/Ref-1 or APE1) plays a major role in the repair of damage caused by chemotherapeutic agents including M and Temozolomide (TMZ), interacts with a number of transcription factors (HIF1-a, p53, AP1, NFkB, etc) to regulate their function through oxidation/reduction (redox) signaling, and is overexpressed in refractory/relapsed MM cells. Furthermore, a reduction in APE1 protein sensitizes MM cells to melphalan indicating that inhibition of this protein may have therapeutic potential in MM. In order to decipher the importance of APE1’s redox and repair functions in MM cells’ response to DNA damage via melphalan and TMZ, we have available to us small molecule APE1 inhibitors that affect only the repair activity or only the redox activity of APE1. The mechanism of action of MLP is primarily via monoadduct leading to DNA interstrand cross-link (ICL) formation which is processed by the Nucleotide Excision Repair (NER) pathway. MLP also causes N7methyl-G and N3methyl-A adduct formation which are repaired by the BER pathway. For these studies, we treated RPMI 8226 cells with several chemotherapeutic agents: M; TMZ, which creates primarily N7methyl-G and N3methyl-A adducts; Methoxyamine (MX), which has been shown to inhibit further processing by the BER pathway; and a small molecule which blocks the redox function of APE1. Our purpose was to overwhelm the DNA repair pathways by causing the accumulation of DNA repair intermediates and inducing apoptosis. M-induced cytotoxicity is enhanced by TMZ (CI=0.08), MX (CI=0.89), and E3330 (CI=0.06), and this effect was synergistic as determined by CalcuSyn software which generates median effect and combinational index (CI) values, with CI<1 indicative of synergy. Using MX to inhibit APE1 in combination with TMZ results in an increase in DNA damage and an increase in apoptosis in 8226 cells. Furthermore, the combination of the redox inhibitor + MX which blocks both functions of APE1 also results in an increase in apoptosis in the MM cells. Further studies include the addition of M to these combinations that are demonstrating an increase in efficacy in MM cells. These results indicate that using these DNA repair-targeted agents in addition to MLP may be a feasible way to increase the effect of the M on MM cells. The potential advantages to patients would be that they would be able to tolerate more treatments and that the combination treatments would be more effective than treatment with M alone. We anticipate that effective modulation of M and/or TMZ will overcome resistance without compromising efficacy and help to alleviate some of the side effects patients have to endure with melphalan treatment. This may be particularly advantageous to the more elderly patients.

scholarly journals Long‐patch DNA repair synthesis during base excision repair in mammalian cells

EMBO Reports ◽  
2003 ◽  
Vol 4 (4) ◽  
pp. 363-367 ◽  
Author(s):  
Ulrike Sattler ◽  
Philippe Frit ◽  
Bernard Salles ◽  
Patrick Calsou
Keyword(s):  
Dna Repair ◽  
Base Excision Repair ◽  
Mammalian Cells ◽  
Excision Repair ◽  
Repair Synthesis ◽  
Base Excision ◽  
Dna Repair Synthesis

scholarly journals Kinetic Modeling Reveals the Roles of Reactive Oxygen Species Scavenging and DNA Repair Processes in Shaping the Dose-Response Curve of KBrO3-Induced DNA Damage

2015 ◽  
Vol 2015 ◽  
pp. 1-12 ◽  
Author(s):  
Maria A. Spassova ◽  
David J. Miller ◽  
Alexander S. Nikolov
Keyword(s):  
Reactive Oxygen Species ◽  
Dna Damage ◽  
Dna Repair ◽  
Base Excision Repair ◽  
Dose Response ◽  
Excision Repair ◽  
Reactive Oxygen ◽  
Oxygen Species ◽  
Repair Processes ◽  
Base Excision

We have developed a kinetic model to investigate how DNA repair processes and scavengers of reactive oxygen species (ROS) can affect the dose-response shape of prooxidant induced DNA damage. We used as an example chemicalKBrO3which is activated by glutathione and forms reactive intermediates that directly interact with DNA to form 8-hydroxy-2-deoxyguanosine DNA adducts (8-OH-dG). The single strand breaks (SSB) that can result from failed base excision repair of these adducts were considered as an effect downstream from 8-OH-dG. We previously demonstrated that, in the presence of effective base excision repair, 8-OH-dG can exhibit threshold-like dose-response dependence, while the downstream SSB can still exhibit a linear dose-response. Here we demonstrate that this result holds for a variety of conditions, including low levels of GSH, the presence of additional SSB repair mechanisms, or a scavenger. It has been shown that melatonin, a terminal scavenger, inhibitsKBrO3-caused oxidative damage. Our modeling revealed that sustained exposure toKBrO3can lead to fast scavenger exhaustion, in which case the dose-response shapes for both endpoints are not substantially affected. The results are important to consider when forming conclusions on a chemical’s toxicity dose dependence based on the dose-response of early genotoxic events.

scholarly journals DNA Excision Repair and DNA Damage-Induced Apoptosis Are Linked to Poly(ADP-Ribosyl)ation but Have Different Requirements for p53

2000 ◽  
Vol 20 (18) ◽  
pp. 6695-6703 ◽  
Author(s):  
Ralph Beneke ◽  
Christoph Geisen ◽  
Branko Zevnik ◽  
Thomas Bauch ◽  
Wolfgang-Ulrich Müller ◽  
...  
Keyword(s):  
Dna Damage ◽  
Cell Death ◽  
Dna Repair ◽  
Zinc Finger Protein ◽  
Excision Repair ◽  
Dna Breaks ◽  
In Vivo Models ◽  
P53 Expression ◽  
Dna Excision Repair ◽  
Parp Activity

ABSTRACT Poly(ADP-ribose) polymerase (PARP) is a DNA binding zinc finger protein that catalyzes the transfer of ADP-ribose residues from NAD+ to itself and different chromatin constituents, forming branched ADP-ribose polymers. The enzymatic activity of PARP is induced upon DNA damage and the PARP protein is cleaved during apoptosis, which suggested a role of PARP in DNA repair and DNA damage-induced cell death. We have generated transgenic mice that lack PARP activity in thymocytes owing to the targeted expression of a dominant negative form of PARP. In the presence of single-strand DNA breaks, the absence of PARP activity correlated with a strongly increased rate of apoptosis compared to cells with intact PARP activity. We found that blockage of PARP activity leads to a drastic increase of p53 expression and activity after DNA damage and correlates with an accelerated onset of Bax expression. DNA repair is almost completely blocked in PARP-deficient thymocytes regardless of p53 status. We found the same increased susceptibility to apoptosis in PARP null mice, a similar inhibition of DNA repair kinetics, and the same upregulation of p53 in response to DNA damage. Thus, based on two different experimental in vivo models, we identify a direct, p53-independent, functional connection between poly(ADP-ribosyl)ation and the DNA excision repair machinery. Furthermore, we propose a p53-dependent link between PARP activity and DNA damage-induced cell death.

Sign in / Sign up

Export Citation Format

Share Document