scholarly journals Aberrant PARP1 Activity Couples DNA Breaks to Deregulated Presynaptic Calcium Signalling and Lethal Seizures

2018 ◽  
Author(s):  
Emilia Komulainen ◽  
Jack Badman ◽  
Stephanie Rey ◽  
Stuart Rulten ◽  
Limei Ju ◽  
...  
Keyword(s):  
Hippocampal Neurons ◽  
Ex Vivo ◽  
Calcium Signalling ◽  
Strand Break ◽  
Parp Inhibition ◽  
Dna Breaks ◽  
Single Strand Break ◽  
Presynaptic Calcium ◽  
Dna Strand

AbstractDefects in DNA single-strand break repair result in cerebellar ataxia which in Xrcc1Nes-Cre mice is promoted by hyperactivity of the DNA strand break sensor protein, Parp1. Here, we show that Parp1 hyperactivity extends beyond the cerebellum in Xrcc1-defective brain, resulting in lethal seizures and shortened lifespan. We demonstrate that aberrant Parp1 activation triggers seizure-like activity in Xrcc1-defective hippocampus ex vivo and aberrant presynaptic calcium signalling in isolated hippocampal neurons in vitro. Moreover, we show that these defects are prevented by Parp1 inhibition and/or deletion. Collectively, these data identify aberrant Parp1 activity at unrepaired DNA breaks as a cell-autonomous source of deregulated presynaptic calcium signalling, and highlight PARP inhibition as a possible therapeutic approach in XRCC1-mutated neurodegenerative disease.SummaryPARP1 activity and presynaptic Ca2+ signalling

scholarly journals Novel PNKP mutations causing defective DNA strand break repair and PARP1 hyperactivity in MCSZ

2019 ◽  
Vol 5 (2) ◽  
pp. e320 ◽  
Author(s):  
Ilona Kalasova ◽  
Hana Hanzlikova ◽  
Neerja Gupta ◽  
Yun Li ◽  
Janine Altmüller ◽  
...  
Keyword(s):  
Topoisomerase I ◽  
Strand Break ◽  
Neurologic Disease ◽  
Dna Breaks ◽  
Novel Mutations ◽  
Molecular Feature ◽  
Single Strand Break ◽  
Dna Strand Break ◽  
Break Repair ◽  
Dna Strand

ObjectiveTo address the relationship between novel mutations in polynucleotide 5'-kinase 3'-phosphatase (PNKP), DNA strand break repair, and neurologic disease.MethodsWe have employed whole-exome sequencing, Sanger sequencing, and molecular/cellular biology.ResultsWe describe here a patient with microcephaly with early onset seizures (MCSZ) from the Indian sub-continent harboring 2 novel mutations in PNKP, including a pathogenic mutation in the fork-head associated domain. In addition, we confirm that MCSZ is associated with hyperactivation of the single-strand break sensor protein protein poly (ADP-ribose) polymerase 1 (PARP1) following the induction of abortive topoisomerase I activity, a source of DNA strand breakage associated previously with neurologic disease.ConclusionsThese data expand the spectrum of PNKP mutations associated with MCSZ and show that PARP1 hyperactivation at unrepaired topoisomerase-induced DNA breaks is a molecular feature of this disease.

scholarly journals In VitroExpansion of Bone Marrow Derived Mesenchymal Stem Cells Alters DNA Double Strand Break Repair of Etoposide Induced DNA Damage

2016 ◽  
Vol 2016 ◽  
pp. 1-11 ◽  
Author(s):  
Ian Hare ◽  
Marieta Gencheva ◽  
Rebecca Evans ◽  
James Fortney ◽  
Debbie Piktel ◽  
...  
Keyword(s):  
Stem Cells ◽  
Mesenchymal Stem Cells ◽  
Ex Vivo ◽  
Strand Break ◽  
Dna Lesions ◽  
Cellular Damage ◽  
Human Mscs ◽  
Dna Breaks ◽  
Double Strand Dna Breaks

