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Sample records for replicating previous results

  1. Effects of 60 Hz electromagnetic fields on early growth in three plant species and a replication of previous results

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    Davis, M.S. [Univ. of Sunderland (United Kingdom). Ecology Centre

    1996-05-01

    In an attempt to replicate the findings of Smith et al., seeds of Raphanus sativus L. (radish), Sinapsis alba L. (mustard), and Hordeum vulgare L. (barley) were grown for between 9 and 21 days in continuous electromagnetic fields (EMFs) at ion-cyclotron resonance conditions for stimulation of Ca{sup 2+} (B{sub H} = 78.3 {micro}T, B{sub HAC} = 40 {micro}T peak-peak at 60 Hz, B{sub v} = 0). On harvesting, radish showed results similar to those of Smith et al. Dry stem weight and plant height were both significantly greater (Mann-Whitney tests, Ps < 0.05) in EMF-exposed plants than in control plants in each EMF experiment. Wet root weight was significantly greater in EMF-exposed plants in two out of three experiments, as were dry leaf weight, dry whole weight, and stem diameter. Dry root weight, wet leaf weight, and wet whole weight were significantly greater in EMF-exposed plants in one of three experiments. All significant differences indicated an increase in weight or size in the EMF-exposed plants. In each of the sham experiments, no differences between exposed and control plants were evident. Mustard plants failed to respond to the EMFs in any of the plant parameters measured. In one experiment, barley similarly failed to respond; but in another showed significantly greater wet root weight and significantly smaller stem diameter and dry seed weight at the end of the experiment in exposed plants compared to control plants. Although these results give no clue about the underlying bioelectromagnetic mechanism, they demonstrate that, at least for one EMF-sensitive biosystem, results can be independently replicated in another laboratory. Such replication is crucial in establishing the validity of bioelectromagnetic science.

  2. Effects of 60 Hz electromagnetic fields on early growth in three plant species and a replication of previous results.

    Science.gov (United States)

    Davies, M S

    1996-01-01

    In an attempt to replicate the findings of Smith et al., seeds of Raphanus sativus L. (radish), Sinapsis alba L. (mustard), and Hordeum vulgare L. (barley) were grown for between 9 and 21 days in continuous electromagnetic fields (EMFs) at "ion-cyclotron resonance" conditions for stimulation of Ca(2+) (B(H) = 78.3 mu T, B(HAC) = 40 mu T peak-peak at 60 Hz, B(V) = 0). On harvesting, radish showed results similar to those of Smith et al. Dry stem weight and plant height were both significantly greater (Mann-Whitney tests, Ps < 0.05) in EMF-exposed plants than in control plants in each EMF experiment. Wet root weight was significantly greater in EMF-exposed plants in two out of three experiments, as were dry leaf weight, dry whole weight, and stem diameter. Dry root weight, wet leaf weight, and wet whole weight were significantly greater in EMF-exposed plants in one of three experiments. All significant differences indicated an increase in weight or size in the EMF-exposed plants. In each of the sham experiments, no differences between exposed and control plants were evident. Mustard plants failed to respond to the EMFs in any of the plant parameters measured. In one experiment, barley similarly failed to respond; but in another showed significantly greater wet root weight and significantly smaller stem diameter and dry seed weight at the end of the experiment in exposed plants compared to control plants. Although these results give no clue about the underlying bioelectromagnetic mechanism, they demonstrate that, at least for one EMF-sensitive biosystem, results can be independently replicated in another laboratory. Such replication is crucial in establishing the validity of bioelectromagnetic science.

  3. The pre-engagement cohabitation effect: a replication and extension of previous findings.

    Science.gov (United States)

    Rhoades, Galena K; Stanley, Scott M; Markman, Howard J

    2009-02-01

    Using a random telephone survey of men and women married within the past 10 years (N = 1,050), the current study replicated previous findings regarding the timing of engagement and the premarital cohabitation effect (see Kline et al., 2004). Those who cohabited before engagement (43.1%) reported lower marital satisfaction, dedication, and confidence as well as more negative communication and greater potential for divorce than those who cohabited only after engagement (16.4%) or not at all until marriage (40.5%). These differences were generally small, but could not be accounted for by length of marriage or by variables often associated with selection into cohabitation (i.e., age, income, education, and religiousness). Similar results were found in a subsample of individuals who cohabited only with the current spouse. There were no significant differences between those who cohabited after engagement and not at all before marriage, supporting a pre-engagement, but not a premarital cohabitation effect.

  4. Abnormal Parietal Brain Function in ADHD: Replication and Extension of Previous EEG Beta Asymmetry Findings

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    T. Sigi eHale

    2014-07-01

    Full Text Available Background: Abundant work indicates ADHD abnormal posterior brain structure and function, including abnormal structural and functional asymmetries and reduced corpus callosum size. However, this literature has attracted considerably less research interest than fronto-striatal findings. Objective: To help address this imbalance, the current study replicates and extends our previous work showing abnormal parietal brain function in ADHD adults during the Conner’s continuous performance test (CPT. Method: Our previous study found that ADHD adults had increased rightward EEG beta (16-21 Hz asymmetry in inferior parietal brain regions during the CPT (p=.00001, and that this metric exhibited a lack of normal correlation (i.e., observed in controls with beta asymmetry at temporal-parietal regions. We re-tested these effects in a new ADHD sample, and with both new and old samples combined. We additionally examined: a EEG asymmetry in multiple frequency bands, b unilateral effects for all asymmetry findings, and c the association between EEG asymmetry and a battery of cognitive tests. Results: We replicated our original findings, again demonstrating abnormal rightward inferior parietal beta asymmetry in adults with ADHD during the CPT, and again this metric exhibited abnormal reduced correlation to temporal-parietal beta asymmetry. Novel analyses also demonstrated a broader pattern of rightward beta and theta asymmetry across inferior, superior, and temporal-parietal brain regions, and showed that rightward parietal asymmetry in ADHD was atypically associated with multiple cognitive tests. Conclusion: Abnormal increased rightward parietal EEG beta asymmetry is an important feature of ADHD. We speculate that this phenotype may occur with any form of impaired capacity for top-down task-directed control over sensory encoding functions, and that it may reflect associated increases of attentional shifting and compensatory sustained/selective attention.

  5. Convergence in the Bilingual Lexicon: A Pre-registered Replication of Previous Studies

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    White, Anne; Malt, Barbara C.; Storms, Gert

    2017-01-01

    Naming patterns of bilinguals have been found to converge and form a new intermediate language system from elements of both the bilinguals’ languages. This converged naming pattern differs from the monolingual naming patterns of both a bilingual’s languages. We conducted a pre-registered replication study of experiments addressing the question whether there is a convergence between a bilingual’s both lexicons. The replication used an enlarged set of stimuli of common household containers, providing generalizability, and more reliable representations of the semantic domain. Both an analysis at the group-level and at the individual level of the correlations between naming patterns reject the two-pattern hypothesis that poses that bilinguals use two monolingual-like naming patterns, one for each of their two languages. However, the results of the original study and the replication comply with the one-pattern hypothesis, which poses that bilinguals converge the naming patterns of their two languages and form a compromise. Since this convergence is only partial the naming pattern in bilinguals corresponds to a moderate version of the one-pattern hypothesis. These findings are further confirmed by a representation of the semantic domain in a multidimensional space and the finding of shorter distances between bilingual category centers than monolingual category centers in this multidimensional space both in the original and in the replication study. PMID:28167921

  6. Result Analysis and Benefits of Detecting Replicate Documents Using MD5 Hash Function

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    Pushpendra Singh Tomar

    2011-12-01

    Full Text Available The definition of what constitutes a replicate has somewhat different interpretations. For instance, some define a replicate as having the exact syntactic terms and sequence, whether having formatting differences or not. In effect, there are either no difference or only formatting differences and the contents of the data are exactly the same. In any case, data replication happens all the time. In large data warehouses, data replication is an inevitable phenomenon as millions of data are gathered at very short intervals. In this paper we provide a detail result analysis on the basis of our approach and the previous one.

  7. Result Analysis and Benefits of Detecting Replicate Documents Using MD5 Hash Function

    Directory of Open Access Journals (Sweden)

    Mr. Pushpendra Singh Tomar

    2011-09-01

    Full Text Available The definition of what constitutes a replicate has somewhat different interpretations. For instance, some define a replicate as having the exact syntactic terms and sequence, whether having formatting differences or not. In effect, there are either no difference or only formatting differences and the contents of the data are exactly the same. In any case, data replication happens all the time. In large data warehouses, data replication is an inevitable phenomenon as millions of data are gathered at very short intervals. In this paper we provide a detail result analysis on the basis of our approach and the previous one.

  8. Effective Teaching and Learning Environments and Principal Self-Efficacy in Oklahoma: Replication of a Previous Study

    Science.gov (United States)

    Berry, Kathryn

    2013-01-01

    The purpose of this study was to replicate a previous study by Smith et al. (2006) that explored principal self-efficacy beliefs for facilitating effective instructional environments at their schools. There has been limited research conducted on principal's self-efficacy, and the studies that have been completed on the topic have not been…

  9. Loss of maintenance DNA methylation results in abnormal DNA origin firing during DNA replication.

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    Haruta, Mayumi; Shimada, Midori; Nishiyama, Atsuya; Johmura, Yoshikazu; Le Tallec, Benoît; Debatisse, Michelle; Nakanishi, Makoto

    2016-01-22

    The mammalian maintenance methyltransferase DNMT1 [DNA (cytosine-5-)-methyltransferase 1] mediates the inheritance of the DNA methylation pattern during replication. Previous studies have shown that depletion of DNMT1 causes a severe growth defect and apoptosis in differentiated cells. However, the detailed mechanisms behind this phenomenon remain poorly understood. Here we show that conditional ablation of Dnmt1 in murine embryonic fibroblasts (MEFs) resulted in an aberrant DNA replication program showing an accumulation of late-S phase replication and causing severely defective growth. Furthermore, we found that the catalytic activity and replication focus targeting sequence of DNMT1 are required for a proper DNA replication program. Taken together, our findings suggest that the maintenance of DNA methylation by DNMT1 plays a critical role in proper regulation of DNA replication in mammalian cells.

  10. Loss of maintenance DNA methylation results in abnormal DNA origin firing during DNA replication

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    Haruta, Mayumi [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Shimada, Midori, E-mail: midorism@med.nagoya-cu.ac.jp [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Nishiyama, Atsuya; Johmura, Yoshikazu [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan); Le Tallec, Benoît; Debatisse, Michelle [Institut Curie, Centre de Recherche, 26 rue d’Ulm, CNRS UMR 3244, 75248 ParisCedex 05 (France); Nakanishi, Makoto, E-mail: mkt-naka@med.nagoya-cu.ac.jp [Department of Cell Biology, Graduate School of Medical Sciences, Nagoya City University, 1 Kawasumi, Mizuho-cho, Mizuho-ku, Nagoya 467-8601 (Japan)

    2016-01-22

    The mammalian maintenance methyltransferase DNMT1 [DNA (cytosine-5-)-methyltransferase 1] mediates the inheritance of the DNA methylation pattern during replication. Previous studies have shown that depletion of DNMT1 causes a severe growth defect and apoptosis in differentiated cells. However, the detailed mechanisms behind this phenomenon remain poorly understood. Here we show that conditional ablation of Dnmt1 in murine embryonic fibroblasts (MEFs) resulted in an aberrant DNA replication program showing an accumulation of late-S phase replication and causing severely defective growth. Furthermore, we found that the catalytic activity and replication focus targeting sequence of DNMT1 are required for a proper DNA replication program. Taken together, our findings suggest that the maintenance of DNA methylation by DNMT1 plays a critical role in proper regulation of DNA replication in mammalian cells. - Highlights: • DNMT1 depletion results in an abnormal DNA replication program. • Aberrant DNA replication is independent of the DNA damage checkpoint in DNMT1cKO. • DNMT1 catalytic activity and RFT domain are required for proper DNA replication. • DNMT1 catalytic activity and RFT domain are required for cell proliferation.

  11. Training Cognitive Functioning in the Elderly--Inability to Replicate Previous Findings.

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    Papalia-Finlay, Diane; And Others

    1980-01-01

    Elderly women volunteers were given a pretest battery of conservation tasks. Conservation scores were the highest yet recorded by elderly participants; consequently, training was not implemented. Results suggest that advanced chronological age does not guarantee poor conservation performance. (Author)

  12. Previous heat shock treatment inhibits Mayaro virus replication in human lung adenocarcinoma (A549) cells.

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    Virgilio, P L; Godinho-Netto, M C; Carvalho Mda, G

    1997-01-01

    Human lung adenocarcinoma cells (A549) were submitted to mild or severe heat shock (42 degrees C or 44 degrees C) for 1 h, while another group of cells was double-heat-shocked (submitted to 42 degrees C for 1 h, returned to 37 degrees C for 3 h, then exposed to 44 degrees C for 1 h). After each heat treatment, the cells were infected with Mayaro virus for 24 h and incubated at 37 degrees C. The results showed that the double-heat-shocked thermotolerant cells exhibited a 10(4)-fold virus titre inhibition, despite the recovery of protein synthesis and original morphology 24 h post-infection. In contrast, cells submitted to mild or severe heat shock exhibited weaker inhibition of Mayaro virus titre (10(2)-fold). The mildly heat-shocked cells also presented a full recovery in protein synthesis, which was not observed in severely heat-shocked cells. These results indicate that exposure of A549 cells to a mild or to a double heat shock treatment before Mayaro virus infection induces an antiviral state.

  13. A Signature of Genomic Instability Resulting from Deficient Replication Licensing

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    Qin, Maochun; Wang, Jianmin; Kunnev, Dimiter; Freeland, Amy

    2017-01-01

    Insufficient licensing of DNA replication origins has been shown to result in genome instability, stem cell deficiency, and cancers. However, it is unclear whether the DNA damage resulting from deficient replication licensing occurs generally or if specific sites are preferentially affected. To map locations of ongoing DNA damage in vivo, the DNAs present in red blood cell micronuclei were sequenced. Many micronuclei are the product of DNA breaks that leave acentromeric remnants that failed to segregate during mitosis and should reflect the locations of breaks. To validate the approach we show that micronuclear sequences identify known common fragile sites under conditions that induce breaks at these locations (hydroxyurea). In MCM2 deficient mice a different set of preferred breakage sites is identified that includes the tumor suppressor gene Tcf3, which is known to contribute to T-lymphocytic leukemias that arise in these mice, and the 45S rRNA gene repeats. PMID:28045896

  14. Panero et al. (2016): Failure to replicate methods caused the failure to replicate results.

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    Kidd, David Comer; Castano, Emanuele

    2017-03-01

    Contrary to Kidd and Castano (2013), Panero et al. (2016) fail to find that reading literary fiction improves performance on an advanced test of theory of mind (ToM), the Reading the Mind in the Eyes Test. However, this commentary shows that the findings presented in Panero et al. (2016) are not reliable due to two striking threats to the internal validity of their studies that were not clearly disclosed or discussed in the manuscript or supplementary materials. First, no effective strategy was implemented to ensure that participants read their assigned texts, and examination of the data revealed many participants whose reading times indicate that they were not exposed to the manipulation. Second, further examination shows that two of the largest studies contributing to Panero et al. (2016) are not valid experiments due to a clear failure of random assignment to conditions. These threats to experimental internal validity make the conclusions presented in Panero et al. (2016) untenable. After removing cases in which participants were not exposed to the manipulation and the data from the two studies without random assignment, an analysis reveals that reading literary fiction improves ToM compared to reading popular genre fiction. This result is consistent with prior studies and indicates that a failure to carefully replicate the methods of Kidd and Castano (2013) led to the failure to replicate Kidd and Castano's (2013) results. (PsycINFO Database Record

  15. Break-seq reveals hydroxyurea-induced chromosome fragility as a result of unscheduled conflict between DNA replication and transcription.

