scholarly journals Specific Human ATR and ATM Inhibitors Modulate Single Strand DNA Formation in Leishmania major Exposed to Oxidative Agent

2022 ◽  
Vol 11 ◽  
Author(s):  
Raíssa Bernardes da Silva ◽  
Willian dos Reis Bertoldo ◽  
Lucila Langoni Naves ◽  
Fernanda Bernadelli de Vito ◽  
Jeziel Dener Damasceno ◽  
...  
Keyword(s):  
Dna Repair ◽  
Ataxia Telangiectasia ◽  
Molecular Mechanisms ◽  
Leishmania Major ◽  
Neglected Tropical Diseases ◽  
Nuclease Activity ◽  
Single Strand ◽  
Specific Inhibition ◽  
Dna Strands ◽  
Repair Pathways

Leishmania parasites are the causative agents of a group of neglected tropical diseases known as leishmaniasis. The molecular mechanisms employed by these parasites to adapt to the adverse conditions found in their hosts are not yet completely understood. DNA repair pathways can be used by Leishmania to enable survival in the interior of macrophages, where the parasite is constantly exposed to oxygen reactive species. In higher eukaryotes, DNA repair pathways are coordinated by the central protein kinases ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3 related (ATR). The enzyme Exonuclease-1 (EXO1) plays important roles in DNA replication, repair, and recombination, and it can be regulated by ATM- and ATR-mediated signaling pathways. In this study, the DNA damage response pathways in promastigote forms of L. major were investigated using bioinformatics tools, exposure of lineages to oxidizing agents and radiation damage, treatment of cells with ATM and ATR inhibitors, and flow cytometry analysis. We demonstrated high structural and important residue conservation for the catalytic activity of the putative LmjEXO1. The overexpression of putative LmjEXO1 made L. major cells more susceptible to genotoxic damage, most likely due to the nuclease activity of this enzyme and the occurrence of hyper-resection of DNA strands. These cells could be rescued by the addition of caffeine or a selective ATM inhibitor. In contrast, ATR-specific inhibition made the control cells more susceptible to oxidative damage in an LmjEXO1 overexpression-like manner. We demonstrated that ATR-specific inhibition results in the formation of extended single-stranded DNA, most likely due to EXO1 nucleasic activity. Antagonistically, ATM inhibition prevented single-strand DNA formation, which could explain the survival phenotype of lineages overexpressing LmjEXO1. These results suggest that an ATM homolog in Leishmania could act to promote end resection by putative LmjEXO1, and an ATR homologue could prevent hyper-resection, ensuring adequate repair of the parasite DNA.

scholarly journals Molecular Mechanisms of the Whole DNA Repair System: A Comparison of Bacterial and Eukaryotic Systems

2010 ◽  
Vol 2010 ◽  
pp. 1-32 ◽  
Author(s):  
Rihito Morita ◽  
Shuhei Nakane ◽  
Atsuhiro Shimada ◽  
Masao Inoue ◽  
Hitoshi Iino ◽  
...  
Keyword(s):  
Dna Repair ◽  
Molecular Mechanisms ◽  
Excision Repair ◽  
Thermus Thermophilus ◽  
Repair System ◽  
System A ◽  
Recombination Repair ◽  
Repair Mechanisms ◽  
Repair Pathways ◽  
Base Excision

DNA is subjected to many endogenous and exogenous damages. All organisms have developed a complex network of DNA repair mechanisms. A variety of different DNA repair pathways have been reported: direct reversal, base excision repair, nucleotide excision repair, mismatch repair, and recombination repair pathways. Recent studies of the fundamental mechanisms for DNA repair processes have revealed a complexity beyond that initially expected, with inter- and intrapathway complementation as well as functional interactions between proteins involved in repair pathways. In this paper we give a broad overview of the whole DNA repair system and focus on the molecular basis of the repair machineries, particularly inThermus thermophilusHB8.