Mesenchymal stem cells (MSCs) are of interest for use in diverse cellular therapies.Ex vivoexpansion of MSCs intended for transplantation must result in generation of cells that maintain fidelity of critical functions. Previous investigations have identified genetic and phenotypic alterations of MSCs within vitropassage, but little is known regarding how culturing influences the ability of MSCs to repair double strand DNA breaks (DSBs), the most severe of DNA lesions. To investigate the response to DSB stress with passagein vitro, primary human MSCs were exposed to etoposide (VP16) at various passages with subsequent evaluation of cellular damage responses and DNA repair. Passage number did not affect susceptibility to VP16 or the incidence and repair kinetics of DSBs. Nonhomologous end joining (NHEJ) transcripts showed little alteration with VP16 exposure or passage; however, homologous recombination (HR) transcripts were reduced following VP16 exposure with this decrease amplified as MSCs were passagedin vitro. Functional evaluations of NHEJ and HR showed that MSCs were unable to activate NHEJ repair following VP16 stress in cells after successive passage. These results indicate thatex vivoexpansion of MSCs alters their ability to perform DSB repair, a necessary function for cells intended for transplantation.

scholarly journals Recombinogenic Flap Ligation Pathway for Intrinsic Repair of Topoisomerase IB-Induced Double-Strand Breaks

2000 ◽  
Vol 20 (21) ◽  
pp. 8059-8068 ◽  
Author(s):  
Chonghui Cheng ◽  
Stewart Shuman
Keyword(s):  
High Efficiency ◽  
Strand Break ◽  
Strand Transfer ◽  
Strand Breaks ◽  
Genomic Deletions ◽  
Topoisomerase Ib ◽  
Dna Strand ◽  
Recombination Pathway

ABSTRACT Topoisomerase IB catalyzes recombinogenic DNA strand transfer reactions in vitro and in vivo. Here we characterize a new pathway of topoisomerase-mediated DNA ligation in vitro (flap ligation) in which vaccinia virus topoisomerase bound to a blunt-end DNA joins the covalently held strand to a 5′ resected end of a duplex DNA containing a 3′ tail. The joining reaction occurs with high efficiency when the sequence of the 3′ tail is complementary to that of the scissile strand immediately 5′ of the cleavage site. A 6-nucleotide segment of complementarity suffices for efficient flap ligation. Invasion of the flap into the duplex apparently occurs while topoisomerase remains bound to DNA, thereby implying a conformational flexibility of the topoisomerase clamp around the DNA target site. The 3′ flap acceptor DNA mimics a processed end in the double-strand-break-repair recombination pathway. Our findings suggest that topoisomerase-induced breaks may be rectified by flap ligation, with ensuing genomic deletions or translocations.

scholarly journals The Functional Consequences of Eukaryotic Topoisomerase 1 Interaction with G-Quadruplex DNA

Genes ◽  
2020 ◽  
Vol 11 (2) ◽  
pp. 193
Author(s):  
Alexandra Berroyer ◽  
Nayun Kim
Keyword(s):  
Topoisomerase I ◽  
Genome Instability ◽  
Strand Break ◽  
Dna Topology ◽  
Single Strand Break ◽  
Dna Structures ◽  
G4 Dna ◽  
G Quadruplex

Topoisomerase I in eukaryotic cells is an important regulator of DNA topology. Its catalytic function is to remove positive or negative superhelical tension by binding to duplex DNA, creating a reversible single-strand break, and finally religating the broken strand. Proper maintenance of DNA topological homeostasis, in turn, is critically important in the regulation of replication, transcription, DNA repair, and other processes of DNA metabolism. One of the cellular processes regulated by the DNA topology and thus by Topoisomerase I is the formation of non-canonical DNA structures. Non-canonical or non-B DNA structures, including the four-stranded G-quadruplex or G4 DNA, are potentially pathological in that they interfere with replication or transcription, forming hotspots of genome instability. In this review, we first describe the role of Topoisomerase I in reducing the formation of non-canonical nucleic acid structures in the genome. We further discuss the interesting recent discovery that Top1 and Top1 mutants bind to G4 DNA structures in vivo and in vitro and speculate on the possible consequences of these interactions.

scholarly journals Mechanistic insight into the role of Poly(ADP-ribosyl)ation in DNA topology modulation and response to DNA damage

