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Sample records for alternative end-joining repair

  1. Double Strand Break Repair, one mechanism can hide another: Alternative non-homologous end joining

    International Nuclear Information System (INIS)

    Rass, E.; Grabarz, A.; Bertrand, P.; Lopez, B.S.

    2012-01-01

    DNA double strand breaks are major cytotoxic lesions encountered by the cells. They can be induced by ionizing radiation or endogenous stress and can lead to genetic instability. Two mechanisms compete for the repair of DNA double strand breaks: homologous recombination and non-homologous end joining (NHEJ). Homologous recombination requires DNA sequences homology and is initiated by single strand resection. Recently, advances have been made concerning the major steps and proteins involved in resection. NHEJ, in contrast, does not require sequence homology. The existence of a DNA double strand break repair mechanism, independent of KU and ligase IV, the key proteins of the canonical non homologous end joining pathway, has been revealed lately and named alternative non homologous end joining. The hallmarks of this highly mutagenic pathway are deletions at repair junctions and frequent use of distal micro-homologies. This mechanism is also initiated by a single strand resection of the break. The aim of this review is firstly to present recent data on single strand resection, and secondly the alternative NHEJ pathway, including a discussion on the fidelity of NHEJ. Based on current knowledge, canonical NHEJ does not appear as an intrinsically mutagenic mechanism, but in contrast, as a conservative one. The structure of broken DNA ends actually dictates the quality repair of the alternative NHEJ and seems the actual responsible for the mutagenesis attributed beforehand to the canonical NHEJ. The existence of this novel DNA double strand breaks repair mechanism needs to be taken into account in the development of radiosensitizing strategies in order to optimise the efficiency of radiotherapy. (authors)

  2. Alternative end-joining pathway(s): bricolage at DNA breaks.

    Science.gov (United States)

    Frit, Philippe; Barboule, Nadia; Yuan, Ying; Gomez, Dennis; Calsou, Patrick

    2014-05-01

    To cope with DNA double strand break (DSB) genotoxicity, cells have evolved two main repair pathways: homologous recombination which uses homologous DNA sequences as repair templates, and non-homologous Ku-dependent end-joining involving direct sealing of DSB ends by DNA ligase IV (Lig4). During the last two decades a third player most commonly named alternative end-joining (A-EJ) has emerged, which is defined as any Ku- or Lig4-independent end-joining process. A-EJ increasingly appears as a highly error-prone bricolage on DSBs and despite expanding exploration, it still escapes full characterization. In the present review, we discuss the mechanism and regulation of A-EJ as well as its biological relevance under physiological and pathological situations, with a particular emphasis on chromosomal instability and cancer. Whether or not it is a genuine DSB repair pathway, A-EJ is emerging as an important cellular process and understanding A-EJ will certainly be a major challenge for the coming years. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  3. Role for Artemis nuclease in the repair of radiation-induced DNA double strand breaks by alternative end joining.

    Science.gov (United States)

    Moscariello, Mario; Wieloch, Radi; Kurosawa, Aya; Li, Fanghua; Adachi, Noritaka; Mladenov, Emil; Iliakis, George

    2015-07-01

    Exposure of cells to ionizing radiation or radiomimetic drugs generates DNA double-strand breaks that are processed either by homologous recombination repair (HRR), or by canonical, DNA-PKcs-dependent non-homologous end-joining (C-NHEJ). Chemical or genetic inactivation of factors involved in C-NHEJ or HRR, but also their local failure in repair proficient cells, promotes an alternative, error-prone end-joining pathway that serves as backup (A-EJ). There is evidence for the involvement of Artemis endonuclease, a protein deficient in a human radiosensitivity syndrome associated with severe immunodeficiency (RS-SCID), in the processing of subsets of DSBs by HRR or C-NHEJ. It is thought that within HRR or C-NHEJ Artemis processes DNA termini at complex DSBs. Whether Artemis has a role in A-EJ remains unknown. Here, we analyze using pulsed-field gel electrophoresis (PFGE) and specialized reporter assays, DSB repair in wild-type pre-B NALM-6 lymphocytes, as well as in their Artemis(-/-), DNA ligase 4(-/-) (LIG4(-/-)), and LIG4(-/-)/Artemis(-/-) double mutant counterparts, under conditions allowing evaluation of A-EJ. Our results substantiate the suggested roles of Artemis in C-NHEJ and HRR, but also demonstrate a role for the protein in A-EJ that is confirmed in Artemis deficient normal human fibroblasts. We conclude that Artemis is a nuclease participating in DSB repair by all major repair pathways. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. FANCD2 Maintains Fork Stability in BRCA1/2-Deficient Tumors and Promotes Alternative End-Joining DNA Repair

    Directory of Open Access Journals (Sweden)

    Zeina Kais

    2016-06-01

    Full Text Available BRCA1/2 proteins function in homologous recombination (HR-mediated DNA repair and cooperate with Fanconi anemia (FA proteins to maintain genomic integrity through replication fork stabilization. Loss of BRCA1/2 proteins results in DNA repair deficiency and replicative stress, leading to genomic instability and enhanced sensitivity to DNA-damaging agents. Recent studies have shown that BRCA1/2-deficient tumors upregulate Polθ-mediated alternative end-joining (alt-EJ repair as a survival mechanism. Whether other mechanisms maintain genomic integrity upon loss of BRCA1/2 proteins is currently unknown. Here we show that BRCA1/2-deficient tumors also upregulate FANCD2 activity. FANCD2 is required for fork protection and fork restart in BRCA1/2-deficient tumors. Moreover, FANCD2 promotes Polθ recruitment at sites of damage and alt-EJ repair. Finally, loss of FANCD2 in BRCA1/2-deficient tumors enhances cell death. These results reveal a synthetic lethal relationship between FANCD2 and BRCA1/2, and they identify FANCD2 as a central player orchestrating DNA repair pathway choice at the replication fork.

  5. Distinct DNA repair pathways involving RecA and nonhomologous end joining in Mycobacterium smegmatis.

    OpenAIRE

    Korycka-Machala, M; Brzostek, A; Rozalska, S; Rumijowska-Galewicz, A; Dziedzic, R; Bowater, R; Dziadek, J

    2006-01-01

    Mycobacterium smegmatis was used to study the relationship between DNA repair processes involving RecA and nonhomologous end joining (NHEJ). The effect of gene deletions in recA and/or in two genes involved in NHEJ (ku and ligD) was tested on the ability of bacteria to join breaks in plasmids transformed into them and in their response to chemicals that damage DNA. The results provide in vivo evidence that only NHEJ is required for the repair of noncompatible DNA ends. By contrast, the respon...

  6. A Role for BLM in Double-Strand Break Repair Pathway Choice: Prevention of CtIP/Mre11-Mediated Alternative Nonhomologous End-Joining

    DEFF Research Database (Denmark)

    Grabarz, Anastazja; Guirouilh-Barbat, Josée; Barascu, Aurelia

    2013-01-01

    The choice of the appropriate double-strand break (DSB) repair pathway is essential for the maintenance of genomic stability. Here, we show that the Bloom syndrome gene product, BLM, counteracts CtIP/MRE11-dependent long-range deletions (>200 bp) generated by alternative end-joining (A-EJ). BLM...... represses A-EJ in an epistatic manner with 53BP1 and RIF1 and is required for ionizing-radiation-induced 53BP1 focus assembly. Conversely, in the absence of 53BP1 or RIF1, BLM promotes formation of A-EJ long deletions, consistent with a role for BLM in DSB end resection. These data highlight a dual role...... for BLM that influences the DSB repair pathway choice: (1) protection against CtIP/MRE11 long-range deletions associated with A-EJ and (2) promotion of DNA resection. These antagonist roles can be regulated, according to cell-cycle stage, by interacting partners such as 53BP1 and TopIII, to avoid...

  7. Distinct DNA repair pathways involving RecA and nonhomologous end joining in Mycobacterium smegmatis.

    Science.gov (United States)

    Korycka-Machala, Malgorzata; Brzostek, Anna; Rozalska, Sylwia; Rumijowska-Galewicz, Anna; Dziedzic, Renata; Bowater, Richard; Dziadek, Jaroslaw

    2006-05-01

    Mycobacterium smegmatis was used to study the relationship between DNA repair processes involving RecA and nonhomologous end joining (NHEJ). The effect of gene deletions in recA and/or in two genes involved in NHEJ (ku and ligD) was tested on the ability of bacteria to join breaks in plasmids transformed into them and in their response to chemicals that damage DNA. The results provide in vivo evidence that only NHEJ is required for the repair of noncompatible DNA ends. By contrast, the response of mycobacteria to mitomycin C preferentially involved a RecA-dependent pathway.

  8. XRCC1 suppresses somatic hypermutation and promotes alternative nonhomologous end joining in Igh genes.

    Science.gov (United States)

    Saribasak, Huseyin; Maul, Robert W; Cao, Zheng; McClure, Rhonda L; Yang, William; McNeill, Daniel R; Wilson, David M; Gearhart, Patricia J

    2011-10-24

    Activation-induced deaminase (AID) deaminates cytosine to uracil in immunoglobulin genes. Uracils in DNA can be recognized by uracil DNA glycosylase and abasic endonuclease to produce single-strand breaks. The breaks are repaired either faithfully by DNA base excision repair (BER) or mutagenically to produce somatic hypermutation (SHM) and class switch recombination (CSR). To unravel the interplay between repair and mutagenesis, we decreased the level of x-ray cross-complementing 1 (XRCC1), a scaffold protein involved in BER. Mice heterozygous for XRCC1 showed a significant increase in the frequencies of SHM in Igh variable regions in Peyer's patch cells, and of double-strand breaks in the switch regions during CSR. Although the frequency of CSR was normal in Xrcc1(+/-) splenic B cells, the length of microhomology at the switch junctions decreased, suggesting that XRCC1 also participates in alternative nonhomologous end joining. Furthermore, Xrcc1(+/-) B cells had reduced Igh/c-myc translocations during CSR, supporting a role for XRCC1 in microhomology-mediated joining. Our results imply that AID-induced single-strand breaks in Igh variable and switch regions become substrates simultaneously for BER and mutagenesis pathways.

  9. Detection and Repair of Ionizing Radiation-Induced DNA Double Strand Breaks: New Developments in Nonhomologous End Joining

    International Nuclear Information System (INIS)

    Wang, Chen; Lees-Miller, Susan P.

    2013-01-01

    DNA damage can occur as a result of endogenous metabolic reactions and replication stress or from exogenous sources such as radiation therapy and chemotherapy. DNA double strand breaks are the most cytotoxic form of DNA damage, and defects in their repair can result in genome instability, a hallmark of cancer. The major pathway for the repair of ionizing radiation-induced DSBs in human cells is nonhomologous end joining. Here we review recent advances on the mechanism of nonhomologous end joining, as well as new findings on its component proteins and regulation

  10. Detection and Repair of Ionizing Radiation-Induced DNA Double Strand Breaks: New Developments in Nonhomologous End Joining

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Chen [Departments of Biochemistry and Molecular Biology and Oncology, and Southern Alberta Cancer Research Institute, University of Calgary, Calgary (Canada); Lees-Miller, Susan P., E-mail: leesmill@ucalgary.ca [Departments of Biochemistry and Molecular Biology and Oncology, and Southern Alberta Cancer Research Institute, University of Calgary, Calgary (Canada)

    2013-07-01

    DNA damage can occur as a result of endogenous metabolic reactions and replication stress or from exogenous sources such as radiation therapy and chemotherapy. DNA double strand breaks are the most cytotoxic form of DNA damage, and defects in their repair can result in genome instability, a hallmark of cancer. The major pathway for the repair of ionizing radiation-induced DSBs in human cells is nonhomologous end joining. Here we review recent advances on the mechanism of nonhomologous end joining, as well as new findings on its component proteins and regulation.

  11. Induction and repair of DNA double strand breaks: The increasing spectrum of non-homologous end joining pathways

    International Nuclear Information System (INIS)

    Mladenov, Emil; Iliakis, George

    2011-01-01

    A defining characteristic of damage induced in the DNA by ionizing radiation (IR) is its clustered character that leads to the formation of complex lesions challenging the cellular repair mechanisms. The most widely investigated such complex lesion is the DNA double strand break (DSB). DSBs undermine chromatin stability and challenge the repair machinery because an intact template strand is lacking to assist restoration of integrity and sequence in the DNA molecule. Therefore, cells have evolved a sophisticated machinery to detect DSBs and coordinate a response on the basis of inputs from various sources. A central function of cellular responses to DSBs is the coordination of DSB repair. Two conceptually different mechanisms can in principle remove DSBs from the genome of cells of higher eukaryotes. Homologous recombination repair (HRR) uses as template a homologous DNA molecule and is therefore error-free; it functions preferentially in the S and G2 phases. Non-homologous end joining (NHEJ), on the other hand, simply restores DNA integrity by joining the two ends, is error prone as sequence is only fortuitously preserved and active throughout the cell cycle. The basis of DSB repair pathway choice remains unknown, but cells of higher eukaryotes appear programmed to utilize preferentially NHEJ. Recent work suggests that when the canonical DNA-PK dependent pathway of NHEJ (D-NHEJ), becomes compromised an alternative NHEJ pathway and not HRR substitutes in a quasi-backup function (B-NHEJ). Here, we outline aspects of DSB induction by IR and review the mechanisms of their processing in cells of higher eukaryotes. We place particular emphasis on backup pathways of NHEJ and summarize their increasing significance in various cellular processes, as well as their potential contribution to carcinogenesis.

  12. Unsolved mystery: the role of BRCA1 in DNA end-joining

    International Nuclear Information System (INIS)

    Saha, Janapriya; Davis, Anthony J.

    2016-01-01

    Heritable mutations in the tumor suppressor gene BRCA1 increase a woman's lifetime risk of developing breast and ovarian cancer. BRCA1's tumor suppressor function is directly linked to its myriad of functions in the cellular response to DNA double-strand breaks (DSBs). BRCA1 interacts with an extensive array of DNA damage responsive proteins and plays important roles in DSB repair, mediated by the homologous recombination pathway, and in the activation of cell cycle checkpoints. However, the role of BRCA1 in the other two DSB repair pathways, classical non-homologous end-joining (C-NHEJ) and alternative NHEJ (A-NHEJ), remains unclear. In this review, we will discuss the current literature on BRCA1's potential role(s) in modulating both C-NHEJ and A-NHEJ. We also present a model showing that BRCA1 contributes to genomic maintenance by promoting precise DNA repair across all cell cycle phases via the direct modulation of DNA end-joining

  13. Processing of DNA double strand breaks by alternative non-homologous end-joining in hyperacetylated chromatin.

    Science.gov (United States)

    Manova, Vasilissa; Singh, Satyendra K; Iliakis, George

    2012-08-22

    Mammalian cells employ at least two subpathways of non-homologous end-joining for the repair of ionizing radiation induced DNA double strand breaks: The canonical DNA-PK-dependent form of non-homologous end-joining (D-NHEJ) and an alternative, slowly operating, error-prone backup pathway (B-NHEJ). In contrast to D-NHEJ, which operates with similar efficiency throughout the cell cycle, B-NHEJ operates more efficiently in G2-phase. Notably, B-NHEJ also shows strong and as of yet unexplained dependency on growth activity and is markedly compromised in serum-deprived cells, or in cells that enter the plateau-phase of growth. The molecular mechanisms underpinning this response remain unknown. Since chromatin structure or changes in chromatin structure are prime candidate-B-NHEJ-modulators, we study here the role of chromatin hyperacetylation, either by HDAC2 knockdown or treatment with the HDAC inhibitor TSA, on the repair by B-NHEJ of IR-induced DSBs. siRNA-mediated knockdown of HDAC2 fails to provoke histone hyperacetylation in Lig4-/- MEFs and has no detectable effect on B-NHEJ function. Treatment with TSA that inhibits multiple HDACs causes efficient, reversible chromatin hyperacetylation in Lig4-/- MEFs, as well as in human HCT116 Lig4-/- cells and the human glioma cell line M059K. The IR yield of DSBs in TSA-treated cells remains similar to that of untreated cells despite the expected chromatin relaxation. In addition, chromatin hyperacetylation leaves unchanged repair of DSBs by B-NHEJ in irradiated exponentially growing, or plateau-phase cells. Notably, under the experimental conditions employed here, chromatin hyperacetylation fails to detectably modulate B-NHEJ in M059K cells as well. In summary, the results show that chromatin acetylation or deacetylation does not affect the kinetics of alternative NHEJ in all types of cells examined both in exponentially growing and serum deprived cultures. We conclude that parameters beyond chromatin acetylation determine B

  14. The mechanism of non-homologous end-joining: a synopsis of synapsis.

    Science.gov (United States)

    Weterings, Eric; van Gent, Dik C

    2004-11-02

    Repair of DNA double-strand breaks (DSBs) by non-homologous end-joining (NHEJ) is required for resistance to genotoxic agents, such as ionizing radiation, but also for proper development of the vertebrate immune system. Much progress has been made in identifying the factors that are involved in this repair pathway. We are now entering the phase in which we begin to understand basic concepts of the reaction mechanism and regulation of non-homologous end-joining. This review concentrates on novel insights into damage recognition and subsequent tethering, processing and joining of DNA ends.

  15. Redundant function of DNA ligase 1 and 3 in alternative end-joining during immunoglobulin class switch recombination.

    Science.gov (United States)

    Masani, Shahnaz; Han, Li; Meek, Katheryn; Yu, Kefei

    2016-02-02

    Nonhomologous end-joining (NHEJ) is the major DNA double-strand break (DSB) repair pathway in mammals and resolves the DSBs generated during both V(D)J recombination in developing lymphocytes and class switch recombination (CSR) in antigen-stimulated B cells. In contrast to the absolute requirement for NHEJ to resolve DSBs associated with V(D)J recombination, DSBs associated with CSR can be resolved in NHEJ-deficient cells (albeit at a reduced level) by a poorly defined alternative end-joining (A-EJ) pathway. Deletion of DNA ligase IV (Lig4), a core component of the NHEJ pathway, reduces CSR efficiency in a mouse B-cell line capable of robust cytokine-stimulated CSR in cell culture. Here, we report that CSR levels are not further reduced by deletion of either of the two remaining DNA ligases (Lig1 and nuclear Lig3) in Lig4(-/-) cells. We conclude that in the absence of Lig4, Lig1, and Lig3 function in a redundant manner in resolving switch region DSBs during CSR.

  16. Mycobacterial nonhomologous end joining mediates mutagenic repair of chromosomal double-strand DNA breaks.

    Science.gov (United States)

    Stephanou, Nicolas C; Gao, Feng; Bongiorno, Paola; Ehrt, Sabine; Schnappinger, Dirk; Shuman, Stewart; Glickman, Michael S

    2007-07-01

    Bacterial nonhomologous end joining (NHEJ) is a recently described DNA repair pathway best characterized in mycobacteria. Bacterial NHEJ proteins LigD and Ku have been analyzed biochemically, and their roles in linear plasmid repair in vivo have been verified genetically; yet the contributions of NHEJ to repair of chromosomal DNA damage are unknown. Here we use an extensive set of NHEJ- and homologous recombination (HR)-deficient Mycobacterium smegmatis strains to probe the importance of HR and NHEJ in repairing diverse types of chromosomal DNA damage. An M. smegmatis Delta recA Delta ku double mutant has no apparent growth defect in vitro. Loss of the NHEJ components Ku and LigD had no effect on sensitivity to UV radiation, methyl methanesulfonate, or quinolone antibiotics. NHEJ deficiency had no effect on sensitivity to ionizing radiation in logarithmic- or early-stationary-phase cells but was required for ionizing radiation resistance in late stationary phase in 7H9 but not LB medium. In addition, NHEJ components were required for repair of I-SceI mediated chromosomal double-strand breaks (DSBs), and in the absence of HR, the NHEJ pathway rapidly mutates the chromosomal break site. The molecular outcomes of NHEJ-mediated chromosomal DSB repair involve predominantly single-nucleotide insertions at the break site, similar to previous findings using plasmid substrates. These findings demonstrate that prokaryotic NHEJ is specifically required for DSB repair in late stationary phase and can mediate mutagenic repair of homing endonuclease-generated chromosomal DSBs.

  17. Lipofection and nucleofection of substrate plasmid can generate widely different readings of DNA end-joining efficiency in different cell lines.

    Science.gov (United States)

    Magin, Simon; Saha, Janapriya; Wang, Minli; Mladenova, Veronika; Coym, Nadine; Iliakis, George

    2013-02-01

    In vivo plasmid end-joining assays are valuable tools for dissecting important qualitative and quantitative aspects of non-homologous end-joining (NHEJ)--a key mechanism for the repair of DNA double-strand breaks (DSBs) in higher eukaryotes. They enable the use of defined DNA ends as substrates for end-joining and the analysis by sequencing of the resulting junctions to identify the repair pathways engaged. Yet, plasmid assays have generated divergent results of end-joining capacity in the same DSB repair mutants when used under different conditions, which implies contributions from undefined and therefore uncontrolled parameters. To help standardize these assays, we searched for parameters underpinning these variations and identified transfection method as an important determinant. Here, we compare a lipid-based transfection method, lipofection, with an electroporation method, nucleofection, and find large, unanticipated and cell line-dependent differences in percent end-joining without recognizable trends. For example, in rodent cells, transfection using lipofection gives nearly WT end-joining in DNA-PKcs mutants and only mildly inhibited end-joining in Lig4 and Ku mutants. In contrast, transfection using nucleofection shows marked end-joining inhibition in all NHEJ mutants tested as compared to the WT. In human HCT116 cells, end-joining after nucleofection is strongly suppressed even in the WT and the differences to the mutants are small. After lipofection, in contrast, end-joining is high in WT cells and markedly suppressed in the mutants. We conclude that better understanding and control of the physicochemical/biological and analytical parameters underpinning these differences will be required to generate with plasmid assays results with quantitative power comparable to that of well-established methods of DSB analysis such as pulsed-field gel electrophoresis or γ-H2AX foci scoring. Until then, caution is needed in the interpretation of the results obtained

  18. Hyperactivation of PARP triggers nonhomologous end-joining in repair-deficient mouse fibroblasts.

    Directory of Open Access Journals (Sweden)

    Natalie R Gassman

    Full Text Available Regulation of poly(ADP-ribose (PAR synthesis and turnover is critical to determining cell fate after genotoxic stress. Hyperactivation of PAR synthesis by poly(ADP-ribose polymerase-1 (PARP-1 occurs when cells deficient in DNA repair are exposed to genotoxic agents; however, the function of this hyperactivation has not been adequately explained. Here, we examine PAR synthesis in mouse fibroblasts deficient in the base excision repair enzyme DNA polymerase β (pol β. The extent and duration of PARP-1 activation was measured after exposure to either the DNA alkylating agent, methyl methanesulfonate (MMS, or to low energy laser-induced DNA damage. There was strong DNA damage-induced hyperactivation of PARP-1 in pol β nullcells, but not in wild-type cells. In the case of MMS treatment, PAR synthesis did not lead to cell death in the pol β null cells, but instead resulted in increased PARylation of the nonhomologous end-joining (NHEJ protein Ku70 and increased association of Ku70 with PARP-1. Inhibition of the NHEJ factor DNA-PK, under conditions of MMS-induced PARP-1 hyperactivation, enhanced necrotic cell death. These data suggest that PARP-1 hyperactivation is a protective mechanism triggering the classical-NHEJ DNA repair pathway when the primary alkylated base damage repair pathway is compromised.

  19. Alternative end-joining catalyzes robust IgH locus deletions and translocations in the combined absence of ligase 4 and Ku70.

    Science.gov (United States)

    Boboila, Cristian; Jankovic, Mila; Yan, Catherine T; Wang, Jing H; Wesemann, Duane R; Zhang, Tingting; Fazeli, Alex; Feldman, Lauren; Nussenzweig, Andre; Nussenzweig, Michel; Alt, Frederick W

    2010-02-16

    Class switch recombination (CSR) in B lymphocytes is initiated by introduction of multiple DNA double-strand breaks (DSBs) into switch (S) regions that flank immunoglobulin heavy chain (IgH) constant region exons. CSR is completed by joining a DSB in the donor S mu to a DSB in a downstream acceptor S region (e.g., S gamma1) by end-joining. In normal cells, many CSR junctions are mediated by classical nonhomologous end-joining (C-NHEJ), which employs the Ku70/80 complex for DSB recognition and XRCC4/DNA ligase 4 for ligation. Alternative end-joining (A-EJ) mediates CSR, at reduced levels, in the absence of C-NHEJ, even in combined absence of Ku70 and ligase 4, demonstrating an A-EJ pathway totally distinct from C-NHEJ. Multiple DSBs are introduced into S mu during CSR, with some being rejoined or joined to each other to generate internal switch deletions (ISDs). In addition, S-region DSBs can be joined to other chromosomes to generate translocations, the level of which is increased by absence of a single C-NHEJ component (e.g., XRCC4). We asked whether ISD and S-region translocations occur in the complete absence of C-NHEJ (e.g., in Ku70/ligase 4 double-deficient B cells). We found, unexpectedly, that B-cell activation for CSR generates substantial ISD in both S mu and S gamma1 and that ISD in both is greatly increased by the absence of C-NHEJ. IgH chromosomal translocations to the c-myc oncogene also are augmented in the combined absence of Ku70 and ligase 4. We discuss the implications of these findings for A-EJ in normal and abnormal DSB repair.

  20. Modeling Non-homologous End Joining

    Science.gov (United States)

    Li, Yongfeng

    2013-01-01

    Non-homologous end joining (NHEJ) is the dominant DNA double strand break (DSB) repair pathway and involves several NHEJ proteins such as Ku, DNA-PKcs, XRCC4, Ligase IV and so on. Once DSBs are generated, Ku is first recruited to the DNA end, followed by other NHEJ proteins for DNA end processing and ligation. Because of the direct ligation of break ends without the need for a homologous template, NHEJ turns out to be an error-prone but efficient repair pathway. Some mechanisms have been proposed of how the efficiency of NHEJ repair is affected. The type of DNA damage is an important factor of NHEJ repair. For instance, the length of DNA fragment may determine the recruitment efficiency of NHEJ protein such as Ku [1], or the complexity of the DNA breaks [2] is accounted for the choice of NHEJ proteins and subpathway of NHEJ repair. On the other hand, the chromatin structure also plays a role of the accessibility of NHEJ protein to the DNA damage site. In this talk, some mathematical models of NHEJ, that consist of series of biochemical reactions complying with the laws of chemical reaction (e.g. mass action, etc.), will be introduced. By mathematical and numerical analysis and parameter estimation, the models are able to capture the qualitative biological features and show good agreement with experimental data. As conclusions, from the viewpoint of modeling, how the NHEJ proteins are recruited will be first discussed for connection between the classical sequential model [4] and recently proposed two-phase model [5]. Then how the NHEJ repair pathway is affected, by the length of DNA fragment [6], the complexity of DNA damage [7] and the chromatin structure [8], will be addressed

  1. Human DNA ligase III bridges two DNA ends to promote specific intermolecular DNA end joining

    Science.gov (United States)

    Kukshal, Vandna; Kim, In-Kwon; Hura, Gregory L.; Tomkinson, Alan E.; Tainer, John A.; Ellenberger, Tom

    2015-01-01

    Mammalian DNA ligase III (LigIII) functions in both nuclear and mitochondrial DNA metabolism. In the nucleus, LigIII has functional redundancy with DNA ligase I whereas LigIII is the only mitochondrial DNA ligase and is essential for the survival of cells dependent upon oxidative respiration. The unique LigIII zinc finger (ZnF) domain is not required for catalytic activity but senses DNA strand breaks and stimulates intermolecular ligation of two DNAs by an unknown mechanism. Consistent with this activity, LigIII acts in an alternative pathway of DNA double strand break repair that buttresses canonical non-homologous end joining (NHEJ) and is manifest in NHEJ-defective cancer cells, but how LigIII acts in joining intermolecular DNA ends versus nick ligation is unclear. To investigate how LigIII efficiently joins two DNAs, we developed a real-time, fluorescence-based assay of DNA bridging suitable for high-throughput screening. On a nicked duplex DNA substrate, the results reveal binding competition between the ZnF and the oligonucleotide/oligosaccharide-binding domain, one of three domains constituting the LigIII catalytic core. In contrast, these domains collaborate and are essential for formation of a DNA-bridging intermediate by adenylated LigIII that positions a pair of blunt-ended duplex DNAs for efficient and specific intermolecular ligation. PMID:26130724

  2. Contribution of canonical nonhomologous end joining to chromosomal rearrangements is enhanced by ATM kinase deficiency.

    Science.gov (United States)

    Bhargava, Ragini; Carson, Caree R; Lee, Gabriella; Stark, Jeremy M

    2017-01-24

    A likely mechanism of chromosomal rearrangement formation involves joining the ends from two different chromosomal double-strand breaks (DSBs). These events could potentially be mediated by either of two end-joining (EJ) repair pathways [canonical nonhomologous end joining (C-NHEJ) or alternative end joining (ALT-EJ)], which cause distinct rearrangement junction patterns. The relative role of these EJ pathways during rearrangement formation has remained controversial. Along these lines, we have tested whether the DNA damage response mediated by the Ataxia Telangiectasia Mutated (ATM) kinase may affect the relative influence of C-NHEJ vs. ALT-EJ on rearrangement formation. We developed a reporter in mouse cells for a 0.4-Mbp deletion rearrangement that is formed by EJ between two DSBs induced by the Cas9 endonuclease. We found that disruption of the ATM kinase causes an increase in the frequency of the rearrangement as well as a shift toward rearrangement junctions that show hallmarks of C-NHEJ. Furthermore, ATM suppresses rearrangement formation in an experimental condition, in which C-NHEJ is the predominant EJ repair event (i.e., expression of the 3' exonuclease Trex2). Finally, several C-NHEJ factors are required for the increase in rearrangement frequency caused by inhibition of the ATM kinase. We also examined ATM effectors and found that H2AX shows a similar influence as ATM, whereas the influence of ATM on this rearrangement seems independent of 53BP1. We suggest that the contribution of the C-NHEJ pathway to the formation of a 0.4-Mbp deletion rearrangement is enhanced in ATM-deficient cells.

  3. Nonhomologous DNA End Joining in Cell-Free Extracts

    Directory of Open Access Journals (Sweden)

    Sheetal Sharma

    2010-01-01

    Full Text Available Among various DNA damages, double-strand breaks (DSBs are considered as most deleterious, as they may lead to chromosomal rearrangements and cancer when unrepaired. Nonhomologous DNA end joining (NHEJ is one of the major DSB repair pathways in higher organisms. A large number of studies on NHEJ are based on in vitro systems using cell-free extracts. In this paper, we summarize the studies on NHEJ performed by various groups in different cell-free repair systems.

  4. Direct Involvement of Retinoblastoma Family Proteins in DNA Repair by Non-homologous End-Joining

    Directory of Open Access Journals (Sweden)

    Rebecca Cook

    2015-03-01

    Full Text Available Deficiencies in DNA double-strand break (DSB repair lead to genetic instability, a recognized cause of cancer initiation and evolution. We report that the retinoblastoma tumor suppressor protein (RB1 is required for DNA DSB repair by canonical non-homologous end-joining (cNHEJ. Support of cNHEJ involves a mechanism independent of RB1’s cell-cycle function and depends on its amino terminal domain with which it binds to NHEJ components XRCC5 and XRCC6. Cells with engineered loss of RB family function as well as cancer-derived cells with mutational RB1 loss show substantially reduced levels of cNHEJ. RB1 variants disabled for the interaction with XRCC5 and XRCC6, including a cancer-associated variant, are unable to support cNHEJ despite being able to confer cell-cycle control. Our data identify RB1 loss as a candidate driver of structural genomic instability and a causative factor for cancer somatic heterogeneity and evolution.

  5. The endless tale of non-homologous end-joining.

    Science.gov (United States)

    Weterings, Eric; Chen, David J

    2008-01-01

    DNA double-strand breaks (DSBs) are introduced in cells by ionizing radiation and reactive oxygen species. In addition, they are commonly generated during V(D)J recombination, an essential aspect of the developing immune system. Failure to effectively repair these DSBs can result in chromosome breakage, cell death, onset of cancer, and defects in the immune system of higher vertebrates. Fortunately, all mammalian cells possess two enzymatic pathways that mediate the repair of DSBs: homologous recombination and non-homologous end-joining (NHEJ). The NHEJ process utilizes enzymes that capture both ends of the broken DNA molecule, bring them together in a synaptic DNA-protein complex, and finally repair the DNA break. In this review, all the known enzymes that play a role in the NHEJ process are discussed and a working model for the co-operation of these enzymes during DSB repair is presented.

  6. Environmental Stress Induces Trinucleotide Repeat Mutagenesis in Human Cells by Alt-Nonhomologous End Joining Repair.

    Science.gov (United States)

    Chatterjee, Nimrat; Lin, Yunfu; Yotnda, Patricia; Wilson, John H

    2016-07-31

    Multiple pathways modulate the dynamic mutability of trinucleotide repeats (TNRs), which are implicated in neurodegenerative disease and evolution. Recently, we reported that environmental stresses induce TNR mutagenesis via stress responses and rereplication, with more than 50% of mutants carrying deletions or insertions-molecular signatures of DNA double-strand break repair. We now show that knockdown of alt-nonhomologous end joining (alt-NHEJ) components-XRCC1, LIG3, and PARP1-suppresses stress-induced TNR mutagenesis, in contrast to the components of homologous recombination and NHEJ, which have no effect. Thus, alt-NHEJ, which contributes to genetic mutability in cancer cells, also plays a novel role in environmental stress-induced TNR mutagenesis. Published by Elsevier Ltd.

  7. DNA double strand break repair in mammalian cells: role of MRE11 and BLM proteins at the initiation of Non Homologous End Joining (NHEJ)

    International Nuclear Information System (INIS)

    Grabarz, Anastazja

    2011-01-01

    DNA double strand breaks (DSBs) are highly cytotoxic lesions, which can lead to genetic rearrangements. Two pathways are responsible for repairing these lesions: homologous recombination (HR) and non homologous end joining (NHEJ). In our laboratory, an intrachromosomal substrate has been established in order to measure the efficiency and the fidelity of NHEJ in living cells (Guirouilh-Barbat 2004). This approach led us to identify a KU-independent alternative pathway, which uses micro homologies in the proximity of the junction to accomplish repair - the alternative NHEJ (Guirouilh-Barbat 2004, Guirouilh-Barbat et Rass 2007). The goal of my thesis consisted in identifying and characterising major actors of this pathway. In the absence of KU, alternative NHEJ would be initiated by ssDNA resection of damaged ends. We showed that the nuclease activity of MRE11 is necessary for this mechanism. MRE11 overexpression leads to a two fold stimulation of NHEJ efficiency, while the extinction of MRE11 by siRNA results in a two fold decrease. Our results demonstrate that the proteins RAD50 and CtIP act in the same pathway as MRE11. Moreover, in cells deficient for XRCC4, MIRIN - an inhibitor of the MRN complex - leads to a decrease in repair efficiency, implicating MRE11 in alternative NHEJ. We also showed that MRE11 can act in an ATM-dependent and independent manner (Rass et Grabarz Nat Struct Mol Biol 2009). The initiation of break resection needs to be pursued by a more extensive degradation of DNA, which is accomplished in yeast by the proteins Exo1 and Sgs1/Dna2. In human cells, in vitro studies have recently proposed a similar model of a two-step break resection. We chose to elucidate the role of one of the human homologs of Sgs1 - the RecQ helicase BLM - in the resection process. Our experiments show, that he absence of BLM decreases the efficiency of end joining by NHEJ, accompanied by an increase in error-prone events, especially long-range deletions (≥200 nt). This

  8. The Ku70/80 ring in Non-Homologous End-Joining

    DEFF Research Database (Denmark)

    Kragelund, Birthe Brandt; Weterings, Eric; Hartmann-Petersen, Rasmus

    2016-01-01

    /80 heterodimer is a key-player in the NHEJ pathway and binds to DNA termini with high affinity, where it helps to protect DNA ends from degradation and to recruit other NHEJ factors required for repair. The mechanism of Ku70/80 detachment from the DNA helix after completion of DNA repair is incompletely......Non-homologous end-joining (NHEJ) is an essential DNA double strand break repair pathway during all cell cycle stages. Deficiency in NHEJ factors can lead to accumulation of unrepaired DNA breaks or faulty DNA repair, which may ultimately result in cell death, senescence or carcinogenesis. The Ku70...... understood. Some data suggest that certain DNA repair factors are ubiquitylated and targeted for proteasomal degradation after repair. Recent studies suggest that Ku80 is conjugated to lysine48-linked ubiquitin chains by the Skp1-Cullin-F-box (SCF) complex and/or the RING finger protein 8 (RNF8) ubiquitin-protein...

  9. Non-homologous end joining mediated DNA repair is impaired in the NUP98-HOXD13 mouse model for myelodysplastic syndrome.

    Science.gov (United States)

    Puthiyaveetil, Abdul Gafoor; Reilly, Christopher M; Pardee, Timothy S; Caudell, David L

    2013-01-01

    Chromosomal translocations typically impair cell differentiation and often require secondary mutations for malignant transformation. However, the role of a primary translocation in the development of collaborating mutations is debatable. To delineate the role of leukemic translocation NUP98-HOXD13 (NHD13) in secondary mutagenesis, DNA break and repair mechanisms in stimulated mouse B lymphocytes expressing NHD13 were analyzed. Our results showed significantly reduced expression of non-homologous end joining (NHEJ)-mediated DNA repair genes, DNA Pkcs, DNA ligase4, and Xrcc4 leading to cell cycle arrest at G2/M phase. Our results showed that expression of NHD13 fusion gene resulted in impaired NHEJ-mediated DNA break repair. Copyright © 2012 Elsevier Ltd. All rights reserved.

  10. Human induced pluripotent cells resemble embryonic stem cells demonstrating enhanced levels of DNA repair and efficacy of nonhomologous end-joining

    Energy Technology Data Exchange (ETDEWEB)

    Fan Jinshui; Robert, Carine [Department of Radiation Oncology, University of Maryland School of Medicine, 655 West Baltimore Street, BRB 7-023A, Baltimore, MD 21201 (United States); Jang, Yoon-Young; Liu Hua; Sharkis, Saul; Baylin, Stephen Bruce [Johns Hopkins University School of Medicine, Department of Oncology, Baltimore, MD 21231-1000 (United States); Rassool, Feyruz Virgilia, E-mail: frassool@som.umaryland.edu [Department of Radiation Oncology, University of Maryland School of Medicine, 655 West Baltimore Street, BRB 7-023A, Baltimore, MD 21201 (United States)

    2011-08-01

    Highlights: {yields} iPSC and hESC demonstrate a similar cell cycle profile, with increased S phase cells and decreased G0/G1. {yields} iPSC and hESC increased ROS and decreased DSBs, compared with differentiated parental cells. {yields} iPSC and hESC demonstrate elevated DSB repair activity, including nonhomologous end-joining, compared with differentiated parental cells. {yields} iPSC however show a partial apoptotic response to DNA damage, compared to hESC. {yields} DNA damage responses may constitute important markers for the efficacy of iPSC reprogramming. - Abstract: To maintain the integrity of the organism, embryonic stem cells (ESC) need to maintain their genomic integrity in response to DNA damage. DNA double strand breaks (DSBs) are one of the most lethal forms of DNA damage and can have disastrous consequences if not repaired correctly, leading to cell death, genomic instability and cancer. How human ESC (hESC) maintain genomic integrity in response to agents that cause DSBs is relatively unclear. Adult somatic cells can be induced to 'dedifferentiate' into induced pluripotent stem cells (iPSC) and reprogram into cells of all three germ layers. Whether iPSC have reprogrammed the DNA damage response is a critical question in regenerative medicine. Here, we show that hESC demonstrate high levels of endogenous reactive oxygen species (ROS) which can contribute to DNA damage and may arise from high levels of metabolic activity. To potentially counter genomic instability caused by DNA damage, we find that hESC employ two strategies: First, these cells have enhanced levels of DNA repair proteins, including those involved in repair of DSBs, and they demonstrate elevated nonhomologous end-joining (NHEJ) activity and repair efficacy, one of the main pathways for repairing DSBs. Second, they are hypersensitive to DNA damaging agents, as evidenced by a high level of apoptosis upon irradiation. Importantly, iPSC, unlike the parent cells they are derived

  11. Human induced pluripotent cells resemble embryonic stem cells demonstrating enhanced levels of DNA repair and efficacy of nonhomologous end-joining

    International Nuclear Information System (INIS)

    Fan Jinshui; Robert, Carine; Jang, Yoon-Young; Liu Hua; Sharkis, Saul; Baylin, Stephen Bruce; Rassool, Feyruz Virgilia

    2011-01-01

    Highlights: → iPSC and hESC demonstrate a similar cell cycle profile, with increased S phase cells and decreased G0/G1. → iPSC and hESC increased ROS and decreased DSBs, compared with differentiated parental cells. → iPSC and hESC demonstrate elevated DSB repair activity, including nonhomologous end-joining, compared with differentiated parental cells. → iPSC however show a partial apoptotic response to DNA damage, compared to hESC. → DNA damage responses may constitute important markers for the efficacy of iPSC reprogramming. - Abstract: To maintain the integrity of the organism, embryonic stem cells (ESC) need to maintain their genomic integrity in response to DNA damage. DNA double strand breaks (DSBs) are one of the most lethal forms of DNA damage and can have disastrous consequences if not repaired correctly, leading to cell death, genomic instability and cancer. How human ESC (hESC) maintain genomic integrity in response to agents that cause DSBs is relatively unclear. Adult somatic cells can be induced to 'dedifferentiate' into induced pluripotent stem cells (iPSC) and reprogram into cells of all three germ layers. Whether iPSC have reprogrammed the DNA damage response is a critical question in regenerative medicine. Here, we show that hESC demonstrate high levels of endogenous reactive oxygen species (ROS) which can contribute to DNA damage and may arise from high levels of metabolic activity. To potentially counter genomic instability caused by DNA damage, we find that hESC employ two strategies: First, these cells have enhanced levels of DNA repair proteins, including those involved in repair of DSBs, and they demonstrate elevated nonhomologous end-joining (NHEJ) activity and repair efficacy, one of the main pathways for repairing DSBs. Second, they are hypersensitive to DNA damaging agents, as evidenced by a high level of apoptosis upon irradiation. Importantly, iPSC, unlike the parent cells they are derived from, mimic hESC in their ROS levels

  12. The Ku80 carboxy terminus stimulates joining and artemis-mediated processing of DNA ends.

    Science.gov (United States)

    Weterings, Eric; Verkaik, Nicole S; Keijzers, Guido; Florea, Bogdan I; Wang, Shih-Ya; Ortega, Laura G; Uematsu, Naoya; Chen, David J; van Gent, Dik C

    2009-03-01

    Repair of DNA double-strand breaks (DSBs) is predominantly mediated by nonhomologous end joining (NHEJ) in mammalian cells. NHEJ requires binding of the Ku70-Ku80 heterodimer (Ku70/80) to the DNA ends and subsequent recruitment of the DNA-dependent protein kinase catalytic subunit (DNA-PK(CS)) and the XRCC4/ligase IV complex. Activation of the DNA-PK(CS) serine/threonine kinase requires an interaction with Ku70/80 and is essential for NHEJ-mediated DSB repair. In contrast to previous models, we found that the carboxy terminus of Ku80 is not absolutely required for the recruitment and activation of DNA-PK(CS) at DSBs, although cells that harbored a carboxy-terminal deletion in the Ku80 gene were sensitive to ionizing radiation and showed reduced end-joining capacity. More detailed analysis of this repair defect showed that DNA-PK(CS) autophosphorylation at Thr2647 was diminished, while Ser2056 was phosphorylated to normal levels. This resulted in severely reduced levels of Artemis nuclease activity in vivo and in vitro. We therefore conclude that the Ku80 carboxy terminus is important to support DNA-PK(CS) autophosphorylation at specific sites, which facilitates DNA end processing by the Artemis endonuclease and the subsequent joining reaction.

  13. Coincident In Vitro Analysis of DNA-PK-Dependent and -Independent Nonhomologous End Joining

    Directory of Open Access Journals (Sweden)

    Cynthia L. Hendrickson

    2010-01-01

    Full Text Available In mammalian cells, DNA double-strand breaks (DSBs are primarily repaired by nonhomologous end joining (NHEJ. The current model suggests that the Ku 70/80 heterodimer binds to DSB ends and recruits DNA-PKcs to form the active DNA-dependent protein kinase, DNA-PK. Subsequently, XRCC4, DNA ligase IV, XLF and most likely, other unidentified components participate in the final DSB ligation step. Therefore, DNA-PK plays a key role in NHEJ due to its structural and regulatory functions that mediate DSB end joining. However, recent studies show that additional DNA-PK-independent NHEJ pathways also exist. Unfortunately, the presence of DNA-PKcs appears to inhibit DNA-PK-independent NHEJ, and in vitro analysis of DNA-PK-independent NHEJ in the presence of the DNA-PKcs protein remains problematic. We have developed an in vitro assay that is preferentially active for DNA-PK-independent DSB repair based solely on its reaction conditions, facilitating coincident differential biochemical analysis of the two pathways. The results indicate the biochemically distinct nature of the end-joining mechanisms represented by the DNA-PK-dependent and -independent NHEJ assays as well as functional differences between the two pathways.

  14. Homologous recombination and non-homologous end-joining repair pathways in bovine embryos with different developmental competence

    Energy Technology Data Exchange (ETDEWEB)

    Henrique Barreta, Marcos [Universidade Federal de Santa Catarina, Campus Universitario de Curitibanos, Curitibanos, SC (Brazil); Laboratorio de Biotecnologia e Reproducao Animal-BioRep, Universidade Federal de Santa Maria, Santa Maria, RS (Brazil); Garziera Gasperin, Bernardo; Braga Rissi, Vitor; Cesaro, Matheus Pedrotti de [Laboratorio de Biotecnologia e Reproducao Animal-BioRep, Universidade Federal de Santa Maria, Santa Maria, RS (Brazil); Ferreira, Rogerio [Centro de Educacao Superior do Oeste-Universidade do Estado de Santa Catarina, Chapeco, SC (Brazil); Oliveira, Joao Francisco de; Goncalves, Paulo Bayard Dias [Laboratorio de Biotecnologia e Reproducao Animal-BioRep, Universidade Federal de Santa Maria, Santa Maria, RS (Brazil); Bordignon, Vilceu, E-mail: vilceu.bordignon@mcgill.ca [Department of Animal Science, McGill University, Ste-Anne-De-Bellevue, QC (Canada)

    2012-10-01

    This study investigated the expression of genes controlling homologous recombination (HR), and non-homologous end-joining (NHEJ) DNA-repair pathways in bovine embryos of different developmental potential. It also evaluated whether bovine embryos can respond to DNA double-strand breaks (DSBs) induced with ultraviolet irradiation by regulating expression of genes involved in HR and NHEJ repair pathways. Embryos with high, intermediate or low developmental competence were selected based on the cleavage time after in vitro insemination and were removed from in vitro culture before (36 h), during (72 h) and after (96 h) the expected period of embryonic genome activation. All studied genes were expressed before, during and after the genome activation period regardless the developmental competence of the embryos. Higher mRNA expression of 53BP1 and RAD52 was found before genome activation in embryos with low developmental competence. Expression of 53BP1, RAD51 and KU70 was downregulated at 72 h and upregulated at 168 h post-insemination in response to DSBs induced by ultraviolet irradiation. In conclusion, important genes controlling HR and NHEJ DNA-repair pathways are expressed in bovine embryos, however genes participating in these pathways are only regulated after the period of embryo genome activation in response to ultraviolet-induced DSBs.

  15. Homologous recombination and non-homologous end-joining repair pathways in bovine embryos with different developmental competence

    International Nuclear Information System (INIS)

    Henrique Barreta, Marcos; Garziera Gasperin, Bernardo; Braga Rissi, Vitor; Cesaro, Matheus Pedrotti de; Ferreira, Rogério; Oliveira, João Francisco de; Gonçalves, Paulo Bayard Dias; Bordignon, Vilceu

    2012-01-01

    This study investigated the expression of genes controlling homologous recombination (HR), and non-homologous end-joining (NHEJ) DNA-repair pathways in bovine embryos of different developmental potential. It also evaluated whether bovine embryos can respond to DNA double-strand breaks (DSBs) induced with ultraviolet irradiation by regulating expression of genes involved in HR and NHEJ repair pathways. Embryos with high, intermediate or low developmental competence were selected based on the cleavage time after in vitro insemination and were removed from in vitro culture before (36 h), during (72 h) and after (96 h) the expected period of embryonic genome activation. All studied genes were expressed before, during and after the genome activation period regardless the developmental competence of the embryos. Higher mRNA expression of 53BP1 and RAD52 was found before genome activation in embryos with low developmental competence. Expression of 53BP1, RAD51 and KU70 was downregulated at 72 h and upregulated at 168 h post-insemination in response to DSBs induced by ultraviolet irradiation. In conclusion, important genes controlling HR and NHEJ DNA-repair pathways are expressed in bovine embryos, however genes participating in these pathways are only regulated after the period of embryo genome activation in response to ultraviolet-induced DSBs.

  16. Lung Basal Stem Cells Rapidly Repair DNA Damage Using the Error-Prone Nonhomologous End-Joining Pathway

    Science.gov (United States)

    Weeden, Clare E.; Chen, Yunshun; Ma, Stephen B.; Hu, Yifang; Ramm, Georg; Sutherland, Kate D.; Smyth, Gordon K.

    2017-01-01

    Lung squamous cell carcinoma (SqCC), the second most common subtype of lung cancer, is strongly associated with tobacco smoking and exhibits genomic instability. The cellular origins and molecular processes that contribute to SqCC formation are largely unexplored. Here we show that human basal stem cells (BSCs) isolated from heavy smokers proliferate extensively, whereas their alveolar progenitor cell counterparts have limited colony-forming capacity. We demonstrate that this difference arises in part because of the ability of BSCs to repair their DNA more efficiently than alveolar cells following ionizing radiation or chemical-induced DNA damage. Analysis of mice harbouring a mutation in the DNA-dependent protein kinase catalytic subunit (DNA-PKcs), a key enzyme in DNA damage repair by nonhomologous end joining (NHEJ), indicated that BSCs preferentially repair their DNA by this error-prone process. Interestingly, polyploidy, a phenomenon associated with genetically unstable cells, was only observed in the human BSC subset. Expression signature analysis indicated that BSCs are the likely cells of origin of human SqCC and that high levels of NHEJ genes in SqCC are correlated with increasing genomic instability. Hence, our results favour a model in which heavy smoking promotes proliferation of BSCs, and their predilection for error-prone NHEJ could lead to the high mutagenic burden that culminates in SqCC. Targeting DNA repair processes may therefore have a role in the prevention and therapy of SqCC. PMID:28125611

  17. The Ku80 carboxy terminus stimulates joining and artemis-mediated processing of DNA ends

    DEFF Research Database (Denmark)

    Weterings, Eric; Verkaik, Nicole S; Keijzers, Guido

    2008-01-01

    Repair of DNA double-strand breaks (DSBs) is predominantly mediated by nonhomologous end joining (NHEJ) in mammalian cells. NHEJ requires binding of the Ku70-Ku80 heterodimer (Ku70/80) to the DNA ends and subsequent recruitment of the DNA-dependent protein kinase catalytic subunit (DNA-PK(CS)) an......Repair of DNA double-strand breaks (DSBs) is predominantly mediated by nonhomologous end joining (NHEJ) in mammalian cells. NHEJ requires binding of the Ku70-Ku80 heterodimer (Ku70/80) to the DNA ends and subsequent recruitment of the DNA-dependent protein kinase catalytic subunit (DNA......-PK(CS)) and the XRCC4/ligase IV complex. Activation of the DNA-PK(CS) serine/threonine kinase requires an interaction with Ku70/80 and is essential for NHEJ-mediated DSB repair. In contrast to previous models, we found that the carboxy terminus of Ku80 is not absolutely required for the recruitment and activation...... was phosphorylated to normal levels. This resulted in severely reduced levels of Artemis nuclease activity in vivo and in vitro. We therefore conclude that the Ku80 carboxy terminus is important to support DNA-PK(CS) autophosphorylation at specific sites, which facilitates DNA end processing by the Artemis...

  18. Requirement for XLF/Cernunnos in alignment-based gap filling by DNA polymerases lambda and mu for nonhomologous end joining in human whole-cell extracts.

    Science.gov (United States)

    Akopiants, Konstantin; Zhou, Rui-Zhe; Mohapatra, Susovan; Valerie, Kristoffer; Lees-Miller, Susan P; Lee, Kyung-Jong; Chen, David J; Revy, Patrick; de Villartay, Jean-Pierre; Povirk, Lawrence F

    2009-07-01

    XLF/Cernunnos is a core protein of the nonhomologous end-joining pathway of DNA double-strand break repair. To better define the role of Cernunnos in end joining, whole-cell extracts were prepared from Cernunnos-deficient human cells. These extracts effected little joining of DNA ends with cohesive 5' or 3' overhangs, and no joining at all of partially complementary 3' overhangs that required gap filling prior to ligation. Assays in which gap-filled but unligated intermediates were trapped using dideoxynucleotides revealed that there was no gap filling on aligned DSB ends in the Cernunnos-deficient extracts. Recombinant Cernunnos protein restored gap filling and end joining of partially complementary overhangs, and stimulated joining of cohesive ends more than twentyfold. XLF-dependent gap filling was nearly eliminated by immunodepletion of DNA polymerase lambda, but was restored by addition of either polymerase lambda or polymerase mu. Thus, Cernunnos is essential for gap filling by either polymerase during nonhomologous end joining, suggesting that it plays a major role in aligning the two DNA ends in the repair complex.

  19. Requirement for XLF/Cernunnos in alignment-based gap filling by DNA polymerases ? and ? for nonhomologous end joining in human whole-cell extracts

    OpenAIRE

    Akopiants, Konstantin; Zhou, Rui-Zhe; Mohapatra, Susovan; Valerie, Kristoffer; Lees-Miller, Susan P.; Lee, Kyung-Jong; Chen, David J.; Revy, Patrick; de Villartay, Jean-Pierre; Povirk, Lawrence F.

    2009-01-01

    XLF/Cernunnos is a core protein of the nonhomologous end-joining pathway of DNA double-strand break repair. To better define the role of Cernunnos in end joining, whole-cell extracts were prepared from Cernunnos-deficient human cells. These extracts effected little joining of DNA ends with cohesive 5? or 3? overhangs, and no joining at all of partially complementary 3? overhangs that required gap filling prior to ligation. Assays in which gap-filled but unligated intermediates were trapped us...

  20. Ectopic expression of RNF168 and 53BP1 increases mutagenic but not physiological non-homologous end joining

    DEFF Research Database (Denmark)

    Zong, Dali; Callén, Elsa; Pegoraro, Gianluca

    2015-01-01

    DNA double strand breaks (DSBs) formed during S phase are preferentially repaired by homologous recombination (HR), whereas G1 DSBs, such as those occurring during immunoglobulin class switch recombination (CSR), are repaired by non-homologous end joining (NHEJ). The DNA damage response proteins ...

  1. Mycobacterium tuberculosis and Mycobacterium marinum non-homologous end-joining proteins can function together to join DNA ends in Escherichia coli.

    Science.gov (United States)

    Wright, Douglas G; Castore, Reneau; Shi, Runhua; Mallick, Amrita; Ennis, Don G; Harrison, Lynn

    2017-03-01

    Mycobacterium tuberculosis and Mycobacterium smegmatis express a Ku protein and a DNA ligase D and are able to repair DNA double strand breaks (DSBs) by non-homologous end-joining (NHEJ). This pathway protects against DNA damage when bacteria are in stationary phase. Mycobacterium marinum is a member of this mycobacterium family and like M. tuberculosis is pathogenic. M. marinum lives in water, forms biofilms and infects fish and frogs. M. marinum is a biosafety level 2 (BSL2) organism as it can infect humans, although infections are limited to the skin. M. marinum is accepted as a model to study mycobacterial pathogenesis, as M. marinum and M. tuberculosis are genetically closely related and have similar mechanisms of survival and persistence inside macrophage. The aim of this study was to determine whether M. marinum could be used as a model to understand M. tuberculosis NHEJ repair. We identified and cloned the M. marinum genes encoding NHEJ proteins and generated E. coli strains that express the M. marinum Ku (Mm-Ku) and ligase D (Mm-Lig) individually or together (LHmKumLig strain) from expression vectors integrated at phage attachment sites in the genome. We demonstrated that Mm-Ku and Mm-Lig are both required to re-circularize Cla I-linearized plasmid DNA in E. coli. We compared repair of strain LHmKumLig with that of an E. coli strain (BWKuLig#2) expressing the M. tuberculosis Ku (Mt-Ku) and ligase D (Mt-Lig), and found that LHmKumLig performed 3.5 times more repair and repair was more accurate than BWKuLig#2. By expressing the Mm-Ku with the Mt-Lig, or the Mt-Ku with the Mm-Lig in E. coli, we have shown that the NHEJ proteins from M. marinum and M. tuberculosis can function together to join DNA DSBs. NHEJ repair is therefore conserved between the two species. Consequently, M. marinum is a good model to study NHEJ repair during mycobacterial pathogenesis. © The Author 2016. Published by Oxford University Press on behalf of the UK Environmental Mutagen

  2. Non-homologous end-joining genes are not inactivated in human radiation-induced sarcomas with genomic instability

    International Nuclear Information System (INIS)

    Lefevre, S.H.; Coquelle, A.; Gonin-Laurent, N.

    2005-01-01

    DNA double-strand break (DSB) repair pathways are implicated in the maintenance of genomic stability. However the alterations of these pathways, as may occur in human tumor cells with strong genomic instability, remain poorly characterized. We analyzed the loss of heterozygosity (LOH) and the presence of mutations for a series of genes implicated in DSB repair by non-homologous end-joining in five radiation-induced sarcomas devoid of both active Tp53 and Rb1. LOH was recurrently observed for 8 of the 9 studied genes (KU70, KU80, XRCC4, LIG4, Artemis, MRE11, RAD50, NBS1) but not for DNA-PKcs. No mutation was found in the remaining allele of the genes with LOH and the mRNA expression did not correlate with the allelic status. Our findings suggest that non-homologous end-joining repair pathway alteration is unlikely to be involved in the high genomic instability observed in these tumors. (author)

  3. Biochemical Kinetics Model of DSB Repair and GammaH2AX FOCI by Non-homologous End Joining

    Science.gov (United States)

    Cucinotta, Francis, A.; Pluth, Janice M.; Anderson, Jennifer A.; Harper, Jane V.; O'Neill, Peter

    2007-01-01

    We developed a biochemical kinetics approach to describe the repair of double strand breaks (DSB) produced by low LET radiation by modeling molecular events associated with the mechanisms of non-homologous end-joining (NHEJ). A system of coupled non-linear ordinary differential equations describes the induction of DSB and activation pathways for major NHEJ components including Ku(sub 70/80), DNA-PK(sub cs), and the Ligase IV-XRCC4 hetero-dimer. The autophosphorylation of DNA-PK(sub cs and subsequent induction of gamma-H2AX foci observed after ionizing radiation exposure were modeled. A two-step model of DNA-PK(sub cs) regulation of repair was developed with the initial step allowing access of other NHEJ components to breaks, and a second step limiting access to Ligase IV-XRCC4. Our model assumes that the transition from the first to second-step depends on DSB complexity, with a much slower-rate for complex DSB. The model faithfully reproduced several experimental data sets, including DSB rejoining as measured by pulsed-field electrophoresis (PFGE), quantification of the induction of gamma-H2AX foci, and live cell imaging of the induction of Ku(sub 70/80). Predictions are made for the behaviors of NHEJ components at low doses and dose-rates, where a steady-state is found at dose-rates of 0.1 Gy/hr or lower.

  4. Altered kinetics of nonhomologous end joining and class switch recombination in ligase IV-deficient B cells.

    Science.gov (United States)

    Han, Li; Yu, Kefei

    2008-11-24

    Immunoglobulin heavy chain class switch recombination (CSR) is believed to occur through the generation and repair of DNA double-strand breaks (DSBs) in the long and repetitive switch regions. Although implied, the role of the major vertebrate DSB repair pathway, nonhomologous end joining (NHEJ), in CSR has been controversial. By somatic gene targeting of DNA ligase IV (Lig4; a key component of NHEJ) in a B cell line (CH12F3) capable of highly efficient CSR in vitro, we found that NHEJ is required for efficient CSR. Disruption of the Lig4 gene in CH12F3 cells severely inhibits the initial rate of CSR and causes a late cell proliferation defect under cytokine stimulation. However, unlike V(D)J recombination, which absolutely requires NHEJ, CSR accumulates to a substantial level in Lig4-null cells. The data revealed a fast-acting NHEJ and a slow-acting alterative end joining of switch region breaks during CSR.

  5. Agrobacterium May Delay Plant Nonhomologous End-Joining DNA Repair via XRCC4 to Favor T-DNA Integration[W

    Science.gov (United States)

    Vaghchhipawala, Zarir E.; Vasudevan, Balaji; Lee, Seonghee; Morsy, Mustafa R.; Mysore, Kirankumar S.

    2012-01-01

    Agrobacterium tumefaciens is a soilborne pathogen that causes crown gall disease in many dicotyledonous plants by transfer of a portion of its tumor-inducing plasmid (T-DNA) into the plant genome. Several plant factors that play a role in Agrobacterium attachment to plant cells and transport of T-DNA to the nucleus have been identified, but the T-DNA integration step during transformation is poorly understood and has been proposed to occur via nonhomologous end-joining (NHEJ)–mediated double-strand DNA break (DSB) repair. Here, we report a negative role of X-RAY CROSS COMPLEMENTATION GROUP4 (XRCC4), one of the key proteins required for NHEJ, in Agrobacterium T-DNA integration. Downregulation of XRCC4 in Arabidopsis and Nicotiana benthamiana increased stable transformation due to increased T-DNA integration. Overexpression of XRCC4 in Arabidopsis decreased stable transformation due to decreased T-DNA integration. Interestingly, XRCC4 directly interacted with Agrobacterium protein VirE2 in a yeast two-hybrid system and in planta. VirE2-expressing Arabidopsis plants were more susceptible to the DNA damaging chemical bleomycin and showed increased stable transformation. We hypothesize that VirE2 titrates or excludes active XRCC4 protein available for DSB repair, thus delaying the closure of DSBs in the chromosome, providing greater opportunity for T-DNA to integrate. PMID:23064322

  6. In vivo blunt-end cloning through CRISPR/Cas9-facilitated non-homologous end-joining

    Science.gov (United States)

    Geisinger, Jonathan M.; Turan, Sören; Hernandez, Sophia; Spector, Laura P.; Calos, Michele P.

    2016-01-01

    The CRISPR/Cas9 system facilitates precise DNA modifications by generating RNA-guided blunt-ended double-strand breaks. We demonstrate that guide RNA pairs generate deletions that are repaired with a high level of precision by non-homologous end-joining in mammalian cells. We present a method called knock-in blunt ligation for exploiting these breaks to insert exogenous PCR-generated sequences in a homology-independent manner without loss of additional nucleotides. This method is useful for making precise additions to the genome such as insertions of marker gene cassettes or functional elements, without the need for homology arms. We successfully utilized this method in human and mouse cells to insert fluorescent protein cassettes into various loci, with efficiencies up to 36% in HEK293 cells without selection. We also created versions of Cas9 fused to the FKBP12-L106P destabilization domain in an effort to improve Cas9 performance. Our in vivo blunt-end cloning method and destabilization-domain-fused Cas9 variant increase the repertoire of precision genome engineering approaches. PMID:26762978

  7. Highly precise and developmentally programmed genome assembly in Paramecium requires ligase IV-dependent end joining.

    Directory of Open Access Journals (Sweden)

    Aurélie Kapusta

    2011-04-01

    Full Text Available During the sexual cycle of the ciliate Paramecium, assembly of the somatic genome includes the precise excision of tens of thousands of short, non-coding germline sequences (Internal Eliminated Sequences or IESs, each one flanked by two TA dinucleotides. It has been reported previously that these genome rearrangements are initiated by the introduction of developmentally programmed DNA double-strand breaks (DSBs, which depend on the domesticated transposase PiggyMac. These DSBs all exhibit a characteristic geometry, with 4-base 5' overhangs centered on the conserved TA, and may readily align and undergo ligation with minimal processing. However, the molecular steps and actors involved in the final and precise assembly of somatic genes have remained unknown. We demonstrate here that Ligase IV and Xrcc4p, core components of the non-homologous end-joining pathway (NHEJ, are required both for the repair of IES excision sites and for the circularization of excised IESs. The transcription of LIG4 and XRCC4 is induced early during the sexual cycle and a Lig4p-GFP fusion protein accumulates in the developing somatic nucleus by the time IES excision takes place. RNAi-mediated silencing of either gene results in the persistence of free broken DNA ends, apparently protected against extensive resection. At the nucleotide level, controlled removal of the 5'-terminal nucleotide occurs normally in LIG4-silenced cells, while nucleotide addition to the 3' ends of the breaks is blocked, together with the final joining step, indicative of a coupling between NHEJ polymerase and ligase activities. Taken together, our data indicate that IES excision is a "cut-and-close" mechanism, which involves the introduction of initiating double-strand cleavages at both ends of each IES, followed by DSB repair via highly precise end joining. This work broadens our current view on how the cellular NHEJ pathway has cooperated with domesticated transposases for the emergence of new

  8. Highly precise and developmentally programmed genome assembly in Paramecium requires ligase IV-dependent end joining.

    Science.gov (United States)

    Kapusta, Aurélie; Matsuda, Atsushi; Marmignon, Antoine; Ku, Michael; Silve, Aude; Meyer, Eric; Forney, James D; Malinsky, Sophie; Bétermier, Mireille

    2011-04-01

    During the sexual cycle of the ciliate Paramecium, assembly of the somatic genome includes the precise excision of tens of thousands of short, non-coding germline sequences (Internal Eliminated Sequences or IESs), each one flanked by two TA dinucleotides. It has been reported previously that these genome rearrangements are initiated by the introduction of developmentally programmed DNA double-strand breaks (DSBs), which depend on the domesticated transposase PiggyMac. These DSBs all exhibit a characteristic geometry, with 4-base 5' overhangs centered on the conserved TA, and may readily align and undergo ligation with minimal processing. However, the molecular steps and actors involved in the final and precise assembly of somatic genes have remained unknown. We demonstrate here that Ligase IV and Xrcc4p, core components of the non-homologous end-joining pathway (NHEJ), are required both for the repair of IES excision sites and for the circularization of excised IESs. The transcription of LIG4 and XRCC4 is induced early during the sexual cycle and a Lig4p-GFP fusion protein accumulates in the developing somatic nucleus by the time IES excision takes place. RNAi-mediated silencing of either gene results in the persistence of free broken DNA ends, apparently protected against extensive resection. At the nucleotide level, controlled removal of the 5'-terminal nucleotide occurs normally in LIG4-silenced cells, while nucleotide addition to the 3' ends of the breaks is blocked, together with the final joining step, indicative of a coupling between NHEJ polymerase and ligase activities. Taken together, our data indicate that IES excision is a "cut-and-close" mechanism, which involves the introduction of initiating double-strand cleavages at both ends of each IES, followed by DSB repair via highly precise end joining. This work broadens our current view on how the cellular NHEJ pathway has cooperated with domesticated transposases for the emergence of new mechanisms

  9. Atomic Structure and Nonhomologous End-Joining Function of the Polymerase Component of Bacterial DNA Ligase D

    Energy Technology Data Exchange (ETDEWEB)

    Zhu,H.; Nandakumar, J.; Aniukwu, J.; Wang, L.; Glickman, M.; Lima, C.; Shuman, S.

    2006-01-01

    DNA ligase D (LigD) is a large polyfunctional protein that participates in a recently discovered pathway of nonhomologous end-joining in bacteria. LigD consists of an ATP-dependent ligase domain fused to a polymerase domain (Pol) and a phosphoesterase module. The Pol activity is remarkable for its dependence on manganese, its ability to perform templated and nontemplated primer extension reactions, and its preference for adding ribonucleotides to blunt DNA ends. Here we report the 1.5- Angstroms crystal structure of the Pol domain of Pseudomonas LigD and its complexes with manganese and ATP-dATP substrates, which reveal a minimized polymerase with a two-metal mechanism and a fold similar to that of archaeal DNA primase. Mutational analysis highlights the functionally relevant atomic contacts in the active site. Although distinct nucleoside conformations and contacts for ATP versus dATP are observed in the cocrystals, the functional analysis suggests that the ATP-binding mode is the productive conformation for dNMP and rNMP incorporation. We find that a mutation of Mycobacterium LigD that uniquely ablates the polymerase activity results in increased fidelity of blunt-end double-strand break repair in vivo by virtue of eliminating nucleotide insertions at the recombination junctions. Thus, LigD Pol is a direct catalyst of mutagenic nonhomologous end-joining in vivo. Our studies underscore a previously uncharacterized role for the primase-like polymerase family in DNA repair.

  10. Expression of Mycobacterium tuberculosis Ku and Ligase D in Escherichia coli results in RecA and RecB-independent DNA end-joining at regions of microhomology.

    Science.gov (United States)

    Malyarchuk, Svitlana; Wright, Douglas; Castore, Reneau; Klepper, Emily; Weiss, Bernard; Doherty, Aidan J; Harrison, Lynn

    2007-10-01

    Unlike Escherichia coli, Mycobacterium tuberculosis (Mt) expresses a Ku-like protein and an ATP-dependent DNA ligase that can perform non-homologous end-joining (NHEJ). We have expressed the Mt-Ku and Mt-Ligase D in E. coli using an arabinose-inducible promoter and expression vectors that integrate into specific sites in the E. coli chromosome. E. coli strains have been generated that express the Mt-Ku and Mt-Ligase D on a genetic background that is wild-type for repair, or deficient in either the RecA or RecB protein. Transformation of these strains with linearized plasmid DNA containing a 2bp overhang has demonstrated that expression of both the Mt-Ku and Mt-Ligase D is required for DNA end-joining and that loss of RecA does not prevent this double-strand break repair. Analysis of the re-joined plasmid has shown that repair is predominantly inaccurate and results in the deletion of sequences. Loss of RecB did not prevent the formation of large deletions, but did increase the amount of end-joining. Sequencing the junctions has revealed that the majority of the ligations occurred at regions of microhomology (1-4bps), eliminating one copy of the homologous sequence at the junction. The Mt-Ku and Mt-Ligase D can therefore function in E. coli to re-circularize linear plasmid.

  11. The Ku70/80 ring in Non-Homologous End-Joining: easy to slip on, hard to remove.

    Science.gov (United States)

    Kragelund, Birthe B; Weterings, Eric; Hartmann-Petersen, Rasmus; Keijzers, Guido

    2016-01-01

    Non-homologous end-joining (NHEJ) is an essential DNA double strand break repair pathway during all cell cycle stages. Deficiency in NHEJ factors can lead to accumulation of unrepaired DNA breaks or faulty DNA repair, which may ultimately result in cell death, senescence or carcinogenesis. The Ku70/80 heterodimer is a key-player in the NHEJ pathway and binds to DNA termini with high affinity, where it helps to protect DNA ends from degradation and to recruit other NHEJ factors required for repair. The mechanism of Ku70/80 detachment from the DNA helix after completion of DNA repair is incompletely understood. Some data suggest that certain DNA repair factors are ubiquitylated and targeted for proteasomal degradation after repair. Recent studies suggest that Ku80 is conjugated to lysine48-linked ubiquitin chains by the Skp1-Cullin-F-box (SCF) complex and/or the RING finger protein 8 (RNF8) ubiquitin-protein ligases, followed by rapid proteasomal degradation. In this review we address the structure and function of the Ku70/80 heterodimer and how ubiquitylation may affect the release of Ku70/80 from chromatin and its subsequent degradation via the ubiquitin-proteasome system.

  12. Chromosome End Repair and Genome Stability in Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Susannah F. Calhoun

    2017-08-01

    Full Text Available The human malaria parasite Plasmodium falciparum replicates within circulating red blood cells, where it is subjected to conditions that frequently cause DNA damage. The repair of DNA double-stranded breaks (DSBs is thought to rely almost exclusively on homologous recombination (HR, due to a lack of efficient nonhomologous end joining. However, given that the parasite is haploid during this stage of its life cycle, the mechanisms involved in maintaining genome stability are poorly understood. Of particular interest are the subtelomeric regions of the chromosomes, which contain the majority of the multicopy variant antigen-encoding genes responsible for virulence and disease severity. Here, we show that parasites utilize a competitive balance between de novo telomere addition, also called “telomere healing,” and HR to stabilize chromosome ends. Products of both repair pathways were observed in response to DSBs that occurred spontaneously during routine in vitro culture or resulted from experimentally induced DSBs, demonstrating that both pathways are active in repairing DSBs within subtelomeric regions and that the pathway utilized was determined by the DNA sequences immediately surrounding the break. In combination, these two repair pathways enable parasites to efficiently maintain chromosome stability while also contributing to the generation of genetic diversity.

  13. Either non-homologous ends joining or homologous recombination is required to repair double-strand breaks in the genome of macrophage-internalized Mycobacterium tuberculosis.

    Science.gov (United States)

    Brzostek, Anna; Szulc, Izabela; Klink, Magdalena; Brzezinska, Marta; Sulowska, Zofia; Dziadek, Jaroslaw

    2014-01-01

    The intracellular pathogen Mycobacterium tuberculosis (Mtb) is constantly exposed to a multitude of hostile conditions and is confronted by a variety of potentially DNA-damaging assaults in vivo, primarily from host-generated antimicrobial toxic radicals. Exposure to reactive nitrogen species and/or reactive oxygen species causes different types of DNA damage, including oxidation, depurination, methylation and deamination, that can result in single- or double-strand breaks (DSBs). These breaks affect the integrity of the whole genome and, when left unrepaired, can lead to cell death. Here, we investigated the role of the DSB repair pathways, homologous recombination (HR) and non-homologous ends joining (NHEJ), in the survival of Mtb inside macrophages. To this end, we constructed Mtb strains defective for HR (ΔrecA), NHEJ [Δ(ku,ligD)], or both DSB repair systems [Δ(ku,ligD,recA)]. Experiments using these strains revealed that either HR or NHEJ is sufficient for the survival and propagation of tubercle bacilli inside macrophages. Inhibition of nitric oxide or superoxide anion production with L-NIL or apocynin, respectively, enabled the Δ(ku,ligD,recA) mutant strain lacking both systems to survive intracellularly. Complementation of the Δ(ku,ligD,recA) mutant with an intact recA or ku-ligD rescued the ability of Mtb to propagate inside macrophages.

  14. Either non-homologous ends joining or homologous recombination is required to repair double-strand breaks in the genome of macrophage-internalized Mycobacterium tuberculosis.

    Directory of Open Access Journals (Sweden)

    Anna Brzostek

    Full Text Available The intracellular pathogen Mycobacterium tuberculosis (Mtb is constantly exposed to a multitude of hostile conditions and is confronted by a variety of potentially DNA-damaging assaults in vivo, primarily from host-generated antimicrobial toxic radicals. Exposure to reactive nitrogen species and/or reactive oxygen species causes different types of DNA damage, including oxidation, depurination, methylation and deamination, that can result in single- or double-strand breaks (DSBs. These breaks affect the integrity of the whole genome and, when left unrepaired, can lead to cell death. Here, we investigated the role of the DSB repair pathways, homologous recombination (HR and non-homologous ends joining (NHEJ, in the survival of Mtb inside macrophages. To this end, we constructed Mtb strains defective for HR (ΔrecA, NHEJ [Δ(ku,ligD], or both DSB repair systems [Δ(ku,ligD,recA]. Experiments using these strains revealed that either HR or NHEJ is sufficient for the survival and propagation of tubercle bacilli inside macrophages. Inhibition of nitric oxide or superoxide anion production with L-NIL or apocynin, respectively, enabled the Δ(ku,ligD,recA mutant strain lacking both systems to survive intracellularly. Complementation of the Δ(ku,ligD,recA mutant with an intact recA or ku-ligD rescued the ability of Mtb to propagate inside macrophages.

  15. Environmentally friendly joining of tubes by their ends

    DEFF Research Database (Denmark)

    Silva, Carlos M.A.; Nielsen, Chris Valentin; Alves, Luis M.

    2015-01-01

    This paper proposes an environmentally friendly joining process for connecting tubes by their ends that has the potential to replace current solutions based on fastened, crimped, welded, brazed or adhesive bonded joints. The process is based on a new type of tubular lap joint produced by local...... of deformation and the process feasibility window, and destructive testing to establish the working limits of tubular lap joints under different type of loading conditions. Results demonstrate that the proposed joining process is a flexible and cost-effective technology for connecting tubes by their ends...