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    Hoffman, Elizabeth A; McCulley, Andrew; Haarer, Brian; Arnak, Remigiusz; Feng, Wenyi

    2015-03-01

    We have previously demonstrated that in Saccharomyces cerevisiae replication, checkpoint inactivation via a mec1 mutation leads to chromosome breakage at replication forks initiated from virtually all origins after transient exposure to hydroxyurea (HU), an inhibitor of ribonucleotide reductase. Here we sought to determine whether all replication forks containing single-stranded DNA gaps have equal probability of producing double-strand breaks (DSBs) when cells attempt to recover from HU exposure. We devised a new methodology, Break-seq, that combines our previously described DSB labeling with next generation sequencing to map chromosome breaks with improved sensitivity and resolution. We show that DSBs preferentially occur at genes transcriptionally induced by HU. Notably, different subsets of the HU-induced genes produced DSBs in MEC1 and mec1 cells as replication forks traversed a greater distance in MEC1 cells than in mec1 cells during recovery from HU. Specifically, while MEC1 cells exhibited chromosome breakage at stress-response transcription factors, mec1 cells predominantly suffered chromosome breakage at transporter genes, many of which are the substrates of those transcription factors. We propose that HU-induced chromosome fragility arises at higher frequency near HU-induced genes as a result of destabilized replication forks encountering transcription factor binding and/or the act of transcription. We further propose that replication inhibitors can induce unscheduled encounters between replication and transcription and give rise to distinct patterns of chromosome fragile sites.

  16. Cascades of genetic instability resulting from compromised break-induced replication.

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    Soumini Vasan

    2014-02-01

    Full Text Available Break-induced replication (BIR is a mechanism to repair double-strand breaks (DSBs that possess only a single end that can find homology in the genome. This situation can result from the collapse of replication forks or telomere erosion. BIR frequently produces various genetic instabilities including mutations, loss of heterozygosity, deletions, duplications, and template switching that can result in copy-number variations (CNVs. An important type of genomic rearrangement specifically linked to BIR is half-crossovers (HCs, which result from fusions between parts of recombining chromosomes. Because HC formation produces a fused molecule as well as a broken chromosome fragment, these events could be highly destabilizing. Here we demonstrate that HC formation results from the interruption of BIR caused by a damaged template, defective replisome or premature onset of mitosis. Additionally, we document that checkpoint failure promotes channeling of BIR into half-crossover-initiated instability cascades (HCC that resemble cycles of non-reciprocal translocations (NRTs previously described in human tumors. We postulate that HCs represent a potent source of genetic destabilization with significant consequences that mimic those observed in human diseases, including cancer.

  17. Previous infection with virulent strains of Newcastle disease virus reduces highly pathogenic avian influenza virus replication, disease, and mortality in chickens.

    Science.gov (United States)

    Costa-Hurtado, Mar; Afonso, Claudio L; Miller, Patti J; Shepherd, Eric; Cha, Ra Mi; Smith, Diane; Spackman, Erica; Kapczynski, Darrell R; Suarez, David L; Swayne, David E; Pantin-Jackwood, Mary J

    2015-09-23

    Highly pathogenic avian influenza virus (HPAIV) and Newcastle disease virus (NDV) are two of the most important viruses affecting poultry worldwide and produce co-infections especially in areas of the world where both viruses are endemic; but little is known about the interactions between these two viruses. The objective of this study was to determine if co-infection with NDV affects HPAIV replication in chickens. Only infections with virulent NDV strains (mesogenic Pigeon/1984 or velogenic CA/2002), and not a lentogenic NDV strain (LaSota), interfered with the replication of HPAIV A/chicken/Queretaro/14588-19/95 (H5N2) when the H5N2 was given at a high dose (10(6.9) EID50) two days after the NDV inoculation, but despite this interference, mortality was still observed. However, chickens infected with the less virulent mesogenic NDV Pigeon/1984 strain three days prior to being infected with a lower dose (10(5.3-5.5) EID50) of the same or a different HPAIV, A/chicken/Jalisco/CPA-12283-12/2012 (H7N3), had reduced HPAIV replication and increased survival rates. In conclusion, previous infection of chickens with virulent NDV strains can reduce HPAIV replication, and consequently disease and mortality. This interference depends on the titer of the viruses used, the virulence of the NDV, and the timing of the infections. The information obtained from these studies helps to understand the possible interactions and outcomes of infection (disease and virus shedding) when HPAIV and NDV co-infect chickens in the field.

  18. Is intelligence enhanced by letter priming? A failure to replicate the results of Ciani and Sheldon (2010).

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    Carlin, Sean P; Standing, Lionel G

    2013-04-01

    Previous research has suggested that motivational processes outside an individual's conscious awareness may be primed so as to enhance or impair cognitive performance. The present study involved a conceptual replication of the 2010 study of Ciani and Sheldon (Experiments 1 and 2), employing the same materials and task, to test whether exposure to the letter A before an analogies test improved performance and the letter F impaired it, relative to the neutral letter J. It also examined the effect of pre-exposing participants before testing to a positive or negative verbal passage concerning letter grades. Priming was not found to have any effect: the participants (N = 116), under both pre-exposure conditions, gave analogies scores which were virtually identical whether they had been primed with A, F, or J, thus contradicting the previous results. It is concluded that there is a pressing need for more replications of priming experiments as well as other studies.

  19. First-trimester screening for trisomy 21 with adjustment for biochemical results of previous pregnancies.

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    Wright, David; Syngelaki, Argyro; Birdir, Cahit; Bedei, Ivonne; Nicolaides, Kypros H

    2011-01-01

    To investigate the effect of associations in serum free β-hCG and PAPP-A between successive pregnancies on the performance of screening for trisomy 21 at 11-13 weeks' gestation. In 8,499 women with two consecutive pregnancies, including 49 women with fetal trisomy 21 in the second pregnancy, the correlation in serum free β-hCG multiples of the median (MoM) and PAPP-A MoM between pregnancies was determined, and the effects of correcting for the correlation on the performance of screening was estimated. There were significant associations between pregnancies in free β-hCG MoM (r = 0.4435) and PAPP-A MoM (r = 0.4796). In screening by maternal age and biochemistry at a risk cutoff of 1 in 100, in the second pregnancies the false-positive rate was 35.5% for those with screen-positive results in the first pregnancy, and this was reduced to 17.1% after adjustment for the results of the first pregnancy. Similarly, in women with screen-negative results in the first pregnancy, adjustment for the results improved the detection rate in the second pregnancy from 66.7 to 81.2%. In screening for trisomy 21, adjustment for the biochemical findings in a previous pregnancy has major effects on individual patient-specific risks, increases the detection rate and reduces the false-positive rate. Copyright © 2011 S. Karger AG, Basel.

  20. Priming in word stem completion tasks: comparison with previous results in word fragment completion tasks.

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    Soler, María J; Dasí, Carmen; Ruiz, Juan C

    2015-01-01

    This study investigates priming in an implicit word stem completion (WSC) task by analyzing the effect of linguistic stimuli characteristics on said task. A total of 305 participants performed a WSC task in two phases (study and test). The test phase included 63 unique-solution stems and 63 multiple-solution stems. Analysis revealed that priming (mean = 0.22) was stronger in the case of multiple-solution stems, indicating that they were not a homogeneous group of stimuli. Thus, further analyses were performed only for the data of the unique-solution stems. Correlations between priming and familiarity, frequency of use, and baseline completion were significant. The less familiar words, which were less frequent, had higher priming values. At the same time, the stems with lower baseline completion generated more priming. A regression analysis showed that baseline completion was the only significant predictor of priming, suggesting that the previous processing of the stimuli had a greater impact on the stimuli with low baseline performance. At the same time, baseline completion showed significant positive correlations with familiarity and frequency of use, and a negative correlation with length. When baseline completion was the dependent variable in the regression analysis, the significant variables in the regression were familiarity and length. These results were compared with those obtained in a study using word fragment completion (WFC) by Soler et al. (2009), in which the same words and procedure were employed. Analysis showed that the variables that correlated with priming were the same as in the WSC task, and that completion baseline was the variable that showed the greatest predictive power of priming. This coincidence of results obtained with WFC and WSC tasks highlights the importance of controlling the characteristics of the stimuli used when exploring the nature of priming.

  1. Treatment of extraskeletal osteosarcoma at a previous injection site resulting in prolonged survival in 1 dog.

    Science.gov (United States)

    Selmic, Laura E; Griffin, Lynn R; Rector, Megan H; Lafferty, Mary; Pool, Roy; Ehrhart, Nicole P

    2016-09-01

    A rare presentation of an extraskeletal osteosarcoma at a previous interscapular injection site in a dog is described. Treatment included surgical excision of the tumor followed by 6 rounds of intravenous carboplatin, oral toceranib, and cyclophosphamide. The dog survived for 20.5 months after diagnosis despite early development of pulmonary metastases.

  2. Organization of Replication of Ribosomal DNA in Saccharomyces cerevisiae

    NARCIS (Netherlands)

    Linskens, Maarten H.K.; Huberman, Joel A.

    1988-01-01

    Using recently developed replicon mapping techniques, we have analyzed the replication of the ribosomal DNA in Saccharomyces cerevisiae. The results show that (i) the functional origin of replication colocalizes with an autonomously replicating sequence element previously mapped to the

  3. Previous antibiotic exposure and antimicrobial resistance in invasive pneumococcal disease: results from prospective surveillance.

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    Kuster, Stefan P; Rudnick, Wallis; Shigayeva, Altynay; Green, Karen; Baqi, Mahin; Gold, Wayne L; Lovinsky, Reena; Muller, Matthew P; Powis, Jeff E; Rau, Neil; Simor, Andrew E; Walmsley, Sharon L; Low, Donald E; McGeer, Allison

    2014-10-01

    Estimating the risk of antibiotic resistance is important in selecting empiric antibiotics. We asked how the timing, number of courses, and duration of antibiotic therapy in the previous 3 months affected antibiotic resistance in isolates causing invasive pneumococcal disease (IPD). We conducted prospective surveillance for IPD in Toronto, Canada, from 2002 to 2011. Antimicrobial susceptibility was measured by broth microdilution. Clinical information, including prior antibiotic use, was collected by chart review and interview with patients and prescribers. Clinical information and antimicrobial susceptibility were available for 4062 (90%) episodes; 1193 (29%) of episodes were associated with receipt of 1782 antibiotic courses in the prior 3 months. Selection for antibiotic resistance was class specific. Time elapsed since most recent antibiotic was inversely associated with resistance (cephalosporins: adjusted odds ratio [OR] per day, 0.98; 95% confidence interval [CI], .96-1.00; P = .02; macrolides: OR, 0.98; 95% CI, .96-.99; P = .005; penicillins: OR [log(days)], 0.62; 95% CI, .44-.89; P = .009; fluoroquinolones: profile penalized-likelihood OR [log(days)], 0.62; 95% CI, .39-1.04; P = .07). Risk of resistance after exposure declined most rapidly for fluoroquinolones and penicillins and reached baseline in 2-3 months. The decline in resistance was slowest for macrolides, and in particular for azithromycin. There was no significant association between duration of therapy and resistance for any antibiotic class. Too few patients received multiple courses of the same antibiotic class to assess the significance of repeat courses. Time elapsed since last exposure to a class of antibiotics is the most important factor predicting antimicrobial resistance in pneumococci. The duration of effect is longer for macrolides than other classes. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved

  4. A Bayesian bird's eye view of ‘Replications of important results in social psychology’

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    Schönbrodt, Felix D.; Yao, Yuling; Gelman, Andrew; Wagenmakers, Eric-Jan

    2017-01-01

    We applied three Bayesian methods to reanalyse the preregistered contributions to the Social Psychology special issue ‘Replications of Important Results in Social Psychology’ (Nosek & Lakens. 2014 Registered reports: a method to increase the credibility of published results. Soc. Psychol. 45, 137–141. (doi:10.1027/1864-9335/a000192)). First, individual-experiment Bayesian parameter estimation revealed that for directed effect size measures, only three out of 44 central 95% credible intervals did not overlap with zero and fell in the expected direction. For undirected effect size measures, only four out of 59 credible intervals contained values greater than 0.10 (10% of variance explained) and only 19 intervals contained values larger than 0.05. Second, a Bayesian random-effects meta-analysis for all 38 t-tests showed that only one out of the 38 hierarchically estimated credible intervals did not overlap with zero and fell in the expected direction. Third, a Bayes factor hypothesis test was used to quantify the evidence for the null hypothesis against a default one-sided alternative. Only seven out of 60 Bayes factors indicated non-anecdotal support in favour of the alternative hypothesis (BF10>3), whereas 51 Bayes factors indicated at least some support for the null hypothesis. We hope that future analyses of replication success will embrace a more inclusive statistical approach by adopting a wider range of complementary techniques. PMID:28280547

  5. Approximation of lateral distribution of atmospheric Cherenkov light at different observation levels. Comparison with previous results

    CERN Document Server

    Mishev, A; Stamenov, J

    2005-01-01

    This work summarizes the results presented at 29th International Cosmic Ray Conference in Pune India. Generally the aim of this work is to obtain the lateral distribution of the atmospheric Cherenkov light in extensive air showers produced by different primary particles in wide energy range and at several observation levels and to fit the obtained lateral distributions. Using one large detector and partially modified CORSIKA code version are obtained the lateral distributions of Cherenkov light flux densities at several observation levels for different particle primaries precisely at 536 g/cm2 Chacaltaya, 700 g/cm2 Moussala and 875 g/cm2 Kartalska field observation levels for hadronic primaries and gamma quanta in the energy range 1011 eV-1016 eV. On the basis of the solution of over-determined inverse problem the approximation of these distributions is obtained. The same model function for all the primaries is used and for the different observation levels. The different model parameters for the different pri...

  6. Slow Replication Fork Velocity of Homologous Recombination-Defective Cells Results from Endogenous Oxidative Stress.

    Science.gov (United States)

    Wilhelm, Therese; Ragu, Sandrine; Magdalou, Indiana; Machon, Christelle; Dardillac, Elodie; Técher, Hervé; Guitton, Jérôme; Debatisse, Michelle; Lopez, Bernard S

    2016-05-01

    Replications forks are routinely hindered by different endogenous stresses. Because homologous recombination plays a pivotal role in the reactivation of arrested replication forks, defects in homologous recombination reveal the initial endogenous stress(es). Homologous recombination-defective cells consistently exhibit a spontaneously reduced replication speed, leading to mitotic extra centrosomes. Here, we identify oxidative stress as a major endogenous source of replication speed deceleration in homologous recombination-defective cells. The treatment of homologous recombination-defective cells with the antioxidant N-acetyl-cysteine or the maintenance of the cells at low O2 levels (3%) rescues both the replication fork speed, as monitored by single-molecule analysis (molecular combing), and the associated mitotic extra centrosome frequency. Reciprocally, the exposure of wild-type cells to H2O2 reduces the replication fork speed and generates mitotic extra centrosomes. Supplying deoxynucleotide precursors to H2O2-exposed cells rescued the replication speed. Remarkably, treatment with N-acetyl-cysteine strongly expanded the nucleotide pool, accounting for the replication speed rescue. Remarkably, homologous recombination-defective cells exhibit a high level of endogenous reactive oxygen species. Consistently, homologous recombination-defective cells accumulate spontaneous γH2AX or XRCC1 foci that are abolished by treatment with N-acetyl-cysteine or maintenance at 3% O2. Finally, oxidative stress stimulated homologous recombination, which is suppressed by supplying deoxynucleotide precursors. Therefore, the cellular redox status strongly impacts genome duplication and transmission. Oxidative stress should generate replication stress through different mechanisms, including DNA damage and nucleotide pool imbalance. These data highlight the intricacy of endogenous replication and oxidative stresses, which are both evoked during tumorigenesis and senescence initiation

  7. Leadership Orientation as a Function of Interpersonal Need Structure: A Replication with Negative Results.

    Science.gov (United States)

    Tucker, Jeffrey H.