Role of EXO1 nuclease activity in genome maintenance, the immune response and tumor suppression in Exo1D173A mice

2021 ◽  
Author(s):  
Shanzhi Wang ◽  
Kyeryoung Lee ◽  
Stephen Gray ◽  
Yongwei Zhang ◽  
Catherine Tang ◽  
...  
Keyword(s):  
Dna Repair ◽  
Somatic Hypermutation ◽  
Nuclease Activity ◽  
Strand Break ◽  
Mutant Form ◽  
Metabolic Processes ◽  
Dna Mismatch ◽  
Antibody Diversification ◽  
Repair Pathways ◽  
V Region

ABSTRACTDNA damage response pathways rely extensively on nuclease activity to process DNA intermediates. Exonuclease 1 (EXO1) is a pleiotropic evolutionary conserved DNA exonuclease involved in various DNA repair pathways, replication, antibody diversification, and meiosis. But, whether EXO1 facilitates these DNA metabolic processes through its enzymatic or scaffolding functions remains unclear. Here we dissect the contribution of EXO1 enzymatic versus scaffolding activity by comparing Exo1DA/DA mice expressing a proven nuclease-dead mutant form of EXO1 to entirely EXO1-deficient Exo1−/− and EXO1 wild type Exo1+/+ mice. We show that Exo1DA/DA and Exo1−/− mice are compromised in canonical DNA repair processing, suggesting that the EXO1 enzymatic role is important for error-free DNA mismatch and double-strand break repair pathways. However, in non-canonical repair pathways, EXO1 appears to have a more nuanced function. Next-generation sequencing of heavy chain V region in B cells showed the mutation spectra of Exo1DA/DA mice to be intermediate between Exo1+/+ and Exo1−/− mice, suggesting that both catalytic and scaffolding roles of EXO1 are important for somatic hypermutation. Similarly, while overall class switch recombination in Exo1DA/DA and Exo1−/− mice was comparably defective, switch-switch junction analysis suggests that EXO1 might fulfill an additional scaffolding function downstream of class switching. In contrast to Exo1−/− mice that are infertile, meiosis progressed normally in Exo1DA/DA and Exo1+/+ cohorts, indicating that a structural but not the nuclease function of EXO1 is critical for meiosis. However, both Exo1DA/DA and Exo1−/− mice displayed similar mortality and cancer predisposition profiles. Taken together, these data demonstrate that EXO1 has both scaffolding and enzymatic functions in distinct DNA repair processes and suggest a more composite and intricate role for EXO1 in DNA metabolic processes and disease.

scholarly journals Recent Perspectives in Radiation-Mediated DNA Damage and Repair: Role of NHEJ and Alternative Pathways

2021 ◽  
Author(s):  
Ajay Kumar Sharma ◽  
Priyanka Shaw ◽  
Aman Kalonia ◽  
M.H. Yashavarddhan ◽  
Pankaj Chaudhary ◽  
...  
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Double Strand Breaks ◽  
Single Strand ◽  
End Joining ◽  
Strand Breaks ◽  
Single Strand Annealing ◽  
Repair Pathways ◽  
Non Homologous End Joining ◽  
Alternative Nhej

Radiation is one of the causative agents for the induction of DNA damage in biological systems. There is various possibility of radiation exposure that might be natural, man-made, intentional, or non-intentional. Published literature indicates that radiation mediated cell death is primarily due to DNA damage that could be a single-strand break, double-strand breaks, base modification, DNA protein cross-links. The double-strand breaks are lethal damage due to the breakage of both strands of DNA. Mammalian cells are equipped with strong DNA repair pathways that cover all types of DNA damage. One of the predominant pathways that operate DNA repair is a non-homologous end-joining pathway (NHEJ) that has various integrated molecules that sense, detect, mediate, and repair the double-strand breaks. Even after a well-coordinated mechanism, there is a strong possibility of mutation due to the flexible nature in joining the DNA strands. There are alternatives to NHEJ pathways that can repair DNA damage. These pathways are alternative NHEJ pathways and single-strand annealing pathways that also displayed a role in DNA repair. These pathways are not studied extensively, and many reports are showing the relevance of these pathways in human diseases. The chapter will very briefly cover the radiation, DNA repair, and Alternative repair pathways in the mammalian system. The chapter will help the readers to understand the basic and applied knowledge of radiation mediated DNA damage and its repair in the context of extensively studied NHEJ pathways and unexplored alternative NHEJ pathways.