  16. Influence of reduced glutathione on end-joining of DNA double-strand breaks: Cytogenetical and molecular approach

    Energy Technology Data Exchange (ETDEWEB)

    Ghoshal, Nitin [Molecular Genetics Laboratory, Department of Biotechnology & Bioinformatics, North-Eastern Hill University, Shillong, Meghalaya-793022 (India); Sharma, Sheetal [Department of Biochemistry, Indian Institute of Science, Bangalore, 560 012 (India); Banerjee, Atanu; Kurkalang, Sillarine [Molecular Genetics Laboratory, Department of Biotechnology & Bioinformatics, North-Eastern Hill University, Shillong, Meghalaya-793022 (India); Raghavan, Sathees C. [Department of Biochemistry, Indian Institute of Science, Bangalore, 560 012 (India); Chatterjee, Anupam, E-mail: chatterjeeanupam@hotmail.com [Molecular Genetics Laboratory, Department of Biotechnology & Bioinformatics, North-Eastern Hill University, Shillong, Meghalaya-793022 (India)

    2017-01-15

    Highlights: • DNA lesions induced by Blem and radiation interact well and form higher frequency of exchange aberrations. • Cellular level of glutathione does influence such interaction of DNA lesions. • Oligomer-based cell-free assay system demonstrated better end-joining efficiency at higher level of endogenous GSH. - Abstract: Radiation induced DNA double-strand breaks (DSB) are the major initial lesions whose misrejoining may lead to exchange aberrations. However, the role of glutathione (GSH), a major cellular thiol, in regulating cell’s sensitivity to DNA damaging agents is not well understood. Influence of endogenous GSH on the efficiency of X-rays and bleomycin (Blem) induced DNA DSBs end-joining has been tested here cytogenetically, in human lymphocytes and Hct116 cells. In another approach, oligomeric DNA (75 bp) containing 5′-compatible and non-compatible overhangs mimicking the endogenous DSB were for rejoining in presence of cell-free extracts from cells having different endogenous GSH levels. Frequency of aberrations, particularly exchange aberrations, was significantly increased when Blem was combined with radiation. The exchange aberration frequency was further enhanced when combined treatment was given at 4 °C since DNA lesions are poorly repaired at 4 °C so that a higher number of DNA breaks persist and interact when shifted from 4 °C to 37 °C. The exchange aberrations increased further when the combined treatment was given to Glutathione-ester (GE) pre-treated cells, indicating more frequent rejoining of DNA lesions in presence of higher cellular GSH. This is further supported by the drastic reduction in frequency of exchange aberrations but significant increase in incidences of deletions when combined treatment was given to GSH-depleted cells. End-joining efficiency of DNA DSBs with compatible ends was better than for non-compatible ends. End-joining efficiency of testicular and MCF7 cell extracts was better than that of lungs and

  17. Non-homologous end joining is the responsible pathway for the repair of fludarabine-induced DNA double strand breaks in mammalian cells

    International Nuclear Information System (INIS)

    Campos-Nebel, Marcelo de; Larripa, Irene; Gonzalez-Cid, Marcela

    2008-01-01

    Fludarabine (FLU), an analogue of adenosine, interferes with DNA synthesis and inhibits the chain elongation leading to replication arrest and DNA double strand break (DSB) formation. Mammalian cells use two main pathways of DSB repair to maintain genomic stability: homologous recombination (HR) and non-homologous end joining (NHEJ). The aim of the present work was to evaluate the repair pathways employed in the restoration of DSB formed following replication arrest induced by FLU in mammalian cells. Replication inhibition was induced in human lymphocytes and fibroblasts by FLU. DSB occurred in a dose-dependent manner on early/middle S-phase cells, as detected by γH2AX foci formation. To test whether conservative HR participates in FLU-induced DSB repair, we measured the kinetics of Rad51 nuclear foci formation in human fibroblasts. There was no significant induction of Rad51 foci after FLU treatment. To further confirm these results, we analyzed the frequency of sister chromatid exchanges (SCE) in both human cells. We did not find increased frequencies of SCE after FLU treatment. To assess the participation of NHEJ pathway in the repair of FLU-induced damage, we used two chemical inhibitors of the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), vanillin and wortmannin. Human fibroblasts pretreated with DNA-PKcs inhibitors showed increased levels of chromosome breakages and became more sensitive to cell death. An active role of NHEJ pathway was also suggested from the analysis of Chinese hamster cell lines. XR-C1 (DNA-PKcs-deficient) and XR-V15B (Ku80-deficient) cells showed hypersensitivity to FLU as evidenced by the increased frequency of chromosome aberrations, decreased mitotic index and impaired survival rates. In contrast, CL-V4B (Rad51C-deficient) and V-C8 (Brca2-deficient) cell lines displayed a FLU-resistant phenotype. Together, our results suggest a major role for NHEJ repair in the preservation of genome integrity against FLU-induced DSB

  18. Non-homologous end joining is the responsible pathway for the repair of fludarabine-induced DNA double strand breaks in mammalian cells

    Energy Technology Data Exchange (ETDEWEB)

    Campos-Nebel, Marcelo de [Departamento de Genetica, Instituto de Investigaciones Hematologicas Mariano R. Castex, Academia Nacional de Medicina, Buenos Aires (Argentina)], E-mail: mnebel@hematologia.anm.edu.ar; Larripa, Irene; Gonzalez-Cid, Marcela [Departamento de Genetica, Instituto de Investigaciones Hematologicas Mariano R. Castex, Academia Nacional de Medicina, Buenos Aires (Argentina)

    2008-11-10

    Fludarabine (FLU), an analogue of adenosine, interferes with DNA synthesis and inhibits the chain elongation leading to replication arrest and DNA double strand break (DSB) formation. Mammalian cells use two main pathways of DSB repair to maintain genomic stability: homologous recombination (HR) and non-homologous end joining (NHEJ). The aim of the present work was to evaluate the repair pathways employed in the restoration of DSB formed following replication arrest induced by FLU in mammalian cells. Replication inhibition was induced in human lymphocytes and fibroblasts by FLU. DSB occurred in a dose-dependent manner on early/middle S-phase cells, as detected by {gamma}H2AX foci formation. To test whether conservative HR participates in FLU-induced DSB repair, we measured the kinetics of Rad51 nuclear foci formation in human fibroblasts. There was no significant induction of Rad51 foci after FLU treatment. To further confirm these results, we analyzed the frequency of sister chromatid exchanges (SCE) in both human cells. We did not find increased frequencies of SCE after FLU treatment. To assess the participation of NHEJ pathway in the repair of FLU-induced damage, we used two chemical inhibitors of the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), vanillin and wortmannin. Human fibroblasts pretreated with DNA-PKcs inhibitors showed increased levels of chromosome breakages and became more sensitive to cell death. An active role of NHEJ pathway was also suggested from the analysis of Chinese hamster cell lines. XR-C1 (DNA-PKcs-deficient) and XR-V15B (Ku80-deficient) cells showed hypersensitivity to FLU as evidenced by the increased frequency of chromosome aberrations, decreased mitotic index and impaired survival rates. In contrast, CL-V4B (Rad51C-deficient) and V-C8 (Brca2-deficient) cell lines displayed a FLU-resistant phenotype. Together, our results suggest a major role for NHEJ repair in the preservation of genome integrity against FLU

  19. Deciphering the Role of Alternative Non-Homologous End Joining (Alt-NHEJ) DNA Repair in Breast Cancer

    Science.gov (United States)

    2017-12-01

    consecutive TTAGGG repeats. To detect rare reads containing fusion junctions, we exploited the novel sequence arrangement created by the ligation of the 39G...journal.pgen.1001005 (2010). 14 van Kregten, M. et al. T-DNA integration in plants results from polymerase-theta-mediated DNA repair. Nat Plants 2

  20. KIN17, XPC, DNA-PKCS and XRCC4 proteins in the cellular response to DNA damages. Relations between nucleotide excision repair and non-homologous end joining in a human syn-genic model

    International Nuclear Information System (INIS)

    Despras, Emmanuelle

    2006-01-01

    The response to genotoxic stress involves many cellular factors in a complex network of mechanisms that aim to preserve the genetic integrity of the organism. These mechanisms enclose the detection and repair of DNA lesions, the regulation of transcription and replication and, eventually, the setting of cell death. Among the nuclear proteins involved in this response, kin17 proteins are zinc-finger proteins conserved through evolution and activated by ultraviolet (UV) or ionizing radiations (IR). We showed that human kin17 protein (HSAkin17) is found in the cell under a soluble form and a form tightly anchored to nuclear structures. A fraction of HSAkin17 protein is directly associated with chromatin. HSAkin17 protein is recruited to nuclear structures 24 hours after treatment with various agents inducing DNA double-strand breaks (DSB) and/or replication forks blockage. Moreover, the reduction of total HSAkin17 protein level sensitizes RKO cells to IR. We also present evidence for the involvement of HSAkin17 protein in DNA replication. This hypothesis was further confirmed by the biochemical demonstration of its belonging to the replication complex. HSAkin17 protein could link DNA replication and DNA repair, a defect in the HSAkin17 pathway leading to an increased radiosensitivity. In a second part, we studied the interactions between two DNA repair mechanisms: nucleotide excision repair (NER) and non-homologous end joining (NHEJ). NER repairs a wide variety of lesions inducing a distortion of the DNA double helix including UV-induced pyrimidine dimers. NHEJ allows the repair of DSB by direct joining of DNA ends. We used a syn-genic model for DNA repair defects based on RNA interference developed in the laboratory. Epstein-Barr virus-derived vectors (pEBV) allow long-term expression of siRNA and specific extinction of the targeted gene. The reduction of the expression of genes involved in NER (XPA and XPC) or NHEJ (DNA-PKcs and XRCC4) leads to the expected

  1. Relative contribution of homologous recombination and non-homologous end-joining to DNA double-strand break repair after oxidative stress in Saccharomyces cerevisiae.

    Science.gov (United States)

    Letavayová, Lucia; Marková, Eva; Hermanská, Katarína; Vlcková, Viera; Vlasáková, Danusa; Chovanec, Miroslav; Brozmanová, Jela

    2006-05-10

    Oxidative damage to DNA seems to be an important factor in developing many human diseases including cancer. It involves base and sugar damage, base-free sites, DNA-protein cross-links and DNA single-strand (SSB) and double-strand (DSB) breaks. Oxidative DSB can be formed in various ways such as their direct induction by the drug or their generation either through attempted and aborted repair of primary DNA lesions or through DNA replication-dependent conversion of SSB. In general, two main pathways are responsible for repairing DSB, homologous recombination (HR) and non-homologous end-joining (NHEJ), with both of them being potential candidates for the repair of oxidative DSB. We have examined relative contribution of HR and NHEJ to cellular response after oxidative stress in Saccharomyces cerevisiae. Therefore, cell survival, mutagenesis and DSB induction and repair in the rad52, yku70 and rad52 yku70 mutants after hydrogen peroxide (H(2)O(2)), menadione (MD) or bleomycin (BLM) exposure were compared to those obtained for the corresponding wild type. We show that MD exposure does not lead to observable DSB induction in yeast, suggesting that the toxic effects of this agent are mediated by other types of DNA damage. Although H(2)O(2) treatment generates some DSB, their yield is relatively low and hence DSB may only partially be responsible for toxicity of H(2)O(2), particularly at high doses of the agent. On the other hand, the basis of the BLM toxicity resides primarily in DSB induction. Both HR and NHEJ act on BLM-induced DSB, although their relative participation in the process is not equal. Based on our results we suggest that the complexity and/or the quality of the BLM-induced DSB might represent an obstacle for the NHEJ pathway.

  2. Cadmium delays non-homologous end joining (NHEJ) repair via inhibition of DNA-PKcs phosphorylation and downregulation of XRCC4 and Ligase IV

    Energy Technology Data Exchange (ETDEWEB)

    Li, Weiwei; Gu, Xueyan; Zhang, Xiaoning; Kong, Jinxin [Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, School of Life Sciences, Lanzhou University, Lanzhou 730000 (China); Ding, Nan [Gansu Key laboratory of Space Radiobiology, Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Qi, Yongmei; Zhang, Yingmei [Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, School of Life Sciences, Lanzhou University, Lanzhou 730000 (China); Wang, Jufang [Gansu Key laboratory of Space Radiobiology, Institute of Modern Physics, Chinese Academy of Sciences, Lanzhou 730000 (China); Huang, Dejun, E-mail: huangdj@lzu.edu.cn [Gansu Key Laboratory of Biomonitoring and Bioremediation for Environmental Pollution, School of Life Sciences, Lanzhou University, Lanzhou 730000 (China)

    2015-09-15

    Highlights: • Cadmium (Cd) exposure delayed the repair of DNA damage induced by X-ray. • Cd exposure altered the phosphorylation of DNA-PKcs on Thr-2609 and Ser-2056 sites. • Cd impaired the formation of XRCC4 and Ligase IV foci, and down-regulated their protein expression. • Zinc mitigated the effects of Cd on DDR by regulating pDNA-PKcs (Thr-2609), XRCC4 and Ligase IV. - Abstract: Although studies have shown that cadmium (Cd) interfered with DNA damage repair (DDR), whether Cd could affect non-homologous end joining (NHEJ) repair remains elusive. To further understand the effect of Cd on DDR, we used X-ray irradiation of Hela cells as an in vitro model system, along with γH2AX and 53BP1 as markers for DNA damage. Results showed that X-ray significantly increased γH2AX and 53BP1 foci in Hela cells (p < 0.01), all of which are characteristic of accrued DNA damage. The number of foci declined rapidly over time (1–8 h postirradiation), indicating an initiation of NHEJ process. However, the disappearance of γH2AX and 53BP1 foci was remarkably slowed by Cd pretreatment (p < 0.01), suggesting that Cd reduced the efficiency of NHEJ. To further elucidate the mechanisms of Cd toxicity, several markers of NHEJ pathway including Ku70, DNA-PKcs, XRCC4 and Ligase IV were examined. Our data showed that Cd altered the phosphorylation of DNA-PKcs, and reduced the expression of both XRCC4 and Ligase IV in irradiated cells. These observations are indicative of the impairment of NHEJ-dependent DNA repair pathways. In addition, zinc (Zn) mitigated the effects of Cd on NHEJ, suggesting that the Cd-induced NHEJ alteration may partly result from the displacement of Zn or from an interference with the normal function of Zn-containing proteins by Cd. Our findings provide a new insight into the toxicity of Cd on NHEJ repair and its underlying mechanisms in human cells.

  3. Analysis of DNA double-strand break repair pathways in mice

    International Nuclear Information System (INIS)

    Brugmans, Linda; Kanaar, Roland; Essers, Jeroen

    2007-01-01

    During the last years significant new insights have been gained into the mechanism and biological relevance of DNA double-strand break (DSB) repair in relation to genome stability. DSBs are a highly toxic DNA lesion, because they can lead to chromosome fragmentation, loss and translocations, eventually resulting in cancer. DSBs can be induced by cellular processes such as V(D)J recombination or DNA replication. They can also be introduced by exogenous agents DNA damaging agents such as ionizing radiation or mitomycin C. During evolution several pathways have evolved for the repair of these DSBs. The most important DSB repair mechanisms in mammalian cells are nonhomologous end-joining and homologous recombination. By using an undamaged repair template, homologous recombination ensures accurate DSB repair, whereas the untemplated nonhomologous end-joining pathway does not. Although both pathways are active in mammals, the relative contribution of the two repair pathways to genome stability differs in the different cell types. Given the potential differences in repair fidelity, it is of interest to determine the relative contribution of homologous recombination and nonhomologous end-joining to DSB repair. In this review, we focus on the biological relevance of DSB repair in mammalian cells and the potential overlap between nonhomologous end-joining and homologous recombination in different tissues

  4. DNA double-strand break response factors influence end-joining features of IgH class switch and general translocation junctions.

    Science.gov (United States)

    Panchakshari, Rohit A; Zhang, Xuefei; Kumar, Vipul; Du, Zhou; Wei, Pei-Chi; Kao, Jennifer; Dong, Junchao; Alt, Frederick W

    2018-01-23

    Ig heavy chain (IgH) class switch recombination (CSR) in B lymphocytes switches IgH constant regions to change antibody functions. CSR is initiated by DNA double-strand breaks (DSBs) within a donor IgH switch (S) region and a downstream acceptor S region. CSR is completed by fusing donor and acceptor S region DSB ends by classical nonhomologous end-joining (C-NHEJ) and, in its absence, by alternative end-joining that is more biased to use longer junctional microhomologies (MHs). Deficiency for DSB response (DSBR) factors, including ataxia telangiectasia-mutated (ATM) and 53BP1, variably impair CSR end-joining, with 53BP1 deficiency having the greatest impact. However, studies of potential impact of DSBR factor deficiencies on MH-mediated CSR end-joining have been technically limited. We now use a robust DSB joining assay to elucidate impacts of deficiencies for DSBR factors on CSR and chromosomal translocation junctions in primary mouse B cells and CH12F3 B-lymphoma cells. Compared with wild-type, CSR and c-myc to S region translocation junctions in the absence of 53BP1, and, to a lesser extent, other DSBR factors, have increased MH utilization; indeed, 53BP1-deficient MH profiles resemble those associated with C-NHEJ deficiency. However, translocation junctions between c-myc DSB and general DSBs genome-wide are not MH-biased in ATM-deficient versus wild-type CH12F3 cells and are less biased in 53BP1- and C-NHEJ-deficient cells than CSR junctions or c-myc to S region translocation junctions. We discuss potential roles of DSBR factors in suppressing increased MH-mediated DSB end-joining and features of S regions that may render their DSBs prone to MH-biased end-joining in the absence of DSBR factors.

  5. Repair of DNA DSB in higher eukaryotes

    International Nuclear Information System (INIS)

    Wang, H.; Perrault, A.R.; Takeda, Y.; Iliakis, G.

    2003-01-01

    Cells of higher eukaryotes process within minutes double strand breaks (DSBs) in their genome using a NHEJ apparatus that engages DNA-PKcs, Ku, DNA ligase IV, XRCC4, and other as of yet unidentified factors. Although chemical inhibition, or mutation, in any of these factors delays processing, cells ultimately remove the majority of DNA DSBs using an alternative pathway operating with slower kinetics. This alternative pathway is active in mutants deficient in genes of the RAD52 epistasis group. We proposed, therefore, that it reflects an alternative form of NHEJ that operates as a backup (B-NHEJ) to the DNA-PK- dependent (D-NHEJ) pathway, rather than homology directed repair of DSBs. We studied the role of Ku and DNA-PKcs in the coordination of these pathways using as a model end joining of restriction endonuclease linearized plasmid DNA in whole cell extracts. Efficient error-free endjoining observed in such in-vitro reactions is strongly inhibited by anti-Ku antibodies. The inhibition requires DNA-PKcs, despite that fact that Ku efficiently binds DNA ends in the presence of antibodies, or in the absence of DNA-PKcs. Strong inhibition of DNA endjoining is also mediated by wortmannin, an inhibitor of DNA-PKcs, in the presence but not in the absence of Ku, and this inhibition can be rescued by pre-incubating the reaction with double stranded oligonucleotides. The results are compatible with a role of Ku in directing endjoining to a DNA-PK dependent pathway, mediated by efficient end binding and productive interactions with DNA-PKcs. On the other hand, efficient end joining is observed in extracts of cells lacking DNA-PKcs, as well as in Ku-depleted extracts sugggesting the operation of alternative pathways. Extracts depleted of Ku and DNA-PKcs rejoin blunt ends, as well as homologous ends with 3' or 5' protruding single strands with similar efficiency, but addition of Ku suppresses joining of blunt ends and homologous ends with 3' overhangs. We propose that the

  6. Sibling rivalry: competition between Pol X family members in V(D)J recombination and general double strand break repair.

    Science.gov (United States)

    Nick McElhinny, Stephanie A; Ramsden, Dale A

    2004-08-01

    The nonhomologous end-joining pathway is a major means for repairing double-strand breaks (DSBs) in all mitotic cell types. This repair pathway is also the only efficient means for resolving DSB intermediates in V(D)J recombination, a lymphocyte-specific genome rearrangement required for assembly of antigen receptors. A role for polymerases in end-joining has been well established. They are a major factor in determining the character of repair junctions but, in contrast to 'core' end-joining factors, typically appear to have a subtle impact on the efficiency of end-joining. Recent work implicates several members of the Pol X family in end-joining and suggests surprising complexity in the control of how these different polymerases are employed in this pathway.

  7. SV40 Utilizes ATM Kinase Activity to Prevent Non-homologous End Joining of Broken Viral DNA Replication Products

    Science.gov (United States)

    Sowd, Gregory A.; Mody, Dviti; Eggold, Joshua; Cortez, David; Friedman, Katherine L.; Fanning, Ellen

    2014-01-01

    Simian virus 40 (SV40) and cellular DNA replication rely on host ATM and ATR DNA damage signaling kinases to facilitate DNA repair and elicit cell cycle arrest following DNA damage. During SV40 DNA replication, ATM kinase activity prevents concatemerization of the viral genome whereas ATR activity prevents accumulation of aberrant genomes resulting from breakage of a moving replication fork as it converges with a stalled fork. However, the repair pathways that ATM and ATR orchestrate to prevent these aberrant SV40 DNA replication products are unclear. Using two-dimensional gel electrophoresis and Southern blotting, we show that ATR kinase activity, but not DNA-PKcs kinase activity, facilitates some aspects of double strand break (DSB) repair when ATM is inhibited during SV40 infection. To clarify which repair factors associate with viral DNA replication centers, we examined the localization of DSB repair proteins in response to SV40 infection. Under normal conditions, viral replication centers exclusively associate with homology-directed repair (HDR) and do not colocalize with non-homologous end joining (NHEJ) factors. Following ATM inhibition, but not ATR inhibition, activated DNA-PKcs and KU70/80 accumulate at the viral replication centers while CtIP and BLM, proteins that initiate 5′ to 3′ end resection during HDR, become undetectable. Similar to what has been observed during cellular DSB repair in S phase, these data suggest that ATM kinase influences DSB repair pathway choice by preventing the recruitment of NHEJ factors to replicating viral DNA. These data may explain how ATM prevents concatemerization of the viral genome and promotes viral propagation. We suggest that inhibitors of DNA damage signaling and DNA repair could be used during infection to disrupt productive viral DNA replication. PMID:25474690

  8. SV40 utilizes ATM kinase activity to prevent non-homologous end joining of broken viral DNA replication products.

    Directory of Open Access Journals (Sweden)

    Gregory A Sowd

    2014-12-01

    Full Text Available Simian virus 40 (SV40 and cellular DNA replication rely on host ATM and ATR DNA damage signaling kinases to facilitate DNA repair and elicit cell cycle arrest following DNA damage. During SV40 DNA replication, ATM kinase activity prevents concatemerization of the viral genome whereas ATR activity prevents accumulation of aberrant genomes resulting from breakage of a moving replication fork as it converges with a stalled fork. However, the repair pathways that ATM and ATR orchestrate to prevent these aberrant SV40 DNA replication products are unclear. Using two-dimensional gel electrophoresis and Southern blotting, we show that ATR kinase activity, but not DNA-PK(cs kinase activity, facilitates some aspects of double strand break (DSB repair when ATM is inhibited during SV40 infection. To clarify which repair factors associate with viral DNA replication centers, we examined the localization of DSB repair proteins in response to SV40 infection. Under normal conditions, viral replication centers exclusively associate with homology-directed repair (HDR and do not colocalize with non-homologous end joining (NHEJ factors. Following ATM inhibition, but not ATR inhibition, activated DNA-PK(cs and KU70/80 accumulate at the viral replication centers while CtIP and BLM, proteins that initiate 5' to 3' end resection during HDR, become undetectable. Similar to what has been observed during cellular DSB repair in S phase, these data suggest that ATM kinase influences DSB repair pathway choice by preventing the recruitment of NHEJ factors to replicating viral DNA. These data may explain how ATM prevents concatemerization of the viral genome and promotes viral propagation. We suggest that inhibitors of DNA damage signaling and DNA repair could be used during infection to disrupt productive viral DNA replication.

  9. Lithium chloride protects retinal neurocytes from nutrient deprivation by promoting DNA non-homologous end-joining

    International Nuclear Information System (INIS)

    Zhuang Jing; Li Fan; Liu Xuan; Liu Zhiping; Lin Jianxian; Ge Yihong; Kaminski, Joseph M.; Summers, James Bradley; Wang Zhichong; Ge Jian; Yu Keming

    2009-01-01

    Lithium chloride is a therapeutic agent for treatment of bipolar affective disorders. Increasing numbers of studies have indicated that lithium has neuroprotective effects. However, the molecular mechanisms underlying the actions of lithium have not been fully elucidated. This study aimed to investigate whether lithium chloride produces neuroprotective function by improving DNA repair pathway in retinal neurocyte. In vitro, the primary cultured retinal neurocytes (85.7% are MAP-2 positive cells) were treated with lithium chloride, then cultured with serum-free media to simulate the nutrient deprived state resulting from ischemic insult. The neurite outgrowth of the cultured cells increased significantly in a dose-dependent manner when exposed to different levels of lithium chloride. Genomic DNA electrophoresis demonstrated greater DNA integrity of retinal neurocytes when treated with lithium chloride as compared to the control. Moreover, mRNA and protein levels of Ligase IV (involved in DNA non-homologous end-joining (NHEJ) pathway) in retinal neurocytes increased with lithium chloride. The end joining activity assay was performed to determine the role of lithium on NHEJ in the presence of extract from retinal neurocytes. The rejoining levels in retinal neurocytes treated with lithium were significantly increased as compared to the control. Furthermore, XRCC4, the Ligase IV partner, and the transcriptional factor, CREB and CTCF, were up-regulated in retinal cells after treating with 1.0 mM lithium chloride. Therefore, our data suggest that lithium chloride protects the retinal neural cells from nutrient deprivation in vitro, which may be similar to the mechanism of cell death in glaucoma. The improvement in DNA repair pathway involving in Ligase IV might have an important role in lithium neuroprotection. This study provides new insights into the neural protective mechanisms of lithium chloride.

  10. Chromosome End Repair and Genome Stability in Plasmodium falciparum.

    Science.gov (United States)

    Calhoun, Susannah F; Reed, Jake; Alexander, Noah; Mason, Christopher E; Deitsch, Kirk W; Kirkman, Laura A

    2017-08-08

    The human malaria parasite Plasmodium falciparum replicates within circulating red blood cells, where it is subjected to conditions that frequently cause DNA damage. The repair of DNA double-stranded breaks (DSBs) is thought to rely almost exclusively on homologous recombination (HR), due to a lack of efficient nonhomologous end joining. However, given that the parasite is haploid during this stage of its life cycle, the mechanisms involved in maintaining genome stability are poorly understood. Of particular interest are the subtelomeric regions of the chromosomes, which contain the majority of the multicopy variant antigen-encoding genes responsible for virulence and disease severity. Here, we show that parasites utilize a competitive balance between de novo telomere addition, also called "telomere healing," and HR to stabilize chromosome ends. Products of both repair pathways were observed in response to DSBs that occurred spontaneously during routine in vitro culture or resulted from experimentally induced DSBs, demonstrating that both pathways are active in repairing DSBs within subtelomeric regions and that the pathway utilized was determined by the DNA sequences immediately surrounding the break. In combination, these two repair pathways enable parasites to efficiently maintain chromosome stability while also contributing to the generation of genetic diversity. IMPORTANCE Malaria is a major global health threat, causing approximately 430,000 deaths annually. This mosquito-transmitted disease is caused by Plasmodium parasites, with infection with the species Plasmodium falciparum being the most lethal. Mechanisms underlying DNA repair and maintenance of genome integrity in P. falciparum are not well understood and represent a gap in our understanding of how parasites survive the hostile environment of their vertebrate and insect hosts. Our work examines DNA repair in real time by using single-molecule real-time (SMRT) sequencing focused on the subtelomeric

  11. BCR/ABL downregulates DNA-PK(CS)-dependent and upregulates backup non-homologous end joining in leukemic cells.

    Science.gov (United States)

    Poplawski, Tomasz; Blasiak, Janusz

    2010-06-01

    Non-homologous end joining (NHEJ) and homologous recombination repair (HRR) are the main mechanisms involved in the processing of DNA double strand breaks (DSBs) in humans. We showed previously that the oncogenic tyrosine kinase BCR/ABL stimulated DSBs repair by HRR. To evaluate the role of BCR/ABL in DSBs repair by NHEJ we examined the ability of leukemic BCR/ABL-expressing cell line BV173 to repair DNA damage induced by two DNA topoisomerase II inhibitors: etoposide and sobuzoxane. DNA lesions induced by sobuzoxane are repaired by a NHEJ pathway which is dependent on the catalytic subunit of protein kinase dependent on DNA (DNA-PK(CS); D-NHEJ), whereas damage evoked by etoposide are repaired by two distinct NHEJ pathways, dependent on or independent of DNA-PK(CS) (backup NHEJ, B-NHEJ). Cells incubated with STI571, a highly specific inhibitor of BCR/ABL, displayed resistance to these agents associated with an accelerated kinetics of DSBs repair, as measured by the neutral comet assay and pulsed field gel electrophoresis. However, in a functional NHEJ assay, cells preincubated with STI571 repaired DSBs induced by a restriction enzyme with a lower efficacy than without the preincubation and addition of wortmannin, a specific inhibitor of DNA-PK(CS), did not change efficacy of the NHEJ reaction. We suggest that BCR/ABL switch on B-NHEJ which is more error-prone then D-NHEJ and in such manner contribute to the increase of the genomic instability of leukemic cells.

  12. Positive selection on the nonhomologous end-joining factor Cernunnos-XLF in the human lineage

    Directory of Open Access Journals (Sweden)

    Jurka Jerzy

    2006-06-01

    Full Text Available Abstract Background Cernunnos-XLF is a nonhomologous end-joining factor that is mutated in patients with a rare immunodeficiency with microcephaly. Several other microcephaly-associated genes such as ASPM and microcephalin experienced recent adaptive evolution apparently linked to brain size expansion in humans. In this study we investigated whether Cernunnos-XLF experienced similar positive selection during human evolution. Results We obtained or reconstructed full-length coding sequences of chimpanzee, rhesus macaque, canine, and bovine Cernunnos-XLF orthologs from sequence databases and sequence trace archives. Comparison of coding sequences revealed an excess of nonsynonymous substitutions consistent with positive selection on Cernunnos-XLF in the human lineage. The hotspots of adaptive evolution are concentrated around a specific structural domain, whose analogue in the structurally similar XRCC4 protein is involved in binding of another nonhomologous end-joining factor, DNA ligase IV. Conclusion Cernunnos-XLF is a microcephaly-associated locus newly identified to be under adaptive evolution in humans, and possibly played a role in human brain expansion. We speculate that Cernunnos-XLF may have contributed to the increased number of brain cells in humans by efficient double strand break repair, which helps to prevent frequent apoptosis of neuronal progenitors and aids mitotic cell cycle progression. Reviewers This article was reviewed by Chris Ponting and Richard Emes (nominated by Chris Ponting, Kateryna Makova, Gáspár Jékely and Eugene V. Koonin.

  13. Comparative analysis of the end-joining activity of several DNA ligases.

    Directory of Open Access Journals (Sweden)

    Robert J Bauer

    Full Text Available DNA ligases catalyze the repair of phosphate backbone breaks in DNA, acting with highest activity on breaks in one strand of duplex DNA. Some DNA ligases have also been observed to ligate two DNA fragments with short complementary overhangs or blunt-ended termini. In this study, several wild-type DNA ligases (phage T3, T4, and T7 DNA ligases, Paramecium bursaria chlorella virus 1 (PBCV1 DNA ligase, human DNA ligase 3, and Escherichia coli DNA ligase were tested for their ability to ligate DNA fragments with several difficult to ligate end structures (blunt-ended termini, 3'- and 5'- single base overhangs, and 5'-two base overhangs. This analysis revealed that T4 DNA ligase, the most common enzyme utilized for in vitro ligation, had its greatest activity on blunt- and 2-base overhangs, and poorest on 5'-single base overhangs. Other ligases had different substrate specificity: T3 DNA ligase ligated only blunt ends well; PBCV1 DNA ligase joined 3'-single base overhangs and 2-base overhangs effectively with little blunt or 5'- single base overhang activity; and human ligase 3 had highest activity on blunt ends and 5'-single base overhangs. There is no correlation of activity among ligases on blunt DNA ends with their activity on single base overhangs. In addition, DNA binding domains (Sso7d, hLig3 zinc finger, and T4 DNA ligase N-terminal domain were fused to PBCV1 DNA ligase to explore whether modified binding to DNA would lead to greater activity on these difficult to ligate substrates. These engineered ligases showed both an increased binding affinity for DNA and increased activity, but did not alter the relative substrate preferences of PBCV1 DNA ligase, indicating active site structure plays a role in determining substrate preference.

  14. The Ku80 Carboxy Terminus Stimulates Joining and Artemis-Mediated Processing of DNA Ends▿

    Science.gov (United States)

    Weterings, Eric; Verkaik, Nicole S.; Keijzers, Guido; Florea, Bogdan I.; Wang, Shih-Ya; Ortega, Laura G.; Uematsu, Naoya; Chen, David J.; van Gent, Dik C.

    2009-01-01

    Repair of DNA double-strand breaks (DSBs) is predominantly mediated by nonhomologous end joining (NHEJ) in mammalian cells. NHEJ requires binding of the Ku70-Ku80 heterodimer (Ku70/80) to the DNA ends and subsequent recruitment of the DNA-dependent protein kinase catalytic subunit (DNA-PKCS) and the XRCC4/ligase IV complex. Activation of the DNA-PKCS serine/threonine kinase requires an interaction with Ku70/80 and is essential for NHEJ-mediated DSB repair. In contrast to previous models, we found that the carboxy terminus of Ku80 is not absolutely required for the recruitment and activation of DNA-PKCS at DSBs, although cells that harbored a carboxy-terminal deletion in the Ku80 gene were sensitive to ionizing radiation and showed reduced end-joining capacity. More detailed analysis of this repair defect showed that DNA-PKCS autophosphorylation at Thr2647 was diminished, while Ser2056 was phosphorylated to normal levels. This resulted in severely reduced levels of Artemis nuclease activity in vivo and in vitro. We therefore conclude that the Ku80 carboxy terminus is important to support DNA-PKCS autophosphorylation at specific sites, which facilitates DNA end processing by the Artemis endonuclease and the subsequent joining reaction. PMID:19103741

  15. Double-Strand DNA Break Repair in Mycobacteria.

    Science.gov (United States)

    Glickman, Michael S

    2014-10-01

    Discontinuity of both strands of the chromosome is a lethal event in all living organisms because it compromises chromosome replication. As such, a diversity of DNA repair systems has evolved to repair double-strand DNA breaks (DSBs). In part, this diversity of DSB repair systems has evolved to repair breaks that arise in diverse physiologic circumstances or sequence contexts, including cellular states of nonreplication or breaks that arise between repeats. Mycobacteria elaborate a set of three genetically distinct DNA repair pathways: homologous recombination, nonhomologous end joining, and single-strand annealing. As such, mycobacterial DSB repair diverges substantially from the standard model of prokaryotic DSB repair and represents an attractive new model system. In addition, the presence in mycobacteria of a DSB repair system that can repair DSBs in nonreplicating cells (nonhomologous end joining) or when DSBs arise between repeats (single-strand annealing) has clear potential relevance to Mycobacterium tuberculosis pathogenesis, although the exact role of these systems in M. tuberculosis pathogenesis is still being elucidated. In this article we will review the genetics of mycobacterial DSB repair systems, focusing on recent insights.

  16. Silencing of end-joining repair for efficient site-specific gene insertion after TALEN/CRISPR mutagenesis in Aedes aegypti.

    Science.gov (United States)

    Basu, Sanjay; Aryan, Azadeh; Overcash, Justin M; Samuel, Glady Hazitha; Anderson, Michelle A E; Dahlem, Timothy J; Myles, Kevin M; Adelman, Zach N

    2015-03-31

    Conventional control strategies for mosquito-borne pathogens such as malaria and dengue are now being complemented by the development of transgenic mosquito strains reprogrammed to generate beneficial phenotypes such as conditional sterility or pathogen resistance. The widespread success of site-specific nucleases such as transcription activator-like effector nucleases (TALENs) and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 in model organisms also suggests that reprogrammable gene drive systems based on these nucleases may be capable of spreading such beneficial phenotypes in wild mosquito populations. Using the mosquito Aedes aegypti, we determined that mutations in the FokI domain used in TALENs to generate obligate heterodimeric complexes substantially and significantly reduce gene editing rates. We found that CRISPR/Cas9-based editing in the mosquito Ae. aegypti is also highly variable, with the majority of guide RNAs unable to generate detectable editing. By first evaluating candidate guide RNAs using a transient embryo assay, we were able to rapidly identify highly effective guide RNAs; focusing germ line-based experiments only on this cohort resulted in consistently high editing rates of 24-90%. Microinjection of double-stranded RNAs targeting ku70 or lig4, both essential components of the end-joining response, increased recombination-based repair in early embryos as determined by plasmid-based reporters. RNAi-based suppression of Ku70 concurrent with embryonic microinjection of site-specific nucleases yielded consistent gene insertion frequencies of 2-3%, similar to traditional transposon- or ΦC31-based integration methods but without the requirement for an initial docking step. These studies should greatly accelerate investigations into mosquito biology, streamline development of transgenic strains for field releases, and simplify the evaluation of novel Cas9-based gene drive systems.

  17. DNA ligase C1 mediates the LigD-independent nonhomologous end-joining pathway of Mycobacterium smegmatis.

    Science.gov (United States)

    Bhattarai, Hitesh; Gupta, Richa; Glickman, Michael S

    2014-10-01

    Nonhomologous end joining (NHEJ) is a recently described bacterial DNA double-strand break (DSB) repair pathway that has been best characterized for mycobacteria. NHEJ can religate transformed linear plasmids, repair ionizing radiation (IR)-induced DSBs in nonreplicating cells, and seal I-SceI-induced chromosomal DSBs. The core components of the mycobacterial NHEJ machinery are the DNA end binding protein Ku and the polyfunctional DNA ligase LigD. LigD has three autonomous enzymatic modules: ATP-dependent DNA ligase (LIG), DNA/RNA polymerase (POL), and 3' phosphoesterase (PE). Although genetic ablation of ku or ligD abolishes NHEJ and sensitizes nonreplicating cells to ionizing radiation, selective ablation of the ligase activity of LigD in vivo only mildly impairs NHEJ of linearized plasmids, indicating that an additional DNA ligase can support NHEJ. Additionally, the in vivo role of the POL and PE domains in NHEJ is unclear. Here we define a LigD ligase-independent NHEJ pathway in Mycobacterium smegmatis that requires the ATP-dependent DNA ligase LigC1 and the POL domain of LigD. Mycobacterium tuberculosis LigC can also support this backup NHEJ pathway. We also demonstrate that, although dispensable for efficient plasmid NHEJ, the activities of the POL and PE domains are required for repair of IR-induced DSBs in nonreplicating cells. These findings define the genetic requirements for a LigD-independent NHEJ pathway in mycobacteria and demonstrate that all enzymatic functions of the LigD protein participate in NHEJ in vivo. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  18. Sirt6 Promotes DNA End Joining in iPSCs Derived from Old Mice

    Directory of Open Access Journals (Sweden)

    Wen Chen

    2017-03-01

    Full Text Available Induced pluripotent stem cells (iPSCs have great potential for treating age-related diseases, but the genome integrity of iPSCs is critically important. Here, we demonstrate that non-homologous end joining (NHEJ, rather than homologous recombination (HR, is less efficient in iPSCs from old mice than young mice. We further find that Sirt6 is downregulated in iPSCs from old mice. Sirt6 directly binds to Ku80 and facilitates the Ku80/DNA-PKcs interaction, thus promoting DNA-PKcs phosphorylation at residue S2056, leading to efficient NHEJ. Rescue experiments show that introducing a combination of Sirt6 and the Yamanaka factors during reprogramming significantly promotes DNA double-strand break (DSB repair by activating NHEJ in iPSCs derived from old mice. Thus, our study suggests a strategy to improve the quality of iPSCs derived from old donors by activating NHEJ and stabilizing the genome.

  19. Development of versatile non-homologous end joining-based knock-in module for genome editing.

    Science.gov (United States)

    Sawatsubashi, Shun; Joko, Yudai; Fukumoto, Seiji; Matsumoto, Toshio; Sugano, Shigeo S

    2018-01-12

    CRISPR/Cas9-based genome editing has dramatically accelerated genome engineering. An important aspect of genome engineering is efficient knock-in technology. For improved knock-in efficiency, the non-homologous end joining (NHEJ) repair pathway has been used over the homology-dependent repair pathway, but there remains a need to reduce the complexity of the preparation of donor vectors. We developed the versatile NHEJ-based knock-in module for genome editing (VIKING). Using the consensus sequence of the time-honored pUC vector to cut donor vectors, any vector with a pUC backbone could be used as the donor vector without customization. Conditions required to minimize random integration rates of the donor vector were also investigated. We attempted to isolate null lines of the VDR gene in human HaCaT keratinocytes using knock-in/knock-out with a selection marker cassette, and found 75% of clones isolated were successfully knocked-in. Although HaCaT cells have hypotetraploid genome composition, the results suggest multiple clones have VDR null phenotypes. VIKING modules enabled highly efficient knock-in of any vectors harboring pUC vectors. Users now can insert various existing vectors into an arbitrary locus in the genome. VIKING will contribute to low-cost genome engineering.

  20. Synthetic lethality between murine DNA repair factors XLF and DNA-PKcs is rescued by inactivation of Ku70

    DEFF Research Database (Denmark)

    Xing, Mengtan; Bjørås, Magnar; Daniel, Jeremy A

    2017-01-01

    DNA double-strand breaks (DSBs) are recognized and repaired by the Classical Non-Homologous End-Joining (C-NHEJ) and Homologous Recombination pathways. C-NHEJ includes the core Ku70 and Ku80 (or Ku86) heterodimer that binds DSBs and thus promotes recruitment of accessory downstream NHEJ factors XLF......, PAXX, DNA-PKcs, Artemis and other core subunits, XRCC4 and DNA Ligase 4 (Lig4). In the absence of core C-NHEJ factors, DNA repair can be performed by Alternative End-Joining, which likely depends on DNA Ligase 1 and DNA Ligase 3. Genetic inactivation of C-NHEJ factors, such as Ku70, Ku80, XLF, PAXX...... with severe apoptosis in the central nervous system. Here, we demonstrate that inactivation of the Ku70 gene rescues the synthetic lethality between XLF and DNA-PKcs, resulting in triple knockout mice that are indistinguishable from Ku70-deficient littermates by size or levels of genomic instability. Moreover...

  1. End-joining inhibition at telomeres requires the translocase and polySUMO-dependent ubiquitin ligase Uls1.

    Science.gov (United States)

    Lescasse, Rachel; Pobiega, Sabrina; Callebaut, Isabelle; Marcand, Stéphane

    2013-03-20

    In eukaryotes, permanent inhibition of the non-homologous end joining (NHEJ) repair pathway at telomeres ensures that chromosome ends do not fuse. In budding yeast, binding of Rap1 to telomere repeats establishes NHEJ inhibition. Here, we show that the Uls1 protein is required for the maintenance of NHEJ inhibition at telomeres. Uls1 protein is a non-essential Swi2/Snf2-related translocase and a Small Ubiquitin-related Modifier (SUMO)-Targeted Ubiquitin Ligase (STUbL) with unknown targets. Loss of Uls1 results in telomere-telomere fusions. Uls1 requirement is alleviated by the absence of poly-SUMO chains and by rap1 alleles lacking SUMOylation sites. Furthermore, Uls1 limits the accumulation of Rap1 poly-SUMO conjugates. We propose that one of Uls1 functions is to clear non-functional poly-SUMOylated Rap1 molecules from telomeres to ensure the continuous efficiency of NHEJ inhibition. Since Uls1 is the only known STUbL with a translocase activity, it can be the general molecular sweeper for the clearance of poly-SUMOylated proteins on DNA in eukaryotes.

  2. AAV-Mediated Gene Targeting Is Significantly Enhanced by Transient Inhibition of Nonhomologous End Joining or the Proteasome In Vivo

    Science.gov (United States)

    Paulk, Nicole K.; Loza, Laura Marquez; Finegold, Milton J.

    2012-01-01

    Abstract Recombinant adeno-associated virus (rAAV) vectors have clear potential for use in gene targeting but low correction efficiencies remain the primary drawback. One approach to enhancing efficiency is a block of undesired repair pathways like nonhomologous end joining (NHEJ) to promote the use of homologous recombination. The natural product vanillin acts as a potent inhibitor of NHEJ by inhibiting DNA-dependent protein kinase (DNA-PK). Using a homology containing rAAV vector, we previously demonstrated in vivo gene repair frequencies of up to 0.1% in a model of liver disease hereditary tyrosinemia type I. To increase targeting frequencies, we administered vanillin in combination with rAAV. Gene targeting frequencies increased up to 10-fold over AAV alone, approaching 1%. Fah−/−Ku70−/− double knockout mice also had increased gene repair frequencies, genetically confirming the beneficial effects of blocking NHEJ. A second strategy, transient proteasomal inhibition, also increased gene-targeting frequencies but was not additive to NHEJ inhibition. This study establishes the benefit of transient NHEJ inhibition with vanillin, or proteasome blockage with bortezomib, for increasing hepatic gene targeting with rAAV. Functional metabolic correction of a clinically relevant disease model was demonstrated and provided evidence for the feasibility of gene targeting as a therapeutic strategy. PMID:22486314

  3. Garcinol, a Histone Acetyltransferase Inhibitor, Radiosensitizes Cancer Cells by Inhibiting Non-Homologous End Joining

    Energy Technology Data Exchange (ETDEWEB)

    Oike, Takahiro [Division of Multistep Carcinogenesis, National Cancer Center Research Institute, Chuo-ku, Tokyo (Japan); Division of Genome Biology, National Cancer Center Research Institute, Chuo-ku, Tokyo (Japan); Department of Radiation Oncology, Gunma University Graduate School of Medicine, Maebashi, Gunma (Japan); Ogiwara, Hideaki [Division of Genome Biology, National Cancer Center Research Institute, Chuo-ku, Tokyo (Japan); Torikai, Kohta [Gunma University Heavy Ion Medical Center, Maebashi, Gunma (Japan); Nakano, Takashi [Department of Radiation Oncology, Gunma University Graduate School of Medicine, Maebashi, Gunma (Japan); Yokota, Jun [Division of Multistep Carcinogenesis, National Cancer Center Research Institute, Chuo-ku, Tokyo (Japan); Kohno, Takashi, E-mail: tkkohno@ncc.go.jp [Division of Genome Biology, National Cancer Center Research Institute, Chuo-ku, Tokyo (Japan)

    2012-11-01

    Purpose: Non-homologous end joining (NHEJ), a major pathway used to repair DNA double-strand breaks (DSBs) generated by ionizing radiation (IR), requires chromatin remodeling at DSB sites through the acetylation of histones by histone acetyltransferases (HATs). However, the effect of compounds with HAT inhibitory activities on the DNA damage response (DDR), including the NHEJ and cell cycle checkpoint, as well as on the radiosensitivity of cancer cells, remains largely unclear. Here, we investigated whether garcinol, a HAT inhibitor found in the rinds of Garcinia indica fruit (called mangosteens), has effects on DDR, and whether it can be used for radiosensitization. Methods and Materials: The following assays were used to examine the effect of garcinol on the inhibition of DSB repair, including the following: a conventional neutral comet assay; a cell-based assay recently developed by us, in which NHEJ repair of DSBs on chromosomal DNA was evaluated; the micrococcal nuclease sensitivity assay; and immunoblotting for autophosphorylation of DNA-dependent protein kinase catalytic subunit (DNA-PKcs). We assessed the effect of garcinol on the cell cycle checkpoint after IR treatment by analyzing the phosphorylation levels of checkpoint kinases CHK1 and CHK2 and histone H3, and by cell cycle profile analysis using flow cytometry. The radiosensitizing effect of garcinol was assessed by a clonogenic survival assay, whereas its effects on apoptosis and senescence were examined by annexin V and senescence-associated {beta}-galactosidase (SA-{beta}-Gal) staining, respectively. Results: We found that garcinol inhibits DSB repair, including NHEJ, without affecting cell cycle checkpoint. Garcinol radiosensitized A549 lung and HeLa cervical carcinoma cells with dose enhancement ratios (at 10% surviving fraction) of 1.6 and 1.5, respectively. Cellular senescence induced by IR was enhanced by garcinol. Conclusion: These results suggest that garcinol is a radiosensitizer that

  4. Xrcc1-dependent and Ku-dependent DNA double-strand break repair kinetics in Arabidopsis plants.

    Science.gov (United States)

    Charbonnel, Cyril; Gallego, Maria E; White, Charles I

    2010-10-01

    Double-strand breakage (DSB) of DNA involves loss of information on the two strands of the DNA fibre and thus cannot be repaired by simple copying of the complementary strand which is possible with single-strand DNA damage. Homologous recombination (HR) can precisely repair DSB using another copy of the genome as template and non-homologous recombination (NHR) permits repair of DSB with little or no dependence on DNA sequence homology. In addition to the well-characterised Ku-dependent non-homologous end-joining (NHEJ) pathway, much recent attention has been focused on Ku-independent NHR. The complex interrelationships and regulation of NHR pathways remain poorly understood, even more so in the case of plants, and we present here an analysis of Ku-dependent and Ku-independent repair of DSB in Arabidopsis thaliana. We have characterised an Arabidopsis xrcc1 mutant and developed quantitative analysis of the kinetics of appearance and loss of γ-H2AX foci as a tool to measure DSB repair in dividing root tip cells of γ-irradiated plants in vivo. This approach has permitted determination of DSB repair kinetics in planta following a short pulse of γ-irradiation, establishing the existence of a Ku-independent, Xrcc1-dependent DSB repair pathway. Furthermore, our data show a role for Ku80 during the first minutes post-irradiation and that Xrcc1 also plays such a role, but only in the absence of Ku. The importance of Xrcc1 is, however, clearly visible at later times in the presence of Ku, showing that alternative end-joining plays an important role in DSB repair even in the presence of active NHEJ. © 2010 The Authors. Journal compilation © 2010 Blackwell Publishing Ltd.

  5. Using Negotiated Joining to Construct and Fill Open-ended Roles in Elite Culinary Groups.

    Science.gov (United States)

    Tan, Vaughn

    2015-03-01

    This qualitative study examines membership processes in groups operating in an uncertain environment that prevents them from fully predefining new members' roles. I describe how nine elite high-end, cutting-edge culinary groups in the U.S. and Europe, ranging from innovative restaurants to culinary R&D groups, use negotiated joining-a previously undocumented process-to systematically construct and fill these emergent, open-ended roles. I show that negotiated joining is a consistently patterned, iterative process that begins with a role that both aspirant and target group explicitly understand to be provisional. This provisional role is then jointly modified and constructed by the aspirant and target group through repeated iterations of proposition, validation through trial and evaluation, and selective integration of validated role components. The initially provisional role stabilizes and the aspirant achieves membership if enough role components are validated; otherwise the negotiated joining process is abandoned. Negotiated joining allows the aspirant and target group to learn if a mutually desirable role is likely and, if so, to construct such a role. In addition, the provisional roles in negotiated joining can support absorptive capacity by allowing novel role components to enter target groups through aspirants' efforts to construct stable roles for themselves, while the internal adjustment involved in integrating newly validated role components can have the unintended side effect of supporting adaptation by providing opportunities for the groups to use these novel role components to modify their role structure and goals to suit a changing and uncertain environment. Negotiated joining thus reveals role ambiguity's hitherto unexamined beneficial consequences and provides a foundation for a contingency theory of new-member acquisition.

  6. DNA-dependent protein kinase in nonhomologous end joining: a lock with multiple keys?

    Science.gov (United States)

    Weterings, Eric; Chen, David J

    2007-10-22

    The DNA-dependent protein kinase (DNA-PK) is one of the central enzymes involved in DNA double-strand break (DSB) repair. It facilitates proper alignment of the two ends of the broken DNA molecule and coordinates access of other factors to the repair complex. We discuss the latest findings on DNA-PK phosphorylation and offer a working model for the regulation of DNA-PK during DSB repair.

  7. DNA apoptosis and stability in B-cell chronic lymphoid leukaemia: implication of the DNA double-strand breaks repair system by non homologous recombination

    International Nuclear Information System (INIS)

    Deriano, L.

    2005-01-01

    After an introduction presenting the diagnosis and treatment of chronic lymphoid leukaemia, its molecular and genetic characteristics, and its cellular origin and clonal evolution, this research thesis describes the apoptosis (definition and characteristics, cancer and chemotherapy, apoptotic ways induced by gamma irradiation), the genotoxic stresses, the different repair mechanisms for different damages, and the DNA repair processes. It reports how human chronic lymphocytic leukaemia B cells can escape DNA damage-induced apoptosis through the non-homologous end-joining DNA repair pathway, and presents non-homologous end-joining DNA repair as a potent mutagenic process in human chronic lymphocytic leukaemia B cells

  8. Double-strand break repair-adox: Restoration of suppressed double-strand break repair during mitosis induces genomic instability.

    Science.gov (United States)

    Terasawa, Masahiro; Shinohara, Akira; Shinohara, Miki

    2014-12-01

    Double-strand breaks (DSBs) are one of the severest types of DNA damage. Unrepaired DSBs easily induce cell death and chromosome aberrations. To maintain genomic stability, cells have checkpoint and DSB repair systems to respond to DNA damage throughout most of the cell cycle. The failure of this process often results in apoptosis or genomic instability, such as aneuploidy, deletion, or translocation. Therefore, DSB repair is essential for maintenance of genomic stability. During mitosis, however, cells seem to suppress the DNA damage response and proceed to the next G1 phase, even if there are unrepaired DSBs. The biological significance of this suppression is not known. In this review, we summarize recent studies of mitotic DSB repair and discuss the mechanisms of suppression of DSB repair during mitosis. DSB repair, which maintains genomic integrity in other phases of the cell cycle, is rather toxic to cells during mitosis, often resulting in chromosome missegregation and aberration. Cells have multiple safeguards to prevent genomic instability during mitosis: inhibition of 53BP1 or BRCA1 localization to DSB sites, which is important to promote non-homologous end joining or homologous recombination, respectively, and also modulation of the non-homologous end joining core complex to inhibit DSB repair. We discuss how DSBs during mitosis are toxic and the multiple safeguard systems that suppress genomic instability. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

  9. De novo CNV formation in mouse embryonic stem cells occurs in the absence of Xrcc4-dependent nonhomologous end joining.

    Directory of Open Access Journals (Sweden)

    Martin F Arlt

    2012-09-01

    Full Text Available Spontaneous copy number variant (CNV mutations are an important factor in genomic structural variation, genomic disorders, and cancer. A major class of CNVs, termed nonrecurrent CNVs, is thought to arise by nonhomologous DNA repair mechanisms due to the presence of short microhomologies, blunt ends, or short insertions at junctions of normal and de novo pathogenic CNVs, features recapitulated in experimental systems in which CNVs are induced by exogenous replication stress. To test whether the canonical nonhomologous end joining (NHEJ pathway of double-strand break (DSB repair is involved in the formation of this class of CNVs, chromosome integrity was monitored in NHEJ-deficient Xrcc4(-/- mouse embryonic stem (ES cells following treatment with low doses of aphidicolin, a DNA replicative polymerase inhibitor. Mouse ES cells exhibited replication stress-induced CNV formation in the same manner as human fibroblasts, including the existence of syntenic hotspot regions, such as in the Auts2 and Wwox loci. The frequency and location of spontaneous and aphidicolin-induced CNV formation were not altered by loss of Xrcc4, as would be expected if canonical NHEJ were the predominant pathway of CNV formation. Moreover, de novo CNV junctions displayed a typical pattern of microhomology and blunt end use that did not change in the absence of Xrcc4. A number of complex CNVs were detected in both wild-type and Xrcc4(-/- cells, including an example of a catastrophic, chromothripsis event. These results establish that nonrecurrent CNVs can be, and frequently are, formed by mechanisms other than Xrcc4-dependent NHEJ.

  10. Non-homologous end joining pathway is the major route of protection against 4β-hydroxywithanolide E-induced DNA damage in MCF-7 cells.

    Science.gov (United States)

    You, B-J; Wu, Y-C; Lee, C-L; Lee, H-Z

    2014-03-01

    4β-Hydroxywithanolide E is a bioactive withanolide extracted from Physalis peruviana. 4β-Hydroxywithanolide E caused reactive oxygen species production and cell apoptosis in human breast cancer MCF-7 cells. We further found that 4β-hydroxywithanolide E induced DNA damage and regulated the DNA damage signaling in MCF-7 cells. The DNA damage sensors and repair proteins act promptly to remove DNA lesions by 4β-hydroxywithanolide E. The ataxia-telangiectasia mutated protein (ATM)-dependent DNA damage signaling pathway is involved in 4β-hydroxywithanolide E-induced apoptosis of MCF-7 cells. Non-homologous end joining pathway, but not homologous recombination, is the major route of protection of MCF-7 cells against 4β-hydroxywithanolide E-induced DNA damage. 4β-Hydroxywithanolide E had no significant impact on the base excision repair pathway. In this study, we examined the 4β-hydroxywithanolide E-induced DNA damage as a research tool in project investigating the DNA repair signaling in breast cancer cells. We also suggest that 4β-hydroxywithanolide E assert its anti-tumor activity in carcinogenic progression and develop into a dietary chemopreventive agent. Copyright © 2014 Elsevier Ltd. All rights reserved.

  11. Molecular mechanism of protein assembly on DNA double-strand breaks in the non-homologous end-joining pathway

    International Nuclear Information System (INIS)

    Yano, Ken-ichi; Morotomi-Yano, Keiko; Adachi, Noritaka; Akiyama, Hidenori

    2009-01-01

    Non-homologous end-joining (NHEJ) is the major repair pathway for DNA double-strand breaks (DSBs) in mammalian species. Upon DSB induction, a living cell quickly activates the NHEJ pathway comprising of multiple molecular events. However, it has been difficult to analyze the initial phase of DSB responses in living cells, primarily due to technical limitations. Recent advances in real-time imaging and site-directed DSB induction using laser microbeam allow us to monitor the spatiotemporal dynamics of NHEJ factors in the immediate-early phase after DSB induction. These new approaches, together with the use of cell lines deficient in each essential NHEJ factor, provide novel mechanistic insights into DSB recognition and protein assembly on DSBs in the NHEJ pathway. In this review, we provide an overview of recent progresses in the imaging analyses of the NHEJ core factors. These studies strongly suggest that the NHEJ core factors are pre-assembled into a large complex on DSBs prior to the progression of the biochemical reactions in the NHEJ pathway. Instead of the traditional step-by-step assembly model from the static view of NHEJ, a novel model for dynamic protein assembly in the NHEJ pathway is proposed. This new model provides important mechanistic insights into the protein assembly at DSBs and the regulation of DSB repair. (author)

  12. Human longevity and variation in DNA damage response and repair

    DEFF Research Database (Denmark)

    Debrabant, Birgit; Soerensen, Mette; Flachsbart, Friederike

    2014-01-01

    others. Data were applied on 592 SNPs from 77 genes involved in nine sub-processes: DNA-damage response, base excision repair (BER), nucleotide excision repair, mismatch repair, non-homologous end-joining, homologous recombinational repair (HRR), RecQ helicase activities (RECQ), telomere functioning...... in genotyping procedures and investigated SNPs, potentially inducing differences in the coverage of gene regions. Specifically, five genes were not covered at all in the German data. Therefore, investigations in additional study populations are needed before final conclusion can be drawn....

  13. A robust network of double-strand break repair pathways governs genome integrity during C. elegans development.

    NARCIS (Netherlands)

    Pontier, D.B.; Tijsterman, M.

    2009-01-01

    To preserve genomic integrity, various mechanisms have evolved to repair DNA double-strand breaks (DSBs). Depending on cell type or cell cycle phase, DSBs can be repaired error-free, by homologous recombination, or with concomitant loss of sequence information, via nonhomologous end-joining (NHEJ)

  14. Agrobacterium tumefaciens T-DNA Integration and Gene Targeting in Arabidopsis thaliana Non-Homologous End-Joining Mutants

    Directory of Open Access Journals (Sweden)

    Qi Jia

    2012-01-01

    Full Text Available In order to study the role of AtKu70 and AtKu80 in Agrobacterium-mediated transformation and gene targeting, plant lines with a T-DNA insertion in AtKu80 or AtKu70 genes were functionally characterized. Such plant lines lacked both subunits, indicating that heterodimer formation between AtKu70 and AtKu80 is needed for the stability of the proteins. Homozygous mutants were phenotypically indistinguishable from wild-type plants and were fertile. However, they were hypersensitive to the genotoxic agent bleomycin, resulting in more DSBs as quantified in comet assays. They had lower end-joining efficiency, suggesting that NHEJ is a critical pathway for DSB repair in plants. Both Atku mutants and a previously isolated Atmre11 mutant were impaired in Agrobacterium T-DNA integration via floral dip transformation, indicating that AtKu70, AtKu80, and AtMre11 play an important role in T-DNA integration in Arabidopsis. The frequency of gene targeting was not significantly increased in the Atku80 and Atku70 mutants, but it was increased at least 10-fold in the Atmre11 mutant compared with the wild type.

  15. Non-homologous end joining dependency of {gamma}-irradiation-induced adaptive frameshift mutation formation in cell cycle-arrested yeast cells

    Energy Technology Data Exchange (ETDEWEB)

    Heidenreich, Erich [Institute of Cancer Research, Division of Molecular Genetics, Medical University of Vienna, Borschkegasse 8a, A-1090 Vienna (Austria)]. E-mail: erich.heidenreich@meduniwien.ac.at; Eisler, Herfried [Institute of Cancer Research, Division of Molecular Genetics, Medical University of Vienna, Borschkegasse 8a, A-1090 Vienna (Austria)

    2004-11-22

    There is a strong selective pressure favoring adaptive mutations which relieve proliferation-limiting adverse living conditions. Due to their importance for evolution and pathogenesis, we are interested in the mechanisms responsible for the formation of such adaptive, gain-of-fitness mutations in stationary-phase cells. During previous studies on the occurrence of spontaneous reversions of an auxotrophy-causing frameshift allele in the yeast Saccharomyces cerevisiae, we noticed that about 50% of the adaptive reversions depended on a functional non-homologous end joining (NHEJ) pathway of DNA double-strand break (DSB) repair. Here, we show that the occasional NHEJ component Pol4, which is the yeast ortholog of mammalian DNA polymerase lambda, is not required for adaptive mutagenesis. An artificially imposed excess of DSBs by {gamma}-irradiation resulted in a dramatic increase in the incidence of adaptive, cell cycle arrest-releasing frameshift reversions. By the use of DNA ligase IV-deficient strains we detected that the majority of the {gamma}-induced adaptive mutations were also dependent on a functional NHEJ pathway. This suggests that the same mutagenic NHEJ mechanism acts on spontaneously arising as well as on ionizing radiation-induced DSBs. Inaccuracy of the NHEJ repair pathway may extensively contribute to the incidence of frameshift mutations in resting (non-dividing) eukaryotic cells, and thus act as a driving force in tumor development.

  16. Non-homologous end joining dependency of γ-irradiation-induced adaptive frameshift mutation formation in cell cycle-arrested yeast cells

    International Nuclear Information System (INIS)

    Heidenreich, Erich; Eisler, Herfried

    2004-01-01

    There is a strong selective pressure favoring adaptive mutations which relieve proliferation-limiting adverse living conditions. Due to their importance for evolution and pathogenesis, we are interested in the mechanisms responsible for the formation of such adaptive, gain-of-fitness mutations in stationary-phase cells. During previous studies on the occurrence of spontaneous reversions of an auxotrophy-causing frameshift allele in the yeast Saccharomyces cerevisiae, we noticed that about 50% of the adaptive reversions depended on a functional non-homologous end joining (NHEJ) pathway of DNA double-strand break (DSB) repair. Here, we show that the occasional NHEJ component Pol4, which is the yeast ortholog of mammalian DNA polymerase lambda, is not required for adaptive mutagenesis. An artificially imposed excess of DSBs by γ-irradiation resulted in a dramatic increase in the incidence of adaptive, cell cycle arrest-releasing frameshift reversions. By the use of DNA ligase IV-deficient strains we detected that the majority of the γ-induced adaptive mutations were also dependent on a functional NHEJ pathway. This suggests that the same mutagenic NHEJ mechanism acts on spontaneously arising as well as on ionizing radiation-induced DSBs. Inaccuracy of the NHEJ repair pathway may extensively contribute to the incidence of frameshift mutations in resting (non-dividing) eukaryotic cells, and thus act as a driving force in tumor development

  17. Mouse embryonic stem cells, but not somatic cells, predominantly use homologous recombination to repair double-strand DNA breaks.

    Science.gov (United States)

    Tichy, Elisia D; Pillai, Resmi; Deng, Li; Liang, Li; Tischfield, Jay; Schwemberger, Sandy J; Babcock, George F; Stambrook, Peter J

    2010-11-01

    Embryonic stem (ES) cells give rise to all cell types of an organism. Since mutations at this embryonic stage would affect all cells and be detrimental to the overall health of an organism, robust mechanisms must exist to ensure that genomic integrity is maintained. To test this proposition, we compared the capacity of murine ES cells to repair DNA double-strand breaks with that of differentiated cells. Of the 2 major pathways that repair double-strand breaks, error-prone nonhomologous end joining (NHEJ) predominated in mouse embryonic fibroblasts, whereas the high fidelity homologous recombinational repair (HRR) predominated in ES cells. Microhomology-mediated end joining, an emerging repair pathway, persisted at low levels in all cell types examined. The levels of proteins involved in HRR and microhomology-mediated end joining were highly elevated in ES cells compared with mouse embryonic fibroblasts, whereas those for NHEJ were quite variable, with DNA Ligase IV expression low in ES cells. The half-life of DNA Ligase IV protein was also low in ES cells. Attempts to increase the abundance of DNA Ligase IV protein by overexpression or inhibition of its degradation, and thereby elevate NHEJ in ES cells, were unsuccessful. When ES cells were induced to differentiate, however, the level of DNA Ligase IV protein increased, as did the capacity to repair by NHEJ. The data suggest that preferential use of HRR rather than NHEJ may lend ES cells an additional layer of genomic protection and that the limited levels of DNA Ligase IV may account for the low level of NHEJ activity.

  18. Current topics in DNA double-strand break repair

    International Nuclear Information System (INIS)

    Kobayashi, Junya; Takata, Minoru; Iwabuchi, Kuniyoshi; Miyagawa, Kiyoshi; Sonoda, Eiichiro; Suzuki, Keiji; Tauchi, Hiroshi

    2008-01-01

    DNA double strand break (DSB) is one of the most critical types of damage which is induced by ionizing radiation. In this review, we summarize current progress in investigations on the function of DSB repair-related proteins. We focused on recent findings in the analysis of the function of proteins such as 53BP1, histone H2AX, Mus81-Eme1, Fanc complex, and UBC13, which are found to be related to homologous recombination repair or to non-homologous end joining. In addition to the function of these proteins in DSB repair, the biological function of nuclear foci formation following DSB induction is discussed. (author)

  19. Scarf Repair of Composite Laminates

    Directory of Open Access Journals (Sweden)

    Xie Zonghong

    2016-01-01

    Full Text Available The use of composite materials, such as carbon-fiber reinforced plastic (CFRP composites, aero-structures has led to an increased need of advanced assembly joining and repair technologies. Adhesive bonded repairs as an alternative to recover full or part of initial strength were investigated. Tests were conducted with the objective of evaluating the effectiveness of techniques used for repairing damage fiber reinforced laminated composites. Failure loads and failure modes were generated and compared with the following parameters: scarf angles, roughness of grind tool and number of external plies. Results showed that scarf angle was the critical parameter and the largest tensile strength was observed with the smallest scarf angle. Besides, the use of external plies at the outer surface could not increase the repairs efficiency for large scarf angle. Preparing the repair surfaces by sanding them with a sander ranging from 60 to 100 grit number had significant effect on the failure load. These results allowed the proposal of design principles for repairing CFRP structures.

  20. Mycobacterial UvrD1 is a Ku-dependent DNA helicase that plays a role in multiple DNA repair events, including double-strand break repair.

    Science.gov (United States)

    Sinha, Krishna Murari; Stephanou, Nicolas C; Gao, Feng; Glickman, Michael S; Shuman, Stewart

    2007-05-18

    Mycobacterium tuberculosis and other bacterial pathogens have a Ku-dependent nonhomologous end joining pathway of DNA double-strand break repair. Here we identify mycobacterial UvrD1 as a novel interaction partner for Ku in a genome-wide yeast two-hybrid screen. UvrD1 per se is a vigorous DNA-dependent ATPase but a feeble DNA helicase. Ku stimulates UvrD1 to catalyze ATP-dependent unwinding of 3'-tailed DNAs. UvrD1, Ku, and DNA form a stable ternary complex in the absence of ATP. The Ku binding determinants are located in the distinctive C-terminal segment of UvrD1. A second mycobacterial paralog, UvrD2, is a vigorous Ku-independent DNA helicase. Ablation of UvrD1 sensitizes Mycobacterium smegmatis to killing by ultraviolet and ionizing radiation and to a single chromosomal break generated by I-SceI endonuclease. The physical and functional interactions of bacterial Ku and UvrD1 highlight the potential for cross-talk between components of nonhomologous end joining and nucleotide excision repair pathways.

  1. DNA repair in Mycobacterium tuberculosis revisited.

    Science.gov (United States)

    Dos Vultos, Tiago; Mestre, Olga; Tonjum, Tone; Gicquel, Brigitte

    2009-05-01

    Our understanding of Mycobacterium tuberculosis DNA repair mechanisms is still poor compared with that of other bacterial organisms. However, the publication of the first complete M. tuberculosis genome sequence 10 years ago boosted the study of DNA repair systems in this organism. A first step in the elucidation of M. tuberculosis DNA repair mechanisms was taken by Mizrahi and Andersen, who identified homologs of genes involved in the reversal or repair of DNA damage in Escherichia coli and related organisms. Genes required for nucleotide excision repair, base excision repair, recombination, and SOS repair and mutagenesis were identified. Notably, no homologs of genes involved in mismatch repair were identified. Novel characteristics of the M. tuberculosis DNA repair machinery have been found over the last decade, such as nonhomologous end joining, the presence of Mpg, ERCC3 and Hlr - proteins previously presumed to be produced exclusively in mammalian cells - and the recently discovered bifunctional dCTP deaminase:dUTPase. The study of these systems is important to develop therapeutic agents that can counteract M. tuberculosis evolutionary changes and to prevent adaptive events resulting in antibiotic resistance. This review summarizes our current understanding of the M. tuberculosis DNA repair system.

  2. Exonuclease 1 and its versatile roles in DNA repair

    DEFF Research Database (Denmark)

    Keijzers, Guido; Liu, Dekang; Rasmussen, Lene Juel

    2016-01-01

    Exonuclease 1 (EXO1) is a multifunctional 5' → 3' exonuclease and a DNA structure-specific DNA endonuclease. EXO1 plays roles in DNA replication, DNA mismatch repair (MMR) and DNA double-stranded break repair (DSBR) in lower and higher eukaryotes and contributes to meiosis, immunoglobulin...... maturation, and micro-mediated end-joining in higher eukaryotes. In human cells, EXO1 is also thought to play a role in telomere maintenance. Mutations in the human EXO1 gene correlate with increased susceptibility to some cancers. This review summarizes recent studies on the enzymatic functions...

  3. 30 CFR 75.1508 - Training and records for examination, maintenance and repair of refuge alternatives and components.

    Science.gov (United States)

    2010-07-01

    ... repair, the person conducting the maintenance or repair shall make a record of all corrective action..., maintenance and repair of refuge alternatives and components. 75.1508 Section 75.1508 Mineral Resources MINE..., maintenance and repair of refuge alternatives and components. (a) Persons examining, maintaining, or repairing...

  4. SAMHD1 Promotes DNA End Resection to Facilitate DNA Repair by Homologous Recombination

    Directory of Open Access Journals (Sweden)

    Waaqo Daddacha

    2017-08-01

    Full Text Available DNA double-strand break (DSB repair by homologous recombination (HR is initiated by CtIP/MRN-mediated DNA end resection to maintain genome integrity. SAMHD1 is a dNTP triphosphohydrolase, which restricts HIV-1 infection, and mutations are associated with Aicardi-Goutières syndrome and cancer. We show that SAMHD1 has a dNTPase-independent function in promoting DNA end resection to facilitate DSB repair by HR. SAMHD1 deficiency or Vpx-mediated degradation causes hypersensitivity to DSB-inducing agents, and SAMHD1 is recruited to DSBs. SAMHD1 complexes with CtIP via a conserved C-terminal domain and recruits CtIP to DSBs to facilitate end resection and HR. Significantly, a cancer-associated mutant with impaired CtIP interaction, but not dNTPase-inactive SAMHD1, fails to rescue the end resection impairment of SAMHD1 depletion. Our findings define a dNTPase-independent function for SAMHD1 in HR-mediated DSB repair by facilitating CtIP accrual to promote DNA end resection, providing insight into how SAMHD1 promotes genome integrity.

  5. RNF4 is required for DNA double-strand break repair in vivo

    DEFF Research Database (Denmark)

    Vyas, R; Kumar, R; Clermont, F

    2013-01-01

    for both homologous recombination (HR) and non-homologous end joining repair. To establish a link between Rnf4 and the DNA damage response (DDR) in vivo, we generated an Rnf4 allelic series in mice. We show that Rnf4-deficiency causes persistent ionizing radiation-induced DNA damage and signaling...

  6. The role of DNA dependent protein kinase in synapsis of DNA ends

    NARCIS (Netherlands)

    E.P.W.C. Weterings (Eric); N.S. Verkaik (Nicole); H.T. Brüggenwirth (Hennie); D.C. van Gent (Dik); J.H.J. Hoeijmakers (Jan)

    2003-01-01

    textabstractDNA dependent protein kinase (DNA-PK) plays a central role in the non-homologous end-joining pathway of DNA double strand break repair. Its catalytic subunit (DNA-PK(CS)) functions as a serine/threonine protein kinase. We show that DNA-PK forms a stable complex at DNA termini that blocks

  7. Repairable-conditionally repairable damage model based on dual Poisson processes.

    Science.gov (United States)

    Lind, B K; Persson, L M; Edgren, M R; Hedlöf, I; Brahme, A

    2003-09-01

    The advent of intensity-modulated radiation therapy makes it increasingly important to model the response accurately when large volumes of normal tissues are irradiated by controlled graded dose distributions aimed at maximizing tumor cure and minimizing normal tissue toxicity. The cell survival model proposed here is very useful and flexible for accurate description of the response of healthy tissues as well as tumors in classical and truly radiobiologically optimized radiation therapy. The repairable-conditionally repairable (RCR) model distinguishes between two different types of damage, namely the potentially repairable, which may also be lethal, i.e. if unrepaired or misrepaired, and the conditionally repairable, which may be repaired or may lead to apoptosis if it has not been repaired correctly. When potentially repairable damage is being repaired, for example by nonhomologous end joining, conditionally repairable damage may require in addition a high-fidelity correction by homologous repair. The induction of both types of damage is assumed to be described by Poisson statistics. The resultant cell survival expression has the unique ability to fit most experimental data well at low doses (the initial hypersensitive range), intermediate doses (on the shoulder of the survival curve), and high doses (on the quasi-exponential region of the survival curve). The complete Poisson expression can be approximated well by a simple bi-exponential cell survival expression, S(D) = e(-aD) + bDe(-cD), where the first term describes the survival of undamaged cells and the last term represents survival after complete repair of sublethal damage. The bi-exponential expression makes it easy to derive D(0), D(q), n and alpha, beta values to facilitate comparison with classical cell survival models.

  8. Alternatives Analysis Amchitka Island Mud Pit Cap Repair, Amchitka, Alaska January 2016

    Energy Technology Data Exchange (ETDEWEB)

    Darr, Paul S. [US Department of Energy, Washington, DC (United States). Office of Legacy Management

    2016-01-01

    The U.S. Department of Energy (DOE) Office of Legacy Management (LM) manages the Nevada Offsites program, which includes a series of reclaimed drilling mud impoundments on Amchitka Island, Alaska (Figure 1). Navarro Research and Engineering, Inc. is the Legacy Management Support contractor (the Contractor) for LM. The Contractor has procured Tetra Tech, Inc. to provide engineering support to the Amchitka mud pit reclamation project. The mud pit caps were damaged during a 7.9-magnitude earthquake that occurred in 2014. The goals of the current project are to investigate conditions at the mud pit impoundments, identify feasible alternatives for repair of the cover systems and the contents, and estimate relative costs of repair alternatives. This report presents descriptions of the sites and past investigations, existing conditions, summaries of various repair/mitigation alternatives, and direct, unburdened, order-of-magnitude (-15% to +50%) associated costs.

  9. Alternative end-joining of DNA breaks

    NARCIS (Netherlands)

    Schendel, Robin van

    2016-01-01

    DNA is arguably the most important molecule found in any organism, as it contains all information to perform cellular functions and enables continuity of species. It is continuously exposed to DNA-damaging agents both from endogenous and exogenous sources. To protect DNA against these sources of DNA

  10. Double strand break repair: two mechanisms in competition but tightly linked to cell cycle

    International Nuclear Information System (INIS)

    Delacote, F.

    2002-11-01

    DNA double strand breaks (DSB) are highly toxic damage although they can be induced to create genetic diversity. Two distinct pathways can repair DSB: Homologous Recombination (HR) and Non Homologous End Joining (NHEJ). If un- or mis-repaired, this damage can lead to cancer. Thus, it is essential to investigate how these two pathways are regulated for DSB repair. NHEJ inhibition leads to HR DSB repair stimulation. However, this channeling to HR is tightly linked to cell cycle since NHEJ and HR are active in G1/early S and late S/G2, respectively. Our results suggest that G1-unrepaired DSB go through S phase to be repaired by HR in G2. Those results allow a better understanding of DSB repair mechanisms regulation. (author)

  11. GANP regulates the choice of DNA repair pathway by DNA-PKcs interaction in AID-dependent IgV region diversification.

    Science.gov (United States)

    Eid, Mohammed Mansour Abbas; Maeda, Kazuhiko; Almofty, Sarah Ameen; Singh, Shailendra Kumar; Shimoda, Mayuko; Sakaguchi, Nobuo

    2014-06-15

    RNA export factor germinal center-associated nuclear protein (GANP) interacts with activation-induced cytidine deaminase (AID) and shepherds it from the cytoplasm to the nucleus and toward the IgV region loci in B cells. In this study, we demonstrate a role for GANP in the repair of AID-initiated DNA damage in chicken DT40 B cells to generate IgV region diversity by gene conversion and somatic hypermutation. GANP plays a positive role in IgV region diversification of DT40 B cells in a nonhomologous end joining-proficient state. DNA-PKcs physically interacts with GANP, and this interaction is dissociated by dsDNA breaks induced by a topoisomerase II inhibitor, etoposide, or AID overexpression. GANP affects the choice of DNA repair mechanism in B cells toward homologous recombination rather than nonhomologous end joining repair. Thus, GANP presumably plays a critical role in protection of the rearranged IgV loci by favoring homologous recombination of the DNA breaks under accelerated AID recruitment. Copyright © 2014 by The American Association of Immunologists, Inc.