    1983-01-01

    To replicate the Kuehl et al. (1975) study, 151 graduate students completed the Leadership Opinion Questionnaire, Least Preferred Co-Worker (LPC) Questionnaire, FIRO-B (Functional Interpersonal Relations Orientation-Behavior), and F-Scale. Data failed to replicate earlier significant correlations between interpersonal needs as measured by the LPC…

  8. Leadership Orientation as a Function of Interpersonal Need Structure: A Replication with Negative Results.

    Science.gov (United States)

    Tucker, Jeffrey H.

    1983-01-01

    To replicate the Kuehl et al. (1975) study, 151 graduate students completed the Leadership Opinion Questionnaire, Least Preferred Co-Worker (LPC) Questionnaire, FIRO-B (Functional Interpersonal Relations Orientation-Behavior), and F-Scale. Data failed to replicate earlier significant correlations between interpersonal needs as measured by the LPC…

  9. Host cell variations resulting from F plasmid-controlled replication of the Escherichia coli chromosome.

    Science.gov (United States)

    Tresguerres, E F; Nieto, C; Casquero, I; Cánovas, J L

    1986-01-01

    Cell size and DNA concentration were measured in Escherichia coli K-12 ET64. This strain carries a dnaA (Ts) mutation that has been suppressed by the insertion of the F plasmid into the chromosome. ET64 can grow in a balanced steady state of exponential growth at the restrictive temperature for its dnaA allele (39 degrees C), in which chromosome replication is controlled by the F plasmid, and at the permissive temperature (30 degrees C), in which chromosome replication is controlled by dnaA-oriC. When cells grown at the indicated temperatures were compared, it was observed that at 39 degrees C, the cell mass increased and the amount of cellular DNA decreased slightly; therefore, the DNA concentration was strongly reduced. These changes can neither be explained by the reduction of the generation time (which is only 10-15%) nor from observed changes in the replication time and in the time between DNA synthesis termination and cell division. Variations were mainly due to the increase in cell mass per origin of replication, at initiation, in cells grown at 39 degrees C. Control of chromosome replication by the F plasmid appears to be the reason for the increase in the initiation mass. Other possible causes, such as the modification of growth temperature, the generation time, or both, were discarded. These observations suggest that at one growth rate, the F plasmid replicates at a particular cell mass to F particle number ratio, and that this ratio is higher than the cell mass to oriC ratio at the initiation of chromosome replication. This fact might be significant to coordinate the replication of two different replicons in the same cell. PMID:3511032

  10. Patterns and predictors of ADHD persistence into adulthood: Results from the National Comorbidity Survey Replication

    Science.gov (United States)

    Kessler, Ronald C.; Adler, Lenard A.; Barkley, Russell; Biederman, Joseph; Conners, C. Keith; Faraone, Stephen V.; Greenhill, Laurence L.; Jaeger, Savina; Secnik, Kristina; Spencer, Thomas; Üstün, T. Bedirhan; Zaslavsky, Alan M.

    2010-01-01

    BACKGROUND Despite growing interest in adult ADHD, little is known about predictors of persistence of childhood cases into adulthood. METHODS A retrospective assessment of childhood ADHD, childhood risk factors, and a screen for adult ADHD were included in a sample of 3197 18–44 year old respondents in the National Comorbidity Survey Replication (NCS-R). Blinded adult ADHD clinical reappraisal interviews were administered to a sub-sample of respondents. Multiple imputation (MI) was used to estimate adult persistence of childhood ADHD. Logistic regression was used to study retrospectively reported childhood predictors of persistence. Potential predictors included socio-demographics, childhood ADHD severity, childhood adversity, traumatic life experiences, and comorbid DSM-IV child-adolescent disorders (anxiety, mood, impulse-control, and substance disorders). RESULTS 36.3% of respondents with retrospectively assessed childhood ADHD were classified by blinded clinical interviews as meeting DSM-IV criteria for current ADHD. Childhood ADHD severity and childhood treatment significantly predicted persistence. Controlling for severity and excluding treatment, none of the other variables significantly predicted persistence even though they were significantly associated with childhood ADHD. CONCLUSIONS No modifiable risk factors were found for adult persistence of ADHD. Further research, ideally based on prospective general population samples, is needed to search for modifiable determinants of adult persistence of ADHD. PMID:15950019

  11. Sleep disturbance and obsessive-compulsive symptoms: Results from the national comorbidity survey replication.

    Science.gov (United States)

    Cox, Rebecca C; Olatunji, Bunmi O

    2016-04-01

    A small body of developing research has found evidence for sleep disturbance in obsessive-compulsive disorder (OCD) and links between sleep disturbance and obsessive-compulsive symptoms (OCS) in unselected samples. However, the link between sleep disturbance and OCS is yet to be examined in a nationally representative sample. Furthermore, the extent to which the link between sleep disturbance and OCS is accounted for by symptoms of depression remains unclear. To address this gap in the literature, the present study examined the relationship between sleep disturbance and OCS in a nationally representative sample. Participants were assessed in the National Comorbidity Survey Replication (NCS-R; n = 2073). Consistent with predictions, results revealed that individuals with sleep disturbance reported increased OCS severity compared to individuals without sleep disturbance. Further, sleep disturbance severity was associated with OCS severity, even when controlling for depression (and other anxiety-related disorders). This study is the first to link sleep disturbance and OCS in a nationally representative sample, and these findings highlight the unique role of sleep disturbance in the experience of OCS. Future research is necessary to delineate specific mechanisms that may account for this relationship.

  12. Great apes can defer exchange: a replication with different results suggesting future oriented behavior.

    Science.gov (United States)

    Osvath, Mathias; Persson, Tomas

    2013-01-01

    The topic of cognitive foresight in non-human animals has received considerable attention in the last decade. The main questions concern whether the animals can prepare for upcoming situations which are, to various degrees, contextually or sensorially detached from the situation in which the preparations are made. Studies on great apes have focused on tool-related tasks, e.g., the ability to select a tool which is functional only in the future. Dufour and Sterck (2008), however, investigated whether chimpanzees were also able to prepare for a future exchange with a human: an object exchanged for a food item. The study included extensive training on the exchangeable item, which is traditionally not compatible with methods for studying planning abilities, as associative learning cannot be precluded. Nevertheless, despite this training, the chimpanzees could not solve the deferred exchange task. Given that great apes can plan for tool use, these results are puzzling. In addition, claims that great ape foresight is highly limited has been based on this study (Suddendorf and Corballis, 2010). Here we partly replicated Dufour and Sterck's study to discern whether temporally deferred and spatially displaced exchange tasks are beyond the capabilities of great apes. In addition to chimpanzees we tested orangutans. One condition followed the one used by Dufour and Sterck, in which the exchange items, functional only in the future, are placed at a location that freely allows for selections by the subjects. In order to test the possibility that the choice set-up could explain the negative results in Dufour and Sterck's study, our second condition followed a method used in the planning study by Osvath and Osvath (2008), where the subjects make a forced one-item-choice from a tray. We found that it is within the capabilities of chimpanzees and orangutans to perform deferred exchange in both conditions.

  13. Great apes can defer exchange: a replication with different results suggesting future oriented behaviour

    Directory of Open Access Journals (Sweden)

    Mathias eOsvath

    2013-10-01

    Full Text Available The topic of cognitive foresight in non-human animals has received considerable attention in the last decade. The main questions concern whether the animals can prepare for upcoming situations which are, to various degrees, contextually or sensorially detached from the situation in which the preparations are made. Studies on great apes have focused on tool-related tasks, e.g. the ability to select a tool which is functional only in the future. Dufour and Sterck (2008, however, investigated whether chimpanzees were also able to prepare for a future exchange with a human: an object exchanged for a food item. The study included extensive training on the exchangeable item, which is traditionally not compatible with methods for studying planning abilities, as associative learning cannot be precluded. Nevertheless, despite this training, the chimpanzees could not solve the deferred exchange task. Given that great apes can plan for tool use, these results are puzzling. In addition, claims that great ape foresight is highly limited has been based on this study (Suddendorf and Corballis, 2010. Here we partly replicated Dufour and Sterck’s study to discern whether temporally deferred and spatially displaced exchange tasks are beyond the capabilities of great apes. In addition to chimpanzees we tested orangutans. One condition followed the one used by Dufour and Sterck, in which the exchange items, functional only in the future, are placed at a location that freely allows for selections by the subjects. In order to test the possibility that the choice set-up could explain the negative results in Dufour and Sterck’s study, our second condition followed a method used in the planning study by Osvath and Osvath (2008, where the subjects make a forced one-item-choice from a tray. We found that it is within the capabilities of chimpanzees and orangutans to perform deferred exchange in both conditions.

  14. The prevalence and correlates of adult ADHD in the United States: Results from the National Comorbidity Survey Replication

    Science.gov (United States)

    Kessler, Ronald C.; Adler, Lenard; Barkley, Russell; Biederman, Joseph; Conners, C. Keith; Demler, Olga; Faraone, Stephen V.; Greenhill, Laurence L.; Howes, Mary J.; Secnik, Kristina; Spencer, Thomas; Ustun, T. Bedirhan; Walters, Ellen E.; Zaslavsky, Alan M.

    2010-01-01

    OBJECTIVE Despite growing interest in adult attention-deficit/hyperactivity disorder (ADHD), little is known about prevalence or correlates. METHODS A screen for adult ADHD was included in a probability sub-sample (n = 3199) of 18–44 year old respondents in the National Comorbidity Survey Replication (NCS-R), a nationally representative household survey that used a lay-administered diagnostic interview to assess a wide range of DSM-IV disorders. Blinded clinical follow-up interviews of adult ADHD were carried out with 154 NCS-R respondents, over-sampling those with a positive screen. Multiple imputation (MI) was used to estimate prevalence and correlates of clinician-assessed adult ADHD. RESULTS Estimated prevalence of current adult ADHD is 4.4%. Significant correlates include being male, previously married, unemployed, and Non-Hispanic White. Adult ADHD is highly comorbid with many other NCS-R/DSM-IV disorders and is associated with substantial role impairment. The majority of cases are untreated, although many obtain treatment for other comorbid mental and substance disorders. CONCLUSIONS Efforts are needed to increase the detection and treatment of adult ADHD. Research is needed to determine whether effective treatment would reduce the onset, persistence, and severity of disorders that co-occur with adult ADHD. PMID:16585449

  15. Modeling inhomogeneous DNA replication kinetics.

    Directory of Open Access Journals (Sweden)

    Michel G Gauthier

    Full Text Available In eukaryotic organisms, DNA replication is initiated at a series of chromosomal locations called origins, where replication forks are assembled proceeding bidirectionally to replicate the genome. The distribution and firing rate of these origins, in conjunction with the velocity at which forks progress, dictate the program of the replication process. Previous attempts at modeling DNA replication in eukaryotes have focused on cases where the firing rate and the velocity of replication forks are homogeneous, or uniform, across the genome. However, it is now known that there are large variations in origin activity along the genome and variations in fork velocities can also take place. Here, we generalize previous approaches to modeling replication, to allow for arbitrary spatial variation of initiation rates and fork velocities. We derive rate equations for left- and right-moving forks and for replication probability over time that can be solved numerically to obtain the mean-field replication program. This method accurately reproduces the results of DNA replication simulation. We also successfully adapted our approach to the inverse problem of fitting measurements of DNA replication performed on single DNA molecules. Since such measurements are performed on specified portion of the genome, the examined DNA molecules may be replicated by forks that originate either within the studied molecule or outside of it. This problem was solved by using an effective flux of incoming replication forks at the model boundaries to represent the origin activity outside the studied region. Using this approach, we show that reliable inferences can be made about the replication of specific portions of the genome even if the amount of data that can be obtained from single-molecule experiments is generally limited.

  16. How many replicate tests do I need?$-$ Variability of cookstove performance and emissions has implications for obtaining useful results

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Yungang [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Environmental Energy Technologies Division; Sohn, Michael D. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Environmental Energy Technologies Division; Gadgil, Ashok J. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Environmental Energy Technologies Division; Wang, Yilun [ISO Innovative Analytics San Francisco, CA (United States); Lask, Kathleen M. [Univ. of California, Berkeley, CA (United States). College of Engineering Applied Science and Technology Program; Kirchstetter, Thomas W. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Environmental Energy Technologies Division

    2013-02-01

    Almost half of the world’s population still cooks on biomass cookstoves of poor efficiency and primitive design, such as three stone fires (TSF). Emissions from biomass cookstoves contribute to adverse health effects and climate change. A number of “improved cookstoves” with higher energy efficiency and lower emissions have been designed and promoted across the world. During the design development, and for selection of a stove for dissemination, the stove performance and emissions are commonly evaluated, communicated and compared using the arithmetic average of replicate tests made using a standardized laboratory-based test, commonly the water boiling test (WBT). However, published literature shows different WBT results reported from different laboratories for the same stove technology. Also, there is no agreement in the literature on how many replicate tests should be performed to ensure “significance” in the reported average performance. This matter has not received attention in the rapidly growing literature on stoves, and yet is crucial for estimating and communicating the performance of a stove, and for comparing the performance between stoves. We present results of statistical analyses using data from a number of replicate tests of performance and emission of the Berkeley-Darfur Stove (BDS) and the TSF under well-controlled laboratory conditions. We observed moderate variability in the test results for the TSF and BDS when measuring several characteristics. Here we focus on two as illustrative: time-to-boil and PM2.5 (particulate matter less than or equal to 2.5 micrometers in diameter) emissions. We demonstrate that interpretation of the results comparing these stoves could be misleading if only a small number of replicates had been conducted. We then describe a practical approach, useful to both stove testers and designers, to assess the number of replicates needed to obtain useful data. Caution should be exercised in attaching high credibility to

  17. Colonic perforation resulting from ingested chicken bone revealing previously undiagnosed colonic adenocarcinoma: report of a case and review of literature.

    Science.gov (United States)

    McGregor, Douglas H; Liu, Xiaoying; Ulusarac, Ozlem; Ponnuru, Kimberly D; Schnepp, Stephanie L

    2011-02-18

    An 86 year old male with a four-day history of nonspecific gastrointestinal symptoms was found on colonoscopy to have evidence of sigmoid colon obstruction and possible perforation. Emergent operative exploration revealed diffuse peritonitis, sigmoid perforation, adjacent dense adhesions, and a foreign body protruding through the perforated area. Pathologic examination showed the foreign body to be a sliver of bone consistent with chicken bone and the sigmoid subacute perforation to be associated distally with a circumferential ulcerated obstructing mass, microscopically seen to be transmurally infiltrating adenocarcinoma, signet-ring cell type. There was extensive acute and organizing peritonitis, 100% Escherichia coli was cultured from peritoneal fluid, and the patient died two days postoperatively with sepsis and hypotension. This appears to be the fifth reported case of colonic perforation resulting from foreign body perforation due to previously undiagnosed adenocarcinoma. The four previously reported cases were all deeply invasive adenocarcinoma of sigmoid colon, and the foreign bodies included three chicken/poultry bones and a metallic staple. These five cases are highly unusual examples of a potentially lethal malignant neoplasm being clinically revealed by a usually (but not always) innocuous event, the ingestion of a small foreign body.

  18. Colonic perforation resulting from ingested chicken bone revealing previously undiagnosed colonic adenocarcinoma: report of a case and review of literature

    Directory of Open Access Journals (Sweden)

    Ulusarac Ozlem

    2011-02-01

    Full Text Available Abstract An 86 year old male with a four-day history of nonspecific gastrointestinal symptoms was found on colonoscopy to have evidence of sigmoid colon obstruction and possible perforation. Emergent operative exploration revealed diffuse peritonitis, sigmoid perforation, adjacent dense adhesions, and a foreign body protruding through the perforated area. Pathologic examination showed the foreign body to be a sliver of bone consistent with chicken bone and the sigmoid subacute perforation to be associated distally with a circumferential ulcerated obstructing mass, microscopically seen to be transmurally infiltrating adenocarcinoma, signet-ring cell type. There was extensive acute and organizing peritonitis, 100% Escherichia coli was cultured from peritoneal fluid, and the patient died two days postoperatively with sepsis and hypotension. This appears to be the fifth reported case of colonic perforation resulting from foreign body perforation due to previously undiagnosed adenocarcinoma. The four previously reported cases were all deeply invasive adenocarcinoma of sigmoid colon, and the foreign bodies included three chicken/poultry bones and a metallic staple. These five cases are highly unusual examples of a potentially lethal malignant neoplasm being clinically revealed by a usually (but not always innocuous event, the ingestion of a small foreign body.