Functional interaction between Fanconi anemia and mTOR pathways during stalled replication fork recovery

2020 ◽  
Author(s):  
Matthew Nolan ◽  
Kenneth Knudson ◽  
Marina K Holz ◽  
Indrajit Chaudhury
Keyword(s):  
Dna Repair ◽  
Dna Replication ◽  
Fanconi Anemia ◽  
Replication Fork ◽  
Genomic Stability ◽  
Repair Pathway ◽  
Functional Interaction ◽  
Mechanistic Target Of Rapamycin ◽  
Dna Strands ◽  
Repair Pathways

We have previously demonstrated that Fanconi Anemia (FA) proteins work in concert with other FA and non-FA proteins to mediate stalled replication fork restart. Previous studies suggest a connection between FA protein FANCD2 and a non-FA protein mechanistic target of rapamycin (mTOR). A recent study showed that mTOR is involved in actin-dependent DNA replication fork restart, suggesting possible roles in FA DNA repair pathway. In this study, we demonstrate that during replication stress mTOR interacts and cooperates with FANCD2 to provide cellular stability, mediates stalled replication fork restart and prevents nucleolytic degradation of the nascent DNA strands. Taken together, this study unravels a novel functional cross-talk between two important mechanisms: mTOR and FA DNA repair pathways that ensure genomic stability.

scholarly journals DNA Repair Repertoire of the Enigmatic Hydra

2021 ◽  
Vol 12 ◽  
Author(s):  
Apurva Barve ◽  
Alisha A. Galande ◽  
Saroj S. Ghaskadbi ◽  
Surendra Ghaskadbi
Keyword(s):  
Dna Repair ◽  
Molecular Mechanisms ◽  
Large Body ◽  
Evolutionary Developmental Biology ◽  
Model Systems ◽  
Repair Capacity ◽  
Apparent Lack ◽  
Dna Repair Capacity ◽  
Repair Pathways ◽  
Repair Genes

Since its discovery by Abraham Trembley in 1744, hydra has been a popular research organism. Features like spectacular regeneration capacity, peculiar tissue dynamics, continuous pattern formation, unique evolutionary position, and an apparent lack of organismal senescence make hydra an intriguing animal to study. While a large body of work has taken place, particularly in the domain of evolutionary developmental biology of hydra, in recent years, the focus has shifted to molecular mechanisms underlying various phenomena. DNA repair is a fundamental cellular process that helps to maintain integrity of the genome through multiple repair pathways found across taxa, from archaea to higher animals. DNA repair capacity and senescence are known to be closely associated, with mutations in several repair pathways leading to premature ageing phenotypes. Analysis of DNA repair in an animal like hydra could offer clues into several aspects including hydra’s purported lack of organismal ageing, evolution of DNA repair systems in metazoa, and alternative functions of repair proteins. We review here the different DNA repair mechanisms known so far in hydra. Hydra genes from various DNA repair pathways show very high similarity with their vertebrate orthologues, indicating conservation at the level of sequence, structure, and function. Notably, most hydra repair genes are more similar to deuterostome counterparts than to common model invertebrates, hinting at ancient evolutionary origins of repair pathways and further highlighting the relevance of organisms like hydra as model systems. It appears that hydra has the full repertoire of DNA repair pathways, which are employed in stress as well as normal physiological conditions and may have a link with its observed lack of senescence. The close correspondence of hydra repair genes with higher vertebrates further demonstrates the need for deeper studies of various repair components, their interconnections, and functions in this early metazoan.

scholarly journals Differential Effects of DNA Double-Strand Break Repair Pathways on Single-Strand and Self-Complementary Adeno-Associated Virus Vector Genomes

2010 ◽  
Vol 84 (17) ◽  
pp. 8673-8682 ◽  
Author(s):  
Marcela P. Cataldi ◽  
Douglas M. McCarty
Keyword(s):  
Dna Repair ◽  
Strand Break ◽  
Single Strand ◽  
Nonhomologous End Joining ◽  
Chromosomal Integration ◽  
End Joining ◽  
Dsb Repair ◽  
Adeno Associated Virus ◽  
Dna Double Strand Break ◽  
Repair Pathways