  12. A data mining approach for classifying DNA repair genes into ageing-related or non-ageing-related

    Directory of Open Access Journals (Sweden)

    Vasieva Olga

    2011-01-01

    Full Text Available Abstract Background The ageing of the worldwide population means there is a growing need for research on the biology of ageing. DNA damage is likely a key contributor to the ageing process and elucidating the role of different DNA repair systems in ageing is of great interest. In this paper we propose a data mining approach, based on classification methods (decision trees and Naive Bayes, for analysing data about human DNA repair genes. The goal is to build classification models that allow us to discriminate between ageing-related and non-ageing-related DNA repair genes, in order to better understand their different properties. Results The main patterns discovered by the classification methods are as follows: (a the number of protein-protein interactions was a predictor of DNA repair proteins being ageing-related; (b the use of predictor attributes based on protein-protein interactions considerably increased predictive accuracy of attributes based on Gene Ontology (GO annotations; (c GO terms related to "response to stimulus" seem reasonably good predictors of ageing-relatedness for DNA repair genes; (d interaction with the XRCC5 (Ku80 protein is a strong predictor of ageing-relatedness for DNA repair genes; and (e DNA repair genes with a high expression in T lymphocytes are more likely to be ageing-related. Conclusions The above patterns are broadly integrated in an analysis discussing relations between Ku, the non-homologous end joining DNA repair pathway, ageing and lymphocyte development. These patterns and their analysis support non-homologous end joining double strand break repair as central to the ageing-relatedness of DNA repair genes. Our work also showcases the use of protein interaction partners to improve accuracy in data mining methods and our approach could be applied to other ageing-related pathways.

  13. Genome engineering with TALENs and ZFNs: repair pathways and donor design.

    Science.gov (United States)

    Carroll, Dana; Beumer, Kelly J

    2014-09-01

    Genome engineering with targetable nucleases depends on cellular pathways of DNA repair after target cleavage. Knowledge of how those pathways work, their requirements and their active factors, can guide experimental design and improve outcomes. While many aspects of both homologous recombination (HR) and nonhomologous end joining (NHEJ) are shared by a broad range of cells and organisms, some features are specific to individual situations. This article reviews the influence of repair mechanisms on the results of gene targeting experiments, with an emphasis on lessons learned from experiments with Drosophila. Copyright © 2014 Elsevier Inc. All rights reserved.

  14. The helicase domain of Polθ counteracts RPA to promote alt-NHEJ.

    Science.gov (United States)

    Mateos-Gomez, Pedro A; Kent, Tatiana; Deng, Sarah K; McDevitt, Shane; Kashkina, Ekaterina; Hoang, Trung M; Pomerantz, Richard T; Sfeir, Agnel

    2017-12-01

    Mammalian polymerase theta (Polθ) is a multifunctional enzyme that promotes error-prone DNA repair by alternative nonhomologous end joining (alt-NHEJ). Here we present structure-function analyses that reveal that, in addition to the polymerase domain, Polθ-helicase activity plays a central role during double-strand break (DSB) repair. Our results show that the helicase domain promotes chromosomal translocations by alt-NHEJ in mouse embryonic stem cells and also suppresses CRISPR-Cas9- mediated gene targeting by homologous recombination (HR). In vitro assays demonstrate that Polθ-helicase activity facilitates the removal of RPA from resected DSBs to allow their annealing and subsequent joining by alt-NHEJ. Consistent with an antagonistic role for RPA during alt-NHEJ, inhibition of RPA1 enhances end joining and suppresses recombination. Taken together, our results reveal that the balance between HR and alt-NHEJ is controlled by opposing activities of Polθ and RPA, providing further insight into the regulation of repair-pathway choice in mammalian cells.

  15. DNA double-strand break repair: a tale of pathway choices

    Institute of Scientific and Technical Information of China (English)

    Jing Li; Xingzhi Xu

    2016-01-01

    Deoxyribonucleic acid double-strand breaks (DSBs) are cytotoxic lesions that must be repaired either through homologous recombination (HR) or non-homologous end-joining (NHEJ) pathways.DSB repair is critical for genome integrity,cellular homeostasis and also constitutes the biological foundation for radiotherapy and the majority of chemotherapy.The choice between HR and NHEJ is a complex yet not completely understood process that will entail more future efforts.Herein we review our current understandings about how the choice is made over an antagonizing balance between p53-binding protein 1 and breast cancer 1 in the context of cell cycle stages,downstream effects,and distinct chromosomal histone marks.These exciting areas of research will surely bring more mechanistic insights about DSB repair and be utilized in the clinical settings.

  16. Repair of endogenous and ionizing radiation-induced DNA damages: mechanisms and biological functions

    International Nuclear Information System (INIS)

    Boiteux, S.

    2002-01-01

    The cellular DNA is continuously exposed to endogenous and exogenous stress. Oxidative stress due to cellular metabolism is the major cause of endogenous DNA damage. On the other hand, ionizing radiation (IR) is an important exogenous stress. Both induce similar DNA damages: damaged bases, abasic sites and strand breakage. Most of these lesions are lethal and/or mutagenic. The survival of the cell is managed by efficient and accurate DNA repair mechanisms that remove lesions before their replication or transcription. DNA repair pathways involved in the removal of IR-induced lesions are briefly described. Base excision repair (BER) is mostly involved in the removal of base damage, abasic sites and single strand breaks. In contrast, DNA double strand breaks are mostly repaired by non-homologous end joining (NHEJ) or homologous recombination (HR). How DNA repair pathways prevent cancer process is also discussed. (author)

  17. Deletion of individual Ku subunits in mice causes an NHEJ-independent phenotype potentially by altering apurinic/apyrimidinic site repair

    NARCIS (Netherlands)

    Y.J. Choi (Yong Jun); H. Li (Han); M.Y. Son (Mi Young); X.-H. Wang (Xiao-Hong); J.L. Fornsaglio (Jamie L.); R.W. Sobol (Robert W.); M. Lee (Moonsook); J. Vijg (Jan); S. Imholz (Sandra); M.E.T. Dollé (Martijn); H. van Steeg (Harry); E. Reiling (Erwin); P. Hasty (Paul)

    2014-01-01

    textabstractKu70 and Ku80 form a heterodimer called Ku that forms a holoenzyme with DNA dependent-protein kinase catalytic subunit (DNA-PKCS) to repair DNA double strand breaks (DSBs) through the nonhomologous end joining (NHEJ) pathway. As expected mutating these genes in mice caused a similar DSB

  18. FAMS DECOMMISSIONING END-STATE ALTERNATIVE EVALUATION

    International Nuclear Information System (INIS)

    Grimm, B; Stephen Chostner, S; Brenda Green, B

    2006-01-01

    Nuclear Material Management (NMM) completed a comprehensive study at the request of the Department of Energy Savannah River Operations Office (DOE-SR) in 2004 (Reference 11.1). The study evaluated the feasibility of removal and/or mitigation of the Pu-238 source term in the F-Area Material Storage (FAMS) facility during on-going material storage operations. The study recommended different options to remove and/or mitigate the Pu-238 source term depending on its location within the facility. During April 2005, the Department of Energy (DOE) sent a letter of direction (LOD) to Washington Savannah River Company (WSRC) directing WSRC to implement a new program direction that would enable an accelerated shutdown and decommissioning of FAMS (Reference 11.2). Further direction in the LOD stated that effective December 1, 2006 the facility will be transitioned to begin deactivation and decommissioning (D and D) activities. To implement the LOD, Site D and D (SDD) and DOE agreed the planning end-state would be demolition of the FAMS structure to the building slab. SDD developed the D and D strategy, preliminary cost and schedule, and issued the deactivation project plan in December 2005 (Reference 11.3). Due to concerns and questions regarding the FAMS planning end-state and in support of the project's Critical Decision 1, an alternative study was performed to evaluate the various decommissioning end-states and the methods by which those end-states are achieved. This report documents the results of the alternative evaluation which was performed in a structured decision-making process as outlined in the E7 Manual, Procedure 2.15, ''Alternative Studies'' (Reference 11.4)

  19. The Impact of Hedgehog Signaling Pathway on DNA Repair Mechanisms in Human Cancer

    International Nuclear Information System (INIS)

    Meng, Erhong; Hanna, Ann; Samant, Rajeev S.; Shevde, Lalita A.

    2015-01-01

    Defined cellular mechanisms have evolved that recognize and repair DNA to protect the integrity of its structure and sequence when encountering assaults from endogenous and exogenous sources. There are five major DNA repair pathways: mismatch repair, nucleotide excision repair, direct repair, base excision repair and DNA double strand break repair (including non-homologous end joining and homologous recombination repair). Aberrant activation of the Hedgehog (Hh) signaling pathway is a feature of many cancer types. The Hh pathway has been documented to be indispensable for epithelial-mesenchymal transition, invasion and metastasis, cancer stemness, and chemoresistance. The functional transcription activators of the Hh pathway include the GLI proteins. Inhibition of the activity of GLI can interfere with almost all DNA repair types in human cancer, indicating that Hh/GLI functions may play an important role in enabling tumor cells to survive lethal types of DNA damage induced by chemotherapy and radiotherapy. Thus, Hh signaling presents an important therapeutic target to overcome DNA repair-enabled multi-drug resistance and consequently increase chemotherapeutic response in the treatment of cancer

  20. The Impact of Hedgehog Signaling Pathway on DNA Repair Mechanisms in Human Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Meng, Erhong; Hanna, Ann; Samant, Rajeev S.; Shevde, Lalita A., E-mail: lsamant@uab.edu [Department of Pathology, Comprehensive Cancer Center, University of Alabama at Birmingham, WTI320D, 1824 6th Avenue South, Birmingham, AL 35233 (United States)

    2015-07-21

    Defined cellular mechanisms have evolved that recognize and repair DNA to protect the integrity of its structure and sequence when encountering assaults from endogenous and exogenous sources. There are five major DNA repair pathways: mismatch repair, nucleotide excision repair, direct repair, base excision repair and DNA double strand break repair (including non-homologous end joining and homologous recombination repair). Aberrant activation of the Hedgehog (Hh) signaling pathway is a feature of many cancer types. The Hh pathway has been documented to be indispensable for epithelial-mesenchymal transition, invasion and metastasis, cancer stemness, and chemoresistance. The functional transcription activators of the Hh pathway include the GLI proteins. Inhibition of the activity of GLI can interfere with almost all DNA repair types in human cancer, indicating that Hh/GLI functions may play an important role in enabling tumor cells to survive lethal types of DNA damage induced by chemotherapy and radiotherapy. Thus, Hh signaling presents an important therapeutic target to overcome DNA repair-enabled multi-drug resistance and consequently increase chemotherapeutic response in the treatment of cancer.

  1. DNA Double Strand Break Response and Limited Repair Capacity in Mouse Elongated Spermatids

    Directory of Open Access Journals (Sweden)

    Emad A. Ahmed

    2015-12-01

    Full Text Available Spermatids are extremely sensitive to genotoxic exposures since during spermiogenesis only error-prone non homologous end joining (NHEJ repair pathways are available. Hence, genomic damage may accumulate in sperm and be transmitted to the zygote. Indirect, delayed DNA fragmentation and lesions associated with apoptotic-like processes have been observed during spermatid elongation, 27 days after irradiation. The proliferating spermatogonia and early meiotic prophase cells have been suggested to retain a memory of a radiation insult leading later to this delayed fragmentation. Here, we used meiotic spread preparations to localize phosphorylate histone H2 variant (γ-H2AX foci marking DNA double strand breaks (DSBs in elongated spermatids. This technique enabled us to determine the background level of DSB foci in elongated spermatids of RAD54/RAD54B double knockout (dko mice, severe combined immunodeficiency SCID mice, and poly adenosine diphosphate (ADP-ribose polymerase 1 (PARP1 inhibitor (DPQ-treated mice to compare them with the appropriate wild type controls. The repair kinetics data and the protein expression patterns observed indicate that the conventional NHEJ repair pathway is not available for elongated spermatids to repair the programmed and the IR-induced DSBs, reflecting the limited repair capacity of these cells. However, although elongated spermatids express the proteins of the alternative NHEJ, PARP1-inhibition had no effect on the repair kinetics after IR, suggesting that DNA damage may be passed onto sperm. Finally, our genetic mutant analysis suggests that an incomplete or defective meiotic recombinational repair of Spo11-induced DSBs may lead to a carry-over of the DSB damage or induce a delayed nuclear fragmentation during the sensitive programmed chromatin remodeling occurring in elongated spermatids.

  2. The C-Terminal Domain of Cernunnos/XLF Is Dispensable for DNA Repair In Vivo▿ †

    Science.gov (United States)

    Malivert, Laurent; Callebaut, Isabelle; Rivera-Munoz, Paola; Fischer, Alain; Mornon, Jean-Paul; Revy, Patrick; de Villartay, Jean-Pierre

    2009-01-01

    The core nonhomologous end-joining DNA repair pathway is composed of seven factors: Ku70, Ku80, DNA-PKcs, Artemis, XRCC4 (X4), DNA ligase IV (L4), and Cernunnos/XLF (Cernunnos). Although Cernunnos and X4 are structurally related and participate in the same complex together with L4, they have distinct functions during DNA repair. L4 relies on X4 but not on Cernunnos for its stability, and L4 is required for optimal interaction of Cernunnos with X4. We demonstrate here, using in vitro-generated Cernunnos mutants and a series of functional assays in vivo, that the C-terminal region of Cernunnos is dispensable for its activity during DNA repair. PMID:19103754

  3. Mycobacteria exploit three genetically distinct DNA double-strand break repair pathways.

    Science.gov (United States)

    Gupta, Richa; Barkan, Daniel; Redelman-Sidi, Gil; Shuman, Stewart; Glickman, Michael S

    2011-01-01

    Bacterial pathogens rely on their DNA repair pathways to resist genomic damage inflicted by the host. DNA double-strand breaks (DSBs) are especially threatening to bacterial viability. DSB repair by homologous recombination (HR) requires nucleases that resect DSB ends and a strand exchange protein that facilitates homology search. RecBCD and RecA perform these functions in Escherichia coli and constitute the major pathway of error-free DSB repair. Mycobacteria, including the human pathogen M. tuberculosis, elaborate an additional error-prone pathway of DSB repair via non-homologous end-joining (NHEJ) catalysed by Ku and DNA ligase D (LigD). Little is known about the relative contributions of HR and NHEJ to mycobacterial chromosome repair, the factors that dictate pathway choice, or the existence of additional DSB repair pathways. Here we demonstrate that Mycobacterium smegmatis has three DSB repair pathway options: HR, NHEJ and a novel mechanism of single-strand annealing (SSA). Inactivation of NHEJ or SSA is compensated by elevated HR. We find that mycobacterial RecBCD does not participate in HR or confer resistance to ionizing radiation (IR), but is required for the RecA-independent SSA pathway. In contrast, the mycobacterial helicase-nuclease AdnAB participates in the RecA-dependent HR pathway, and is a major determinant of resistance to IR and oxidative DNA damage. These findings reveal distinctive features of mycobacterial DSB repair, most notably the dedication of the RecBCD and AdnAB helicase-nuclease machines to distinct repair pathways. © 2010 Blackwell Publishing Ltd.

  4. An Alternative Front End Analysis Strategy for Complex Systems

    Science.gov (United States)

    2014-12-01

    missile ( ABM ) system . Patriot is employed in the field through a battalion echelon organizational structure. The line battery is the basic building...Research Report 1981 An Alternative Front End Analysis Strategy for Complex Systems M. Glenn Cobb U.S. Army Research Institute...NUMBER W5J9CQ11D0003 An Alternative Front End Analysis Strategy for Complex Systems 5b. PROGRAM ELEMENT NUMBER 633007 6

  5. Beyond repair foci: DNA double-strand break repair in euchromatic and heterochromatic compartments analyzed by transmission electron microscopy.

    Directory of Open Access Journals (Sweden)

    Yvonne Lorat

    Full Text Available DNA double-strand breaks (DSBs generated by ionizing radiation pose a serious threat to the preservation of genetic and epigenetic information. The known importance of local chromatin configuration in DSB repair raises the question of whether breaks in different chromatin environments are recognized and repaired by the same repair machinery and with similar efficiency. An essential step in DSB processing by non-homologous end joining is the high-affinity binding of Ku70-Ku80 and DNA-PKcs to double-stranded DNA ends that holds the ends in physical proximity for subsequent repair.Using transmission electron microscopy to localize gold-labeled pKu70 and pDNA-PKcs within nuclear ultrastructure, we monitored the formation and repair of actual DSBs within euchromatin (electron-lucent and heterochromatin (electron-dense in cortical neurons of irradiated mouse brain.While DNA lesions in euchromatin (characterized by two pKu70-gold beads, reflecting the Ku70-Ku80 heterodimer are promptly sensed and rejoined, DNA packaging in heterochromatin appears to retard DSB processing, due to the time needed to unravel higher-order chromatin structures. Complex pKu70-clusters formed in heterochromatin (consisting of 4 or ≥ 6 gold beads may represent multiple breaks in close proximity caused by ionizing radiation of highly-compacted DNA. All pKu70-clusters disappeared within 72 hours post-irradiation, indicating efficient DSB rejoining. However, persistent 53BP1 clusters in heterochromatin (comprising ≥ 10 gold beads, occasionally co-localizing with γH2AX, but not pKu70 or pDNA-PKcs, may reflect incomplete or incorrect restoration of chromatin structure rather than persistently unrepaired DNA damage.Higher-order organization of chromatin determines the accessibility of DNA lesions to repair complexes, defining how readily DSBs are detected and processed. DNA lesions in heterochromatin appear to be more complex, with multiple breaks in spatial vicinity inducing

  6. TEMS: an alternative method for the repair of benign recto-vaginal fistulae.

    Science.gov (United States)

    Darwood, R J; Borley, N R

    2008-07-01

    The repair of high recto-vaginal fistula can be challenging since access may be limited via the endo-anal approach yet the alternative trans-abdominal route carries significant morbidity. We report the use of TEMS to repair a recto-vaginal fistula following anterior resection and pelvic radiotherapy. The patient was placed prone and a 25 cm rectoscope was inserted. A proximally based mucosal advancement flap was raised to repair the fistula. The patient was discharged 2 days later and a contrast study confirmed closure of the fistula. COMPARISON WITH OTHER TECHNIQUES: TEMS allows excellent visualisation of a rectovaginal fistula compared to standard endo-anal or trans-vaginal techniques. The morbidity is lower than the trans-abdominal route. TEMS is a useful technique for the repair of benign recto-vaginal fistula and has distinct advantages over conventional techniques.

  7. Regulation of homologous recombination repair protein Rad51 by Ku70

    International Nuclear Information System (INIS)

    Du Liqing; Liu Qiang; Wang Yan; Xu Chang; Cao Jia; Fu Yue; Chen Fenghua; Fan Feiyue

    2013-01-01

    Objective: To explore the regulative effect of non-homologous end joining (NHEJ)protein Ku70 on homologous recombination repair protein Rad51, and to investigate the synergistic mechanism of homologous recombination repair in combination with NHEJ. Methods: Observed Rad51 protein expression after transfect Ku70 small interfering RNA or Ku70 plasmid DNA into tumor cells using Western blot. Results: Expression of Rad51 was obviously reduced after pretreated with Ku70 small interfering RNA. And with the increasing expression of Ku70 protein after transfection of Ku70 plasmid DNA PGCsi3.0-hKu70 into tumor cell lines, the Rad51 protein expression was increased. Conclusion: Ku70 protein has regulating effect on gene expression of Rad51, and it might participate in the collaboration between homologous recombination repair and NHEJ. (authors)

  8. Chernobyl risk reduction via ventilation stack repair: a successful international project

    International Nuclear Information System (INIS)

    Gore, B.F.; Glukhov, A.Y.; Schmidt, J.P.; Vargo, G.I.; Osterloh, J.V.; Kupny, V.I.; Korneev, A.A.; Matvejev, Y.A.

    1999-01-01

    The recently completed project to repair the ChNPP Unit 3/4 ventilation stack is the first international project providing a direct physical improvement and safety upgrade at the Shelter (Ukrytie) facility. The 1986 explosion at Chernobyl NPP Unit 4 severely damaged the ventilation stack framework of bracing and its foundation. At one node, where six elements of the framework joined together, one element was completely missing one element was torn from the node and left hanging from its other end, and one element was badly bent and dented. Foundation supports were knocked measurably off plumb. Repair of the ChNPP ventilation stack is the first international project providing direct physical improvement and safety upgrade at the Shelter facility. The repairs have restored the stack to full design strength, from an emergency condition with a high probability of collapse

  9. Screening of Pesticides with the Potential of Inducing DSB and Successive Recombinational Repair

    Directory of Open Access Journals (Sweden)

    Karen Suárez-Larios

    2017-01-01

    Full Text Available A study was realized to ascertain whether eight selected pesticides would induce double strand breaks (DSB in lymphocyte cultures and whether this damage would induce greater levels of proteins Rad51 participating in homologous recombination or of p-Ku80 participating in nonhomologous end joining. Only five pesticides were found to induce DSB of which only glyphosate and paraoxon induced a significant increase of p-Ku80 protein, indicating that nonhomologous end joining recombinational DNA repair system would be activated. The type of gamma-H2AX foci observed was comparable to that induced by etoposide at similar concentrations. These results are of importance since these effects occurred at low concentrations in the micromolar range, in acute treatments to the cells. Effects over longer exposures in actual environmental settings are expected to produce cumulative damage if repeated events of recombination take place over time.

  10. Bruce unit 1 moderator to end shield cooling leak repairs

    Energy Technology Data Exchange (ETDEWEB)

    Boucher, P; Ashton, A [Ontario Hydro, Tiverton, ON (Canada). Bruce Nuclear Generating Station-A

    1996-12-31

    In October 1994, a leak developed between the heavy water Moderator System and the light water End Shield Cooling System at Ontario Hydro`s Bruce A Generating Station Unit 1. The interface between these two systems consists of numerous reactor components all within the reactor vessel. This paper describes the initial discovery and determination of the leak source. The techniques used to pinpoint the leak location are described. The repair strategies and details are outlined. Flushing and refilling of the Moderator system are discussed. The current status of the Unit 1 End Shield Cooling System is given with possible remedial measures for clean-up. Recommendations and observations are provided for future references. (author). 7 figs.

  11. Sensitization to radiation and alkylating agents by inhibitors of poly(ADP-ribose) polymerase is enhanced in cells deficient in DNA double-strand break repair.

    Science.gov (United States)

    Löser, Dana A; Shibata, Atsushi; Shibata, Akiko K; Woodbine, Lisa J; Jeggo, Penny A; Chalmers, Anthony J

    2010-06-01

    As single agents, chemical inhibitors of poly(ADP-ribose) polymerase (PARP) are nontoxic and have clinical efficacy against BRCA1- and BRCA2-deficient tumors. PARP inhibitors also enhance the cytotoxicity of ionizing radiation and alkylating agents but will only improve clinical outcomes if tumor sensitization exceeds effects on normal tissues. It is unclear how tumor DNA repair proficiency affects the degree of sensitization. We have previously shown that the radiosensitizing effect of PARP inhibition requires DNA replication and will therefore affect rapidly proliferating tumors more than normal tissues. Because many tumors exhibit defective DNA repair, we investigated the impact of double-strand break (DSB) repair integrity on the sensitizing effects of the PARP inhibitor olaparib. Sensitization to ionizing radiation and the alkylating agent methylmethane sulfonate was enhanced in DSB repair-deficient cells. In Artemis(-/-) and ATM(-/-) mouse embryo fibroblasts, sensitization was replication dependent and associated with defective repair of replication-associated damage. Radiosensitization of Ligase IV(-/-) mouse embryo fibroblasts was independent of DNA replication and is explained by inhibition of "alternative" end joining. After methylmethane sulfonate treatment, PARP inhibition promoted replication-independent accumulation of DSB, repair of which required Ligase IV. Our findings predict that the sensitizing effects of PARP inhibitors will be more pronounced in rapidly dividing and/or DNA repair defective tumors than normal tissues and show their potential to enhance the therapeutic ratio achieved by conventional DNA-damaging agents.

  12. DNA repair: Dynamic defenders against cancer and aging

    Energy Technology Data Exchange (ETDEWEB)

    Fuss, Jill O.; Cooper, Priscilla K.

    2006-04-01

    (UV) component of sunlight. NER can be divided into two classes based on where the repair occurs. NER occurring in DNA that is not undergoing transcription (i.e., most of the genome) is called global genome repair (GGR or GGNER), while NER taking place in the transcribed strand of active genes is called transcription-coupled repair (TCR or TC-NER). We will explore NER in more detail below. Mismatch repair (MMR) is another type of excision repair that specifically removes mispaired bases resulting from replication errors. DNA damage can also result in breaks in the DNA backbone, in one or both strands. Single-strand breaks (SSBs) are efficiently repaired by a mechanism that shares common features with the later steps in BER. Double-strand breaks (DSBs) are especially devastating since by definition there is no intact complementary strand to serve as a template for repair, and even one unrepaired DSB can be lethal [3]. In cells that have replicated their DNA prior to cell division, the missing information can be supplied by the duplicate copy, or sister chromatid, and DSBs in these cells are faithfully repaired by homologous recombination involving the exchange of strands of DNA between the two copies. However, most cells in the body are non-dividing, and in these cells the major mechanism for repairing DSBs is by non-homologous end joining (NHEJ), which as the name implies involves joining two broken DNA ends together without a requirement for homologous sequence and which therefore has a high potential for loss of genetic information.

  13. Prospects of joining multi-material structures

    Science.gov (United States)

    Sankaranarayanan, R.; Hynes, N. Rajesh Jesudoss

    2018-05-01

    Spring up trends and necessities make the pipelines for the brand new Technologies. The same way, Multimaterial structures emerging as fruitful alternatives for the conventional structures in the manufacturing sector. Especially manufacturing of transport vehicles is placing a perfect platform for these new structures. Bonding or joining technology plays a crucial role in the field of manufacturing for sustainability. These latest structures are purely depending on such joining technologies so that multi-material structuring can be possible practically. The real challenge lies on joining dissimilar materials of different properties and nature. Escalation of thermoplastic usage in large structural components also faces similar ambiguity for joining multi-material structures. Adhesive bonding, mechanical fastening and are the answering technologies for multi-material structures. This current paper analysis the prospects of these bonding technologies to meet the challenges of tomorrow.

  14. Advanced manufacturing technologies modern machining, advanced joining, sustainable manufacturing

    CERN Document Server

    2017-01-01

    This book provides details and collective information on working principle, process mechanism, salient features, and unique applications of various advanced manufacturing techniques and processes belong. The book is divided in three sessions covering modern machining methods, advanced repair and joining techniques and, finally, sustainable manufacturing. The latest trends and research aspects of those fields are highlighted.

  15. DNA repair in neurons: So if they don't divide what's to repair?

    International Nuclear Information System (INIS)

    Fishel, Melissa L.; Vasko, Michael R.; Kelley, Mark R.

    2007-01-01

    Neuronal DNA repair remains one of the most exciting areas for investigation, particularly as a means to compare the DNA repair response in mitotic (cancer) vs. post-mitotic (neuronal) cells. In addition, the role of DNA repair in neuronal cell survival and response to aging and environmental insults is of particular interest. DNA damage caused by reactive oxygen species (ROS) such as generated by mitochondrial respiration includes altered bases, abasic sites, and single- and double-strand breaks which can be prevented by the DNA base excision repair (BER) pathway. Oxidative stress accumulates in the DNA of the human brain over time especially in the mitochondrial DNA (mtDNA) and is proposed to play a critical role in aging and in the pathogenesis of several neurological disorders including Parkinson's disease, ALS, and Alzheimer's diseases. Because DNA damage accumulates in the mtDNA more than nuclear DNA, there is increased interest in DNA repair pathways and the consequence of DNA damage in the mitochondria of neurons. The type of damage that is most likely to occur in neuronal cells is oxidative DNA damage which is primarily removed by the BER pathway. Following the notion that the bulk of neuronal DNA damage is acquired by oxidative DNA damage and ROS, the BER pathway is a likely area of focus for neuronal studies of DNA repair. BER variations in brain aging and pathology in various brain regions and tissues are presented. Therefore, the BER pathway is discussed in greater detail in this review than other repair pathways. Other repair pathways including direct reversal, nucleotide excision repair (NER), mismatch repair (MMR), homologous recombination and non-homologous end joining are also discussed. Finally, there is a growing interest in the role that DNA repair pathways play in the clinical arena as they relate to the neurotoxicity and neuropathy associated with cancer treatments. Among the numerous side effects of cancer treatments, major clinical effects

  16. Telomeres and genomic damage repair. Their implication in human pathology

    International Nuclear Information System (INIS)

    Perez, Maria del R.; Dubner, Diana; Michelin, Severino; Gisone, Pablo; Carosella, Edgardo D.

    2002-01-01

    Telomeres, functional complexed that protect eukaryotic chromosome ends, participate in the regulation of cell proliferation and could play a role in the stabilization of genomic regions in response to genotoxic stress. Their significance in human pathology becomes evident in several diseases sharing genomic instability as a common trait, in which alterations of the telomere metabolism have been demonstrated. Many of them are also associated with hypersensitivity to ionizing radiation and cancer susceptibility. Besides the specific proteins belonging to the telomeric complex, other proteins involved in the DNA repair machinery, such as ATM, BRCA1, BRCA2, PARP/tankyrase system, DNA-PK and RAD50-MRE11-NBS1 complexes, are closely related with the telomere. This suggests that the telomere sequesters DNA repair proteins for its own structure maintenance, with could also be released toward damaged sites in the genomic DNA. This communication describes essential aspects of telomere structure and function and their links with homologous recombination, non-homologous end-joining (NHEJ), V(D)J system and mismatch-repair (MMR). Several pathological conditions exhibiting alterations in some of these mechanisms are also considered. The cell response to ionizing radiation and its relationship with the telomeric metabolism is particularly taken into account as a model for studying genotoxicity. (author)

  17. Repair pathways for heavy ion-induced complex DNA double strand breaks

    International Nuclear Information System (INIS)

    Yajima, Hirohiko; Nakajima, Nakako; Hirakawa, Hirokazu; Murakami, Takeshi; Okayasu, Ryuichi; Fujimori, Akira

    2012-01-01

    DNA double strand break (DSB) induced by ionizing radiation (IR) is a deleterious damage leading to cell death and genome instability if not properly repaired. It is well known that DSB is repaired by two major pathways, non-homologous end-joining (NHEJ) and homologous recombination (HR). It is also known that NHEJ is dominant throughout the cell cycle after X- or gamma-ray irradiation in mammalian cells, Meanwhile, it is thought that heavy-ion radiation (e.g., carbon-ions, iron-ions) gives rise to clustered DNA damages consisting of not only strand breaks but also aberrant bases in the vicinity of DSBs (complex DSBs). Our previous work suggested that the efficiency of NHEJ is diminished for repair of complex DSBs induced by heavy-ion radiation. We thought that this difficulty in NHEJ process associated with heavy ion induced complex DNA damage might be extended to HR process in cells exposed to heavy ions. In order to find out if this notion is true or not, exposed human cells to X-rays and heavy-ions, and studied HR associated processes at the molecular level. Our result indicates that complex DSBs induced by heavy ions effectively evoke DNA end resection activity during the HR process. Together with our results, a relevant recent progress in the field of DNA DSB repair will be discussed. (author)

  18. An alternate end design for SSC dipoles

    International Nuclear Information System (INIS)

    Peters, C.; Caspi, S.; Taylor, C.

    1989-02-01

    Experience in the SSC dipole program has shown that fabrication of cylindrical coil ends is difficult. Cable stiffness requires large forces to maintain the proper position of the conductors in the end during winding. After winding, the coil ends remain distorted nd significant motion of the need conductors is required to force the coil end into the molding cavity. Local mechanical stresses are high during this process and extra pieces of insulation are required to prevent turn-to-turn shorts from developing during the winding and molding steps. Prior to assembly the coil end is compressed in a mold cavity and injected with a filler material to correct surface irregularities and fill voids in the end. LBL has developed an alternate design which permits the conductors to be wound over the end using minimal force and technician coerosion. The conductors are placed on a conical surface where the largest diameter over the outer layer conductors is 10 cm. No coil end spaces or insulation pieces between turns are required. The conductor geometry was analytically optimized to meet SSC multipole requirements for the ends. The first 1-m dipole utilizing this end geometry has been constructed and successfully tested. Design and construction data are presented. Also model test results, including training and multipole measurements of the end are given. 1 ref., 12 figs., 3 tabs

  19. The role of DNA dependent protein kinase in synapsis of DNA ends.

    Science.gov (United States)

    Weterings, Eric; Verkaik, Nicole S; Brüggenwirth, Hennie T; Hoeijmakers, Jan H J; van Gent, Dik C

    2003-12-15

    DNA dependent protein kinase (DNA-PK) plays a central role in the non-homologous end-joining pathway of DNA double strand break repair. Its catalytic subunit (DNA-PK(CS)) functions as a serine/threonine protein kinase. We show that DNA-PK forms a stable complex at DNA termini that blocks the action of exonucleases and ligases. The DNA termini become accessible after autophosphorylation of DNA-PK(CS), which we demonstrate to require synapsis of DNA ends. Interestingly, the presence of DNA-PK prevents ligation of the two synapsed termini, but allows ligation to another DNA molecule. This alteration of the ligation route is independent of the type of ligase that we used, indicating that the intrinsic architecture of the DNA-PK complex itself is not able to support ligation of the synapsed DNA termini. We present a working model in which DNA-PK creates a stable molecular bridge between two DNA ends that is remodeled after DNA-PK autophosphorylation in such a way that the extreme termini become accessible without disrupting synapsis. We infer that joining of synapsed DNA termini would require an additional protein factor.

  20. Alternative electro-chemically based processing routes for joining of plasma facing components

    Energy Technology Data Exchange (ETDEWEB)

    Krauss, Wolfgang, E-mail: wolfgang.krauss@kit.edu [Karlsruhe Institute of Technology (KIT), Institute for Materials Research III, Hermann-von-Helmholtz-Platz 1 76344 Eggenstein-Leopoldshafen (Germany); Lorenz, Julia; Holstein, Nils; Konys, Juergen [Karlsruhe Institute of Technology (KIT), Institute for Materials Research III, Hermann-von-Helmholtz-Platz 1 76344 Eggenstein-Leopoldshafen (Germany)

    2011-10-15

    Tungsten is considered in fusion technology as functional and structural material in the area of blanket and divertor for future application in DEMO. The KIT design of a He-cooled divertor includes joints between W and W-alloys as well as of W with Eurofer-steel. The main challenges range from expansion mismatch problem for tungsten/steel joints over metallurgical reactions with brittle phase formation to crack stopping ability and excellent surface wetting. These requirements were only met partly and insufficiently in the past e.g. by direct Cu-casting of tungsten onto steel. Both, the joining needs and the observed failure scenarios of conventionally joined components initiated the development of improved joining technologies based on electro-chemical processing routes. As electrolytes aqueous and aprotic, water free, system are integrated into this development line. In the first step principle requirements are presented to guarantee a reproducible and adherent deposition of scales based on Ni and Cu acting as inter layers and filler, respectively, to generate a real metallurgical bonding as demonstrate by 1100 deg. C joining tests. The development field aprotic systems based on ionic liquids is discussed with respect to enable development of refractory metal based fillers with focus high temperature W-W brazing.

  1. How additive manufacturing enables more sustainable end-user maintenance, repair and overhaul (MRO) strategies

    NARCIS (Netherlands)

    Wits, Wessel Willems; Reyes Garcia, José Roberto; Jauregui Becker, Juan Manuel

    2016-01-01

    This paper projects how Maintenance, Repair and Overhaul (MRO) strategies can be optimized to the specific needs of end-users using Additive Manufacturing (AM) technologies. AM can significantly reduce the design and production times for customized parts. This leads to key advantages for MRO

  2. DNA Polymerases λ and β: The Double-Edged Swords of DNA Repair

    Directory of Open Access Journals (Sweden)

    Elisa Mentegari

    2016-08-01

    Full Text Available DNA is constantly exposed to both endogenous and exogenous damages. More than 10,000 DNA modifications are induced every day in each cell’s genome. Maintenance of the integrity of the genome is accomplished by several DNA repair systems. The core enzymes for these pathways are the DNA polymerases. Out of 17 DNA polymerases present in a mammalian cell, at least 13 are specifically devoted to DNA repair and are often acting in different pathways. DNA polymerases β and λ are involved in base excision repair of modified DNA bases and translesion synthesis past DNA lesions. Polymerase λ also participates in non-homologous end joining of DNA double-strand breaks. However, recent data have revealed that, depending on their relative levels, the cell cycle phase, the ratio between deoxy- and ribo-nucleotide pools and the interaction with particular auxiliary proteins, the repair reactions carried out by these enzymes can be an important source of genetic instability, owing to repair mistakes. This review summarizes the most recent results on the ambivalent properties of these enzymes in limiting or promoting genetic instability in mammalian cells, as well as their potential use as targets for anticancer chemotherapy.

  3. DNA Polymerases λ and β: The Double-Edged Swords of DNA Repair.

    Science.gov (United States)

    Mentegari, Elisa; Kissova, Miroslava; Bavagnoli, Laura; Maga, Giovanni; Crespan, Emmanuele

    2016-08-31

    DNA is constantly exposed to both endogenous and exogenous damages. More than 10,000 DNA modifications are induced every day in each cell's genome. Maintenance of the integrity of the genome is accomplished by several DNA repair systems. The core enzymes for these pathways are the DNA polymerases. Out of 17 DNA polymerases present in a mammalian cell, at least 13 are specifically devoted to DNA repair and are often acting in different pathways. DNA polymerases β and λ are involved in base excision repair of modified DNA bases and translesion synthesis past DNA lesions. Polymerase λ also participates in non-homologous end joining of DNA double-strand breaks. However, recent data have revealed that, depending on their relative levels, the cell cycle phase, the ratio between deoxy- and ribo-nucleotide pools and the interaction with particular auxiliary proteins, the repair reactions carried out by these enzymes can be an important source of genetic instability, owing to repair mistakes. This review summarizes the most recent results on the ambivalent properties of these enzymes in limiting or promoting genetic instability in mammalian cells, as well as their potential use as targets for anticancer chemotherapy.

  4. Double strand break rejoining by the Ku-dependent mechanism of non-homologous end-joining

    International Nuclear Information System (INIS)

    Jeggo, P.; Singleton, B.; Beamish, H.; Priestley, A.

    1999-01-01

    The DNA-dependent protein kinase functions in the repair of DNA double strand breaks (DSBs) and in V(D)J recombination. To gain insight into the function of DNA-PK in this process we have carried out a mutation analysis of Ku80 and DNA-PKcs. Mutations at multiple sites within the N-terminal two thirds of Ku80 result in loss of Ku70/80 interaction, loss of DNA end-binding activity and inability to complement Ku80 defective cell fines. In contrast, mutations in the carboxy terminal region of the protein do not impair DNA end-binding activity but decrease the ability of Ku to activate DNA-PK. To gain insight into important functional domains within DNA-PKcs, we have analysed defective mutants, including the mouse scid cell line, and the rodent mutants, ire-20 and V-3. Mutational changes in the carboxy terminal region have been identified in all cases. Our results strongly suggest that the C-terminus of DNA-PKcs is required for kinase activity. (author)

  5. Synapsis-defective mutants reveal a correlation between chromosome conformation and the mode of double-strand break repair during Caenorhabditis elegans meiosis.

    Science.gov (United States)

    Smolikov, Sarit; Eizinger, Andreas; Hurlburt, Allison; Rogers, Eric; Villeneuve, Anne M; Colaiácovo, Mónica P

    2007-08-01

    SYP-3 is a new structural component of the synaptonemal complex (SC) required for the regulation of chromosome synapsis. Both chromosome morphogenesis and nuclear organization are altered throughout the germlines of syp-3 mutants. Here, our analysis of syp-3 mutants provides insights into the relationship between chromosome conformation and the repair of meiotic double-strand breaks (DSBs). Although crossover recombination is severely reduced in syp-3 mutants, the production of viable offspring accompanied by the disappearance of RAD-51 foci suggests that DSBs are being repaired in these synapsis-defective mutants. Our studies indicate that once interhomolog recombination is impaired, both intersister recombination and nonhomologous end-joining pathways may contribute to repair during germline meiosis. Moreover, our studies suggest that the conformation of chromosomes may influence the mode of DSB repair employed during meiosis.

  6. Impact of DNA3'pp5'G capping on repair reactions at DNA 3' ends.

    Science.gov (United States)

    Das, Ushati; Chauleau, Mathieu; Ordonez, Heather; Shuman, Stewart

    2014-08-05

    Many biological scenarios generate "dirty" DNA 3'-PO4 ends that cannot be sealed by classic DNA ligases or extended by DNA polymerases. The noncanonical ligase RtcB can "cap" these ends via a unique chemical mechanism entailing transfer of GMP from a covalent RtcB-GMP intermediate to a DNA 3'-PO4 to form DNA3'pp5'G. Here, we show that capping protects DNA 3' ends from resection by Escherichia coli exonucleases I and III and from end-healing by T4 polynucleotide 3' phosphatase. By contrast, the cap is an effective primer for DNA synthesis. E. coli DNA polymerase I and Mycobacterium DinB1 extend the DNAppG primer to form an alkali-labile DNApp(rG)pDNA product. The addition of dNTP depends on pairing of the cap guanine with an opposing cytosine in the template strand. Aprataxin, an enzyme implicated in repair of A5'pp5'DNA ends formed during abortive ligation by classic ligases, is highly effective as a DNA 3' decapping enzyme, converting DNAppG to DNA3'p and GMP. We conclude that the biochemical impact of DNA capping is to prevent resection and healing of a 3'-PO4 end, while permitting DNA synthesis, at the price of embedding a ribonucleotide and a pyrophosphate linkage in the repaired strand. Aprataxin affords a means to counter the impact of DNA capping.

  7. Effects of different brazing and welding methods on the fracture load of various orthodontic joining configurations.

    Science.gov (United States)

    Bock, Jens J; Bailly, Jacqueline; Fuhrmann, Robert A

    2009-06-01

    The aim of this study was to compare the fracture load of different joints made by conventional brazing, tungston inert gas (TIG) and laser welding. Six standardized joining configurations of spring hard quality orthodontic wire were investigated: end-to-end, round, cross, 3 mm length, 9 mm length and 6.5 mm to orthodontic band. The joints were made by five different methods: brazing with universal silver solder, two TIG and two laser welding devices. The fracture loads were measured with a universal testing machine (Zwick 005). Data were analysed with the Mann-Whitney-Wilcoxon and Kruskal-Wallis tests. The significance level was set at Pwelding (Pwelding (826 N). No differences between the various TIG or laser welding devices were demonstrated, although it was not possible to join an orthodontic wire to an orthodontic band using TIG welding. For orthodontic purposes laser and TIG welding are solder free alternatives. TIG welding and laser welding showed similar results. The laser technique is an expensive, but sophisticated and simple method.

  8. Ubiquitin-specific protease 5 is required for the efficient repair of DNA double-strand breaks.

    Directory of Open Access Journals (Sweden)

    Satoshi Nakajima

    Full Text Available During the DNA damage response (DDR, ubiquitination plays an important role in the recruitment and regulation of repair proteins. However, little is known about elimination of the ubiquitination signal after repair is completed. Here we show that the ubiquitin-specific protease 5 (USP5, a deubiquitinating enzyme, is involved in the elimination of the ubiquitin signal from damaged sites and is required for efficient DNA double-strand break (DSB repair. Depletion of USP5 sensitizes cells to DNA damaging agents, produces DSBs, causes delayed disappearance of γH2AX foci after Bleocin treatment, and influences DSB repair efficiency in the homologous recombination pathway but not in the non-homologous end joining pathway. USP5 co-localizes to DSBs induced by laser micro-irradiation in a RAD18-dependent manner. Importantly, polyubiquitin chains at sites of DNA damage remained for longer periods in USP5-depleted cells. Our results show that disassembly of polyubiquitin chains by USP5 at sites of damage is important for efficient DSB repair.

  9. Numt-mediated double-strand break repair mitigates deletions during primate genome evolution.

    Directory of Open Access Journals (Sweden)

    Einat Hazkani-Covo

    2008-10-01

    Full Text Available Non-homologous end joining (NHEJ is the major mechanism of double-strand break repair (DSBR in mammalian cells. NHEJ has traditionally been inferred from experimental systems involving induced double strand breaks (DSBs. Whether or not the spectrum of repair events observed in experimental NHEJ reflects the repair of natural breaks by NHEJ during chromosomal evolution is an unresolved issue. In primate phylogeny, nuclear DNA sequences of mitochondrial origin, numts, are inserted into naturally occurring chromosomal breaks via NHEJ. Thus, numt integration sites harbor evidence for the mechanisms that act on the genome over evolutionary timescales. We have identified 35 and 55 lineage-specific numts in the human and chimpanzee genomes, respectively, using the rhesus monkey genome as an outgroup. One hundred and fifty two numt-chromosome fusion points were classified based on their repair patterns. Repair involving microhomology and repair leading to nucleotide additions were detected. These repair patterns are within the experimentally determined spectrum of classical NHEJ, suggesting that information from experimental systems is representative of broader genetic loci and end configurations. However, in incompatible DSBR events, small deletions always occur, whereas in 54% of numt integration events examined, no deletions were detected. Numts show a statistically significant reduction in deletion frequency, even in comparison to DSBR involving filler DNA. Therefore, numts show a unique mechanism of integration via NHEJ. Since the deletion frequency during numt insertion is low, native overhangs of chromosome breaks are preserved, allowing us to determine that 24% of the analyzed breaks are cohesive with overhangs of up to 11 bases. These data represent, to the best of our knowledge, the most comprehensive description of the structure of naturally occurring DSBs. We suggest a model in which the sealing of DSBs by numts, and probably by other filler

  10. DNA damage repair and radiosensitivity

    International Nuclear Information System (INIS)

    Suzuki, Norio

    2003-01-01

    Tailored treatment is not new in radiotherapy; it has been the major subject for the last 20-30 years. Radiation responses and RBE (relative biological effectiveness) depend on assay systems, endpoints, type of tissues and tumors, radiation quality, dose rate, dose fractionation, physiological and environmental factors etc, Latent times to develop damages also differ among tissues and endpoints depending on doses and radiation quality. Recent progress in clarification of radiation induced cell death, especially of apoptotic cell death, is quite important for understanding radiosensitivity of tumor cure process as well as of tumorigenesis. Apoptotic cell death as well as dormant cells had been unaccounted and missed into a part of reproductive cell death. Another area of major progress has been made in clarifying repair mechanisms of radiation damage, i.e., non-homologous end joining (NHEJ) and homologous recombinational repair (HRR). New approaches and developments such as cDNA or protein micro arrays and so called informatics in addition to basic molecular biological analysis are expected to aid identifying molecules and their roles in signal transduction pathways, which are multi-factorial and interactive each other being involved in radiation responses. (authors)

  11. Guardians of the mycobacterial genome: A review on DNA repair systems in Mycobacterium tuberculosis.

    Science.gov (United States)

    Singh, Amandeep

    2017-12-01

    The genomic integrity of Mycobacterium tuberculosis is continuously threatened by the harsh survival conditions inside host macrophages, due to immune and antibiotic stresses. Faithful genome maintenance and repair must be accomplished under stress for the bacillus to survive in the host, necessitating a robust DNA repair system. The importance of DNA repair systems in pathogenesis is well established. Previous examination of the M. tuberculosis genome revealed homologues of almost all the major DNA repair systems, i.e. nucleotide excision repair (NER), base excision repair (BER), homologous recombination (HR) and non-homologous end joining (NHEJ). However, recent developments in the field have pointed to the presence of novel proteins and pathways in mycobacteria. Homologues of archeal mismatch repair proteins were recently reported in mycobacteria, a pathway previously thought to be absent. RecBCD, the major nuclease-helicase enzymes involved in HR in E. coli, were implicated in the single-strand annealing (SSA) pathway. Novel roles of archeo-eukaryotic primase (AEP) polymerases, previously thought to be exclusive to NHEJ, have been reported in BER. Many new proteins with a probable role in DNA repair have also been discovered. It is now realized that the DNA repair systems in M. tuberculosis are highly evolved and have redundant backup mechanisms to mend the damage. This review is an attempt to summarize our current understanding of the DNA repair systems in M. tuberculosis.

  12. Stripped-down DNA repair in a highly reduced parasite

    Directory of Open Access Journals (Sweden)

    Fast Naomi M

    2007-03-01

    Full Text Available Abstract Background Encephalitozoon cuniculi is a member of a distinctive group of single-celled parasitic eukaryotes called microsporidia, which are closely related to fungi. Some of these organisms, including E. cuniculi, also have uniquely small genomes that are within the prokaryotic range. Thus, E. cuniculi has undergone a massive genome reduction which has resulted in a loss of genes from diverse biological pathways, including those that act in DNA repair. DNA repair is essential to any living cell. A loss of these mechanisms invariably results in accumulation of mutations and/or cell death. Six major pathways of DNA repair in eukaryotes include: non-homologous end joining (NHEJ, homologous recombination repair (HRR, mismatch repair (MMR, nucleotide excision repair (NER, base excision repair (BER and methyltransferase repair. DNA polymerases are also critical players in DNA repair processes. Given the close relationship between microsporidia and fungi, the repair mechanisms present in E. cuniculi were compared to those of the yeast Saccharomyces cerevisiae to ascertain how the process of genome reduction has affected the DNA repair pathways. Results E. cuniculi lacks 16 (plus another 6 potential absences of the 56 DNA repair genes sought via BLASTP and PSI-BLAST searches. Six of 14 DNA polymerases or polymerase subunits are also absent in E. cuniculi. All of these genes are relatively well conserved within eukaryotes. The absence of genes is not distributed equally among the different repair pathways; some pathways lack only one protein, while there is a striking absence of many proteins that are components of both double strand break repair pathways. All specialized repair polymerases are also absent. Conclusion Given the large number of DNA repair genes that are absent from the double strand break repair pathways, E. cuniculi is a prime candidate for the study of double strand break repair with minimal machinery. Strikingly, all of the

  13. Involvement of DNA repair in telomere maintenance and chromosomal instability in human cells

    International Nuclear Information System (INIS)

    Ayouaz, Ali

    2008-01-01

    Telomeres are a major actor of cell immortalization, precursor of a carcinogenesis process. Thus, it appears that the maintenance of telomeres is crucial in the implementation of carcinogenesis process. Due to their structures and under some conditions, telomeres can be assimilated in some respects to chromosomal breakages. Within this perspective, this research thesis aims at determining under which circumstances telomeres can be taken as targets by DNA repair mechanisms. More precisely, the author addressed the respective contributions of two repair mechanisms (the Non-Homologous End-Joining or NHEJ, and Homologous Recombination or HR) in the maintenance of telomere integrity. The author first discusses knowledge related to the interaction between chromosomal extremities and repair mechanisms. Then, he defines the behaviour of these mechanisms with respect to telomeres. He shows that, in absence of recombination mechanisms, the integrity of telomeres is not affected. Finally, he reports the attempt to determine their respective contributions in telomeric homeostasis [fr

  14. The DNA-dependent protein kinase: a multifunctional protein kinase with roles in DNA double strand break repair and mitosis

    Science.gov (United States)

    Jette, Nicholas; Lees-Miller, Susan P.

    2015-01-01

    The DNA-dependent protein kinase (DNA-PK) is a serine/threonine protein kinase composed of a large catalytic subunit (DNA-PKcs) and the Ku70/80 heterodimer. Over the past two decades, significant progress has been made in elucidating the role of DNA-PK in non-homologous end joining (NHEJ), the major pathway for repair of ionizing radiation-induced DNA double strand breaks in human cells and recently, additional roles for DNA-PK have been reported. In this review, we will describe the biochemistry, structure and function of DNA-PK, its roles in DNA double strand break repair and its newly described roles in mitosis and other cellular processes. PMID:25550082

  15. DNA repair in neurons: So if they don't divide what's to repair?

    Energy Technology Data Exchange (ETDEWEB)

    Fishel, Melissa L. [Department of Pediatrics (Section of Hematology/Oncology), Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 1044 W. Walnut, Room 302C, Indianapolis, IN 46202 (United States); Vasko, Michael R. [Department of Pharmacology and Toxicology, Indiana University School of Medicine, 1044 W. Walnut St., Indianapolis, IN 46202 (United States); Kelley, Mark R. [Department of Pediatrics (Section of Hematology/Oncology), Herman B Wells Center for Pediatric Research, Indiana University School of Medicine, 1044 W. Walnut, Room 302C, Indianapolis, IN 46202 (United States) and Department of Pharmacology and Toxicology, Indiana University School of Medicine, 1044 W. Walnut St., Indianapolis, IN 46202 (United States) and Department of Biochemistry and Molecular Biology, Indiana University School of Medicine, 1044 W. Walnut, Room 302C, Indianapolis, IN 46202 (United States)]. E-mail: mkelley@iupui.edu

    2007-01-03

    Neuronal DNA repair remains one of the most exciting areas for investigation, particularly as a means to compare the DNA repair response in mitotic (cancer) vs. post-mitotic (neuronal) cells. In addition, the role of DNA repair in neuronal cell survival and response to aging and environmental insults is of particular interest. DNA damage caused by reactive oxygen species (ROS) such as generated by mitochondrial respiration includes altered bases, abasic sites, and single- and double-strand breaks which can be prevented by the DNA base excision repair (BER) pathway. Oxidative stress accumulates in the DNA of the human brain over time especially in the mitochondrial DNA (mtDNA) and is proposed to play a critical role in aging and in the pathogenesis of several neurological disorders including Parkinson's disease, ALS, and Alzheimer's diseases. Because DNA damage accumulates in the mtDNA more than nuclear DNA, there is increased interest in DNA repair pathways and the consequence of DNA damage in the mitochondria of neurons. The type of damage that is most likely to occur in neuronal cells is oxidative DNA damage which is primarily removed by the BER pathway. Following the notion that the bulk of neuronal DNA damage is acquired by oxidative DNA damage and ROS, the BER pathway is a likely area of focus for neuronal studies of DNA repair. BER variations in brain aging and pathology in various brain regions and tissues are presented. Therefore, the BER pathway is discussed in greater detail in this review than other repair pathways. Other repair pathways including direct reversal, nucleotide excision repair (NER), mismatch repair (MMR), homologous recombination and non-homologous end joining are also discussed. Finally, there is a growing interest in the role that DNA repair pathways play in the clinical arena as they relate to the neurotoxicity and neuropathy associated with cancer treatments. Among the numerous side effects of cancer treatments, major

  16. Summary of Dissimilar Metal Joining Trials Conducted by Edison Welding Institute

    Energy Technology Data Exchange (ETDEWEB)

    MJ Lambert

    2005-11-18

    Under the direction of the NASA-Glenn Research Center, the Edison Welding Institute (EWI) in Columbus, OH performed a series of non-fusion joining experiments to determine the feasibility of joining refractory metals or refractory metal alloys to Ni-based superalloys. Results, as reported by EWI, can be found in the project report for EWI Project 48819GTH (Attachment A, at the end of this document), dated October 10, 2005. The three joining methods used in this investigation were inertia welding, magnetic pulse welding, and electro-spark deposition joining. Five materials were used in these experiments: Mo-47Re, T-111, Hastelloy X, Mar M-247 (coarse-grained, 0.5 mm to several millimeter average grain size), and Mar M-247 (fine-grained, approximately 50 {micro}m average grain size). Several iterative trials of each material combination with each joining method were performed to determine the best practice joining method. Mo-47Re was found to be joined easily to Hastelloy X via inertia welding, but inertia welding of the Mo-alloy to both Mar M-247 alloys resulted in inconsistent joint strength and large reaction layers between the two metals. T-111 was found to join well to Hastelloy X and coarse-grained Mar M-247 via inertia welding, but joining to fine-grained Mar M-247 resulted in low joint strength. Magnetic pulse welding (MPW) was only successful in joining T-111 tubing to Hastelloy X bar stock. The joint integrity and reaction layer between the metals were found to be acceptable. This single joining trial, however, caused damage to the electromagnetic concentrators used in this process. Subsequent design efforts to eliminate the problem resulted in a loss of power imparted to the accelerating work piece, and results could not be reproduced. Welding trials of Mar M-247 to T-111 resulted in catastrophic failure of the bar stock, even at lower power. Electro-spark deposition joining of Mo-47Re, in which the deposited material was Hastelloy X, did not have a

  17. Summary of Dissimilar Metal Joining Trials Conducted by Edison Welding Institute

    International Nuclear Information System (INIS)

    MJ Lambert

    2005-01-01

    Under the direction of the NASA-Glenn Research Center, the Edison Welding Institute (EWI) in Columbus, OH performed a series of non-fusion joining experiments to determine the feasibility of joining refractory metals or refractory metal alloys to Ni-based superalloys. Results, as reported by EWI, can be found in the project report for EWI Project 48819GTH (Attachment A, at the end of this document), dated October 10, 2005. The three joining methods used in this investigation were inertia welding, magnetic pulse welding, and electro-spark deposition joining. Five materials were used in these experiments: Mo-47Re, T-111, Hastelloy X, Mar M-247 (coarse-grained, 0.5 mm to several millimeter average grain size), and Mar M-247 (fine-grained, approximately 50 (micro)m average grain size). Several iterative trials of each material combination with each joining method were performed to determine the best practice joining method. Mo-47Re was found to be joined easily to Hastelloy X via inertia welding, but inertia welding of the Mo-alloy to both Mar M-247 alloys resulted in inconsistent joint strength and large reaction layers between the two metals. T-111 was found to join well to Hastelloy X and coarse-grained Mar M-247 via inertia welding, but joining to fine-grained Mar M-247 resulted in low joint strength. Magnetic pulse welding (MPW) was only successful in joining T-111 tubing to Hastelloy X bar stock. The joint integrity and reaction layer between the metals were found to be acceptable. This single joining trial, however, caused damage to the electromagnetic concentrators used in this process. Subsequent design efforts to eliminate the problem resulted in a loss of power imparted to the accelerating work piece, and results could not be reproduced. Welding trials of Mar M-247 to T-111 resulted in catastrophic failure of the bar stock, even at lower power. Electro-spark deposition joining of Mo-47Re, in which the deposited material was Hastelloy X, did not have a

  18. FANCD2 Binds CtIP and Regulates DNA-End Resection during DNA Interstrand Crosslink Repair

    Directory of Open Access Journals (Sweden)

    Junya Unno

    2014-05-01

    Full Text Available The Fanconi anemia (FA pathway is critically involved in the maintenance of hematopoietic stem cells and the suppression of carcinogenesis. A key FA protein, FANCD2, is monoubiquitinated and accumulates in chromatin in response to DNA interstrand crosslinks (ICLs, where it coordinates DNA repair through mechanisms that are still poorly understood. Here, we report that CtIP protein directly interacts with FANCD2. A region spanning amino acids 166 to 273 of CtIP and monoubiquitination of FANCD2 are both essential for the FANCD2-CtIP interaction and mitomycin C (MMC-induced CtIP foci. Remarkably, both FANCD2 and CtIP are critical for MMC-induced RPA2 hyperphosphorylation, an event that accompanies end resection of double-strand breaks. Collectively, our results reveal a role of monoubiquitinated FANCD2 in end resection that depends on its binding to CtIP during ICL repair.

  19. The Heterochromatic Barrier to DNA Double Strand Break Repair: How to Get the Entry Visa

    Directory of Open Access Journals (Sweden)

    Aaron A. Goodarzi

    2012-09-01

    Full Text Available Over recent decades, a deep understanding of pathways that repair DNA double strand breaks (DSB has been gained from biochemical, structural, biophysical and cellular studies. DNA non-homologous end-joining (NHEJ and homologous recombination (HR represent the two major DSB repair pathways, and both processes are now well understood. Recent work has demonstrated that the chromatin environment at a DSB significantly impacts upon DSB repair and that, moreover, dramatic modifications arise in the chromatin surrounding a DSB. Chromatin is broadly divided into open, transcriptionally active, euchromatin (EC and highly compacted, transcriptionally inert, heterochromatin (HC, although these represent extremes of a spectrum. The HC superstructure restricts both DSB repair and damage response signaling. Moreover, DSBs within HC (HC-DSBs are rapidly relocalized to the EC-HC interface. The damage response protein kinase, ataxia telangiectasia mutated (ATM, is required for HC-DSB repair but is dispensable for the relocalization of HC-DSBs. It has been proposed that ATM signaling enhances HC relaxation in the DSB vicinity and that this is a prerequisite for HC-DSB repair. Hence, ATM is essential for repair of HC-DSBs. Here, we discuss how HC impacts upon the response to DSBs and how ATM overcomes the barrier that HC poses to repair.

  20. More efficient repair of DNA double-strand breaks in skeletal muscle stem cells compared to their committed progeny

    Directory of Open Access Journals (Sweden)

    Leyla Vahidi Ferdousi

    2014-11-01

    Full Text Available The loss of genome integrity in adult stem cells results in accelerated tissue aging and is possibly cancerogenic. Adult stem cells in different tissues appear to react robustly to DNA damage. We report that adult skeletal stem (satellite cells do not primarily respond to radiation-induced DNA double-strand breaks (DSBs via differentiation and exhibit less apoptosis compared to other myogenic cells. Satellite cells repair these DNA lesions more efficiently than their committed progeny. Importantly, non-proliferating satellite cells and post-mitotic nuclei in the fiber exhibit dramatically distinct repair efficiencies. Altogether, reduction of the repair capacity appears to be more a function of differentiation than of the proliferation status of the muscle cell. Notably, satellite cells retain a high efficiency of DSB repair also when isolated from the natural niche. Finally, we show that repair of DSB substrates is not only very efficient but, surprisingly, also very accurate in satellite cells and that accurate repair depends on the key non-homologous end-joining factor DNA-PKcs.

  1. More efficient repair of DNA double-strand breaks in skeletal muscle stem cells compared to their committed progeny.

    Science.gov (United States)

    Vahidi Ferdousi, Leyla; Rocheteau, Pierre; Chayot, Romain; Montagne, Benjamin; Chaker, Zayna; Flamant, Patricia; Tajbakhsh, Shahragim; Ricchetti, Miria

    2014-11-01

    The loss of genome integrity in adult stem cells results in accelerated tissue aging and is possibly cancerogenic. Adult stem cells in different tissues appear to react robustly to DNA damage. We report that adult skeletal stem (satellite) cells do not primarily respond to radiation-induced DNA double-strand breaks (DSBs) via differentiation and exhibit less apoptosis compared to other myogenic cells. Satellite cells repair these DNA lesions more efficiently than their committed progeny. Importantly, non-proliferating satellite cells and post-mitotic nuclei in the fiber exhibit dramatically distinct repair efficiencies. Altogether, reduction of the repair capacity appears to be more a function of differentiation than of the proliferation status of the muscle cell. Notably, satellite cells retain a high efficiency of DSB repair also when isolated from the natural niche. Finally, we show that repair of DSB substrates is not only very efficient but, surprisingly, also very accurate in satellite cells and that accurate repair depends on the key non-homologous end-joining factor DNA-PKcs. Copyright © 2014. Published by Elsevier B.V.

  2. 40 CFR 61.243-2 - Alternative standards for valves in VHAP service-skip period leak detection and repair.

    Science.gov (United States)

    2010-07-01

    ... VHAP service-skip period leak detection and repair. 61.243-2 Section 61.243-2 Protection of Environment... AIR POLLUTANTS National Emission Standard for Equipment Leaks (Fugitive Emission Sources) § 61.243-2 Alternative standards for valves in VHAP service—skip period leak detection and repair. (a)(1) An owner or...

  3. Comparison of initial DNA (Chromosome) damage/repair in cells exposed to heavy ion particles and X-rays

    International Nuclear Information System (INIS)

    Okayasu, Ryuichi; Okada, Maki; Noguchi, Mitsuho; Saito, Shiori; Okabe, Atsushi; Takakura, Kahoru

    2005-01-01

    We have studied cell survival and chromosome damage/repair in normal and non homologous end-joining (NHEJ) deficient human cells exposed to carbon ions (290 MeV/u, ∼70 keV/um), iron ions (500 MeV/u, ∼200 keV/um) and X-rays. In order to examine the effect of heavy ion on double strand break (DSB) repair machinery, the auto-phosphorylation of DNA-PKcs was also investigated. The important discoveries made during this period are: 200 keV/um iron irradiation induced additional molecular damage beyond that 70 keV/um carbon did. Iron irradiation not only caused an inefficient G1 chromosome repair, but also induced non-repairable DSB/chromosome damage. The auto-phosphorylation of DNA-PKcs was significantly affected by high linear energy transfer (LET) irradiation when compared to X-rays. These results indicate NHEJ machinery was markedly disturbed by high LET radiation when compared to low LET radiation. (author)

  4. Small molecules, inhibitors of DNA-PK, targeting DNA repair and beyond

    Directory of Open Access Journals (Sweden)

    David eDavidson

    2013-01-01

    Full Text Available Many current chemotherapies function by damaging genomic DNA in rapidly dividing cells ultimately leading to cell death. This therapeutic approach differentially targets cancer cells that generally display rapid cell division compared to normal tissue cells. However, although these treatments are initially effective in arresting tumor growth and reducing tumor burden, resistance and disease progression eventually occur. A major mechanism underlying this resistance is increased levels of cellular DNA repair. Most cells have complex mechanisms in place to repair DNA damage that occurs due to environmental exposures or normal metabolic processes. These systems, initially overwhelmed when faced with chemotherapy induced DNA damage, become more efficient under constant selective pressure and as a result chemotherapies become less effective. Thus, inhibiting DNA repair pathways using target specific small molecule inhibitors may overcome cellular resistance to DNA damaging chemotherapies. Non-homologous end joining (NHEJ a major mechanism for the repair of double strand breaks (DSB in DNA is regulated in part by the serine/threonine kinase, DNA dependent protein kinase (DNA-PK. The DNA-PK holoenzyme acts as a scaffold protein tethering broken DNA ends and recruiting other repair molecules. It also has enzymatic activity that may be involved in DNA damage signaling. Because of its’ central role in repair of DSBs, DNA-PK has been the focus of a number of small molecule studies. In these studies specific DNA-PK inhibitors have shown efficacy in synergizing chemotherapies in vitro. However, compounds currently known to specifically inhibit DNA-PK are limited by poor pharmacokinetics: these compounds have poor solubility and have high metabolic lability in vivo leading to short serum half-lives. Future improvement in DNA-PK inhibition will likely be achieved by designing new molecules based on the recently reported crystallographic structure of DNA

  5. Loss of CHD1 causes DNA repair defects and enhances prostate cancer therapeutic responsiveness

    DEFF Research Database (Denmark)

    Kari, Vijayalakshmi; Mansour, Wael Yassin; Raul, Sanjay Kumar

    2016-01-01

    The CHD1 gene, encoding the chromo-domain helicase DNA-binding protein-1, is one of the most frequently deleted genes in prostate cancer. Here, we examined the role of CHD1 in DNA double-strand break (DSB) repair in prostate cancer cells. We show that CHD1 is required for the recruitment of Ct......-homologous end joining. Together, we provide evidence for a previously unknown role of CHD1 in DNA DSB repair via HR and show that CHD1 depletion sensitizes cells to PARP inhibitors, which has potential therapeutic relevance. Our findings suggest that CHD1 deletion, like BRCA1/2 mutation in ovarian cancer, may...... serve as a marker for prostate cancer patient stratification and the utilization of targeted therapies such as PARP inhibitors, which specifically target tumors with HR defects....

  6. Chrysler Upset Protrusion Joining Techniques for Joining Dissimilar Metals

    Energy Technology Data Exchange (ETDEWEB)

    Logan, Stephen [FCA US LLC, Auburn Hills, MI (United States)

    2017-09-28

    The project goal was to develop and demonstrate a robust, cost effective, and versatile joining technique, known as Upset Protrusion Joining (UPJ), for joining challenging dissimilar metal com-binations, especially those where one of the metals is a die cast magnesium (Mg) component. Since two of the key obstacles preventing more widespread use of light metals (especially in high volume automotive applications) are 1) a lack of robust joining techniques and 2) susceptibility to galvanic corrosion, and since the majority of the joint combinations evaluated in this project include die cast Mg (the lightest structural metal) as one of the two materials being joined, and since die casting is the most common and cost effective process for producing Mg components, then successful project completion provides a key enabler to high volume application of lightweight materials, thus potentially leading to reduced costs, and encouraging implementation of lightweight multi-material vehicles for significant reductions in energy consumption and reduced greenhouse gas emissions. Eco-nomic benefits to end-use consumers are achieved primarily via the reduction in fuel consumption. Unlike currently available commercial processes, the UPJ process relies on a very robust mechanical joint rather than intermetallic bonding, so the more cathodic material can be coated prior to joining, thus creating a robust isolation against galvanic attack on the more anodic material. Additionally, since the UPJ protrusion is going through a hole that can be pre-drilled or pre-punched prior to coating, the UPJ process is less likely to damage the coating when the joint is being made. Further-more, since there is no additional cathodic material (such as a steel fastener) used to create the joint, there is no joining induced galvanic activity beyond that of the two parent materials. In accordance with its originally proposed plan, this project has successfully developed process variants of UPJ to enable

  7. Differentiation of Human Induced Pluripotent or Embryonic Stem Cells Decreases the DNA Damage Repair by Homologous Recombination

    Directory of Open Access Journals (Sweden)

    Kalpana Mujoo

    2017-11-01

    Full Text Available The nitric oxide (NO-cyclic GMP pathway contributes to human stem cell differentiation, but NO free radical production can also damage DNA, necessitating a robust DNA damage response (DDR to ensure cell survival. How the DDR is affected by differentiation is unclear. Differentiation of stem cells, either inducible pluripotent or embryonic derived, increased residual DNA damage as determined by γ-H2AX and 53BP1 foci, with increased S-phase-specific chromosomal aberration after exposure to DNA-damaging agents, suggesting reduced homologous recombination (HR repair as supported by the observation of decreased HR-related repair factor foci formation (RAD51 and BRCA1. Differentiated cells also had relatively increased fork stalling and R-loop formation after DNA replication stress. Treatment with NO donor (NOC-18, which causes stem cell differentiation has no effect on double-strand break (DSB repair by non-homologous end-joining but reduced DSB repair by HR. Present studies suggest that DNA repair by HR is impaired in differentiated cells.

  8. Distinct roles of FANCO/RAD51C in DNA damage signaling and repair: implications for fanconi anemia and breast cancer susceptibility

    International Nuclear Information System (INIS)

    Nagaraju, G.; Somyajit, K.; Subramanya, S.

    2012-01-01

    Unrepaired or misrepaired chromosomal double-strand breaks (DSBs) can cause gross chromosomal rearrangements which eventually can lead to tumorigenesis through inactivation of tumor suppressor genes or activation of oncogenes. There are two major mechanisms of DSB repair: non-homologous end joining (NHEJ) and homologous recombination (HR). DSBs that are generated during S and G2 phase of the cell are preferentially repaired by sister chromatid recombination (SCR), an HR pathway that utilizes neighboring sister chromatid as a template. Since the copied information is accurate, SCR is potentially an error-free pathway. HR also plays a critical role in the repair of daughter strand gaps (DSGs) that arise as a result of replication fork stalling and facilitates replication fork recovery. Furthermore, in collaboration with nucleotide excision repair and translesion synthesis, HR is involved in the repair of DNA interstrand cross-links (ICLs). Thus, HR is important for the maintenance of genome integrity and its dysfunction can lead to various genetic disorders and cancer

  9. Transition piece for joining together tubular pieces

    International Nuclear Information System (INIS)

    Holko, K.H.

    1981-01-01

    A transition piece for joining together tubular pieces formed respectively from a low alloy or carbon steel and a high temperature alloy containing at least 16% chromium includes a plurality of tubular parts welded together and formed from materials of selected composition with a maximum chromium content difference of 5% between adjacent parts when the chromium content of each part is below 10% and a maximum chromium difference of 7% between adjacent parts when the chromium content of either part is above 10%. The transition parts are also graded as to such characteristics as thermal expansion coefficient. The transition parts at opposite ends of the transition joint have chromium percentages similar to the tubular pieces to which they are to be joined. The parts may be joined by fusion and/or friction welding and parts may be formed by fusion weld deposition. (author)

  10. Maintenance & Repair Cost Calculation and Assessment of Resale Value for Different Alternative Commercial Vehicle Powertrain Technologies

    OpenAIRE

    Kleiner, Florian; Friedrich, Horst E.

    2017-01-01

    For detailed evaluation of the Total Cost of Ownership, expenditures for Maintenance & Repair as well as the resale value are important to consider and should not be neglected. However, information on Maintenance & Repair costs as well as residual values for commercial vehicles with alternative powertrains is missing and data on this issue is rare. There is a lack of information and consolidated knowledge. In order to enable a holistic cost assessment for commercial vehicles, a comprehensive ...

  11. Joining of parts via magnetic heating of metal aluminum powders

    Science.gov (United States)

    Baker, Ian

    2013-05-21

    A method of joining at least two parts includes steps of dispersing a joining material comprising a multi-phase magnetic metal-aluminum powder at an interface between the at least two parts to be joined and applying an alternating magnetic field (AMF). The AMF has a magnetic field strength and frequency suitable for inducing magnetic hysteresis losses in the metal-aluminum powder and is applied for a period that raises temperature of the metal-aluminum powder to an exothermic transformation temperature. At the exothermic transformation temperature, the metal-aluminum powder melts and resolidifies as a metal aluminide solid having a non-magnetic configuration.

  12. NONPROCESS SOLVENT USE IN THE FURNITURE REFINISHING AND REPAIR INDUSTRY: EVALUATION OF ALTERNATIVE CHEMICAL STRIPPERS

    Science.gov (United States)

    The report gives results of an evaluation of the feasibility of using alternatives to high volatile organic compound/hazardous air pollutant (VOC/HAP) solvent-based, chemical strippers that are currently used in the furniture repair and refinishing industry to remove both traditi...

  13. An efficient system for deletion of large DNA fragments in Escherichia coli via introduction of both Cas9 and the non-homologous end joining system from Mycobacterium smegmatis.

    Science.gov (United States)

    Zheng, Xuan; Li, Shi-Yuan; Zhao, Guo-Ping; Wang, Jin

    2017-04-15

    Accompanied with the internal non-homologous end joining (NHEJ) system, Cas9 can be used to easily inactivate a gene or delete a fragment through introduction of DNA double-stranded breaks (DSBs) in eukaryotic cells. While in most prokaryotes (e.g. Escherichia coli), due to the lack of NHEJ, homologous recombination (HR) is required for repair of DSBs, which is less convenient. Here, a markerless system was developed for rapid gene inactivation or fragment deletion in E. coli via introduction of both Cas9 and a bacterial NHEJ system. Three bacterial NHEJ systems, i.e. Mycobacterium smegmatis (Msm), Mycobacterium tuberculosis (Mtb) and Bacillus subtilis (Bs), were tested in E. coli, and the MsmNHEJ system showed the best efficiency. With the employment of Cas9 and MsmNHEJ, we efficiently mutated lacZ gene, deleted glnALG operon and two large DNA fragments (67 kb and 123 kb) in E. coli, respectively. Moreover, the system was further designed to allow for continuous inactivation of genes or deletion of DNA fragments in E. coli. We envision this system can be extended to other bacteria, especially those with low HR efficiency. Copyright © 2017 Elsevier Inc. All rights reserved.

  14. Altered Hematopoiesis in Mice Lacking DNA Polymerase μ Is Due to Inefficient Double-Strand Break Repair

    Science.gov (United States)

    Lucas, Daniel; Escudero, Beatriz; Ligos, José Manuel; Segovia, Jose Carlos; Estrada, Juan Camilo; Terrados, Gloria; Blanco, Luis; Samper, Enrique; Bernad, Antonio

    2009-01-01

    Polymerase mu (Polμ) is an error-prone, DNA-directed DNA polymerase that participates in non-homologous end-joining (NHEJ) repair. In vivo, Polμ deficiency results in impaired Vκ-Jκ recombination and altered somatic hypermutation and centroblast development. In Polμ−/− mice, hematopoietic development was defective in several peripheral and bone marrow (BM) cell populations, with about a 40% decrease in BM cell number that affected several hematopoietic lineages. Hematopoietic progenitors were reduced both in number and in expansion potential. The observed phenotype correlates with a reduced efficiency in DNA double-strand break (DSB) repair in hematopoietic tissue. Whole-body γ-irradiation revealed that Polμ also plays a role in DSB repair in non-hematopoietic tissues. Our results show that Polμ function is required for physiological hematopoietic development with an important role in maintaining early progenitor cell homeostasis and genetic stability in hematopoietic and non-hematopoietic tissues. PMID:19229323

  15. Trex2 enables spontaneous sister chromatid exchanges without facilitating DNA double-strand break repair.

    Science.gov (United States)

    Dumitrache, Lavinia C; Hu, Lingchuan; Son, Mi Young; Li, Han; Wesevich, Austin; Scully, Ralph; Stark, Jeremy; Hasty, Paul

    2011-08-01

    Trex2 is a 3' → 5' exonuclease that removes 3'-mismatched sequences in a biochemical assay; however, its biological function remains unclear. To address biology we previously generated trex2(null) mouse embryonic stem (ES) cells and expressed in these cells wild-type human TREX2 cDNA (Trex2(hTX2)) or cDNA with a single-amino-acid change in the catalytic domain (Trex2(H188A)) or in the DNA-binding domain (Trex2(R167A)). We found the trex2(null) and Trex2(H188A) cells exhibited spontaneous broken chromosomes and trex2(null) cells exhibited spontaneous chromosomal rearrangements. We also found ectopically expressed human TREX2 was active at the 3' ends of I-SceI-induced chromosomal double-strand breaks (DSBs). Therefore, we hypothesized Trex2 participates in DNA DSB repair by modifying 3' ends. This may be especially important for ends with damaged nucleotides. Here we present data that are unexpected and prompt a new model. We found Trex2-altered cells (null, H188A, and R167A) were not hypersensitive to camptothecin, a type-1 topoisomerase inhibitor that induces DSBs at replication forks. In addition, Trex2-altered cells were not hypersensitive to γ-radiation, an agent that causes DSBs throughout the cell cycle. This observation held true even in cells compromised for one of the two major DSB repair pathways: homology-directed repair (HDR) or nonhomologous end joining (NHEJ). Trex2 deletion also enhanced repair of an I-SceI-induced DSB by both HDR and NHEJ without affecting pathway choice. Interestingly, however, trex2(null) cells exhibited reduced spontaneous sister chromatid exchanges (SCEs) but this was not due to a defect in HDR-mediated crossing over. Therefore, reduced spontaneous SCE could be a manifestation of the same defect that caused spontaneous broken chromosomes and spontaneous chromosomal rearrangements. These unexpected data suggest Trex2 does not enable DSB repair and prompt a new model that posits Trex2 suppresses the formation of broken

  16. A cell cycle-dependent regulatory circuit composed of 53BP1-RIF1 and BRCA1-CtIP controls DNA repair pathway choice.

    Science.gov (United States)

    Escribano-Díaz, Cristina; Orthwein, Alexandre; Fradet-Turcotte, Amélie; Xing, Mengtan; Young, Jordan T F; Tkáč, Ján; Cook, Michael A; Rosebrock, Adam P; Munro, Meagan; Canny, Marella D; Xu, Dongyi; Durocher, Daniel

    2013-03-07

    DNA double-strand break (DSB) repair pathway choice is governed by the opposing activities of 53BP1 and BRCA1. 53BP1 stimulates nonhomologous end joining (NHEJ), whereas BRCA1 promotes end resection and homologous recombination (HR). Here we show that 53BP1 is an inhibitor of BRCA1 accumulation at DSB sites, specifically in the G1 phase of the cell cycle. ATM-dependent phosphorylation of 53BP1 physically recruits RIF1 to DSB sites, and we identify RIF1 as the critical effector of 53BP1 during DSB repair. Remarkably, RIF1 accumulation at DSB sites is strongly antagonized by BRCA1 and its interacting partner CtIP. Lastly, we show that depletion of RIF1 is able to restore end resection and RAD51 loading in BRCA1-depleted cells. This work therefore identifies a cell cycle-regulated circuit, underpinned by RIF1 and BRCA1, that governs DSB repair pathway choice to ensure that NHEJ dominates in G1 and HR is favored from S phase onward. Copyright © 2013 Elsevier Inc. All rights reserved.