  19. Temperature sensitive influenza A virus genome replication results from low thermal stability of polymerase-cRNA complexes

    Directory of Open Access Journals (Sweden)

    Tiley Laurence S

    2006-08-01

    Full Text Available Abstract Background The RNA-dependent RNA polymerase of Influenza A virus is a determinant of viral pathogenicity and host range that is responsible for transcribing and replicating the negative sense segmented viral genome (vRNA. Transcription produces capped and polyadenylated mRNAs whereas genome replication involves the synthesis of an alternative plus-sense transcript (cRNA with unmodified termini that is copied back to vRNA. Viral mRNA transcription predominates at early stages of viral infection, while later, negative sense genome replication is favoured. However, the "switch" that regulates the transition from transcription to replication is poorly understood. Results We show that temperature strongly affects the balance between plus and minus-sense RNA synthesis with high temperature causing a large decrease in vRNA accumulation, a moderate decrease in cRNA levels but (depending on genome segment either increased or unchanged levels of mRNA. We found no evidence implicating cellular heat shock protein activity in this effect despite the known association of hsp70 and hsp90 with viral polymerase components. Temperature-shift experiments indicated that polymerase synthesised at 41°C maintained transcriptional activity even though genome replication failed. Reduced polymerase association with viral RNA was seen in vivo and in confirmation of this, in vitro binding assays showed that temperature increased the rate of dissociation of polymerase from both positive and negative sense promoters. However, the interaction of polymerase with the cRNA promoter was particularly heat labile, showing rapid dissociation even at 37°C. This suggested that vRNA synthesis fails at elevated temperatures because the polymerase does not bind the promoter. In support of this hypothesis, a mutant cRNA promoter with vRNA-like sequence elements supported vRNA synthesis at higher temperatures than the wild-type promoter. Conclusion The differential stability of

  20. Sofosbuvir Inhibits Hepatitis E Virus Replication In Vitro and Results in an Additive Effect When Combined With Ribavirin.

    Science.gov (United States)

    Dao Thi, Viet Loan; Debing, Yannick; Wu, Xianfang; Rice, Charles M; Neyts, Johan; Moradpour, Darius; Gouttenoire, Jérôme

    2016-01-01

    Infection with hepatitis E virus genotype 3 may result in chronic hepatitis in immunocompromised patients. Reduction of immunosuppression or treatment with ribavirin or pegylated interferon-α can result in viral clearance. However, safer and more effective treatment options are needed. Here, we show that sofosbuvir inhibits the replication of hepatitis E virus genotype 3 both in subgenomic replicon systems as well as a full-length infectious clone. Moreover, the combination of sofosbuvir and ribavirin results in an additive antiviral effect. Sofosbuvir may be considered as an add-on therapy to ribavirin for the treatment of chronic hepatitis E in immunocompromised patients.

  1. Selective replication of oncolytic virus M1 results in a bystander killing effect that is potentiated by Smac mimetics.

    Science.gov (United States)

    Cai, Jing; Lin, Yuan; Zhang, Haipeng; Liang, Jiankai; Tan, Yaqian; Cavenee, Webster K; Yan, Guangmei

    2017-06-27

    Oncolytic virotherapy is a treatment modality that uses native or genetically modified viruses that selectively replicate in and kill tumor cells. Viruses represent a type of pathogen-associated molecular pattern and thereby induce the up-regulation of dozens of cytokines via activating the host innate immune system. Second mitochondria-derived activator of caspases (Smac) mimetic compounds (SMCs), which antagonize the function of inhibitor of apoptosis proteins (IAPs) and induce apoptosis, sensitize tumor cells to multiple cytokines. Therefore, we sought to determine whether SMCs sensitize tumor cells to cytokines induced by the oncolytic M1 virus, thus enhancing a bystander killing effect. Here, we report that SMCs potentiate the oncolytic effect of M1 in vitro, in vivo, and ex vivo. This strengthened oncolytic efficacy resulted from the enhanced bystander killing effect caused by the M1 virus via cytokine induction. Through a microarray analysis and subsequent validation using recombinant cytokines, we identified IL-8, IL-1A, and TRAIL as the key cytokines in the bystander killing effect. Furthermore, SMCs increased the replication of M1, and the accumulation of virus protein induced irreversible endoplasmic reticulum stress- and c-Jun N-terminal kinase-mediated apoptosis. Nevertheless, the combined treatment with M1 and SMCs had little effect on normal and human primary cells. Because SMCs selectively and significantly enhance the bystander killing effect and the replication of oncolytic virus M1 specifically in cancer cells, this combined treatment may represent a promising therapeutic strategy.

  2. Passive acoustic monitoring using a towed hydrophone array results in identification of a previously unknown beaked whale habitat.

    Science.gov (United States)

    Yack, Tina M; Barlow, Jay; Calambokidis, John; Southall, Brandon; Coates, Shannon

    2013-09-01

    Beaked whales are diverse and species rich taxa. They spend the vast majority of their time submerged, regularly diving to depths of hundreds to thousands of meters, typically occur in small groups, and behave inconspicuously at the surface. These factors make them extremely difficult to detect using standard visual survey methods. However, recent advancements in acoustic detection capabilities have made passive acoustic monitoring (PAM) a viable alternative. Beaked whales can be discriminated from other odontocetes by the unique characteristics of their echolocation clicks. In 2009 and 2010, PAM methods using towed hydrophone arrays were tested. These methods proved highly effective for real-time detection of beaked whales in the Southern California Bight (SCB) and were subsequently implemented in 2011 to successfully detect and track beaked whales during the ongoing Southern California Behavioral Response Study. The three year field effort has resulted in (1) the successful classification and tracking of Cuvier's (Ziphius cavirostris), Baird's (Berardius bairdii), and unidentified Mesoplodon beaked whale species and (2) the identification of areas of previously unknown beaked whale habitat use. Identification of habitat use areas will contribute to a better understanding of the complex relationship between beaked whale distribution, occurrence, and preferred habitat characteristics on a relatively small spatial scale. These findings will also provide information that can be used to promote more effective management and conservation of beaked whales in the SCB, a heavily used Naval operation and training region.

  3. Experimental depletion of CD8+ cells in acutely SIVagm-Infected African Green Monkeys results in increased viral replication

    Directory of Open Access Journals (Sweden)

    Apetrei Cristian

    2010-05-01

    Full Text Available Abstract Background In vivo CD8+ cell depletions in pathogenic SIV infections identified a key role for cellular immunity in controlling viral load (VL and disease progression. However, similar studies gave discordant results in chronically-infected SMs, leading some authors to propose that in natural hosts, SIV replication is independent of cellular immunity. To assess the role of cellular immune responses in the control of SIV replication in natural hosts, we investigated the impact of CD8+ cell depletion during acute SIV infection in AGMs. Results Nine AGMs were infected with SIVagm.sab and were followed up to day 225 p.i. Four were intravenously infused with the cM-T807 antibody on days 0 (50 mg/kg, 6, and 13 (10 mg/kg, respectively post infection (p.i.. CD8+ cells were depleted for up to 28 days p.i. in peripheral blood and LNs in all treated AGMs. Partial CD8+ T cell depletion occurred in the intestine. SIVagm VLs peaked at similar levels in both groups (107-108 RNA copies/ml. However, while VLs were controlled in undepleted AGMs, reaching set-point levels (104-105 RNA copies/ml by day 28 p.i., high VLs (>106 RNA copies/ml were maintained by day 21 p.i. in CD8-depleted AGMs. By day 42 p.i., VLs were comparable between the two groups. The levels of immune activation and proliferation remained elevated up to day 72 p.i. in CD8-depleted AGMs and returned to preinfection levels in controls by day 28 p.i. None of the CD8-depleted animals progressed to AIDS. Conclusion CD8+ cells are responsible for a partial control of postacute viral replication in SIVagm.sab-infected AGMs. In contrast to macaques, the SIVagm-infected AGMs are able to control viral replication after recovery of the CD8+ T cells and avoid disease progression.

  4. Bendamustine compared with chlorambucil in previously untreated patients with chronic lymphocytic leukaemia: updated results of a randomized phase III trial.

    Science.gov (United States)

    Knauf, Wolfgang U; Lissitchkov, Toshko; Aldaoud, Ali; Liberati, Anna M; Loscertales, Javier; Herbrecht, Raoul; Juliusson, Gunnar; Postner, Gerhard; Gercheva, Liana; Goranov, Stefan; Becker, Martin; Fricke, Hans-Joerg; Huguet, Francoise; Del Giudice, Ilaria; Klein, Peter; Merkle, Karlheinz; Montillo, Marco

    2012-10-01

    The efficacy of bendamustine versus chlorambucil in a phase III trial of previously untreated patients with Binet stage B/C chronic lymphocytic leukaemia (CLL) was re-evaluated after a median observation time of 54 months in May 2010. Overall survival (OS) was analysed for the first time. At follow-up, investigator-assessed complete response (CR) rate (21·0% vs 10·8%), median progression-free survival (21·2 vs 8·8 months; P chlorambucil. OS was not different between groups for all patients or those ≤65 years, >65 years, responders and non-responders. However, patients with objective response or a CR experienced a significantly longer OS than non-responders or those without a CR. Significantly more patients on chlorambucil progressed to second/further lines of treatment compared with those on bendamustine (78·3% vs 63·6%; P = 0·004). The benefits of bendamustine over chlorambucil were achieved without reducing quality of life. In conclusion, bendamustine is significantly more effective than chlorambucil in previously untreated CLL patients, with the achievement of a CR or objective response appearing to prolong OS. Bendamustine should be considered as a preferred first-line option over chlorambucil for CLL patients ineligible for fludarabine, cyclophosphamide and rituximab. © 2012 Blackwell Publishing Ltd.

  5. Cyclophilin A binds to the viral RNA and replication proteins, resulting in inhibition of tombusviral replicase assembly.

    Science.gov (United States)

    Kovalev, Nikolay; Nagy, Peter D

    2013-12-01

    Replication of plus-stranded RNA viruses is greatly affected by numerous host-encoded proteins that act as restriction factors. Cyclophilins, which are a large family of cellular prolyl isomerases, have been found to inhibit Tomato bushy stunt tombusvirus (TBSV) replication in a Saccharomyces cerevisiae model based on genome-wide screens and global proteomics approaches. In this report, we further characterize single-domain cyclophilins, including the mammalian cyclophilin A and plant Roc1 and Roc2, which are orthologs of the yeast Cpr1p cyclophilin, a known inhibitor of TBSV replication in yeast. We found that recombinant CypA, Roc1, and Roc2 strongly inhibited TBSV replication in a cell-free replication assay. Additional in vitro studies revealed that CypA, Roc1, and Roc2 cyclophilins bound to the viral replication proteins, and CypA and Roc1 also bound to the viral RNA. These interactions led to inhibition of viral RNA recruitment, the assembly of the viral replicase complex, and viral RNA synthesis. A catalytically inactive mutant of CypA was also able to inhibit TBSV replication in vitro due to binding to the replication proteins and the viral RNA. Overexpression of CypA and its mutant in yeast or plant leaves led to inhibition of tombusvirus replication, confirming that CypA is a restriction factor for TBSV. Overall, the current work has revealed a regulatory role for the cytosolic single-domain Cpr1-like cyclophilins in RNA virus replication.

  6. Bomb-Pulse Chlorine-36 at the Proposed Yucca Mountain Repository Horizon: An Investigation of Previous Conflicting Results and Collection of New Data

    Energy Technology Data Exchange (ETDEWEB)

    Cizdziel, James

    2006-07-31

    yielded 1160 ± 50 (1σ) x 10-15. Three samples collected from Alcove 6 averaged 490 ± 100 (1σ) x10-15; a sample from Sundance Fault resulted in a ratio of 920 ± 60 (1σ) x10-15, and a sample from the Bow Ridge Fault produced 530 ± 20 (1σ) x10-15. The results are significant because: 1) they tend to be lower than LANL data for comparable samples, albeit in agreement with the range of data produced in the area, and 2) they show that a bomb-pulse 36Cl/Cl ratio was measured in rock collected at the repository horizon level by a second and independent group of investigators (UNLV). Because of time UNLV was not able to replicate the results, and these few data points are insufficient to draw major and definitive conclusions. Leachates of soil samples collected from the surface above the ESF yielded several ratios with bomb-pulse 36Cl, particularly for samples encompassing the wetting front. Soil samples collected above the south ramp, where there was limited soil coverage due to a large amount of rock outcrop, had relatively large ratios ranging from 2170 ± 110 (1σ) x10-15 to 5670 ± 350 (1σ) x10-15. Soil samples from profiles from above the north ramp ranged from 820 ± 70 (1σ) x10-15 to 2390 ± 160 (1σ) x10-15, which compare favorably with previous measurements near the site. Water seepage into the ESF south ramp and 36Cl standards made from NIST material were also analyzed. The standards were produced to have nominal 36Cl/Cl ratios (10-15) of 500, 2,500 and 10,000 and the results showed good agreement with the calculated ratios. The seepage samples ranged between 680 ± 40 (1σ) x10-15 to 1110 ± 40 (1σ) x10-15, consistent with that found for modern meteoric water, with a small bomb-pulse component. Bomb-pulse 36Cl may not have been incorporated in this fast-path water because the surface above the infiltration zone consists mostly of outcrop and the flow pathways have probably mostly been leached. 99Tc was measured in five of nine leaches of ESF rock but poor

  7. Two-stage replication of previous genome-wide association studies of AS3MT-CNNM2-NT5C2 gene cluster region in a large schizophrenia case-control sample from Han Chinese population.

    Science.gov (United States)

    Guan, Fanglin; Zhang, Tianxiao; Li, Lu; Fu, Dongke; Lin, Huali; Chen, Gang; Chen, Teng

    2016-10-01

    Schizophrenia is a devastating psychiatric condition with high heritability. Replicating the specific genetic variants that increase susceptibility to schizophrenia in different populations is critical to better understand schizophrenia. CNNM2 and NT5C2 are genes recently identified as susceptibility genes for schizophrenia in Europeans, but the exact mechanism by which these genes confer risk for schizophrenia remains unknown. In this study, we examined the potential for genetic susceptibility to schizophrenia of a three-gene cluster region, AS3MT-CNNM2-NT5C2. We implemented a two-stage strategy to conduct association analyses of the targeted regions with schizophrenia. A total of 8218 individuals were recruited, and 45 pre-selected single nucleotide polymorphisms (SNPs) were genotyped. Both single-marker and haplotype-based analyses were conducted in addition to imputation analysis to increase the coverage of our genetic markers. Two SNPs, rs11191419 (OR=1.24, P=7.28×10(-5)) and rs11191514 (OR=1.24, P=0.0003), with significant independent effects were identified. These results were supported by the data from both the discovery and validation stages. Further haplotype and imputation analyses also validated these results, and bioinformatics analyses indicated that CALHM1, which is located approximately 630kb away from CNNM2, might be a susceptible gene for schizophrenia. Our results provide further support that AS3MT, CNNM2 and CALHM1 are involved with the etiology and pathogenesis of schizophrenia, suggesting these genes are potential targets of interest for the improvement of disease management and the development of novel pharmacological strategies.

  8. Nonrecurrent PMP22-RAI1 contiguous gene deletions arise from replication-based mechanisms and result in Smith-Magenis syndrome with evident peripheral neuropathy.