Mutagenesis ◽  
2019 ◽  
Vol 35 (1) ◽  
pp. 107-118
Author(s):  
Bakhyt T Matkarimov ◽  
Dmitry O Zharkov ◽  
Murat K Saparbaev
Keyword(s):  
Dna Damage ◽  
Dna Repair ◽  
Genotoxic Stress ◽  
Strand Break ◽  
Dna Supercoiling ◽  
Dna Strand Breaks ◽  
Chromosomal Dna ◽  
Dna Breaks ◽  
Strand Breaks ◽  
Dna Strand

Abstract Genotoxic stress generates single- and double-strand DNA breaks either through direct damage by reactive oxygen species or as intermediates of DNA repair. Failure to detect and repair DNA strand breaks leads to deleterious consequences such as chromosomal aberrations, genomic instability and cell death. DNA strand breaks disrupt the superhelical state of cellular DNA, which further disturbs the chromatin architecture and gene activity regulation. Proteins from the poly(ADP-ribose) polymerase (PARP) family, such as PARP1 and PARP2, use NAD+ as a substrate to catalyse the synthesis of polymeric chains consisting of ADP-ribose units covalently attached to an acceptor molecule. PARP1 and PARP2 are regarded as DNA damage sensors that, upon activation by strand breaks, poly(ADP-ribosyl)ate themselves and nuclear acceptor proteins. Noteworthy, the regularly branched structure of poly(ADP-ribose) polymer suggests that the mechanism of its synthesis may involve circular movement of PARP1 around the DNA helix, with a branching point in PAR corresponding to one complete 360° turn. We propose that PARP1 stays bound to a DNA strand break end, but rotates around the helix displaced by the growing poly(ADP-ribose) chain, and that this rotation could introduce positive supercoils into damaged chromosomal DNA. This topology modulation would enable nucleosome displacement and chromatin decondensation around the lesion site, facilitating the access of DNA repair proteins or transcription factors. PARP1-mediated DNA supercoiling can be transmitted over long distances, resulting in changes in the high-order chromatin structures. The available structures of PARP1 are consistent with the strand break-induced PAR synthesis as a driving force for PARP1 rotation around the DNA axis.

scholarly journals Homologous Recombination, but Not DNA Repair, Is Reduced in Vertebrate Cells Deficient in RAD52

1998 ◽  
Vol 18 (11) ◽  
pp. 6430-6435 ◽  
Author(s):  
Yuko Yamaguchi-Iwai ◽  
Eiichiro Sonoda ◽  
Jean-Marie Buerstedde ◽  
Olga Bezzubova ◽  
Ciaran Morrison ◽  
...  
Keyword(s):  
Genetic Recombination ◽  
Strand Break ◽  
Intermediate Step ◽  
Immunoglobulin Gene ◽  
Γ Irradiation ◽  
Focus Formation ◽  
Dna Damages ◽  
Dna Strand Exchange ◽  
Dna Strand

ABSTRACT Rad52 plays a pivotal role in double-strand break (DSB) repair and genetic recombination in Saccharomyces cerevisiae, where mutation of this gene leads to extreme X-ray sensitivity and defective recombination. Yeast Rad51 and Rad52 interact, as do their human homologues, which stimulates Rad51-mediated DNA strand exchange in vitro, suggesting that Rad51 and Rad52 act cooperatively. To define the role of Rad52 in vertebrates, we generatedRAD52 −/− mutants of the chicken B-cell line DT40. Surprisingly, RAD52 −/− cells were not hypersensitive to DNA damages induced by γ-irradiation, methyl methanesulfonate, or cis-platinum(II)diammine dichloride (cisplatin). Intrachromosomal recombination, measured by immunoglobulin gene conversion, and radiation-induced Rad51 nuclear focus formation, which is a putative intermediate step during recombinational repair, occurred as frequently inRAD52 −/− cells as in wild-type cells. Targeted integration frequencies, however, were consistently reduced inRAD52 −/− cells, showing a clear role for Rad52 in genetic recombination. These findings reveal striking differences between S. cerevisiae and vertebrates in the functions of RAD51 and RAD52.

scholarly journals PARP inhibition versus PARP-1 silencing: different outcomes in terms of single-strand break repair and radiation susceptibility