  17. RECQ HELICASE RECQL4 PARTICIPATES IN NON-HOMOLOGOUS END JOINING AND INTERACTS WITH THE KU COMPLEX

    DEFF Research Database (Denmark)

    Shamanna, Raghavendra A; Singh, Dharmendra Kumar; Lu, Huiming

    2014-01-01

    -irradiation and resulted in accumulation of 53BP1 foci after irradiation, indicating defects in the processing of DSB. We find that RECQL4 interacts with the Ku70/Ku80 heterodimer, part of the DNA-dependent protein kinase (DNA-PK) complex, via its N-terminal domain. Further, RECQL4 stimulates higher order DNA binding...... of Ku70/Ku80 to a blunt end DNA substrate. Taken together, these results implicate that RECQL4 participates in the NHEJ pathway of DSB repair via a functional interaction with the Ku70/Ku80 complex. This is the first study to provide both in vitro and in vivo evidence for a role of a RecQ helicase...

  18. DNA repair goes hip-hop: SMARCA and CHD chromatin remodellers join the break dance.

    Science.gov (United States)

    Rother, Magdalena B; van Attikum, Haico

    2017-10-05

    Proper signalling and repair of DNA double-strand breaks (DSB) is critical to prevent genome instability and diseases such as cancer. The packaging of DNA into chromatin, however, has evolved as a mere obstacle to these DSB responses. Posttranslational modifications and ATP-dependent chromatin remodelling help to overcome this barrier by modulating nucleosome structures and allow signalling and repair machineries access to DSBs in chromatin. Here we recap our current knowledge on how ATP-dependent SMARCA- and CHD-type chromatin remodellers alter chromatin structure during the signalling and repair of DSBs and discuss how their dysfunction impacts genome stability and human disease.This article is part of the themed issue 'Chromatin modifiers and remodellers in DNA repair and signalling'. © 2017 The Authors.

  19. DNA-PK, ATM and ATR collaboratively regulate p53-RPA interaction to facilitate homologous recombination DNA repair.

    Science.gov (United States)

    Serrano, M A; Li, Z; Dangeti, M; Musich, P R; Patrick, S; Roginskaya, M; Cartwright, B; Zou, Y

    2013-05-09

    Homologous recombination (HR) and nonhomologous end joining (NHEJ) are two distinct DNA double-stranded break (DSB) repair pathways. Here, we report that DNA-dependent protein kinase (DNA-PK), the core component of NHEJ, partnering with DNA-damage checkpoint kinases ataxia telangiectasia mutated (ATM) and ATM- and Rad3-related (ATR), regulates HR repair of DSBs. The regulation was accomplished through modulation of the p53 and replication protein A (RPA) interaction. We show that upon DNA damage, p53 and RPA were freed from a p53-RPA complex by simultaneous phosphorylations of RPA at the N-terminus of RPA32 subunit by DNA-PK and of p53 at Ser37 and Ser46 in a Chk1/Chk2-independent manner by ATR and ATM, respectively. Neither the phosphorylation of RPA nor of p53 alone could dissociate p53 and RPA. Furthermore, disruption of the release significantly compromised HR repair of DSBs. Our results reveal a mechanism for the crosstalk between HR repair and NHEJ through the co-regulation of p53-RPA interaction by DNA-PK, ATM and ATR.

  20. Identification of the DNA repair defects in a case of Dubowitz syndrome.

    Directory of Open Access Journals (Sweden)

    Jingyin Yue

    Full Text Available Dubowitz Syndrome is an autosomal recessive disorder with a unique set of clinical features including microcephaly and susceptibility to tumor formation. Although more than 140 cases of Dubowitz syndrome have been reported since 1965, the genetic defects of this disease has not been identified. In this study, we systematically analyzed the DNA damage response and repair capability of fibroblasts established from a Dubowitz Syndrome patient. Dubowitz syndrome fibroblasts are hypersensitive to ionizing radiation, bleomycin, and doxorubicin. However, they have relatively normal sensitivities to mitomycin-C, cisplatin, and camptothecin. Dubowitz syndrome fibroblasts also have normal DNA damage signaling and cell cycle checkpoint activations after DNA damage. These data implicate a defect in repair of DNA double strand break (DSB likely due to defective non-homologous end joining (NHEJ. We further sequenced several genes involved in NHEJ, and identified a pair of novel compound mutations in the DNA Ligase IV gene. Furthermore, expression of wild type DNA ligase IV completely complement the DNA repair defects in Dubowitz syndrome fibroblasts, suggesting that the DNA ligase IV mutation is solely responsible for the DNA repair defects. These data suggests that at least subset of Dubowitz syndrome can be attributed to DNA ligase IV mutations.

  1. Nrf2 facilitates repair of radiation induced DNA damage through homologous recombination repair pathway in a ROS independent manner in cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Jayakumar, Sundarraj; Pal, Debojyoti; Sandur, Santosh K., E-mail: sskumar@barc.gov.in

    2015-09-15

    Highlights: • Nrf2 inhibition in A549 cells led to attenuated DNA repair and radiosensitization. • Influence of Nrf2 on DNA repair is not linked to its antioxidant function. • Nrf2 influences DNA repair through homologous recombination (HR) repair pathway. • Many genes involved in HR pathway show ARE sequences in their upstream region. - Abstract: Nrf2 is a redox sensitive transcription factor that is involved in the co-ordinated transcription of genes involved in redox homeostasis. But the role of Nrf2 in DNA repair is not investigated in detail. We have employed A549 and MCF7 cells to study the role of Nrf2 on DNA repair by inhibiting Nrf2 using all-trans retinoic acid (ATRA) or by knock down approach prior to radiation exposure (4 Gy). DNA damage and repair analysis was studied by γH2AX foci formation and comet assay. Results suggested that the inhibition of Nrf2 in A549 or MCF7 cells led to significant slowdown in DNA repair as compared to respective radiation controls. The persistence of residual DNA damage even in the presence of free radical scavenger N-acetyl cysteine, suggested that the influence of Nrf2 on DNA repair was not linked to its antioxidant functions. Further, its influence on non-homologous end joining repair pathway was studied by inhibiting both Nrf2 and DNA-PK together. This led to synergistic reduction of survival fraction, indicating that Nrf2 may not be influencing the NHEJ pathway. To investigate the role of homologous recombination repair (HR) pathway, RAD51 foci formation was monitored. There was a significant reduction in the foci formation in cells treated with ATRA or shRNA against Nrf2 as compared to their respective radiation controls. Further, Nrf2 inhibition led to significant reduction in mRNA levels of RAD51. BLAST analysis was also performed on upstream regions of DNA repair genes to identify antioxidant response element and found that many repair genes that are involved in HR pathway may be regulated by Nrf2

  2. Functional intersection of ATM and DNA-dependent protein kinase catalytic subunit in coding end joining during V(D)J recombination

    DEFF Research Database (Denmark)

    Lee, Baeck-Seung; Gapud, Eric J; Zhang, Shichuan

    2013-01-01

    V(D)J recombination is initiated by the RAG endonuclease, which introduces DNA double-strand breaks (DSBs) at the border between two recombining gene segments, generating two hairpin-sealed coding ends and two blunt signal ends. ATM and DNA-dependent protein kinase catalytic subunit (DNA-PKcs) ar......V(D)J recombination is initiated by the RAG endonuclease, which introduces DNA double-strand breaks (DSBs) at the border between two recombining gene segments, generating two hairpin-sealed coding ends and two blunt signal ends. ATM and DNA-dependent protein kinase catalytic subunit (DNA......-PKcs) are serine-threonine kinases that orchestrate the cellular responses to DNA DSBs. During V(D)J recombination, ATM and DNA-PKcs have unique functions in the repair of coding DNA ends. ATM deficiency leads to instability of postcleavage complexes and the loss of coding ends from these complexes. DNA...... when ATM is present and its kinase activity is intact. The ability of ATM to compensate for DNA-PKcs kinase activity depends on the integrity of three threonines in DNA-PKcs that are phosphorylation targets of ATM, suggesting that ATM can modulate DNA-PKcs activity through direct phosphorylation of DNA...

  3. The DNA-dependent protein kinase: a multifunctional protein kinase with roles in DNA double strand break repair and mitosis

    OpenAIRE

    Jette, Nicholas; Lees-Miller, Susan P.

    2014-01-01

    The DNA-dependent protein kinase (DNA-PK) is a serine/threonine protein kinase composed of a large catalytic subunit (DNA-PKcs) and the Ku70/80 heterodimer. Over the past two decades, significant progress has been made in elucidating the role of DNA-PK in non-homologous end joining (NHEJ), the major pathway for repair of ionizing radiation-induced DNA double strand breaks in human cells and recently, additional roles for DNA-PK have been reported. In this review, we will describe the biochemi...

  4. Genomic survey and expression analysis of DNA repair genes in the genus Leptospira.

    Science.gov (United States)

    Martins-Pinheiro, Marinalva; Schons-Fonseca, Luciane; da Silva, Josefa B; Domingos, Renan H; Momo, Leonardo Hiroyuki Santos; Simões, Ana Carolina Quirino; Ho, Paulo Lee; da Costa, Renata M A

    2016-04-01

    Leptospirosis is an emerging zoonosis with important economic and public health consequences and is caused by pathogenic leptospires. The genus Leptospira belongs to the order Spirochaetales and comprises saprophytic (L. biflexa), pathogenic (L. interrogans) and host-dependent (L. borgpetersenii) members. Here, we present an in silico search for DNA repair pathways in Leptospira spp. The relevance of such DNA repair pathways was assessed through the identification of mRNA levels of some genes during infection in animal model and after exposition to spleen cells. The search was performed by comparison of available Leptospira spp. genomes in public databases with known DNA repair-related genes. Leptospires exhibit some distinct and unexpected characteristics, for instance the existence of a redundant mechanism for repairing a chemically diverse spectrum of alkylated nucleobases, a new mutS-like gene and a new shorter version of uvrD. Leptospira spp. shares some characteristics from Gram-positive, as the presence of PcrA, two RecQ paralogs and two SSB proteins; the latter is considered a feature shared by naturally competent bacteria. We did not find a significant reduction in the number of DNA repair-related genes in both pathogenic and host-dependent species. Pathogenic leptospires were enriched for genes dedicated to base excision repair and non-homologous end joining. Their evolutionary history reveals a remarkable importance of lateral gene transfer events for the evolution of the genus. Up-regulation of specific DNA repair genes, including components of SOS regulon, during infection in animal model validates the critical role of DNA repair mechanisms for the complex interplay between host/pathogen.

  5. A newly identified DNA ligase of Saccharomyces cerevisiae involved in RAD52-independent repair of DNA double-strand breaks

    Science.gov (United States)

    Schär, Primo; Herrmann, Gernot; Daly, Graham; Lindahl, Tomas

    1997-01-01

    Eukaryotic DNA ligases are ATP-dependent DNA strand-joining enzymes that participate in DNA replication, repair, and recombination. Whereas mammalian cells contain several different DNA ligases, encoded by at least three distinct genes, only one DNA ligase has been detected previously in either budding yeast or fission yeast. Here, we describe a newly identified nonessential Saccharomyces cerevisiae gene that encodes a DNA ligase distinct from the CDC9 gene product. This DNA ligase shares significant amino acid sequence homology with human DNA ligase IV; accordingly, we designate the yeast gene LIG4. Recombinant LIG4 protein forms a covalent enzyme-AMP complex and can join a DNA single-strand break in a DNA/RNA hybrid duplex, the preferred substrate in vitro. Disruption of the LIG4 gene causes only marginally increased cellular sensitivity to several DNA damaging agents, and does not further sensitize cdc9 or rad52 mutant cells. In contrast, lig4 mutant cells have a 1000-fold reduced capacity for correct recircularization of linearized plasmids by illegitimate end-joining after transformation. Moreover, homozygous lig4 mutant diploids sporulate less efficiently than isogenic wild-type cells, and show retarded progression through meiotic prophase I. Spore viability is normal, but lig4 mutants appear to produce a higher proportion of tetrads with only three viable spores. The mutant phenotypes are consistent with functions of LIG4 in an illegitimate DNA end-joining pathway and ensuring efficient meiosis. PMID:9271115

  6. In-Network Processing of an Iceberg Join Query in Wireless Sensor Networks Based on 2-Way Fragment Semijoins

    Directory of Open Access Journals (Sweden)

    Hyunchul Kang

    2015-03-01

    Full Text Available We investigate the in-network processing of an iceberg join query in wireless sensor networks (WSNs. An iceberg join is a special type of join where only those joined tuples whose cardinality exceeds a certain threshold (called iceberg threshold are qualified for the result. Processing such a join involves the value matching for the join predicate as well as the checking of the cardinality constraint for the iceberg threshold. In the previous scheme, the value matching is carried out as the main task for filtering non-joinable tuples while the iceberg threshold is treated as an additional constraint. We take an alternative approach, meeting the cardinality constraint first and matching values next. In this approach, with a logical fragmentation of the join operand relations on the aggregate counts of the joining attribute values, the optimal sequence of 2-way fragment semijoins is generated, where each fragment semijoin employs a Bloom filter as a synopsis of the joining attribute values. This sequence filters non-joinable tuples in an energy-efficient way in WSNs. Through implementation and a set of detailed experiments, we show that our alternative approach considerably outperforms the previous one.

  7. In-Network Processing of an Iceberg Join Query in Wireless Sensor Networks Based on 2-Way Fragment Semijoins

    Science.gov (United States)

    Kang, Hyunchul

    2015-01-01

    We investigate the in-network processing of an iceberg join query in wireless sensor networks (WSNs). An iceberg join is a special type of join where only those joined tuples whose cardinality exceeds a certain threshold (called iceberg threshold) are qualified for the result. Processing such a join involves the value matching for the join predicate as well as the checking of the cardinality constraint for the iceberg threshold. In the previous scheme, the value matching is carried out as the main task for filtering non-joinable tuples while the iceberg threshold is treated as an additional constraint. We take an alternative approach, meeting the cardinality constraint first and matching values next. In this approach, with a logical fragmentation of the join operand relations on the aggregate counts of the joining attribute values, the optimal sequence of 2-way fragment semijoins is generated, where each fragment semijoin employs a Bloom filter as a synopsis of the joining attribute values. This sequence filters non-joinable tuples in an energy-efficient way in WSNs. Through implementation and a set of detailed experiments, we show that our alternative approach considerably outperforms the previous one. PMID:25774710

  8. Precision Joining Center

    Science.gov (United States)

    Powell, J. W.; Westphal, D. A.

    1991-08-01

    A workshop to obtain input from industry on the establishment of the Precision Joining Center (PJC) was held on July 10-12, 1991. The PJC is a center for training Joining Technologists in advanced joining techniques and concepts in order to promote the competitiveness of U.S. industry. The center will be established as part of the DOE Defense Programs Technology Commercialization Initiative, and operated by EG&G Rocky Flats in cooperation with the American Welding Society and the Colorado School of Mines Center for Welding and Joining Research. The overall objectives of the workshop were to validate the need for a Joining Technologists to fill the gap between the welding operator and the welding engineer, and to assure that the PJC will train individuals to satisfy that need. The consensus of the workshop participants was that the Joining Technologist is a necessary position in industry, and is currently used, with some variation, by many companies. It was agreed that the PJC core curriculum, as presented, would produce a Joining Technologist of value to industries that use precision joining techniques. The advantage of the PJC would be to train the Joining Technologist much more quickly and more completely. The proposed emphasis of the PJC curriculum on equipment intensive and hands-on training was judged to be essential.

  9. Development of joining techniques for fabrication of fuel rod simulators

    International Nuclear Information System (INIS)

    Moorhead, A.J.; McCulloch, R.W.; Reed, R.W.; Woodhouse, J.J.

    1980-10-01

    Much of the safety-related thermal-hydraulic tests on nuclear reactors are conducted not in the reactor itself, but in mockup segments of a core that uses resistance-heated fuel rod simulators (FRS) in place of the radioactive fuel rods. Laser welding and furnace brazing techniques are described for joining subassemblies for FRS that have survived up to 1000 h steady-state operation at 700 to 1100 0 C cladding temperatures and over 5000 thermal transients, ranging from 10 to 100 0 C/s. A pulsed-laser welding procedure that includes use of small-diameter filler wire is used to join one end of a resistance heating element of Pt-8 W, Fe-22 Cr-5.5 Al-0.5 Co, or 80 Ni-20 Cr (wt %) to a tubular conductor of an appropriate intermediate material. The other end of the heating element is laser welded to an end plug, which in turn is welded to a central conductor rod

  10. The two faces of plan repair

    NARCIS (Netherlands)

    Van der Krogt, R.P.J.; De Weerdt, M.M.

    2004-01-01

    Plan repair has two faces. Alternately, a plan repair method looks like a planning method, or looks like a method that does exactly the opposite, i.e., removing actions from a plan. We propose a general framework for plan repair that shows the relation between these two alternating steps. Any plan

  11. A quantitative model of the major pathways for radiation-induced DNA double-strand break repair

    International Nuclear Information System (INIS)

    Belov, O.V.; Krasavin, E.A.; Lyashko, M.S.; Batmunkh, M.; Sweilam, N.H.

    2014-01-01

    We have developed a model approach to simulate the major pathways of DNA double-strand break (DSB) repair in mammalian and human cells. The proposed model shows a possible mechanistic explanation of the basic regularities of DSB processing through the nonhomologous end-joining (NHEJ), homologous recombination (HR), and single-strand annealing (SSA). It reconstructs the time-courses of radiation-induced foci specific to particular repair processes including the major intermediate stages. The model is validated for ionizing radiations of a wide range of linear energy transfer (0.2-236 keV/μm) including a relatively broad spectrum of heavy ions. The appropriate set of reaction rate constants was suggested to satisfy the kinetics of DSB rejoining for the considered types of exposure. The simultaneous assessment of three repair pathways allows one to describe their possible biological relations in response to radiation. With the help of the proposed approach, we reproduce several experimental data sets on γ-H2AX foci remaining in different types of cells including those defective in NHEJ, HR, or SSA functions.

  12. A new incomplete-repair model based on a ''reciprocal-time'' pattern of sublethal damage repair

    International Nuclear Information System (INIS)

    Dale, R.G.; Fowler, J.F.

    1999-01-01

    A radiobiological model for closely spaced non-instantaneous radiation fractions is presented, based on the premise that the time process of sublethal damage (SLD) repair is 'reciprocal-time' (second order), rather than exponential (first order), in form. The initial clinical implications of such an incomplete-repair model are assessed. A previously derived linear-quadratic-based model was revised to take account of the possibility that SLD may repair with time such that the fraction of an element of initial damage remaining at time t is given as 1/(1+zt), where z is an appropriate rate constant; z is the reciprocal of the first half-time (τ) of repair. The general equation so derived for incomplete repair is applicable to all types of radiotherapy delivered at high, low and medium dose-rate in fractions delivered at regular time intervals. The model allows both the fraction duration and interfraction intervals to vary between zero and infinity. For any given value of z, reciprocal repair is associated with an apparent 'slowing-down' in the SLD repair rate as treatment proceeds. The instantaneous repair rates are not directly governed by total dose or dose per fraction, but are influenced by the treatment duration and individual fraction duration. Instantaneous repair rates of SLD appear to be slower towards the end of a continuous treatment, and are also slower following 'long' fractions than they are following 'short' fractions. The new model, with its single repair-rate parameter, is shown to be capable of providing a degree of quantitative explanation for some enigmas that have been encountered in clinical studies. A single-component reciprocal repair process provides an alternative explanation for the apparent existence of a range of repair rates in human tissues, and which have hitherto been explained by postulating the existence of a multi-exponential repair process. The build-up of SLD over extended treatments is greater than would be inferred using a

  13. The indirect effect of radiation reduces the repair fidelity of NHEJ as verified in repair deficient CHO cell lines exposed to different radiation qualities and potassium bromate

    International Nuclear Information System (INIS)

    Bajinskis, Ainars; Olsson, Gunilla; Harms-Ringdahl, Mats

    2012-01-01

    The complexity of DNA lesions induced by ionizing radiation is mainly dependent on radiation quality, where the indirect action of radiation may contribute to different extent depending on the type of radiation under study. The effect of indirect action of radiation can be investigated by using agents that induce oxidative DNA damage or by applying free radical scavengers. The aim of this study was to investigate the role of the indirect effect of radiation for the repair fidelity of non-homologous end-joining (NHEJ), homologous recombination repair (HRR) and base excision repair (BER) when DNA damage of different complexity was induced by gamma radiation, alpha particles or from base damages (8-oxo-dG) induced by potassium bromate (KBrO 3 ). CHO cells lines deficient in XRCC3 (HRR) irs1SF, XRCC7 (NHEJ) V3-3 and XRCC1 (BER) EM9 were irradiated in the absence or presence of the free radical scavenger dimethyl sulfoxide (DMSO). The endpoints investigated included rate of cell proliferation by the DRAG assay, clonogenic cell survival and the level of primary DNA damage by the comet assay. The results revealed that the indirect effect of low-LET radiation significantly reduced the repair fidelity of both NHEJ and HRR pathways. For high-LET radiation the indirect effect of radiation also significantly reduced the repair fidelity for the repair deficient cell lines. The results suggest further that the repair fidelity of the error prone NHEJ repair pathway is more impaired by the indirect effect of high-LET radiation relative to the other repair pathways studied. The response to bromate observed for the two DSB repair deficient cell lines strongly support earlier studies that bromate induces complex DNA damages. The significantly reduced repair fidelity of irs1SF and V3-3 suggests that NHEJ as well as HRR are needed for the repair, and that complex DSBs are formed after bromate exposure.

  14. The indirect effect of radiation reduces the repair fidelity of NHEJ as verified in repair deficient CHO cell lines exposed to different radiation qualities and potassium bromate.

    Science.gov (United States)

    Bajinskis, Ainars; Olsson, Gunilla; Harms-Ringdahl, Mats

    2012-03-01

    The complexity of DNA lesions induced by ionizing radiation is mainly dependent on radiation quality, where the indirect action of radiation may contribute to different extent depending on the type of radiation under study. The effect of indirect action of radiation can be investigated by using agents that induce oxidative DNA damage or by applying free radical scavengers. The aim of this study was to investigate the role of the indirect effect of radiation for the repair fidelity of non-homologous end-joining (NHEJ), homologous recombination repair (HRR) and base excision repair (BER) when DNA damage of different complexity was induced by gamma radiation, alpha particles or from base damages (8-oxo-dG) induced by potassium bromate (KBrO(3)). CHO cells lines deficient in XRCC3 (HRR) irs1SF, XRCC7 (NHEJ) V3-3 and XRCC1 (BER) EM9 were irradiated in the absence or presence of the free radical scavenger dimethyl sulfoxide (DMSO). The endpoints investigated included rate of cell proliferation by the DRAG assay, clonogenic cell survival and the level of primary DNA damage by the comet assay. The results revealed that the indirect effect of low-LET radiation significantly reduced the repair fidelity of both NHEJ and HRR pathways. For high-LET radiation the indirect effect of radiation also significantly reduced the repair fidelity for the repair deficient cell lines. The results suggest further that the repair fidelity of the error prone NHEJ repair pathway is more impaired by the indirect effect of high-LET radiation relative to the other repair pathways studied. The response to bromate observed for the two DSB repair deficient cell lines strongly support earlier studies that bromate induces complex DNA damages. The significantly reduced repair fidelity of irs1SF and V3-3 suggests that NHEJ as well as HRR are needed for the repair, and that complex DSBs are formed after bromate exposure. Copyright © 2011 Elsevier B.V. All rights reserved.

  15. The indirect effect of radiation reduces the repair fidelity of NHEJ as verified in repair deficient CHO cell lines exposed to different radiation qualities and potassium bromate

    Energy Technology Data Exchange (ETDEWEB)

    Bajinskis, Ainars, E-mail: ainars.bajinskis@gmt.su.se [Centre for Radiation Protection Research, Department of Genetics, Microbiology and Toxicology, Stockholm University, S-10691 Stockholm (Sweden); Olsson, Gunilla; Harms-Ringdahl, Mats [Centre for Radiation Protection Research, Department of Genetics, Microbiology and Toxicology, Stockholm University, S-10691 Stockholm (Sweden)

    2012-03-01

    The complexity of DNA lesions induced by ionizing radiation is mainly dependent on radiation quality, where the indirect action of radiation may contribute to different extent depending on the type of radiation under study. The effect of indirect action of radiation can be investigated by using agents that induce oxidative DNA damage or by applying free radical scavengers. The aim of this study was to investigate the role of the indirect effect of radiation for the repair fidelity of non-homologous end-joining (NHEJ), homologous recombination repair (HRR) and base excision repair (BER) when DNA damage of different complexity was induced by gamma radiation, alpha particles or from base damages (8-oxo-dG) induced by potassium bromate (KBrO{sub 3}). CHO cells lines deficient in XRCC3 (HRR) irs1SF, XRCC7 (NHEJ) V3-3 and XRCC1 (BER) EM9 were irradiated in the absence or presence of the free radical scavenger dimethyl sulfoxide (DMSO). The endpoints investigated included rate of cell proliferation by the DRAG assay, clonogenic cell survival and the level of primary DNA damage by the comet assay. The results revealed that the indirect effect of low-LET radiation significantly reduced the repair fidelity of both NHEJ and HRR pathways. For high-LET radiation the indirect effect of radiation also significantly reduced the repair fidelity for the repair deficient cell lines. The results suggest further that the repair fidelity of the error prone NHEJ repair pathway is more impaired by the indirect effect of high-LET radiation relative to the other repair pathways studied. The response to bromate observed for the two DSB repair deficient cell lines strongly support earlier studies that bromate induces complex DNA damages. The significantly reduced repair fidelity of irs1SF and V3-3 suggests that NHEJ as well as HRR are needed for the repair, and that complex DSBs are formed after bromate exposure.

  16. Mutagenic effects of carbon ions near the range end in plants

    Energy Technology Data Exchange (ETDEWEB)

    Hase, Yoshihiro, E-mail: hase.yoshihiro@jaea.go.jp [Ion Beam Mutagenesis Research Group, Quantum Beam Science Directorate, Japan Atomic Energy Agency, 1233 Watanuki, Takasaki, Gunma 370-1292 (Japan); Yoshihara, Ryouhei; Nozawa, Shigeki; Narumi, Issay [Ion Beam Mutagenesis Research Group, Quantum Beam Science Directorate, Japan Atomic Energy Agency, 1233 Watanuki, Takasaki, Gunma 370-1292 (Japan)

    2012-03-01

    To gain insight into the mutagenic effects of accelerated heavy ions in plants, the mutagenic effects of carbon ions near the range end (mean linear energy transfer (LET): 425 keV/{mu}m) were compared with the effects of carbon ions penetrating the seeds (mean LET: 113 keV/{mu}m). Mutational analysis by plasmid rescue of Escherichia coli rpsL from irradiated Arabidopsis plants showed a 2.7-fold increase in mutant frequency for 113 keV/{mu}m carbon ions, whereas no enhancement of mutant frequency was observed for carbon ions near the range end. This suggested that carbon ions near the range end induced mutations that were not recovered by plasmid rescue. An Arabidopsis DNA ligase IV mutant, deficient in non-homologous end-joining repair, showed hyper-sensitivity to both types of carbon-ion irradiation. The difference in radiation sensitivity between the wild type and the repair-deficient mutant was greatly diminished for carbon ions near the range end, suggesting that these ions induce irreparable DNA damage. Mutational analysis of the Arabidopsis GL1 locus showed that while the frequency of generation of glabrous mutant sectors was not different between the two types of carbon-ion irradiation, large deletions (>{approx}30 kb) were six times more frequently induced by carbon ions near the range end. When 352 keV/{mu}m neon ions were used, these showed a 6.4 times increase in the frequency of induced large deletions compared with the 113 keV/{mu}m carbon ions. We suggest that the proportion of large deletions increases with LET in plants, as has been reported for mammalian cells. The nature of mutations induced in plants by carbon ions near the range end is discussed in relation to mutation detection by plasmid rescue and transmissibility to progeny.

  17. Direct and inverted repeats elicit genetic instability by both exploiting and eluding DNA double-strand break repair systems in mycobacteria.

    Directory of Open Access Journals (Sweden)

    Ewelina A Wojcik

    Full Text Available Repetitive DNA sequences with the potential to form alternative DNA conformations, such as slipped structures and cruciforms, can induce genetic instability by promoting replication errors and by serving as a substrate for DNA repair proteins, which may lead to DNA double-strand breaks (DSBs. However, the contribution of each of the DSB repair pathways, homologous recombination (HR, non-homologous end-joining (NHEJ and single-strand annealing (SSA, to this sort of genetic instability is not fully understood. Herein, we assessed the genome-wide distribution of repetitive DNA sequences in the Mycobacterium smegmatis, Mycobacterium tuberculosis and Escherichia coli genomes, and determined the types and frequencies of genetic instability induced by direct and inverted repeats, both in the presence and in the absence of HR, NHEJ, and SSA. All three genomes are strongly enriched in direct repeats and modestly enriched in inverted repeats. When using chromosomally integrated constructs in M. smegmatis, direct repeats induced the perfect deletion of their intervening sequences ~1,000-fold above background. Absence of HR further enhanced these perfect deletions, whereas absence of NHEJ or SSA had no influence, suggesting compromised replication fidelity. In contrast, inverted repeats induced perfect deletions only in the absence of SSA. Both direct and inverted repeats stimulated excision of the constructs from the attB integration sites independently of HR, NHEJ, or SSA. With episomal constructs, direct and inverted repeats triggered DNA instability by activating nucleolytic activity, and absence of the DSB repair pathways (in the order NHEJ>HR>SSA exacerbated this instability. Thus, direct and inverted repeats may elicit genetic instability in mycobacteria by 1 directly interfering with replication fidelity, 2 stimulating the three main DSB repair pathways, and 3 enticing L5 site-specific recombination.

  18. Direct and inverted repeats elicit genetic instability by both exploiting and eluding DNA double-strand break repair systems in mycobacteria.

    Science.gov (United States)

    Wojcik, Ewelina A; Brzostek, Anna; Bacolla, Albino; Mackiewicz, Pawel; Vasquez, Karen M; Korycka-Machala, Malgorzata; Jaworski, Adam; Dziadek, Jaroslaw

    2012-01-01

    Repetitive DNA sequences with the potential to form alternative DNA conformations, such as slipped structures and cruciforms, can induce genetic instability by promoting replication errors and by serving as a substrate for DNA repair proteins, which may lead to DNA double-strand breaks (DSBs). However, the contribution of each of the DSB repair pathways, homologous recombination (HR), non-homologous end-joining (NHEJ) and single-strand annealing (SSA), to this sort of genetic instability is not fully understood. Herein, we assessed the genome-wide distribution of repetitive DNA sequences in the Mycobacterium smegmatis, Mycobacterium tuberculosis and Escherichia coli genomes, and determined the types and frequencies of genetic instability induced by direct and inverted repeats, both in the presence and in the absence of HR, NHEJ, and SSA. All three genomes are strongly enriched in direct repeats and modestly enriched in inverted repeats. When using chromosomally integrated constructs in M. smegmatis, direct repeats induced the perfect deletion of their intervening sequences ~1,000-fold above background. Absence of HR further enhanced these perfect deletions, whereas absence of NHEJ or SSA had no influence, suggesting compromised replication fidelity. In contrast, inverted repeats induced perfect deletions only in the absence of SSA. Both direct and inverted repeats stimulated excision of the constructs from the attB integration sites independently of HR, NHEJ, or SSA. With episomal constructs, direct and inverted repeats triggered DNA instability by activating nucleolytic activity, and absence of the DSB repair pathways (in the order NHEJ>HR>SSA) exacerbated this instability. Thus, direct and inverted repeats may elicit genetic instability in mycobacteria by 1) directly interfering with replication fidelity, 2) stimulating the three main DSB repair pathways, and 3) enticing L5 site-specific recombination.

  19. Genetic variations in DNA repair genes, radiosensitivity to cancer and susceptibility to acute tissue reactions in radiotherapy-treated cancer patients

    International Nuclear Information System (INIS)

    Chistiakov, Dimitry A.; Voronova, Natalia V.; Chistiakov, Pavel A.

    2008-01-01

    Ionizing radiation is a well established carcinogen for human cells. At low doses, radiation exposure mainly results in generation of double strand breaks (DSBs). Radiation-related DSBs could be directly linked to the formation of chromosomal rearrangements as has been proven for radiation-induced thyroid tumors. Repair of DSBs presumably involves two main pathways, non-homologous end joining (NHEJ) and homologous recombination (HR). A number of known inherited syndromes, such as ataxia telangiectasia, ataxia-telangiectasia like-disorder, radiosensitive severe combined immunodeficiency, Nijmegen breakage syndrome, and LIG4 deficiency are associated with increased radiosensitivity and/or cancer risk. Many of them are caused by mutations in DNA repair genes. Recent studies also suggest that variations in the DNA repair capacity in the general population may influence cancer susceptibility. In this paper, we summarize the current status of DNA repair proteins as potential targets for radiation-induced cancer risk. We will focus on genetic alterations in genes involved in HR- and NHEJ-mediated repair of DSBs, which could influence predisposition to radiation-related cancer and thereby explain interindividual differences in radiosensitivity or radioresistance in a general population

  20. Genetic variations in DNA repair genes, radiosensitivity to cancer and susceptibility to acute tissue reactions in radiotherapy-treated cancer patients

    Energy Technology Data Exchange (ETDEWEB)

    Chistiakov, Dimitry A. (Dept. of Pathology, Univ. of Pittsburgh, Pittsburgh (US)); Voronova, Natalia V. (Dept. of Molecular Diagnostics, National Research Center GosNIIgenetika, Moscow (RU)); Chistiakov, Pavel A. (Dept. of Radiology, Cancer Research Center, Moscow (RU))

    2008-06-15

    Ionizing radiation is a well established carcinogen for human cells. At low doses, radiation exposure mainly results in generation of double strand breaks (DSBs). Radiation-related DSBs could be directly linked to the formation of chromosomal rearrangements as has been proven for radiation-induced thyroid tumors. Repair of DSBs presumably involves two main pathways, non-homologous end joining (NHEJ) and homologous recombination (HR). A number of known inherited syndromes, such as ataxia telangiectasia, ataxia-telangiectasia like-disorder, radiosensitive severe combined immunodeficiency, Nijmegen breakage syndrome, and LIG4 deficiency are associated with increased radiosensitivity and/or cancer risk. Many of them are caused by mutations in DNA repair genes. Recent studies also suggest that variations in the DNA repair capacity in the general population may influence cancer susceptibility. In this paper, we summarize the current status of DNA repair proteins as potential targets for radiation-induced cancer risk. We will focus on genetic alterations in genes involved in HR- and NHEJ-mediated repair of DSBs, which could influence predisposition to radiation-related cancer and thereby explain interindividual differences in radiosensitivity or radioresistance in a general population

  1. DNA repair systems and the pathogenesis of Mycobacterium tuberculosis: varying activities at different stages of infection.

    Science.gov (United States)

    Gorna, Alina E; Bowater, Richard P; Dziadek, Jaroslaw

    2010-05-25

    Mycobacteria, including most of all MTB (Mycobacterium tuberculosis), cause pathogenic infections in humans and, during the infectious process, are exposed to a range of environmental insults, including the host's immune response. From the moment MTB is exhaled by infected individuals, through an active and latent phase in the body of the new host, until the time they reach the reactivation stage, MTB is exposed to many types of DNA-damaging agents. Like all cellular organisms, MTB has efficient DNA repair systems, and these are believed to play essential roles in mycobacterial pathogenesis. As different stages of infection have great variation in the conditions in which mycobacteria reside, it is possible that different repair systems are essential for progression to specific phases of infection. MTB possesses homologues of DNA repair systems that are found widely in other species of bacteria, such as nucleotide excision repair, base excision repair and repair by homologous recombination. MTB also possesses a system for non-homologous end-joining of DNA breaks, which appears to be widespread in prokaryotes, although its presence is sporadic within different species within a genus. However, MTB does not possess homologues of the typical mismatch repair system that is found in most bacteria. Recent studies have demonstrated that DNA repair genes are expressed differentially at each stage of infection. In the present review, we focus on different DNA repair systems from mycobacteria and identify questions that remain in our understanding of how these systems have an impact upon the infection processes of these important pathogens.

  2. DNA repair and cytokines: TGF-beta, IL-6, and thrombopoietin as different biomarkers of radioresistance

    Directory of Open Access Journals (Sweden)

    Francesca Bianca Aiello

    2016-07-01

    Full Text Available Double strand breaks (DSBs induced by radiotherapy are highly cytotoxic lesions, leading to chromosomal aberrations and cell death. ATM-dependent DNA-damage response, non-homologous end joining, and homologous recombination pathways coordinately contribute to repairing DSBs in higher eukaryotes. It is known that the expression of DSB repair genes is increased in tumors which is one of the main reasons for radioresistance. The inhibition of DSB repair pathways may be useful to increase tumor cell radiosensitivity and may target stem cell-like cancer cells, known to be the most radioresistant tumor components. Commonly overexpressed in neoplastic cells, cytokines confer radioresistance by promoting proliferation, survival, invasion, and angiogenesis. Unfortunately, tumor irradiation increases the expression of various cytokines displaying these effects, including transforming growth factor-beta and interlukin-6. Recently the capabilities of these cytokines to support DNA repair pathways and the ATM-dependent DNA response have been demonstrated. Thrombopoietin, essential for megakaryopoiesis and very important for hematopoietic stem cell homeostasis, has also been found to promote DNA repair in a highly selective manner. These findings reveal a novel mechanism underlying cytokine-related radioresistance, which may be clinically relevant. Therapies targeting specific cytokines may be used to improve radiosensitivity. Specific inhibitors may be chosen in consideration of different tumor microenvironments. Thrombopoietin may be useful in fending off irradiation-induced loss of hematopoietic stem cells.

  3. DNA repair-related genes in sugarcane expressed sequence tags (ESTs

    Directory of Open Access Journals (Sweden)

    R.M.A. Costa

    2001-12-01

    Full Text Available There is much interest in the identification and characterization of genes involved in DNA repair because of their importance in the maintenance of the genome integrity. The high level of conservation of DNA repair genes means that these genetic elements may be used in phylogenetic studies as a source of information on the genetic origin and evolution of species. The mechanisms by which damaged DNA is repaired are well understood in bacteria, yeast and mammals, but much remains to be learned as regards plants. We identified genes involved in DNA repair mechanisms in sugarcane using a similarity search of the Brazilian Sugarcane Expressed Sequence Tag (SUCEST database against known sequences deposited in other public databases (National Center of Biotechnology Information (NCBI database and the Munich Information Center for Protein Sequences (MIPS Arabidopsis thaliana database. This search revealed that most of the various proteins involved in DNA repair in sugarcane are similar to those found in other eukaryotes. However, we also identified certain intriguing features found only in plants, probably due to the independent evolution of this kingdom. The DNA repair mechanisms investigated include photoreactivation, base excision repair, nucleotide excision repair, mismatch repair, non-homologous end joining, homologous recombination repair and DNA lesion tolerance. We report the main differences found in the DNA repair machinery in plant cells as compared to other organisms. These differences point to potentially different strategies plants employ to deal with DNA damage, that deserve further investigation.A identificação e caracterização de genes envolvidos com reparo de DNA são de grande interesse, dada a sua importância na manutenção da integridade genômica. Além disso, a alta conservação dos genes de reparo de DNA faz com que possam ser utilizados como fonte de informação no que diz respeito à origem e evolução das esp

  4. RNF138 joins the HR team

    DEFF Research Database (Denmark)

    Bekker-Jensen, Simon; Mailand, Niels

    2015-01-01

    Two studies show that the E3 ubiquitin ligase RNF138 is recruited to DNA double-strand break sites, where it ubiquitylates key repair factors to promote DNA-end resection and homologous recombination. These findings add insights into the multilayered regulatory mechanisms underlying DNA double-st...

  5. Ku80-deleted cells are defective at base excision repair

    International Nuclear Information System (INIS)

    Li, Han; Marple, Teresa; Hasty, Paul

    2013-01-01

    Graphical abstract: - Highlights: • Ku80-deleted cells are hypersensitive to ROS and alkylating agents. • Cells deleted for Ku80, but not Ku70 or Lig4, have reduced BER capacity. • OGG1 rescues hypersensitivity to H 2 O 2 and paraquat in Ku80-mutant cells. • Cells deleted for Ku80, but not Lig4, are defective at repairing AP sites. • Cells deleted for Ku80, but not Lig4 or Brca2 exon 27, exhibit increased PAR. - Abstract: Ku80 forms a heterodimer with Ku70, called Ku, that repairs DNA double-strand breaks (DSBs) via the nonhomologous end joining (NHEJ) pathway. As a consequence of deleting NHEJ, Ku80-mutant cells are hypersensitive to agents that cause DNA DSBs like ionizing radiation. Here we show that Ku80 deletion also decreased resistance to ROS and alkylating agents that typically cause base lesions and single-strand breaks (SSBs). This is unusual since base excision repair (BER), not NHEJ, typically repairs these types of lesions. However, we show that deletion of another NHEJ protein, DNA ligase IV (Lig4), did not cause hypersensitivity to these agents. In addition, the ROS and alkylating agents did not induce γ-H2AX foci that are diagnostic of DSBs. Furthermore, deletion of Ku80, but not Lig4 or Ku70, reduced BER capacity. Ku80 deletion also impaired BER at the initial lesion recognition/strand scission step; thus, involvement of a DSB is unlikely. Therefore, our data suggests that Ku80 deletion impairs BER via a mechanism that does not repair DSBs

  6. Ku80-deleted cells are defective at base excision repair

    Energy Technology Data Exchange (ETDEWEB)

    Li, Han [The University of Texas Health Science Center at San Antonio, The Institute of Biotechnology, The Department of Molecular Medicine, 15355 Lambda Drive, San Antonio, TX 78245-3207 (United States); Tumor Suppression Group, Spanish National Cancer Research Centre (CNIO), Madrid 28029 (Spain); Marple, Teresa [The University of Texas Health Science Center at San Antonio, The Institute of Biotechnology, The Department of Molecular Medicine, 15355 Lambda Drive, San Antonio, TX 78245-3207 (United States); Hasty, Paul, E-mail: hastye@uthscsa.edu [The University of Texas Health Science Center at San Antonio, The Institute of Biotechnology, The Department of Molecular Medicine, 15355 Lambda Drive, San Antonio, TX 78245-3207 (United States); Tumor Suppression Group, Spanish National Cancer Research Centre (CNIO), Madrid 28029 (Spain)

    2013-05-15

    Graphical abstract: - Highlights: • Ku80-deleted cells are hypersensitive to ROS and alkylating agents. • Cells deleted for Ku80, but not Ku70 or Lig4, have reduced BER capacity. • OGG1 rescues hypersensitivity to H{sub 2}O{sub 2} and paraquat in Ku80-mutant cells. • Cells deleted for Ku80, but not Lig4, are defective at repairing AP sites. • Cells deleted for Ku80, but not Lig4 or Brca2 exon 27, exhibit increased PAR. - Abstract: Ku80 forms a heterodimer with Ku70, called Ku, that repairs DNA double-strand breaks (DSBs) via the nonhomologous end joining (NHEJ) pathway. As a consequence of deleting NHEJ, Ku80-mutant cells are hypersensitive to agents that cause DNA DSBs like ionizing radiation. Here we show that Ku80 deletion also decreased resistance to ROS and alkylating agents that typically cause base lesions and single-strand breaks (SSBs). This is unusual since base excision repair (BER), not NHEJ, typically repairs these types of lesions. However, we show that deletion of another NHEJ protein, DNA ligase IV (Lig4), did not cause hypersensitivity to these agents. In addition, the ROS and alkylating agents did not induce γ-H2AX foci that are diagnostic of DSBs. Furthermore, deletion of Ku80, but not Lig4 or Ku70, reduced BER capacity. Ku80 deletion also impaired BER at the initial lesion recognition/strand scission step; thus, involvement of a DSB is unlikely. Therefore, our data suggests that Ku80 deletion impairs BER via a mechanism that does not repair DSBs.

  7. Fast and scalable inequality joins

    KAUST Repository

    Khayyat, Zuhair

    2016-09-07

    Inequality joins, which is to join relations with inequality conditions, are used in various applications. Optimizing joins has been the subject of intensive research ranging from efficient join algorithms such as sort-merge join, to the use of efficient indices such as (Formula presented.)-tree, (Formula presented.)-tree and Bitmap. However, inequality joins have received little attention and queries containing such joins are notably very slow. In this paper, we introduce fast inequality join algorithms based on sorted arrays and space-efficient bit-arrays. We further introduce a simple method to estimate the selectivity of inequality joins which is then used to optimize multiple predicate queries and multi-way joins. Moreover, we study an incremental inequality join algorithm to handle scenarios where data keeps changing. We have implemented a centralized version of these algorithms on top of PostgreSQL, a distributed version on top of Spark SQL, and an existing data cleaning system, Nadeef. By comparing our algorithms against well-known optimization techniques for inequality joins, we show our solution is more scalable and several orders of magnitude faster. © 2016 Springer-Verlag Berlin Heidelberg

  8. Repair by genetic recombination in bacteria: overview

    International Nuclear Information System (INIS)

    Howard-Flanders, P.

    1975-01-01

    DNA molecules that have been damaged in both strands at the same level are not subject to repair by excision but instead can be repaired through recombination with homologous molecules. Examples of two-strand damage include postreplication gaps opposite pyrimidine dimers, two-strand breaks produced by x-rays, and chemically induced interstrand cross-links. In ultraviolet-irradiated bacteria, and newly synthesized DNA is of length equal to the interdimer spacing. With continued incubation, this low-molecular-weight DNA is joined into high-molecular-weight chains (postreplication repair), a process associated with sister exchanges in bacteria. Recombination is initiated by pyrimidine dimers opposite postreplication gaps and by interstrand cross-links that have been cut by excision enzymes. The free ends at the resulting gaps presumably initiate the exchanges. Postreplication repair in Escherichia coli occurs in recB - and recC - but is greatly slowed in recF - mutants. RecB and recC are the structural genes for exonuclease V, which digests two-stranded DNA by releasing oligonucleotides first from one strand and then from the other. The postreplication sister exchanges in ultraviolet-irradiated bacteria result in the distribution of pyrimidine dimers between parental and daughter strands, indicating that long exchanges involving both strands of each duplex occur. The R1 restriction endonuclease from E. coli has been used to cut the DNA of a bacterial drug-resistance transfer factor with one nuclease-sensitive site, and also DNA from the frog Xenopus enriched for ribosomal 18S and 28S genes. The fragments were annealed with the cut plasmid DNA and ligated, producing a new larger plasmid carrying the eukaryotic rDNA and able to infect and replicate in E. coli

  9. Loss of Ubr2, an E3 ubiquitin ligase, leads to chromosome fragility and impaired homologous recombinational repair

    International Nuclear Information System (INIS)

    Ouyang, Yan; Kwon, Yong Tae; An, Jee Young; Eller, Danny; Tsai, S.-C.; Diaz-Perez, Silvia; Troke, Joshua J.; Teitell, Michael A.; Marahrens, York

    2006-01-01

    The N-end rule pathway of protein degradation targets proteins with destabilizing N-terminal residues. Ubr2 is one of the E3 ubiquitin ligases of the mouse N-end rule pathway. We have previously shown that Ubr2 -/- male mice are infertile, owing to the arrest of spermatocytes between the leptotene/zygotene and pachytene of meiosis I, the failure of chromosome pairing, and subsequent apoptosis. Here, we report that mouse fibroblast cells derived from Ubr2 -/- embryos display genome instability. The frequency of chromosomal bridges and micronuclei were much higher in Ubr2 -/- fibroblasts than in +/+ controls. Metaphase chromosome spreads from Ubr2 -/- cells revealed a high incidence of spontaneous chromosomal gaps, indicating chromosomal fragility. These fragile sites were generally replicated late in S phase. Ubr2 -/- cells were hypersensitive to mitomycin C, a DNA cross-linking agent, but displayed normal sensitivity to gamma-irradiation. A reporter assay showed that Ubr2 -/- cells are significantly impaired in the homologous recombination repair of a double strand break. In contrast, Ubr2 -/- cells appeared normal in an assay for non-homologous end joining. Our results therefore unveil the role of the ubiquitin ligase Ubr2 in maintaining genome integrity and in homologous recombination repair

  10. Loss of Ubr2, an E3 ubiquitin ligase, leads to chromosome fragility and impaired homologous recombinational repair

    Energy Technology Data Exchange (ETDEWEB)

    Ouyang, Yan [Department of Human Genetics, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Kwon, Yong Tae [Center for Pharmacogenetics and Department of Pharmaceutical Sciences, University of Pittsburgh, Pittsburgh, PA 15261 (United States); An, Jee Young [Center for Pharmacogenetics and Department of Pharmaceutical Sciences, University of Pittsburgh, Pittsburgh, PA 15261 (United States); Eller, Danny [Department of Human Genetics, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Tsai, S.-C. [Department of Human Genetics, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Diaz-Perez, Silvia [Department of Human Genetics, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Troke, Joshua J. [Department of Pathology and Laboratory Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Teitell, Michael A. [Department of Pathology and Laboratory Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States); Marahrens, York [Department of Human Genetics, David Geffen School of Medicine at UCLA, Los Angeles, CA 90095 (United States)]. E-mail: ymarahrens@mednet.ucla.edu

    2006-04-11

    The N-end rule pathway of protein degradation targets proteins with destabilizing N-terminal residues. Ubr2 is one of the E3 ubiquitin ligases of the mouse N-end rule pathway. We have previously shown that Ubr2{sup -/-} male mice are infertile, owing to the arrest of spermatocytes between the leptotene/zygotene and pachytene of meiosis I, the failure of chromosome pairing, and subsequent apoptosis. Here, we report that mouse fibroblast cells derived from Ubr2{sup -/-} embryos display genome instability. The frequency of chromosomal bridges and micronuclei were much higher in Ubr2{sup -/-} fibroblasts than in +/+ controls. Metaphase chromosome spreads from Ubr2{sup -/-} cells revealed a high incidence of spontaneous chromosomal gaps, indicating chromosomal fragility. These fragile sites were generally replicated late in S phase. Ubr2{sup -/-} cells were hypersensitive to mitomycin C, a DNA cross-linking agent, but displayed normal sensitivity to gamma-irradiation. A reporter assay showed that Ubr2{sup -/-} cells are significantly impaired in the homologous recombination repair of a double strand break. In contrast, Ubr2{sup -/-} cells appeared normal in an assay for non-homologous end joining. Our results therefore unveil the role of the ubiquitin ligase Ubr2 in maintaining genome integrity and in homologous recombination repair.

  11. Performance experiments with alternative advanced teleoperator control modes for a simulated solar maximum satellite repair

    Science.gov (United States)

    Das, H.; Zak, H.; Kim, W. S.; Bejczy, A. K.; Schenker, P. S.

    1992-01-01

    Experiments are described which were conducted at the JPL Advanced Teleoperator Lab to demonstrate and evaluate the effectiveness of various teleoperator control modes in the performance of a simulated Solar Max Satellite Repair (SMSR) task. THe SMSR was selected as a test because it is very rich in performance capability requirements and it actually has been performed by two EVA astronauts in the Space Shuttle Bay in 1984. The main subtasks are: thermal blanket removal; installation of a hinge attachment for electrical panel opening; opening of electrical panel; removal of electrical connectors; relining of cable bundles; replacement of electrical panel; securing parts and cables; re-mate electrical connectors; closing of electrical panel; and reinstating thermal blanket. The current performance experiments are limited to thermal blanket cutting, electrical panel unbolting and handling electrical bundles and connectors. In one formal experiment even different control modes were applied to the unbolting and reinsertion of electrical panel screws subtasks. The seven control modes are alternative combinations of manual position and rate control with force feedback and remote compliance referenced to force-torque sensor information. Force-torque sensor and end effector position data and task completion times were recorded for analysis and quantification of operator performance.

  12. Mycobacterium smegmatis Ku binds DNA without free ends.

    Science.gov (United States)

    Kushwaha, Ambuj K; Grove, Anne

    2013-12-01

    Ku is central to the non-homologous end-joining pathway of double-strand-break repair in all three major domains of life, with eukaryotic homologues being associated with more diversified roles compared with prokaryotic and archaeal homologues. Ku has a conserved central 'ring-shaped' core domain. While prokaryotic homologues lack the N- and C-terminal domains that impart functional diversity to eukaryotic Ku, analyses of Ku from certain prokaryotes such as Pseudomonas aeruginosa and Mycobacterium smegmatis have revealed the presence of distinct C-terminal extensions that modulate DNA-binding properties. We report in the present paper that the lysine-rich C-terminal extension of M. smegmatis Ku contacts the core protein domain as evidenced by an increase in DNA-binding affinity and a decrease in thermal stability and intrinsic tryptophan fluorescence upon its deletion. Ku deleted for this C-terminus requires free DNA ends for binding, but translocates to internal DNA sites. In contrast, full-length Ku can directly bind DNA without free ends, suggesting that this property is conferred by its C-terminus. Such binding to internal DNA sites may facilitate recruitment to sites of DNA damage. The results of the present study also suggest that extensions beyond the shared core domain may have independently evolved to expand Ku function.

  13. Numerical modelling in friction lap joining of aluminium alloy and carbon-fiber-reinforced-plastic sheets

    Science.gov (United States)

    Das, A.; Bang, H. S.; Bang, H. S.

    2018-05-01

    Multi-material combinations of aluminium alloy and carbon-fiber-reinforced-plastics (CFRP) have gained attention in automotive and aerospace industries to enhance fuel efficiency and strength-to-weight ratio of components. Various limitations of laser beam welding, adhesive bonding and mechanical fasteners make these processes inefficient to join metal and CFRP sheets. Friction lap joining is an alternative choice for the same. Comprehensive studies in friction lap joining of aluminium to CFRP sheets are essential and scare in the literature. The present work reports a combined theoretical and experimental study in joining of AA5052 and CFRP sheets using friction lap joining process. A three-dimensional finite element based heat transfer model is developed to compute the temperature fields and thermal cycles. The computed results are validated extensively with the corresponding experimentally measured results.

  14. Time-lapse crystallography snapshots of a double-strand break repair polymerase in action.

    Science.gov (United States)

    Jamsen, Joonas A; Beard, William A; Pedersen, Lars C; Shock, David D; Moon, Andrea F; Krahn, Juno M; Bebenek, Katarzyna; Kunkel, Thomas A; Wilson, Samuel H

    2017-08-15

    DNA polymerase (pol) μ is a DNA-dependent polymerase that incorporates nucleotides during gap-filling synthesis in the non-homologous end-joining pathway of double-strand break repair. Here we report time-lapse X-ray crystallography snapshots of catalytic events during gap-filling DNA synthesis by pol μ. Unique catalytic intermediates and active site conformational changes that underlie catalysis are uncovered, and a transient third (product) metal ion is observed in the product state. The product manganese coordinates phosphate oxygens of the inserted nucleotide and PP i . The product metal is not observed during DNA synthesis in the presence of magnesium. Kinetic analyses indicate that manganese increases the rate constant for deoxynucleoside 5'-triphosphate insertion compared to magnesium. The likely product stabilization role of the manganese product metal in pol μ is discussed. These observations provide insight on structural attributes of this X-family double-strand break repair polymerase that impact its biological function in genome maintenance.DNA polymerase (pol) μ functions in DNA double-strand break repair. Here the authors use time-lapse X-ray crystallography to capture the states of pol µ during the conversion from pre-catalytic to product complex and observe a third transiently bound metal ion in the product state.

  15. Machining, joining and modifications of advanced materials

    CERN Document Server

    Altenbach, Holm

    2016-01-01

    This book presents the latest advances in mechanical and materials engineering applied to the machining, joining and modification of modern engineering materials. The contributions cover the classical fields of casting, forming and injection moulding as representative manufacturing methods, whereas additive manufacturing methods (rapid prototyping and laser sintering) are treated as more innovative and recent technologies that are paving the way for the manufacturing of shapes and features that traditional methods are unable to deliver. The book also explores water jet cutting as an innovative cutting technology that avoids the heat build-up typical of classical mechanical cutting. It introduces readers to laser cutting as an alternative technology for the separation of materials, and to classical bonding and friction stir welding approaches in the context of joining technologies. In many cases, forming and machining technologies require additional post-treatment to achieve the required level of surface quali...

  16. Pharmacodynamic Assay Panel for Monitoring Phospho-Signaling Networks | Office of Cancer Clinical Proteomics Research

    Science.gov (United States)

    The DNA damage response (DDR) is a highly regulated signal transduction network that orchestrates the temporal and spatial organization of protein complexes required to repair (or tolerate) DNA damage (e.g., nucleotide excision repair, base excision repair, homologous recombination, non-homologous end joining, post-replication repair).

  17. Joining of advanced materials

    CERN Document Server

    Messler, Robert W

    1993-01-01

    Provides an unusually complete and readable compilation of the primary and secondary options for joining conventional materials in non-conventional ways. Provides unique coverage of adhesive bonding using both organic and inorganic adhesives, cements and mortars. Focuses on materials issues without ignoring issues related to joint design, production processing, quality assurance, process economics, and joining performance in service.Joining of advanced materials is a unique treatment of joining of both conventional and advanced metals andalloys, intermetallics, ceramics, glasses, polymers, a

  18. The Use of Explosive Forming for Fastening and Joining Structural and Pressure Components

    Science.gov (United States)

    Schroeder, J. W.

    1985-01-01

    Explosive expansion of tubes into tubesheets has been used for over 20 years in the fabrication and repair of shell and tube heat exchangers. The use of explosives to perform these expansions has offered several distinct advantages over other methods. First, the process is fast and economical and can be performed with minimal training of personnel. Secondly, explosive forming does not cause the deleterious metallurgical effects which often result from other forming operations. In addition, the process can be performed remotely without the need for sophisticated handling equipment. The expansion of tubes into tubesheets is only one of many possible fastening and joining applications for which explosive forming can be used to achieve highly successful results. The explosive forming process and where it has been used are described. In addition, some possible adaptations to other joining applications are identified and discussed.

  19. Joining by plastic deformation

    DEFF Research Database (Denmark)

    Mori, Ken-ichiro; Bay, Niels; Fratini, Livan

    2013-01-01

    As the scale and complexity of products such as aircraft and cars increase, demand for new functional processes to join mechanical parts grows. The use of plastic deformation for joining parts potentially offers improved accuracy, reliability and environmental safety as well as creating opportuni......As the scale and complexity of products such as aircraft and cars increase, demand for new functional processes to join mechanical parts grows. The use of plastic deformation for joining parts potentially offers improved accuracy, reliability and environmental safety as well as creating...

  20. Ceramic joining

    Energy Technology Data Exchange (ETDEWEB)

    Loehman, R.E. [Sandia National Lab., Albuquerque, NM (United States)

    1996-04-01

    This paper describes the relation between reactions at ceramic-metal interfaces and the development of strong interfacial bonds in ceramic joining. Studies on a number of systems are described, including silicon nitrides, aluminium nitrides, mullite, and aluminium oxides. Joints can be weakened by stresses such as thermal expansion mismatch. Ceramic joining is used in a variety of applications such as solid oxide fuel cells.

  1. Heterogenous mismatch-repair status in colorectal cancer

    DEFF Research Database (Denmark)

    Joost, Patrick; Veurink, Nynke; Holck, Susanne

    2014-01-01

    BACKGROUND: Immunohistochemical staining for mismatch repair proteins is efficient and widely used to identify mismatch repair defective tumors. The tumors typically show uniform and widespread loss of MMR protein staining. We identified and characterized colorectal cancers with alternative......, heterogenous mismatch repair protein staining in order to delineate expression patterns and underlying mechanisms. METHODS: Heterogenous staining patterns that affected at least one of the mismatch repair proteins MLH1, PMS2, MSH2 and MSH6 were identified in 14 colorectal cancers. Based on alternative....... CONCLUSIONS: Heterogenous mismatch repair status can be demonstrated in colorectal cancer. Though rare, attention to this phenomenon is recommended since it corresponds to differences in mismatch repair status that are relevant for correct classification. VIRTUAL SLIDES: The virtual slide(s) for this article...

  2. In Vitro Expansion of Bone Marrow Derived Mesenchymal Stem Cells Alters DNA Double Strand Break Repair of Etoposide Induced DNA Damage

    Directory of Open Access Journals (Sweden)

    Ian Hare

    2016-01-01

    Full Text Available Mesenchymal stem cells (MSCs are of interest for use in diverse cellular therapies. Ex vivo expansion 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 with in vitro passage, 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 passage in 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 passaged in 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 that ex vivo expansion of MSCs alters their ability to perform DSB repair, a necessary function for cells intended for transplantation.

  3. In Vitro Expansion of Bone Marrow Derived Mesenchymal Stem Cells Alters DNA Double Strand Break Repair of Etoposide Induced DNA Damage.

    Science.gov (United States)

    Hare, Ian; Gencheva, Marieta; Evans, Rebecca; Fortney, James; Piktel, Debbie; Vos, Jeffrey A; Howell, David; Gibson, Laura F

    2016-01-01

    Mesenchymal stem cells (MSCs) are of interest for use in diverse cellular therapies. Ex vivo expansion 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 with in vitro passage, 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 passage in 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 passaged in 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 that ex vivo expansion of MSCs alters their ability to perform DSB repair, a necessary function for cells intended for transplantation.

  4. Weld repair of creep damaged steels

    International Nuclear Information System (INIS)

    Croker, A.B.L.; Harrison, R.P.; Moss, C.J.

    1995-01-01

    A cooperative research centre project 'Welding of Thermally Modified Structures' was commenced in June 1993 with support from ANSTO, CSIRO, BHP, University of Wollongong and the CRC for Materials, Welding and Joining. The main aims of the project are to quantify the effects of performing repair welds on materials which have operated for extended periods at elevated temperature. Welding is an increasingly used method for performing repairs, replacements, retrofits and modifications to elevated temperature plant, however, the effects of these repairs on the ultimate life of a component are poorly understood. This paper presents details of the three ex-service materials chosen for the project, a carbon steel and two alloy steels. Work is also presented on development of new methods of assessing materials and components both destructively, along with new methods of modelling welded components in high temperature service. 6 figs, 3 tabs

  5. Joining SiC/SiC composites for fusion applications

    International Nuclear Information System (INIS)

    Henager, C.J.

    2007-01-01

    Full text of publication follows: The use of SiC-reinforced composites for fusion reactors or other nuclear applications will likely require some form of joining in order to form large structural or functional sections. Joints must be able to allow load transfer to the composite material so that the fiber reinforcements are able to carry their designed loads and the joint should not fail in a brittle manner. In addition, joints must be tolerant of radiation damage similar to the composite material to maintain the structural integrity of the joined section. Other requirements of interest are low activation joints and ease of joining application and processing, which are a difficult set of criteria to meet. Research at PNNL has been directed at high-strength joints using solid-state displacement reactions and pre-ceramic polymer joints that are easy to apply and that could be used in field repair situations. This research will report on the further development and testing of such joints using a double notch shear (DNS) specimen that is suitable for in-reactor testing. The results reveal that the solid-state joints are stronger than the polymer-based joints but require high-temperature, high-pressure processing. The polymer-based joints can be applied with reduced pressure and temperature processing compared to the solid-state joints. The polymer-based joints may have adequate strength for some applications where ease of application and in situ processing are required. Although irradiation testing is not reported in this study the use of the DNS specimen is discussed with regard to future neutron irradiations. (authors)

  6. Deficiency of double-strand DNA break repair does not impair Mycobacterium tuberculosis virulence in multiple animal models of infection.

    Science.gov (United States)

    Heaton, Brook E; Barkan, Daniel; Bongiorno, Paola; Karakousis, Petros C; Glickman, Michael S

    2014-08-01

    Mycobacterium tuberculosis persistence within its human host requires mechanisms to resist the effector molecules of host immunity, which exert their bactericidal effects through damaging pathogen proteins, membranes, and DNA. Substantial evidence indicates that bacterial pathogens, including M. tuberculosis, require DNA repair systems to repair the DNA damage inflicted by the host during infection, but the role of double-strand DNA break (DSB) repair systems is unclear. Double-strand DNA breaks are the most cytotoxic form of DNA damage and must be repaired for chromosome replication to proceed. M. tuberculosis elaborates three genetically distinct DSB repair systems: homologous recombination (HR), nonhomologous end joining (NHEJ), and single-strand annealing (SSA). NHEJ, which repairs DSBs in quiescent cells, may be particularly relevant to M. tuberculosis latency. However, very little information is available about the phenotype of DSB repair-deficient M. tuberculosis in animal models of infection. Here we tested M. tuberculosis strains lacking NHEJ (a Δku ΔligD strain), HR (a ΔrecA strain), or both (a ΔrecA Δku strain) in C57BL/6J mice, C3HeB/FeJ mice, guinea pigs, and a mouse hollow-fiber model of infection. We found no difference in bacterial load, histopathology, or host mortality between wild-type and DSB repair mutant strains in any model of infection. These results suggest that the animal models tested do not inflict DSBs on the mycobacterial chromosome, that other repair pathways can compensate for the loss of NHEJ and HR, or that DSB repair is not required for M. tuberculosis pathogenesis. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  7. A New Approach to Joining Dissimilar Ceramic Oxides for Chemical Sensors

    International Nuclear Information System (INIS)

    Zhuiykov, Serge

    2009-01-01

    Conventional joining of dissimilar oxides for sensing electrodes (SE) of chemical sensors has been pivotal to the development of various sensors and is vital to their further development. However, it is shown that the uncertainty (of a fundamental nature) in the properties of dissimilar oxides in SE causes the determination of their sensing characteristics to be ambiguous. Characteristics are different for such controlled parameters as pyrolysis temperature, crystal structure, particle's morphology and size, chemical and phase composition, the coefficient of thermal expansion (CTE), surface architecture, the bulk and surface stoichiometry and type and conductivity of additives. Here, we provide an alternative approach for joining dissimilar metal-oxides for chemical sensors SE. The approach relies on the development of at least one transient liquid oxide phase on the ceramic-SE interface. These results constitute key points relevant to selection oxides for joining, sintering temperatures and heating/cooling temperature rates.

  8. Inducible error-prone repair in Escherichia coli

    International Nuclear Information System (INIS)

    Sedgwick, S.G.

    1975-01-01

    A hypothesis that ultraviolet-induced mutagenesis arises from the induction of an error-prone mode of postreplication repair that requires the exrA + recA + genotype has been tested with alkaline sucrose gradient centrifugation coupled with assays of fixation determined by loss of photoreversibility. The inhibitor of protein synthesis, chloramphenicol, added before irradiation, prevented a small amount of postreplication repair and completely eliminated mutation fixation in E. coli WP2/sub s/ uvrA. However, chloramphenicol did not affect strand joining: in uvrA bacteria allowed 20 min of growth between irradiation and antibiotic treatment; in nonmutable uvrA exrA bacteria; and in urvA tif bacteria grown at 42 0 for 70 min before irradiation. These observations indicate that an inducible product is involved in a fraction of postreplication repair and is responsible for induced mutagenesis. (auth)

  9. Performance of electro-plated and joined components for divertor application

    International Nuclear Information System (INIS)

    Krauss, Wolfgang; Lorenz, Julia; Konys, Jürgen

    2013-01-01

    Highlights: • Active interlayers of Ni and Pd were electroplated on W to assist joining. • Demonstrator types of W-steel and W–W joints were successfully fabricated. • Diffusion processes increase operation temperature above brazing temperature. • Ni electro-plating is less sensitive to variation of deposition parameters than Pd. • Shear tests showed values in resistance comparable to those of commercial fillers. -- Abstract: A general challenge in divertor development, independently of design type and cooling medium water or helium, is the reliable and adapted joining of components. Depending on the design variants, the characteristics of the joints will be focused on functional or structural behavior to guarantee e.g. good thermal conductivity and sufficient mechanical strength. All variants will have in common that tungsten is the plasma facing material. Thus, material combinations to be joined will range from Cu base over steel to tungsten. Especially tungsten shows lacks in adapted joining due to its metallurgical behavior ranging from immiscibility over bad wetting up to brittle intermetallic phase formation. Joining assisted by electro-chemical deposition of functional and filler layers showed that encouraging progress was achieved in wetting applying nickel interlayers. Nickel proved to be a good reference material but alternative elements (e.g. Pd, Fe) may be more attractive in fusion to manufacture suitable joints. Replacing of Ni as activator element by Pd for W/W or W/steel joints was achieved and joining with Cu-filler was successfully performed. Manufactured joints were characterized applying metallurgical testing and SEM/EDX analyses demonstrating the applicability of Pd activator. First shear tests showed that the joints exhibit mechanical stability sufficient for technical application

  10. Plasma facing parts and repairing method

    International Nuclear Information System (INIS)

    Fuse, Toshiaki; Tachikawa, Nobuo.

    1994-01-01

    Plasma facing parts of the present invention are constituted by joining an armour comprising a material having a high melting point and a cooling member comprising copper or the like. A metal member having good solderability with the cooling member is disposed on the joined surface of the armor member. In addition, the joined surface of the cooling member is provided with a barrier layer for preventing invasion of a solder. A solder having a low melting point is interposed between the armour and the cooling member. If they are heated entirely, the solder having low melting point is melted, so that the metal member having good solderability disposed on the armor member is soldered with the barrier layer for the cooling member. Upon exchange of the armour, the joint is heated again. Then, the solder having a low melting point is melted and the armour member and the cooling member are separated. If a solder is put on the cooling member and a new armour is placed and then heated, repairing is completed. (I.S.)

  11. Repair of oxidative DNA base damage in the host genome influences the HIV integration site sequence preference.

    Directory of Open Access Journals (Sweden)

    Geoffrey R Bennett

    Full Text Available Host base excision repair (BER proteins that repair oxidative damage enhance HIV infection. These proteins include the oxidative DNA damage glycosylases 8-oxo-guanine DNA glycosylase (OGG1 and mutY homolog (MYH as well as DNA polymerase beta (Polβ. While deletion of oxidative BER genes leads to decreased HIV infection and integration efficiency, the mechanism remains unknown. One hypothesis is that BER proteins repair the DNA gapped integration intermediate. An alternative hypothesis considers that the most common oxidative DNA base damages occur on guanines. The subtle consensus sequence preference at HIV integration sites includes multiple G:C base pairs surrounding the points of joining. These observations suggest a role for oxidative BER during integration targeting at the nucleotide level. We examined the hypothesis that BER repairs a gapped integration intermediate by measuring HIV infection efficiency in Polβ null cell lines complemented with active site point mutants of Polβ. A DNA synthesis defective mutant, but not a 5'dRP lyase mutant, rescued HIV infection efficiency to wild type levels; this suggested Polβ DNA synthesis activity is not necessary while 5'dRP lyase activity is required for efficient HIV infection. An alternate hypothesis that BER events in the host genome influence HIV integration site selection was examined by sequencing integration sites in OGG1 and MYH null cells. In the absence of these 8-oxo-guanine specific glycosylases the chromatin elements of HIV integration site selection remain the same as in wild type cells. However, the HIV integration site sequence preference at G:C base pairs is altered at several positions in OGG1 and MYH null cells. Inefficient HIV infection in the absence of oxidative BER proteins does not appear related to repair of the gapped integration intermediate; instead oxidative damage repair may participate in HIV integration site preference at the sequence level.

  12. Join Operations in Temporal Databases

    DEFF Research Database (Denmark)

    Gao, D.; Jensen, Christian Søndergaard; Snodgrass, R.T.

    2005-01-01

    Joins are arguably the most important relational operators. Poor implementations are tantamount to computing the Cartesian product of the input relations. In a temporal database, the problem is more acute for two reasons. First, conventional techniques are designed for the evaluation of joins...... with equality predicates rather than the inequality predicates prevalent in valid-time queries. Second, the presence of temporally varying data dramatically increases the size of a database. These factors indicate that specialized techniques are needed to efficiently evaluate temporal joins. We address...... this need for efficient join evaluation in temporal databases. Our purpose is twofold. We first survey all previously proposed temporal join operators. While many temporal join operators have been defined in previous work, this work has been done largely in isolation from competing proposals, with little...

  13. Laser-activated solid protein bands for peripheral nerve repair: an vivo study.

    Science.gov (United States)

    Lauto, A; Trickett, R; Malik, R; Dawes, J M; Owen, E R

    1997-01-01

    Severed tibial nerves in rats were repaired using a novel technique, utilizing a semiconductor diode-laser-activated protein solder applied longitudinally across the join. Welding was produced by selective laser denaturation of solid solder bands containing the dye indocyanine green. An in vivo study, using 48 adult male Wistar rats, compared conventional microsuture-repaired tibial nerves with laser solder-repaired nerves. Nerve repairs were characterised immediately after surgery and after 3 months. Successful regeneration with average compound muscle action potentials of 2.5 +/- 0.5 mV and 2.7 +/- 0.3 mV (mean and standard deviation) was demonstrated for the laser-soldered nerves and the sutured nerves, respectively. Histopathology confirmed comparable regeneration of axons in laser- and suture-operated nerves. The laser-based nerve repair technique was easier and faster than microsuture repair, minimising manipulation damage to the nerve.

  14. End of Life Care

    Science.gov (United States)

    ... Related Topics Choosing Wisely Join our e-newsletter! Aging & Health A to Z End of Life Care ... be used. Ice chips, popsicles, moist swabs, or artificial saliva can help prevent the mouth from becoming ...

  15. Prognostic factors in sensory recovery after digital nerve repair

    OpenAIRE

    Bulut, Tugrul; Akgun, Ulas; Citlak, Atilla; Aslan, Cihan; Sener, Ufuk; Sener, Muhittin

    2018-01-01

    Objective: The prognostic factors that affect sensory nerve recovery after digital nerve repair are variable because of nonhomogeneous data, subjective tests, and different assessment/scoring methods. The aim of this study was to evaluate the success of sensory nerve recovery after digital nerve repair and to investigate the prognostic factors in sensorial healing.Methods: Ninety-six digital nerve repairs of 63 patients were retrospectively evaluated. All nerves were repaired with end-to-end ...

  16. Reward optimization of a repairable system

    International Nuclear Information System (INIS)

    Castro, I.T.; Perez-Ocon, R.

    2006-01-01

    This paper analyzes a system subject to repairable and non-repairable failures. Non-repairable failures lead to replacement of the system. Repairable failures, first lead to repair but they lead to replacement after a fixed number of repairs. Operating and repair times follow phase type distributions (PH-distributions) and the pattern of the operating times is modelled by a geometric process. In this context, the problem is to find the optimal number of repairs, which maximizes the long-run average reward per unit time. To this end, the optimal number is determined and it is obtained by efficient numerical procedures

  17. Reward optimization of a repairable system

    Energy Technology Data Exchange (ETDEWEB)

    Castro, I.T. [Departamento de Matematicas, Facultad de Veterinaria, Universidad de Extremadura, Avenida de la Universidad, s/n. 10071 Caceres (Spain)]. E-mail: inmatorres@unex.es; Perez-Ocon, R. [Departamento de Estadistica e Investigacion Operativa, Facultad de Ciencias, Universidad de Granada, Avenida de Severo Ochoa, s/n. 18071 Granada (Spain)]. E-mail: rperezo@ugr.es

    2006-03-15

    This paper analyzes a system subject to repairable and non-repairable failures. Non-repairable failures lead to replacement of the system. Repairable failures, first lead to repair but they lead to replacement after a fixed number of repairs. Operating and repair times follow phase type distributions (PH-distributions) and the pattern of the operating times is modelled by a geometric process. In this context, the problem is to find the optimal number of repairs, which maximizes the long-run average reward per unit time. To this end, the optimal number is determined and it is obtained by efficient numerical procedures.

  18. Friction riveting as an alternative mechanical fastening to join engineering plastics

    Science.gov (United States)

    Gagliardi, Francesco; Conte, Romina; Bentrovato, Renato; Simeoli, Giorgio; Russo, Pietro; Ambrogio, Giuseppina

    2018-05-01

    Friction Rivecting is a quite new joining process to connect multi-material structures. In brief, a metallic rivet is dipped rotating inside matrixes, usually made of plastics, increasing its original diameter. The use of high-performance plastics is more suitable being their higher mechanical and thermal properties important to avoid material degradation and to allow strong part connections. High-speed friction welding system has been usually used to perform the process. In the work here proposed, the joints have been achieved by means of a traditional milling machine and the attention has been focused on a widely used engineering plastic, i.e. polyamide 6 (PA6) with and without glass fiber reinforcement. A specific speed multiplier has been attached into the mandrel of the used machine to increase the reachable rotational speed. Moreover, rivets made of Titanium Grade 2 and of an Aluminum Alloy, the AA-6060, are utilized. The influence that the heating and the forging length can have on the quality of the obtained junctions, considering a fixed joining depth, has been tested and investigated. The performed connections have been judged by tensile tests, which were set to quantify the maximum strength of the joints for a transverse speed of 1,0 mm/min. Barreling effect can be observed close to the tip, which loses the initial shape of a cylinder characterized by straight vertical walls. Finally, the possible degradation of the polymer, due to temperature increment, has been also evaluated close to the working zone. According to that, it has to be highlighted that the process needs a heating balance, which is necessary to get sound joints. The compromise has, on one side, to allow the rivet penetration and deformation, and on the other side, to avoid the degradation of the polymer, which would affect its properties and a proper rivet deformation.