    Science.gov (United States)

    Yuan, Bo; Neira, Juanita; Gu, Shen; Harel, Tamar; Liu, Pengfei; Briceño, Ignacio; Elsea, Sarah H; Gómez, Alberto; Potocki, Lorraine; Lupski, James R

    2016-10-01

    Hereditary neuropathy with liability to pressure palsies (HNPP) and Smith-Magenis syndrome (SMS) are genomic disorders associated with deletion copy number variants involving chromosome 17p12 and 17p11.2, respectively. Nonallelic homologous recombination (NAHR)-mediated recurrent deletions are responsible for the majority of HNPP and SMS cases; the rearrangement products encompass the key dosage-sensitive genes PMP22 and RAI1, respectively, and result in haploinsufficiency for these genes. Less frequently, nonrecurrent genomic rearrangements occur at this locus. Contiguous gene duplications encompassing both PMP22 and RAI1, i.e., PMP22-RAI1 duplications, have been investigated, and replication-based mechanisms rather than NAHR have been proposed for these rearrangements. In the current study, we report molecular and clinical characterizations of six subjects with the reciprocal phenomenon of deletions spanning both genes, i.e., PMP22-RAI1 deletions. Molecular studies utilizing high-resolution array comparative genomic hybridization and breakpoint junction sequencing identified mutational signatures that were suggestive of replication-based mechanisms. Systematic clinical studies revealed features consistent with SMS, including features of intellectual disability, speech and gross motor delays, behavioral problems and ocular abnormalities. Five out of six subjects presented clinical signs and/or objective electrophysiologic studies of peripheral neuropathy. Clinical profiling may improve the clinical management of this unique group of subjects, as the peripheral neuropathy can be more severe or of earlier onset as compared to SMS patients having the common recurrent deletion. Moreover, the current study, in combination with the previous report of PMP22-RAI1 duplications, contributes to the understanding of rare complex phenotypes involving multiple dosage-sensitive genes from a genetic mechanistic standpoint.

  9. Replication of five prostate cancer loci identified in an Asian population – Results from the NCI Breast and Prostate Cancer Cohort Consortium (BPC3)

    Science.gov (United States)

    Lindstrom, Sara; Schumacher, Fredrick R.; Campa, Daniele; Albanes, Demetrius; Andriole, Gerald; Berndt, Sonja I.; Bueno-de-Mesquita, H. Bas; Chanock, Stephen J.; Diver, W. Ryan; Ganziano, J. Michael; Gapstur, Susan M.; Giovannucci, Edward; Haiman, Christopher A.; Henderson, Brian; Hunter, David J; Johansson, Mattias; Kolonel, Laurence N.; Le Marchand, Loic; Ma, Jing; Stampfer, Meir; Stevens, Victoria L.; Trichopoulos, Dimitrios; Virtamo, Jarmo; Willett, Walter C.; Yeager, Meredith; Hsing, Ann W.; Kraft, Peter

    2011-01-01

    Background A recent Genome-Wide Association Study (GWAS) of prostate cancer in a Japanese population identified five novel regions not previously discovered in other ethnicities. In this study, we attempt to replicate these five loci in a series of nested prostate cancer case-control studies of European ancestry. Methods We genotyped five SNPs: rs13385191 (chromosome 2p24), rs12653946 (5p15), rs1983891 (6p21), rs339331 (6p22) and rs9600079 (13q22), in 7,956 prostate cancer cases and 8,148 controls from a series of nested case-control studies within the NCI Breast and Prostate Cancer Cohort Consortium (BPC3). We tested each SNP for association with prostate cancer risk and assessed if associations differed with respect to disease severity and age of onset. Results Four SNPs (rs13385191, rs12653946, rs1983891 and rs339331) were significantly associated with prostate cancer risk (p-values ranging from 0.01 to 1.1×10-5). Allele frequencies and odds ratios were overall lower in our population of European descent compared to the discovery Asian population. SNP rs13385191 (C2orf43) was only associated with low-stage disease (p=0.009, case-only test). No other SNP showed association with disease severity or age of onset. We did not replicate the 13q22 SNP, rs9600079 (p=0.62). Conclusions Four SNPs associated with prostate cancer risk in an Asian population are also associated with prostate cancer risk in men of European descent. Impact This study illustrates the importance of evaluation of prostate cancer risk markers across ethnic groups. PMID:22056501

  10. Guns, Impulsive Angry Behavior, and Mental Disorders: Results from the National Comorbidity Survey Replication (NCS-R).

    Science.gov (United States)

    Swanson, Jeffrey W; Sampson, Nancy A; Petukhova, Maria V; Zaslavsky, Alan M; Appelbaum, Paul S; Swartz, Marvin S; Kessler, Ronald C

    2015-06-01

    Analyses from the National Comorbidity Study Replication provide the first nationally representative estimates of the co-occurrence of impulsive angry behavior and possessing or carrying a gun among adults with and without certain mental disorders and demographic characteristics. The study found that a large number of individuals in the United States self-report patterns of impulsive angry behavior and also possess firearms at home (8.9%) or carry guns outside the home (1.5%). These data document associations of numerous common mental disorders and combinations of angry behavior with gun access. Because only a small proportion of persons with this risky combination have ever been involuntarily hospitalized for a mental health problem, most will not be subject to existing mental health-related legal restrictions on firearms resulting from a history of involuntary commitment. Excluding a large proportion of the general population from gun possession is also not likely to be feasible. Behavioral risk-based approaches to firearms restriction, such as expanding the definition of gun-prohibited persons to include those with violent misdemeanor convictions and multiple DUI convictions, could be a more effective public health policy to prevent gun violence in the population.

  11. Should Excessive Worry Be Required for a Diagnosis of Generalized Anxiety Disorder? Results from the US National Comorbidity Survey Replication

    Science.gov (United States)

    Ruscio, Ayelet Meron; Lane, Michael; Roy-Byrne, Peter; Stang, Paul E.; Stein, Dan J.; Wittchen, Hans-Ulrich; Kessler, Ronald C.

    2007-01-01

    Background Excessive worry is required by DSM-IV, but not ICD-10, for a diagnosis of generalized anxiety disorder (GAD). No large-scale epidemiological study has ever examined the implications of this requirement for estimates of prevalence, severity, or correlates of GAD. Methods Data were analyzed from the US National Comorbidity Survey Replication, a nationally representative, face-to-face survey of adults in the US household population that was fielded in 2001–2003. DSM-IV GAD was assessed with Version 3.0 of the WHO Composite International Diagnostic Interview. Non-excessive worriers meeting all other DSM-IV criteria for GAD were compared with respondents who met full GAD criteria as well as with other survey respondents to consider the implications of removing the excessiveness requirement. Results The estimated lifetime prevalence of GAD increases by approximately 40% when the excessiveness requirement is removed. Excessive GAD begins earlier in life, has a more chronic course, and is associated with greater symptom severity and psychiatric comorbidity than non-excessive GAD. However, non-excessive cases nonetheless evidence substantial persistence and impairment of GAD as well as significantly elevated comorbidity compared to respondents without GAD. Non-excessive cases also have socio-demographic characteristics and familial aggregation of GAD comparable to excessive cases. Conclusions Although individuals who meet all criteria for GAD other than excessiveness have a somewhat milder presentation than those with excessive worry, their syndromes are sufficiently similar to those with excessive worry to warrant a GAD diagnosis. PMID:16300690

  12. Experimental and calculation results of the integral reflood test QUENCH-15 with ZIRLO{sup TM} cladding tubes in comparison with results of previous QUENCH tests

    Energy Technology Data Exchange (ETDEWEB)

    Stuckert, J., E-mail: juri.stuckert@kit.edu [Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany); Birchley, J. [Paul Scherrer Institut (PSI), CH-5232 Villigen PSI (Switzerland); Grosse, M. [Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany); Jaeckel, B. [Paul Scherrer Institut (PSI), CH-5232 Villigen PSI (Switzerland); Steinbrueck, M. [Karlsruhe Institute of Technology (KIT), Hermann-von-Helmholtz-Platz 1, D-76344 Eggenstein-Leopoldshafen (Germany)

    2011-08-15

    thickness of inner oxide layer reaches 20% of that of the outer oxide layer. The measured hydrogen release during the QUENCH-15 test was 41 g in the pre-oxidation and transient phases and 7 g in the quench phase which are comparable with those in QUENCH-06, i.e. 32 g and 4 g, respectively. Post-test calculations were performed using a version of SCDAP/RELAP5/MOD3.2. The calculation results support the heuristic observation that there was no major difference between the influence of Zircaloy-4, M5 or ZIRLO{sup TM} for the beyond-design basis accident present conditions here studied.

  13. Minichromosome replication in vitro: inhibition of re-replication by replicatively assembled nucleosomes.

    Science.gov (United States)

    Krude, T; Knippers, R

    1994-08-19

    Single-stranded circular DNA, containing the SV40 origin sequence, was used as a template for complementary DNA strand synthesis in cytosolic extracts from HeLa cells. In the presence of the replication-dependent chromatin assembly factor CAF-1, defined numbers of nucleosomes were assembled during complementary DNA strand synthesis. These minichromosomes were then induced to semiconservatively replicate by the addition of the SV40 initiator protein T antigen (re-replication). The results indicate that re-replication of minichromosomes appears to be inhibited by two independent mechanisms. One acts at the initiation of minichromosome re-replication, and the other affects replicative chain elongation. To directly demonstrate the inhibitory effect of replicatively assembled nucleosomes, two types of minichromosomes were prepared: (i) post-replicative minichromosomes were assembled in a reaction coupled to replication as above; (ii) pre-replicative minichromosomes were assembled independently of replication on double-stranded DNA. Both types of minichromosomes were used as templates for DNA replication under identical conditions. Replicative fork movement was found to be impeded only on post-replicative minichromosome templates. In contrast, pre-replicative minichromosomes allowed one unconstrained replication cycle, but re-replication was inhibited due to a block in fork movement. Thus, replicatively assembled chromatin may have a profound influence on the re-replication of DNA.

  14. Replication Research and Special Education

    Science.gov (United States)

    Travers, Jason C.; Cook, Bryan G.; Therrien, William J.; Coyne, Michael D.

    2016-01-01

    Replicating previously reported empirical research is a necessary aspect of an evidence-based field of special education, but little formal investigation into the prevalence of replication research in the special education research literature has been conducted. Various factors may explain the lack of attention to replication of special education…

  15. Replication Research and Special Education

    Science.gov (United States)

    Travers, Jason C.; Cook, Bryan G.; Therrien, William J.; Coyne, Michael D.

    2016-01-01

    Replicating previously reported empirical research is a necessary aspect of an evidence-based field of special education, but little formal investigation into the prevalence of replication research in the special education research literature has been conducted. Various factors may explain the lack of attention to replication of special education…

  16. Database Replication

    CERN Document Server

    Kemme, Bettina

    2010-01-01

    Database replication is widely used for fault-tolerance, scalability and performance. The failure of one database replica does not stop the system from working as available replicas can take over the tasks of the failed replica. Scalability can be achieved by distributing the load across all replicas, and adding new replicas should the load increase. Finally, database replication can provide fast local access, even if clients are geographically distributed clients, if data copies are located close to clients. Despite its advantages, replication is not a straightforward technique to apply, and

  17. Role of previous hospitalization in clinically-significant MRSA infection among HIV-infected inpatients: results of a case-control study

    Directory of Open Access Journals (Sweden)

    Festa Anna

    2007-04-01

    Full Text Available Abstract Background HIV-infected subjects have high incidence rates of Staphylococcus aureus infections, with both methicillin-susceptible and methicillin-resistant (MRSA strains. Possible explanations could include the high burden of colonization, the behavioral risk factors, and the frequent exposures to health care facilities of HIV-infected patients. The purpose of the study was to assess the risk factors for clinically- significant methicillin-resistant Staphylococcus aureus (CS-MRSA infections in HIV-infected patients admitted to Infectious Diseases Units. Methods From January 1, 2002 to December 31, 2005, we conducted a retrospective case-control (1:2 study. We identified all the cases of CS-MRSA infections in HIV-infected patients admitted to the National Institute for Infectious Diseases (INMI "Lazzaro Spallanzani" in the 4-year study period. A conditional logistic regression model was used to identify risk factors for CS-MRSA infection. Results We found 27 CS-MRSA infections, i.e. 0.9 CS-MRSA infections per 100 HIV-infected individuals cared for in our Institute. At multivariate analysis, independent predictors of CS-MRSA infection were cumulative hospital stay, invasive procedures in the previous year, and low CD4 cell count. Particularly, the risk for CS-MRSA increased by 14% per an increase of 5 days hospitalization in the previous year. Finally, we identified a low frequency of community-acquired MRSA infections (only 1 of 27; 3.7% among HIV-infected patients. Conclusion Clinicians should be aware of the risk for CS-MRSA infection in the clinical management of HIV-infected patients, especially in those patients with a low CD4 cell count, longer previous hospital stay, and previous invasive procedures.

  18. Role of previous hospitalization in clinically-significant MRSA infection among HIV-infected inpatients: results of a case-control study

    Science.gov (United States)

    Drapeau, Cecilia MJ; Angeletti, Claudio; Festa, Anna; Petrosillo, Nicola

    2007-01-01

    Background HIV-infected subjects have high incidence rates of Staphylococcus aureus infections, with both methicillin-susceptible and methicillin-resistant (MRSA) strains. Possible explanations could include the high burden of colonization, the behavioral risk factors, and the frequent exposures to health care facilities of HIV-infected patients. The purpose of the study was to assess the risk factors for clinically- significant methicillin-resistant Staphylococcus aureus (CS-MRSA) infections in HIV-infected patients admitted to Infectious Diseases Units. Methods From January 1, 2002 to December 31, 2005, we conducted a retrospective case-control (1:2) study. We identified all the cases of CS-MRSA infections in HIV-infected patients admitted to the National Institute for Infectious Diseases (INMI) "Lazzaro Spallanzani" in the 4-year study period. A conditional logistic regression model was used to identify risk factors for CS-MRSA infection. Results We found 27 CS-MRSA infections, i.e. 0.9 CS-MRSA infections per 100 HIV-infected individuals cared for in our Institute. At multivariate analysis, independent predictors of CS-MRSA infection were cumulative hospital stay, invasive procedures in the previous year, and low CD4 cell count. Particularly, the risk for CS-MRSA increased by 14% per an increase of 5 days hospitalization in the previous year. Finally, we identified a low frequency of community-acquired MRSA infections (only 1 of 27; 3.7%) among HIV-infected patients. Conclusion Clinicians should be aware of the risk for CS-MRSA infection in the clinical management of HIV-infected patients, especially in those patients with a low CD4 cell count, longer previous hospital stay, and previous invasive procedures. PMID:17470274

  19. Predicting Psychiatric Rehabilitation Outcome Using Demographic Characteristics: A Replication

    Science.gov (United States)

    Anthony, William A.; Buell, Gregory J.

    1974-01-01

    Replication was undertaken of a recent study conducted by Buell and Anthony which had found that recidivism and posthospital employment could be predicted by a single demographic variable, number of previous hospitalizations and employment history, respectively. Results of the replication were consistent for posthospital employment but not for…

  20. Perfectionism in Gifted Adolescents: A Replication and Extension

    Science.gov (United States)

    Margot, Kelly C.; Rinn, Anne N.