ABSTRACT The linear DNA genomes of recombinant adeno-associated virus (rAAV) gene delivery vectors are acted upon by multiple DNA repair and recombination pathways upon release into the host nucleus, resulting in circularization, concatemer formation, or chromosomal integration. We have compared the fates of single-strand rAAV (ssAAV) and self-complementary AAV (scAAV) genomes in cell lines deficient in each of three signaling factors, ATM, ATR, and DNA-PKCS, orchestrating major DNA double-strand break (DSB) repair pathways. In cells deficient in ATM, transduction as scored by green fluorescent protein (GFP) expression is increased relative to that in wild-type (wt) cells by 2.6-fold for ssAAV and 6.6-fold for scAAV vectors, arguing against a mechanism related to second-strand synthesis. The augmented transduction is not reflected in Southern blots of nuclear vector DNA, suggesting that interactions with ATM lead to silencing in normal cells. The additional functional genomes in ATM−/− cells remain linear, and the number of circularized genomes is not affected by the mutation, consistent with compartmentalization of genomes into different DNA repair pathways. A similar effect is observed in ATR-deficient cells but is specific for ssAAV vector. Conversely, a large decrease in transduction is observed in cells deficient in DNA-PKCS, which is involved in DSB repair by nonhomologous end joining rather than homologous recombination. The mutations also have differential effects on chromosomal integration of ssAAV versus scAAV vector genomes. Integration of ssAAV was specifically reduced in ATM−/− cells, while scAAV integration was more profoundly inhibited in DNA-PKCS −/− cells. Taken together, the results suggest that productive rAAV genome circularization is mediated primarily by nonhomologous end joining.

scholarly journals Archaeal DNA Repair Mechanisms

Biomolecules ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. 1472
Author(s):  
Craig J. Marshall ◽  
Thomas J. Santangelo
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Molecular Mechanisms ◽  
Genomic Integrity ◽  
Dna Stability ◽  
Dna Repair Mechanisms ◽  
Environmental Extremes ◽  
Repair Mechanisms ◽  
Repair Pathways ◽  
Damage Types

Archaea often thrive in environmental extremes, enduring levels of heat, pressure, salinity, pH, and radiation that prove intolerable to most life. Many environmental extremes raise the propensity for DNA damaging events and thus, impact DNA stability, placing greater reliance on molecular mechanisms that recognize DNA damage and initiate accurate repair. Archaea can presumably prosper in harsh and DNA-damaging environments in part due to robust DNA repair pathways but surprisingly, no DNA repair pathways unique to Archaea have been described. Here, we review the most recent advances in our understanding of archaeal DNA repair. We summarize DNA damage types and their consequences, their recognition by host enzymes, and how the collective activities of many DNA repair pathways maintain archaeal genomic integrity.

scholarly journals DNA Repair Endonucleases

2015 ◽  
Vol 20 (7) ◽  
pp. 829-841 ◽  
Author(s):  
Rachel Doherty ◽  
Srinivasan Madhusudan
Keyword(s):  
Dna Repair ◽  
Genomic Dna ◽  
Genomic Stability ◽  
Coping Mechanisms ◽  
New Strategy ◽  
Dna Strands ◽  
Repair Pathways ◽  
Dna Ends ◽  
Successful Repair ◽  
Damaged Dna

Genomic DNA is constantly exposed to endogenous and exogenous damaging agents. To overcome these damaging effects and maintain genomic stability, cells have robust coping mechanisms in place, including repair of the damaged DNA. There are a number of DNA repair pathways available to cells dependent on the type of damage induced. The removal of damaged DNA is essential to allow successful repair. Removal of DNA strands is achieved by nucleases. Exonucleases are those that progressively cut from DNA ends, and endonucleases make single incisions within strands of DNA. This review focuses on the group of endonucleases involved in DNA repair pathways, their mechanistic functions, roles in cancer development, and how targeting these enzymes is proving to be an exciting new strategy for personalized therapy in cancer.

scholarly journals Cancer Stem Cells and Radioresistance: Rho/ROCK Pathway Plea Attention

2016 ◽  
Vol 2016 ◽  
pp. 1-7 ◽  
Author(s):  
Annapurna Pranatharthi ◽  
Cecil Ross ◽  
Sweta Srivastava
Keyword(s):  
Cervical Cancer ◽  
Stem Cells ◽  
Dna Repair ◽  
Molecular Mechanisms ◽  
Tumor Resistance ◽  
Tumor Relapse ◽  
Important Target ◽  
Cervical Cancer Cells ◽  
Repair Pathways