2008 ◽  
Vol 36 (13) ◽  
pp. 4454-4464 ◽  
Author(s):  
C. Godon ◽  
F. P. Cordelieres ◽  
D. Biard ◽  
N. Giocanti ◽  
F. Megnin-Chanet ◽  
...  
Keyword(s):  
Strand Break ◽  
Single Strand ◽  
Parp Inhibition ◽  
Single Strand Break ◽  
Single Strand Break Repair ◽  
Break Repair ◽  
Parp 1

scholarly journals Antileukemic Efficacy in Vitro of Talazoparib and APE1 Inhibitor III Combined with Decitabine in Myeloid Malignancies

Cancers ◽  
2019 ◽  
Vol 11 (10) ◽  
pp. 1493 ◽  
Author(s):  
Kohl ◽  
Flach ◽  
Naumann ◽  
Brendel ◽  
Kleiner ◽  
...  
Keyword(s):  
Base Excision Repair ◽  
Excision Repair ◽  
Donor Cell ◽  
Strand Break ◽  
Low Doses ◽  
Single Strand Break ◽  
Single Strand Break Repair ◽  
Myeloid Leukemias ◽  
Base Excision

Malignant hematopoietic cells of myelodysplastic syndromes (MDS)/chronic myelomonocytic leukemias (CMML) and acute myeloid leukemias (AML) may be vulnerable to inhibition of poly(ADP ribose) polymerase 1/2 (PARP1/2) and apurinic/apyrimidinic endonuclease 1 (APE1). PARP1/2 and APE1 are critical enzymes involved in single-strand break repair and base excision repair, respectively. Here, we investigated the cytotoxic efficacy of talazoparib and APE1 inhibitor III, inhibitors of PARP1/2 and APE1, in primary CD34+ MDS/CMML cell samples (n = 8; 4 MDS and 4 CMML) and in primary CD34+ or CD34− AML cell samples (n = 18) in comparison to healthy CD34+ donor cell samples (n = 8). Strikingly, talazoparib and APE1 inhibitor III demonstrated critical antileukemic efficacy in selected MDS/CMML and AML cell samples. Low doses of talazoparib and APE1 inhibitor III further increased the cytotoxic efficacy of decitabine in MDS/CMML and AML cells. Moreover, low doses of APE1 inhibitor III increased the cytotoxic efficacy of talazoparib in MDS/CMML and AML cells. In summary, talazoparib and APE1 inhibitor III demonstrated substantial antileukemic efficacy as single agents, in combination with decitabine, and combined with each other. Hence, our findings support further investigation of these agents in sophisticated clinical trials.

scholarly journals Induction of CAF-1 Expression in Response to DNA Strand Breaks in Quiescent Human Cells

2006 ◽  
Vol 26 (5) ◽  
pp. 1839-1849 ◽  
Author(s):  
Arman Nabatiyan ◽  
Dávid Szüts ◽  
Torsten Krude
Keyword(s):  
Excision Repair ◽  
Dna Strand Breaks ◽  
Significant Loss ◽  
Human Cells ◽  
Dna Breaks ◽  
Strand Breaks ◽  
Quiescent Cells ◽  
Dna Strand

ABSTRACT Genome stability in eukaryotic cells is maintained through efficient DNA damage repair pathways, which have to access and utilize chromatin as their natural template. Here we investigate the role of chromatin assembly factor 1 (CAF-1) and its interacting protein, PCNA, in the response of quiescent human cells to DNA double-strand breaks (DSBs). The expression of CAF-1 and PCNA is dramatically induced in quiescent cells upon the generation of DSBs by the radiomimetic drug bleocin (a bleomycin compound) or by ionizing radiation. This induction depends on DNA-PK. CAF-1 and PCNA are recruited to damaged chromatin undergoing DNA repair of single- and double-strand DNA breaks by the base excision repair and nonhomologous end-joining pathways, respectively, in the absence of extensive DNA synthesis. CAF-1 prepared from repair-proficient quiescent cells after induction by bleocin mediates nucleosome assembly in vitro. Depletion of CAF-1 by RNA interference in bleocin-treated quiescent cells in vivo results in a significant loss of cell viability and an accumulation of DSBs. These results support a novel and essential role for CAF-1 in the response of quiescent human cells to DSBs, possibly by reassembling chromatin following repair of DNA strand breaks.

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