    2016-01-01

    To provide further generalizability for the results garnered by two previous studies, the authors conducted a methodological replication. In addition to adding to the body of replication research done with gifted students, the purpose of this study was to examine perfectionism differences among gifted adolescents in regards to gender, birth order,…

  1. Risk for High Depressive Symptoms in Diagnosed and Previously Undetected Diabetes: 5-Year Follow-Up Results of the Heinz Nixdorf Recall Study

    Science.gov (United States)

    Icks, Andrea; Albers, Bernd; Haastert, Burkhard; Pechlivanis, Sonali; Pundt, Noreen; Slomiany, Uta; Erbel, Raimund; Jöckel, Karl-Heinz; Kruse, Johannes; Kulzer, Bernd; Nowotny, Bettina; Herder, Christian; Giani, Guido; Moebus, Susanne

    2013-01-01

    Objective The objective of this study was to determine the risk for the development of high depressive symptoms in study participants with diagnosed and previously undetected diabetes mellitus compared to those without diabetes in a prospective population-based cohort study in Germany. Methods We estimated the 5-year cumulative incidence of high depressive symptoms in participants without high depressive symptoms at baseline (n = 3,633, 51.4% men, mean age (SD) 59.1 (7.6) years, 7.0% diagnosed diabetes, 5.3% previously undetected diabetes) from the population-based Heinz Nixdorf Recall study. Diabetes was assessed by self-report, medication, and blood glucose. High depressive symptoms were assessed using CES-D. We calculated odds ratios and their corresponding 95% confidence interval, using multiple logistic regression analyses. Result Cumulative 5-year incidences (95% CI) of high depressive symptoms in participants with diagnosed, undetected, and without diabetes were 7.1 (4.2–10.9), 4.1 (1.8–8.0), and 6.5 (5.6–7.4), respectively. The age-sex-adjusted OR for developing high depressive symptoms was 1.22 (0.74–2.03) in participants with diagnosed compared to those without diabetes, and 1.00 (0.59–1.68) after adjustment for BMI, physical activity, education, stroke, and myocardial infarction. The age-sex adjusted OR for developing high depressive symptoms in participants with previously undetected diabetes compared to those without diabetes was 0.72; 0.35–1.48; and fully adjusted 0.62; 0.30–1.30. Conclusion We found no significant associations, maybe due to low power. However, our results are in line with a recent meta-analysis suggesting that risk of developing high depressive symptoms in patients with diagnosed diabetes may be moderately higher than in those without diabetes, and that comorbidity may explain in part this association. In participants with previously undetected diabetes, this first longitudinal study indicates that the risk is not

  2. Risk for high depressive symptoms in diagnosed and previously undetected diabetes: 5-year follow-up results of the Heinz Nixdorf Recall study.

    Directory of Open Access Journals (Sweden)

    Andrea Icks

    Full Text Available OBJECTIVE: The objective of this study was to determine the risk for the development of high depressive symptoms in study participants with diagnosed and previously undetected diabetes mellitus compared to those without diabetes in a prospective population-based cohort study in Germany. METHODS: We estimated the 5-year cumulative incidence of high depressive symptoms in participants without high depressive symptoms at baseline (n = 3,633, 51.4% men, mean age (SD 59.1 (7.6 years, 7.0% diagnosed diabetes, 5.3% previously undetected diabetes from the population-based Heinz Nixdorf Recall study. Diabetes was assessed by self-report, medication, and blood glucose. High depressive symptoms were assessed using CES-D. We calculated odds ratios and their corresponding 95% confidence interval, using multiple logistic regression analyses. RESULT: Cumulative 5-year incidences (95% CI of high depressive symptoms in participants with diagnosed, undetected, and without diabetes were 7.1 (4.2-10.9, 4.1 (1.8-8.0, and 6.5 (5.6-7.4, respectively. The age-sex-adjusted OR for developing high depressive symptoms was 1.22 (0.74-2.03 in participants with diagnosed compared to those without diabetes, and 1.00 (0.59-1.68 after adjustment for BMI, physical activity, education, stroke, and myocardial infarction. The age-sex adjusted OR for developing high depressive symptoms in participants with previously undetected diabetes compared to those without diabetes was 0.72; 0.35-1.48; and fully adjusted 0.62; 0.30-1.30. CONCLUSION: We found no significant associations, maybe due to low power. However, our results are in line with a recent meta-analysis suggesting that risk of developing high depressive symptoms in patients with diagnosed diabetes may be moderately higher than in those without diabetes, and that comorbidity may explain in part this association. In participants with previously undetected diabetes, this first longitudinal study indicates that the risk is not

  3. Bomb-Pulse Chlorine-36 At The Proposed Yucca Mountain Repository Horizon: An Investigation Of Previous Conflicting Results And Collection Of New Data

    Energy Technology Data Exchange (ETDEWEB)

    J. Cizdziel

    2006-07-28

    Previous studies by scientists at Los Alamos National Laboratory (LANL) found elevated ratios of chlorine-36 to total chloride ({sup 36}Cl/Cl) in samples of rock collected from the Exploratory Studies Facility (ESF) and the Enhanced Characterization of the Repository Block (ECRB) at Yucca Mountain as the tunnels were excavated. The data were interpreted as an indication that fluids containing 'bomb-pulse' {sup 36}Cl reached the repository horizon in the {approx}50 years since the peak period of above-ground nuclear testing. Moreover, the data support the concept that so-called fast pathways for infiltration not only exist but are active, possibly through a combination of porous media, faults and/or other geologic features. Due to the significance of {sup 36}Cl data to conceptual models of unsaturated zone flow and transport, the United States Geological Survey (USGS) was requested by the Department of Energy (DOE) to design and implement a study to validate the LANL findings. The USGS chose to drill new boreholes at select locations across zones where bomb-pulse ratios had previously been identified. The drill cores were analyzed at Lawrence Livermore National Laboratory (LLNL) for {sup 36}Cl/Cl using both active and passive leaches, with the USGS/LLNL concluding that the active leach extracted too much rock-Cl and the passive leach did not show bomb-pulse ratios. Because consensus was not reached between the USGS/LLNL and LANL on several fundamental points, including the conceptual strategy for sampling, interpretation and use of tritium ({sup 3}H) data, and the importance and interpretation of blanks, in addition to the presence or absence of bomb-pulse {sup 36}Cl, an evaluation by an independent entity, the University of Nevada, Las Vegas (UNLV), using new samples was initiated. This report is the result of that study. The overall objectives of the UNLV study were to investigate the source or sources of the conflicting results from the previous

  4. LHCb Data Replication During SC3

    CERN Multimedia

    Smith, A

    2006-01-01

    LHCb's participation in LCG's Service Challenge 3 involves testing the bulk data transfer infrastructure developed to allow high bandwidth distribution of data across the grid in accordance with the computing model. To enable reliable bulk replication of data, LHCb's DIRAC system has been integrated with gLite's File Transfer Service middleware component to make use of dedicated network links between LHCb computing centres. DIRAC's Data Management tools previously allowed the replication, registration and deletion of files on the grid. For SC3 supplementary functionality has been added to allow bulk replication of data (using FTS) and efficient mass registration to the LFC replica catalog.Provisional performance results have shown that the system developed can meet the expected data replication rate required by the computing model in 2007. This paper details the experience and results of integration and utilisation of DIRAC with the SC3 transfer machinery.

  5. Relationship between mental disorders/suicidality and three sexual behaviors: results from the National Comorbidity Survey Replication.

    Science.gov (United States)

    Mota, Natalie P; Cox, Brian J; Katz, Laurence Y; Sareen, Jitender

    2010-06-01

    The present study examined the relationship between sexual behaviors and mental disorders and suicidality in the National Comorbidity Survey Replication, a representative sample of adults ages 18 years and older (N = 5,692). The World Health Organization Composite International Diagnostic Interview was used to make DSM-IV based disorder diagnoses. Participants were also asked about suicidality and sexual behaviors. Multiple logistic regression analyses adjusted for sociodemographic variables were used to examine the relationships of three sexual behaviors (age of first intercourse, number of past year partners, and past year condom use) with 15 mental disorders (clustered into any mood, anxiety, substance use, and disruptive behavior groups) and suicidality (ideation and attempts). Compared to ages 15-17, those with age of first intercourse between 12 and 14 had increased rates of lifetime disruptive behavior, substance use, and any mental disorder, and suicidal ideation and attempts (adjusted odds ratio (AOR) range, 1.46-2.01). Those with age of first intercourse between ages 18-25 and 26-35 were at decreased likelihood of several lifetime disorder groups (AOR range, 0.19-0.81). Individuals who had two or more sexual partners in the past year had increased rates of all past year disorder groups examined (AOR range, 1.44-5.01). Never married participants who rarely/never used condoms were more likely than those who always used condoms to experience any mood, substance use, and any mental disorder, and suicide attempts (AOR range, 1.77-8.13). Future research should longitudinally examine these associations and account better for possible familial and personality confounders.

  6. [The HBeAg/antiHBe seroconversion as a result of lamivudine treatment in children with chronic hepatitis B unresponsive to previous interferon alpha therapy].

    Science.gov (United States)

    Lebensztejn, Dariusz Marek; Skiba, Elzbieta; Sobaniec-Lotowska, Maria; Kaczmarski, Maciej

    2004-06-01

    The aim of the study was evaluation the HBeAg/antiHBe seroconversion frequency as a result of lamivudine treatment in children who are nonresponders to previous IFN-alpha therapy. The observation was carried out on 41 children, aged 4-17 years, with biopsy-proven chronic hepatitis B (HBeAg+) treated with lamivudine 3-4 mg/kg/d (max. 100 mg/d) for 12 months. After 6 months of lamivudine therapy 59.3% children normalized GPT activity and only 1 child (2.5%) lost HBeAg. At the end of 12 months of therapy 81.5% normalized GPT activity and 4 of them (10%) lost HBeAg and seroconverted to antiHBe. None of treated children lost HBsAg. The age, sex, pretreatment GPT activity and active histological disease were not predictors of lamivudine-induced HBeAg loss. There were no side effects of lamivudine therapy except one boy who had severe thrombocytopenia. The HBeAg/antiHBe seroconversion rate after one year trial of lamivudine in children with chronic hepatitis B unresponsive to previous IFN alpha therapy was 10%. The age, sex, pretreatment GPT activity and active histologic disease were not predictors of lamivudine-induced HBeAg loss.

  7. The Role of Core Needle Biopsy for Thyroid Nodules with Initially Indeterminate Results on Previous Fine-Needle Aspiration: A Systematic Review and Meta-Analysis.

    Science.gov (United States)

    Suh, C H; Baek, J H; Park, C; Choi, Y J; Lee, J H

    2017-07-01

    Sonography-guided fine-needle aspiration leads to relatively frequent cases of indeterminate cytology for the diagnosis of thyroid nodules. Our aim was to evaluate the efficacy and safety of core needle biopsy for the examination of thyroid nodules with initially indeterminate results on fine-needle aspiration. A computerized search of the MEDLINE and Embase databases was performed to identify relevant original articles. Studies investigating the use of core needle biopsy for thyroid nodules with initially indeterminate results on previous fine-needle aspiration were eligible for inclusion. The pooled proportions for nondiagnostic results, inconclusive results, malignancy on core needle biopsy, the ability of core needle biopsy to diagnose malignancy, and the related complications of the procedure were analyzed. The meta-analytic pooling was based on a random-effects model. Nine eligible studies, involving 2240 patients with 2245 thyroid nodules, were included. The pooled proportion for nondiagnostic results was 1.8% (95% CI, 0.4%-3.2%), and the pooled proportion for inconclusive results was 25.1% (95% CI, 15.4%-34.9%). The pooled proportion for malignancy was 18.9% (95% CI, 8.4%-29.5%). With regard to the diagnostic performance for malignancy, the sensitivity of core needle biopsy varied, ranging from 44.7% to 85.0%, but the specificity was 100% in all cases. No major complications of core needle biopsy were observed. The relatively small number of included studies and retrospective nature were limitations. Core needle biopsy has low nondiagnostic result rates and high specificity for the diagnosis of malignancy. It is a safe diagnostic technique with a higher diagnostic yield, especially when molecular testing is not available or fine-needle aspiration did not yield enough cells for molecular testing. © 2017 by American Journal of Neuroradiology.

  8. Replication data collection highlights value in diversity of replication attempts

    Science.gov (United States)

    DeSoto, K. Andrew; Schweinsberg, Martin

    2017-01-01

    Researchers agree that replicability and reproducibility are key aspects of science. A collection of Data Descriptors published in Scientific Data presents data obtained in the process of attempting to replicate previously published research. These new replication data describe published and unpublished projects. The different papers in this collection highlight the many ways that scientific replications can be conducted, and they reveal the benefits and challenges of crucial replication research. The organizers of this collection encourage scientists to reuse the data contained in the collection for their own work, and also believe that these replication examples can serve as educational resources for students, early-career researchers, and experienced scientists alike who are interested in learning more about the process of replication. PMID:28291224

  9. Autocatalytic replication and homochirality in biopolymers: is homochirality a requirement of life or a result of it?

    Science.gov (United States)

    Wu, Meng; Walker, Sara I; Higgs, Paul G

    2012-09-01

    A key step in the origin of life is the establishment of autocatalytic cycles controlled by biopolymer catalysts. These catalysts (either ribozymes or proteins) are composed of homochiral monomers. Homochirality in living systems is maintained because biopolymers are asymmetric in their catalysis and synthesize molecules of their own handedness. Asymmetric autocatalysis is also possible with small molecules, as demonstrated by the Soai reaction, but it is rare. As far as we know, single nucleotides and amino acids are not autocatalytic. The observation that organic molecules in meteorites can have an enantiomeric excess of a few percent suggests that the prebiotic mixture may have had a partial chiral bias that was caused by external physical influences. Here, we consider the way that such a partial prebiotic bias would influence the origin of ribozymes in an RNA world scenario. We have previously shown how a transition to a living state can occur in a model for RNA polymerization. Here, we add chirality to the problem by considering simultaneous synthesis and polymerization of left- and right-handed monomers. The two chemical synthesis rates may be equal or unequal, due to physical or chemical effects prior to the origin of life. We determine the stationary states of this reaction system. The nonliving state is racemic, or slightly biased. There are two living states that are almost completely homochiral, whether or not the nonliving state is biased. It is a feature of our model that, for some regions of parameter space, living and nonliving states are both found to be stable under the same conditions. The origin of life therefore involves a stochastic transition between the nonliving and living states. Our model extends previous theories by treating the origin of life and the origin of chirality as aspects of the same model.

  10. Replication studies in longevity

    DEFF Research Database (Denmark)

    Varcasia, O; Garasto, S; Rizza, T

    2001-01-01

    In Danes we replicated the 3'APOB-VNTR gene/longevity association study previously carried out in Italians, by which the Small alleles (less than 35 repeats) had been identified as frailty alleles for longevity. In Danes, neither genotype nor allele frequencies differed between centenarians and 20...

  11. Thirty-five-year results after Charnley total hip arthroplasty in patients less than fifty years old. A concise follow-up of previous reports.

    Science.gov (United States)

    Warth, Lucian C; Callaghan, John J; Liu, Steve S; Klaassen, Alison L; Goetz, Devon D; Johnston, Richard C

    2014-11-05

    We report the updated results for a previously described cohort of patients who were less than fifty years old at the time of the index Charnley total hip arthroplasty with cement. The original cohort consisted of ninety-three consecutive hips in sixty-nine patients. The patients were followed for a minimum of thirty-five years after surgery or until death. At the latest follow-up evaluation, there were forty-one total hip replacements (44%) in thirty-two living patients. Thirty-four (37%) of the ninety-three total hip replacements in the original cohort had been revised or removed. Twenty acetabular (22%) and seven femoral (8%) components had been revised for aseptic loosening. Since the twenty-five-year follow-up, the average six-minute-walk distance decreased from 395 m to 171 m, and this decrease correlated with increasing comorbidity. This study demonstrates the durability of cemented total hip replacements in a young patient population. Although 63% (fifty-nine) of the ninety-three original hip replacements were functioning at the latest follow-up or at the time of death, a significant decrease in activity level was seen over time (p < 0.001). Of the forty-one original implants in the patients who were alive at the time of the thirty-five-year follow-up, only 46% (nineteen) were retained. Copyright © 2014 by The Journal of Bone and Joint Surgery, Incorporated.