Radiation is the most potent mode of cancer therapy; however, resistance to radiation therapy results in tumor relapse and subsequent fatality. The cancer stem cell (CSC), which has better DNA repair capability, has been shown to contribute to tumor resistance and is an important target for treatment. Signaling molecules such as Notch, Wnt, and DNA repair pathways regulate molecular mechanisms in CSCs; however, none of them have been translated into therapeutic targets. The RhoGTPases and their effector ROCK-signaling pathway, though important for tumor progression, have not been well studied in the context of radioresistance. There are reports that implicate RhoA in radioresistance. ROCK2 has also been shown to interact with BRCA2 in the regulation of cell division. Incidentally, statins (drug for cardiovascular ailment) are functional inhibitors of RhoGTPases. Studies suggest that patients on statins have a better prognosis in cancers. Data from our lab suggest that ROCK signaling regulates radioresistance in cervical cancer cells. Collectively, these findings suggest that Rho/ROCK signaling may be important for radiation resistance. In this review, we enumerate the role of Rho/ROCK signaling in stemness and radioresistance and highlight the need to explore these molecules for a better understanding of radioresistance and development of therapeutics.

scholarly journals Human Papillomaviruses Preferentially Recruit DNA Repair Factors to Viral Genomes for Rapid Repair and Amplification

mBio ◽  
2018 ◽  
Vol 9 (1) ◽  
Author(s):  
Kavi Mehta ◽  
Laimonis Laimins
Keyword(s):  
Dna Repair ◽  
High Risk ◽  
Ataxia Telangiectasia ◽  
Human Papillomaviruses ◽  
Dna Breaks ◽  
Ataxia Telangiectasia Mutated ◽  
Genome Amplification ◽  
Viral Genomes ◽  
Repair Pathways ◽  
E6 And E7

ABSTRACT High-risk human papillomaviruses (HPVs) activate the ataxia telangiectasia mutated-dependent (ATM) DNA damage response as well as the ataxia telangiectasia mutated-dependent DNA-related (ATR) pathway in the absence of external DNA damaging agents for differentiation-dependent genome amplification. Through the use of comet assays and pulsed-field gel electrophoresis, our studies showed that these pathways are activated in response to DNA breaks induced by the viral proteins E6 and E7 alone and independently of viral replication. The majority of these virally induced DNA breaks are present in cellular DNAs and only minimally in HPV episomes. Treatment of HPV-positive cells with inhibitors of both ATM and ATR leads to the generation of DNA breaks and the fragmentation of viral episomes, indicating that DNA breaks are introduced into HPV genomes. These breaks, however, are rapidly repaired through the preferential recruitment of homologous recombination repair enzymes, such as RAD51 and BRCA1, to viral genomes at the expense of cellular DNAs. When HPV-positive cells are treated with hydroxyurea, this recruitment of RAD51 and BRCA1 to viral genomes is greatly enhanced with little recruitment to damaged cellular DNAs and with retention of the ability of viral genomes to amplify. Overall, our studies demonstrated that human papillomaviruses induce breaks into cellular and viral DNAs and that the preferential repair of these lesions in viral episomes leads to genome amplification. IMPORTANCE High-risk human papillomaviruses (HPVs) are the etiologic agents of cervical cancer and are linked to the development of many other anogenital and oropharyngeal cancers. Replication of high-risk HPVs requires the activation of the ataxia telangiectasia-mutated (ATM) and ATM- and Rad3-related (ATR) DNA repair pathways. Our studies have shown that HPVs activate these pathways by inducing double-strand breaks primarily in cellular DNAs and minimally in viral genomes. Breaks are induced in viral genomes but are rapidly repaired through the preferential recruitment of homologous repair factors such as RAD51 and BRCA1 to HPV episomes. The preferential repair of breaks in viral genomes leads to amplification. Our study identified a novel mechanism by which human papillomaviruses manipulate DNA repair pathways to productively replicate viral genomes. The induction of genetic instability in cellular DNAs likely contributes to the generation of mutations that lead to the development of cancers.

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