  19. DNA repair pathways involved in determining the level of cytotoxicity of environmentally relevant UV radiation

    International Nuclear Information System (INIS)

    Carpenter, L.

    2000-01-01

    The sensitivity of cell lines with defects in various DNA repair processes to different wavelengths of UV has been assessed in order to determine the importance of these repair pathways to the cytotoxicity of UV light. The cell lines used in this work were xrs-6 (a Chinese Hamster Ovary (CHO) cell line) mutant for XRCC5/Ku80, EM9 a CHO cell line mutant for XRCC1, UV61 a CHO cell line mutant for ERCC6/CSB, and E3p53-/-, a mouse embryonic fibroblast cell line null for p53. Xrs-6 (defective in Non Homologous End-Joining) was found to be sensitive to the cytotoxic effects of broadband UVA, but not narrowband UVA or narrowband UVB. EM9 (defective in Base Excision Repair/Single-Strand Break Repair) was not sensitive to the cytotoxic effects of both broadband and narrowband UVA, narrowband UVB or narrowband UVC. UV61 (defective in the Transcription Coupled Repair branch of Nucleotide Excision Repair) was sensitive to the cytotoxic effects of narrowband UVA, UVB and UVC. E3p53-/- was sensitive to the cytotoxic effects of narrowband UVA and UVB. Broadband UVA was found to induce high levels of chromosomal damage in xrs-6, as quantified by the micronucleus assay, most likely as a result of this cell lines inability to repair DNA double strand breaks. EM9 was found to be defective in the repair of broadband UVA-induced single strand breaks, as measured by the alkaline gel electrophoresis ('comet') assay. UV61 was unable to repair broadband UVB-induced DNA damage as measured by the alkaline gel electrophoresis ('comet') assay. These results provide evidence that: 1. DNA double-strand breaks contribute to the cytotoxicity of UVA to a greater extent than single-strand breaks. 2. Repair mechanisms that operate in response to UVA may be coupled to transcription. 3. UVB may directly induce SSBs. 4. P53 is involved in the response of the cell to both UVA and UVB radiation. (author)

  20. Blind-date Conversation Joining

    Directory of Open Access Journals (Sweden)

    Luca Cesari

    2013-07-01

    Full Text Available We focus on a form of joining conversations among multiple parties in service-oriented applications where a client may asynchronously join an existing conversation without need to know in advance any information about it. More specifically, we show how the correlation mechanism provided by orchestration languages enables a form of conversation joining that is completely transparent to clients and that we call 'blind-date joining'. We provide an implementation of this strategy by using the standard orchestration language WS-BPEL. We then present its formal semantics by resorting to COWS, a process calculus specifically designed for modelling service-oriented applications. We illustrate our approach by means of a simple, but realistic, case study from the online games domain.

  1. The RSF1 histone-remodelling factor facilitates DNA double-strand break repair by recruiting centromeric and Fanconi Anaemia proteins.

    Directory of Open Access Journals (Sweden)

    Fabio Pessina

    2014-05-01

    Full Text Available ATM is a central regulator of the cellular responses to DNA double-strand breaks (DSBs. Here we identify a biochemical interaction between ATM and RSF1 and we characterise the role of RSF1 in this response. The ATM-RSF1 interaction is dependent upon both DSBs and ATM kinase activity. Together with SNF2H/SMARCA5, RSF1 forms the RSF chromatin-remodelling complex. Although RSF1 is specific to the RSF complex, SNF2H/SMARCA5 is a catalytic subunit of several other chromatin-remodelling complexes. Although not required for checkpoint signalling, RSF1 is required for efficient repair of DSBs via both end-joining and homology-directed repair. Specifically, the ATM-dependent recruitment to sites of DSBs of the histone fold proteins CENPS/MHF1 and CENPX/MHF2, previously identified at centromeres, is RSF1-dependent. In turn these proteins recruit and regulate the mono-ubiquitination of the Fanconi Anaemia proteins FANCD2 and FANCI. We propose that by depositing CENPS/MHF1 and CENPX/MHF2, the RSF complex either directly or indirectly contributes to the reorganisation of chromatin around DSBs that is required for efficient DNA repair.

  2. Laparoscopic repair of postoperative perineal hernia.

    LENUS (Irish Health Repository)

    Ryan, Stephen

    2010-01-01

    Perineal hernias are infrequent complications following abdominoperineal operations. Various approaches have been described for repair of perineal hernias including open transabdominal, transperineal or combined abdominoperineal repairs. The use of laparoscopic transabdominal repair of perineal hernias is not well-described. We present a case report demonstrating the benefits of laparoscopic repair of perineal hernia following previous laparoscopic abdominoperineal resection (APR) using a nonabsorbable mesh to repair the defect. We have demonstrated that the use of laparoscopy with repair of the pelvic floor defect using a non absorbable synthetic mesh offers an excellent alternative with many potential advantages over open transabdominal and transperineal repairs.

  3. Dynamic dependence on ATR and ATM for double-strand break repair in human embryonic stem cells and neural descendants.

    Science.gov (United States)

    Adams, Bret R; Golding, Sarah E; Rao, Raj R; Valerie, Kristoffer

    2010-04-02

    The DNA double-strand break (DSB) is the most toxic form of DNA damage. Studies aimed at characterizing DNA repair during development suggest that homologous recombination repair (HRR) is more critical in pluripotent cells compared to differentiated somatic cells in which nonhomologous end joining (NHEJ) is dominant. We have characterized the DNA damage response (DDR) and quality of DNA double-strand break (DSB) repair in human embryonic stem cells (hESCs), and in vitro-derived neural cells. Resolution of ionizing radiation-induced foci (IRIF) was used as a surrogate for DSB repair. The resolution of gamma-H2AX foci occurred at a slower rate in hESCs compared to neural progenitors (NPs) and astrocytes perhaps reflective of more complex DSB repair in hESCs. In addition, the resolution of RAD51 foci, indicative of active homologous recombination repair (HRR), showed that hESCs as well as NPs have high capacity for HRR, whereas astrocytes do not. Importantly, the ATM kinase was shown to be critical for foci formation in astrocytes, but not in hESCs, suggesting that the DDR is different in these cells. Blocking the ATM kinase in astrocytes not only prevented the formation but also completely disassembled preformed repair foci. The ability of hESCs to form IRIF was abrogated with caffeine and siRNAs targeted against ATR, implicating that hESCs rely on ATR, rather than ATM for regulating DSB repair. This relationship dynamically changed as cells differentiated. Interestingly, while the inhibition of the DNA-PKcs kinase (and presumably non-homologous endjoining [NHEJ]) in astrocytes slowed IRIF resolution it did not in hESCs, suggesting that repair in hESCs does not utilize DNA-PKcs. Altogether, our results show that hESCs have efficient DSB repair that is largely ATR-dependent HRR, whereas astrocytes critically depend on ATM for NHEJ, which, in part, is DNA-PKcs-independent.

  4. Lightning Fast and Space Efficient Inequality Joins

    KAUST Repository

    Khayyat, Zuhair

    2015-09-01

    Inequality joins, which join relational tables on inequality conditions, are used in various applications. While there have been a wide range of optimization methods for joins in database systems, from algorithms such as sort-merge join and band join, to various indices such as B+-tree,R*-tree and Bitmap, inequality joins have received little attention and queries containing such joins are usually very slow. In this paper, we introduce fast inequality join algorithms. We put columns to be joined in sorted arrays and we use permutation arrays to encode positions of tuples in one sorted array w.r.t. the other sorted array. In contrast to sort-merge join, we use space effcient bit-arrays that enable optimizations, such as Bloom filter indices, for fast computation of the join results. We have implemented a centralized version of these algorithms on top of PostgreSQL, and a distributed version on top of Spark SQL. We have compared against well known optimization techniques for inequality joins and show that our solution is more scalable and several orders of magnitude faster.

  5. Lightning fast and space efficient inequality joins

    KAUST Repository

    Khayyat, Zuhair

    2015-09-01

    Inequality joins, which join relational tables on inequality conditions, are used in various applications. While there have been a wide range of optimization methods for joins in database systems, from algorithms such as sort-merge join and band join, to various indices such as B+-tree, R*-tree and Bitmap, inequality joins have received little attention and queries containing such joins are usually very slow. In this paper, we introduce fast inequality join algorithms. We put columns to be joined in sorted arrays and we use permutation arrays to encode positions of tuples in one sorted array w.r.t. the other sorted array. In contrast to sort-merge join, we use space efficient bit-arrays that enable optimizations, such as Bloom filter indices, for fast computation of the join results. We have implemented a centralized version of these algorithms on top of PostgreSQL, and a distributed version on top of Spark SQL. We have compared against well known optimization techniques for inequality joins and show that our solution is more scalable and several orders of magnitude faster.

  6. Lightning fast and space efficient inequality joins

    KAUST Repository

    Khayyat, Zuhair; Lucia, William; Singh, Meghna; Ouzzani, Mourad; Papotti, Paolo; Quiané -Ruiz, Jorge-Arnulfo; Tang, Nan; Kalnis, Panos

    2015-01-01

    Inequality joins, which join relational tables on inequality conditions, are used in various applications. While there have been a wide range of optimization methods for joins in database systems, from algorithms such as sort-merge join and band join, to various indices such as B+-tree, R*-tree and Bitmap, inequality joins have received little attention and queries containing such joins are usually very slow. In this paper, we introduce fast inequality join algorithms. We put columns to be joined in sorted arrays and we use permutation arrays to encode positions of tuples in one sorted array w.r.t. the other sorted array. In contrast to sort-merge join, we use space efficient bit-arrays that enable optimizations, such as Bloom filter indices, for fast computation of the join results. We have implemented a centralized version of these algorithms on top of PostgreSQL, and a distributed version on top of Spark SQL. We have compared against well known optimization techniques for inequality joins and show that our solution is more scalable and several orders of magnitude faster.

  7. FIXED ASSETS WITH AN OPEN-ENDED USEFUL LIFE AS A NEW OBJECT OF ACCOUNTING IN THE SHIP REPAIR INDUSTRY

    Directory of Open Access Journals (Sweden)

    Olga Zharikova

    2015-09-01

    Full Text Available The industry-specific factors that contribute to the allocation of a new object of accounting in property, plant and equipment of the ship repair industry organizations are revealed. Specificity the salvaging operation of hydraulic structures, which affect the organization and methodology of objects` accounting, is described. The definition of «fixed assets with an open-ended useful life» is proposed.

  8. DNA-synthesis inhibition and repair DNA-synthesis in CHO Ade- C cells: An alternative approach to genotoxicity testing

    International Nuclear Information System (INIS)

    Slamenova, D.; Papsova, E.; Gabelova, A.; Dusinska, M.; Collins, A.; Wsolova, L.

    1997-01-01

    We describe an alternative assay to determine genotoxicity. Its main feature is that it combines two measures in a single experiment; the inhibition of replicative DNA synthesis together with the stimulation of DNA repair. We show that, in tests of four different genotoxic agents, the assay gives results that are entirely consistent with what is known about the mode of action of these agents. In addition, we have demonstrated that chemical carcinogens requiring metabolic activation can be examined using a standard procedure of incubation with a microsomal activating fraction. We consider the combined assay for DNA synthesis inhibition and repair synthesis to be a useful way for the rapid pre-screening of chemicals suspected of genotoxic activity on the level of mammalian cells. (author)

  9. Studies on the repair of double strand break of DNA and cellular carcinogenesis, and consideration on the concept of extinction of nuclear power

    International Nuclear Information System (INIS)

    Teraoka, Hirobumi

    2013-01-01

    This paper describes the relationship between the repair of double strand break (DSB) of DNA and cellular carcinogenesis mainly on author's investigations, and his recent thought aiming at the extinction of nuclear power. The molecular repairing system is explained about DNA DSB induced by radiation and chemicals. When DSB occurs, nucleosome consisting from 4 core-histones participates to link the broken ends and then repair mechanisms of homologous recombination (HRR) and non-homologous end joining (NHEJ) begin to work. The latter is dominant in mammalians. Thus the genetic defect in these systems of DSB response and repair is a course of disorders such as ataxia telangiectasia (AT) (DSB sensor defect), genetic breast cancer (HRR defect), and radiosensitive-severe combined immunodeficiency (RS-SCID) (NHEJ defect), all of which result in cancer formation. NHEJ repair is known to be error-prone. Against multi-step carcinogenesis where accumulated gene mutations lead to the cancer formation, the author thinks chromosomal instability is one of important carcinogenic causes: the instability can be a trigger of producing cancer stem cells because the cells can be yielded from mouse embryonic stem cells where DSB is shown to participate in the process. Low dose radiation produces a small amount of DSB, to which the repair response is less sensitive at G2/M checkpoint, ultimately leading to genomic instability. Considering effects of the low dose radiation exposure above, and of the internal exposure to 3 H-thymidine beta ray in cells, of indoor Rn participating 16% of lung cancer incidence (Canadian epidemiological data) and so on, together with moral and social responsibility of scientist and technologist, the author says to have attained to the concept of the ''Extinction of Nuclear Power''. (T.T)

  10. Joining uranium to steel

    International Nuclear Information System (INIS)

    Perkins, M.A.

    1976-05-01

    A method has been devised which will allow the joining of uranium to steel by fusion welding through the use of an intermediate material. Uranium-0.5 titanium was joined to AISI 304L stainless steel by using a vanadium insert. Also, a method is now available for selecting possible filler metals when two entirely dissimilar metals need to be joined. This method allows a quantitative ranking to be made of the possible filler metals and thus the most likely candidate can be selected

  11. Authenticated join processing in outsourced databases

    KAUST Repository

    Yang, Yin

    2009-01-01

    Database outsourcing requires that a query server constructs a proof of result correctness, which can be verified by the client using the data owner\\'s signature. Previous authentication techniques deal with range queries on a single relation using an authenticated data structure (ADS). On the other hand, authenticated join processing is inherently more complex than ranges since only the base relations (but not their combination) are signed by the owner. In this paper, we present three novel join algorithms depending on the ADS availability: (i) Authenticated Indexed Sort Merge Join (AISM), which utilizes a single ADS on the join attribute, (ii) Authenticated Index Merge Join (AIM) that requires an ADS (on the join attribute) for both relations, and (iii) Authenticated Sort Merge Join (ASM), which does not rely on any ADS. We experimentally demonstrate that the proposed methods outperform two benchmark algorithms, often by several orders of magnitude, on all performance metrics, and effectively shift the workload to the outsourcing service. Finally, we extend our techniques to complex queries that combine multi-way joins with selections and projections. ©2009 ACM.

  12. Authenticated join processing in outsourced databases

    KAUST Repository

    Yang, Yin; Papadias, Dimitris; Papadopoulos, Stavros; Kalnis, Panos

    2009-01-01

    Database outsourcing requires that a query server constructs a proof of result correctness, which can be verified by the client using the data owner's signature. Previous authentication techniques deal with range queries on a single relation using an authenticated data structure (ADS). On the other hand, authenticated join processing is inherently more complex than ranges since only the base relations (but not their combination) are signed by the owner. In this paper, we present three novel join algorithms depending on the ADS availability: (i) Authenticated Indexed Sort Merge Join (AISM), which utilizes a single ADS on the join attribute, (ii) Authenticated Index Merge Join (AIM) that requires an ADS (on the join attribute) for both relations, and (iii) Authenticated Sort Merge Join (ASM), which does not rely on any ADS. We experimentally demonstrate that the proposed methods outperform two benchmark algorithms, often by several orders of magnitude, on all performance metrics, and effectively shift the workload to the outsourcing service. Finally, we extend our techniques to complex queries that combine multi-way joins with selections and projections. ©2009 ACM.

  13. In normal human fibroblasts variation in DSB repair capacity cannot be ascribed to radiation-induced changes in the localisation, expression or activity of major NHEJ proteins

    DEFF Research Database (Denmark)

    Kasten-Pisula, Ulla; Vronskaja, Svetlana; Overgaard, Jens

    2008-01-01

    in the activity of the DNA-PK complex induced upon irradiation. CONCLUSIONS: For normal human fibroblasts, the level or activity of NHEJ proteins measured prior to or after irradiation cannot be used to predict the DSB repair capacity or cellular radiosensitivity. Udgivelsesdato: 2008-Mar......BACKGROUND AND PURPOSE: The aim of the present study was to test whether for normal human fibroblasts the variation in double-strand break (DSB) repair capacity results from radiation-induced differences in localisation, expression or activity of major non-homologous end-joining (NHEJ) proteins....... MATERIALS AND METHODS: Experiments were performed with 11 normal human fibroblast strains AF01-11. NHEJ proteins were determined by Western blot and DNA-PK activity by pulldown-assay. RESULTS: The four NHEJ proteins tested (Ku70, Ku80, XRCC4 and DNA-PKcs) were found to be localised almost exclusively...

  14. The ’TIS’ Intelligent Gateway Computer: An Alternative to the ’Doomsday Scenario’,

    Science.gov (United States)

    1983-02-01

    century. The traditional intermediary, the librarian, or information specialist, will have joined the fate of dinosaurs by that time, Lewis said, unless...others in similar work environments, (Refs. 2-5) also by Lewis who offers it as a partial alternative to the extinction of the traditional information...1979 1960 1981 1982 Year The underlying reasons probably are that major end users at LLNL: 1) Don’t have the time to bother with routine searching- 2

  15. Joining Movement Sequences: Modified Dynamic Movement Primitives for Robotics Applications Exemplified on Handwriting

    DEFF Research Database (Denmark)

    Kulvicius, Tomas; Ning, K.; Tamosiunaite, M.

    2012-01-01

    to simulated handwriting generation, which are also shown on a robot, where an adaptive algorithm is used to learn trajectories from human demonstration. These results demonstrate that the new method is a feasible alternative for joining of movement sequences, which has a high potential for all robotics...

  16. Web-ADARE: A Web-Aided Data Repairing System

    KAUST Repository

    Gu, Binbin

    2017-03-08

    Data repairing aims at discovering and correcting erroneous data in databases. In this paper, we develop Web-ADARE, an end-to-end web-aided data repairing system, to provide a feasible way to involve the vast data sources on the Web in data repairing. Our main attention in developing Web-ADARE is paid on the interaction problem between web-aided repairing and rule-based repairing, in order to minimize the Web consultation cost while reaching predefined quality requirements. The same interaction problem also exists in crowd-based methods but this is not yet formally defined and addressed. We first prove in theory that the optimal interaction scheme is not feasible to be achieved, and then propose an algorithm to identify a scheme for efficient interaction by investigating the inconsistencies and the dependencies between values in the repairing process. Extensive experiments on three data collections demonstrate the high repairing precision and recall of Web-ADARE, and the efficiency of the generated interaction scheme over several baseline ones.

  17. Web-ADARE: A Web-Aided Data Repairing System

    KAUST Repository

    Gu, Binbin; Li, Zhixu; Yang, Qiang; Xie, Qing; Liu, An; Liu, Guanfeng; Zheng, Kai; Zhang, Xiangliang

    2017-01-01

    Data repairing aims at discovering and correcting erroneous data in databases. In this paper, we develop Web-ADARE, an end-to-end web-aided data repairing system, to provide a feasible way to involve the vast data sources on the Web in data repairing. Our main attention in developing Web-ADARE is paid on the interaction problem between web-aided repairing and rule-based repairing, in order to minimize the Web consultation cost while reaching predefined quality requirements. The same interaction problem also exists in crowd-based methods but this is not yet formally defined and addressed. We first prove in theory that the optimal interaction scheme is not feasible to be achieved, and then propose an algorithm to identify a scheme for efficient interaction by investigating the inconsistencies and the dependencies between values in the repairing process. Extensive experiments on three data collections demonstrate the high repairing precision and recall of Web-ADARE, and the efficiency of the generated interaction scheme over several baseline ones.

  18. ATM loss leads to synthetic lethality in BRCA1 BRCT mutant mice associated with exacerbated defects in homology-directed repair.

    Science.gov (United States)

    Chen, Chun-Chin; Kass, Elizabeth M; Yen, Wei-Feng; Ludwig, Thomas; Moynahan, Mary Ellen; Chaudhuri, Jayanta; Jasin, Maria

    2017-07-18

    BRCA1 is essential for homology-directed repair (HDR) of DNA double-strand breaks in part through antagonism of the nonhomologous end-joining factor 53BP1. The ATM kinase is involved in various aspects of DNA damage signaling and repair, but how ATM participates in HDR and genetically interacts with BRCA1 in this process is unclear. To investigate this question, we used the Brca1 S1598F mouse model carrying a mutation in the BRCA1 C-terminal domain of BRCA1. Whereas ATM loss leads to a mild HDR defect in adult somatic cells, we find that ATM inhibition leads to severely reduced HDR in Brca1 S1598F cells. Consistent with a critical role for ATM in HDR in this background, loss of ATM leads to synthetic lethality of Brca1 S1598F mice. Whereas both ATM and BRCA1 promote end resection, which can be regulated by 53BP1, 53bp1 deletion does not rescue the HDR defects of Atm mutant cells, in contrast to Brca1 mutant cells. These results demonstrate that ATM has a role in HDR independent of the BRCA1-53BP1 antagonism and that its HDR function can become critical in certain contexts.

  19. Turbo Charge CPU Utilization in Fork/Join Using the ManagedBlocker

    CERN Multimedia

    CERN. Geneva

    2017-01-01

    Fork/Join is a framework for parallelizing calculations using recursive decomposition, also called divide and conquer. These algorithms occasionally end up duplicating work, especially at the beginning of the run. We can reduce wasted CPU cycles by implementing a reserved caching scheme. Before a task starts its calculation, it tries to reserve an entry in the shared map. If it is successful, it immediately begins. If not, it blocks until the other thread has finished its calculation. Unfortunately this might result in a significant number of blocked threads, decreasing CPU utilization. In this talk we will demonstrate this issue and offer a solution in the form of the ManagedBlocker. Combined with the Fork/Join, it can keep parallelism at the desired level.

  20. A novel conduit-based coaptation device for primary nerve repair.

    Science.gov (United States)

    Bamba, Ravinder; Riley, D Colton; Kelm, Nathaniel D; Cardwell, Nancy; Pollins, Alonda C; Afshari, Ashkan; Nguyen, Lyly; Dortch, Richard D; Thayer, Wesley P

    2018-06-01

    Conduit-based nerve repairs are commonly used for small nerve gaps, whereas primary repair may be performed if there is no tension on nerve endings. We hypothesize that a conduit-based nerve coaptation device will improve nerve repair outcomes by avoiding sutures at the nerve repair site and utilizing the advantages of a conduit-based repair. The left sciatic nerves of female Sprague-Dawley rats were transected and repaired using a novel conduit-based device. The conduit-based device group was compared to a control group of rats that underwent a standard end-to-end microsurgical repair of the sciatic nerve. Animals underwent behavioral assessments at weekly intervals post-operatively using the sciatic functional index (SFI) test. Animals were sacrificed at four weeks to obtain motor axon counts from immunohistochemistry. A sub-group of animals were sacrificed immediately post repair to obtain MRI images. SFI scores were superior in rats which received conduit-based repairs compared to the control group. Motor axon counts distal to the injury in the device group at four weeks were statistically superior to the control group. MRI tractography was used to demonstrate repair of two nerves using the novel conduit device. A conduit-based nerve coaptation device avoids sutures at the nerve repair site and leads to improved outcomes in a rat model. Conduit-based nerve repair devices have the potential to standardize nerve repairs while improving outcomes.

  1. Parallel trajectory similarity joins in spatial networks

    KAUST Repository

    Shang, Shuo

    2018-04-04

    The matching of similar pairs of objects, called similarity join, is fundamental functionality in data management. We consider two cases of trajectory similarity joins (TS-Joins), including a threshold-based join (Tb-TS-Join) and a top-k TS-Join (k-TS-Join), where the objects are trajectories of vehicles moving in road networks. Given two sets of trajectories and a threshold θ, the Tb-TS-Join returns all pairs of trajectories from the two sets with similarity above θ. In contrast, the k-TS-Join does not take a threshold as a parameter, and it returns the top-k most similar trajectory pairs from the two sets. The TS-Joins target diverse applications such as trajectory near-duplicate detection, data cleaning, ridesharing recommendation, and traffic congestion prediction. With these applications in mind, we provide purposeful definitions of similarity. To enable efficient processing of the TS-Joins on large sets of trajectories, we develop search space pruning techniques and enable use of the parallel processing capabilities of modern processors. Specifically, we present a two-phase divide-and-conquer search framework that lays the foundation for the algorithms for the Tb-TS-Join and the k-TS-Join that rely on different pruning techniques to achieve efficiency. For each trajectory, the algorithms first find similar trajectories. Then they merge the results to obtain the final result. The algorithms for the two joins exploit different upper and lower bounds on the spatiotemporal trajectory similarity and different heuristic scheduling strategies for search space pruning. Their per-trajectory searches are independent of each other and can be performed in parallel, and the mergings have constant cost. An empirical study with real data offers insight in the performance of the algorithms and demonstrates that they are capable of outperforming well-designed baseline algorithms by an order of magnitude.

  2. Parallel trajectory similarity joins in spatial networks

    KAUST Repository

    Shang, Shuo; Chen, Lisi; Wei, Zhewei; Jensen, Christian S.; Zheng, Kai; Kalnis, Panos

    2018-01-01

    The matching of similar pairs of objects, called similarity join, is fundamental functionality in data management. We consider two cases of trajectory similarity joins (TS-Joins), including a threshold-based join (Tb-TS-Join) and a top-k TS-Join (k-TS-Join), where the objects are trajectories of vehicles moving in road networks. Given two sets of trajectories and a threshold θ, the Tb-TS-Join returns all pairs of trajectories from the two sets with similarity above θ. In contrast, the k-TS-Join does not take a threshold as a parameter, and it returns the top-k most similar trajectory pairs from the two sets. The TS-Joins target diverse applications such as trajectory near-duplicate detection, data cleaning, ridesharing recommendation, and traffic congestion prediction. With these applications in mind, we provide purposeful definitions of similarity. To enable efficient processing of the TS-Joins on large sets of trajectories, we develop search space pruning techniques and enable use of the parallel processing capabilities of modern processors. Specifically, we present a two-phase divide-and-conquer search framework that lays the foundation for the algorithms for the Tb-TS-Join and the k-TS-Join that rely on different pruning techniques to achieve efficiency. For each trajectory, the algorithms first find similar trajectories. Then they merge the results to obtain the final result. The algorithms for the two joins exploit different upper and lower bounds on the spatiotemporal trajectory similarity and different heuristic scheduling strategies for search space pruning. Their per-trajectory searches are independent of each other and can be performed in parallel, and the mergings have constant cost. An empirical study with real data offers insight in the performance of the algorithms and demonstrates that they are capable of outperforming well-designed baseline algorithms by an order of magnitude.

  3. Constraints in the use of repair half times and mathematical modelling for the clinical application of HDR and PDR treatment schedules as an alternative for LDR brachytherapy

    International Nuclear Information System (INIS)

    Pop, L.A.M.; Broek, J.F.C.M. van den; Visser, A.G.; Kogel, A.J. van der

    1996-01-01

    Using theoretical models based on radiobiological principles for the design of new treatment schedules for HDR and PDR brachytherapy, it is important to realise the impact of assumptions regarding the kinetics of repair. Extrapolations based on longer repair half times in a continuous LDR reference scheme may lead to the calculation of dangerously high doses for alternative HDR and PDR treatment schedules. We used the clinical experience obtained with conventional ERT and LDR brachytherapy in head and neck cancer as a clinical guideline to check the impact of the radiobiological parameters used. Biologically equivalent dose (BED) values for the in clinical practice of LDR brachytherapy recommended dose of 65-70 Gy (prescribed at a dose rate between 30-50 cGy/h) are calculated as a function of the repair half time. These BED values are compared with the biological effect of a clinical reference dose of conventional ERT with 2 Gy/day and complete repair between the fractions. From this comparison of LDR and ERT treatment schedules, a range of values for the repair half times of acute or late responding tissues is demarcated with a reasonable fit to the clinical data. For the acute effects (or tumor control) the best fits are obtained for repair half times of about 0.5 h, while for late effects the repair half times are at least 1 h and can be as high as 3 h. Within these ranges of repair half times for acute and late effects, the outcome of 'alternative' HDR or PDR treatment schedules are discussed. It is predominantly the late reacting normal tissue with the longer repair half time for which problems will be encountered and no or only marginal gain is to be expected of decreasing the dose rate per pulse in PDR brachytherapy

  4. On the Expressiveness of Joining

    Directory of Open Access Journals (Sweden)

    Thomas Given-Wilson

    2015-08-01

    Full Text Available The expressiveness of communication primitives has been explored in a common framework based on the pi-calculus by considering four features: synchronism (asynchronous vs synchronous, arity (monadic vs polyadic data, communication medium (shared dataspaces vs channel-based, and pattern-matching (binding to a name vs testing name equality vs intensionality. Here another dimension coordination is considered that accounts for the number of processes required for an interaction to occur. Coordination generalises binary languages such as pi-calculus to joining languages that combine inputs such as the Join Calculus and general rendezvous calculus. By means of possibility/impossibility of encodings, this paper shows coordination is unrelated to the other features. That is, joining languages are more expressive than binary languages, and no combination of the other features can encode a joining language into a binary language. Further, joining is not able to encode any of the other features unless they could be encoded otherwise.

  5. CRISPR/Cas-mediated knock-in via non-homologous end-joining in the crustacean Daphnia magna.

    Science.gov (United States)

    Kumagai, Hitoshi; Nakanishi, Takashi; Matsuura, Tomoaki; Kato, Yasuhiko; Watanabe, Hajime

    2017-01-01

    The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated system (Cas) is widely used for mediating the knock-in of foreign DNA into the genomes of various organisms. Here, we report a process of CRISPR/Cas-mediated knock-in via non-homologous end joining by the direct injection of Cas9/gRNA ribonucleoproteins (RNPs) in the crustacean Daphnia magna, which is a model organism for studies on toxicology, ecology, and evolution. First, we confirmed the cleavage activity of Cas9 RNPs comprising purified Cas9 proteins and gRNAs in D. magna. We used a gRNA that targets exon 10 of the eyeless gene. Cas9 proteins were incubated with the gRNAs and the resulting Cas9 RNPs were injected into D. magna eggs, which led to a typical phenotype of the eyeless mutant, i.e., eye deformity. The somatic and heritable mutagenesis efficiencies were up to 96% and 40%, respectively. Second, we tested the CRISPR/Cas-mediated knock-in of a plasmid by the injection of Cas9 RNPs. The donor DNA plasmid harboring the fluorescent reporter gene was designed to contain the gRNA recognition site. The co-injection of Cas9 RNPs together with the donor DNAs resulted in generation of one founder animal that produced fluorescent progenies. This transgenic Daphnia had donor DNA at the targeted genomic site, which suggested the concurrent cleavage of the injected plasmid DNA and genomic DNA. Owing to its simplicity and ease of experimental design, we suggest that the CRISPR/Cas-mediated knock-in method represents a promising tool for studying functional genomics in D. magna.

  6. BRCA1-associated exclusion of 53BP1 from DNA damage sites underlies temporal control of DNA repair

    Science.gov (United States)

    Chapman, J. Ross; Sossick, Alex J.; Boulton, Simon J.; Jackson, Stephen P.

    2012-01-01

    Summary Following irradiation, numerous DNA-damage-responsive proteins rapidly redistribute into microscopically visible subnuclear aggregates, termed ionising-radiation-induced foci (IRIF). How the enrichment of proteins on damaged chromatin actually relates to DNA repair remains unclear. Here, we use super-resolution microscopy to examine the spatial distribution of BRCA1 and 53BP1 proteins within single IRIF at subdiffraction-limit resolution, yielding an unprecedented increase in detail that was not previously apparent by conventional microscopy. Consistent with a role for 53BP1 in promoting DNA double-strand break repair by non-homologous end joining, 53BP1 enrichment in IRIF is most prominent in the G0/G1 cell cycle phases, where it is enriched in dense globular structures. By contrast, as cells transition through S phase, the recruitment of BRCA1 into the core of IRIF is associated with an exclusion of 53BP1 to the focal periphery, leading to an overall reduction of 53BP1 occupancy at DNA damage sites. Our data suggest that the BRCA1-associated IRIF core corresponds to chromatin regions associated with repair by homologous recombination, and the enrichment of BRCA1 in IRIF represents a temporal switch in the DNA repair program. We propose that BRCA1 antagonises 53BP1-dependent DNA repair in S phase by inhibiting its interaction with chromatin proximal to damage sites. Furthermore, the genomic instability exhibited by BRCA1-deficient cells might result from a failure to efficiently exclude 53BP1 from such regions during S phase. PMID:22553214

  7. Genetic variation in DNA repair gene XRCC7 (G6721T) and susceptibility to breast cancer.

    Science.gov (United States)

    Nasiri, Meysam; Saadat, Iraj; Omidvari, Shahpour; Saadat, Mostafa

    2012-08-15

    The human XRCC7 is a DNA double-strand break (DSBs) repair gene, involved in non-homologous end joining (NHEJ). It is speculated that DNA DSBs repair have an important role during development of breast cancer. The human XRCC7 is a NHEJ DSBs repair gene. Genetic variation G6721T of XRCC7 (rs7003908) is located in the intron 8 of the gene. This polymorphism may regulate splicing and cause mRNA instability. In the present study, we specifically investigated whether common G6721T genetic variant of XRCC7 was associated with an altered risk of breast cancer. The present study included 362 females with breast cancer. Age frequency-matched controls (362 persons) were randomly selected from the healthy female blood donors, according to the age distribution of the cases. Using RFLP-PCR based method, the polymorphism of XRCC7 was determined. The TG (OR=1.20, 95% CI: 0.83-1.74, P=0.320) and TT (OR=1.01, 95% CI: 0.67-1.53, P=0.933) genotypes had no significant effect on risk of breast cancer, in comparison with the GG genotype. Our present findings indicate that the TT and TG genotypes were not associated with an altered breast cancer risk. Copyright © 2012 Elsevier B.V. All rights reserved.

  8. Common Influence Join

    DEFF Research Database (Denmark)

    Yiu, Man Lung; Mamoulis, Nikos; Karras, Panagiotis

    2008-01-01

    We identify and formalize a novel join operator for two spatial pointsets P and Q. The common influence join (CIJ) returns the pairs of points (p,q),p isin P,q isin Q, such that there exists a location in space, being closer to p than to any other point in P and at the same time closer to q than ......-demand, is very efficient in practice, incurring only slightly higher I/O cost than the theoretical lower bound cost for the problem....

  9. Speeding Up Neighbour-Joining Tree Construction

    DEFF Research Database (Denmark)

    Brodal, Gerth Stølting; Fagerberg, Rolf; Mailund, Thomas

    A widely used method for constructing phylogenetic trees is the neighbour-joining method of Saitou and Nei. We develope heuristics for speeding up the neighbour-joining method which generate the same phylogenetic trees as the original method. All heuristics are based on using a quad-tree to guide...... the search for the next pair of nodes to join, but di#er in the information stored in quad-tree nodes, the way the search is performed, and in the way the quad-tree is updated after a join. We empirically evaluate the performance of the heuristics on distance matrices obtained from the Pfam collection...... of alignments, and compare the running time with that of the QuickTree tool, a well-known and widely used implementation of the standard neighbour-joining method. The results show that the presented heuristics can give a significant speed-up over the standard neighbour-joining method, already for medium sized...

  10. Role of Double-Strand Break End-Tethering during Gene Conversion in Saccharomyces cerevisiae.

    Directory of Open Access Journals (Sweden)

    Suvi Jain

    2016-04-01

    Full Text Available Correct repair of DNA double-strand breaks (DSBs is critical for maintaining genome stability. Whereas gene conversion (GC-mediated repair is mostly error-free, repair by break-induced replication (BIR is associated with non-reciprocal translocations and loss of heterozygosity. We have previously shown that a Recombination Execution Checkpoint (REC mediates this competition by preventing the BIR pathway from acting on DSBs that can be repaired by GC. Here, we asked if the REC can also determine whether the ends that are engaged in a GC-compatible configuration belong to the same break, since repair involving ends from different breaks will produce potentially deleterious translocations. We report that the kinetics of repair are markedly delayed when the two DSB ends that participate in GC belong to different DSBs (termed Trans compared to the case when both DSB ends come from the same break (Cis. However, repair in Trans still occurs by GC rather than BIR, and the overall efficiency of repair is comparable. Hence, the REC is not sensitive to the "origin" of the DSB ends. When the homologous ends for GC are in Trans, the delay in repair appears to reflect their tethering to sequences on the other side of the DSB that themselves recombine with other genomic locations with which they share sequence homology. These data support previous observations that the two ends of a DSB are usually tethered to each other and that this tethering facilitates both ends encountering the same donor sequence. We also found that the presence of homeologous/repetitive sequences in the vicinity of a DSB can distract the DSB end from finding its bona fide homologous donor, and that inhibition of GC by such homeologous sequences is markedly increased upon deleting Sgs1 but not Msh6.

  11. Efficient Joins with Compressed Bitmap Indexes

    Energy Technology Data Exchange (ETDEWEB)

    Computational Research Division; Madduri, Kamesh; Wu, Kesheng

    2009-08-19

    We present a new class of adaptive algorithms that use compressed bitmap indexes to speed up evaluation of the range join query in relational databases. We determine the best strategy to process a join query based on a fast sub-linear time computation of the join selectivity (the ratio of the number of tuples in the result to the total number of possible tuples). In addition, we use compressed bitmaps to represent the join output compactly: the space requirement for storing the tuples representing the join of two relations is asymptotically bounded by min(h; n . cb), where h is the number of tuple pairs in the result relation, n is the number of tuples in the smaller of the two relations, and cb is the cardinality of the larger column being joined. We present a theoretical analysis of our algorithms, as well as experimental results on large-scale synthetic and real data sets. Our implementations are efficient, and consistently outperform well-known approaches for a range of join selectivity factors. For instance, our count-only algorithm is up to three orders of magnitude faster than the sort-merge approach, and our best bitmap index-based algorithm is 1.2x-80x faster than the sort-merge algorithm, for various query instances. We achieve these speedups by exploiting several inherent performance advantages of compressed bitmap indexes for join processing: an implicit partitioning of the attributes, space-efficiency, and tolerance of high-cardinality relations.

  12. Low concentrations of antimony impair DNA damage signaling and the repair of radiation-induced DSB in HeLa S3 cells.

    Science.gov (United States)

    Koch, Barbara; Maser, Elena; Hartwig, Andrea

    2017-12-01

    Antimony is utilized in a large variety of industrial applications, leading to significant environmental and occupational exposure. Mainly based on animal experiments, the IARC and MAK Commission have classified antimony and its inorganic compounds as Group 2B or 2 carcinogens, respectively. However, the underlying mode(s) of action are still largely unknown. In the present study, we investigated the impact of non-cytotoxic up to cytotoxic concentrations of SbCl 3 on DNA DSB repair and cell cycle control in HeLa S3 cells. We induced DSB by γ-irradiation and analyzed inhibitory actions of antimony on potential molecular targets of the DSB repair machinery. Antimony disturbed cell cycle control, affecting phosphorylation of Chk1. Furthermore, the repair of DSB was impaired in the presence of antimony, as monitored by pulsed-field gel electrophoresis and γH2AX foci formation of cells in G1 and G2 phase. Specifically, BRCA1 and RAD51 were identified as molecular targets. Our results point towards an interference with both non-homologous end-joining (NHEJ) and homologous recombination (HR), and inhibitory effects may be explained by interactions with critical cysteine groups; this needs to be further investigated. Altogether, the results provide further evidence for the impairment of DNA repair processes as one underlying mechanism in antimony-induced carcinogenicity.

  13. Effects of expression level of DNA repair-related genes involved in the NHEJ pathway on radiation-induced cognitive impairment

    International Nuclear Information System (INIS)

    Zhang Liyuan; Chen Liesong; Sun Rui; Ji Shengjun; Ding Yanyan; Wu Jia; Tian Ye

    2013-01-01

    Cranial radiation therapy can induce cognitive decline. Impairments of hippocampal neurogenesis are thought to be a paramountly important mechanism underlying radiation-induced cognitive dysfunction. In the mature nervous system, DNA double-strand breaks (DSBs) are mainly repaired by non-homologous end-joining (NHEJ) pathways. It has been demonstrated that NHEJ deficiencies are associated with impaired neurogenesis. In our study, rats were randomly divided into five groups to be irradiated by single doses of 0 (control), 0 (anesthesia control), 2, 10, and 20 Gy, respectively. The cognitive function of the irradiated rats was measured by open field, Morris water maze and passive avoidance tests. Real-time PCR was also used to detect the expression level of DNA DSB repair-related genes involved in the NHEJ pathway, such as XRCC4, XRCC5 and XRCC6, in the hippocampus. The influence of different radiation doses on cognitive function in rats was investigated. From the results of the behavior tests, we found that rats receiving 20 Gy irradiation revealed poorer learning and memory, while no significant loss of learning and memory existed in rats receiving irradiation from 0-10 Gy. The real-time PCR and Western blot results showed no significant difference in the expression level of DNA repair-related genes between the 10 and 20 Gy groups, which may help to explain the behavioral results, id est (i.e.) DNA damage caused by 0-10 Gy exposure was appropriately repaired, however, damage induced by 20 Gy exceeded the body's maximum DSB repair ability. Ionizing radiation-induced cognitive impairments depend on the radiation dose, and more directly on the body's own ability to repair DNA DSBs via the NHEJ pathway. (author)

  14. Welding and Joining of Titanium Aluminides

    Science.gov (United States)

    Cao, Jian; Qi, Junlei; Song, Xiaoguo; Feng, Jicai

    2014-01-01

    Welding and joining of titanium aluminides is the key to making them more attractive in industrial fields. The purpose of this review is to provide a comprehensive overview of recent progress in welding and joining of titanium aluminides, as well as to introduce current research and application. The possible methods available for titanium aluminides involve brazing, diffusion bonding, fusion welding, friction welding and reactive joining. Of the numerous methods, solid-state diffusion bonding and vacuum brazing have been most heavily investigated for producing reliable joints. The current state of understanding and development of every welding and joining method for titanium aluminides is addressed respectively. The focus is on the fundamental understanding of microstructure characteristics and processing–microstructure–property relationships in the welding and joining of titanium aluminides to themselves and to other materials. PMID:28788113

  15. Fumarase is involved in DNA double-strand break resection through a functional interaction with Sae2

    DEFF Research Database (Denmark)

    Leshets, Michael; Ramamurthy, Dharanidharan; Lisby, Michael

    2018-01-01

    One of the most severe forms of DNA damage is the double-strand break (DSB). Failure to properly repair the damage can cause mutation, gross chromosomal rearrangements and lead to the development of cancer. In eukaryotes, homologous recombination (HR) and non-homologous end joining (NHEJ) are the......One of the most severe forms of DNA damage is the double-strand break (DSB). Failure to properly repair the damage can cause mutation, gross chromosomal rearrangements and lead to the development of cancer. In eukaryotes, homologous recombination (HR) and non-homologous end joining (NHEJ......) are the main DSB repair pathways. Fumarase is a mitochondrial enzyme which functions in the tricarboxylic acid cycle. Intriguingly, the enzyme can be readily detected in the cytosolic compartment of all organisms examined, and we have shown that cytosolic fumarase participates in the DNA damage response...

  16. Dynamic dependence on ATR and ATM for double-strand break repair in human embryonic stem cells and neural descendants.

    Directory of Open Access Journals (Sweden)

    Bret R Adams

    2010-04-01

    Full Text Available The DNA double-strand break (DSB is the most toxic form of DNA damage. Studies aimed at characterizing DNA repair during development suggest that homologous recombination repair (HRR is more critical in pluripotent cells compared to differentiated somatic cells in which nonhomologous end joining (NHEJ is dominant. We have characterized the DNA damage response (DDR and quality of DNA double-strand break (DSB repair in human embryonic stem cells (hESCs, and in vitro-derived neural cells. Resolution of ionizing radiation-induced foci (IRIF was used as a surrogate for DSB repair. The resolution of gamma-H2AX foci occurred at a slower rate in hESCs compared to neural progenitors (NPs and astrocytes perhaps reflective of more complex DSB repair in hESCs. In addition, the resolution of RAD51 foci, indicative of active homologous recombination repair (HRR, showed that hESCs as well as NPs have high capacity for HRR, whereas astrocytes do not. Importantly, the ATM kinase was shown to be critical for foci formation in astrocytes, but not in hESCs, suggesting that the DDR is different in these cells. Blocking the ATM kinase in astrocytes not only prevented the formation but also completely disassembled preformed repair foci. The ability of hESCs to form IRIF was abrogated with caffeine and siRNAs targeted against ATR, implicating that hESCs rely on ATR, rather than ATM for regulating DSB repair. This relationship dynamically changed as cells differentiated. Interestingly, while the inhibition of the DNA-PKcs kinase (and presumably non-homologous endjoining [NHEJ] in astrocytes slowed IRIF resolution it did not in hESCs, suggesting that repair in hESCs does not utilize DNA-PKcs. Altogether, our results show that hESCs have efficient DSB repair that is largely ATR-dependent HRR, whereas astrocytes critically depend on ATM for NHEJ, which, in part, is DNA-PKcs-independent.

  17. Genome Editing in Escherichia coli with Cas9 and synthetic CRISPRs

    Energy Technology Data Exchange (ETDEWEB)

    Peng, Ze; Richardson, Sarah; Robinson, David; Deutsch, Samuel; Cheng, Jan-Fang

    2014-03-14

    Recently, the Cas9-CRISPR system has proven to be a useful tool for genome editing in eukaryotes, which repair the double stranded breaks made by Cas9 with non-homologous end joining or homologous recombination. Escherichia coli lacks non-homologous end joining and has a very low homologous recombination rate, effectively rendering targeted Cas9 activity lethal. We have developed a heat curable, serializable, plasmid based system for selectionless Cas9 editing in arbitrary E. coli strains that uses synthetic CRISPRs for targeting and -red to effect repairs of double stranded breaks. We have demonstrated insertions, substitutions, and multi-target deletions with our system, which we have tested in several strains.

  18. Welding and Joining of Titanium Aluminides

    Directory of Open Access Journals (Sweden)

    Jian Cao

    2014-06-01

    Full Text Available Welding and joining of titanium aluminides is the key to making them more attractive in industrial fields. The purpose of this review is to provide a comprehensive overview of recent progress in welding and joining of titanium aluminides, as well as to introduce current research and application. The possible methods available for titanium aluminides involve brazing, diffusion bonding, fusion welding, friction welding and reactive joining. Of the numerous methods, solid-state diffusion bonding and vacuum brazing have been most heavily investigated for producing reliable joints. The current state of understanding and development of every welding and joining method for titanium aluminides is addressed respectively. The focus is on the fundamental understanding of microstructure characteristics and processing–microstructure–property relationships in the welding and joining of titanium aluminides to themselves and to other materials.

  19. Chromosomal Integrity after UV Irradiation Requires FANCD2-Mediated Repair of Double Strand Breaks.

    Science.gov (United States)

    Federico, María Belén; Vallerga, María Belén; Radl, Analía; Paviolo, Natalia Soledad; Bocco, José Luis; Di Giorgio, Marina; Soria, Gastón; Gottifredi, Vanesa

    2016-01-01

    Fanconi Anemia (FA) is a rare autosomal recessive disorder characterized by hypersensitivity to inter-strand crosslinks (ICLs). FANCD2, a central factor of the FA pathway, is essential for the repair of double strand breaks (DSBs) generated during fork collapse at ICLs. While lesions different from ICLs can also trigger fork collapse, the contribution of FANCD2 to the resolution of replication-coupled DSBs generated independently from ICLs is unknown. Intriguingly, FANCD2 is readily activated after UV irradiation, a DNA-damaging agent that generates predominantly intra-strand crosslinks but not ICLs. Hence, UV irradiation is an ideal tool to explore the contribution of FANCD2 to the DNA damage response triggered by DNA lesions other than ICL repair. Here we show that, in contrast to ICL-causing agents, UV radiation compromises cell survival independently from FANCD2. In agreement, FANCD2 depletion does not increase the amount of DSBs generated during the replication of UV-damaged DNA and is dispensable for UV-induced checkpoint activation. Remarkably however, FANCD2 protects UV-dependent, replication-coupled DSBs from aberrant processing by non-homologous end joining, preventing the accumulation of micronuclei and chromatid aberrations including non-homologous chromatid exchanges. Hence, while dispensable for cell survival, FANCD2 selectively safeguards chromosomal stability after UV-triggered replication stress.

  20. Understanding the role of RecN in DSB repair pathway in Deinococcus radiodurans

    International Nuclear Information System (INIS)

    Pellegrino, S.

    2012-01-01

    Deinococcus radiodurans is a Gram-positive bacterium known for its extreme resistance to a broad variety of DNA damaging agents. Among these, Ionizing Radiations and desiccation are the most harmful for the cell, since they introduce breaks in the genome. Double Strand Breaks (DSB) are particularly hazardous for the cell and they need to be repaired very efficiently, in order to avoid mutations leading to altered, if not lethal, phenotypes. Homologous Recombination (HR) is the most efficient mechanism by which DSBs are repaired. D. radiodurans is able to completely restore its genome in only 3 hours, and it accomplishes the entire process through the RecFOR pathway. In order to be repaired, DSBs first need to be recognized. The protein believed to be responsible for this important step that takes place soon after the damage occurs in the cell, is RecN. RecN is recruited at the early stages of DNA repair and in vivo studies have demonstrated its propensity to localize to discrete foci. In vitro studies also suggest that RecN possesses a DNA end-joining activity previously observed for SMC proteins (such as cohesin), which are structurally related to RecN. Several structural studies have been carried out on the SMC-like protein, Rad50, but so far no structural information is available for RecN. The work presented here focused on the structural characterization of RecN and its constitutive domains. We obtained crystal structures of three partially overlapping constructs of RecN and Small Angle X-ray Scattering was performed on the individual domains and the full-length protein. The study of RecN in solution complemented our crystallographic study and enabled us to build a reliable, atomic model of the full-length protein. Mutations were designed and the mutant RecN proteins were produced in order to characterize the ATP hydrolysis activity of RecN, which is a conserved feature of this family of proteins. Extensive biochemical studies were carried out on wild-type and

  1. Ring-constrained Join

    DEFF Research Database (Denmark)

    Yiu, Man Lung; Karras, Panagiotis; Mamoulis, Nikos

    2008-01-01

    . This new operation has important applications in decision support, e.g., placing recycling stations at fair locations between restaurants and residential complexes. Clearly, RCJ is defined based on a geometric constraint but not on distances between points. Thus, our operation is fundamentally different......We introduce a novel spatial join operator, the ring-constrained join (RCJ). Given two sets P and Q of spatial points, the result of RCJ consists of pairs (p, q) (where p ε P, q ε Q) satisfying an intuitive geometric constraint: the smallest circle enclosing p and q contains no other points in P, Q...

  2. Performance Comparison of the Optimized Inverted Joined Wing Airplane Concept and Classical Configuration Airplanes

    Directory of Open Access Journals (Sweden)

    Sieradzki Adam

    2016-09-01

    Full Text Available The joined wing concept is an unconventional airplane configuration, known since the mid-twenties of the last century. It has several possible advantages, like reduction of the induced drag and weight due to the closed wing concept. The inverted joined wing variant is its rarely considered version, with the front wing being situated above the aft wing. The following paper presents a performance prediction of the recently optimized configuration of this airplane. Flight characteristics obtained numerically were compared with the performance of two classical configuration airplanes of similar category. Their computational fluid dynamics (CFD models were created basing on available documentation, photographs and some inverse engineering methods. The analysis included simulations performed for a scale of 3-meter wingspan inverted joined wing demonstrator and also for real-scale manned airplanes. Therefore, the results of CFD calculations allowed us to assess the competitiveness of the presented concept, as compared to the most technologically advanced airplanes designed and manufactured to date. At the end of the paper, the areas where the inverted joined wing is better than conventional airplane were predicted and new research possibilities were described.

  3. Polychlorinated biphenyl quinone induces oxidative DNA damage and repair responses: The activations of NHEJ, BER and NER via ATM-p53 signaling axis

    Energy Technology Data Exchange (ETDEWEB)

    Dong, Hui; Shi, Qiong; Song, Xiufang; Fu, Juanli; Hu, Lihua; Xu, Demei; Su, Chuanyang; Xia, Xiaomin; Song, Erqun; Song, Yang, E-mail: songyangwenrong@hotmail.com

    2015-07-01

    Our previous studies demonstrated that polychlorinated biphenyl (PCB) quinone induced oxidative DNA damage in HepG2 cells. To promote genomic integrity, DNA damage response (DDR) coordinates cell-cycle transitions, DNA repair and apoptosis. PCB quinone-induced cell cycle arrest and apoptosis have been documented, however, whether PCB quinone insult induce DNA repair signaling is still unknown. In this study, we identified the activation of DDR and corresponding signaling events in HepG2 cells upon the exposure to a synthetic PCB quinone, PCB29-pQ. Our data illustrated that PCB29-pQ induces the phosphorylation of p53, which was mediated by ataxia telangiectasia mutated (ATM) protein kinase. The observed phosphorylated histone H2AX (γ-H2AX) foci and the elevation of 8-hydroxy-2′-deoxyguanosine (8-OHdG) indicated that DDR was stimulated by PCB29-pQ treatment. Additionally, we found PCB29-pQ activates non-homologous end joining (NHEJ), base excision repair (BER) and nucleotide excision repair (NER) signalings. However, these repair pathways are not error-free processes and aberrant repair of DNA damage may cause the potential risk of carcinogenesis and mutagenesis. - Highlights: • Polychlorinated biphenyl quinone induces oxidative DNA damage in HepG2 cells. • The elevation of γ-H2AX and 8-OHdG indicates the activation of DNA damage response. • ATM-p53 signaling acts as the DNA damage sensor and effector. • Polychlorinated biphenyl quinone activates NHEJ, BER and NER signalings.

  4. Anatomic deltoid ligament repair with anchor-to-post suture reinforcement: technique tip.

    Science.gov (United States)

    Lack, William; Phisitkul, Phinit; Femino, John E

    2012-01-01

    The deltoid ligament is the primary ligamentous stabilizer of the ankle joint. Both superficial and deep components of the ligament can be disrupted with a rotational ankle fracture, chronic ankle instability, or in late stage adult acquired flatfoot deformity. The role of deltoid ligament repair in these conditions has been limited and its contribution to arthritis is largely unknown. Neglect of the deltoid ligament in the treatment of ankle injuries may be due to difficulties in diagnosis and lack of an effective method for repair. Most acute repair techniques address the superficial deltoid ligament with direct end-to-end repair, fixation through bone tunnels, or suture anchor repair of avulsion injuries. Deep deltoid ligament repair has been described using direct end-to-end repair with sutures, as well as by autograft and allograft tendon reconstruction utilizing various techniques. Newer tenodesis techniques have been described for late reconstruction of both deep and superficial components in patients with stage 4 adult acquired flatfoot deformity. We describe a technique that provides anatomic ligament-to-bone repair of the superficial and deep bundles of the deltoid ligament while reducing the talus toward the medial malleolar facet of the tibiotalar joint with anchor-to-post reinforcement of the ligamentous repair. This technique may protect and allow the horizontally oriented fibers of the deep deltoid ligament to heal with the appropriate resting length while providing immediate stability of the construct.

  5. Gastroesophageal junction adenocarcinoma displays abnormalities in homologous recombination and nucleotide excision repair

    Directory of Open Access Journals (Sweden)

    Dewalt RI

    2014-02-01

    Full Text Available Robin I Dewalt,1 Kenneth A Kesler,2 Zane T Hammoud,3 LeeAnn Baldridge,4 Eyas M Hattab,4 Shadia I Jalal1,5 1Division of Hematology/Oncology, Department of Medicine, 2Cardiothoracic Division, Department of Surgery, Indiana University School of Medicine, Indianapolis, IN, USA; 3Henry Ford Hospital, Detroit, MI, USA; 4Department of Pathology and Laboratory Medicine, Indiana University School of Medicine, Indianapolis, IN, USA; 5Indiana University Melvin and Bren Simon Cancer Center, Indianapolis, IN, USA Objective: Esophageal adenocarcinoma (EAC continues to be a disease associated with high mortality. Among the factors leading to poor outcomes are innate resistance to currently available therapies, advanced stage at diagnosis, and complex biology. Platinum and ionizing radiation form the backbone of treatment for the majority of patients with EAC. Of the multiple processes involved in response to platinum chemotherapy or ionizing radiation, deoxyribonucleic acid (DNA repair has been a major player in cancer sensitivity to these agents. DNA repair defects have been described in various malignancies. The purpose of this study was to determine whether alterations in DNA repair are present in EAC compared with normal gastroesophageal tissues. Methods: We analyzed the expression of genes involved in homologous recombination (HR, nonhomologous end-joining, and nucleotide excision repair (NER pathways in 12 EAC tumor samples with their matched normal counterparts. These pathways were chosen because they are the main pathways involved in the repair of platinum- or ionizing-radiation-induced damage. In addition, abnormalities in these pathways have not been well characterized in EAC. Results: We identified increased expression of at least one HR gene in eight of the EAC tumor samples. Alterations in the expression of EME1, a structure-specific endonuclease involved in HR, were the most prevalent, with messenger (mRNA overexpression in six of the EAC samples

  6. Constitutional chromothripsis rearrangements involve clustered double-stranded DNA breaks and nonhomologous repair mechanisms.

    Science.gov (United States)

    Kloosterman, Wigard P; Tavakoli-Yaraki, Masoumeh; van Roosmalen, Markus J; van Binsbergen, Ellen; Renkens, Ivo; Duran, Karen; Ballarati, Lucia; Vergult, Sarah; Giardino, Daniela; Hansson, Kerstin; Ruivenkamp, Claudia A L; Jager, Myrthe; van Haeringen, Arie; Ippel, Elly F; Haaf, Thomas; Passarge, Eberhard; Hochstenbach, Ron; Menten, Björn; Larizza, Lidia; Guryev, Victor; Poot, Martin; Cuppen, Edwin

    2012-06-28

    Chromothripsis represents a novel phenomenon in the structural variation landscape of cancer genomes. Here, we analyze the genomes of ten patients with congenital disease who were preselected to carry complex chromosomal rearrangements with more than two breakpoints. The rearrangements displayed unanticipated complexity resembling chromothripsis. We find that eight of them contain hallmarks of multiple clustered double-stranded DNA breaks (DSBs) on one or more chromosomes. In addition, nucleotide resolution analysis of 98 breakpoint junctions indicates that break repair involves nonhomologous or microhomology-mediated end joining. We observed that these eight rearrangements are balanced or contain sporadic deletions ranging in size between a few hundred base pairs and several megabases. The two remaining complex rearrangements did not display signs of DSBs and contain duplications, indicative of rearrangement processes involving template switching. Our work provides detailed insight into the characteristics of chromothripsis and supports a role for clustered DSBs driving some constitutional chromothripsis rearrangements. Copyright © 2012 The Authors. Published by Elsevier Inc. All rights reserved.

  7. Major Roles for Pyrimidine Dimers, Nucleotide Excision Repair, and ATR in the Alternative Splicing Response to UV Irradiation

    Directory of Open Access Journals (Sweden)

    Manuel J. Muñoz

    2017-03-01

    Full Text Available We have previously found that UV irradiation promotes RNA polymerase II (RNAPII hyperphosphorylation and subsequent changes in alternative splicing (AS. We show now that UV-induced DNA damage is not only necessary but sufficient to trigger the AS response and that photolyase-mediated removal of the most abundant class of pyrimidine dimers (PDs abrogates the global response to UV. We demonstrate that, in keratinocytes, RNAPII is the target, but not a sensor, of the signaling cascade initiated by PDs. The UV effect is enhanced by inhibition of gap-filling DNA synthesis, the last step in the nucleotide excision repair pathway (NER, and reduced by the absence of XPE, the main NER sensor of PDs. The mechanism involves activation of the protein kinase ATR that mediates the UV-induced RNAPII hyperphosphorylation. Our results define the sequence UV-PDs-NER-ATR-RNAPII-AS as a pathway linking DNA damage repair to the control of both RNAPII phosphorylation and AS regulation.

  8. Oxidative DNA Damage in Neurons: Implication of Ku in Neuronal Homeostasis and Survival

    Directory of Open Access Journals (Sweden)

    Daniela De Zio

    2012-01-01

    Full Text Available Oxidative DNA damage is produced by reactive oxygen species (ROS which are generated by exogenous and endogenous sources and continuously challenge the cell. One of the most severe DNA lesions is the double-strand break (DSB, which is mainly repaired by nonhomologous end joining (NHEJ pathway in mammals. NHEJ directly joins the broken ends, without using the homologous template. Ku70/86 heterodimer, also known as Ku, is the first component of NHEJ as it directly binds DNA and recruits other NHEJ factors to promote the repair of the broken ends. Neurons are particularly metabolically active, displaying high rates of transcription and translation, which are associated with high metabolic and mitochondrial activity as well as oxygen consumption. In such a way, excessive oxygen radicals can be generated and constantly attack DNA, thereby producing several lesions. This condition, together with defective DNA repair systems, can lead to a high accumulation of DNA damage resulting in neurodegenerative processes and defects in neurodevelopment. In light of recent findings, in this paper, we will discuss the possible implication of Ku in neurodevelopment and in mediating the DNA repair dysfunction observed in certain neurodegenerations.

  9. Comparison of Direct Side-to-End and End-to-End Hypoglossal-Facial Anastomosis for Facial Nerve Repair.

    Science.gov (United States)

    Samii, Madjid; Alimohamadi, Maysam; Khouzani, Reza Karimi; Rashid, Masoud Rafizadeh; Gerganov, Venelin

    2015-08-01

    The hypoglossal facial anastomosis (HFA) is the gold standard for facial reanimation in patients with severe facial nerve palsy. The major drawbacks of the classic HFA technique are lingual morbidities due to hypoglossal nerve transection. The side-to-end HFA is a modification of the classic technique with fewer tongue-related morbidities. In this study we compared the outcome of the classic end-to-end and the direct side-to-end HFA surgeries performed at our center in regards to the facial reanimation success rate and tongue-related morbidities. Twenty-six successive cases of HFA were enrolled. In 9 of them end-to-end anastomoses were performed, and 17 had direct side-to-end anastomoses. The House-Brackmann (HB) and Pitty and Tator (PT) scales were used to document surgical outcome. The hemiglossal atrophy, swallowing, and hypoglossal nerve function were assessed at follow-up. The original pathology was vestibular schwannoma in 15, meningioma in 4, brain stem glioma in 4, and other pathologies in 3. The mean interval between facial palsy and HFA was 18 months (range: 0-60). The median follow-up period was 20 months. The PT grade at follow-up was worse in patients with a longer interval from facial palsy and HFA (P value: 0.041). The lesion type was the only other factor that affected PT grade (the best results in vestibular schwannoma and the worst in the other pathologies group, P value: 0.038). The recovery period for facial tonicity was longer in patients with radiation therapy before HFA (13.5 vs. 8.5 months) and those with a longer than 2-year interval from facial palsy to HFA (13.5 vs. 8.5 months). Although no significant difference between the side-to-end and the end-to-end groups was seen in terms of facial nerve functional recovery, patients from the side-to-end group had a significantly lower rate of lingual morbidities (tongue hemiatrophy: 100% vs. 5.8%, swallowing difficulty: 55% vs. 11.7%, speech disorder 33% vs. 0%). With the side-to-end HFA

  10. Welding repair of a dissimilar weld and respective consequences for other German plants

    International Nuclear Information System (INIS)

    Brummer, G.; Dauwel, W.; Wesseling, U.; Ilg, U.; Lauer, P.; Widera, M.; Wachter, O.

    2002-01-01

    During a regular refueling outage in a German nuclear power plant in year 2000, additional non-destructive examinations have been performed on request of the Authority, to fulfill some recommendations of the independent experts with regard to the retrospective application of the Basic Safety Concept for the ferritic main coolant piping of this plant. During these inspections, indications were found in a dissimilar weld between one of the fifteen MCL (main coolant lines) nozzles and the ECC (emergency core cooling) system piping. By means of on-site metallography and laboratory investigations on three boat samples taken from this weld, it could be shown that the indications were due to hot cracking in the surface layer of the weld. In the course of these investigations, at three locations at the circumference of the weld, dis-bonding defects were found between the ferritic base metal of the nozzle and the austenitic weld butter, which has been applied to join the nozzle to the austenitic safe-end. According to the results of the extensive investigations, the dis-bonding occurred during the manufacturing process after stress-relief heat-treatment of the buttering during the welding of the austenitic safe-end to the butter material. There was no evidence for any crack growth during operation of the plant. Due to the large size of the boat-samples, a weld repair was mandatory. This repair has been performed using the so-called temper-bead technique as specified in the ASME Code, without subsequent stress relief heat treatment, using an advanced automatic orbital TIG welding process. The welding has been successfully performed without the need of further repair work. For those dissimilar welds, all other plants, except one, had used Inconel welding material for buttering the ferritic nozzle instead of stainless steel welding metal. For metallurgical reasons, dis-bonding along the fusion line for Inconel buttered dissimilar welds is unlikely to occur. Nevertheless all

  11. The time course of repair of ultraviolet-induced DNA damage; implications for the structural organization of repair

    International Nuclear Information System (INIS)

    Collins, A.; Squires, S.

    1986-01-01

    Alternative molecular mechanisms can be envisaged for the cellular repair of UV-damaged DNA. In the 'random collision' model, DNA damage distributed throughout the genome is recognised and repaired by a process of random collision between DNA damage and repair enzymes. The other model assumes a 'processive' mechanism, whereby DNA is scanned for damage by a repair complex moving steadily along its length. Random collision should result in a declining rate of repair with time as the concentration of lesions in the DNA falls; but the processive model predicts a constant rate until scanning is complete. The authors have examined the time course of DNA repair in human fibroblasts given low doses of UV light. Using 3 distinct assays, the authors find no sign of a constant repair rate after 4 J/m 2 or less, even when the first few hours after irradiation are examined. Thus DNA repair is likely to depend on random collision. (Auth.)

  12. Reduction of spontaneous somatic mutation frequency by a low-dose X irradiation of Drosophila larvae and possible involvement of DNA single-strand damage repair.

    Science.gov (United States)

    Koana, Takao; Takahashi, Takashi; Tsujimura, Hidenobu

    2012-03-01

    The third instar larvae of Drosophila were irradiated with X rays, and the somatic mutation frequency in their wings was measured after their eclosion. In the flies with normal DNA repair and apoptosis functions, 0.2 Gy irradiation at 0.05 Gy/min reduced the frequency of the so-called small spot (mutant cell clone with reduced reproductive activity) compared with that in the sham-irradiated flies. When apoptosis was suppressed using the baculovirus p35 gene, the small spot frequency increased four times in the sham-irradiated control group, but the reduction by the 0.2-Gy irradiation was still evident. In a non-homologous end joining-deficient mutant, the small spot frequency was also reduced by 0.2 Gy radiation. In a mutant deficient in single-strand break repair, no reduction in the small spot frequency by 0.2 Gy radiation was observed, and the small spot frequency increased with the radiation dose. Large spot (mutant cell clone with normal reproductive activity) frequency was not affected by suppression of apoptosis and increased monotonically with radiation dose in wild-type larvae and in mutants for single- or double-strand break repair. It is hypothesized that some of the small spots resulted from single-strand damage and, in wild-type larvae, 0.2 Gy radiation activated the normal single-strand break repair gene, which reduced the background somatic mutation frequency.

  13. Exonuclease 1 is a critical mediator of survival during DNA double strand break repair in nonquiescent hematopoietic stem and progenitor cells.

    Science.gov (United States)

    Desai, Amar; Qing, Yulan; Gerson, Stanton L

    2014-02-01

    Hematopoietic stem cell (HSC) populations require DNA repair pathways to maintain their long-term survival and reconstitution capabilities, but mediators of these processes are still being elucidated. Exonuclease 1 (Exo1) participates in homologous recombination (HR) and Exo1 loss results in impaired 5' HR end resection. We use cultured Exo1(mut) fibroblasts and bone marrow to demonstrate that loss of Exo1 function results in defective HR in cycling cells. Conversely, in Exo1(mut) mice HR is not required for maintenance of quiescent HSCs at steady state, confirming the steady state HSC reliance on nonhomologous end joining (NHEJ). Exo1(mut) mice sustained serial repopulation, displayed no defect in competitive repopulation or niche occupancy, and exhibited no increased sensitivity to whole body ionizing radiation. However, when Exo1(mut) HSCs were pushed into cell cycle in vivo with 5-fluorouracil or poly IC, the hematopoietic population became hypersensitive to IR, resulting in HSC defects and animal death. We propose Exo1-mediated HR is dispensable for stem cell function in quiescent HSC, whereas it is essential to HSC response to DNA damage processing after cell cycle entry, and its loss is not compensated by intact NHEJ. In HSCs, the maintenance of stem cell function after DNA damage is dependent on the DNA repair capacity, segregated by active versus quiescent points in cell cycle. © AlphaMed Press.

  14. Potts shunt in a child with end-stage pulmonary hypertension after late repair of ventricular septal defect

    DEFF Research Database (Denmark)

    Petersen, Cecilie; Helvind, Morten; Jensen, Tim

    2013-01-01

    We report on a 10-year-old boy with medically refractory pulmonary arterial hypertension (PAH) and end-stage right heart failure after closure of a ventricular septal defect. The boy was a candidate for lung transplantation (LTX), but an alternative option was to create an Eisenmenger physiology ...

  15. Trajectory similarity join in spatial networks

    KAUST Repository

    Shang, Shuo

    2017-09-07

    The matching of similar pairs of objects, called similarity join, is fundamental functionality in data management. We consider the case of trajectory similarity join (TS-Join), where the objects are trajectories of vehicles moving in road networks. Thus, given two sets of trajectories and a threshold θ, the TS-Join returns all pairs of trajectories from the two sets with similarity above θ. This join targets applications such as trajectory near-duplicate detection, data cleaning, ridesharing recommendation, and traffic congestion prediction. With these applications in mind, we provide a purposeful definition of similarity. To enable efficient TS-Join processing on large sets of trajectories, we develop search space pruning techniques and take into account the parallel processing capabilities of modern processors. Specifically, we present a two-phase divide-and-conquer algorithm. For each trajectory, the algorithm first finds similar trajectories. Then it merges the results to achieve a final result. The algorithm exploits an upper bound on the spatiotemporal similarity and a heuristic scheduling strategy for search space pruning. The algorithm\\'s per-trajectory searches are independent of each other and can be performed in parallel, and the merging has constant cost. An empirical study with real data offers insight in the performance of the algorithm and demonstrates that is capable of outperforming a well-designed baseline algorithm by an order of magnitude.

  16. An inverse switch in DNA base excision and strand break repair contributes to melphalan resistance in multiple myeloma cells.

    Directory of Open Access Journals (Sweden)

    Mirta M L Sousa

    Full Text Available Alterations in checkpoint and DNA repair pathways may provide adaptive mechanisms contributing to acquired drug resistance. Here, we investigated the levels of proteins mediating DNA damage signaling and -repair in RPMI8226 multiple myeloma cells and its Melphalan-resistant derivative 8226-LR5. We observed markedly reduced steady-state levels of DNA glycosylases UNG2, NEIL1 and MPG in the resistant cells and cross-resistance to agents inducing their respective DNA base lesions. Conversely, repair of alkali-labile sites was apparently enhanced in the resistant cells, as substantiated by alkaline comet assay, autoribosylation of PARP-1, and increased sensitivity to PARP-1 inhibition by 4-AN or KU58684. Reduced base-excision and enhanced single-strand break repair would both contribute to the observed reduction in genomic alkali-labile sites, which could jeopardize productive processing of the more cytotoxic Melphalan-induced interstrand DNA crosslinks (ICLs. Furthermore, we found a marked upregulation of proteins in the non-homologous end-joining (NHEJ pathway of double-strand break (DSB repair, likely contributing to the observed increase in DSB repair kinetics in the resistant cells. Finally, we observed apparent upregulation of ATR-signaling and downregulation of ATM-signaling in the resistant cells. This was accompanied by markedly increased sensitivity towards Melphalan in the presence of ATR-, DNA-PK, or CHK1/2 inhibitors whereas no sensitizing effect was observed subsequent to ATM inhibition, suggesting that replication blocking lesions are primary triggers of the DNA damage response in the Melphalan resistant cells. In conclusion, Melphalan resistance is apparently contributed by modulation of the DNA damage response at multiple levels, including downregulation of specific repair pathways to avoid repair intermediates that could impair efficient processing of cytotoxic ICLs and ICL-induced DSBs. This study has revealed several novel

  17. Recrafting the neighbor-joining method

    Directory of Open Access Journals (Sweden)

    Pedersen Christian NS

    2006-01-01

    Full Text Available Abstract Background The neighbor-joining method by Saitou and Nei is a widely used method for constructing phylogenetic trees. The formulation of the method gives rise to a canonical Θ(n3 algorithm upon which all existing implementations are based. Results In this paper we present techniques for speeding up the canonical neighbor-joining method. Our algorithms construct the same phylogenetic trees as the canonical neighbor-joining method. The best-case running time of our algorithms are O(n2 but the worst-case remains O(n3. We empirically evaluate the performance of our algoritms on distance matrices obtained from the Pfam collection of alignments. The experiments indicate that the running time of our algorithms evolve as Θ(n2 on the examined instance collection. We also compare the running time with that of the QuickTree tool, a widely used efficient implementation of the canonical neighbor-joining method. Conclusion The experiments show that our algorithms also yield a significant speed-up, already for medium sized instances.

  18. Willmore energy for joining of carbon nanostructures

    Science.gov (United States)

    Sripaturad, P.; Alshammari, N. A.; Thamwattana, N.; McCoy, J. A.; Baowan, D.

    2018-06-01

    Numerous types of carbon nanostructure have been found experimentally, including nanotubes, fullerenes and nanocones. These structures have applications in various nanoscale devices and the joining of these structures may lead to further new configurations with more remarkable properties and applications. The join profile between different carbon nanostructures in a symmetric configuration may be modelled using the calculus of variations. In previous studies, carbon nanostructures were assumed to deform according to perfect elasticity, thus the elastic energy, depending only on the axial curvature, was used to determine the join profile consisting of a finite number of discrete bonds. However, one could argue that the relevant energy should also involve the rotational curvature, especially when its size is comparable to the axial curvature. In this paper, we use the Willmore energy, a natural generalisation of the elastic energy that depends on both the axial and rotational curvatures. Catenoids are absolute minimisers of this energy and pieces of these may be used to join various nanostructures. We focus on the cases of joining a fullerene to a nanotube and joining two fullerenes along a common axis. By comparing our results with the earlier work, we find that both energies give similar joining profiles. Further work on other configurations may reveal which energy provides a better model.

  19. Expanding the CRISPR/Cas9 toolkit for Pichia pastoris with efficient donor integration and alternative resistance markers.

    Science.gov (United States)

    Weninger, Astrid; Fischer, Jasmin E; Raschmanová, Hana; Kniely, Claudia; Vogl, Thomas; Glieder, Anton

    2018-04-01

    Komagataella phaffii (syn. Pichia pastoris) is one of the most commonly used host systems for recombinant protein expression. Achieving targeted genetic modifications had been hindered by low frequencies of homologous recombination (HR). Recently, a CRISPR/Cas9 genome editing system has been implemented for P. pastoris enabling gene knockouts based on indels (insertion, deletions) via non-homologous end joining (NHEJ) at near 100% efficiency. However, specifically integrating homologous donor cassettes via HR for replacement studies had proven difficult resulting at most in ∼20% correct integration using CRISPR/Cas9. Here, we demonstrate the CRISPR/Cas9 mediated integration of markerless donor cassettes at efficiencies approaching 100% using a ku70 deletion strain. The Ku70p is involved in NHEJ repair and lack of the protein appears to favor repair via HR near exclusively. While the absolute number of transformants in the Δku70 strain is reduced, virtually all surviving transformants showed correct integration. In the wildtype strain, markerless donor cassette integration was also improved up to 25-fold by placing an autonomously replicating sequence (ARS) on the donor cassette. Alternative strategies for improving donor cassette integration using a Cas9 nickase variant or reducing off targeting associated toxicity using a high fidelity Cas9 variant were so far not successful in our hands in P. pastoris. Furthermore we provide Cas9/gRNA expression plasmids with a Geneticin resistance marker which proved to be versatile tools for marker recycling. The reported CRSIPR-Cas9 tools can be applied for modifying existing production strains and also pave the way for markerless whole genome modification studies in P. pastoris. © 2017 The Authors. Journal of Cellular Biochemistry Published by Wiley Periodicals, Inc.

  20. Repair kinetics of DNA double-strand breaks and incidence of apoptosis in mouse neural stem/progenitor cells and their differentiated neurons exposed to ionizing radiation.

    Science.gov (United States)

    Kashiwagi, Hiroki; Shiraishi, Kazunori; Sakaguchi, Kenta; Nakahama, Tomoya; Kodama, Seiji

    2018-05-01

    Neuronal loss leads to neurodegenerative disorders, including Alzheimer's disease, Parkinson's disease and Huntington's disease. Because of their long lifespans, neurons are assumed to possess highly efficient DNA repair ability and to be able to protect themselves from deleterious DNA damage such as DNA double-strand breaks (DSBs) produced by intrinsic and extrinsic sources. However, it remains largely unknown whether the DSB repair ability of neurons is more efficient compared with that of other cells. Here, we investigated the repair kinetics of X-ray-induced DSBs in mouse neural cells by scoring the number of phosphorylated 53BP1 foci post irradiation. We found that p53-independent apoptosis was induced time dependently during differentiation from neural stem/progenitor cells (NSPCs) into neurons in culture for 48 h. DSB repair in neurons differentiated from NSPCs in culture was faster than that in mouse embryonic fibroblasts (MEFs), possibly due to the higher DNA-dependent protein kinase activity, but it was similar to that in NSPCs. Further, the incidence of p53-dependent apoptosis induced by X-irradiation in neurons was significantly higher than that in NSPCs. This difference in response of X-ray-induced apoptosis between neurons and NSPCs may reflect a difference in the fidelity of non-homologous end joining or a differential sensitivity to DNA damage other than DSBs.