  12. Late tamoxifen in patients previously operated for breast cancer without postoperative tamoxifen: 5-year results of a single institution randomised study

    Directory of Open Access Journals (Sweden)

    Bidoli Ettore

    2010-05-01

    Full Text Available Abstract Background A population of breast cancer patients exists who, for various reasons, never received adjuvant post-operative tamoxifen (TAM. This study was aimed to evaluate the role of late TAM in these patients. Methods From 1997 to 2003, patients aged 35 to 75 years, operated more than 2 years previously for monolateral breast cancer without adjuvant TAM, with no signs of metastases and no contraindication to TAM were randomized to TAM 20 mg/day orally for 2 years or follow-up alone. Events were categorized as locoregional relapse, distant metastases, metachronous breast cancer, tumours other than breast cancer and death from any causes, whichever occurred first. The sample size (197 patients per arm, plus 10% allowance was based on the assumption of a 30% decrease in the number of events occurring at a rate of 5% annually in the 10 years following randomization. Four hundred and thirty-three patients were randomized in the study (TAM 217, follow-up 216. Patients characteristics (TAM/follow-up included: median age 55/55 years, median time from surgery 25/25 months (range, 25-288/25-294, in situ carcinoma 18/24, oestrogen receptor (ER positive in 75/68, negative in 70/57, unknown in 72/91 patients. Previous adjuvant treatment included chemotherapy in 131/120 and an LHRH analogue in 11/13 patients. Results Thirty-six patients prematurely discontinued TAM after a median of 1 month, mostly because of subjective intolerance. Eighty-three events (TAM 39, follow-up 44 occurred: locoregional relapse in 10/8, distant metastases in 14/16, metachronous breast cancer in 4/10, other tumours in 11/10 patients. Less ER-positive secondary breast cancers occurred in the TAM treated patients than in follow-up patients (1 vs 10, p = 0.005. Event-free survival was similar in both groups of patients. Conclusions This 5-year analysis revealed significantly less metachronous ER-positive breast cancers in the TAM treated patients. No other statistically

  13. Abiotic self-replication.

    Science.gov (United States)

    Meyer, Adam J; Ellefson, Jared W; Ellington, Andrew D

    2012-12-18

    The key to the origins of life is the replication of information. Linear polymers such as nucleic acids that both carry information and can be replicated are currently what we consider to be the basis of living systems. However, these two properties are not necessarily coupled. The ability to mutate in a discrete or quantized way, without frequent reversion, may be an additional requirement for Darwinian evolution, in which case the notion that Darwinian evolution defines life may be less of a tautology than previously thought. In this Account, we examine a variety of in vitro systems of increasing complexity, from simple chemical replicators up to complex systems based on in vitro transcription and translation. Comparing and contrasting these systems provides an interesting window onto the molecular origins of life. For nucleic acids, the story likely begins with simple chemical replication, perhaps of the form A + B → T, in which T serves as a template for the joining of A and B. Molecular variants capable of faster replication would come to dominate a population, and the development of cycles in which templates could foster one another's replication would have led to increasingly complex replicators and from thence to the initial genomes. The initial genomes may have been propagated by RNA replicases, ribozymes capable of joining oligonucleotides and eventually polymerizing mononucleotide substrates. As ribozymes were added to the genome to fill gaps in the chemistry necessary for replication, the backbone of a putative RNA world would have emerged. It is likely that such replicators would have been plagued by molecular parasites, which would have been passively replicated by the RNA world machinery without contributing to it. These molecular parasites would have been a major driver for the development of compartmentalization/cellularization, as more robust compartments could have outcompeted parasite-ridden compartments. The eventual outsourcing of metabolic

  14. Mirror activity in the human brain while observing hand movements: a comparison between EEG desynchronization in the mu-range and previous fMRI results.

    Science.gov (United States)

    Perry, Anat; Bentin, Shlomo

    2009-07-28

    Mu (mu) rhythms are EEG oscillations between 8-13 Hz distinguished from alpha by having more anterior distribution and being desynchronized by motor rather than visual activity. Evidence accumulating during the last decade suggests that the desynchronization of mu rhythms (mu suppression) might be also a manifestation of a human Mirror Neuron System (MNS). To further explore this hypothesis we used a paradigm that, in a previous fMRI study, successfully activated this putative MNS in humans. Our direct goal was to provide further support for a link between modulation of mu rhythms and the MNS, by finding parallels between the reported patterns of fMRI activations and patterns of mu suppression. The EEG power in the mu range has been recorded while participants passively observed either a left or a right hand, reaching to and grasping objects, and compared it with that recorded while participants observed the movement of a ball, and while observing static grasping scenes or still objects. Mirroring fMRI results (Shmuelof, L., Zohary, E., 2005. Dissociation between ventral and dorsal fMRI activation during object and action recognition. Neuron 47, 457-470), mu suppression was larger in the hemisphere contra-lateral to the moving hand and larger when the hands grasped different objects in different ways than when the movement was repetitive. No suppression was found while participants observed still objects but mu suppression was also found while seeing static grasping postures. These data are discussed in light of similar parallels between modulations of alpha waves and fMRI while recording EEG in the magnet. The present data support a link between mu suppression and a human MNS.

  15. Effect of posterior multilevel vertebral osteotomies on coronal and sagittal balance in fused scoliosis deformity caused by previous surgery: preliminary results.

    Science.gov (United States)

    Yang, Jae Hyuk; Suh, Seung Woo; Cho, Won Tae; Hwang, Jin Ho; Hong, Jae Young; Modi, Hitesh N

    2014-10-15

    Prospective case series study. To study the effect of posterior multilevel vertebral osteotomy (posterior crack osteotomy) on coronal and sagittal balance in patients with the fusion mass over the spine caused by previous surgery. Few studies have investigated revisional scoliosis surgery with the fusion mass using osteotomy. Among patients who had a history of prior surgery for scoliosis correction and posterior fusion, those showing progression of the curve postoperatively due to nonunion, implant failure, or adding-on phenomenon were enrolled. All patients were treated using posterior crack osteotomy. For clinical evaluation, the pre- and postoperative Gross Motor Function Classification System score for walking status and the Berg balanced scale were used. For radiological evaluation, pre- and postoperative Cobb angle, and coronal and sagittal balance factors were used. Ten patients (5 males and 5 females) were enrolled. The preoperative diagnosis was neuromuscular scoliosis (3 cases), syndromic scoliosis (1 case), congenital scoliosis (5 cases), and neurofibromatosis (1 case). Osteotomies were performed at 3.3±1.3 levels on average. Pre- and postoperative Cobb angles were 70.8°±30.0° and 28.1°±20.0° (P=0.002 (0.97)), respectively. In pre- and postoperative evaluation of coronal balance, the coronal balance, clavicle angle, and T1-tilt angle were 36.8±27.1 mm and 10.4±8.5 mm, 6.7°±8.0° and 3.3°±1.5°, and 7.8°±19.0° and 4.7°±2.1°, respectively (P=0.002, 0.002, 0.002). In pre- and postoperative evaluation of sagittal balance, the spinal vertical axis, thoracic kyphosis, and lumbar alignments were 25.1±37.8 mm and 14.1±21.8 mm, 33.5°±51.1° and 29.7°±27.4°, and 45.7°±34.8° and 48.9°±23.1° (P=0.002, 0.169, 0.169). The walking and functional statuses did not change (P=0.317, 0.932). Although pulmonary and gastrointestinal complications were noted, the patients were discharged without complications. Posterior crack osteotomy can be

  16. Office and ambulatory blood pressure control with a fixed-dose combination of candesartan and hydrochlorothiazide in previously uncontrolled hypertensive patients: results of CHILI CU Soon

    Directory of Open Access Journals (Sweden)

    Bramlage P

    2011-12-01

    Full Text Available Thomas Mengden1, Reinhold Hübner2, Peter Bramlage31Kerckhoff-Klinik GmbH, Bad Nauheim, 2Takeda Pharma GmbH, Aachen, 3Institut für Kardiovaskuläre Pharmakologie und Epidemiologie, Mahlow, GermanyBackground: Fixed-dose combinations of candesartan 32 mg and hydrochlorothiazide (HCTZ have been shown to be effective in clinical trials. Upon market entry we conducted a noninterventional study to document the safety and effectiveness of this fixed-dose combination in an unselected population in primary care and to compare blood pressure (BP values obtained during office measurement (OBPM with ambulatory blood pressure measurement (ABPM.Methods: CHILI CU Soon was a prospective, noninterventional, noncontrolled, open-label, multicenter study with a follow-up of at least 10 weeks. High-risk patients aged ≥18 years with previously uncontrolled hypertension were started on candesartan 32 mg in a fixed-dose combination with either 12.5 mg or 25 mg HCTZ. OBPM and ABPM reduction and adverse events were documented.Results: A total of 4131 patients (52.8% male with a mean age of 63.0 ± 11.0 years were included. BP was 162.1 ± 14.8/94.7 ± 9.2 mmHg during office visits at baseline. After 10 weeks of candesartan 32 mg/12.5 mg or 25 mg HCTZ, mean BP had lowered to 131.7 ± 10.5/80.0 ± 6.6 mmHg (P < 0.0001 for both comparisons. BP reduction was comparable irrespective of prior or concomitant medication. In patients for whom physicians regarded an ABPM to be necessary (because of suspected noncontrol over 24 hours, ABP at baseline was 158.2/93.7 mmHg during the day and 141.8/85.2 mmHg during the night. At the last visit, BP had significantly reduced to 133.6/80.0 mmHg and 121.0/72.3 mmHg, respectively, resulting in 20.8% being normotensive over 24 hours (<130/80 mmHg. The correlation between OBPM and ABPM was good (r = 0.589 for systolic BP and r = 0.389 for diastolic BP during the day. Of those who were normotensive upon OBPM, 35.1% had high ABPM during the

  17. Absence of Non-histone Protein Complexes at Natural Chromosomal Pause Sites Results in Reduced Replication Pausing in Aging Yeast Cells

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    Marleny Cabral

    2016-11-01

    Full Text Available There is substantial evidence that genomic instability increases during aging. Replication pausing (and stalling at difficult-to-replicate chromosomal sites may induce genomic instability. Interestingly, in aging yeast cells, we observed reduced replication pausing at various natural replication pause sites (RPSs in ribosomal DNA (rDNA and non-rDNA locations (e.g., silent replication origins and tRNA genes. The reduced pausing occurs independent of the DNA helicase Rrm3p, which facilitates replication past these non-histone protein-complex-bound RPSs, and is independent of the deacetylase Sir2p. Conditions of caloric restriction (CR, which extend life span, also cause reduced replication pausing at the 5S rDNA and at tRNA genes. In aged and CR cells, the RPSs are less occupied by their specific non-histone protein complexes (e.g., the preinitiation complex TFIIIC, likely because members of these complexes have primarily cytosolic localization. These conditions may lead to reduced replication pausing and may lower replication stress at these sites during aging.

  18. Dynamics of Escherichia coli Chromosome Segregation during Multifork Replication

    DEFF Research Database (Denmark)

    Nielsen, Henrik Jørck; Youngren, Brenda; Hansen, Flemming G.

    2007-01-01

    Slowly growing Escherichia coli cells have a simple cell cycle, with replication and progressive segregation of the chromosome completed before cell division. In rapidly growing cells, initiation of replication occurs before the previous replication rounds are complete. At cell division...

  19. Drug-Induced QT Prolongation as a Result of an Escitalopram Overdose in a Patient with Previously Undiagnosed Congenital Long QT Syndrome

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    Paul Singh

    2014-01-01

    Full Text Available We present a case of drug-induced QT prolongation caused by an escitalopram overdose in a patient with previously undiagnosed congenital LQTS. A 15-year-old Caucasian female presented following a suicide attempt via an escitalopram overdose. The patient was found to have a prolonged QT interval with episodes of torsades de pointes. The patient was admitted to the telemetry unit and treated. Despite the resolution of the torsades de pointes, she continued to demonstrate a persistently prolonged QT interval. She was seen by the cardiology service and diagnosed with congenital long QT syndrome. This case illustrates the potential for an escitalopram overdose to cause an acute QT prolongation in a patient with congenital LQTS and suggests the importance of a screening electrocardiogram prior to the initiation of SSRIs, especially in patients at high risk for QT prolongation.

  20. The influence of previous infections and antichlamydia pneumoniae seropositivity on functional outcome in ischemic stroke patients: results from the IN2 study.

    Science.gov (United States)

    Consoli, Domenico; Vidale, Simone; Aguglia, Umberto; Bassi, Pietro; Cavallini, Anna; Consoli, Arturo; Galati, Franco; Guidetti, Donata; Micieli, Giuseppe; Neri, Giuseppe; Rasura, Maurizia; Sterzi, Roberto; Toni, Danilo; Inzitari, Domenico

    2015-05-01

    Although a growing attention is being paid to acute ischemic stroke patients, the correlation between clinical outcome and infectious events in this population has been poorly investigated. 749 ischemic stroke (mean age 71 years old, males 56%) patients were enrolled in this prospective case-control study by 11 Italian Stroke Units. Demographic data, vascular risk factors, previous infections and post-stroke infections (PSIs) were recorded. Blood samples were collected and the enzyme-linked immunoassay was chosen to measure Chlamydia pneumoniae IgG and IgA plasma antibodies (antibody titers were classified with specific cut-off levels: IgA > 1:16 and IgG > 1:64). Early poor outcome was defined as mRS score >2 at discharge, while poor outcome at 6-month follow-up. Univariate and multivariate analyses were performed. Median NIHSS was 7, IgA and IgG antichlamydia pneumoniae seropositivities were observed in 308 (37.1%) and 207 (23.6%) patients, respectively. Multivariate analyses showed significant correlations between PSIs and NIHSS (RR: 1.06; 95% CI 1.02-1.09; p < 0.001) and PSIs and IgA antichlamydia pneumoniae seropositivity (RR: 3.84; 95% CI 2.53-5.84; p < 0.001). Significant disability was associated with baseline NIHSS (RR: 1.32; 95% CI 1.16-1.50; p < 0.001), IgA (RR: 2.67; 95% CI 1.06-6.70; p = 0.035) and IgG antichlamydia (RR: 5.75; 95% CI 1.83-18.03; p = 0.003) seropositivity and atrial fibrillation (RR: 2.58; 95% CI 1.81-3.67; p < 0.001). While previous infections were not associated with functional outcome, antichlamydia antibodies play a negative role in ischemic stroke patients. Preventive strategies may reduce the stroke burden and improve the clinical outcome.

  1. BAY 81-8973 safety and efficacy for prophylaxis and treatment of bleeds in previously treated children with severe haemophilia A: results of the LEOPOLD Kids Trial.

    Science.gov (United States)

    Ljung, R; Kenet, G; Mancuso, M E; Kaleva, V; Rusen, L; Tseneklidou-Stoeter, D; Michaels, L A; Shah, A; Hong, W; Maas Enriquez, M

    2016-05-01

    BAY 81-8973, a full-length, unmodified, recombinant factor VIII (FVIII) in development for treatment of haemophilia A, has the same primary amino acid sequence as Bayer's sucrose-formulated recombinant FVIII but is produced with more advanced manufacturing technologies. To demonstrate safety and efficacy of BAY 81-8973 for prophylaxis and treatment of bleeds in previously treated children. In this phase III, multicentre, open-label, nonrandomized study, boys aged ≤12 years with severe haemophilia A and ≥50 exposure days (EDs) to FVIII products received prophylaxis with BAY 81-8973 25-50 IU kg(-1) ≥2 times weekly for ≥50 EDs. The efficacy endpoint was annualized number of total bleeds. Adverse events (AEs) and immunogenicity were assessed. Fifty-one patients were treated (age: 2× per week (57%) regimen at study start. Median [quartile 1; quartile 3 (Q1; Q3)] annualized number of bleeds for the combined age groups was 1.90 (0; 6.02) for total bleeds, 0 (0; 2.01) for joint bleeds and 0 (0; 0) for spontaneous bleeds. Median (Q1; Q3) annualized number of total bleeds within 48 h of previous prophylaxis infusion was 1.88 (0; 3.97) for children aged <6 years and 0 (0; 1.96) for children aged 6-<12 years. No drug-related serious AEs or inhibitors were reported. Prophylaxis with BAY 81-8973 using individualized prophylaxis regimens of 2× per week, 3× per week and every-other-day infusions was efficacious in prevention and treatment of bleeds in children with severe haemophilia A. Treatment with BAY 81-8973 was well tolerated. © 2015 John Wiley & Sons Ltd.