  1. To repair the fault or end the acid reign?

    Science.gov (United States)

    Modlin, I M

    1995-01-01

    Despite the vast effort and expense devoted to the elucidation of the cause of esophagogastro-duodenal ulcer disease, relatively minimal progress has been made towards the understanding of causation. Since earliest times, it has been recognized that milk, chalk powder, or charcoal ameliorate the disease process and its symptoms. In addition, the avoidance of acidic or spicy foods provides some relief. Thus ulcer disease has been ascribed to acid or the consequences of hyperacidity. Reams of data and countless meetings have purported to confirm and support this viewpoint. Surprisingly, the co-secretion in the stomach of the powerful proteolytic enzyme pepsin has been virtually ignored as a pathogenetic agent. Marshalling distant antipodean resources, a novel bacteria, H. pylori, was identified as a significant causative agent in ulcer disease. To the amazement of nobody but gastroenterologists, it became apparent that there might be more than one cause for ulcer disease. Subsequently, both corporations and physicians seized the reins of multi-variant antibiotic therapy as a panacea for the third millennium treatment of peptic ulcer disease. Only a brave few have raised the issues of possible abnormalities in intrinsic mucosal function which might generate a locus minoris resistentiae. Defective mucosal repair mechanisms have barely been evaluated, since the regulation of normal mucosal healing is so poorly understood. Nevertheless, consideration of the therapeutic potential of mucosal protection has found support at both an intellectual and a clinical level. The more exciting recent possibility of the local delivery of growth factors which might promote healing has provided a unique opportunity for further therapeutic advance. Indeed in the future the exogenous regulation of mucosal repair may provide a milieu conductive to the resolution of an old but ill-understood problem. It is certainly apparent that processes beyond parietal cell proton secretion are critical

  2. Role of XRCC4 phosphorylation by DNA-PK in the regulation of NHEJ repair pathway of DNA double strand break

    International Nuclear Information System (INIS)

    Sharma, Mukesh Kumar; Imamichi, Shoji; Fukuchi, Mikoto; Kamdar, Radhika P.; Sicheng, Liu; Wanotayan, Rujira; Matsumoto, Yoshihisa

    2014-01-01

    Non-homologous end-joining (NHEJ) is the predominant pathway of DNA double strand breaks in higher eukaryotes and is active throughout the cell cycle. NHEJ repair includes many factors as Ku70/86, DNA-PKcs, XRCC4-Ligase IV complex and XLF (also known as Cernunnos). In these factors, DNA-PKcs acts as central regulator in NHEJ repair. It recruited at the DNA damages site after DNA damage and after association with Ku its kinase activity is activated. It phosphorylates many of important NHEJ proteins in vitro including XRCC4, Ku 70/86, Artemis, and even DNA-PKcs but till now, very less studies have been done to know the role and significance of phosphorylation in the NHEJ repair. Studies by other researchers identified various phosphorylation sites in XRCC4 by DNA-PK using mass spectrometry but these phosphorylation sites were shown to be dispensable for DSB repair. In the present investigation, we identified 3 serine and one new threonine phosphorylation sites in XRCC4 protein by DNA-PK. In vivo phosphorylation at these sites was verified by generating phosphorylation specific antibodies and the requirement for DNA-PK therein was verified by using DNA-PK inhibitor and DNA-PK proficient and deficient cell lines in response to radiation and zeocin treatment. We have also found that phosphorylation at these sites showed dose dependency in response to radiation treatment. The two serine and one threonine phosphorylation site is also biological important as their mutation into alanine significantly elevated radiosensitivity as measured by colony formation assay. Neutral comet assay showed delayed kinetics in DSB repair of these mutants. Furthermore, we have found a protein, with putative DSB repair function, which interacts with domain including the phosphorylation sites.These results indicate that these phosphorylation sites would mediate functional link between XRCC4 and DNA-PK. (author)

  3. Diverless pipeline repair system for deep water

    Energy Technology Data Exchange (ETDEWEB)

    Spinelli, Carlo M. [Eni Gas and Power, Milan (Italy); Fabbri, Sergio; Bachetta, Giuseppe [Saipem/SES, Venice (Italy)

    2009-07-01

    SiRCoS (Sistema Riparazione Condotte Sottomarine) is a diverless pipeline repair system composed of a suite of tools to perform a reliable subsea pipeline repair intervention in deep and ultra deep water which has been on the ground of the long lasting experience of Eni and Saipem in designing, laying and operating deep water pipelines. The key element of SiRCoS is a Connection System comprising two end connectors and a repair spool piece to replace a damaged pipeline section. A Repair Clamp with elastomeric seals is also available for pipe local damages. The Connection System is based on pipe cold forging process, consisting in swaging the pipe inside connectors with suitable profile, by using high pressure seawater. Three swaging operations have to be performed to replace the damaged pipe length. This technology has been developed through extensive theoretical work and laboratory testing, ending in a Type Approval by DNV over pipe sizes ranging from 20 inches to 48 inches OD. A complete SiRCoS system has been realised for the Green Stream pipeline, thoroughly tested in workshop as well as in shallow water and is now ready, in the event of an emergency situation.The key functional requirements for the system are: diverless repair intervention and fully piggability after repair. Eni owns this technology and is now available to other operators under Repair Club arrangement providing stand-by repair services carried out by Saipem Energy Services. The paper gives a description of the main features of the Repair System as well as an insight into the technological developments on pipe cold forging reliability and long term duration evaluation. (author)

  4. MICROWAVE JOINING OF ALUMINA CERAMIC AND HYDROXYLAPATITE BIOCERAMIC

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    Microwave joining is a rapid developmental new techniqu e in recent years.This paper introduces a new microwave joining equipment which was made by our lab,succeeds in alumina ceramic-hydroxylapatite bioceramic j o in in the equipment, and analyzes the join situation of join boundary by using s canni ng electron microscope(SEM),this paper analyzes the mechanism of microwave joini ng also.

  5. Repair of Defective Composite Resin Restoration: Current Trend ...

    African Journals Online (AJOL)

    Background: Repair of defective composite resins restorations is being increasingly recognized as a viable alternative to replacement. there is however no consensus yet on the treatment protocol. Objective: To determine the views and practice of specialists in Conservative Dentistry in Nigeria as regard to repair procedure ...

  6. Capacity of ultraviolet-induced DNA repair in human glioma cells

    Energy Technology Data Exchange (ETDEWEB)

    Itoh, Hiroji

    1987-04-01

    A DNA repair abnormality is likely related to an increased incidence of neoplasms in several autosomal recessive diseases such as xeroderma pigmentosum, Fanconi's anemia, Bloom's syndrome and ataxia telangiectasia. In human glioma cells, however, there are only a few reports on DNA repair. In this study, an ultraviolet (UV)-induced DNA repair was examined systematically in many human glioma cells. Two human malignant glioma cell lines (MMG-851, U-251-MG) and 7 human glioma cell strains (4, benign; 3, malignant) of short term culture, in which glial fibrillary acidic protein (GFAP) staining were positive, were used. To investigate the capacity of DNA repair, UV sensitivity was determined by colony formation; excision repair by autoradiography and Cytosine Arabinoside (Ara-C) assay; and post-replication repair by the joining rate of newly synthesized DNA. As a result, the colony-forming abilities of malignant glioma cell lines were lower than those of normal human fibroblasts, but no difference was found between two malignant glioma cell lines. The excision repair of the malignant group (2 cell lines and 3 cell strains) was apparently lower than that of the benign group (4 cell strains). In two malignant glioma cell lines, the excision repair of MMG-851 was lower than that of U-251-MG, and the post-replication repair of MMG-851 was higher than that of U-251-MG. These results were considered to correspond well with colony-forming ability. The results indicate that there are some differences in each human malignant glioma cell in its UV-induced DNA repair mechanism, and that the excision repair of the malignant glioma cells is apparently lower than that of the benign glioma cells. These findings may be useful for diagnosis and treatment.

  7. Pathways for double-strand break repair in genetically unstable Z-DNA-forming sequences.

    Science.gov (United States)

    Kha, Diem T; Wang, Guliang; Natrajan, Nithya; Harrison, Lynn; Vasquez, Karen M

    2010-05-14

    DNA can adopt many structures that differ from the canonical B-form, and several of these non-canonical DNA structures have been implicated in genetic instability associated with human disease. Earlier, we found that Z-DNA causes DNA double-strand breaks (DSBs) in mammalian cells that can result in large-scale deletions and rearrangements. In contrast, the same Z-DNA-forming CG repeat in Escherichia coli resulted in only small contractions or expansions within the repeat. This difference in the Z-DNA-induced mutation spectrum between mammals and bacteria might be due to different mechanisms for DSB repair; in mammalian cells, non-homologous end-joining (NHEJ) is a major DSB repair pathway, while E. coli do not contain this system and typically use homologous recombination (HR) to process DSBs. To test the extent to which the different DSB repair pathways influenced the Z-DNA-induced mutagenesis, we engineered bacterial E.coli strains to express an inducible NHEJ system, to mimic the situation in mammalian cells. Mycobacterium tuberculosis NHEJ proteins Ku and ligase D (LigD) were expressed in E.coli cells in the presence or absence of HR, and the Z-DNA-induced mutations were characterized. We found that the presence of the NHEJ mechanism markedly shifted the mutation spectrum from small deletions/insertions to large-scale deletions (from 2% to 24%). Our results demonstrate that NHEJ plays a role in the generation of Z-DNA-induced large-scale deletions, suggesting that this pathway is associated with DNA structure-induced destabilization of genomes from prokaryotes to eukaryotes. (c) 2010 Elsevier Ltd. All rights reserved.

  8. Transient elevation of glycolysis confers radio-resistance by facilitating DNA repair in cells

    International Nuclear Information System (INIS)

    Bhatt, Anant Narayan; Chauhan, Ankit; Khanna, Suchit; Rai, Yogesh; Singh, Saurabh; Soni, Ravi; Kalra, Namita; Dwarakanath, Bilikere S

    2015-01-01

    Cancer cells exhibit increased glycolysis for ATP production (the Warburg effect) and macromolecular biosynthesis; it is also linked with therapeutic resistance that is generally associated with compromised respiratory metabolism. Molecular mechanisms underlying radio-resistance linked to elevated glycolysis remain incompletely understood. We stimulated glycolysis using mitochondrial respiratory modifiers (MRMs viz. di-nitro phenol, DNP; Photosan-3, PS3; Methylene blue, MB) in established human cell lines (HEK293, BMG-1 and OCT-1). Glucose utilization and lactate production, levels of glucose transporters and glycolytic enzymes were investigated as indices of glycolysis. Clonogenic survival, DNA repair and cytogenetic damage were studied as parameters of radiation response. MRMs induced the glycolysis by enhancing the levels of two important regulators of glucose metabolism GLUT-1 and HK-II and resulted in 2 fold increase in glucose consumption and lactate production. This increase in glycolysis resulted in resistance against radiation-induced cell death (clonogenic survival) in different cell lines at an absorbed dose of 5 Gy. Inhibition of glucose uptake and glycolysis (using fasentin, 2-deoxy-D-glucose and 3-bromopyruvate) in DNP treated cells failed to increase the clonogenic survival of irradiated cells, suggesting that radio-resistance linked to inhibition of mitochondrial respiration is glycolysis dependent. Elevated glycolysis also facilitated rejoining of radiation-induced DNA strand breaks by activating both non-homologous end joining (NHEJ) and homologous recombination (HR) pathways of DNA double strand break repair leading to a reduction in radiation-induced cytogenetic damage (micronuclei formation) in these cells. These findings suggest that enhanced glycolysis generally observed in cancer cells may be responsible for the radio-resistance, partly by enhancing the repair of DNA damage

  9. In-situ change and repairing method of armour tile made of carbon fiber composite material in divertor

    International Nuclear Information System (INIS)

    Ishiyama, Shintaro.

    1994-01-01

    A joint portion of a damaged armour tile of a carbon fiber composite material and a divertor substrate is locally heated spontaneously to re-melt the soldering. Then, the damaged tile is removed and the portion where the tile is removed is heated again to melt the soldering, then a tile for exchange is joined. Alternatively, a thermosetting type adhesive is coated on the surface of the damaged armour tile made of carbon fiber composite material on the divertor, and a tile for repairing is adhered thereon then the joint surface is locally heated to cure the adhesive. For local heating, for example, high frequency heating or dielectric heating is used. It is preferably conducted by remote handling by using robot arms under vacuum in an vacuum vessel of the thermonuclear device. The operations of the heating and pressurization for the joint surface are preferably repeated for several times. (N.H.)

  10. Evaluation of different end-of-life management alternatives for used natural cork stoppers through life cycle assessment.

    Science.gov (United States)

    Demertzi, Martha; Dias, Ana Cláudia; Matos, Arlindo; Arroja, Luís Manuel

    2015-12-01

    An important aspect of sustainable development is the implementation of effective and sustainable waste management strategies. The present study focuses on a Life Cycle Assessment (LCA) approach to different waste management strategies for natural cork stoppers, namely incineration at a municipal solid waste incinerator, landfilling in a sanitary landfill, and recycling. In the literature, there are no LCA studies analyzing in detail the end-of-life stage of natural cork stoppers as well as other cork products. In addition, cork is usually treated as wood at the end-of-life stage. Thus, the outcome of this study can provide an important insight into this matter. The results showed that different management alternatives, namely incineration and recycling, could be chosen depending on the impact category considered. The former alternative presented the best environmental results in the impact categories of climate change, ozone depletion and acidification, while the latter for photochemical ozone formation and mineral and fossil resource depletion. The landfilling alternative did not present the best environmental performance in any of the impact categories. However, when the biogenic carbon dioxide emission was assessed for the climate change category, the landfilling alternative was found to be the most effective since most of the biogenic carbon would be permanently stored in the cork products and not emitted into the atmosphere. A sensitivity analysis was performed and the results showed that there are various parameters that can significantly influence the results (e.g., carbon content in cork and decay rate of cork in the landfill). Thus, LCA studies should include a detailed description concerning their assumptions when the end-of-life stage is included in the boundaries since they can influence the results, and furthermore, to facilitate the comparison of different end-of-life scenarios. The present study and the obtained results could be useful for the

  11. Cdk1 Restrains NHEJ through Phosphorylation of XRCC4-like Factor Xlf1

    Directory of Open Access Journals (Sweden)

    Pierre Hentges

    2014-12-01

    Full Text Available Eukaryotic cells use two principal mechanisms for repairing DNA double-strand breaks (DSBs: homologous recombination (HR and nonhomologous end-joining (NHEJ. DSB repair pathway choice is strongly regulated during the cell cycle. Cyclin-dependent kinase 1 (Cdk1 activates HR by phosphorylation of key recombination factors. However, a mechanism for regulating the NHEJ pathway has not been established. Here, we report that Xlf1, a fission yeast XLF ortholog, is a key regulator of NHEJ activity in the cell cycle. We show that Cdk1 phosphorylates residues in the C terminus of Xlf1 over the course of the cell cycle. Mutation of these residues leads to the loss of Cdk1 phosphorylation, resulting in elevated levels of NHEJ repair in vivo. Together, these data establish that Xlf1 phosphorylation by Cdc2Cdk1 provides a molecular mechanism for downregulation of NHEJ in fission yeast and indicates that XLF is a key regulator of end-joining processes in eukaryotic organisms.

  12. Conjoint Forming - Technologies for Simultaneous Forming and Joining

    International Nuclear Information System (INIS)

    Groche, P; Wohletz, S; Mann, A; Krech, M; Monnerjahn, V

    2016-01-01

    The market demand for new products optimized for e. g. lightweight applications or smart components leads to new challenges in production engineering. Hybrid structures represent one promising approach. They aim at higher product performance by using a suitable combination of different materials. The developments of hybrid structures stimulate the research on joining of dissimilar materials. Since they allow for joining dissimilar materials without external heating technologies based on joining by plastic deformation seem to be of special attractiveness. The paper at hand discusses the conjoint forming approach. This approach combines forming and joining in one process. Two or more workpieces are joined while at least one workpiece is plastically deformed. After presenting the fundamental joining mechanisms, the conjoint forming approach is discussed comprehensively. Examples of conjoint processes demonstrate the effectiveness and reveal the underlying phenomena. (paper)

  13. Chromosomal Integrity after UV Irradiation Requires FANCD2-Mediated Repair of Double Strand Breaks.

    Directory of Open Access Journals (Sweden)

    María Belén Federico

    2016-01-01

    Full Text Available Fanconi Anemia (FA is a rare autosomal recessive disorder characterized by hypersensitivity to inter-strand crosslinks (ICLs. FANCD2, a central factor of the FA pathway, is essential for the repair of double strand breaks (DSBs generated during fork collapse at ICLs. While lesions different from ICLs can also trigger fork collapse, the contribution of FANCD2 to the resolution of replication-coupled DSBs generated independently from ICLs is unknown. Intriguingly, FANCD2 is readily activated after UV irradiation, a DNA-damaging agent that generates predominantly intra-strand crosslinks but not ICLs. Hence, UV irradiation is an ideal tool to explore the contribution of FANCD2 to the DNA damage response triggered by DNA lesions other than ICL repair. Here we show that, in contrast to ICL-causing agents, UV radiation compromises cell survival independently from FANCD2. In agreement, FANCD2 depletion does not increase the amount of DSBs generated during the replication of UV-damaged DNA and is dispensable for UV-induced checkpoint activation. Remarkably however, FANCD2 protects UV-dependent, replication-coupled DSBs from aberrant processing by non-homologous end joining, preventing the accumulation of micronuclei and chromatid aberrations including non-homologous chromatid exchanges. Hence, while dispensable for cell survival, FANCD2 selectively safeguards chromosomal stability after UV-triggered replication stress.

  14. Mutagenic repair of double-stranded DNA breaks in vaccinia virus genomes requires cellular DNA ligase IV activity in the cytosol.

    Science.gov (United States)

    Luteijn, Rutger David; Drexler, Ingo; Smith, Geoffrey L; Lebbink, Robert Jan; Wiertz, Emmanuel J H J

    2018-04-20

    Poxviruses comprise a group of large dsDNA viruses that include members relevant to human and animal health, such as variola virus, monkeypox virus, cowpox virus and vaccinia virus (VACV). Poxviruses are remarkable for their unique replication cycle, which is restricted to the cytoplasm of infected cells. The independence from the host nucleus requires poxviruses to encode most of the enzymes involved in DNA replication, transcription and processing. Here, we use the CRISPR/Cas9 genome engineering system to induce DNA damage to VACV (strain Western Reserve) genomes. We show that targeting CRISPR/Cas9 to essential viral genes limits virus replication efficiently. Although VACV is a strictly cytoplasmic pathogen, we observed extensive viral genome editing at the target site; this is reminiscent of a non-homologous end-joining DNA repair mechanism. This pathway was not dependent on the viral DNA ligase, but critically involved the cellular DNA ligase IV. Our data show that DNA ligase IV can act outside of the nucleus to allow repair of dsDNA breaks in poxvirus genomes. This pathway might contribute to the introduction of mutations within the genome of poxviruses and may thereby promote the evolution of these viruses.

  15. Knock-in of large reporter genes in human cells via CRISPR/Cas9-induced homology-dependent and independent DNA repair.

    Science.gov (United States)

    He, Xiangjun; Tan, Chunlai; Wang, Feng; Wang, Yaofeng; Zhou, Rui; Cui, Dexuan; You, Wenxing; Zhao, Hui; Ren, Jianwei; Feng, Bo

    2016-05-19

    CRISPR/Cas9-induced site-specific DNA double-strand breaks (DSBs) can be repaired by homology-directed repair (HDR) or non-homologous end joining (NHEJ) pathways. Extensive efforts have been made to knock-in exogenous DNA to a selected genomic locus in human cells; which, however, has focused on HDR-based strategies and was proven inefficient. Here, we report that NHEJ pathway mediates efficient rejoining of genome and plasmids following CRISPR/Cas9-induced DNA DSBs, and promotes high-efficiency DNA integration in various human cell types. With this homology-independent knock-in strategy, integration of a 4.6 kb promoterless ires-eGFP fragment into the GAPDH locus yielded up to 20% GFP+ cells in somatic LO2 cells, and 1.70% GFP+ cells in human embryonic stem cells (ESCs). Quantitative comparison further demonstrated that the NHEJ-based knock-in is more efficient than HDR-mediated gene targeting in all human cell types examined. These data support that CRISPR/Cas9-induced NHEJ provides a valuable new path for efficient genome editing in human ESCs and somatic cells. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  16. Overlapping sphincteroplasty and posterior repair.

    Science.gov (United States)

    Crane, Andrea K; Myers, Erinn M; Lippmann, Quinn K; Matthews, Catherine A

    2014-12-01

    Knowledge of how to anatomically reconstruct extensive posterior-compartment defects is variable among gynecologists. The objective of this video is to demonstrate an effective technique of overlapping sphincteroplasty and posterior repair. In this video, a scripted storyboard was constructed that outlines the key surgical steps of a comprehensive posterior compartment repair: (1) surgical incision that permits access to posterior compartment and perineal body, (2) dissection of the rectovaginal space up to the level of the cervix, (3) plication of the rectovaginal muscularis, (4) repair of internal and external anal sphincters, and (5) reconstruction of the perineal body. Using a combination of graphic illustrations and live video footage, tips on repair are highlighted. The goals at the end of repair are to: (1) have improved vaginal caliber, (2) increase rectal tone along the entire posterior vaginal wall, (3) have the posterior vaginal wall at a perpendicular plane to the perineal body, (4) reform the hymenal ring, and (5) not have an overly elongated perineal body. This video provides a step-by-step guide on how to perform an overlapping sphincteroplasty and posterior repair.

  17. Mountain Plains Learning Experience Guide: Electric Motor Repair.

    Science.gov (United States)

    Ziller, T.

    This Electric Motor Repair Course is designed to provide the student with practical information for winding, repairing, and troubleshooting alternating current and direct current motors, and controllers. The course is comprised of eight units: (1) Electric Motor Fundamentals, (2) Rewinding, (3) Split-phase Induction Motors, (4) Capacitor Motors,…

  18. Suppression of DNA-dependent protein kinase sensitize cells to radiation without affecting DSB repair

    International Nuclear Information System (INIS)

    Gustafsson, Ann-Sofie; Abramenkovs, Andris; Stenerlöw, Bo

    2014-01-01

    Highlights: • We reduced the level of DNA-PKcs with siRNA and examined cells after γ-irradiation. • Low DNA-PKcs levels lead to radiosensitivity but did not affect repair of DSB. • Low DNA-PKcs levels may block progression of mitosis. • DNA-PKcs role in mitotic progression is independent of its role in DSB repair. • We suggest different mechanisms by which loss of DNA-PKcs function sensitize cells. - Abstract: Efficient and correct repair of DNA double-strand break (DSB) is critical for cell survival. Defects in the DNA repair may lead to cell death, genomic instability and development of cancer. The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is an essential component of the non-homologous end joining (NHEJ) which is the major DSB repair pathway in mammalian cells. In the present study, by using siRNA against DNA-PKcs in four human cell lines, we examined how low levels of DNA-PKcs affected cellular response to ionizing radiation. Decrease of DNA-PKcs levels by 80–95%, induced by siRNA treatment, lead to extreme radiosensitivity, similar to that seen in cells completely lacking DNA-PKcs and low levels of DNA-PKcs promoted cell accumulation in G2/M phase after irradiation and blocked progression of mitosis. Surprisingly, low levels of DNA-PKcs did not affect the repair capacity and the removal of 53BP1 or γ-H2AX foci and rejoining of DSB appeared normal. This was in strong contrast to cells completely lacking DNA-PKcs and cells treated with the DNA-PKcs inhibitor NU7441, in which DSB repair were severely compromised. This suggests that there are different mechanisms by which loss of DNA-PKcs functions can sensitize cells to ionizing radiation. Further, foci of phosphorylated DNA-PKcs (T2609 and S2056) co-localized with DSB and this was independent of the amount of DNA-PKcs but foci of DNA-PKcs was only seen in siRNA-treated cells. Our study emphasizes on the critical role of DNA-PKcs for maintaining survival after radiation exposure

  19. Suppression of DNA-dependent protein kinase sensitize cells to radiation without affecting DSB repair

    Energy Technology Data Exchange (ETDEWEB)

    Gustafsson, Ann-Sofie, E-mail: ann-sofie.gustafsson@bms.uu.se; Abramenkovs, Andris; Stenerlöw, Bo

    2014-11-15

    Highlights: • We reduced the level of DNA-PKcs with siRNA and examined cells after γ-irradiation. • Low DNA-PKcs levels lead to radiosensitivity but did not affect repair of DSB. • Low DNA-PKcs levels may block progression of mitosis. • DNA-PKcs role in mitotic progression is independent of its role in DSB repair. • We suggest different mechanisms by which loss of DNA-PKcs function sensitize cells. - Abstract: Efficient and correct repair of DNA double-strand break (DSB) is critical for cell survival. Defects in the DNA repair may lead to cell death, genomic instability and development of cancer. The catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is an essential component of the non-homologous end joining (NHEJ) which is the major DSB repair pathway in mammalian cells. In the present study, by using siRNA against DNA-PKcs in four human cell lines, we examined how low levels of DNA-PKcs affected cellular response to ionizing radiation. Decrease of DNA-PKcs levels by 80–95%, induced by siRNA treatment, lead to extreme radiosensitivity, similar to that seen in cells completely lacking DNA-PKcs and low levels of DNA-PKcs promoted cell accumulation in G2/M phase after irradiation and blocked progression of mitosis. Surprisingly, low levels of DNA-PKcs did not affect the repair capacity and the removal of 53BP1 or γ-H2AX foci and rejoining of DSB appeared normal. This was in strong contrast to cells completely lacking DNA-PKcs and cells treated with the DNA-PKcs inhibitor NU7441, in which DSB repair were severely compromised. This suggests that there are different mechanisms by which loss of DNA-PKcs functions can sensitize cells to ionizing radiation. Further, foci of phosphorylated DNA-PKcs (T2609 and S2056) co-localized with DSB and this was independent of the amount of DNA-PKcs but foci of DNA-PKcs was only seen in siRNA-treated cells. Our study emphasizes on the critical role of DNA-PKcs for maintaining survival after radiation exposure

  20. Individual capacity for DNA repair and maintenance of genomic integrity: a fertile ground for studies in the field of assisted reproduction

    Directory of Open Access Journals (Sweden)

    Radoslava Vazharova

    2016-05-01

    Full Text Available Many factors may affect the chances for successful pregnancy, especially at a later age. Fertility evaluations including genetic analysis are recommended to couples that have not achieved pregnancy within 6–12 months of unprotected intercourse. This review discusses some of the common polymorphisms in genes coding for proteins functioning in DNA damage identification and repair and maintenance of genomic integrity that may affect the chances of success in natural conception as well as in assisted reproduction (AR. Common polymorphisms in genes coding for proteins functioning in DNA damage identification and repair and maintenance of genomic integrity may affect the chances of success in assisted reproduction as well as in natural conception. The effects of carriership of different alleles of key genes of DNA repair may have differential effects in men and women and at different ages, suggesting complex interactions with the mechanisms controlling cell and tissue aging and programmed cell death. Future studies in the field are needed in order to elucidate the genotype–phenotype relationships and to translate the knowledge about individual repair capacity and maintenance of genomic integrity to potential clinical applications. Abbreviations: aCGH: microarray-based comparative genomic hybridization; AR: assisted reproduction; ATM: ataxia-telangiectasia mutated; ATP: adenosine triphosphate; BER: base excision repair; BFE: basic fertility evaluation; DMSO: dimethyl sulfoxide; FSH: follicle-stimulating hormone; GNRHR: gonadotropin-releasing hormone receptor; HMG: high-mobility group; ICSI: intracytoplasmic sperm injection; IUI: intrauterine insemination; IVF: in vitro fertilization; LH: luteinizing hormone; LIF: leukaemia inhibitory factor; MTR: methionine synthase; MTRR: methionine synthase reductase; NGS: next-generation sequencing; NER: nucleotide excision repair; NHEJ: non-homologous end joining; PAH: polycyclic aromatic hydrocarbons; PCOS

  1. Reoperation of Anastomotic Stricture after Oesophageal Atresia Repair: An Uncommon Event

    Directory of Open Access Journals (Sweden)

    A L Azakpa

    2017-01-01

    Full Text Available Oesophageal atresia is a common malformation in which the survival rate in developed countries is around 90%, while its mortality remains very high in developing countries. Oesophageal stricture post-oesophageal atresia repair is traditionally treated by non-surgical approach. However, surgical resection of the oesophageal stricture may be necessary after the failure of dilations. We report one case of refractory oesophageal stricture post-EA repair in a 3-year-old girl, who underwent oesophageal atresia Type III repair at 11-day-old. We performed an end-to-end oesophageal anastomosis with tracheal oesophageal fistula closure by extra-pleural approach. The patient was lost to follow-up for 3 years. She was seen later for anastomotic oesophageal stricture with the failure of oesophageal dilatations. Surgical resection of oesophageal stricture was performed with end-to-end oesophageal anastomosis.

  2. The contribution of alu elements to mutagenic DNA double-strand break repair.

    Science.gov (United States)

    Morales, Maria E; White, Travis B; Streva, Vincent A; DeFreece, Cecily B; Hedges, Dale J; Deininger, Prescott L

    2015-03-01

    Alu elements make up the largest family of human mobile elements, numbering 1.1 million copies and comprising 11% of the human genome. As a consequence of evolution and genetic drift, Alu elements of various sequence divergence exist throughout the human genome. Alu/Alu recombination has been shown to cause approximately 0.5% of new human genetic diseases and contribute to extensive genomic structural variation. To begin understanding the molecular mechanisms leading to these rearrangements in mammalian cells, we constructed Alu/Alu recombination reporter cell lines containing Alu elements ranging in sequence divergence from 0%-30% that allow detection of both Alu/Alu recombination and large non-homologous end joining (NHEJ) deletions that range from 1.0 to 1.9 kb in size. Introduction of as little as 0.7% sequence divergence between Alu elements resulted in a significant reduction in recombination, which indicates even small degrees of sequence divergence reduce the efficiency of homology-directed DNA double-strand break (DSB) repair. Further reduction in recombination was observed in a sequence divergence-dependent manner for diverged Alu/Alu recombination constructs with up to 10% sequence divergence. With greater levels of sequence divergence (15%-30%), we observed a significant increase in DSB repair due to a shift from Alu/Alu recombination to variable-length NHEJ which removes sequence between the two Alu elements. This increase in NHEJ deletions depends on the presence of Alu sequence homeology (similar but not identical sequences). Analysis of recombination products revealed that Alu/Alu recombination junctions occur more frequently in the first 100 bp of the Alu element within our reporter assay, just as they do in genomic Alu/Alu recombination events. This is the first extensive study characterizing the influence of Alu element sequence divergence on DNA repair, which will inform predictions regarding the effect of Alu element sequence divergence on both

  3. Ku-mediated coupling of DNA cleavage and repair during programmed genome rearrangements in the ciliate Paramecium tetraurelia.

    Directory of Open Access Journals (Sweden)

    Antoine Marmignon

    2014-08-01

    Full Text Available During somatic differentiation, physiological DNA double-strand breaks (DSB can drive programmed genome rearrangements (PGR, during which DSB repair pathways are mobilized to safeguard genome integrity. Because of their unique nuclear dimorphism, ciliates are powerful unicellular eukaryotic models to study the mechanisms involved in PGR. At each sexual cycle, the germline nucleus is transmitted to the progeny, but the somatic nucleus, essential for gene expression, is destroyed and a new somatic nucleus differentiates from a copy of the germline nucleus. In Paramecium tetraurelia, the development of the somatic nucleus involves massive PGR, including the precise elimination of at least 45,000 germline sequences (Internal Eliminated Sequences, IES. IES excision proceeds through a cut-and-close mechanism: a domesticated transposase, PiggyMac, is essential for DNA cleavage, and DSB repair at excision sites involves the Ligase IV, a specific component of the non-homologous end-joining (NHEJ pathway. At the genome-wide level, a huge number of programmed DSBs must be repaired during this process to allow the assembly of functional somatic chromosomes. To understand how DNA cleavage and DSB repair are coordinated during PGR, we have focused on Ku, the earliest actor of NHEJ-mediated repair. Two Ku70 and three Ku80 paralogs are encoded in the genome of P. tetraurelia: Ku70a and Ku80c are produced during sexual processes and localize specifically in the developing new somatic nucleus. Using RNA interference, we show that the development-specific Ku70/Ku80c heterodimer is essential for the recovery of a functional somatic nucleus. Strikingly, at the molecular level, PiggyMac-dependent DNA cleavage is abolished at IES boundaries in cells depleted for Ku80c, resulting in IES retention in the somatic genome. PiggyMac and Ku70a/Ku80c co-purify as a complex when overproduced in a heterologous system. We conclude that Ku has been integrated in the Paramecium

  4. Analysis of a damaged and repaired pre-stressed concrete bridge girder by vehicle impact and effectiveness of repair procedure

    OpenAIRE

    Domínguez Mayans, Félix

    2014-01-01

    This thesis aims to study the structural consequences of the damages produced by vehicle impact in a pres-stressed concrete bridge girder and the repair procedure in a real case-study damaged after the bridge was opened to service. From the analysis of the situation of the beam and its damage state, a study of the repair actions carried out on this beam has been analyzed in order to determine the efficiency of the repair and if other alternatives are possible or more efficient. A stat...

  5. Sustainability assessment of alternative end-uses for disused areas based on multi-criteria decision-making method.

    Science.gov (United States)

    De Feo, Giovanni; De Gisi, Sabino; De Vita, Sabato; Notarnicola, Michele

    2018-08-01

    The main aim of this study was to define and apply a multidisciplinary and multi-criteria approach to sustainability in evaluating alternative end-uses for disused areas. Taking into account the three pillars of sustainability (social, economic and environmental dimension) as well as the need for stakeholders to have new practical instruments, the innovative approach consists of four modules stated (i) sociological, (ii) economic, (iii) environmental and (iv) multi-criteria assessment. By means of a case study on a small Municipality in Southern Italy, three end-uses alternatives, representing three essential services for citizens, were selected: Municipal gym; Market area; Municipal Solid Waste (MSW) separate collection centre. The sociological module was useful to select the most socially sound alternative by means of a consultative referendum, simulated with the use of a structured questionnaire administered to a sample of the population. The economic evaluation was conducted defining the bill of quantities with regarding to six main items (soil handling, landfill disposal tax, public services, structure and services, completion work, equipment and furnishings). The environmental evaluation was performed applying the Delphi method with local technicians who were involved in a qualitative-quantitative evaluation of the three alternatives with regarding to eight possible environmental impacts (landscape impact, soil handling, odour, traffic, noise, atmospheric pollution, wastewater, waste). Finally, the Simple Additive Weighting was used as multi-criteria technique to define alternatives priorities. The obtained results showed how the multi-criteria analysis is a useful decision support tool able to identify transparently and efficiently the most sustainable solutions to a complex social problem. Copyright © 2018 Elsevier B.V. All rights reserved.

  6. Distinct kinetics of human DNA ligases I, IIIalpha, IIIbeta, and IV reveal direct DNA sensing ability and differential physiological functions in DNA repair

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xi; Ballin, Jeff D.; Della-Maria, Julie; Tsai, Miaw-Sheue; White, Elizabeth J.; Tomkinson, Alan E.; Wilson, Gerald M.

    2009-05-11

    The three human LIG genes encode polypeptides that catalyze phosphodiester bond formation during DNA replication, recombination and repair. While numerous studies have identified protein partners of the human DNA ligases (hLigs), there has been little characterization of the catalytic properties of these enzymes. In this study, we developed and optimized a fluorescence-based DNA ligation assay to characterize the activities of purified hLigs. Although hLigI joins DNA nicks, it has no detectable activity on linear duplex DNA substrates with short, cohesive single-strand ends. By contrast, hLigIII{beta} and the hLigIII{alpha}/XRCC1 and hLigIV/XRCC4 complexes are active on both nicked and linear duplex DNA substrates. Surprisingly, hLigIV/XRCC4, which is a key component of the major non-homologous end joining (NHEJ) pathway, is significantly less active than hLigIII on a linear duplex DNA substrate. Notably, hLigIV/XRCC4 molecules only catalyze a single ligation event in the absence or presence of ATP. The failure to catalyze subsequent ligation events reflects a defect in the enzyme-adenylation step of the next ligation reaction and suggests that, unless there is an in vivo mechanism to reactivate DNA ligase IV/XRCC4 following phosphodiester bond formation, the cellular NHEJ capacity will be determined by the number of adenylated DNA ligaseIV/XRCC4 molecules.

  7. Joining technology—A challenge for the use of SiC components in HTRs

    Energy Technology Data Exchange (ETDEWEB)

    Herrmann, M., E-mail: marion.herrmann@tu-dresden.de; Meisel, P.; Lippmann, W.; Hurtado, A.

    2016-09-15

    The availability of suitable joining technologies is paramount to the further advancement of ceramic components and their use in HTRs. Among other joining technologies, a modified brazing technology using a laser beam for heating the components to be joined has been developed at TU Dresden. The laser-induced heating behavior of the ceramic material is determined by the interactions between the material and the laser beam. This was shown in two different silicon carbide materials (SSiC and SiC{sub f}-reinforced ceramic material) using a diode laser with wavelengths 808 nm and 940 nm. The laser-based technique was illustrated by three different examples: sealing of monolithic SiC with a pin configuration for fuel claddings, sealing of SiC heat pipes with a length of 1 m, and demonstration of the transferability of the laser technique to fiber-reinforced components by means of a SiC{sub f}/SiCN material. Because the covalent bonding of SiC does not allow conventional welding, much research has been devoted to developing alternative filler systems. Glass or glass–ceramic fillers enable the tailoring of properties such as CTE and viscosity. Glasses are thermally stable up to their glass transition temperatures. It was shown that the crystallization of the yttrium aluminosilicate glass composition of the present work allows it to be used at 1050 °C without any significant changes occurring in braze tightness. For the SiC heat pipes with sodium as the working fluid, a sodium-resistant metal braze consisting of Ni–Ti–Si was formed. The long-term resistance of this filler to sodium at 800 °C was proven. The results demonstrate the possibility of using the laser-based joining technique for the joining of different SiC materials as well as for different brazing materials.

  8. Thin layer joining by gas adsorption

    Energy Technology Data Exchange (ETDEWEB)

    Taga, Yasunori, E-mail: y-taga@isc.chubu.ac.jp; Fukumura, Toshio

    2014-10-01

    Highlights: • We report thin layer molecular joining between glass and COP by gas adsorption. Thickness of joining layer is 1–2 nm and joining process was carried out at low temperature at about 100 °C. • Adhesion strength measured by 180 degree peel test revealed to be 1–10 N/25 mm and the joined stack showed high durability for practical use. - Abstract: Attempt has been made to join borosilicate glass and cycloolefin (COP) polymer film by using gas adsorption method. After corona plasma treat, COP was exposed to (3-glycidoxypropyl) trimethoxysilane (GPS) and glass to (3-aminopropyl) triethoxysilane (APS) both in air atmosphere, resulting in co-adsorption of water vapor in the atmosphere and organosilane gases. Surface characterization of plasma treated and gas adsorbed surfaces was carried out by X-ray photoelectron spectroscopy (XPS) using Mg Kα X-ray source. Joining was carried out by a roll laminator after contact of both surfaces at room temperature, followed by annealing at 130 °C for 10 min. Adhesion strength was evaluated by 180 degree peel test based on ASTM D-903 and durability was examined under the conditions of 60 °C and 95% RH. It was found that after plasma treatment, complex functional groups such as C-H, C-O, C=O, O-C=O and CO{sub 3} were found on COP and O-H on glass. Thickness of GPS gas adsorption layer on COP was evaluated by the XPS to be at least 1.1 nm by taking inelastic mean free path of Si{sub 2p} photoelectron into consideration. Joining force was found to be more than 5 N/25 mm corresponding to almost equal to COP bulk tensile strength. In addition, durability of this adhesion strength remained unchanged over 2000 h even after exposure to the durability test conditions of 60 °C and 95% RH. The results can be explained in terms of formation of H-H hydrogen bonding and Si-O covalent bonding via silanols will be made at the interface as a result of lamination and annealing processes. In conclusion, ultrathin joining method

  9. Characterization of the roles of the catalytic domains of Mycobacterium tuberculosis ligase D in Ku-dependent error-prone DNA end joining.

    Science.gov (United States)

    Wright, Douglas; DeBeaux, Austin; Shi, Runhua; Doherty, Aidan J; Harrison, Lynn

    2010-09-01

    We previously established an Escherichia coli strain capable of re-circularizing linear plasmid DNA by expressing the Mycobacterium tuberculosis Ku (Mt-Ku) and Mycobacterium tuberculosis ligase D (Mt-LigD) proteins from the E.coli chromosome. Repair was predominately mutagenic due to deletions at the termini. We hypothesized that these deletions could be due to a nuclease activity of Mt-LigD that was previously detected in vitro. Mt-LigD has three domains: an N-terminal polymerase domain (PolDom), a central domain with 3'-phosphoesterase and nuclease activity and a C-terminal ligase domain (LigDom). We generated bacterial strains expressing Mt-Ku and mutant versions of Mt-LigD. Plasmid re-circularization experiments in bacteria showed that the PolDom alone had no re-circularization activity. However, an increase in the total and accurate repair was found when the central domain was deleted. This provides further evidence that this central domain does have nuclease activity that can generate deletions during repair. Deletion of only the PolDom of Mt-LigD resulted in a complete loss of accurate repair and a significant reduction in total repair. This is in agreement with published in vitro work indicating that the PolDom is the major Mt-Ku-binding site. Interestingly, the LigDom alone was able to re-circularize plasmid DNA but only in an Mt-Ku-dependent manner, suggesting a potential second site for Ku-LigD interaction. This work has increased our understanding of the mutagenic repair by Mt-Ku and Mt-LigD and has extended the in vitro biochemical experiments by examining the importance of the Mt-LigD domains during repair in bacteria.

  10. Spatio-temporal joins on symbolic indoor tracking data

    DEFF Research Database (Denmark)

    Lu, Hua; Yang, Bin; Jensen, Christian S.

    2011-01-01

    and studies probabilistic, spatio-temporal joins on historical indoor tracking data. Two meaningful types of join are defined. They return object pairs that satisfy spatial join predicates either at a time point or during a time interval. The predicates considered include “same X,” where X is a semantic...

  11. Ceramic restoration repair: report of two cases

    Directory of Open Access Journals (Sweden)

    Luís Henrique Araújo Raposo

    2009-04-01

    Full Text Available The esthetic and functional rehabilitation of patients with multiple missing teeth can be performed with several techniques and materials. Ceramic restorations provide reliable masticatory function and good esthetics. However, fracture can occur in some cases due to their brittle behavior. In some cases, the replacement of an extensive prosthesis is a problem due to the high treatment cost. In this paper, two cases are presented, in which fractures occurred in extensive metal-ceramic fixed partial dentures, and their replacement was not possible. Ceramic repair was chosen and the sequences of treatment with and without presence of the ceramic fragment are also discussed. The cases illustrate that, in some situations, fractured metal-ceramic partial dentures can be successfully repaired when prosthetic replacement is not a choice. Prosthodontists must use alternatives that allow a reliable repair to extensive metal-ceramic fixed partial dentures. Surface preparation of the ceramic with hydrofluoric acid in conjunction with a silane coupling agent is essential for a predictable bonding of composite resin. The repair performed with composite resin is an esthetic and functional alternative when extensive fixed partial dentures cannot be replaced.

  12. Boolean Operations, Joins, and the Extended Low Hierarchy

    OpenAIRE

    Hemaspaandra, Lane A.; Jiang, Zhigen; Rothe, Joerg; Watanabe, Osamu

    1999-01-01

    We prove that the join of two sets may actually fall into a lower level of the extended low hierarchy than either of the sets. In particular, there exist sets that are not in the second level of the extended low hierarchy, EL_2, yet their join is in EL_2. That is, in terms of extended lowness, the join operator can lower complexity. Since in a strong intuitive sense the join does not lower complexity, our result suggests that the extended low hierarchy is unnatural as a complexity measure. We...

  13. DNA-PK inhibition causes a low level of H2AX phosphorylation and homologous recombination repair in Medaka (Oryzias latipes) cells

    International Nuclear Information System (INIS)

    Urushihara, Yusuke; Kobayashi, Junya; Matsumoto, Yoshihisa; Komatsu, Kenshi; Oda, Shoji; Mitani, Hiroshi

    2012-01-01

    Highlights: ► We investigated the effect of DNA-PK inhibition on DSB repair using fish cells. ► A radiation sensitive mutant RIC1 strain showed a low level of DNA-PK activity. ► DNA-PK dysfunction leads defects in HR repair and DNA-PKcs autophosphorylation. ► DNA-PK dysfunction leads a slight increase in the number of 53BP1 foci after DSBs. ► DNA-PK dysfunction leads an alternative NHEJ that depends on 53BP1. -- Abstract: Nonhomologous end joining (NHEJ) and homologous recombination (HR) are known as DNA double-strand break (DSB) repair pathways. It has been reported that DNA-PK, a member of PI3 kinase family, promotes NHEJ and aberrant DNA-PK causes NHEJ deficiency. However, in this study, we demonstrate that a wild-type cell line treated with DNA-PK inhibitor and a mutant cell line with dysfunctional DNA-PK showed decreased HR efficiency in fish cells (Medaka, Oryzias latipes). Previously, we reported that the radiation-sensitive mutant RIC1 strain has a defect in the Histone H2AX phosphorylation after γ-irradiation. Here, we showed that a DNA-PK inhibitor, NU7026, treatment resulted in significant reduction in the number of γH2AX foci after γ-irradiation in wild-type cells, but had no significant effect in RIC1 cells. In addition, RIC1 cells showed significantly lower levels of DNA-PK kinase activity compared with wild-type cells. We investigated NHEJ and HR efficiency after induction of DSBs. Wild-type cells treated with NU7026 and RIC1 cells showed decreased HR efficiency. These results indicated that aberrant DNA-PK causes the reduction in the number of γH2AX foci and HR efficiency in RIC1 cells. We performed phosphorylated DNA-PKcs (Thr2609) and 53BP1 focus assay after γ-irradiation. RIC1 cells showed significant reduction in the number of phosphorylated DNA-PKcs foci and no deference in the number of 53BP1 foci compared with wild-type cells. These results suggest that low level of DNA-PK activity causes aberrant DNA-PKcs autophosphorylation

  14. Augmented Versus Nonaugmented Repair of Acute Achilles Tendon Rupture: A Systematic Review and Meta-analysis.

    Science.gov (United States)

    Zhang, Yi-Jun; Zhang, Chi; Wang, Quan; Lin, Xiang-Jin

    2017-04-01

    Although simple end-to-end repair of the Achilles tendon is common, many augmented repair protocols have been implemented for acute Achilles tendon rupture. However, whether augmented repair is better than nonaugmented repair of an acute Achilles tendon rupture is still unknown. To conduct a meta-analysis to determine whether augmented surgical repair of an acute Achilles tendon rupture improved subjective patient satisfaction without an increase in rerupture rates. Secondary outcomes assessed included infections, ankle range of motion, calf muscle strength, and minor complications. Meta-analysis. A systematic literature search of peer-reviewed articles was conducted to identify all randomized controlled trials (RCTs) comparing augmented repair and nonaugmented repair for acute Achilles tendon rupture from January 1980 to August 2016 in the electronic databases of PubMed, Web of Science (SCI-E/SSCI/A&HCI), and EMBASE. The keywords (Achilles tendon rupture) AND (surg* OR operat* OR repair* OR augment* OR non-augment* OR end-to-end OR sutur*) were combined, and results were limited to human RCTs and controlled clinical trials published in the English language. Four RCTs involving 169 participants were eligible for inclusion; 83 participants were treated with augmented repair and 86 were treated with nonaugmented repair. Augmented repair led to similar responses when compared with nonaugmented repair for acute Achilles tendon rupture (93% vs 90%, respectively; P = .53). The rerupture rates showed no significant difference for augmented versus nonaugmented repair (7.2% vs 9.3%, respectively; P = .69). No differences in superficial and deep infections occurred in augmented (7 infections) and nonaugmented (8 infections) repair groups during postoperative follow-up ( P = .89). The average incisional infection rate was 8.4% with augmented repair and 9.3% with nonaugmented repair. No significant differences in other complications were found between augmented (7.2%) and

  15. Machine vision method for online surface inspection of easy open can ends

    Science.gov (United States)

    Mariño, Perfecto; Pastoriza, Vicente; Santamaría, Miguel

    2006-10-01

    Easy open can end manufacturing process in the food canning sector currently makes use of a manual, non-destructive testing procedure to guarantee can end repair coating quality. This surface inspection is based on a visual inspection made by human inspectors. Due to the high production rate (100 to 500 ends per minute) only a small part of each lot is verified (statistical sampling), then an automatic, online, inspection system, based on machine vision, has been developed to improve this quality control. The inspection system uses a fuzzy model to make the acceptance/rejection decision for each can end from the information obtained by the vision sensor. In this work, the inspection method is presented. This surface inspection system checks the total production, classifies the ends in agreement with an expert human inspector, supplies interpretability to the operators in order to find out the failure causes and reduce mean time to repair during failures, and allows to modify the minimum can end repair coating quality.

  16. Joining elements of silicon carbide

    International Nuclear Information System (INIS)

    Olson, B.A.

    1979-01-01

    A method of joining together at least two silicon carbide elements (e.g.in forming a heat exchanger) is described, comprising subjecting to sufficiently non-oxidizing atmosphere and sufficiently high temperature, material placed in space between the elements. The material consists of silicon carbide particles, carbon and/or a precursor of carbon, and silicon, such that it forms a joint joining together at least two silicon carbide elements. At least one of the elements may contain silicon. (author)

  17. Laboratory testing of closure cap repair techniques

    International Nuclear Information System (INIS)

    Persoff, P.; Moridis, G.; Tuck, D.M.

    1996-10-01

    Landfill design requires a low permeability closure cap as well as a low permeability liner. The Savannah River Site, in South Carolina, has approximately 85 acres of mixed waste landfills covered with compacted kaolin clay. Maintaining low permeability of the clay cap requires both that the permeability of the compacted clay itself remain low and that the integrity of the barrier be maintained. Barrier breaches typically result from penetration by roots or animals, and especially cracks caused by uneven settling or desiccation. In this study, clay layers, 0.81 m in diameter and 7.6 cm thick, were compacted in 7 lysimeters to simulate closure caps. The hydraulic conductivity of each layer was measured, and the compacted clay layers (CCL's) were cracked by drying. Then various repair techniques were applied and the effectiveness of each repair was assessed by remeasuring the hydraulic conductivity. Finally the repaired CCL was again dried and measured to determine how the repair responded to the conditions that caused the original failure. For a full report of this investigation see Persoff et al. Six repair techniques have been tested, four of which involve the use of injectable barrier liquids colloidal silica (CS) and polysiloxane (PSX) described below: (I) covering the crack with a bentonite geosynthetic clay liner (GCL), (ii) recompaction of new kaolinite at STD+3 moisture content joined to existing kaolinite that had dried and shrunk, (iii) direct injection of colloidal silica to a crack, (iv) injection of colloidal silica (CS) to wells in an overlying sand layer, (v) direct injection of polysiloxane to a crack, and (vi), injection of polysiloxane (PSX) to wells in an overlying soil layer

  18. Recrafting the Neighbor-Joining Method

    DEFF Research Database (Denmark)

    Mailund; Brodal, Gerth Stølting; Fagerberg, Rolf

    2006-01-01

    Background: The neighbor-joining method by Saitou and Nei is a widely used method for constructing phylogenetic trees. The formulation of the method gives rise to a canonical Θ(n3) algorithm upon which all existing implementations are based. Methods: In this paper we present techniques for speeding...... up the canonical neighbor-joining method. Our algorithms construct the same phylogenetic trees as the canonical neighbor-joining method. The best-case running time of our algorithms are O(n2) but the worst-case remains O(n3). We empirically evaluate the performance of our algoritms on distance...... matrices obtained from the Pfam collection of alignments. Results: The experiments indicate that the running time of our algorithms evolve as Θ(n2) on the examined instance collection. We also compare the running time with that of the QuickTree tool, a widely used efficient implementation of the canonical...

  19. A novel small molecule inhibitor of the DNA repair protein Ku70/80.

    Science.gov (United States)

    Weterings, Eric; Gallegos, Alfred C; Dominick, Lauren N; Cooke, Laurence S; Bartels, Trace N; Vagner, Josef; Matsunaga, Terry O; Mahadevan, Daruka

    2016-07-01

    Non-Homologous End-Joining (NHEJ) is the predominant pathway for the repair of DNA double strand breaks (DSBs) in human cells. The NHEJ pathway is frequently upregulated in several solid cancers as a compensatory mechanism for a separate DSB repair defect or for innate genomic instability, making this pathway a powerful target for synthetic lethality approaches. In addition, NHEJ reduces the efficacy of cancer treatment modalities which rely on the introduction of DSBs, like radiation therapy or genotoxic chemotherapy. Consequently, inhibition of the NHEJ pathway can modulate a radiation- or chemo-refractory disease presentation. The Ku70/80 heterodimer protein plays a pivotal role in the NHEJ process. It possesses a ring-shaped structure with high affinity for DSBs and serves as the first responder and central scaffold around which the rest of the repair complex is assembled. Because of this central position, the Ku70/80 dimer is a logical target for the disruption of the entire NHEJ pathway. Surprisingly, specific inhibitors of the Ku70/80 heterodimer are currently not available. We here describe an in silico, pocket-based drug discovery methodology utilizing the crystal structure of the Ku70/80 heterodimer. We identified a novel putative small molecule binding pocket and selected several potential inhibitors by computational screening. Subsequent biological screening resulted in the first identification of a compound with confirmed Ku-inhibitory activity in the low micro-molar range, capable of disrupting the binding of Ku70/80 to DNA substrates and impairing Ku-dependent activation of another NHEJ factor, the DNA-PKCS kinase. Importantly, this compound synergistically sensitized human cell lines to radiation treatment, indicating a clear potential to diminish DSB repair. The chemical scaffold we here describe can be utilized as a lead-generating platform for the design and development of a novel class of anti-cancer agents. Copyright © 2016 Elsevier B.V. All

  20. Biological radiolesions and repair

    International Nuclear Information System (INIS)

    Laskowski, W.

    1981-01-01

    In 7 chapters, the book answers the following questions: 1) What reactions are induced in biological matter by absorption of radiation energy. 2) In what parts of the cell do the radiation-induced reactions with detectable biological effects occur. 3) In which way are these cell components changed by different qualities of radiation. 4) What are the cell mechanisms by which radiation-induced changes can be repaired. 5) What is the importance of these repair processes for man, his life and evolution. At the end of each chapter, there is a bibliography of relevant publications in this field. (orig./MG) [de

  1. Advances in appendage joining techniques for PHWR fuel cladding

    International Nuclear Information System (INIS)

    Desai, P.B.; Ray, T.K.; Date, V.G.; Purushotham, D.S.C.

    1995-01-01

    This paper describes work carried out at the BARC on the development of a technique to join tiny appendages (spacers and bearing pads) to thin cladding (before loading of UO 2 pellets) by resistance welding for PHWR fuel assemblies. The work includes qualifying the process for production environment, designing prototype equipment for regular production and quality monitoring. In the first phase of development, welding of appendages on UO 2 loaded elements was successfully developed, and is being used in production. Welding of appendages on to empty clad tubes is a superior technique for several reasons. Many problems associated with development of welding on empty tubes were resolved. work was initiated, in the second phase of the development task, to select a suitable technique to join appendages on empty clad tubes without any collapse of thin clad. Several alternatives were reviewed and assessed such as laser, full face welding, shim welding and shrink fitting ring spacers. Selection of a method using a mandrel and a modified electrode geometry was fully developed. Results were optimized and process development successfully completed. Appropriate weld monitoring techniques were also reviewed for their adaptation. This technique is useful for 19, 22 as well as 37 element assemblies. (author)

  2. Join or be excluded from biomedicine? JOINS and Post-colonial Korea.

    Science.gov (United States)

    Ma, Eunjeong

    2015-04-01

    This paper discusses re-emergence of Korean medicine(s) in the global context with a focus on a natural drug JOINS, a highly contentious drug regarding its legal status. By following through its life world, the paper contends that the drug is the embodiment of the postcolonial anxiety that crosses the intersections between the aspiring nation and globalizing strategies of the bio-pharmaceutical industry. JOINS is a natural drug prescribed to alleviate the symptoms of degenerative arthritis. SK Chemicals, Ltd., a giant domestic pharmaceutical company developed the drug by utilizing the knowledge of traditional pharmacopeia and put it on the market in 2001. In the domestic market, the drug is treated as a prescription drug, implying that Western medicine-trained doctors (as opposed to Korean medicine doctors) are entitled to prescribe drugs. It also indicates that the drug has undergone a series of lab tests such as toxicity, efficacy, and clinical trials in compliance with regulatory guidelines. However, the domestic standards are not rigorous enough to satisfy international standards, so that it is exported as a nutritional supplement abroad. The government, the pharmaceutical industry, and the Western medicine profession are happy with how the drug stands domestically and internationally. Rather, it is Korean doctors who try to disrupt the status quo and reclaim their rights to traditional knowledge, who have been alienated from the pharmaceuticalization of traditional knowledge. Thus, the JOINS tablet embodies the complex web of modern Korean society, professional interests, the pharmaceutical industry, and globalization.

  3. Peaceful power today and tomorrow: Policy alternatives for the back-end

    International Nuclear Information System (INIS)

    Bedore, J.M.

    1985-01-01

    Atoms for Peace is thirty years old. Twenty-six nations currently operate large-scale commercial reactors for uranium electricity generation, and another five countries seem likely to join this group soon. The author reviews international policy options regarding two of the perceived major problems of civil nuclear power in many parts of the world: nuclear waste and potential nuclear weapons proliferation

  4. Mitochondrial respiratory modifiers confer survival advantage by facilitating DNA repair in cancer cells

    International Nuclear Information System (INIS)

    Chauhan, Ankit; Khanna, Suchit; Singh, Saurabh; Rai, Yogesh; Soni, Ravi; Kalra, Namita; Dwarakanath, B.S.; Bhatt, Anant Narayan

    2014-01-01

    High rate of aerobic glycolysis (Warburg effect), one of the primary hallmarks of cancer cells, acquired during the multistep development of tumors is also responsible for therapeutic resistance. Underlying this hallmark is the compromised respiratory metabolism that contributes to the acquisition of the glycolytic phenotype for sustained ATP production and cell proliferation. Nevertheless, the exact mechanisms underlying the glycolysis-linked radio-resistance in cancer cells remain elusive. In this study, we transiently elevated glycolysis by treating human cell lines (HEK293, BMG-1 and OCT-1) with mitochondrial respiratory modifiers (MRMs) viz. 2,4-dinitrophenol, Photosan-3, and Methylene blue to examine if transient stimulation of glycolysis before irradiation using MRMs is sufficient to confer radioresistance. Treatment with MRMs led to a significant (two-fold) increase in glucose consumption and lactate production together with a robust increase in the protein levels of two key regulators of glucose metabolism, i.e. GLUT-1 and HK-II. MRMs also enhanced the clonogenic survival and facilitated DNA repair by activating both non-homologous end joining (NHEJ) and homologous recombination (HR) pathways of DNA double strand break repair leading to reduction in radiation-induced cytogenetic damage (micronuclei formation) in these cells. Inhibition of glucose uptake by inhibitors like 2-deoxy-D-glucose (2-DG), 3-bromo pyruvate (3-BP) and fasentin under conditions of stimulated glycolysis not only reversed the effect but also sensitized the cells to radiation more profoundly. The inhibition of glycolysis using 2-DG also reduced the levels of Ku 70 (NHEJ) and Rad-51 (HR) proteins. Thus, our results suggest that enhanced glycolysis in cancer cells may confer radio-resistance and offers survival advantage partly by enhancing the repair of DNA damage. (author)

  5. Stent fractures in the Hemobahn/Viabahn stent graft after endovascular popliteal aneurysm repair

    NARCIS (Netherlands)

    Tielliu, Ignace F. J.; Zeebregts, Clark J.; Vourliotakis, George; Bekkema, Foppe; van den Dungen, Jan J. A. M.; Prins, Ted R.; Verhoeven, Eric L. G.

    Objective: During the last decade, endovascular repair of popliteal artery aneurysms (PAAs) has become a valid alternative to open repair. This study analyzes the incidence and origin of stein graft fractures after endovascular repair, its impact on patency, and strategies to prevent fractures.

  6. On some steady-state characteristics of systems with gradual repair

    International Nuclear Information System (INIS)

    Finkelstein, Maxim; Ludick, Zani

    2014-01-01

    We consider a repairable system with continuous output that alternates between states of operation and repair. The output of the system in the operating state is represented by a continuous, decreasing function of time. We assume that during the repair state, the system can produce output that is modelled by an increasing stochastic process. The repair action gradually restores the output of the system to its initial level and it returns to the operating state. We obtain and analyse expressions for the generalized availability and related characteristics of systems with gradual repair and consider several meaningful examples

  7. Survey of Active Structural Control and Repair Using Piezoelectric Patches

    Directory of Open Access Journals (Sweden)

    Ahmed Abuzaid

    2015-05-01

    Full Text Available The piezoelectric actuator has gained popularity over the last few years. Attention has been directed towards the study of their electromechanical response in active repair and the control of damaged structures. This has been made possible through the development of various numerical and analytical techniques for such studies. The shift of focus towards the piezoelectric based approaches has been due to their advantages, which include strategic cost benefits in maintenance, as well as an increase in the life cycle of the repaired structures. Furthermore, adhesively bonded joints are widely used in the manufacturing and repairing of structures in many industries, especially automotive and aerospace engineering. This is due to the requirement for lightweight materials as well as the potential adhesive used to join materials with different characteristics. The piezoelectric actuator has also shown the capacity in controlling and lowering the shear stress concentration and joint edge peel in adhesively bonded joint systems. The structure’s control of stress and repair can generally be viewed as a reinforcement that influences the structure’s damage tolerance. Therefore, the interest of this review is on the applications of the piezoelectric actuators in both structural damage and the bonded adhesive joint system. The specific goal is to recognize the contemporary scientific challenges, including future opportunities.

  8. Police must join the fast track to end AIDS by 2030.

    Science.gov (United States)

    Crofts, Nick; Patterson, David

    2016-01-01

    World leaders have committed to ending AIDS by 2030, but stigma and discrimination remain significant obstacles. In particular, police are critical, front-line determinants of risk for many people living with HIV (PLHIV) and members of other key affected populations (KAPs). The negative impact of adverse police behaviours and practices on HIV risk is well documented, and these risks undermine global efforts to end AIDS. Far less well documented, and less common, are attempts to ameliorate this impact by working to change police behaviours. This Special Issue seeks to help redress this imbalance by presenting a selection of original, provocative and important interventions from academics, police officers and other stakeholders concerned with documenting the potential for constructive, progressive and evidence-based approaches to the reduction of HIV risk. We recommend urgent boosting of efforts and funding to engage police in the HIV response. Among other strategies, this needs sustainable funding of programmes and their evaluation, and increased and continuing advocacy and education at all levels to match policy and law reform.

  9. JOINING DISSIMILAR MATERIALS USING FRICTION STIR SCRIBE TECHNIQUE

    Energy Technology Data Exchange (ETDEWEB)

    Upadhyay, Piyush; Hovanski, Yuri; Jana, Saumyadeep; Fifield, Leonard S.

    2016-09-01

    Development of robust and cost effective method of joining dissimilar materials can provide a critical pathway to enable widespread use of multi-material design and components in mainstream industrial applications. The use of multi-material components such as Steel-Aluminum, Aluminum-Polymer allows design engineers to optimize material utilization based on service requirements and often lead weight and cost reductions. However producing an effective joint between materials with vastly different thermal, microstructural and deformation response is highly problematic using conventional joining and /or fastening methods. This is especially challenging in cost sensitive high volume markets that largely rely on low–cost joining solutions. Friction Stir Scribe technology was developed to meet the demands of joining materials with drastically different properties and melting regimes. The process enables joining of light metals like Magnesium and Aluminum to high temperature materials like Steels and Titanium. Additionally viable joints between polymer composites and metal can also be made using this method. This paper will present state of the art, progress made and challenges associated with this innovative derivative of Friction Stir welding in reference to joining dissimilar metals and polymer/metal combinations.

  10. Joining Dissimilar Materials Using Friction Stir Scribe Technique

    Energy Technology Data Exchange (ETDEWEB)

    Upadhyay, Piyush [Pacific Northwest National Laboratory, Richland 99352, WA e-mail: piyush.upadhyay@pnnl.gov; Hovanski, Yuri [Pacific Northwest National Laboratory, Richland 99352, WA; Jana, Saumyadeep [Pacific Northwest National Laboratory, Richland 99352, WA; Fifield, Leonard S. [Pacific Northwest National Laboratory, Richland 99352, WA

    2016-10-03

    Development of a robust and cost-effective method of joining dissimilar materials could provide a critical pathway to enable widespread use of multi-material designs and components in mainstream industrial applications. The use of multi-material components such as steel-aluminum and aluminum-polymer would allow design engineers to optimize material utilization based on service requirements and could often lead to weight and cost reductions. However, producing an effective joint between materials with vastly different thermal, microstructural, and deformation responses is highly problematic using conventional joining and/or fastening methods. This is especially challenging in cost sensitive, high volume markets that largely rely on low cost joining solutions. Friction stir scribe technology was developed to meet the demands of joining materials with drastically different properties and melting regimes. The process enables joining of light metals like magnesium and aluminum to high temperature materials like steel and titanium. Viable joints between polymer composites and metal can also be made using this method. This paper will present the state of the art, progress made, and challenges associated with this innovative derivative of friction stir welding in reference to joining dissimilar metals and polymer/metal combinations.

  11. GFR Decline as an Alternative End Point to Kidney Failure in Clinical Trials : A Meta-analysis of Treatment Effects From 37 Randomized Trials

    NARCIS (Netherlands)

    Inker, Lesley A.; Lambers Heerspink, Hiddo J.; Mondal, Hasi; Schmid, Christopher H.; Tighiouart, Hocine; Noubary, Farzad; Coresh, Josef; Greene, Tom; Levey, Andrew S.

    2014-01-01

    Background: There is increased interest in using alternative end points for trials of kidney disease progression. The currently established end points of end-stage renal disease and doubling of serum creatinine level, equivalent to a 57% decline in estimated glomerular filtration rate (eGFR), are

  12. Joining of porous silicon carbide bodies

    Science.gov (United States)

    Bates, Carl H.; Couhig, John T.; Pelletier, Paul J.

    1990-05-01

    A method of joining two porous bodies of silicon carbide is disclosed. It entails utilizing an aqueous slip of a similar silicon carbide as was used to form the porous bodies, including the sintering aids, and a binder to initially join the porous bodies together. Then the composite structure is subjected to cold isostatic pressing to form a joint having good handling strength. Then the composite structure is subjected to pressureless sintering to form the final strong bond. Optionally, after the sintering the structure is subjected to hot isostatic pressing to further improve the joint and densify the structure. The result is a composite structure in which the joint is almost indistinguishable from the silicon carbide pieces which it joins.

  13. WRN Exonuclease Structure, Molecular Mechanism, and DNA EndProcessing Role

    Energy Technology Data Exchange (ETDEWEB)

    Perry, J. Jefferson P.; Yannone, Steven M.; Holden, Lauren G.; Hitomi, Chiharu; Asaithamby, Aroumougame; Han, Seungil; Cooper, PriscillaK.; Chen, David J.; Tainer, John A.

    2006-02-15

    WRN is unique among the five human RecQ DNA helicases by having a functional exonuclease domain (WRN-exo) and being defective in the premature aging and cancer-related disorder Werner syndrome. Here, we characterize WRN-exo crystal structures, biochemical activity and participation in DNA end-joining. Metal ion complex structures, active site mutations and activity assays reveal a two-metal-ion mediated nuclease mechanism. The DNA end-binding Ku70/80 complex specifically stimulates WRN-exo activity, and structure-based mutational inactivation of WRN-exo alters DNA end-joining in human cells. We furthermore establish structural and biochemical similarities of WRN-exo to DnaQ family replicative proofreading exonucleases, with WRN-specific adaptations consistent with dsDNA specificity and functionally important conformational changes. These results indicate WRN-exo is a human DnaQ family member and support analogous proof-reading activities that are stimulated by Ku70/80 with implications for WRN functions in age related pathologies and maintenance of genomic integrity.

  14. Hands-on repair of component with a lowered pool water level

    International Nuclear Information System (INIS)

    Perfect, J.F.

    1984-01-01

    The repair of a broken positioner mechanism on a TRIGA Reactor neutron collimator, with a lowered pool water level, presented a unique challenge. Radiation dose measurements were made which indicated the repair could be done safely with only 3 feet of water providing shielding over the reactor core. The entire repair project went quite well, due in large part to extensive preplanning. Repair was done safely and radiation exposures were kept well below allowable levels. Savings were significant using this method of repair compared to the alternative of dismantling the facility. (author)

  15. Hands-on repair of component with a lowered pool water level

    International Nuclear Information System (INIS)

    Perfect, J.F.

    1984-01-01

    The repair of a broken positioner mechanism on a TRIGA Reactor neutron collimator, with a lowered pool water level, presented a unique challenge. Radiation dose measurements were made which indicated the repair could be done safetly with only 3 feet of water providing shielding over the reactor core. The entire repair project went quite well, due in large part to extensive preplanning. Repair was done safely and radiation exposures were kept well below allowable levels. Savings were significant using this method of repair compared to the alternative of dismantling the facility

  16. Efficient string similarity join in multi-core and distributed systems.

    Directory of Open Access Journals (Sweden)

    Cairong Yan

    Full Text Available In big data area a significant challenge about string similarity join is to find all similar pairs more efficiently. In this paper, we propose a parallel processing framework for efficient string similarity join. First, the input is split into some disjoint small subsets according to the joint frequency distribution and the interval distribution of strings. Then the filter-verification strategy is adopted in the computation of string similarity for each subset so that the number of candidate pairs is reduced before an effective pruning strategy is used to improve the performance. Finally, the operation of string join is executed in parallel. Para-Join algorithm based on the multi-threading technique is proposed to implement the framework in a multi-core system while Pada-Join algorithm based on Spark platform is proposed to implement the framework in a cluster system. We prove that Para-Join and Pada-Join cannot only avoid reduplicate computation but also ensure the completeness of the result. Experimental results show that Para-Join can achieve high efficiency and significantly outperform than state-of-the-art approaches, meanwhile, Pada-Join can work on large datasets.

  17. High levels of BRC4 induced by a Tet-On 3G system suppress DNA repair and impair cell proliferation in vertebrate cells.

    Science.gov (United States)

    Abe, Takuya; Branzei, Dana

    2014-10-01

    Transient induction or suppression of target genes is useful to study the function of toxic or essential genes in cells. Here we apply a Tet-On 3G system to DT40 lymphoma B cell lines, validating it for three different genes. Using this tool, we then show that overexpression of the chicken BRC4 repeat of the tumor suppressor BRCA2 impairs cell proliferation and induces chromosomal breaks. Mechanistically, high levels of BRC4 suppress double strand break-induced homologous recombination, inhibit the formation of RAD51 recombination repair foci, reduce cellular resistance to DNA damaging agents and induce a G2 damage checkpoint-mediated cell-cycle arrest. The above phenotypes are mediated by BRC4 capability to bind and inhibit RAD51. The toxicity associated with BRC4 overexpression is exacerbated by chemotherapeutic agents and reversed by RAD51 overexpression, but it is neither aggravated nor suppressed by a deficit in the non-homologous end-joining pathway of double strand break repair. We further find that the endogenous BRCA2 mediates the cytotoxicity associated with BRC4 induction, thus underscoring the possibility that BRC4 or other domains of BRCA2 cooperate with ectopic BRC4 in regulating repair activities or mitotic cell division. In all, the results demonstrate the utility of the Tet-On 3G system in DT40 research and underpin a model in which BRC4 role on cell proliferation and chromosome repair arises primarily from its suppressive role on RAD51 functions. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

  18. Induction of DNA deletions after UV-light irradiation in yeast Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    Stepanova, A.N.; Koltovaya, N.A.

    2008-01-01

    We study mutagenic action of such a damaging agent as UV light, which can lead to DNA double-strand breaks (DSB). DNA deletions and gross rearrangements occur in process of DSB repair. We show that UV light induces deletion and rearrangement very efficiently. Analysis of efficacy of different types of repair shows that cell tries to repair DSBs with a combination of both homologous recombination (HR) and nonhomologous end joining (NHEJ) if available and that DSB repair by HR is more effective than by NHEJ in growing culture of haploid yeast

  19. Endovascular repair of para-anastomotic aortoiliac aneurysms.

    LENUS (Irish Health Repository)

    Tsang, Julian S

    2009-11-01

    The purpose of this study is to evaluate the use of endovascular stent grafts in the treatment of para-anastomotic aneurysms (PAAs) as an alternative to high-risk open surgical repair. We identified all patients with previous open aortic aneurysm repair who underwent infrarenal endovascular aneurysm repair (EVAR) at our institution from June 1998 to April 2007. Patient demographics, previous surgery, and operative complications were recorded. One hundred forty-eight patients underwent EVAR during the study period and 11 patients had previous aortic surgery. Of these 11 redo patients, the mean age was 62 years at initial surgery and 71 years at EVAR. All patients were male. Initial open repair was for rupture in five (45%) patients. The average time between initial and subsequent reintervention was 9 years. All patients were ASA Grade III or IV. Fifty-five percent of the PAAs involved the iliac arteries, 36% the abdominal aorta, and 9% were aortoiliac. Ten patients had endovascular stent-grafts inserted electively, and one patient presented with a contained leak. Aorto-uni-iliac stent-grafts were deployed in seven patients, and bifurcated stent-grafts in four patients. A 100% successful deployment rate was achieved. Perioperative mortality was not seen and one patient needed surgical reintervention to correct an endoleak. Endovascular repair of PAAs is safe and feasible. It is a suitable alternative and has probably now become the treatment of choice in the management of PAAs.

  20. A Modeling Approach for Plastic-Metal Laser Direct Joining

    Science.gov (United States)

    Lutey, Adrian H. A.; Fortunato, Alessandro; Ascari, Alessandro; Romoli, Luca

    2017-09-01

    Laser processing has been identified as a feasible approach to direct joining of metal and plastic components without the need for adhesives or mechanical fasteners. The present work sees development of a modeling approach for conduction and transmission laser direct joining of these materials based on multi-layer optical propagation theory and numerical heat flow simulation. The scope of this methodology is to predict process outcomes based on the calculated joint interface and upper surface temperatures. Three representative cases are considered for model verification, including conduction joining of PBT and aluminum alloy, transmission joining of optically transparent PET and stainless steel, and transmission joining of semi-transparent PA 66 and stainless steel. Conduction direct laser joining experiments are performed on black PBT and 6082 anticorodal aluminum alloy, achieving shear loads of over 2000 N with specimens of 2 mm thickness and 25 mm width. Comparison with simulation results shows that consistently high strength is achieved where the peak interface temperature is above the plastic degradation temperature. Comparison of transmission joining simulations and published experimental results confirms these findings and highlights the influence of plastic layer optical absorption on process feasibility.

  1. Sutureless liver repair and hemorrhage control using laser-mediated fusion of human albumin as a solder.

    Science.gov (United States)

    Wadia, Y; Xie, H; Kajitani, M

    2001-07-01

    Major liver trauma has a high mortality because of immediate exsanguination and a delayed morbidity from septicemia, peritonitis, biliary fistulae, and delayed secondary hemorrhage. We evaluated laser soldering using liquid albumin for welding liver injuries. Fourteen lacerations (6 x 2 cm) and 13 nonanatomic resection injuries (raw surface, 8 x 2 cm) were repaired. An 805-nm laser was used to weld 53% liquid albumin-indocyanine green solder to the liver surface, reinforcing it by welding a free autologous omental scaffold. The animals were heparinized and hepatic inflow occlusion was used for vascular control. For both laceration and resection injuries, 16 soldering repairs were evaluated acutely at 3 hours. Eleven animals were evaluated chronically, two at 2 weeks and nine at 4 weeks. All 27 laser mediated-liver repairs had minimal blood loss compared with the suture controls. No dehiscence, hemorrhage, or bile leakage was seen in any of the laser repairs after 3 hours. All 11 chronic repairs healed without complication. This modality effectively seals the liver surface, joins lacerations with minimal thermal injury, and works independently of the patient's coagulation status.

  2. Enrichment of G2/M cell cycle phase in human pluripotent stem cells enhances HDR-mediated gene repair with customizable endonucleases.

    Science.gov (United States)

    Yang, Diane; Scavuzzo, Marissa A; Chmielowiec, Jolanta; Sharp, Robert; Bajic, Aleksandar; Borowiak, Malgorzata

    2016-02-18

    Efficient gene editing is essential to fully utilize human pluripotent stem cells (hPSCs) in regenerative medicine. Custom endonuclease-based gene targeting involves two mechanisms of DNA repair: homology directed repair (HDR) and non-homologous end joining (NHEJ). HDR is the preferred mechanism for common applications such knock-in, knock-out or precise mutagenesis, but remains inefficient in hPSCs. Here, we demonstrate that synchronizing synchronizing hPSCs in G2/M with ABT phase increases on-target gene editing, defined as correct targeting cassette integration, 3 to 6 fold. We observed improved efficiency using ZFNs, TALENs, two CRISPR/Cas9, and CRISPR/Cas9 nickase to target five genes in three hPSC lines: three human embryonic stem cell lines, neural progenitors and diabetic iPSCs. neural progenitors and diabetic iPSCs. Reversible synchronization has no effect on pluripotency or differentiation. The increase in on-target gene editing is locus-independent and specific to the cell cycle phase as G2/M phase enriched cells show a 6-fold increase in targeting efficiency compared to cells in G1 phase. Concurrently inhibiting NHEJ with SCR7 does not increase HDR or improve gene targeting efficiency further, indicating that HR is the major DNA repair mechanism after G2/M phase arrest. The approach outlined here makes gene editing in hPSCs a more viable tool for disease modeling, regenerative medicine and cell-based therapies.

  3. Turbine repair process, repaired coating, and repaired turbine component

    Science.gov (United States)

    Das, Rupak; Delvaux, John McConnell; Garcia-Crespo, Andres Jose

    2015-11-03

    A turbine repair process, a repaired coating, and a repaired turbine component are disclosed. The turbine repair process includes providing a turbine component having a higher-pressure region and a lower-pressure region, introducing particles into the higher-pressure region, and at least partially repairing an opening between the higher-pressure region and the lower-pressure region with at least one of the particles to form a repaired turbine component. The repaired coating includes a silicon material, a ceramic matrix composite material, and a repaired region having the silicon material deposited on and surrounded by the ceramic matrix composite material. The repaired turbine component a ceramic matrix composite layer and a repaired region having silicon material deposited on and surrounded by the ceramic matrix composite material.

  4. QuickJoin—Fast Neighbour-Joining Tree Reconstruction

    DEFF Research Database (Denmark)

    Mailund; Pedersen, Christian N. Storm

    2004-01-01

    We have built a tool for fast construction of very large phylogenetic trees. The tool uses heuristics for speeding up the neighbour-joining algorithm—while still constructing the same tree as the original neighbour-joining algorithm—making it possible to construct trees for ~8000 species in less...

  5. Fermilab Friends for Science Education | Join Us

    Science.gov (United States)

    Fermilab Friends for Science Education FFSE Home About Us Join Us Support Us Contact Us Join Us improving science (science, technology, engineering and mathematics) education. Your donation allows us to membership dues allow us to create new, innovative science education programs, making the best use of unique

  6. Potentials of Industrie 4.0 and Machine Learning for Mechanical Joining

    OpenAIRE

    Jäckel, Mathias

    2017-01-01

    -Sensitivity analysis of the influence of component properties and joining parameters on the joining result for self-pierce riveting -Possibilities to link mechanical joining technologies with the automotive process chain for quality and flexibility improvements -Potential of using machine learning to reduce automotive product development cycles in relation to mechanical joining -Datamining for machine learning at mechanical joining

  7. Repairs in 104 Gy/h?

    International Nuclear Information System (INIS)

    Anon.

    1994-01-01

    In 1989 it was found that in each unit of the MAPS Candu nuclear power station in India the centre portion of the heavy water inlet manifold opposite the 300 mm inlet pipe had torn away. Equipment for remote inspection, repair and removal of debris in an area with constricted and difficult access and radiation fields of about 10 4 Gy/h was developed by Ricardo Hitec of the United Kingdom. This consists of a work performing manipulator, TV viewing systems and a posting tube manipulator for insertion of tools and debris containers into the calandria. A variety of special end effector and tools were also developed jointly with AEA Technology. A first repair campaign was carried out on Unit 1 in 1991. Following a detailed TV survey of the damage a reappraisal of the situation was undertaken and a programme of equipment enhancement carried out. In July 1992 a second repair campaign took place on Unit 2. The difficulties encountered and the degree of success achieved are described. Work proceeded at an intensive level for 14 days when the campaign was ended in view of the exhaustion both of personnel and the equipment. Although more work could have been done a major improvement had been achieved. (UK)

  8. Factors influencing the tensile strength of repaired Achilles tendon: a biomechanical experiment study.

    Science.gov (United States)

    Jielile, Jiasharete; Bai, Jing Ping; Sabirhazi, Gulnur; Redat, Darebai; Yilihamu, Tuoheti; Xinlin, Baoltri; Hu, Geyang; Tang, Bin; Liang, Bing; Sun, Qi

    2010-10-01

    Operative treatment has been advocated as the method of choice to repair Achilles tendon rupture as surgery results in reduced re-rupture rate and faster rehabilitation. Many surgical techniques have been introduced allowing for postoperative early motion of the ankle joint. However, it is currently very difficult for surgeons to determine the optimal treatment conditions for ruptured Achilles tendon with an increasing number of end-to-end suture methods, suture materials, and epitenon suture techniques. In the present biomechanical experiment study based on an orthogonal design, thirty-two New Zealand white rabbits received Achilles tendon tenotomy and subsequent operative treatment to repair the tendon employing four end-to-end suture methods, four suture materials, and four epitenon suture techniques. The tensile strength of the repaired Achilles tendon was investigated at four rehabilitation periods, and in comparison with the results of another sixteen rabbits with normal Achilles tendons. The end-to-end suture method contributed most to the final Achilles tendon tensile strength in addition to rehabilitation period, with the highest values occurring with the use of the parachute-like ("Pa" bone) suture method. The other two factors, namely, suture material and epitenon suture technique, had relatively little influence on the results. The parachute-like ("Pa" bone) surgical technique is superior to the other three end-to-end suture methods, with enhanced tensile strength of the repaired tendon. This method allows for postoperative early kinesitherapy of the ankle and knee joints. Therefore, this technique is highly recommended in clinical situations for treatment of ruptured Achilles tendon. 2010 Elsevier Ltd. All rights reserved.

  9. New Joining Technology for Optimized Metal/Composite Assemblies

    Directory of Open Access Journals (Sweden)

    Holger Seidlitz

    2014-01-01

    Full Text Available The development of a new joining technology, which is used to manufacture high strength hybrid constructions with thermoplastic composites (FRP and metals, is introduced. Similar to natural regulation effects at trees, fibers around the FRP joint become aligned along the lines of force and will not be destroyed by the joining process. This is achieved by the local utilization of the specific plastic flow properties of the FRT and metal component. Compared with usual joining methods—such as flow drill screws, blind and self-piercing rivets—noticeably higher tensile properties can be realized through the novel process management. The load-bearing capability increasing effect could be proved on hybrid joints with hot-dip galvanized steel HX420LAD and orthotropic glass—as well as carbon—fiber reinforced plastics. The results, which were determined in tensile-shear and cross-shear tests according to DIN EN ISO 14273 and DIN EN ISO 14272, are compared with holding loads of established joining techniques with similar joining point diameter and material combinations.

  10. Domain structure of a NHEJ DNA repair ligase from Mycobacterium tuberculosis.

    Science.gov (United States)

    Pitcher, Robert S; Tonkin, Louise M; Green, Andrew J; Doherty, Aidan J

    2005-08-19

    A prokaryotic non-homologous end-joining (NHEJ) system for the repair of DNA double-strand breaks (DSBs), composed of a Ku homodimer (Mt-Ku) and a multidomain multifunctional ATP-dependent DNA ligase (Mt-Lig), has been described recently in Mycobacterium tuberculosis. Mt-Lig exhibits polymerase and nuclease activity in addition to DNA ligation activity. These functions were ascribed to putative polymerase, nuclease and ligase domains that together constitute a monomeric protein. Here, the separate polymerase, nuclease and ligase domains of Mt-Lig were cloned individually, over-expressed and the soluble proteins purified to homogeneity. The polymerase domain demonstrated DNA-dependent RNA primase activity, catalysing the synthesis of unprimed oligoribonucleotides on single-stranded DNA templates. The polymerase domain can also extend DNA in a template-dependent manner. This activity was eliminated when the catalytic aspartate residues were replaced with alanine. The ligase domain catalysed the sealing of nicked double-stranded DNA designed to mimic a DSB, consistent with the role of Mt-Lig in NHEJ. Deletion of the active-site lysine residue prevented the formation of an adenylated ligase complex and consequently thwarted ligation. The nuclease domain did not function independently as a 3'-5' exonuclease. DNA-binding assays revealed that both the polymerase and ligase domains bind DNA in vitro, the latter with considerably higher affinity. Mt-Ku directly stimulated the polymerase and nuclease activities of Mt-Lig. The polymerase domain bound Mt-Ku in vitro, suggesting it may recruit Mt-Lig to Ku-bound DNA in vivo. Consistent with these data, Mt-Ku stimulated the primer extension activity of the polymerase domain, suggestive of a functional interaction relevant to NHEJ-mediated DSB repair processes.

  11. Ionizing radiation sensitivity of DNA polymerase lambda-deficient cells.

    NARCIS (Netherlands)

    Vermeulen, C.; Bertocci, B.; Begg, A.C.; Vens, C.

    2007-01-01

    Ionizing radiation induces a diverse spectrum of DNA lesions, including strand breaks and oxidized bases. In mammalian cells, ionizing radiation-induced lesions are targets of non-homologous end joining, homologous recombination, and base excision repair. In vitro assays show a potential involvement

  12. Microwave joining of SiC ceramics and composites

    Energy Technology Data Exchange (ETDEWEB)

    Ahmad, I.; Silberglitt, R.; Tian, Y.L. [FM Technologies, Inc., Fairfax, VA (United States); Katz, J.D. [Los Alamos National Lab., NM (United States)

    1997-04-01

    Potential applications of SiC include components for advanced turbine engines, tube assemblies for radiant burners and petrochemical processing and heat exchangers for high efficiency electric power generation systems. Reliable methods for joining SiC are required in order to cost-effectively fabricate components for these applications from commercially available shapes and sizes. This manuscript reports the results of microwave joining experiments performed using two different types of SiC materials. The first were on reaction bonded SiC, and produced joints with fracture toughness equal to or greater than that of the base material over an extended range of joining temperatures. The second were on continuous fiber-reinforced SiC/SiC composite materials, which were successfully joined with a commercial active brazing alloy, as well as by using a polymer precursor.

  13. Repair & Strengthening of Distressed/Damaged Ends of Prestressed Beams with FRP Composites

    Science.gov (United States)

    2018-02-01

    Over the past few decades, fiber reinforced polymer (FRP) composites have emerged as a lightweight and efficient material used for the repair and retrofit of concrete infrastructures. FRP can be applied to concrete as either externally bonded laminat...

  14. Enabling Dissimilar Material Joining Using Friction Stir Scribe Technology

    Energy Technology Data Exchange (ETDEWEB)

    Hovanski, Yuri; Upadyay, Piyush; Kleinbaum, Sarah; Carlson, Blair; Boettcher, Eric; Ruokolainen, Robert

    2017-04-05

    One challenge in adapting welding processes to dissimilar material joining is the diversity of melting temperatures of the different materials. Although the use of mechanical fasteners and adhesives have mostly paved the way for near-term implementation of dissimilar material systems, these processes only accentuate the need for low-cost welding processes capable of joining dissimilar material components regardless of alloy, properties, or melting temperature. Friction stir scribe technology was developed to overcome the challenges of joining dissimilar material components where melting temperatures vary greatly, and properties and/or chemistry are not compatible with more traditional welding processes. Although the friction stir scribe process is capable of joining dissimilar metals and metal/polymer systems, a more detailed evaluation of several aluminum/steel joints is presented herein to demonstrate the ability to both chemically and mechanically join dissimilar materials.

  15. Chromosome aberration model combining radiation tracks, chromatin structure, DSB repair and chromatin mobility

    International Nuclear Information System (INIS)

    Friedland, W.; Kundrat, P.

    2015-01-01

    The module that simulates the kinetics and yields of radiation-induced chromosome aberrations within the biophysical code PARTRAC is described. Radiation track structures simulated by Monte Carlo methods are overlapped with multi-scale models of DNA and chromatin to assess the resulting DNA damage. Spatial mobility of individual DNA ends from double-strand breaks is modelled simultaneously with their processing by the non-homologous end-joining enzymes. To score diverse types of chromosome aberrations, the joined ends are classified regarding their original chromosomal location, orientation and the involvement of centromeres. A comparison with experimental data on dicentrics induced by gamma and alpha particles shows that their relative dose dependence is predicted correctly, although the absolute yields are overestimated. The critical model assumptions on chromatin mobility and on the initial damage recognition and chromatin remodelling steps and their future refinements to solve this issue are discussed. (authors)

  16. ROCLA robots repaired after tough times

    CERN Multimedia

    Patrice Loïezi

    2004-01-01

    The team of five welders and five mechanics, represented by Pascal Mésenge (right) and Fabrice Multon (left), worked during two week-ends to repair the two ROCLA. At the centre, Oliver Boettcher, technical manager for the robot.

  17. Enhancement of postreplication repair in Chinese hamster cells

    International Nuclear Information System (INIS)

    D'Ambrosio, S.M.; Setlow, R.B.

    1976-01-01

    Alkaline sedimentation profiles of pulse-labeled DNA from Chinese hamster cells showed that DNA from cells treated with N-acetoxy-acetylaminofluorene or ultraviolet radiation was made in segments smaller than those from untreated cells. Cells treated with a small dose (2.5 μM) of N-acetoxy-acetylaminofluorene or(2.5 J . m -2 ) 254-nm radiation, several hours before a larger dose (7 to 10 μM) of N-acetoxy-acetylaminofluorene or 5.0 J . m -2 of 254-nm radiation, also synthesized small DNA after the second dose. However, the rate at which this small DNA was joined together into parental size was appreciably greater than in absence of the small dose. This enhancement of postreplication repair (as a result of the initial small dose) was not observed when cells were incubated with cycloheximide between the two treatments. The results suggest that N-acetoxy-acetylaminofluorene and ultraviolet-damaged DNA from Chinese hamster cells are repaired by similar postreplicative mechanisms that require de novo protein synthesis for enhancement

  18. Prognostic factors in sensory recovery after digital nerve repair.

    Science.gov (United States)

    Bulut, Tuğrul; Akgün, Ulaş; Çıtlak, Atilla; Aslan, Cihan; Şener, Ufuk; Şener, Muhittin

    2016-01-01

    The prognostic factors that affect sensory nerve recovery after digital nerve repair are variable because of nonhomogeneous data, subjective tests, and different assessment/scoring methods. The aim of this study was to evaluate the success of sensory nerve recovery after digital nerve repair and to investigate the prognostic factors in sensorial healing. Ninety-six digital nerve repairs of 63 patients were retrospectively evaluated. All nerves were repaired with end-to-end neurorraphy. The static two-point discrimination (s2PD) and Semmes Weinstein monofilament (SWM) tests were performed to evaluate sensory recovery. The association between prognostic factors such as gender, age, involved digit, time from injury to repair, length of follow-up, smoking, concomitant injuries, type of injury, and sensory recovery results were assessed. The s2PD test demonstrated excellent results in 26 nerves (27%), good results in 61 nerves (64%), and poor results in 9 nerves (9%). The results of the SWM test according to Imai classification showed that 31 nerves (32%) were normal, light touch was diminished in 38 nerves (40%), protective sensation was diminished in 17 nerves (18%), loss of protective sensation occurred in 5 nerves (5%), and 5 nerves (5%) were anesthetic. There was a negative relationship between age, smoking, concomitant injuries, and sensory recovery. Our results demonstrate that concomitant tendon, bone and vascular injuries, older age, and smoking were associated with worse sensory nerve recovery results. However, all digital nerve injuries should be repaired, regardless of these prognostic factors.

  19. Regulation of DNA repair mechanism in human glioma xenograft cells both in vitro and in vivo in nude mice.

    Science.gov (United States)

    Ponnala, Shivani; Veeravalli, Krishna Kumar; Chetty, Chandramu; Dinh, Dzung H; Rao, Jasti S

    2011-01-01

    Glioblastoma Multiforme (GBM) is the most lethal form of brain tumor. Efficient DNA repair and anti-apoptotic mechanisms are making glioma treatment difficult. Proteases such as MMP9, cathepsin B and urokinase plasminogen activator receptor (uPAR) are over expressed in gliomas and contribute to enhanced cancer cell proliferation. Non-homologous end joining (NHEJ) repair mechanism plays a major role in double strand break (DSB) repair in mammalian cells. Here we show that silencing MMP9 in combination with uPAR/cathepsin B effects NHEJ repair machinery. Expression of DNA PKcs and Ku70/80 at both mRNA and protein levels in MMP9-uPAR (pMU) and MMP9-cathepsin B (pMC) shRNA-treated glioma xenograft cells were reduced. FACS analysis showed an increase in apoptotic peak and proliferation assays revealed a significant reduction in the cell population in pMU- and pMC-treated cells compared to untreated cells. We hypothesized that reduced NHEJ repair led to DSBs accumulation in pMU- and pMC-treated cells, thereby initiating cell death. This hypothesis was confirmed by reduced Ku70/Ku80 protein binding to DSB, increased comet tail length and elevated γH2AX expression in treated cells compared to control. Immunoprecipitation analysis showed that EGFR-mediated lowered DNA PK activity in treated cells compared to controls. Treatment with pMU and pMC shRNA reduced the expression of DNA PKcs and ATM, and elevated γH2AX levels in xenograft implanted nude mice. Glioma cells exposed to hypoxia and irradiation showed DSB accumulation and apoptosis after pMU and pMC treatments compared to respective controls. Our results suggest that pMU and pMC shRNA reduce glioma proliferation by DSB accumulation and increase apoptosis under normoxia, hypoxia and in combination with irradiation. Considering the radio- and chemo-resistant cancers favored by hypoxia, our study provides important therapeutic potential of MMP9, uPAR and cathepsin B shRNA in the treatment of glioma from clinical stand

  20. Regulation of DNA repair mechanism in human glioma xenograft cells both in vitro and in vivo in nude mice.

    Directory of Open Access Journals (Sweden)

    Shivani Ponnala

    Full Text Available Glioblastoma Multiforme (GBM is the most lethal form of brain tumor. Efficient DNA repair and anti-apoptotic mechanisms are making glioma treatment difficult. Proteases such as MMP9, cathepsin B and urokinase plasminogen activator receptor (uPAR are over expressed in gliomas and contribute to enhanced cancer cell proliferation. Non-homologous end joining (NHEJ repair mechanism plays a major role in double strand break (DSB repair in mammalian cells.Here we show that silencing MMP9 in combination with uPAR/cathepsin B effects NHEJ repair machinery. Expression of DNA PKcs and Ku70/80 at both mRNA and protein levels in MMP9-uPAR (pMU and MMP9-cathepsin B (pMC shRNA-treated glioma xenograft cells were reduced. FACS analysis showed an increase in apoptotic peak and proliferation assays revealed a significant reduction in the cell population in pMU- and pMC-treated cells compared to untreated cells. We hypothesized that reduced NHEJ repair led to DSBs accumulation in pMU- and pMC-treated cells, thereby initiating cell death. This hypothesis was confirmed by reduced Ku70/Ku80 protein binding to DSB, increased comet tail length and elevated γH2AX expression in treated cells compared to control. Immunoprecipitation analysis showed that EGFR-mediated lowered DNA PK activity in treated cells compared to controls. Treatment with pMU and pMC shRNA reduced the expression of DNA PKcs and ATM, and elevated γH2AX levels in xenograft implanted nude mice. Glioma cells exposed to hypoxia and irradiation showed DSB accumulation and apoptosis after pMU and pMC treatments compared to respective controls.Our results suggest that pMU and pMC shRNA reduce glioma proliferation by DSB accumulation and increase apoptosis under normoxia, hypoxia and in combination with irradiation. Considering the radio- and chemo-resistant cancers favored by hypoxia, our study provides important therapeutic potential of MMP9, uPAR and cathepsin B shRNA in the treatment of glioma from

  1. Implementing optical fibres for the structural health monitoring of composite patch repaired structures

    DEFF Research Database (Denmark)

    Karatzas, Vasileios; Kotsidis, Elias A.; Tsouvalis, Nicholas G.

    2017-01-01

    Structural health monitoring is increasingly being implemented to improve the level of safety of structures and to reduce inspection and repair costs by allowing for correct planning of these actions, if needed. Composite patch repairing presents an appealing alternative to traditional repair...... methods as it enables the reduction of closedown time and the mitigation of complications associated with traditional repair methods. As reinforcement with the use of composite patches is predominantly performed at defected structures, the urge to monitor the performance of the repair becomes even greater...

  2. On the Complexity of Inner Product Similarity Join

    DEFF Research Database (Denmark)

    Ahle, Thomas Dybdahl; Pagh, Rasmus; Razenshteyn, Ilya

    2016-01-01

    A number of tasks in classification, information retrieval, recommendation systems, and record linkage reduce to the core problem of inner product similarity join (IPS join): identifying pairs of vectors in a collection that have a sufficiently large inner product. IPS join is well understood when...... bounds for (A)LSH-based algorithms. In particular, we show that asymmetry can be avoided by relaxing the LSH definition to only consider the collision probability of distinct elements. A new indexing method for IPS based on linear sketches, implying that our hardness results are not far from being tight...

  3. Comparison of Early Active Tension and Immobilization Protocols in Rehabilitation of the Hand, Following Zone 5-6 Extensor Tendon Repair

    Directory of Open Access Journals (Sweden)

    Fereidoun Layeghi

    2013-07-01

    Full Text Available Objective: The objective of the study was to assess and compare Early Active Tension and Immobilization Protocols in Rehabilitation of the Hand, Following Zone 5-6 Extensor Tendon Repair. Materials & Methods: From the 21 patients enrolled in a randomized controlled trial study, 18 cases with average age of 32. 43 years completed the protocol. A total of 41 extensor tendons from 27 fingers of the subjects were studied. A surgeon repaired all tendons in zone 5 and/or 6 by an end to end epitenon running technique using prolen 0.3. For rehabilitation, subjects were randomly assigned into two “Immobilization” and “early active tension” group. Edema, metacarpophalangeal flexion, combined flexion and extension of wrist and fingers, total active motion of fingers and metacarpal extensor lag were assessed 6 weeks after surgery. All aforementioned variables and grip strength were reassessed 12 weeks after surgery. Results: Compared to traditional immobilization protocol, early active tension could significantly improve hand edema, metacarpal flexion, combined flexion and extension of wrist and fingers, total active motion of fingers and grip strength (P0.05. Conclusion: Our results demonstrate that in compliant patients, early active tension could be an alternative method for traditional immobilization protocol, after surgical repair of extensor tendon injuries in zone 5 and/or 6.

  4. Factors influencing the end of the service life of protective gloves used in car repair shops: a preliminary report.

    Science.gov (United States)

    Emilia, Irzmańska; Agnieszka, Stefko; Katarzyna, Dyńska-Kukulska

    2015-01-01

    The paper presents the results of an end-of-service-life study on 2 kinds of protective gloves designed for workplaces in which workers are exposed to mineral oils and mechanical factors. The authors developed their own end-of-service-life study method that takes into account factors occurring during real-life use of protective gloves. The examined gloves were subjected to mechanical, chemical, and physical factors. The objective of the study was to compare the protective in new gloves subjected to a laboratory simulation test and in gloves used at workplaces in car repair shops. A further goal was to design a glove assessment procedure that would ensure comprehensive analysis of the actual level of performance provided by gloves exposed to selected chemical and mechanical factors as well as subjected to the influence of temperature and humidity, mechanical damage, and chemical degradation of material. The results lead to the conclusion that simultaneous exposure of protective gloves to mechanical, chemical, and physical factors significantly decreases their performance levels. Furthermore, similar performance levels were obtained for gloves subjected to laboratory simulation tests and for those used in the workplace.

  5. Effects of processing parameters on Be/CuCrZr joining

    International Nuclear Information System (INIS)

    Park, Jeong-Yong; Lee, Jung-Suk; Choi, Byung-Kwon; Park, Sang-Yun; Hong, Bong Guen; Jeong, Yong Hwan; Jung, Ki-Jung

    2007-01-01

    A joining of Be/CuCrZr has been considered as the key technology for the fabrication of the ITER first wall. Among the joining methods, Hot isostatic pressing (HIP), which is one of the diffusion bonding methods, is the most feasible method to join the Be and CuCrZr alloy. In the HIP joining of Be and CuCrZr, the interlayer was used to prevent the formation of brittle intermetallic compounds in the interface. Therefore, it is crucial to select a suitable interlayer for a joining of Be and CuCrZr. On the other hand, the diffusion between Be and CuCrZr would be enhanced with an increase of the HIP joining temperature, thereby increasing the joint strength. However, the HIP joining temperature is limited by the mechanical properties of CuCrZr. During the fabrication process of the ITER first wall, CuCrZr is subjected to several thermal cycles including a solution annealing, a cooling and an aging. The HIP joining of Be and CuCrZr corresponds to the aging of CuCrZr. The HIP joining at a higher temperature would cause a degradation of the mechanical properties of CuCrZr by an overaging effect although it is preferable for an improvement of the joint strength. In this study, the effect of the cooling rate on the mechanical properties of aged CuCrZr was investigated to find the maximum HIP temperature without a degradation of the mechanical properties of CuCrZr

  6. Crypt base columnar stem cells in small intestines of mice are radioresistant

    NARCIS (Netherlands)

    Hua, G.; Thin, T.H.; Feldman, R.; Haimovitz-Friedman, A.; Clevers, H.; Fuks, Z.; Kolesnick, R.

    2012-01-01

    BACKGROUND & AIMS: Adult stem cells have been proposed to be quiescent and radiation resistant, repairing DNA double-strand breaks by nonhomologous end joining. However, the population of putative small intestinal stem cells (ISCs) at position +4 from the crypt base contradicts this model, in that

  7. Arabidopsis DNA polymerase lambda mutant is mildly sensitive to DNA double strand breaks but defective in integration of a transgene

    Czech Academy of Sciences Publication Activity Database

    Furukawa, T.; Angelis, Karel; Britt, A.B.

    2015-01-01

    Roč. 6, MAY 27 (2015) ISSN 1664-462X R&D Projects: GA ČR GA13-06595S Institutional support: RVO:61389030 Keywords : DNA polymerase * DNA repair * Non homologous end joining Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 4.495, year: 2015

  8. Repair replication in permeabilized Escherichia coli

    International Nuclear Information System (INIS)

    Masker, W.E.; Simon, T.J.; Hanawalt, P.C.

    1975-01-01

    We have examined the modes of DNA synthesis in Escherichia coli strains made permeable to nucleoside triphosphates by treatment with toluene. In this quasi in vitro system, polymerase-I-deficient mutants exhibit a nonconservative mode of synthesis with properties expected for the resynthesis step of excision-repair. This uv-stimulated DNA synthesis can be performed by either DNA polymerase II or III and it also requires the uvrA gene product. It requires the four deoxynucleoside triphosphates; but, in contrast to the semiconservative mode, the ATP requirement can be partially satisfied by other nucleoside triphosphates. The ATP-dependent recBC nuclease is not involved. The observed uv-stimulated mode of DNA synthesis may be part of an alternate excision-repair mechanism which supplements or complements DNA-polymerase-I-dependent repair in vivo

  9. Towards mastering CRISPR-induced gene knock-in in plants: Survey of key features and focus on the model Physcomitrella patens.

    Science.gov (United States)

    Collonnier, Cécile; Guyon-Debast, Anouchka; Maclot, François; Mara, Kostlend; Charlot, Florence; Nogué, Fabien

    2017-05-15

    Beyond its predominant role in human and animal therapy, the CRISPR-Cas9 system has also become an essential tool for plant research and plant breeding. Agronomic applications rely on the mastery of gene inactivation and gene modification. However, if the knock-out of genes by non-homologous end-joining (NHEJ)-mediated repair of the targeted double-strand breaks (DSBs) induced by the CRISPR-Cas9 system is rather well mastered, the knock-in of genes by homology-driven repair or end-joining remains difficult to perform efficiently in higher plants. In this review, we describe the different approaches that can be tested to improve the efficiency of CRISPR-induced gene modification in plants, which include the use of optimal transformation and regeneration protocols, the design of appropriate guide RNAs and donor templates and the choice of nucleases and means of delivery. We also present what can be done to orient DNA repair pathways in the target cells, and we show how the moss Physcomitrella patens can be used as a model plant to better understand what DNA repair mechanisms are involved, and how this knowledge could eventually be used to define more performant strategies of CRISPR-induced gene knock-in. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. Behaviour of UV-sensitive mutants of Proteus mirabilis to repair incision breaks

    International Nuclear Information System (INIS)

    Stoerl, K.; Mund, C.

    1977-01-01

    In U.V.-sensitive mutants of P. mirabilis with the phenotype HCR, REC and EXR single-strand breaks appeared immediately after UV-irradiation. The behaviour of REC- and EXR-mutants was similar to the wildtype. The number of incision breaks observed by sedimentation analysis in these strains was very low. They could be joined during the excision repair process. From the ability of REC- and EXR-strains to rejoin most of the induced single-strand breaks it can be concluded that these strains have approximately the same capacity for excision repair as the wildtype. HCR-mutants of P. mirabilis produced single-strand breaks after UV-irradiation in contrast to HCR-mutants of E. coli. Therefore we suggest that HCR-mutants of P. mirabilis are not completely inhibited in the incision step. The single-strand breaks introduced in the DNA at the beginning of the repair process were not rejoined during further incubation. Experiments with toluenized cells led to the same results. The newly synthesized daughter DNA-strands of UV-irradiated HCR-mutants were of low molecular weight in comparison with those from unirradiated control cells during the repair period. This result is in agreement with the incapability of HCR-mutants to remove the pyrimidine dimers from the parental template strand. (author)

  11. A biomechanical evaluation of all-inside 2-stitch meniscal repair devices with matched inside-out suture repair.

    Science.gov (United States)

    Ramappa, Arun J; Chen, Alvin; Hertz, Benjamin; Wexler, Michael; Grimaldi Bournissaint, Leandro; DeAngelis, Joseph P; Nazarian, Ara

    2014-01-01

    Many all-inside suture-based devices are currently available, including the Meniscal Cinch, FasT-Fix, Ultra FasT-Fix, RapidLoc, MaxFire, and CrossFix System. These different devices have been compared in various configurations, but to our knowledge, the Sequent meniscal repair device, which applies running sutures, has not been compared with the Ultra FasT-Fix, nor has it been compared with its suture, No. 0 Hi-Fi, using an inside-out repair technique. To assess the quality of the meniscal repair, all new devices should be compared with the gold standard: the inside-out repair. To that end, this study aims to compare the biomechanical characteristics of running sutures delivered by the Sequent meniscal repair device with 2 vertical mattress sutures applied using the Ultra FasT-Fix device and with 2 vertical mattress sutures using an inside-out repair technique with No. 0 Hi-Fi suture. Controlled laboratory study. Paired (medial and lateral), fresh-frozen porcine menisci were randomly assigned to 1 of 3 groups: Sequent (n = 17), Ultra FasT-Fix (n = 19), and No. 0 Hi-Fi inside-out repair (n = 20). Bucket-handle tears were created in all menisci and were subjected to repair according to their grouping. Once repaired, the specimens were subjected to cyclic loading (100, 300, and 500 cycles), followed by loading to failure. The Sequent and Ultra FasT-Fix device repairs and the suture repair exhibited low initial displacements. The Sequent meniscal repair device demonstrated the lowest displacement in response to cyclic loading. No. 0 Hi-Fi suture yielded the highest load to failure. With the development of the next generation of all-inside meniscal repair devices, surgeons may use these findings to select the method best suited for their patients. The Sequent meniscal repair device displays the least amount of displacement during cyclic loading but has a similar failure load to other devices.

  12. Inspection and repair of steam generator tubing with a robot

    International Nuclear Information System (INIS)

    Boehm, H.H.; Foerch, H.

    1985-01-01

    During inspection and repair of steam generator tubing, radiation exposure to personnel is an unrequested endowment. To combat this intrinsic handicap, a robot has been designed for deployment in all operations inside the steam generator water chamber. This measure drastically reduces entering time and also improves inspection capabilities with regard to the accuracy and reproduction of the desired tube address. The inherent flexibility of the robot allows for performing various inspection and repair techniques: eddy-current testing of tubing; ultrasonic testing of tubing; visual examination of tube ends; profilometry measurements; tube plugging; plug removal; tube extraction; sleeving of tubes; tube end repair; chemical cleaning; and thermal treatment. Plant experience has highlighted the following features of the robot: 1) short installation and demounting periods; 2) installation independent of manhole location; 3) installation possible from outside the steam generator; 4) only one relocation required to address all the tube positions; 5) fast and highly accurate positioning; 6) operational surveillance not required; and 7) drastic reduction of radiation exposure to personnel during repair work

  13. Repair process and a repaired component

    Energy Technology Data Exchange (ETDEWEB)

    Roberts, III, Herbert Chidsey; Simpson, Stanley F.

    2018-02-20

    Matrix composite component repair processes are disclosed. The matrix composite repair process includes applying a repair material to a matrix composite component, securing the repair material to the matrix composite component with an external securing mechanism and curing the repair material to bond the repair material to the matrix composite component during the securing by the external securing mechanism. The matrix composite component is selected from the group consisting of a ceramic matrix composite, a polymer matrix composite, and a metal matrix composite. In another embodiment, the repair process includes applying a partially-cured repair material to a matrix composite component, and curing the repair material to bond the repair material to the matrix composite component, an external securing mechanism securing the repair material throughout a curing period, In another embodiment, the external securing mechanism is consumed or decomposed during the repair process.

  14. Joining of Gamma Titanium Aluminides

    National Research Council Canada - National Science Library

    Baeslack, William

    2002-01-01

    .... Although organized and presented by joining process, many of the observations made and relationships developed, particularly those regarding the weldability and welding metallurgy of gamma titanium...

  15. [Early prenatal diagnosis of diprosopic syncephalic joined twins].

    Science.gov (United States)

    Picaud, A; Nlome-Nze, A R; Engongha-Beka, T; Ogowet-Igumu, N

    1990-06-01

    The authors summarize the case of diprosopic syncephalic joined twins diagnosed at 22 weeks of pregnancy by ultrasonography performed because of hydramnios. The rate of separation anomalies of monozygotic twins is assessed by a review of the literature: from 1 to twenty to fifty thousands for joined twins to 1 per cent fifty thousand to fifteen millions for diprosopus. The etiology is the result of a late division of the egg between D12 and D16. Often an encephalic diprosopic joined twins cause elevated levels of maternal serum alpha protein. Early ultrasonography permits to consider a vaginal therapeutic abortion.

  16. A link between double-strand break-related repair and V(D)J recombination: the scid mutation

    International Nuclear Information System (INIS)

    Hendrickson, E.A.; Qin, X.Q.; Bump, E.A.; Schatz, D.G.; Oettinger, M.; Weaver, D.T.

    1991-01-01

    We show here that mammalian site-specific recombination and DNA-repair pathways share a common factor. The effects of DNA-damaging agents on cell lines derived from mice homozygous for the scid (severe combined immune deficiency) mutation were studied. Surprisingly, all scid cell lines exhibited a profound hypersensitivity to DNA-damaging agents that caused double-strand breaks (x-irradiation and bleomycin) but not to other chemicals that caused single-strand breaks or cross-links. Neutral filter elution assays demonstrated that the x-irradiation hypersensitivity could be correlated with a deficiency in repairing double-strand breaks. These data suggest that the scid gene product is involved in two pathways: DNA repair of random double-strand breaks and the site-specific and lymphoid-restricted variable-(diversity)-joining [V(D)J] DNA rearrangement process. We propose that the scid gene product performs a similar function in both pathways and may be a ubiquitous protein

  17. Transcription-associated processes cause DNA double-strand breaks and translocations in neural stem/progenitor cells.

    Science.gov (United States)

    Schwer, Bjoern; Wei, Pei-Chi; Chang, Amelia N; Kao, Jennifer; Du, Zhou; Meyers, Robin M; Alt, Frederick W

    2016-02-23

    High-throughput, genome-wide translocation sequencing (HTGTS) studies of activated B cells have revealed that DNA double-strand breaks (DSBs) capable of translocating to defined bait DSBs are enriched around the transcription start sites (TSSs) of active genes. We used the HTGTS approach to investigate whether a similar phenomenon occurs in primary neural stem/progenitor cells (NSPCs). We report that breakpoint junctions indeed are enriched around TSSs that were determined to be active by global run-on sequencing analyses of NSPCs. Comparative analyses of transcription profiles in NSPCs and B cells revealed that the great majority of TSS-proximal junctions occurred in genes commonly expressed in both cell types, possibly because this common set has higher transcription levels on average than genes transcribed in only one or the other cell type. In the latter context, among all actively transcribed genes containing translocation junctions in NSPCs, those with junctions located within 2 kb of the TSS show a significantly higher transcription rate on average than genes with junctions in the gene body located at distances greater than 2 kb from the TSS. Finally, analysis of repair junction signatures of TSS-associated translocations in wild-type versus classical nonhomologous end-joining (C-NHEJ)-deficient NSPCs reveals that both C-NHEJ and alternative end-joining pathways can generate translocations by joining TSS-proximal DSBs to DSBs on other chromosomes. Our studies show that the generation of transcription-associated DSBs is conserved across divergent cell types.

  18. INDUCTION HEATING IN HISTORY AND DEVELOPMENT. APPLICATION IN MODERN TRANSPORT REPAIRING TECHNOLOGIES

    Directory of Open Access Journals (Sweden)

    Yu. Batyhin

    2017-06-01

    Full Text Available The technologies used in repair of vehicles were analyzed in the given paper. The shortcomings of the mechanical repair methods in question can be solved by using induction heating. Analysis of the stages of development and implementation of induction heating in industries showed effective performance of this technology and its opportunities for further improvement. An alternative repair technique, which consists in using induction heating, was proposed.

  19. Patient selection, echocardiographic screening and treatment strategies for interventional tricuspid repair using the edge-to-edge repair technique.

    Science.gov (United States)

    Hausleiter, Jörg; Braun, Daniel; Orban, Mathias; Latib, Azeem; Lurz, Philipp; Boekstegers, Peter; von Bardeleben, Ralph Stephan; Kowalski, Marek; Hahn, Rebecca T; Maisano, Francesco; Hagl, Christian; Massberg, Steffen; Nabauer, Michael

    2018-04-24

    Severe tricuspid regurgitation (TR) has long been neglected despite its well known association with mortality. While surgical mortality rates remain high in isolated tricuspid valve surgery, interventional TR repair is rapidly evolving as an alternative to cardiac surgery in selected patients at high surgical risk. Currently, interventional edge-to-edge repair is the most frequently applied technique for TR repair even though the device has not been developed for this particular indication. Due to the inherent differences in tricuspid and mitral valve anatomy and pathology, percutaneous repair of the tricuspid valve is challenging due to a variety of factors including the complexity and variability of tricuspid valve anatomy, echocardiographic visibility of the valve leaflets, and device steering to the tricuspid valve. Furthermore, it remains to be clarified which patients are suitable for a percutaneous tricuspid repair and which features predict a successful procedure. On the basis of the available experience, we describe criteria for patient selection including morphological valve features, a standardized process for echocardiographic screening, and a strategy for clip placement. These criteria will help to achieve standardization of valve assessment and the procedural approach, and to further develop interventional tricuspid valve repair using either currently available devices or dedicated tricuspid edge-to-edge repair devices in the future. In summary, this manuscript will provide guidance for patient selection and echocardiographic screening when considering edge-to-edge repair for severe TR.

  20. Overview of existing cartilage repair technology.

    Science.gov (United States)

    McNickle, Allison G; Provencher, Matthew T; Cole, Brian J

    2008-12-01

    Currently, autologous chondrocyte implantation and osteochondral grafting bridge the gap between palliation of cartilage injury and resurfacing via arthroplasty. Emerging technologies seek to advance first generation techniques and accomplish several goals including predictable outcomes, cost-effective technology, single-stage procedures, and creation of durable repair tissue. The biologic pipeline represents a variety of technologies including synthetics, scaffolds, cell therapy, and cell-infused matrices. Synthetic constructs, an alternative to biologic repair, resurface a focal chondral defect rather than the entire joint surface. Scaffolds are cell-free constructs designed as a biologic "net" to augment marrow stimulation techniques. Minced cartilage technology uses stabilized autologous or allogeneic fragments in 1-stage transplantation. Second and third generation cell-based methods include alternative membranes, chondrocyte seeding, and culturing onto scaffolds. Despite the promising early results of these products, significant technical obstacles remain along with unknown long-term durability. The vast array of developing technologies has exceptional promise and the potential to revolutionize the cartilage treatment algorithm within the next decade.

  1. Engineering a Nickase on the Homing Endonuclease I-DmoI Scaffold

    DEFF Research Database (Denmark)

    Molina, Rafael; Marcaida, María José; Redondo, Pilar

    2015-01-01

    strand break could be an approach to reduce the toxicity associated with non-homologous end joining by promoting the use of homologous recombination to repair the cleavage of a single DNA break. Taking advantage of the sequential DNA cleavage mechanism of I-DmoI LAGLIDADG homing endonuclease, we have......Homing endonucleases are useful tools for genome modification because of their capability to recognize and cleave specifically large DNA targets. These endonucleases generate a DNA double strand break that can be repaired by the DNA damage response machinery. The break can be repaired by homologous...

  2. Joining NbTi superconductors by ultrasonic welding

    International Nuclear Information System (INIS)

    Hafstrom, J.W.; Killpatrick, D.H.; Niemann, R.C.; Purcell, J.R.; Thresh, H.R.

    1976-01-01

    An important consideration in the design and construction of large, high-field, superconducting magnets is the capability to fabricate reliable, high-strength, low-resistance joints. A process for joining NbTi, copper stabilized, superconducting composites by ultrasonic welding is described. This process yields a joint strength comparable to that of the superconducting composite and a resistivity significantly lower than achieved by conventional soft soldering. The superconducting properties of the composite are not affected by the joining process. Scarfing the joint to maintain a constant conductor cross section does not degrade its electrical or mechanical properties. The application of the ultrasonic joining process, including process control, scarfing, and NDT procedures, in the construction of the superconducting magnet (U.S. SCMS) for the joint U. S. -- Soviet MHD program is described

  3. A successful double-layer facial nerve repair: A case presentation

    Directory of Open Access Journals (Sweden)

    Mehmet Dadaci

    2015-04-01

    Full Text Available The best method to repair the facial nerve is to perform the primary repair soon after the injury, without any tension in the nerve ends. We present a case of patient who had a full-thickness facial nerve cut at two different levels. The patient underwent primary repair, recovered almost completely in the fourth postoperative month, and had full movement in mimic muscles. Despite lower success rates in double-level cuts, performing appropriate primary repair at an appropriate time can reverse functional losses at early stages, and lead to recovery without any complications. [Hand Microsurg 2015; 4(1.000: 24-27

  4. Localization Microscopy Analyses of MRE11 Clusters in 3D-Conserved Cell Nuclei of Different Cell Lines

    Directory of Open Access Journals (Sweden)

    Marion Eryilmaz

    2018-01-01

    Full Text Available In radiation biophysics, it is a subject of nowadays research to investigate DNA strand break repair in detail after damage induction by ionizing radiation. It is a subject of debate as to what makes up the cell’s decision to use a certain repair pathway and how the repair machinery recruited in repair foci is spatially and temporarily organized. Single-molecule localization microscopy (SMLM allows super-resolution analysis by precise localization of single fluorescent molecule tags, resulting in nuclear structure analysis with a spatial resolution in the 10 nm regime. Here, we used SMLM to study MRE11 foci. MRE11 is one of three proteins involved in the MRN-complex (MRE11-RAD50-NBS1 complex, a prominent DNA strand resection and broken end bridging component involved in homologous recombination repair (HRR and alternative non-homologous end joining (a-NHEJ. We analyzed the spatial arrangements of antibody-labelled MRE11 proteins in the nuclei of a breast cancer and a skin fibroblast cell line along a time-course of repair (up to 48 h after irradiation with a dose of 2 Gy. Different kinetics for cluster formation and relaxation were determined. Changes in the internal nano-scaled structure of the clusters were quantified and compared between the two cell types. The results indicate a cell type-dependent DNA damage response concerning MRE11 recruitment and cluster formation. The MRE11 data were compared to H2AX phosphorylation detected by γH2AX molecule distribution. These data suggested modulations of MRE11 signal frequencies that were not directly correlated to DNA damage induction. The application of SMLM in radiation biophysics offers new possibilities to investigate spatial foci organization after DNA damaging and during subsequent repair.

  5. Localization Microscopy Analyses of MRE11 Clusters in 3D-Conserved Cell Nuclei of Different Cell Lines.

    Science.gov (United States)

    Eryilmaz, Marion; Schmitt, Eberhard; Krufczik, Matthias; Theda, Franziska; Lee, Jin-Ho; Cremer, Christoph; Bestvater, Felix; Schaufler, Wladimir; Hausmann, Michael; Hildenbrand, Georg

    2018-01-22

    In radiation biophysics, it is a subject of nowadays research to investigate DNA strand break repair in detail after damage induction by ionizing radiation. It is a subject of debate as to what makes up the cell's decision to use a certain repair pathway and how the repair machinery recruited in repair foci is spatially and temporarily organized. Single-molecule localization microscopy (SMLM) allows super-resolution analysis by precise localization of single fluorescent molecule tags, resulting in nuclear structure analysis with a spatial resolution in the 10 nm regime. Here, we used SMLM to study MRE11 foci. MRE11 is one of three proteins involved in the MRN-complex (MRE11-RAD50-NBS1 complex), a prominent DNA strand resection and broken end bridging component involved in homologous recombination repair (HRR) and alternative non-homologous end joining (a-NHEJ). We analyzed the spatial arrangements of antibody-labelled MRE11 proteins in the nuclei of a breast cancer and a skin fibroblast cell line along a time-course of repair (up to 48 h) after irradiation with a dose of 2 Gy. Different kinetics for cluster formation and relaxation were determined. Changes in the internal nano-scaled structure of the clusters were quantified and compared between the two cell types. The results indicate a cell type-dependent DNA damage response concerning MRE11 recruitment and cluster formation. The MRE11 data were compared to H2AX phosphorylation detected by γH2AX molecule distribution. These data suggested modulations of MRE11 signal frequencies that were not directly correlated to DNA damage induction. The application of SMLM in radiation biophysics offers new possibilities to investigate spatial foci organization after DNA damaging and during subsequent repair.

  6. Incontinence after primary repair of obstetric anal sphincter tears is related to relative length of reconstructed external sphincter

    DEFF Research Database (Denmark)

    Norderval, S; Røssaak, K.; Markskog, A

    2012-01-01

    To determine if anatomic primary repair with end-to-end reconstruction of the external anal sphincter (EAS) in its full length combined with separate repair of coexisting internal anal sphincter (IAS) tear, when present, results in less incontinence and better anal sphincter integrity compared...

  7. DNA end resection by CtIP and exonuclease 1 prevents genomic instability

    DEFF Research Database (Denmark)

    Eid, Wassim; Steger, Martin; El-Shemerly, Mahmoud

    2010-01-01

    End resection of DNA-which is essential for the repair of DNA double-strand breaks (DSBs) by homologous recombination-relies first on the partnership between MRE11-RAD50-NBS1 (MRN) and CtIP, followed by a processive step involving helicases and exonucleases such as exonuclease 1 (EXO1). In this s......End resection of DNA-which is essential for the repair of DNA double-strand breaks (DSBs) by homologous recombination-relies first on the partnership between MRE11-RAD50-NBS1 (MRN) and CtIP, followed by a processive step involving helicases and exonucleases such as exonuclease 1 (EXO1...... of DNA-PK-dependent radial chromosome formation. Thus, our study identifies new functions of CtIP and EXO1 in DNA end resection and provides new information on the regulation of DSB repair pathways, which is a key factor in the maintenance of genome integrity....

  8. Mouse RAD54 affects DNA double-strand break repair and sister chromatid exchange

    NARCIS (Netherlands)

    H.B. Beverloo (Berna); R.D. Johnson (Roger); M. Jasin (Maria); R. Kanaar (Roland); J.H.J. Hoeijmakers (Jan); M.L.G. Dronkert (Mies)

    2000-01-01

    textabstractCells can achieve error-free repair of DNA double-strand breaks (DSBs) by homologous recombination through gene conversion with or without crossover. In contrast, an alternative homology-dependent DSB repair pathway, single-strand annealing (SSA), results in deletions. In this study, we

  9. Targeting DNA double strand break repair with hyperthermia and DNA-PKcs inhibition to enhance the effect of radiation treatment.

    Science.gov (United States)

    van Oorschot, Bregje; Granata, Giovanna; Di Franco, Simone; Ten Cate, Rosemarie; Rodermond, Hans M; Todaro, Matilde; Medema, Jan Paul; Franken, Nicolaas A P

    2016-10-04

    Radiotherapy is based on the induction of lethal DNA damage, primarily DNA double-strand breaks (DSB). Efficient DSB repair via Non-Homologous End Joining or Homologous Recombination can therefore undermine the efficacy of radiotherapy. By suppressing DNA-DSB repair with hyperthermia (HT) and DNA-PKcs inhibitor NU7441 (DNA-PKcsi), we aim to enhance the effect of radiation.The sensitizing effect of HT for 1 hour at 42°C and DNA-PKcsi [1 μM] to radiation treatment was investigated in cervical and breast cancer cells, primary breast cancer sphere cells (BCSCs) enriched for cancer stem cells, and in an in vivo human tumor model. A significant radio-enhancement effect was observed for all cell types when DNA-PKcsi and HT were applied separately, and when both were combined, HT and DNA-PKcsi enhanced radio-sensitivity to an even greater extent. Strikingly, combined treatment resulted in significantly lower survival rates, 2 to 2.5 fold increase in apoptosis, more residual DNA-DSB 6 h post treatment and a G2-phase arrest. In addition, tumor growth analysis in vivo showed significant reduction in tumor growth and elevated caspase-3 activity when radiation was combined with HT and DNA-PKcsi compared to radiation alone. Importantly, no toxic side effects of HT or DNA-PKcsi were found.In conclusion, inhibiting DNA-DSB repair using HT and DNA-PKcsi before radiotherapy leads to enhanced cytotoxicity in cancer cells. This effect was even noticed in the more radio-resistant BCSCs, which are clearly sensitized by combined treatment. Therefore, the addition of HT and DNA-PKcsi to conventional radiotherapy is promising and might contribute to more efficient tumor control and patient outcome.

  10. Development of a Dynamic, End-to-End Free Piston Stirling Convertor Model

    Science.gov (United States)

    Regan, Timothy F.; Gerber, Scott S.; Roth, Mary Ellen

    2003-01-01

    A dynamic model for a free-piston Stirling convertor is being developed at the NASA Glenn Research Center. The model is an end-to-end system model that includes the cycle thermodynamics, the dynamics, and electrical aspects of the system. The subsystems of interest are the heat source, the springs, the moving masses, the linear alternator, the controller and the end-user load. The envisioned use of the model will be in evaluating how changes in a subsystem could affect the operation of the convertor. The model under development will speed the evaluation of improvements to a subsystem and aid in determining areas in which most significant improvements may be found. One of the first uses of the end-to-end model will be in the development of controller architectures. Another related area is in evaluating changes to details in the linear alternator.

  11. Autologous hematopoietic stem cell transplantation in lymphoma patients is associated with a decrease in the double strand break repair capacity of peripheral blood lymphocytes.

    Science.gov (United States)

    Lacoste, Sandrine; Bhatia, Smita; Chen, Yanjun; Bhatia, Ravi; O'Connor, Timothy R

    2017-01-01

    Patients who undergo autologous hematopoietic stem cell transplantation (aHCT) for treatment of a relapsed or refractory lymphoma are at risk of developing therapy related- myelodysplasia/acute myeloid leukemia (t-MDS/AML). Part of the risk likely resides in inherent interindividual differences in their DNA repair capacity (DRC), which is thought to influence the effect chemotherapeutic treatments have on the patient's stem cells prior to aHCT. Measuring DRC involves identifying small differences in repair proficiency among individuals. Initially, we investigated the cell model in healthy individuals (primary lymphocytes and/or lymphoblastoid cell lines) that would be appropriate to measure genetically determined DRC using host-cell reactivation assays. We present evidence that interindividual differences in DRC double-strand break repair (by non-homologous end-joining [NHEJ] or single-strand annealing [SSA]) are better preserved in non-induced primary lymphocytes. In contrast, lymphocytes induced to proliferate are required to assay base excision (BER) or nucleotide excision repair (NER). We established that both NHEJ and SSA DRCs in lymphocytes of healthy individuals were inversely correlated with the age of the donor, indicating that DSB repair in lymphocytes is likely not a constant feature but rather something that decreases with age (~0.37% NHEJ DRC/year). To investigate the predictive value of pre-aHCT DRC on outcome in patients, we then applied the optimized assays to the analysis of primary lymphocytes from lymphoma patients and found that individuals who later developed t-MDS/AML (cases) were indistinguishable in their DRC from controls who never developed t-MDS/AML. However, when DRC was investigated shortly after aHCT in the same individuals (21.6 months later on average), aHCT patients (both cases and controls) showed a significant decrease in DSB repair measurements. The average decrease of 6.9% in NHEJ DRC observed among aHCT patients was much higher

  12. Autologous hematopoietic stem cell transplantation in lymphoma patients is associated with a decrease in the double strand break repair capacity of peripheral blood lymphocytes.

    Directory of Open Access Journals (Sweden)

    Sandrine Lacoste

    Full Text Available Patients who undergo autologous hematopoietic stem cell transplantation (aHCT for treatment of a relapsed or refractory lymphoma are at risk of developing therapy related- myelodysplasia/acute myeloid leukemia (t-MDS/AML. Part of the risk likely resides in inherent interindividual differences in their DNA repair capacity (DRC, which is thought to influence the effect chemotherapeutic treatments have on the patient's stem cells prior to aHCT. Measuring DRC involves identifying small differences in repair proficiency among individuals. Initially, we investigated the cell model in healthy individuals (primary lymphocytes and/or lymphoblastoid cell lines that would be appropriate to measure genetically determined DRC using host-cell reactivation assays. We present evidence that interindividual differences in DRC double-strand break repair (by non-homologous end-joining [NHEJ] or single-strand annealing [SSA] are better preserved in non-induced primary lymphocytes. In contrast, lymphocytes induced to proliferate are required to assay base excision (BER or nucleotide excision repair (NER. We established that both NHEJ and SSA DRCs in lymphocytes of healthy individuals were inversely correlated with the age of the donor, indicating that DSB repair in lymphocytes is likely not a constant feature but rather something that decreases with age (~0.37% NHEJ DRC/year. To investigate the predictive value of pre-aHCT DRC on outcome in patients, we then applied the optimized assays to the analysis of primary lymphocytes from lymphoma patients and found that individuals who later developed t-MDS/AML (cases were indistinguishable in their DRC from controls who never developed t-MDS/AML. However, when DRC was investigated shortly after aHCT in the same individuals (21.6 months later on average, aHCT patients (both cases and controls showed a significant decrease in DSB repair measurements. The average decrease of 6.9% in NHEJ DRC observed among aHCT patients was

  13. Use of orthogonal field alternational gel electrophoresis (OFAGE) for studying DNA double strand breakage and repair

    International Nuclear Information System (INIS)

    Contopoulou, C.R.; Cook, V.; Mortimer, R.K.

    1987-01-01

    The study of DNA double strand breakage and repair has normally been carried by using neutral sucrose gradient or neutral elution techniques. The authors have applied OFAGE procedures to study x-ray induced double strand breaks and repair. Breakage of chromosomes is seen by a decrease in intensity of individual chromosome bands; as expected, this decrease becomes more pronounced as chromosome size increases. The fragments of broken chromosomes appears as a broad smear in the size range 100 kb to 1000 kb. Following repair, these fragments partially disappear and the chromosomal bands increase in intensity. In four repair deficient mutants, rad51, rad52, rad54, rad55, no increase in chromosomal band intensity was seen. These results have been confirmed by blotting for a specific chromosome

  14. Differential gene expression in a DNA double-strand-break repair mutant XRS-5 defective in Ku80. Analysis by cDNA microarray

    International Nuclear Information System (INIS)

    Chan, John Y.H.; Chen, Lung-Kun; Chang, Jui-Feng

    2001-01-01

    The ability of cells to rejoin DNA double-strand breaks (DSBs) usually correlates with their radiosensitivity. This correlation has been demonstrated in radiosensitive cells, including the Chinese hamster ovary mutant XRS-5. XRS-5 is defective in a DNA end-binding protein, Ku80, which is a component of a DNA-dependent protein kinase complex used for joining strand breaks. However, Ku80-deficient cells are known to be retarded in cell proliferation and growth as well as other yet to be identified defects. Using custom-made 600-gene cDNA microarray filters, we found differential gene expressions between the wild-type and XRS-5 cells. Defective Ku80 apparently affects the expression of several repair genes, including topoisomerase-I and -IIA, ERCC5, MLH1, and ATM. In contrast, other DNA repair-associated genes, such as GADD45A, EGR1 MDM2 and p53, were not affected. In addition, for large numbers of growth-associated genes, such as cyclins and clks, the growth factors and cytokines were also affected. Down-regulated expression was also found in several categories of seemingly unrelated genes, including apoptosis, angiogenesis, kinase and signaling, phosphatase, stress protein, proto-oncogenes and tumor suppressors, transcription and translation factors. A RT-PCR analysis confirmed that the XRS-5 cells used were defective in Ku80 expression. The diversified groups of genes being affected could mean that Ku80, a multi-functional DNA-binding protein, not only affects DNA repair, but is also involved in transcription regulation. Our data, taken together, indicate that there are specific genes being modulated in Ku80- deficient cells, and that some of the DNA repair pathways and other biological functions are apparently linked, suggesting that a defect in one gene could have global effects on many other processes. (author)

  15. Differential gene expression in a DNA double-strand-break repair mutant XRS-5 defective in Ku80. Analysis by cDNA microarray

    Energy Technology Data Exchange (ETDEWEB)

    Chan, John Y.H.; Chen, Lung-Kun; Chang, Jui-Feng [National Yang Ming Univ., Taipei, Taiwan (China). Inst. of Radiological Sciences] (and others)

    2001-12-01

    The ability of cells to rejoin DNA double-strand breaks (DSBs) usually correlates with their radiosensitivity. This correlation has been demonstrated in radiosensitive cells, including the Chinese hamster ovary mutant XRS-5. XRS-5 is defective in a DNA end-binding protein, Ku80, which is a component of a DNA-dependent protein kinase complex used for joining strand breaks. However, Ku80-deficient cells are known to be retarded in cell proliferation and growth as well as other yet to be identified defects. Using custom-made 600-gene cDNA microarray filters, we found differential gene expressions between the wild-type and XRS-5 cells. Defective Ku80 apparently affects the expression of several repair genes, including topoisomerase-I and -IIA, ERCC5, MLH1, and ATM. In contrast, other DNA repair-associated genes, such as GADD45A, EGR1 MDM2 and p53, were not affected. In addition, for large numbers of growth-associated genes, such as cyclins and clks, the growth factors and cytokines were also affected. Down-regulated expression was also found in several categories of seemingly unrelated genes, including apoptosis, angiogenesis, kinase and signaling, phosphatase, stress protein, proto-oncogenes and tumor suppressors, transcription and translation factors. A RT-PCR analysis confirmed that the XRS-5 cells used were defective in Ku80 expression. The diversified groups of genes being affected could mean that Ku80, a multi-functional DNA-binding protein, not only affects DNA repair, but is also involved in transcription regulation. Our data, taken together, indicate that there are specific genes being modulated in Ku80- deficient cells, and that some of the DNA repair pathways and other biological functions are apparently linked, suggesting that a defect in one gene could have global effects on many other processes. (author)

  16. Repair of abasic sites in DNA

    Energy Technology Data Exchange (ETDEWEB)

    Dianov, Grigory L.; Sleeth, Kate M.; Dianova, Irina I.; Allinson, Sarah L

    2003-10-29

    Repair of both normal and reduced AP sites is activated by AP endonuclease, which recognizes and cleaves a phosphodiester bond 5' to the AP site. For a short period of time an incised AP site is occupied by poly(ADP-ribose) polymerase and then DNA polymerase {beta} adds one nucleotide into the repair gap and simultaneously removes the 5'-sugar phosphate. Finally, the DNA ligase III/XRCC1 complex accomplishes repair by sealing disrupted DNA ends. However, long-patch BER pathway, which is involved in the removal of reduced abasic sites, requires further DNA synthesis resulting in strand displacement and the generation of a damage-containing flap that is later removed by the flap endonuclease. Strand-displacement DNA synthesis is accomplished by DNA polymerase {delta}/{epsilon} and DNA ligase I restores DNA integrity. DNA synthesis by DNA polymerase {delta}/{epsilon} is dependent on proliferating cell nuclear antigen, which also stimulates the DNA ligase I and flap endonuclease. These repair events are supported by multiple protein-protein interactions.

  17. One and a half ventricular repair as an alternative for hypoplastic right ventricle.

    Science.gov (United States)

    Maluf, Miguel Angel; Carvalho, Antonio Carlos; Carvalho, Werther Brunow

    2010-01-01

    Patients with complex congenital heart disease, characterized by right ventricle hypoplasia, had a palliative surgical option with one and a half ventricular repair. From July 2001 to March 2009, nine patients (mean age 5.2 years, range 3 to 9 years) with hypoplastic right ventricle, underwent correction with one and a half ventricle technique. Preoperative diagnoses included: pulmonary atresia with intact ventricular septum, in six and Ebstein's anomaly, in three cases. Six patients had bidirectional cavo-pulmonary shunt (Glenn operation) previously. The surgical approach was performed with cardiopulmonary bypass to correct intracardiac defects: atrial septal defect closure (nine cases); right ventricle outlet tract reconstruction with porcine pulmonary prosthesis (seven cases); tricuspid valvuloplasty (three cases). There was one (11.1%) hospital death. All the patients left the hospital in good clinical conditions. One patient presented pulmonary stenosis at distal prosthesis anastomosis and needed surgical correction. There was one (12.5%) late deaths after reoperation. At mean follow-up of 39.8 months (range 16 months to 8.4 years) seven patients are alive in functional class I (NYHA). Surgical treatment of congenital cardiac anomalies in the presence of a hypoplastic right ventricle by means of one and a half ventricle repair has the advantages of reducing the surgical risk of biventricular repair compared to the Fontan circulation; it maintains a low right atrium pressure, a pulsatile pulmonary blood flow and improves the systemic oxygen saturation with short and medium-term promising results. Longer follow-up is needed to prove the efficacy of such a repair in the long term.

  18. The Ku Heterodimer and the Metabolism of Single-Ended DNA Double-Strand Breaks

    Directory of Open Access Journals (Sweden)

    Alessia Balestrini

    2013-06-01

    Full Text Available Single-ended double-strand breaks (DSBs are a common form of spontaneous DNA break, generated when the replisome encounters a discontinuity in the DNA template. Given their prevalence, understanding the mechanisms governing the fate(s of single-ended DSBs is important. We describe the influence of the Ku heterodimer and Mre11 nuclease activity on processing of single-ended DSBs. Separation-of-function alleles of yku70 were derived that phenocopy Ku deficiency with respect to single-ended DSBs but remain proficient for NHEJ. The Ku mutants fail to regulate Exo1 activity, and bypass the requirement for Mre11 nuclease activity in the repair of camptothecin-induced single-ended DSBs. Ku mutants exhibited reduced affinity for DNA ends, manifest as both reduced end engagement and enhanced probability of diffusing inward on linear DNA. This study reveals an interplay between Ku and Mre11 in the metabolism of single-ended DSBs that is distinct from repair pathway choice at double-ended DSBs.

  19. Significant accumulation of persistent organic pollutants and dysregulation in multiple DNA damage repair pathways in the electronic-waste-exposed populations

    Energy Technology Data Exchange (ETDEWEB)

    He, Xiaobo; Jing, Yaqing; Wang, Jianhai; Li, Keqiu [Basic Medical College, Tianjin Medical University, Tianjin 300070 (China); Yang, Qiaoyun [Department of Occupational and Environmental Health, School of Public Health, Tianjin Medical University, Tianjin 300070 (China); Zhao, Yuxia [Basic Medical College, Tianjin Medical University, Tianjin 300070 (China); Li, Ran [State Key Joint Laboratory for Environmental Simulation and Pollution Control, College of Environmental Sciences and Engineering and Center for Environment and Health, Peking University, Beijing 100871 (China); Ge, Jie [Department of Breast Surgery, Tianjin Medical University Cancer Institute and Hospital, Tianjin 300060 (China); Key Laboratory of Breast Cancer Prevention and Treatment of the Ministry of Education, Tianjin Medical University Cancer Institute and Hospital, Tianjin 300060 (China); Qiu, Xinghua, E-mail: xhqiu@pku.edu.cn [State Key Joint Laboratory for Environmental Simulation and Pollution Control, College of Environmental Sciences and Engineering and Center for Environment and Health, Peking University, Beijing 100871 (China); Li, Guang, E-mail: lig@tijmu.edu.cn [Basic Medical College, Tianjin Medical University, Tianjin 300070 (China)

    2015-02-15

    Electronic waste (e-waste) has created a worldwide environmental and health problem, by generating a diverse group of hazardous compounds such as persistent organic pollutants (POPs). Our previous studies demonstrated that populations from e-waste exposed region have a significantly higher level of chromosomal aberrancy and incidence of DNA damage. In this study, we further demonstrated that various POPs persisted at a significantly higher concentration in the exposed group than those in the unexposed group. The level of reactive oxygen species and micronucleus rate were also significantly elevated in the exposed group. RNA sequencing analysis revealed 31 genes in DNA damage responses and repair pathways that were differentially expressed between the two groups (Log 2 ratio >1 or <−1). Our data demonstrated that both females and males of the exposed group have activated a series of DNA damage response genes; however many important DNA repair pathways have been dysregulated. Expressions of NEIL1/3 and RPA3, which are critical in initiating base pair and nucleotide excision repairs respectively, have been downregulated in both females and males of the exposed group. In contrast, expression of RNF8, an E3 ligase involved in an error prone non-homologous end joining repair for DNA double strand break, was upregulated in both genders of the exposed group. The other genes appeared to be differentially expressed only when the males or females of the two groups were compared respectively. Importantly, the expression of cell cycle regulatory gene CDC25A that has been implicated in multiple kinds of malignant transformation was significantly upregulated among the exposed males while downregulated among the exposed females. In conclusion, our studies have demonstrated significant correlations between e-waste disposing and POPs accumulation, DNA lesions and dysregulation of multiple DNA damage repair mechanisms in the residents of the e-waste exposed region. - Highlights:

  20. DNA repair in ultraviolet-irradiated spores of Bacillus subtilis

    International Nuclear Information System (INIS)

    Wang, T.C.V.

    1976-01-01

    It has been shown previously by others that at least two independent repair mechanisms are present in Bacillus subtilis for removing ''spore photoproduct'' from DNA of ultraviolet (254 nm)-irradiated spores after germination. One of these, designated as ''spore repair,'' is shown in this study to restore ''spore photoproduct'' to two thymine residues, leaving the DNA backbone intact at the end of the process in vivo. The circumstances under which this repair can occur and some characteristics of its energy requirements have been clarified. The second repair process is identified as excision repair, which can excise both ''spore photoproduct'' from DNA of irradiated spores and cyclobutane-type pyrimidine dimers from DNA of irradiated vegetative cells. In this study it is shown that the gene hcr 1 affects an enzyme activity for the incision step initiating this repair, while the gene hcr 42 affects a step subsequent to incision in the mechanism. In addition a third, independent repair system, termed ''germinative excision repair,'' is discovered and shown to be specific for excising only cyclobutane-type pyrimidine dimers but not ''spore photoproduct.'' This repair system is responsible for the observed high ultraviolet-resistance and temporary capacity for host cell reactivation on recently germinated spores of Bacillus subtilis HCR - strains

  1. Impact of radiotherapy on PBMCs DNA repair capacity - Use of a multiplexed functional repair assay

    International Nuclear Information System (INIS)

    Sauvaigo, S.; Sarrazy, F.; Breton, J.; Caillat, S.; Chapuis, V.

    2012-01-01

    Radiation therapy is an essential part of cancer treatment as about 50% of patients will receive radiations at least once. Significant broad variation in radiosensitivity has been demonstrated in patients. About 5-10% of patients develop acute toxicity after radiotherapy. Therefore there is a need for the identification of markers able to predict the occurrence of adverse effects and thus adapt the radiotherapy regimen for radiosensitive patients. As a first step toward this goal, and considering the DNA repair defects associated with hypersensitivity radiation syndromes, we investigated the DNA repair phenotype of patients receiving radiotherapy. More precisely, we used a functional repair assay on support to follow the evolution of the glycosylases/AP endonuclease activities of PBMCs extracts of a series of patients during the time course of radiotherapy. For each patient, we collected one PBMCs sample before the first radiotherapy application (S1) and three samples after (S2 to S4) (one day and one week after application 1, and one at the end of the radiotherapy protocol). These four samples have been analysed for 11 donors. Clustering analyses of the results demonstrated a great heterogeneity of responses among the patients. Interestingly, this heterogeneity decreased between S1 and S4 where only 2 classes of patients remained if we except one patient that exhibited an atypical DNA repair phenotype. Furthermore, we showed that repair of several oxidized bases significantly increased between S1 and S3 or S4 (8oxoG, thymine glycol, A paired with 8oxoG), suggesting an adaptation of patients repair systems to the oxidative stress generated by the ionising radiations. Our preliminary results provided evidence that the DNA repair phenotype was impacted by the radiotherapy regimen. Further characterization of patients with known repair defects are needed to determine if atypical repair phenotypes could be associated with radiotherapy complications. Finally

  2. Ageing airplane repair assessment program for Airbus A300

    Science.gov (United States)

    Gaillardon, J. M.; Schmidt, HANS-J.; Brandecker, B.

    1992-01-01

    This paper describes the current status of the repair categorization activities and includes all details about the methodologies developed for determination of the inspection program for the skin on pressurized fuselages. For inspection threshold determination two methods are defined based on fatigue life approach, a simplified and detailed method. The detailed method considers 15 different parameters to assess the influences of material, geometry, size location, aircraft usage, and workmanship on the fatigue life of the repair and the original structure. For definition of the inspection intervals a general method is developed which applies to all concerned repairs. For this the initial flaw concept is used by considering 6 parameters and the detectable flaw sizes depending on proposed nondestructive inspection methods. An alternative method is provided for small repairs allowing visual inspection with shorter intervals.

  3. Development of Joining Technologies for the ITER Blanket First Wall

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Byoung Kwon; Jung, Yang Il; Park, Dong Jun; Kim, Hyun Gil; Park, Sang Yoon; Park, Jeong Yong; Jeong, Yong Hwan; Lee, Dong Won; Kim, Suk Kwon [KAERI, Daejeon (Korea, Republic of)

    2011-01-15

    The design of the ITER blanket first wall includes the Beryllium amour tiles joined to CuCrZr heat sink with stainless steel cooling tubes. For the ITER application, the Be/CuCrZr/SS joint was proposed as a first wall material. The joining of Be/CuCrZr as well as CuCrZr/SS was generally carried out by using a hot isostatic pressing (CuC) in many countries. The joining strength for Be/CuCrZr is relatively lower than that for CuCrZr/SS, since we usually forms surface oxides (BeO) and brittle a metallics with Cu. Therefore, the joining technology for the Be/CuCrZr joint has been investigated. Be is apt to adsorb oxygen in an air atmosphere, so we should be etched to eliminate the surface pre-oxide using a chemical solution and Ar ions in a vacuum chamber. Then we is coated with a first was to prevent further oxidation. The kinds of a first we are chosen to be able to enhance the joining strength as inhibiting excessive be diffusion. The performance of the Be/CuCrZr/SS joint used for the ITER first wall is primarily dependent on the joining strength of the Be/CuCrZr interface. The Cr/Cu and Ti/Cr/Cu interlayers enabled the successful joining of be tile to CuCrZr plate. Moreover, ion-beam assisted deposition (IBAD) increased joining strength of the Be/CuCrZr joint mock-ups. IBAD induced the increased packing of depositing atoms, which resulted in denser and more adhesive interlayers. The interlayers formed by IBAD process revealed about 40% improved resistance to the scratch test. It is suggested that the improved adhesion of coating interlayers enabled tight joining of Be and CuCrZr blocks. As compared to without IBAD coating, the shear strength as well as the 4-point bend strength were increased more than 20% depending on interlayer types and coating conditions

  4. Development of Joining Technologies for the ITER Blanket First Wall

    International Nuclear Information System (INIS)

    Choi, Byoung Kwon; Jung, Yang Il; Park, Dong Jun; Kim, Hyun Gil; Park, Sang Yoon; Park, Jeong Yong; Jeong, Yong Hwan; Lee, Dong Won; Kim, Suk Kwon

    2011-01-01

    The design of the ITER blanket first wall includes the Beryllium amour tiles joined to CuCrZr heat sink with stainless steel cooling tubes. For the ITER application, the Be/CuCrZr/SS joint was proposed as a first wall material. The joining of Be/CuCrZr as well as CuCrZr/SS was generally carried out by using a hot isostatic pressing (CuC) in many countries. The joining strength for Be/CuCrZr is relatively lower than that for CuCrZr/SS, since we usually forms surface oxides (BeO) and brittle a metallics with Cu. Therefore, the joining technology for the Be/CuCrZr joint has been investigated. Be is apt to adsorb oxygen in an air atmosphere, so we should be etched to eliminate the surface pre-oxide using a chemical solution and Ar ions in a vacuum chamber. Then we is coated with a first was to prevent further oxidation. The kinds of a first we are chosen to be able to enhance the joining strength as inhibiting excessive be diffusion. The performance of the Be/CuCrZr/SS joint used for the ITER first wall is primarily dependent on the joining strength of the Be/CuCrZr interface. The Cr/Cu and Ti/Cr/Cu interlayers enabled the successful joining of be tile to CuCrZr plate. Moreover, ion-beam assisted deposition (IBAD) increased joining strength of the Be/CuCrZr joint mock-ups. IBAD induced the increased packing of depositing atoms, which resulted in denser and more adhesive interlayers. The interlayers formed by IBAD process revealed about 40% improved resistance to the scratch test. It is suggested that the improved adhesion of coating interlayers enabled tight joining of Be and CuCrZr blocks. As compared to without IBAD coating, the shear strength as well as the 4-point bend strength were increased more than 20% depending on interlayer types and coating conditions

  5. Inference of Large Phylogenies Using Neighbour-Joining

    DEFF Research Database (Denmark)

    Simonsen, Martin; Mailund, Thomas; Pedersen, Christian Nørgaard Storm

    2011-01-01

    The neighbour-joining method is a widely used method for phylogenetic reconstruction which scales to thousands of taxa. However, advances in sequencing technology have made data sets with more than 10,000 related taxa widely available. Inference of such large phylogenies takes hours or days using...... the Neighbour-Joining method on a normal desktop computer because of the O(n^3) running time. RapidNJ is a search heuristic which reduce the running time of the Neighbour-Joining method significantly but at the cost of an increased memory consumption making inference of large phylogenies infeasible. We present...... two extensions for RapidNJ which reduce the memory requirements and \\makebox{allows} phylogenies with more than 50,000 taxa to be inferred efficiently on a desktop computer. Furthermore, an improved version of the search heuristic is presented which reduces the running time of RapidNJ on many data...

  6. Temporalis myofascial repair of traumatic defects of the anterior fossa. Technical note.

    Science.gov (United States)

    Gillespie, R P; Shagets, F W; de los Reyes, R A

    1986-06-01

    Bilateral temporalis myofascial flaps in continuity with frontal periosteum can be used in repairing extensive dural and bone defects of the anterior cranial fossa floor. The technique of preserving and using this flap is described and offers an alternative to the use of frontal pericranial tissue for repair of anterior dural defects.

  7. Structure of a preternary complex involving a prokaryotic NHEJ DNA polymerase.

    Science.gov (United States)

    Brissett, Nigel C; Martin, Maria J; Pitcher, Robert S; Bianchi, Julie; Juarez, Raquel; Green, Andrew J; Fox, Gavin C; Blanco, Luis; Doherty, Aidan J

    2011-01-21

    In many prokaryotes, a specific DNA primase/polymerase (PolDom) is required for nonhomologous end joining (NHEJ) repair of DNA double-strand breaks (DSBs). Here, we report the crystal structure of a catalytically active conformation of Mycobacterium tuberculosis PolDom, consisting of a polymerase bound to a DNA end with a 3' overhang, two metal ions, and an incoming nucleotide but, significantly, lacking a primer strand. This structure represents a polymerase:DNA complex in a preternary intermediate state. This polymerase complex occurs in solution, stabilizing the enzyme on DNA ends and promoting nucleotide extension of short incoming termini. We also demonstrate that the invariant Arg(220), contained in a conserved loop (loop 2), plays an essential role in catalysis by regulating binding of a second metal ion in the active site. We propose that this NHEJ intermediate facilitates extension reactions involving critically short or noncomplementary DNA ends, thus promoting break repair and minimizing sequence loss during DSB repair. Copyright © 2011 Elsevier Inc. All rights reserved.

  8. Quantifying alternative splicing from paired-end RNA-sequencing data

    OpenAIRE

    Rossell, David; Stephan-Otto Attolini, Camille; Kroiss, Manuel; Stöcker, Almond

    2014-01-01

    RNA-sequencing has revolutionized biomedical research and, in particular, our ability to study gene alternative splicing. The problem has important implications for human health, as alternative splicing may be involved in malfunctions at the cellular level and multiple diseases. However, the high-dimensional nature of the data and the existence of experimental biases pose serious data analysis challenges. We find that the standard data summaries used to study alternative splicing are severely...

  9. Poorly understood and often miscategorized congenital umbilical cord hernia: an alternative repair method.

    Science.gov (United States)

    İnce, E; Temiz, A; Ezer, S S; Gezer, H Ö; Hiçsönmez, A

    2017-06-01

    Umbilical cord hernia is poorly understood and often miscategorized as "omphalocele minor". Careless clamping of the cord leads to iatrogenic gut injury in the situation of umbilical cord hernia. This study aimed to determine the characteristics and outcomes of umbilical cord hernias. We also highlight an alternative repair method for umbilical cord hernias. We recorded 15 cases of umbilical cord hernias over 10 years. The patients' data were retrospectively reviewed, and preoperative preparation of the newborn, gestational age, birth weight, other associated malformations, surgical technique used, enteral nutrition, and length of hospitalization were recorded. This study included 15 neonates with umbilical cord hernias. The mean gestational age at the time of referral was 38.2 ± 2.1

  10. Possible role of repetitious DNA in recombinatory joining during chromosome rearrangement in Drosophila melanogaster

    International Nuclear Information System (INIS)

    Lee, C.S.

    1975-01-01

    It is postulated that certain repetitious DNA components play a role in the recombination processes during chromosome rearrangements. When the distribution of silver grain densities after the in situ hybridization of repetitious DNA and the distribution of chromosome breaks due to x-irradiation are compared, a strong correlation is found for the euchromatic portion of the D. melanogaster salivary X chromosome. These observations justify the postulate above that certain repetitious DNA provides homologous regions in the DNA of broken chromosome ends necessary for proper recombinatory joining. (U.S.)

  11. Identification of auxotrophic mutants of the yeast Kluyveromyces marxianus by non-homologous end joining-mediated integrative transformation with genes from Saccharomyces cerevisiae.

    Science.gov (United States)

    Yarimizu, Tohru; Nonklang, Sanom; Nakamura, Junpei; Tokuda, Shuya; Nakagawa, Takaaki; Lorreungsil, Sasithorn; Sutthikhumpha, Surasit; Pukahuta, Charida; Kitagawa, Takao; Nakamura, Mikiko; Cha-Aim, Kamonchai; Limtong, Savitree; Hoshida, Hisashi; Akada, Rinji

    2013-12-01

    The isolation and application of auxotrophic mutants for gene manipulations, such as genetic transformation, mating selection and tetrad analysis, form the basis of yeast genetics. For the development of these genetic methods in the thermotolerant fermentative yeast Kluyveromyces marxianus, we isolated a series of auxotrophic mutants with defects in amino acid or nucleic acid metabolism. To identify the mutated genes, linear DNA fragments of nutrient biosynthetic pathway genes were amplified from Saccharomyces cerevisiae chromosomal DNA and used to directly transform the K. marxianus auxotrophic mutants by random integration into chromosomes through non-homologous end joining (NHEJ). The appearance of transformant colonies indicated that the specific S. cerevisiae gene complemented the K. marxianus mutant. Using this interspecific complementation approach with linear PCR-amplified DNA, we identified auxotrophic mutations of ADE2, ADE5,7, ADE6, HIS2, HIS3, HIS4, HIS5, HIS6, HIS7, LYS1, LYS2, LYS4, LYS9, LEU1, LEU2, MET2, MET6, MET17, TRP3, TRP4 and TRP5 without the labour-intensive requirement of plasmid construction. Mating, sporulation and tetrad analysis techniques for K. marxianus were also established. With the identified auxotrophic mutant strains and S. cerevisiae genes as selective markers, NHEJ-mediated integrative transformation with PCR-amplified DNA is an attractive system for facilitating genetic analyses in the yeast K. marxianus. Copyright © 2013 John Wiley & Sons, Ltd.

  12. Draft environmental statement related to steam-generator repair at Point Beach Nuclear Plant, Unit No. 1. Docket No. 50-266

    International Nuclear Information System (INIS)

    1983-07-01

    The staff has considered the environmental impacts and economic costs of the proposed steam generator repair at the Point Beach Nuclear Plant Unit No. 1 along with reasonable alternatives to the proposed action. The staff has concluded that the proposed repair will not significantly affect the quality of the human environment and that there are no preferable alternatives to the proposed action. Furthermore, any impacts from the repair program are outweighed by its benefits

  13. Final environmental statement related to steam-generator repair at Point Beach Nuclear Plant, Unit No. 1 (Docket No. 50-266)

    International Nuclear Information System (INIS)

    1983-09-01

    The staff hhas considered the environental impacts and economic costs of the proposed steam generator repair at the Point Beach Nuclear Plant, Unit No. 1 along with reasonable alternatives to the proposed action. The staff has concluded that the proposed repair will not significantly affect the quality of the human environment and that there are no preferable alternatives to the proposed action. Furthermore, any impacts from the repair program are outweighed by its benefits

  14. A technique for introducing looped sutures in flexor tendon repair

    Directory of Open Access Journals (Sweden)

    Kamath B

    2006-01-01

    Full Text Available Stronger flexor tendon repairs facilitate early active motion therapy protocols. Core sutures using looped suture material provide 1 ½ to twice the strength of Kessler′s technique (with four strand and six strand Tsuge technique respectively. The technique is well-described and uses preformed looped sutures (supramid. This is not available in many countries and we describe a technique whereby looped sutures can be introduced in flexor tendon repair by the use of 23 G hypodermic needle and conventional 4.0 or 5.0 sutures. This is an alternative when the custom made preformed sutures are not available. This can be practiced in zone 3 to zone 5 repairs. Technical difficulties limit its use in zone 2 repairs.

  15. Researching the Ends of Identity: Birth and Death on Social Media

    OpenAIRE

    Tama Leaver

    2015-01-01

    This paper argues that expanding the scope of social media studies to examine birth and early life at one end, and death and memorialisation at the other, demonstrates that social media is never just about an individual, but also the way individuals are always already joined together as families, groups, communities and more. Mapping these ends of identity also reveals more of the nuances of everyday social media use and its impact.

  16. Is Conventional Open Repair for Abdominal Aortic Aneurysm Feasible in Nonagenarians?

    Science.gov (United States)

    Uehara, Kyokun; Matsuda, Hitoshi; Inoue, Yosuke; Omura, Atsushi; Seike, Yoshimasa; Sasaki, Hiroaki; Kobayashi, Junjiro

    2017-09-25

    Background : Although endovascular repair for abdominal aortic aneurysm has been found to be beneficial in very elderly patients, some patients have contraindications to this procedure. For nonagenarians, the results of open repair remain unclear. The purpose of this study was to compare the outcomes of open vs. endovascular repair for abdominal aortic aneurysm in nonagenarian patients. Methods and Results : Fourteen patients undergoing open surgical repair and 24 undergoing endovascular repair for abdominal aortic aneurysm were evaluated. There was no significant difference in early mortality between the open and endovascular groups (0% vs. 4.1%, p=0.16). The open repair group required much longer hospital stays (26.4 vs. 10.6 days, respectively, p=0.003). Finally, 12 patients (86%) undergoing open repair vs. 21 (88%) undergoing endovascular repair returned home (p=0.49). During a mean follow-up period of 23.4±23.5 months, cumulative estimated 1- and 3-year survival rates were 90.0% and 48.0%, respectively in the open repair group and 90.6% and 54.9%, respectively in the endovascular repair group (p=0.51). Conclusion : Although endovascular repair for abdominal aortic aneurysm was superior in terms of recovery, the results of conventional open repair were acceptable even in nonagenarian patients. Open repair remains an alternative for patients with contraindications to endovascular repair.

  17. Joining of thin thickness SiCf/SiC composites: research of a joining composition and of an associated elaboration process

    International Nuclear Information System (INIS)

    Jacques, E.

    2012-01-01

    The present work is part of the Fourth Generation Fast Reactors program. One of the key issues is the joining of the SiC f /SiC ceramic matrix composites (CMC) to seal the combustible cladding. At the present time, no chemical composition as a joint is refractive enough to face the expected operating temperatures. The aims of this study are the following: (1) the identification of a joining composition and its associated elaboration process answering the specifications, (2) the validation of a local and fast heating process and (3) the definition of mechanical tests to characterise these joinings. We describe the methodology and the results for joining SiC and SiC f /SiC substrates at solid and liquid state using metallic silicides. Joint integrity and joint strength can be improved by adding small SiC particles to the silicides powders. Chemical reactivity, wettability tests and thermomechanical properties analysis have been carried out on the joints. Cross sections of the assembly were prepared to study the joint/substrate chemical bonding, the cracking and the crack deflection in the vicinity of the interface. Also, one of the challenge consists in using a local heating at a high temperature (around 1800 C) for a short time to avoid the degradation of the composite structure. The assemblies have been then performed in an inductive furnace but in order to prepare the joining technology, trials of local heating have also been investigated with a CO 2 laser beam and a microwave generator. Finally, descriptions of the 4-points bending mechanical test used and the associated results are presented. (author) [fr

  18. Microstructure and properties of ceramics and composites joined by plastic deformation.

    Energy Technology Data Exchange (ETDEWEB)

    Goretta, K. C.; Singh, D.; Chen, N.; Gutierrez-Mora, F.; Lorenzo-Martin, M. de la, Cinta; Dominguez-Rodriguez, A.; Routbort, J. L.; Energy Systems; Univ. of Seville

    2008-12-01

    A review is presented of the design of suitable materials systems for joining by high-temperature plastic deformation, details of the joining techniques, microstructures and properties of the resulting composite bodies, and prospects and limitation for this type of joining technology. Joining parameters and resulting forms are discussed for Al{sub 2}O{sub 3}/mullite particulate composites, Y{sub 2}O{sub 3}-stabilized ZrO{sub 2} particulate/Al{sub 2}O{sub 3} particulate and whisker-reinforced composites, hydroxyapatite bioceramics, La{sub 0.85}Sr{sub 0.15}MnO{sub 3} electronic ceramics, MgF{sub 2} optical ceramics, and Ni{sub 3}Al intermetallics. Results are contrasted with those obtained by other methods of joining brittle, high-temperature materials, with special focus on durability and mechanical properties.

  19. Microstructure and properties of ceramics and composites joined by plastic deformation

    Energy Technology Data Exchange (ETDEWEB)

    Goretta, K.C. [Argonne National Laboratory, Argonne, IL 60439-4838 (United States)], E-mail: ken.goretta@aoard.af.mil; Singh, D.; Chen Nan [Argonne National Laboratory, Argonne, IL 60439-4838 (United States); Gutierrez-Mora, F.; Cinta Lorenzo-Martin, M. de la [Argonne National Laboratory, Argonne, IL 60439-4838 (United States); University of Seville, Seville 41080 (Spain); Dominguez-Rodriguez, A. [University of Seville, Seville 41080 (Spain); Routbort, J.L. [Argonne National Laboratory, Argonne, IL 60439-4838 (United States)

    2008-12-20

    A review is presented of the design of suitable materials systems for joining by high-temperature plastic deformation, details of the joining techniques, microstructures and properties of the resulting composite bodies, and prospects and limitation for this type of joining technology. Joining parameters and resulting forms are discussed for Al{sub 2}O{sub 3}/mullite particulate composites, Y{sub 2}O{sub 3}-stabilized ZrO{sub 2} particulate/Al{sub 2}O{sub 3} particulate and whisker-reinforced composites, hydroxyapatite bioceramics, La{sub 0.85}Sr{sub 0.15}MnO{sub 3} electronic ceramics, MgF{sub 2} optical ceramics, and Ni{sub 3}Al intermetallics. Results are contrasted with those obtained by other methods of joining brittle, high-temperature materials, with special focus on durability and mechanical properties.

  20. Pulmonary stenosis and pulmonary regurgitation: both ends of the spectrum in residual hemodynamic impairment after tetralogy of Fallot repair

    Science.gov (United States)

    Park, Han Ki

    2013-01-01

    Repair of tetralogy of Fallot (TOF) has shown excellent outcomes. However it leaves varying degrees of residual hemodynamic impairment, with severe pulmonary stenosis (PS) and free pulmonary regurgitation (PR) at both ends of the spectrum. Since the 1980s, studies evaluating late outcomes after TOF repair revealed the adverse impacts of residual chronic PR on RV volume and function; thus, a turnaround of operational strategies has occurred from aggressive RV outflow tract (RVOT) reconstruction for complete relief of RVOT obstruction to conservative RVOT reconstruction for limiting PR. This transformation has raised the question of how much residual PS after conservative RVOT reconstruction is acceptable. Besides, as pulmonary valve replacement (PVR) increases in patients with RV deterioration from residual PR, there is concern regarding when it should be performed. Regarding residual PS, several studies revealed that PS in addition to PR was associated with less PR and a small RV volume. This suggests that PS combined with PR makes RV diastolic property to protect against dilatation through RV hypertrophy and supports conservative RVOT enlargement despite residual PS. Also, several studies have revealed the pre-PVR threshold of RV parameters for the normalization of RV volume and function after PVR, and based on these results, the indications for PVR have been revised. Although there is no established strategy, better understanding of RV mechanics, development of new surgical and interventional techniques, and evidence for the effect of PVR on RV reverse remodeling and its late outcome will aid us to optimize the management of TOF. PMID:23807889

  1. Pulmonary stenosis and pulmonary regurgitation: both ends of the spectrum in residual hemodynamic impairment after tetralogy of Fallot repair.

    Science.gov (United States)

    Yoo, Byung Won; Park, Han Ki

    2013-06-01

    Repair of tetralogy of Fallot (TOF) has shown excellent outcomes. However it leaves varying degrees of residual hemodynamic impairment, with severe pulmonary stenosis (PS) and free pulmonary regurgitation (PR) at both ends of the spectrum. Since the 1980s, studies evaluating late outcomes after TOF repair revealed the adverse impacts of residual chronic PR on RV volume and function; thus, a turnaround of operational strategies has occurred from aggressive RV outflow tract (RVOT) reconstruction for complete relief of RVOT obstruction to conservative RVOT reconstruction for limiting PR. This transformation has raised the question of how much residual PS after conservative RVOT reconstruction is acceptable. Besides, as pulmonary valve replacement (PVR) increases in patients with RV deterioration from residual PR, there is concern regarding when it should be performed. Regarding residual PS, several studies revealed that PS in addition to PR was associated with less PR and a small RV volume. This suggests that PS combined with PR makes RV diastolic property to protect against dilatation through RV hypertrophy and supports conservative RVOT enlargement despite residual PS. Also, several studies have revealed the pre-PVR threshold of RV parameters for the normalization of RV volume and function after PVR, and based on these results, the indications for PVR have been revised. Although there is no established strategy, better understanding of RV mechanics, development of new surgical and interventional techniques, and evidence for the effect of PVR on RV reverse remodeling and its late outcome will aid us to optimize the management of TOF.

  2. Pulmonary stenosis and pulmonary regurgitation: both ends of the spectrum in residual hemodynamic impairment after tetralogy of Fallot repair

    Directory of Open Access Journals (Sweden)

    Byung Won Yoo

    2013-06-01

    Full Text Available Repair of tetralogy of Fallot (TOF has shown excellent outcomes. However it leaves varying degrees of residual hemodynamic impairment, with severe pulmonary stenosis (PS and free pulmonary regurgitation (PR at both ends of the spectrum. Since the 1980s, studies evaluating late outcomes after TOF repair revealed the adverse impacts of residual chronic PR on RV volume and function; thus, a turnaround of operational strategies has occurred from aggressive RV outflow tract (RVOT reconstruction for complete relief of RVOT obstruction to conservative RVOT reconstruction for limiting PR. This transformation has raised the question of how much residual PS after conservative RVOT reconstruction is acceptable. Besides, as pulmonary valve replacement (PVR increases in patients with RV deterioration from residual PR, there is concern regarding when it should be performed. Regarding residual PS, several studies revealed that PS in addition to PR was associated with less PR and a small RV volume. This suggests that PS combined with PR makes RV diastolic property to protect against dilatation through RV hypertrophy and supports conservative RVOT enlargement despite residual PS. Also, several studies have revealed the pre-PVR threshold of RV parameters for the normalization of RV volume and function after PVR, and based on these results, the indications for PVR have been revised. Although there is no established strategy, better understanding of RV mechanics, development of new surgical and interventional techniques, and evidence for the effect of PVR on RV reverse remodeling and its late outcome will aid us to optimize the management of TOF.

  3. Strategies to Improve Regeneration of the Soft Palate Muscles After Cleft Palate Repair

    Science.gov (United States)

    Carvajal Monroy, Paola L.; Grefte, Sander; Kuijpers-Jagtman, Anne Marie; Wagener, Frank A.D.T.G.

    2012-01-01

    Children with a cleft in the soft palate have difficulties with speech, swallowing, and sucking. These patients are unable to separate the nasal from the oral cavity leading to air loss during speech. Although surgical repair ameliorates soft palate function by joining the clefted muscles of the soft palate, optimal function is often not achieved. The regeneration of muscles in the soft palate after surgery is hampered because of (1) their low intrinsic regenerative capacity, (2) the muscle properties related to clefting, and (3) the development of fibrosis. Adjuvant strategies based on tissue engineering may improve the outcome after surgery by approaching these specific issues. Therefore, this review will discuss myogenesis in the noncleft and cleft palate, the characteristics of soft palate muscles, and the process of muscle regeneration. Finally, novel therapeutic strategies based on tissue engineering to improve soft palate function after surgical repair are presented. PMID:22697475

  4. Joining of Aluminium Alloy Sheets by Rectangular Mechanical Clinching

    International Nuclear Information System (INIS)

    Abe, Y.; Mori, K.; Kato, T.

    2011-01-01

    A mechanical clinching has the advantage of low running costs. However, the joint strength is not high. To improve the maximum load of the joined sheets by a mechanical clinching, square and rectangular mechanical clinching were introduced. In the mechanical clinching, the two sheets are mechanically joined by forming an interlock between the lower and upper sheets by the punch and die. The joined length with the interlock was increased by the rectangular punch and die. The deforming behaviours of the sheets in the mechanical clinching were investigated, and then the interlock in the sheets had distribution in the circumference of the projection. Although the interlocks were formed in both projection side and diagonal, the interlock in the diagonal was smaller because of the long contact length between the lower sheet and the die cavity surface. The maximum load of the joined sheets by the rectangular mechanical clinching was two times larger than the load by the round mechanical clinching.

  5. End-to-side neurorraphy: a long-term study of neural regeneration in a rat model.

    Science.gov (United States)

    Tarasidis, G; Watanabe, O; Mackinnon, S E; Strasberg, S R; Haughey, B H; Hunter, D A

    1998-10-01

    This study evaluated long-term reinnervation of an end-to-side neurorraphy and the resultant functional recovery in a rat model. The divided distal posterior tibial nerve was repaired to the side of an intact peroneal nerve. Control groups included a cut-and-repair of the posterior tibial nerve and an end-to-end repair of the peroneal nerve to the posterior tibial nerve. Evaluations included walking-track analysis, nerve conduction studies, muscle mass measurements, retrograde nerve tracing, and histologic evaluation. Walking tracks indicated poor recovery of posterior tibial nerve function in the experimental group. No significant difference in nerve conduction velocities was seen between the experimental and control groups. Gastrocnemius muscle mass measurements revealed no functional recovery in the experimental group. Similarly, retrograde nerve tracing revealed minimal motor neuron staining in the experimental group. However, some sensory staining was seen within the dorsal root ganglia of the end-to-side group. Histologic study revealed minimal myelinated axonal regeneration in the experimental group as compared with findings in the other groups. These results suggest that predominantly sensory regeneration occurs in an end-to-side neurorraphy at an end point of 6 months.

  6. Fast and scalable inequality joins

    KAUST Repository

    Khayyat, Zuhair; Lucia, William; Singh, Meghna; Ouzzani, Mourad; Papotti, Paolo; Quiané -Ruiz, Jorge Arnulfo; Tang, Nan; Kalnis, Panos

    2016-01-01

    an incremental inequality join algorithm to handle scenarios where data keeps changing. We have implemented a centralized version of these algorithms on top of PostgreSQL, a distributed version on top of Spark SQL, and an existing data cleaning system, Nadeef

  7. Laser repair welding of molds with various pulse shapes

    Directory of Open Access Journals (Sweden)

    M. Pleterski

    2010-01-01

    Full Text Available Repair welding of cold-work tool steels with conventional methods is very difficult due to cracking during remelting or cladding and is generally performed with preheating. As an alternative, repair welding with laser technology has recently been used. This paper presents the influence of different pulse shapes on welding of such tools with the pulsed Nd:YAG laser. Repair welding tests were carried out on AISI D2 tool steel, quenched and tempered to hardness of 56 HRc, followed by microstructural analysis and investigation of defects with scanning electron microscopy. Test results suggest that it is possible to obtain sound welds without preheating, with the right selection of welding parameters and appropriate pulse shape.

  8. The Ku heterodimer and the metabolism of single-ended DNA double-strand breaks.

    Science.gov (United States)

    Balestrini, Alessia; Ristic, Dejan; Dionne, Isabelle; Liu, Xiao Z; Wyman, Claire; Wellinger, Raymund J; Petrini, John H J

    2013-06-27

    Single-ended double-strand breaks (DSBs) are a common form of spontaneous DNA break, generated when the replisome encounters a discontinuity in the DNA template. Given their prevalence, understanding the mechanisms governing the fate(s) of single-ended DSBs is important. We describe the influence of the Ku heterodimer and Mre11 nuclease activity on processing of single-ended DSBs. Separation-of-function alleles of yku70 were derived that phenocopy Ku deficiency with respect to single-ended DSBs but remain proficient for NHEJ. The Ku mutants fail to regulate Exo1 activity, and bypass the requirement for Mre11 nuclease activity in the repair of camptothecin-induced single-ended DSBs. Ku mutants exhibited reduced affinity for DNA ends, manifest as both reduced end engagement and enhanced probability of diffusing inward on linear DNA. This study reveals an interplay between Ku and Mre11 in the metabolism of single-ended DSBs that is distinct from repair pathway choice at double-ended DSBs. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  9. 49 CFR 192.283 - Plastic pipe: Qualifying joining procedures.

    Science.gov (United States)

    2010-10-01

    ... 49 Transportation 3 2010-10-01 2010-10-01 false Plastic pipe: Qualifying joining procedures. 192... Materials Other Than by Welding § 192.283 Plastic pipe: Qualifying joining procedures. (a) Heat fusion... for making plastic pipe joints by a heat fusion, solvent cement, or adhesive method, the procedure...

  10. C-terminal low-complexity sequence repeats of Mycobacterium smegmatis Ku modulate DNA binding.

    Science.gov (United States)

    Kushwaha, Ambuj K; Grove, Anne

    2013-01-24

    Ku protein is an integral component of the NHEJ (non-homologous end-joining) pathway of DSB (double-strand break) repair. Both eukaryotic and prokaryotic Ku homologues have been characterized and shown to bind DNA ends. A unique feature of Mycobacterium smegmatis Ku is its basic C-terminal tail that contains several lysine-rich low-complexity PAKKA repeats that are absent from homologues encoded by obligate parasitic mycobacteria. Such PAKKA repeats are also characteristic of mycobacterial Hlp (histone-like protein) for which they have been shown to confer the ability to appose DNA ends. Unexpectedly, removal of the lysine-rich extension enhances DNA-binding affinity, but an interaction between DNA and the PAKKA repeats is indicated by the observation that only full-length Ku forms multiple complexes with a short stem-loop-containing DNA previously designed to accommodate only one Ku dimer. The C-terminal extension promotes DNA end-joining by T4 DNA ligase, suggesting that the PAKKA repeats also contribute to efficient end-joining. We suggest that low-complexity lysine-rich sequences have evolved repeatedly to modulate the function of unrelated DNA-binding proteins.

  11. Scarless Cas9 Assisted Recombineering (no‐SCAR) in Escherichia coli, an Easy‐to‐Use System for Genome Editing

    OpenAIRE

    Reisch, Christopher R; Jones, Kristala L.

    2018-01-01

    The discovery and development of genome editing systems that leverage the site‐specific DNA endonuclease system CRISPR/Cas9 has fundamentally changed the ease and speed of genome editing in many organisms. In eukaryotes, the CRISPR/Cas9 system utilizes a “guide” RNA to enable the Cas9 nuclease to make a double‐strand break at a particular genome locus, which is repaired by non‐homologous end joining (NHEJ) repair enzymes, often generating random mutations in the process. A specific alteration...

  12. Superplasticity and joining of zirconia-based ceramics

    International Nuclear Information System (INIS)

    Dominguez-Rodriguez, A.; Gutierrez-Mora, F.; Jimenez-Melendo, M.; Chaim, R.; Routbort, J. L.

    1999-01-01

    Steady-state creep and joining of alumina/zirconia composites containing alumina volume fractions of 20, 60, and 85% have been investigated between 1,250 and 1,350 C. Superplasticity of these compounds is controlled by grain-boundary sliding and the creep rate is a function of alumina volume fraction, not grain size. Using the principles of superplasticity, pieces of the composite have been joined by applying the stress required to achieve 5 to 10% strain to form a strong interface at temperatures as low as 1,200 C

  13. Superplasticity and joining of zirconia-based ceramics

    International Nuclear Information System (INIS)

    Gutierrez-Mora, F.; Dominguez-Rodriguez, A.; Jimenez-Melendo, M.; Chaim, R.; Ravi, G.B.; Routbort, J.L.

    2000-01-01

    Steady-state creep and joining of alumina/zirconia composites containing alumina volume fractions of 20, 60 and 85% have been investigated between 1,250 and 1,350 C. Superplasticity of these compounds is controlled by grain-boundary sliding and the creep rate is a function of alumina volume fraction, not grain size. Using the principles of superplasticity, pieces of the composite have been joined by applying the stress required to achieve 5 to 10% strain to form a strong interface at temperatures as low as 1,200 C

  14. Three dimensional multi-pass repair weld simulations

    International Nuclear Information System (INIS)

    Elcoate, C.D.; Dennis, R.J.; Bouchard, P.J.; Smith, M.C.

    2005-01-01

    Full 3-dimensional (3-D) simulation of multi-pass weld repairs is now feasible and practical given the development of improved analysis tools and significantly greater computer power. This paper presents residual stress results from 3-D finite element (FE) analyses simulating a long (arc length of 62 deg. ) and a short (arc length of 20 deg. ) repair to a girth weld in a 19.6 mm thick, 432 mm outer diameter cylindrical test component. Sensitivity studies are used to illustrate the importance of weld bead inter-pass temperature assumptions and to show where model symmetry can be used to reduce the analysis size. The predicted residual stress results are compared with measured axial, hoop and radial through-wall profiles in the heat affected zone of the test component repairs. A good overall agreement is achieved between neutron diffraction and deep hole drilling measurements and the prediction at the mid-length position of the short repair. These results demonstrate that a coarse 3-D FE model, using a 'block-dumped' weld bead deposition approach (rather than progressively depositing weld metal), can accurately capture the important components of a short repair weld residual stress field. However, comparisons of measured with predicted residual stress at mid-length and stop-end positions in the long repair are less satisfactory implying some shortcomings in the FE modelling approach that warrant further investigation

  15. Development of a procedure for forming assisted thermal joining of tubes

    Science.gov (United States)

    Chen, Hui; Löbbe, Christian; Staupendahl, Daniel; Tekkaya, A. Erman

    2018-05-01

    With the demand of lightweight design in the automotive industry, not only the wall-thicknesses of tubular components of the chassis or spaceframe are continuously decreased. Also the thicknesses of exhaust system parts are reduced to save material and mass. However, thinner tubular parts bring about additional challenges in joining. Welding or brazing methods, which are utilized in joining tubes with specific requirements concerning leak tightness, are sensitive to the gap between the joining partners. Furthermore, a large joining area is required to ensure the durability of the joint. The introduction of a forming step in the assembled state prior to thermal joining can define and control the gap for subsequent brazing or welding. The mechanical pre-joint resulting from the previously described calibration step also results in easier handling of the tubes prior to thermal joining. In the presented investigation, a spinning process is utilized to produce force-fit joints of varying lengths and diameter reduction and form-fit joints with varying geometrical attributes. The spinning process facilitates a high formability and geometrical flexibility, while at the achievable precision is high and the process forces are low. The strength of the joints is used to evaluate the joint quality. Finally, a comparison between joints produced by forming with subsequent brazing and original tube is conducted, which presents the high performance of the developed procedure for forming assisted thermal joining.

  16. Model Checking Processes Specified In Join-Calculus Algebra

    Directory of Open Access Journals (Sweden)

    Sławomir Piotr Maludziński

    2014-01-01

    Full Text Available This article presents a model checking tool used to verify concurrent systems specified in join-calculus algebra. The temporal properties of systems under verification are expressed in CTL logic. Join-calculus algebra with its operational semantics defined by the chemical abstract machine serves as the basic method for the specification of concurrent systems and their synchronization mechanisms, and allows the examination of more complex systems.

  17. Molecular dynamics simulation of joining process of Ag-Au nanowires and mechanical properties of the hybrid nanojoint

    Directory of Open Access Journals (Sweden)

    Su Ding

    2015-05-01

    Full Text Available The nanojoining process of Ag-Au hybrid nanowires at 800K was comprehensively studied by virtue of molecular dynamics (MD simulation. Three kinds of configurations including end-to-end, T-like and X-like were built in the simulation aiming to understand the nanojoining mechanism. The detailed dynamic evolution of atoms, crystal structure transformation and defects development during the nanojoining processes were performed. The results indicate that there are two stages in the nanojoining process of Ag-Au nanowires which are atom diffusion and new bonds formation. Temperature is a key parameter affecting both stages ascribed to the energy supply and the optimum temperature for Ag-Au nanojoint with diameter of 4.08 nm has been discussed. The mechanical properties of the nanojoint were examined with simulation of tensile test on the end-to-end joint. It was revealed that the nanojoint was strong enough to resist fracture at the joining area.

  18. Characterization of Genomic Deletion Efficiency Mediated by Clustered Regularly Interspaced Palindromic Repeats (CRISPR)/Cas9 Nuclease System in Mammalian Cells*♦

    Science.gov (United States)

    Canver, Matthew C.; Bauer, Daniel E.; Dass, Abhishek; Yien, Yvette Y.; Chung, Jacky; Masuda, Takeshi; Maeda, Takahiro; Paw, Barry H.; Orkin, Stuart H.

    2014-01-01

    The clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated (Cas) 9 nuclease system has provided a powerful tool for genome engineering. Double strand breaks may trigger nonhomologous end joining repair, leading to frameshift mutations, or homology-directed repair using an extrachromosomal template. Alternatively, genomic deletions may be produced by a pair of double strand breaks. The efficiency of CRISPR/Cas9-mediated genomic deletions has not been systematically explored. Here, we present a methodology for the production of deletions in mammalian cells, ranging from 1.3 kb to greater than 1 Mb. We observed a high frequency of intended genomic deletions. Nondeleted alleles are nonetheless often edited with inversions or small insertion/deletions produced at CRISPR recognition sites. Deleted alleles also typically include small insertion/deletions at predicted deletion junctions. We retrieved cells with biallelic deletion at a frequency exceeding that of probabilistic expectation. We demonstrate an inverse relationship between deletion frequency and deletion size. This work suggests that CRISPR/Cas9 is a robust system to produce a spectrum of genomic deletions to allow investigation of genes and genetic elements. PMID:24907273

  19. MMEJ-assisted gene knock-in using TALENs and CRISPR-Cas9 with the PITCh systems.

    Science.gov (United States)

    Sakuma, Tetsushi; Nakade, Shota; Sakane, Yuto; Suzuki, Ken-Ichi T; Yamamoto, Takashi

    2016-01-01

    Programmable nucleases enable engineering of the genome by utilizing endogenous DNA double-strand break (DSB) repair pathways. Although homologous recombination (HR)-mediated gene knock-in is well established, it cannot necessarily be applied in every cell type and organism because of variable HR frequencies. We recently reported an alternative method of gene knock-in, named the PITCh (Precise Integration into Target Chromosome) system, assisted by microhomology-mediated end-joining (MMEJ). MMEJ harnesses independent machinery from HR, and it requires an extremely short homologous sequence (5-25 bp) for DSB repair, resulting in precise gene knock-in with a more easily constructed donor vector. Here we describe a streamlined protocol for PITCh knock-in, including the design and construction of the PITCh vectors, and their delivery to either human cell lines by transfection or to frog embryos by microinjection. The construction of the PITCh vectors requires only a few days, and the entire process takes ∼ 1.5 months to establish knocked-in cells or ∼ 1 week from injection to early genotyping in frog embryos.

  20. Biomaterial-mediated strategies targeting vascularization for bone repair.

    Science.gov (United States)

    García, José R; García, Andrés J

    2016-04-01

    Repair of non-healing bone defects through tissue engineering strategies remains a challenging feat in the clinic due to the aversive microenvironment surrounding the injured tissue. The vascular damage that occurs following a bone injury causes extreme ischemia and a loss of circulating cells that contribute to regeneration. Tissue-engineered constructs aimed at regenerating the injured bone suffer from complications based on the slow progression of endogenous vascular repair and often fail at bridging the bone defect. To that end, various strategies have been explored to increase blood vessel regeneration within defects to facilitate both tissue-engineered and natural repair processes. Developments that induce robust vascularization will need to consolidate various parameters including optimization of embedded therapeutics, scaffold characteristics, and successful integration between the construct and the biological tissue. This review provides an overview of current strategies as well as new developments in engineering biomaterials to induce reparation of a functional vascular supply in the context of bone repair.

  1. Aberrant repair and fibrosis development in skeletal muscle

    Directory of Open Access Journals (Sweden)

    Mann Christopher J

    2011-05-01

    Full Text Available Abstract The repair process of damaged tissue involves the coordinated activities of several cell types in response to local and systemic signals. Following acute tissue injury, infiltrating inflammatory cells and resident stem cells orchestrate their activities to restore tissue homeostasis. However, during chronic tissue damage, such as in muscular dystrophies, the inflammatory-cell infiltration and fibroblast activation persists, while the reparative capacity of stem cells (satellite cells is attenuated. Abnormal dystrophic muscle repair and its end stage, fibrosis, represent the final common pathway of virtually all chronic neurodegenerative muscular diseases. As our understanding of the pathogenesis of muscle fibrosis has progressed, it has become evident that the muscle provides a useful model for the regulation of tissue repair by the local microenvironment, showing interplay among muscle-specific stem cells, inflammatory cells, fibroblasts and extracellular matrix components of the mammalian wound-healing response. This article reviews the emerging findings of the mechanisms that underlie normal versus aberrant muscle-tissue repair.

  2. Glass-Metal Joining in Nuclear Environment: the State of the Art

    International Nuclear Information System (INIS)

    Jacobs, M.

    2007-01-01

    Full text of publication follows: In the ITER fusion machine and in material testing fission reactors, it is not possible to avoid the use of non-metallic materials like glass for example. There is therefore a need to apply metal to glass joints. This problem arose already at the beginning of the 19. century when the electric light bulb was invented. Nowadays this type of glass-metal joint is very successful and widely used in the electronic industry. In the case of ITER and material testing reactors, glass-metal joints are necessary for the fixation of the optical windows and optical fibres to a metal structure to perform diagnostics. These types of joints are still difficult to make and their behaviour is not fully understood. A joint between glass and metal for a nuclear or fusion application has indeed to resist high temperatures and high neutron fluences, while keeping a good mechanical strength and remaining leak tight. These characteristics are difficult to obtain under these severe conditions. This paper presents an overview of the different joining technologies that can be used to join glass to metal in a severe nuclear environment. The working mechanism of the technologies are explained, together with their respective advantages and drawbacks. Three different types of joining are discussed: fastening, liquid phase joining and solid phase joining. Fastening is a mechanical attachment technique, not achieving easily hermetic seals. Liquid and solid phase joining on the other hand form a real bond, what makes the joint much stronger. The most important technologies using liquid phase joining are adhesive bonding, fusion welding and brazing. In the case of the solid phase joining the choices are ultrasonic torsion welding, diffusion bonding and electrostatic bonding. If it is usually not possible to join the glass directly to the metal, an interlayer must be used. One speaks then of indirect joining. The paper will conclude with a discussion on the best

  3. DNA excision repair in permeable human fibroblasts

    International Nuclear Information System (INIS)

    Kaufmann, W.K.; Bodell, W.J.; Cleaver, J.E.

    1983-01-01

    U.v. irradiation of confluent human fibroblasts activated DNA repair, aspects of which were characterized in the cells after they were permeabilized. Incubation of intact cells for 20 min between irradiation and harvesting was necessary to obtain a maximum rate of reparative DNA synthesis. Cells harvested immediately after irradiation before repair was initiated displayed only a small stimulation of DNA synthesis, indicating that permeable cells have a reduced capacity to recognize pyrimidine dimers and activate repair. The distribution of sizes of DNA strands labeled during 10 min of reparative DNA synthesis resembled that of parental DNA. However, during a 60-min incubation of permeable cells at 37 degrees C, parental DNA and DNA labeled by reparative DNA synthesis were both cleaved to smaller sizes. Cleavage also occurred in unirradiated cells, indicating that endogenous nuclease was active during incubation. Repair patches synthesized in permeable cells displayed increased sensitivity to digestion by micrococcal nuclease. However, the change in sensitivity during a chase with unlabeled DNA precursors was small, suggesting that reassembly of nucleosome structure at sites of repair was impaired. To examine whether this deficiency was due to a preponderance of incomplete or unligated repair patches, 3H-labeled (repaired) DNA was purified, then digested with exonuclease III and nuclease S1 to probe for free 3' ends and single-stranded regions. About 85% of the [3H]DNA synthesized during a 10-min pulse resisted digestion, suggesting that a major fraction of the repair patches that were filled were also ligated. U.v. light-activated DNA synthesis in permeable cells, therefore, appears to represent the continuation of reparative gap-filling at sites of excision repair activated within intact cells. Gap-filling and ligation were comparatively efficient processes in permeable cells

  4. Taiwan university joins search for antimatter

    CERN Multimedia

    Chiu, Y

    2002-01-01

    National Cheng Kung University announced yesterday that it will be joining the Alpha Magnetic Spectrometer, an international scientific research project aimed at looking for antimatter in space (1 page).

  5. High-energy electron beams for ceramic joining

    Science.gov (United States)

    Turman, Bob N.; Glass, S. J.; Halbleib, J. A.; Helmich, D. R.; Loehman, Ron E.; Clifford, Jerome R.

    1995-03-01

    Joining of structural ceramics is possible using high melting point metals such as Mo and Pt that are heated with a high energy electron beam, with the potential for high temperature joining. A 10 MeV electron beam can penetrate through 1 cm of ceramic, offering the possibility of buried interface joining. Because of transient heating and the lower heat capacity of the metal relative to the ceramic, a pulsed high power beam has the potential for melting the metal without decomposing or melting the ceramic. We have demonstrated the feasibility of the process with a series of 10 MeV, 1 kW electron beam experiments. Shear strengths up to 28 MPa have been measured. This strength is comparable to that reported in the literature for bonding silicon nitride (Si3N4) to molybdenum with copper-silver-titanium braze, but weaker than that reported for Si3N4 - Si3N4 with gold-nickel braze. The bonding mechanism appears to be formation of a thin silicide layer. Beam damage to the Si3N4 was also assessed.

  6. Top scientists join Stephen Hawking at Perimeter Institute

    Science.gov (United States)

    Banks, Michael

    2009-03-01

    Nine leading researchers are to join Stephen Hawking as visiting fellows at the Perimeter Institute for Theoretical Physics in Ontario, Canada. The researchers, who include string theorists Leonard Susskind from Stanford University and Asoka Sen from the Harisch-Chandra Research Institute in India, will each spend a few months of the year at the institute as "distinguished research chairs". They will be joined by another 30 scientists to be announced at a later date.

  7. Pulsed Joining Of Body-In-White Components

    Energy Technology Data Exchange (ETDEWEB)

    Bonnen, John [Ford Motor Company, Dearborn, MI (United States)

    2014-09-30

    The objective of this project was to develop cost efficient high quality pulsed welding (PW) technology for joining 6xxx Aluminum and High Strength Steel (with tensile strength above 580MPa) components enabling broad usage of hydroformed parts and leading to substantial weight reduction of cars and trucks to reduce US demand on petroleum, lower carbon emissions and energy expenditures. In general, pulsed welding is a form of impact welding where two dissimilar metal pieces are joined by accelerating one to velocities exceeding 300m/s at which point the first piece strikes the second and forms a weld. In this work, two methods were used to accelerate the flyer material: Electro-Magnetic (EM) pulse and Electro-Hydraulic (EH) pulse launching. The advantage of pulsed welding techniques is that welds can be formed between two materials that cannot otherwise be welded: high strength aluminum and high strength steel. The technical objectives of the project included: 1) developing cost affordable production feasible tooling design for PW of 6xxx aluminum to High Strength steel with strengths above 580MPa; 2) demonstrating that fabricated joints can exceed the required service load strength initially at the coupon level and then at the component level; 3) developing fundamental understanding of the mechanisms of joint formation and conditions leading to formation of high quality PW joint; and 4) creating a numerical model predicting the tooling and electric discharge parameters necessary for the joint formation and that satisfy the targeted strength parameters. The project successfully developed: 1) EM and EH pulsed welds between high strength aluminum with tensile strengths exceeding 240MPa and steels exceeding 580MPa; 2) pulsed welds of extrusions with strengths exceeding project requirements; 3) EM and EH flyer launch models and 4) weld interface formation models. However, the grant holder, Ford Motor Company, could see no path to commercialization and the work was

  8. ROMANIA'S MACROECONOMIC ACHIEVEMENTS FOR JOINING THE UNIQUE EUROPEAN CURENCY

    Directory of Open Access Journals (Sweden)

    SILVIA POPESCU

    2011-04-01

    Full Text Available The Romanian government has announced plans to join the eurozone by 2015. Currently, the leu is not yet part of ERM II but plans to join in 2010-2012. The economic advantages of the monetary union grow with expansion of the Euro zone. There is also a high level of skepticism; the main fear about the Euro is the inflation –that is considerable promoted by the Euro currency’s exchange rate in comparison with 2002; another restraint is due to member states inability to establish their own interest rates. The IMF arose the option of joining the Euro zone criteria relaxing. A one-sided Euro’s joining was suggested by International Monetary Fund on March-April 2009, in a confidential report mentioned by The Financial Times as the emergent states in Central and Eastern Europe to be able to pass to the unique currency, but not being represented in the Central European Bank Board. By its side, CEB considers that emergent states of the European Union must not pass to the unique currency unilaterally, because such a fact could under-mine the trust in Euro currency worldwide. This option would hardly deepen the macroeconomic controversies inside the Euro zone and would contradict the previous conditions already imposed. An acceptable solution could be the fastening of emergent countries joining the Exchange Rate Mechanism 2, after they are aware of risks arisen by such a step. The European Commission endorses in the Convergence Report on 2010 that Romania doesn’t meet any criteria needed by passing to the unique European currency, respectively: prices stability; budget position of the government; stability of exchange rate; interest convergence on long run and there are also law impediments. Our paper discusses arguments for a faster passing to the Euro currency versus arguments for a late joining the Euro currency in Romania.

  9. Microwave joining of SiC

    Energy Technology Data Exchange (ETDEWEB)

    Silberglitt, R.; Ahmad, I. [FM Technologies, Inc., Fairfax, VA (United States); Black, W.M. [George Mason Univ., Fairfax, VA (United States)] [and others

    1995-05-01

    The purpose of this work is to optimize the properties of SiC-SiC joints made using microwave energy. The current focus is on optimization of time-temperature profiles, production of SiC from chemical precursors, and design of new applicators for joining of long tubes.

  10. Joining of SiCf/SiC composites for thermonuclear fusion reactors

    International Nuclear Information System (INIS)

    Ferraris, M.; Badini, C.; Montorsi, M.; Appendino, P.; Scholz, H.W.

    1994-01-01

    Due to their favourable radiological behaviour, SiC f /SiC composites are promising structural materials for future use in fusion reactors. A problem to cope with is the joining of the ceramic composite material (CMC) to itself for more complex structures. Maintenance concepts for a reactor made of SiC f /SiC will demand a method of joining. The joining agents should comply with the low-activation approach of the base material. With the acceptable elements Si and Mg, sandwich structures of composite/metal/composite were prepared in Ar atmosphere at temperatures just above the melting points of the metals. Another promising route is the use of joining agents of boro-silicate glasses: their composition can be tailored to obtain softening temperatures of interest for fusion applications. The glassy joint can be easily ceramised to improve thermomechanical properties. The joining interfaces were investigated by SEM-EDS, XRD and mechanical tests. ((orig.))

  11. Why join the Staff Association

    CERN Multimedia

    Association du personnel

    2011-01-01

    Becoming a member of the Staff Association (SA) is above all a personal choice, showing that the joining person’s commitment and adherence to values such as solidarity, social cohesion, etc.In September, the SA launches a membership campaign to convince a maximum number to join, to inform, arouse interest and support. Posters, emails and individual contacts are part of the campaign programme, just like this editorial. As far as individual contacts are concerned, we ask you to give time and lend an ear to the delegates of your department in the Staff Council, who will approach you, in order to make an open and constructive discussion possible. Do not hesitate to ask questions and let them know your thoughts about the SA, as (constructive) criticism enables us to progress. The Staff Association and its role of collective representation The Staff Association, via its delegates, represents collectively all staff of the Organization before the Director-General and Member States. To do this, staff rep...

  12. Flexible Friction Stir Joining Technology

    Energy Technology Data Exchange (ETDEWEB)

    Feng, Zhili [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Lim, Yong Chae [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States); Mahoney, Murray [MegaStir Technologies LLC, Orem, UT (United States); Sanderson, Samuel [MegaStir Technologies LLC, Orem, UT (United States); Larsen, Steve [MegaStir Technologies LLC, Orem, UT (United States); Steel, Russel [MegaStir Technologies LLC, Orem, UT (United States); Fleck, Dale [MegaStir Technologies LLC, Orem, UT (United States); Fairchild, Doug P [ExxonMobil, Upstream Research Company (URC), Houston, TX (United States); Wasson, Andrew J [ExxonMobil, Upstream Research Company (URC), Houston, TX (United States); Babb, Jon [MegaStir Technologies LLC, Orem, UT (United States); Higgins, Paul [MegaStir Technologies LLC, Orem, UT (United States)

    2015-07-23

    Reported herein is the final report on a U.S. Department of Energy (DOE) Advanced Manufacturing Office (AMO) project with industry cost-share that was jointly carried out by Oak Ridge National Laboratory (ORNL), ExxonMobil Upstream Research Company (ExxonMobil), and MegaStir Technologies (MegaStir). The project was aimed to advance the state of the art of friction stir welding (FSW) technology, a highly energy-efficient solid-state joining process, for field deployable, on-site fabrications of large, complex and thick-sectioned structures of high-performance and high-temperature materials. The technology innovations developed herein attempted to address two fundamental shortcomings of FSW: 1) the inability for on-site welding and 2) the inability to weld thick section steels, both of which have impeded widespread use of FSW in manufacturing. Through this work, major advance has been made toward transforming FSW technology from a “specialty” process to a mainstream materials joining technology to realize its pervasive energy, environmental, and economic benefits across industry.

  13. DNA double strand break repair is enhanced by P53 following induction by DNA damage and is dependent on the C-terminal domain of P53

    International Nuclear Information System (INIS)

    Wei Tang; Powell, Simon N.

    1996-01-01

    Purpose: The tumor suppressor gene p53 can mediate cell cycle arrest or apoptosis in response to DNA damage. Accumulating evidence suggests that it may also directly or indirectly influence the DNA repair machinery. In the present study, we investigated whether p53, induced by DNA damage, could enhance the rejoining of double-strand DNA breaks. Materials and Methods: DNA double-strand breaks (dsb) were made by restriction enzyme digestion of a plasmid, between a promoter and a 'reporter' gene: luciferase (LUC) or chloramphenicol acetyl-transferase (CAT). Linear or circular plasmid DNA (LUC or CAT) was co-transfected with circular β-Gal plasmid (to normalize for uptake) into mouse embryonic fibroblasts genetically matched to be (+/+) or (-/-) for p53. Their ability to rejoin linearized plasmid was measured by the luciferase or CAT activity detected in rescued plasmids. The activity detected in cells transfected with linear plasmid was scored relative to the activity detected in cells transfected with circular plasmid. Results: Ionizing radiation (IR, 2 Gy) enhanced the dsb repair activity in wild type p53 cells; however, p53 null cells lose this effect, indicating that the enhancement of dsb repair was p53-dependent. REF cells with dominant-negative mutant p53 showed a similar induction compared with the parental REF cells with wild-type p53. This ala-143 mutant p53 prevents cell cycle arrest and transactivation of p21 WAF1/cip1) following IR, indicating that the p53-dependent enhancement of DNA repair is distinct from transactivation. Immortalized murine embryonic fibroblasts, 10(1)VasK1 cells, which express p53 cDNA encoding a temperature-sensitive mutant in the DNA sequence specific binding domain (ala135 to val135) with an alternatively spliced C-terminal domain (ASp53: amino-acids 360-381) and, 10(1)Val5 cells, which express the normal spliced p53 (NSp53) with the same temperature-sensitive mutant were compared. It was found that 10(1)VasK1 cells showed no DNA

  14. Development of reconstitution method for surveillance specimens using surface activated joining

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, Terumi; Kaihara, Shoichiro; Yoshida, Kazuo; Sato, Akira [Ishikawajima-Harima Heavy Industries Co. Ltd., Tokyo (Japan); Onizawa, Kunio; Nishiyama, Yutaka; Fukaya, Kiyoshi; Suzuki, Masahide

    1996-03-01

    Evaluation of embrittlement of reactor vessel steel due to irradiation requires surveillance tests. However, many surveillance specimens are necessary for nuclear plants life extension. Therefore, a specimen reconstitution technique has become important to provide the many specimens for continued surveillance. A surface activated joining (SAJ) method has been developed to join various materials together at low temperatures with little deformation, and is useful to bond irradiated specimens. To assess the validity of this method, Charpy impact tests were carried out, and the characteristics caused by heating during joining were measured. The test results showed the Charpy impact values were almost the same as base materials, and surface activated joining reduced heat affected zone to less than 2 mm. (author).

  15. The stem cell secretome and its role in brain repair.

    Science.gov (United States)

    Drago, Denise; Cossetti, Chiara; Iraci, Nunzio; Gaude, Edoardo; Musco, Giovanna; Bachi, Angela; Pluchino, Stefano

    2013-12-01

    Compelling evidence exists that non-haematopoietic stem cells, including mesenchymal (MSCs) and neural/progenitor stem cells (NPCs), exert a substantial beneficial and therapeutic effect after transplantation in experimental central nervous system (CNS) disease models through the secretion of immune modulatory or neurotrophic paracrine factors. This paracrine hypothesis has inspired an alternative outlook on the use of stem cells in regenerative neurology. In this paradigm, significant repair of the injured brain may be achieved by injecting the biologics secreted by stem cells (secretome), rather than implanting stem cells themselves for direct cell replacement. The stem cell secretome (SCS) includes cytokines, chemokines and growth factors, and has gained increasing attention in recent years because of its multiple implications for the repair, restoration or regeneration of injured tissues. Thanks to recent improvements in SCS profiling and manipulation, investigators are now inspired to harness the SCS as a novel alternative therapeutic option that might ensure more efficient outcomes than current stem cell-based therapies for CNS repair. This review discusses the most recent identification of MSC- and NPC-secreted factors, including those that are trafficked within extracellular membrane vesicles (EVs), and reflects on their potential effects on brain repair. It also examines some of the most convincing advances in molecular profiling that have enabled mapping of the SCS. Copyright © 2013 The Authors. Published by Elsevier Masson SAS.. All rights reserved.

  16. Social Pharmacy and Clinical Pharmacy-Joining Forces.

    Science.gov (United States)

    Almarsdottir, Anna Birna; Granas, Anne Gerd

    2015-12-22

    This commentary seeks to define the areas of social pharmacy and clinical pharmacy to uncover what they have in common and what still sets them apart. Common threats and challenges of the two areas are reviewed in order to understand the forces in play. Forces that still keep clinical and social pharmacy apart are university structures, research traditions, and the management of pharmacy services. There are key (but shrinking) differences between clinical and social pharmacy which entail the levels of study within pharmaceutical sciences, the location in which the research is carried out, the choice of research designs and methods, and the theoretical foundations. Common strengths and opportunities are important to know in order to join forces. Finding common ground can be developed in two areas: participating together in multi-disciplinary research, and uniting in a dialogue with internal and external key players in putting forth what is needed for the profession of pharmacy. At the end the question is posed, "What's in a name?" and we argue that it is important to emphasize what unifies the families of clinical pharmacy and social pharmacy for the benefit of both fields, pharmacy in general, and society at large.

  17. Comparative analysis of the practice of China and Russia joining the WTO

    Directory of Open Access Journals (Sweden)

    Anastasia Sergeyevna Loginova

    2015-09-01

    Full Text Available Objective to examine the positive and negative experience of China39s joining the World Trade Organization WTO that joined it on terms similar to the Russian ones for the best understanding of the advantages and disadvantages of joining and building the appropriate policy. Methods the objective of the work has been achieved through the application of both general and specific scientific methods. In particular the analysis and synthesis allowed to study the social and economic context of the state in the framework of joining the WTO the consequences of such membership and to identify the key areas for improvement of this step for Russia. Statistical techniques were used in the process of gathering information about the key economic indicators. The fundamental method was comparativelegal which made it possible to draw parallels in joining the WTO by various states. Results basing on the measures taken by China both before and after joining the WTO as well as evaluation of their implication the conclusion was made about its successful experience. The actions before joining were very effective. In particular they are applicable to the fields of agriculture automotive industry investment policy etc. The measures of nonaddress subsidizing maintaining the socioeconomic stability in the country the export orientation of the economy along with functioning of special economic zones with joint ventures are in the authors39 opinion constructive actions for Russiarsquos adaptation to the WTO. Scientific novelty for the first time the thesis is put forward about the similarity of the Russia and China conditions when joining the WTO. The analysis is made of the complex measures preceding and following the joining. The necessity is grounded of reflecting the China39s positive and negative experiences in the Russian policy. Practical significance the main provisions and conclusions can be used in the research activity in addressing the issues of Russiarsquos

  18. Additive Manufacturing in Offsite Repair of Consumer Electronics

    Science.gov (United States)

    Chekurov, Sergei; Salmi, Mika

    Spare parts for products that are at the end of their life cycles, but still under warranty, are logistically difficult because they are commonly not stored in the central warehouse. These uncommon spare parts occupy valuable space in smaller inventories and take a long time to be transported to the point of need, thus delaying the repair process. This paper proposes that storing the spare parts on a server and producing them with additive manufacturing (AM) on demand can shorten the repair cycle by simplifying the logistics. Introducing AM in the repair supply chain lowers the number of products that need to be reimbursed to the customer due to lengthy repairs, improves the repair statistics of the repair shops, and reduces the number of items that are held in stock. For this paper, the functionality of the concept was verified by reverse engineering a memory cover of a portable computer and laser sintering it from polyamide 12. The additively manufactured component fit well and the computer operated normally after the replacement. The current spare part supply chain model and models with AM machinery located at the repair shop, the centralized spare part provider, and the original equipment manufacturer were provided. The durations of the repair process in the models were compared by simulating two scenarios with the Monte Carlo method. As the biggest improvement, the model with the AM machine in the repair shop reduced the duration of the repair process from 14 days to three days. The result points to the conclusion that placing the machine as close to the need as possible is the best option, if there is enough demand. The spare parts currently compatible with AM are plastic components without strict surface roughness requirements, but more spare parts will become compatible with the development of AM.

  19. Draft environmental statement related to steam generator repair at H.B. Robinson Steam Electric Plant Unit No. 2, (Docket No. 50-261)

    International Nuclear Information System (INIS)

    1983-09-01

    The staff has considered the environmental impacts and economic costs of the proposed steam generator repair at the H.B. Robinson Steam Electric Plant Unit No. 2 along with reasonable alternatives to the proposed action. The staff has concluded that the proposed repair will not significantly affect the quality of the human environment and that there are no preferable alternatives to the proposed action. Furthermore, any impacts from the repair program are outweighted by its benefits

  20. Joining of aluminum sheet and glass fiber reinforced polymer using extruded pins

    Science.gov (United States)

    Conte, Romina; Buhl, Johannes; Ambrogio, Giuseppina; Bambach, Markus

    2018-05-01

    The present contribution proposes a new approach for joining sheet metal and fiber reinforced composites. The joining process draws upon a Friction Stir Forming (FSF) process, which is performed on the metal sheet to produce slender pins. These pins are used to pierce through the composite. Joining is complete by forming a locking head out of the part if the pin sticks out of the composite. Pins of different diameters and lengths were produced from EN AW-1050 material, which were joined to glass fiber reinforced polyamide-6. The strength of the joint has been experimentally tested in order to understand the effect of the process temperature on the pins strength and therefore on the joining. The results demonstrate the feasibility of this new technique, which uses no excess material.

  1. Repair of fractured abutment teeth under pre-existing crowns: An alternative approach

    Directory of Open Access Journals (Sweden)

    Kennedy Mascarenhas

    2013-01-01

    Full Text Available This article describes a technique for repair of abutment tooth which fractured during removal of a provisional restoration before bisque trial. The technique uses plastic templates to fabricate new composite core foundation for the existing crowns. This technique helps the dentist to rebuild the core in a single appointment.

  2. Related issues in repair of bile duct injury and traumatic biliary stricture

    Directory of Open Access Journals (Sweden)

    WANG Shuguang

    2017-02-01

    Full Text Available Inappropriate treatment of bile duct injury and traumatic biliary stricture may cause serious consequences such as recurrent cholangitis, formation of hepatolithiasis, and biliary cirrhosis. This article elaborates on the influencing factors for the effect of the repair of bile duct injury and traumatic biliary stricture, repair principles, timing of repair or reconstruction, and related methods and techniques. It is pointed out that if there is no significant local infection and the bile duct wall defect is <2 cm, end-to-end anastomosis should be used for repair; if the bile duct wall defect is >2 cm, Roux-en-Y hepaticojejunostomy should be used for reconstruction. If the upper wall of the bile duct had a large defect and the lower wall has an integral structure, pedicled umbilical vein graft, pedicled jejunal wall seromuscular flap, or gastric wall seromuscular flap should be used for repair. The patients with severe congestion and edema at the site of injury should be treated with sufficient external drainage of the injured bile duct and then selective repair or reconstruction. Patients with hepatic duct stenosis in the liver lobe or hepatic segments and liver tissue atrophy can be treated with hepalobectomy or segmental hepatectomy. The key to successful repair is exposure and removal of high hilar bile duct stricture, while segmental hepatectomy of the Ⅳb segment can fully expose the left and right hepatic pedicles and help with the incision of the left and right hepatic ducts and secondary hepatic ducts, and therefore, it is a good method for exposing high bile duct stricture.

  3. Pain and convalescence following laparoscopic ventral hernia repair

    DEFF Research Database (Denmark)

    Eriksen, Jens Ravn

    Severe pain is usual after laparoscopic ventral hernia repair (LVHR). Mesh fixation with titanium tacks may play a key role in the development of acute and chronic pain and alternative fixation methods should therefore be investigated. This PhD thesis was based on three studies and aimed too: 1) ...

  4. Testing of self-repairing composite airplane components by use of CAI and the release of the repair chemicals from carefully inserted small tubes

    Science.gov (United States)

    Dry, Carolyn

    2007-04-01

    The research on self repair of airplane components, under an SBIR phase II with Wright Patterson Air Force Base, has investigated the attributes and best end use applications for such a technology. These attributes include issues related to manufacturability, cost, potential benefits such as weight reduction, and cost reduction. The goal of our research has been to develop self-repairing composites with unique strength for air vehicles. Our revolutionary approach involves the autonomous release of repair chemicals from within the composite matrix itself. The repair agents are contained in hollow, structural fibers that are embedded within the matrix. Under stress, the composite senses external environmental factors and reacts by releasing the repair agents from within the hollow vessels. This autonomous response occurs wherever and whenever cracking, debonding or other matrix damage transpires. Superior performance over the life of the composite is achieved through this self-repairing mechanism. The advantages to the military would be safely executed missions, fewer repairs and eventually lighter vehicles. In particular the research has addressed the issues by correlating the impact of the various factors, such as 1) delivery vessel placement, shape/size and effect on composite strength, chemicals released and their effect on the matrix, release trigger and efficacy and any impact on matrix properties 2) impact of composite processing methods that involve heat and pressure on the repair vessels. Our self repairing system can be processed at temperatures of 300-350F, repairs in less than 30 seconds and does not damage the composite by repair fiber insertion or chemical release. Scaling up and manufacture of components has revealed that anticipating potential problems allowed us to avoid those associated with processing temperatures and pressures. The presentation will focus on compression after impact testing and the placement of repair fibers/tubes into prepreg

  5. Recent progress in micro and nano-joining

    Energy Technology Data Exchange (ETDEWEB)

    Zhou, Y; Hu, A; Khan, M I; Wu, W; Tam, B; Yavuz, M [Centre for Advanced Materials Joining Department of Mechanical and Mechatronics Engineering, University of Waterloo 200 University Avenue West, Waterloo, N2L 3G1 (Canada)], E-mail: nzhou@uwaterloo.ca

    2009-05-01

    Micro and nano-joining has been identified as a key enabling technology in the construction of micromechanical and microelectronic devices. The current article reviews recent progress in micro and nano-joining. In particular, laser micro-welding (LMW) of crossed 316 LVM stainless steel (SS) wire was compared to conventional resistance micro-welding (RMW) and was successfully employed in welding a Pt-Ir /SS dissimilar combination. Welding of Au nanoparticles was realized using femtosecond laser irradiation and its application in the surface enhanced Raman spectroscopy was investigated. Brazing between carbon nanotube (CNT) bundles and Ni electrodes was attained in vacuum, resulting in the development of a novel CNT filament of incandescent lamps.

  6. RPA accumulation during class switch recombination represents 5'-3' DNA-end resection during the S-G2/M phase of the cell cycle.

    Science.gov (United States)

    Yamane, Arito; Robbiani, Davide F; Resch, Wolfgang; Bothmer, Anne; Nakahashi, Hirotaka; Oliveira, Thiago; Rommel, Philipp C; Brown, Eric J; Nussenzweig, Andre; Nussenzweig, Michel C; Casellas, Rafael

    2013-01-31

    Activation-induced cytidine deaminase (AID) promotes chromosomal translocations by inducing DNA double-strand breaks (DSBs) at immunoglobulin (Ig) genes and oncogenes in the G1 phase. RPA is a single-stranded DNA (ssDNA)-binding protein that associates with resected DSBs in the S phase and facilitates the assembly of factors involved in homologous repair (HR), such as Rad51. Notably, RPA deposition also marks sites of AID-mediated damage, but its role in Ig gene recombination remains unclear. Here, we demonstrate that RPA associates asymmetrically with resected ssDNA in response to lesions created by AID, recombination-activating genes (RAG), or other nucleases. Small amounts of RPA are deposited at AID targets in G1 in an ATM-dependent manner. In contrast, recruitment in the S-G2/M phase is extensive, ATM independent, and associated with Rad51 accumulation. In the S-G2/M phase, RPA increases in nonhomologous-end-joining-deficient lymphocytes, where there is more extensive DNA-end resection. Thus, most RPA recruitment during class switch recombination represents salvage of unrepaired breaks by homology-based pathways during the S-G2/M phase of the cell cycle. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  7. Autophosphorylation of DNA-PKCS regulates its dynamics at DNA double-strand breaks.

    Science.gov (United States)

    Uematsu, Naoya; Weterings, Eric; Yano, Ken-ichi; Morotomi-Yano, Keiko; Jakob, Burkhard; Taucher-Scholz, Gisela; Mari, Pierre-Olivier; van Gent, Dik C; Chen, Benjamin P C; Chen, David J

    2007-04-23

    The DNA-dependent protein kinase catalytic subunit (DNA-PK(CS)) plays an important role during the repair of DNA double-strand breaks (DSBs). It is recruited to DNA ends in the early stages of the nonhomologous end-joining (NHEJ) process, which mediates DSB repair. To study DNA-PK(CS) recruitment in vivo, we used a laser system to introduce DSBs in a specified region of the cell nucleus. We show that DNA-PK(CS) accumulates at DSB sites in a Ku80-dependent manner, and that neither the kinase activity nor the phosphorylation status of DNA-PK(CS) influences its initial accumulation. However, impairment of both of these functions results in deficient DSB repair and the maintained presence of DNA-PK(CS) at unrepaired DSBs. The use of photobleaching techniques allowed us to determine that the kinase activity and phosphorylation status of DNA-PK(CS) influence the stability of its binding to DNA ends. We suggest a model in which DNA-PK(CS) phosphorylation/autophosphorylation facilitates NHEJ by destabilizing the interaction of DNA-PK(CS) with the DNA ends.

  8. Effects of Smoking on Hand Tendon Repair: Scientific Study & Literature Review

    OpenAIRE

    Samona J; Samona S; Gilin M; Carwile D; Persons S; AG Dass

    2017-01-01

    Purpose: The clinical effect of smoking on after tendon repair in smokers vs non-smokers was investigated, via exploring effects on flexor and tendon repair, and incidence of wound complications. Methods: Retrospective review of patients treated by 1 orthopedic surgeon, fellowship trained in hand surgery, and 3 occupational therapists working together in 1 physical therapy institution. 56 patients (20 smokers and 36 non-smokers) with acute traumatic tendon lacerations, fixed via direct (end-t...

  9. DNA repair , cell repair and radiosensitivity

    International Nuclear Information System (INIS)

    Zhestyanikov, V.D.

    1983-01-01

    Data obtained in laboratory of radiation cytology and literature data testifying to a considerable role of DNA repair in cell sensitivity to radiation and chemical DNA-tropic agents have been considered. Data pointing to the probability of contribution of inducible repair of DNA into plant cells sensitivity to X-rays are obtained. Certain violations of DNA repair do not result in the increase of radiosensitivity. It is assumed that in the cases unknown mechanisms of DNA repair operate

  10. The Role of Transesophageal Echocardiography in Endovascular Repair of Traumatic Aortic Transection

    Directory of Open Access Journals (Sweden)

    Swathy B

    2014-09-01

    Full Text Available Traumatic rupture of the thoracic aorta is a leading cause of death, following major blunt trauma, and endovascular repair has evolved as a viable alternative to open repair. This report highlights the role of transesophageal echocardiography as a valuable imaging tool for locating the exact position of the lesion, guiding placement of the endograft, detecting leaks around it and supplementing information derived from angiography during endograft deployment.

  11. Laparoscopic repair for vesicouterine fistulae

    Directory of Open Access Journals (Sweden)

    Rafael A. Maioli

    2015-10-01

    . Discussion: Laparoscopy has advantages over open surgery in that it is associated with less pain, shorter length of hospital stay, better cosmesis, quicker recovery, and equal efficacy. Although cases of VUF are rarely noted, the laparoscopic skill obtained through other urological procedures suggest, that laparoscopic repair may be the procedure of choice for such cases (2. The reported operative time for the laparoscopic repair of VUF in the literature varies between 140 and 220 min (3. However, laparoscopic techniques should be considered as a mode of abdominal access and should not influence the method of surgical repair. Surgical success should depend on the adherence to good technique rather than the approach. Hence, this method appears to be a viable alternative for surgeons experienced with laparoscopic suturing techniques. Conclusion: Laparoscopic repair appears to be a viable alternative for surgeons experienced with laparoscopic suturing techniques.

  12. Role of DNA repair in repair of cytogenetic damages. Slowly repaired DNA injuries involved in cytogenetic damages repair

    International Nuclear Information System (INIS)

    Zaichkina, S.I.; Rozanova, O.M.; Aptikaev, G.F.; Ganassi, E.Eh.

    1989-01-01

    Caffeine was used to study the kinetics of cytogenetic damages repair in Chinese hamster fibroblasts. Its half-time (90 min) was shown to correlate with that of repair of slowly repaired DNA damages. The caffeine-induced increase in the number of irreparable DNA damages, attributed to inhibition of double-strand break repair, is in a quantitative correlation with the effect of the cytogenetic damage modification

  13. [Preliminary investigation of treatment of ulnar nerve defect by end-to-side neurorrhaphy].

    Science.gov (United States)

    Luo, Y; Wang, T; Fang, H

    1997-11-01

    In the repair of the defect of peripheral nerve, it was necessary to find an operative method with excellent therapeutic effect but simple technique. Based on the experimental study, one case of old injury of the ulnar nerve was treated by end-to-side neurorraphy with the intact median nerve. In this case the nerve defect was over 3 cm and unable to be sutured directly. The patient was followed up for fourteen months after the operation. The recovery of the sensation and the myodynamia was evaluated. The results showed that: the sensation and the motor function innervated by ulnar nerve were recovered. The function of the hand was almost recovered to be normal. It was proved that the end-to-side neurorraphy between the distal stump with the intact median nerve to repair the defect of the ulnar nerve was a new operative procedure for nerve repair. Clinically it had good effect with little operative difficulty. This would give a bright prospect to repair of peripheral nerve defect in the future.

  14. Cyclic Testing of the 6-Strand Tang and Modified Lim-Tsai Flexor Tendon Repair Techniques.

    Science.gov (United States)

    Kang, Gavrielle Hui-Ying; Wong, Yoke-Rung; Lim, Rebecca Qian-Ru; Loke, Austin Mun-Kitt; Tay, Shian-Chao

    2018-03-01

    In this study, we compared the Tang repair technique with the 6-strand modified Lim-Tsai repair technique under cyclic testing conditions. Twenty fresh-frozen porcine flexor tendons were randomized into 2 groups for repair with either the modified Lim-Tsai or the Tang technique using Supramid 4-0 core sutures and Ethilon 6-0 epitendinous running suture. The repaired tendons were subjected to 2 stage cyclic loading. The survival rate and gap formation at the repair site were recorded. Tendons repaired by the Tang technique achieved an 80% survival rate. None of the modified Lim-Tsai repairs survived. The mean gap formed at the end of 1000 cycles was 1.09 mm in the Tang repairs compared with 4.15 mm in the modified Lim-Tsai repairs. The Tang repair is biomechanically stronger than the modified Lim-Tsai repair under cyclic loading. The Tang repair technique may exhibit a higher tolerance for active mobilization after surgery with less propensity for gap formation. Copyright © 2018 American Society for Surgery of the Hand. Published by Elsevier Inc. All rights reserved.

  15. Zinc finger nuclease-mediated precision genome editing of an endogenous gene in hexaploid bread wheat (Triticum aestivum) using a DNA repair template.

    Science.gov (United States)

    Ran, Yidong; Patron, Nicola; Kay, Pippa; Wong, Debbie; Buchanan, Margaret; Cao, Ying-Ying; Sawbridge, Tim; Davies, John P; Mason, John; Webb, Steven R; Spangenberg, German; Ainley, William M; Walsh, Terence A; Hayden, Matthew J

    2018-05-07

    Sequence-specific nucleases have been used to engineer targeted genome modifications in various plants. While targeted gene knockouts resulting in loss of function have been reported with relatively high rates of success, targeted gene editing using an exogenously supplied DNA repair template and site-specific transgene integration has been more challenging. Here, we report the first application of zinc finger nuclease (ZFN)-mediated, nonhomologous end-joining (NHEJ)-directed editing of a native gene in allohexaploid bread wheat to introduce, via a supplied DNA repair template, a specific single amino acid change into the coding sequence of acetohydroxyacid synthase (AHAS) to confer resistance to imidazolinone herbicides. We recovered edited wheat plants having the targeted amino acid modification in one or more AHAS homoalleles via direct selection for resistance to imazamox, an AHAS-inhibiting imidazolinone herbicide. Using a cotransformation strategy based on chemical selection for an exogenous marker, we achieved a 1.2% recovery rate of edited plants having the desired amino acid change and a 2.9% recovery of plants with targeted mutations at the AHAS locus resulting in a loss-of-function gene knockout. The latter results demonstrate a broadly applicable approach to introduce targeted modifications into native genes for nonselectable traits. All ZFN-mediated changes were faithfully transmitted to the next generation. © 2018 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

  16. Joining of Dissimilar alloy Sheets (Al 6063&AISI 304 during Resistance Spot Welding Process: A Feasibility Study for Automotive industry

    Directory of Open Access Journals (Sweden)

    Reddy Sreenivasulu

    2014-12-01

    Full Text Available Present design trends in automotive manufacture have shifted emphasis to alternative lightweight materials in order to achieve higher fuel efficiency and to bring down vehicle emission. Although some other joining techniques are more and more being used, spot welding still remains the primary joining method in automobile manufacturing so far. Spot welds for automotive applications should have a sufficiently large diameter, so that nugget pullout mode is the dominant failure mode. Interfacial mode is unacceptable due to its low load carrying and energy absorption capability. Strength tests with different static loading were performed in, to reveal the failure mechanisms for the lap-shear geometry and the cross-tension geometry. Based on the literature survey performed, venture into this work was amply motivated by the fact that a little research work has been conducted to joining of dissimilar materials like non ferrous to ferrous. Most of the research works concentrated on joining of different materials like steel to steel or aluminium alloy to aluminium alloy by resistance spot welding. In this work, an experimental study on the resistance spot weldability of aluminium alloy (Al 6063 and austenitic stainless steel (AISI304 sheets, which are lap joined by using a pedestal type resistance spot welding machine. Welding was conducted using a 45-deg truncated cone copper electrode with 10-mm face diameter. The weld nugget diameter, force estimation under lap shear test and T – peel test were investigated using digital type tensometer attached with capacitive displacement transducer (Mikrotech, Bangalore, Model: METM2000ER1. The results shows that joining of Al 6063 and AISI 304 thin sheets by RSW method are feasible for automotive structural joints where the loads are below 1000N act on them, it is observed that by increasing the spots per unit length, then the joint with standing strength to oppose failure is also increased linearly incase of

  17. Concordant and opposite roles of DNA-PK and the "facilitator of chromatin transcription" (FACT in DNA repair, apoptosis and necrosis after cisplatin

    Directory of Open Access Journals (Sweden)

    Calkins Anne S

    2011-06-01

    Full Text Available Abstract Background Platinum-containing chemotherapy produces specific DNA damage and is used to treat several human solid tumors. Tumors initially sensitive to platinum-based drugs frequently become resistant. Inhibition of DNA repair is a potential strategy to enhance cisplatin effectiveness. After cisplatin treatment, a balance between repair and apoptosis determines whether cancer cells proliferate or die. DNA-dependent protein kinase (DNA-PK binds to DNA double strand breaks (DSBs through its Ku subunits and initiates non-homologous end joining. Inhibition of DNA-PK sensitizes cancer cells to cisplatin killing. The goal of this study is to elucidate the mechanism underlying the effects of DNA-PK on cisplatin sensitivity. Results Silencing the expression of the catalytic subunit of DNA-PK (DNA-PKcs increased sensitivity to cisplatin and decreased the appearance of γH2AX after cisplatin treatment. We purified DNA-PK by its Ku86 subunit and identified interactors by tandem mass spectrometry before and after cisplatin treatment. The structure specific recognition protein 1 (SSRP1, Spt16 and γH2AX appeared in the Ku86 complex 5 hours after cisplatin treatment. SSRP1 and Spt16 form the facilitator of chromatin transcription (FACT. The cisplatin-induced association of FACT with Ku86 and γH2AX was abrogated by DNase treatment. In living cells, SSRP1 and Ku86 were recruited at sites of DSBs induced by laser beams. Silencing SSRP1 expression increased sensitivity to cisplatin and decreased γH2AX appearance. However, while silencing SSRP1 in cisplatin-treated cells increased both apoptosis and necrosis, DNA-PKcs silencing, in contrast, favored necrosis over apoptosis. Conclusions DNA-PK and FACT both play roles in DNA repair. Therefore both are putative targets for therapeutic inhibition. Since DNA-PK regulates apoptosis, silencing DNA-PKcs redirects cells treated with cisplatin toward necrosis. Silencing FACT however, allows both apoptosis and

  18. Salvage Flexor Hallucis Longus Transfer for a Failed Achilles Repair: Endoscopic Technique.

    Science.gov (United States)

    Gonçalves, Sérgio; Caetano, Rubén; Corte-Real, Nuno

    2015-10-01

    Flexor hallucis longus (FHL) transfer is a well-established treatment option in failed Achilles tendon (AT) repair and has been routinely performed as an open procedure. We detail the surgical steps needed to perform an arthroscopic transfer of the FHL for a chronic AT rupture. The FHL tendon is harvested as it enters in its tunnel beneath the sustentaculum tali; a tunnel is then drilled in the calcaneus as near to the AT footprint as possible. By use of a suture-passing device, the free end of the FHL is advanced to the plantar aspect of the foot. After adequate tension is applied to the construct, the tendon is fixed in place with an interference screw in an inside-out fashion. This minimally invasive approach is a safe and valid alternative to classic open procedures with the obvious advantages of preserving the soft-tissue envelope and using a biologically intact tendon.

  19. Cancer and non-cancer risk at low doses of radiation: biological basis of radiation-environment interplay

    International Nuclear Information System (INIS)

    Sasaki, Masao S.

    2013-01-01

    Cancer and non-cancer risk at low doses of ionizing radiation remains poorly defined due to ambiguity at low doses caused by limitations in statistical power and information available on interplay with environment. To deal with these problems, a novel non-parametric statistics was developed based on artificial neural networks theorem and applied to cancer and non-cancer risk in A-bomb survivors. The analysis revealed several unique features at low doses that could not be accounted for by nominal radiation dose alone. They include (1) threshold that varies with organ, gender and age, including cardiovascular diseases, (2) prevalence of infectious diseases, and (3) suppression of pathogenesis of HTLV1. The threshold is unique as it is manifested as negative excess relative risk, a reduction of spontaneous rate at low doses. The response is consistent with currently emerging laboratory data on DNA double-strand break (DSB) repair pathway choice and its sustainability as epigenetic memory in accordance with histone code theory. In response to DSB, of radiation or DNA replication arrest origin, distinct and competitively operating repair pathways are instigated. Activation by low doses of restitution-directed canonical non-homologous end-joining (C-NHEJ) suppresses both error-prone alternative end-joining (Alt-NHEJ) and homologous recombination (HR). The latter two present major pathways to mutagenesis at stalled replication folk associated with endogenous and exogenous genotoxin such as tobacco smoke metabolites and AID-associated somatic hypermutation and class switch recombination in Ig gene. Suppression of these error-prone pathways by low doses of low LET radiation is consistent with the reduction of cancer occurrence by environmental genotoxin, immunodiversity and stable integration of retrovirus DNA, providing a significant modulator of dose linearity at low doses. Whole picture may bring about a new landscape of cancer and non-cancer molecular epidemiology which

  20. Laser microbeams for DNA damage induction, optical tweezers for the search on blood pressure relaxing drugs: contributions to ageing research

    Science.gov (United States)

    Grigaravicius, P.; Monajembashi, S.; Hoffmann, M.; Altenberg, B.; Greulich, K. O.

    2009-08-01

    One essential cause of human ageing is the accumulation of DNA damages during lifetime. Experimental studies require quantitative induction of damages and techniques to visualize the subsequent DNA repair. A new technique, the "immuno fluorescent comet assay", is used to directly visualize DNA damages in the microscope. Using DNA repair proteins fluorescently labeled with green fluorescent protein, it could be shown that the repair of the most dangerous DNA double strand breaks starts with the inaccurate "non homologous end joining" pathway and only after 1 - 1 ½ minutes may switch to the more accurate "homologous recombination repair". One might suggest investigating whether centenarians use "homologous recombination repair" differently from those ageing at earlier years and speculate whether it is possible, for example by nutrition, to shift DNA repair to a better use of the error free pathway and thus promote healthy ageing. As a complementary technique optical tweezers, and particularly its variant "erythrocyte mediated force application", is used to simulate the effects of blood pressure on HUVEC cells representing the inner lining of human blood vessels. Stimulating one cell induces in the whole neighbourhood waves of calcium and nitric oxide, known to relax blood vessels. NIFEDIPINE and AMLODIPINE, both used as drugs in the therapy of high blood pressure, primarily a disease of the elderly, prolong the availability of nitric oxide. This partially explains their mode of action. In contrast, VERAPAMILE, also a blood pressure reducing drug, does not show this effect, indicating that obviously an alternative mechanism must be responsible for vessel relaxation.