  2. Lack of association of variants previously associated with anti-TNF medication response in rheumatoid arthritis patients: results from a homogeneous Greek population.

    Science.gov (United States)

    Zervou, Maria I; Myrthianou, Efsevia; Flouri, Irene; Plant, Darren; Chlouverakis, Gregory; Castro-Giner, Francesc; Rapsomaniki, Panayiota; Barton, Anne; Boumpas, Dimitrios T; Sidiropoulos, Prodromos; Goulielmos, George N

    2013-01-01

    Treatment strategies blocking tumor necrosis factor (anti-TNF) have proven very successful in patients with rheumatoid arthritis (RA), showing beneficial effects in approximately 50-60% of the patients. However, a significant subset of patients does not respond to anti-TNF agents, for reasons that are still unknown. The aim of this study was to validate five single nucleotide polymorphisms (SNPs) of PTPRC, CD226, AFF3, MyD88 and CHUK gene loci that have previously been reported to predict anti-TNF outcome. In addition, two markers of RA susceptibility, namely TRAF1/C5 and STAT4 were assessed, in a cohort of anti-TNF-treated RA patients, from the homogeneous Greek island of Crete, Greece. The RA patient cohort consisted of 183 patients treated with either of 3 anti-TNF biologic agents (infliximab, adalimumab and etanercept) from the Clinic of Rheumatology of the University Hospital of Crete. The SNPs were genotyped by TaqMan assays or following the Restriction Fragments Length Polymorphisms (RFLPs) approach. Disease activity score in 28 joints (DAS28) at baseline and after 6 months were available for all patients and analysis of good versus poor response at 6 months was performed for each SNP. None of the 7 genetic markers correlated with treatment response. We conclude that the gene polymorphisms under investigation are not strongly predictive of anti-TNF response in RA patients from Greece.

  3. Lack of association of variants previously associated with anti-TNF medication response in rheumatoid arthritis patients: results from a homogeneous Greek population.

    Directory of Open Access Journals (Sweden)

    Maria I Zervou

    Full Text Available Treatment strategies blocking tumor necrosis factor (anti-TNF have proven very successful in patients with rheumatoid arthritis (RA, showing beneficial effects in approximately 50-60% of the patients. However, a significant subset of patients does not respond to anti-TNF agents, for reasons that are still unknown. The aim of this study was to validate five single nucleotide polymorphisms (SNPs of PTPRC, CD226, AFF3, MyD88 and CHUK gene loci that have previously been reported to predict anti-TNF outcome. In addition, two markers of RA susceptibility, namely TRAF1/C5 and STAT4 were assessed, in a cohort of anti-TNF-treated RA patients, from the homogeneous Greek island of Crete, Greece. The RA patient cohort consisted of 183 patients treated with either of 3 anti-TNF biologic agents (infliximab, adalimumab and etanercept from the Clinic of Rheumatology of the University Hospital of Crete. The SNPs were genotyped by TaqMan assays or following the Restriction Fragments Length Polymorphisms (RFLPs approach. Disease activity score in 28 joints (DAS28 at baseline and after 6 months were available for all patients and analysis of good versus poor response at 6 months was performed for each SNP. None of the 7 genetic markers correlated with treatment response. We conclude that the gene polymorphisms under investigation are not strongly predictive of anti-TNF response in RA patients from Greece.

  4. Culture Negative Listeria monocytogenes Meningitis Resulting in Hydrocephalus and Severe Neurological Sequelae in a Previously Healthy Immunocompetent Man with Penicillin Allergy

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    Shahin Gaini

    2015-01-01

    Full Text Available A previously healthy 74-year-old Caucasian man with penicillin allergy was admitted with evolving headache, confusion, fever, and neck stiffness. Treatment for bacterial meningitis with dexamethasone and monotherapy ceftriaxone was started. The cerebrospinal fluid showed negative microscopy for bacteria, no bacterial growth, and negative polymerase chain reaction for bacterial DNA. The patient developed hydrocephalus on a second CT scan of the brain on the 5th day of admission. An external ventricular catheter was inserted and Listeria monocytogenes grew in the cerebrospinal fluid from the catheter. The patient had severe neurological sequelae. This case report emphasises the importance of covering empirically for Listeria monocytogenes in all patients with penicillin allergy with suspected bacterial meningitis. The case also shows that it is possible to have significant infection and inflammation even with negative microscopy, negative cultures, and negative broad range polymerase chain reaction in cases of Listeria meningitis. Follow-up spinal taps can be necessary to detect the presence of Listeria monocytogenes.

  5. Missing osteoconductive effect of a resorbable PEO/PBT copolymer in human bone defects: a clinically relevant pilot study with contrary results to previous animal studies.

    Science.gov (United States)

    Roessler, M; Wilke, A; Griss, P; Kienapfel, H

    2000-01-01

    PEO/PBT 70/30 (POLYACTIVE(R) 70/30), a degradable porous copolymer with elastic properties, was found to be osteoconductive in many animal studies. The aim of this study was to determine the osteoconductive effect in a human paired control iliac defect model. In seven patients undergoing anterior spinal interbody fusion surgery, two bicortical iliac defects for autograft harvesting were created. The defect size was identical for both defects measuring about 40 x 15 mm (group I). One defect was filled with the degradable implants, whereas the remaining one was left untreated as a control. The defect site for treatment was chosen randomly. In three further patients, only one defect measuring about 40 x 35 mm was created (group II). All patients were examined clinically and radiologically by spiral-CT after 1, 6, 12, 24, and 52 weeks. Three-dimensional reconstructions as well as CT-volumetric measurements using 1 mm sections were used as evaluation methods. In group I, a two-tailed paired t-test showed that the treated defects had significantly less formation of new bone than the untreated ones (p < 0.05 after 12 weeks, p < 0.01 after 52 weeks). Also, in group II, not much bone ingrowth could be observed. The histological evaluation of one patient in group I revealed no bone within the pores, and a fibrous layer between bone and implant was always present. Therefore, PEO/PBT 70/30 cannot be recommended as a bone substitute for clinical use. Differences in bone regeneration between humans and certain animal species as well as inapplicable defect models in previous animal studies are discussed as possible reasons for the failure.

  6. A comparison of results from previous and present investigations of benthic macroinvertebrates in the small and shallow Lake Shoji, Fuji Five Lakes, Japan

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    Kimio Hirabayashi

    2012-10-01

    Full Text Available In order to clarify the current status of the benthic community of Lake Shoji, and to examine the difference with the last quantitative data of the lake by Miyadi (1932 and Kitagawa (1973, the distribution of benthic macroinvertebrates in this lake was studied. Also, a comparison of the benthic fauna and density was made between the present and previous studies, with a discussion concerning the succession of benthic macroinvertebrates in relation to changes in the lake bottom environment. On March 1, 2010, and September 7, 2011, bathymetrical sampling surveys were carried out using a standard Ekman-Birge grab at each of the 7 locations in Lake Shoji. The average densities of the benthic communities for all the stations were 4,107 inds. m-2, comprised principally of oligochaetes 50.2%, chironomids 48.6%, and chaoborid 1.2% in March 2010, whereas in September 2011 the average density was 1,185 inds. m-2, and mostly comprised of oligochaetes 86.6% and chironomids 13.4%. On the other hand, the average biomasses (wet weight of these benthic macroinvertebrates in March 2010 were composed of 33.3% oligochaetes, 62.8% chironomid larvae, and 3.9% chaoborid larvae, as opposed to 48.8% oligochaetes and 51.2% chironomid larvae, respectively, in September 2011. Propsilocerus akamusi (Tokunaga, 1938 and Procladius sp. of chironomid larvae were the dominant species, making up 10.0% and 17.6%, respectively, of the total chironomid fauna in density, and 83.2% and 9.2% in biomass in March 2010. The mean density of P. akamusi was about 2.5 times higher than that reported by Kitagawa in the 1970s. In recent years, the number of P. akamusi larvae has tended to increase; they are widely distributed, and the anoxic-layer and anaerobic-layer are thickening, especially during summer to late fall in Lake Shoji, suggesting that the lake is in the process of eutrophication.doi: 10.5324/fn.v31i0.1365.Published online: 17 October 2012

  7. Can GSTM1 and GSTT1 polymorphisms predict clinical outcomes of chemotherapy in gastric and colorectal cancers? A result based on the previous reports

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    Liu H

    2016-06-01

    Full Text Available Haixia Liu,1,* Wei Shi,2,* Lianli Zhao,3 Dianlu Dai,4 Jinghua Gao,5 Xiangjun Kong6 1Department of Ultrasound, 2Office of Medical Statistics, 3Human Resource Department, 4Department of Surgical Oncology, 5Department of Medical Oncology, 6Central Laboratory, Cangzhou Central Hospital, Yunhe District, Cangzhou, People’s Republic of China *These authors contributed equally to this study and should be considered cofirst authors Background: Gastric and colorectal cancers remain the major causes of cancer-related death. Although chemotherapy improves the prognosis of the patients with gastrointestinal cancers, some patients do not benefit from therapy and are exposed to the adverse effects. The polymorphisms in genes including GSTM1 and GSTT1 have been explored to predict therapeutic efficacy; however, the results were inconsistent and inconclusive. Materials and methods: A systematic review and meta-analysis was performed by searching relevant studies about the association between the GSTM1 and GSTT1 polymorphisms and chemotherapy efficacy in gastrointestinal cancers in databases such as PubMed, EMBASE, Web of Science, Chinese National Knowledge Infrastructure, and Wanfang database up to January 10, 2016. Subgroup analyses were also performed according to ethnicity, cancer type, evaluation criteria, study type, chemotherapy type, and age. Results: A total of 19 articles containing 3,217 cases were finally included. Overall analysis suggested that no significance was found between overall toxicity, neurotoxicity, neutropenia, gastrointestinal toxicity, tumor response, and progression-free survival, and the polymorphisms in GSTM1 and GSTT1, while GSTM1 polymorphism associated with overall survival (OS; hazard ratio =1.213, 95% confidence interval =1.060–1.388, P=0.005. Subgroup analyses suggested that neurotoxicity was associated with GSTM1 polymorphism in the Asian population, neutropenia was associated with GSTM1 polymorphism in palliative

  8. Personality and Academic Motivation: Replication, Extension, and Replication

    Science.gov (United States)

    Jones, Martin H.; McMichael, Stephanie N.

    2015-01-01

    Previous work examines the relationships between personality traits and intrinsic/extrinsic motivation. We replicate and extend previous work to examine how personality may relate to achievement goals, efficacious beliefs, and mindset about intelligence. Approximately 200 undergraduates responded to the survey with a 150 participants replicating…

  9. Intravenous C.E.R.A. maintains stable haemoglobin levels in patients on dialysis previously treated with darbepoetin alfa: results from STRIATA, a randomized phase III study

    Science.gov (United States)

    Canaud, Bernard; Mingardi, Giulio; Braun, Johann; Aljama, Pedro; Kerr, Peter G.; Locatelli, Francesco; Villa, Giuseppe; Van Vlem, Bruno; McMahon, Alan W.; Kerloëguen, Cécile; Beyer, Ulrich

    2008-01-01

    Background. Extending the administration interval of erythropoiesis-stimulating agents (ESAs) represents an opportunity to improve the efficiency of anaemia management in patients with chronic kidney disease (CKD). However, effective haemoglobin (Hb) maintenance can be challenging with epoetin alfa and epoetin beta administered at extended intervals. C.E.R.A., a continuous erythropoietin receptor activator, has a unique pharmacologic profile and long half-life (∼130 h), allowing administration at extended intervals. Phase III results have demonstrated that C.E.R.A. administered once every 4 weeks effectively maintains stable Hb levels in patients with CKD on dialysis. Methods. STRIATA (Stabilizing haemoglobin TaRgets in dialysis following IV C.E.R.A. Treatment for Anaemia) was a multicentre, open-label randomized phase III study to evaluate the efficacy and safety of intravenous C.E.R.A. administered once every 2 weeks (Q2W) for Hb maintenance following direct conversion from darbepoetin alfa (DA). Adult patients on dialysis receiving stable intravenous DA once weekly (QW) or Q2W were randomized (1:1) to continue their current DA regimen (n = 156) or receive intravenous C.E.R.A. Q2W (n = 157) for 52 weeks. Doses were adjusted to maintain Hb levels within ± 1.0 g/dl of baseline and between 10.0 and 13.5 g/dl. The primary endpoint was the mean Hb change between baseline and the evaluation period (weeks 29–36). Results. Most patients (>80%) received DA QW before randomization. The mean (95% CI) difference between C.E.R.A. and DA in the primary endpoint was 0.18 g/dl (−0.05, 0.41), within a pre-defined non-inferiority limit. C.E.R.A. was clinically non-inferior to DA (P < 0.0001) in maintaining Hb levels. Both treatments were well tolerated. Conclusions. Stable Hb levels were successfully maintained in patients on haemodialysis directly converted to Q2W intravenous C.E.R.A. from DA. PMID:18586762

  10. Replication of biotinylated human immunodeficiency viruses.

    Science.gov (United States)

    Belshan, Michael; Matthews, John M; Madson, Christian J

    2011-01-01

    Previous work demonstrated recently the adaptation of the Escherichia coli biotin ligase BirA - biotin acceptor sequence (BAS) labeling system to produce human immunodeficiency virus type 1 viruses with biotinylated integrase (NLXIN(B)) and matrix (NLXMA(B)) proteins (Belshan et al., 2009). This report describes the construction of an HIV permissive cell line stably expressing BirA (SupT1.BirA). Consistent with the results in the previous report, NLXMA(B) replicated similar to wild-type levels and expressed biotinylated Gag and MA proteins in the SupT1.BirA cells, whereas the replication of NLXIN(B) was reduced severely. Three additional HIV type 2 (HIV-2) viruses were constructed with the BAS inserted into the vpx and vpr accessory genes. Two BAS insertions were made into the C-terminal half of the Vpx, including one internal insertion, and one at the N-terminus of Vpr. All three viruses were replication competent in the SupT1.BirA cells and their target proteins biotinylated efficiently and incorporated into virions. These results demonstrate the potential utility of the biotinylation system to label and capture HIV protein complexes in the context of replicating virus.

  11. Constructing a paleo-DEM in an urban area by the example of the city of Aachen, Germany: Methods and previous results

    Science.gov (United States)

    Pröschel, Bernhard; Lehmkuhl, Frank

    2017-04-01

    Reconstructing paleo-landscapes in urban areas is always a special challenge since the research area often witnessed constant human impact over long time periods. Dense building development is a major difficulty, particularly in regard to accessibility to in-situ soils and archaeological findings. It is therefore necessary to use data from various sources and combine methods from different fields to gain a detailed picture of the former topography. The area, which is occupied by the city of Aachen today, looks back on a long history of human influence. Traces of human activity can be dated back to Neolithic time. The first architectural structures and the first road network were built by the Romans about 2000 years ago. From then on, the area of Aachen was more or less continuously inhabited forming today's city. This long history is represented by archaeological findings throughout the city. Several meters of settlement deposits, covering different eras, are present in many locations. Therefore, it can be assumed that the modern topography significantly differs from the pre-roman topography. The main objective of this project is a reconstruction of the paleo-topography of Aachen in order to gain new insights on the spatial preconditions that the first settlers found. Moreover, further attention is given to the question whether and to what extent a paleo-DEM can help to clarify specific open archaeological and historical questions. The main database for the reconstruction are the archaeological excavation reports of the past 150 years, provided by municipal and regional archives. After analyzing these written accounts, we linked this information to drill data, provided by the Geological Service of North Rhine-Westphalia. Together with additional sources like geological and hydrological maps, we generated a GIS-based terrain model. The result is a high-resolution terrain model, representing the undisturbed pre-roman topography of the inner city of Aachen without any

  12. Medroxyprogesterone Acetate Regulates HIV-1 Uptake and Transcytosis but Not Replication in Primary Genital Epithelial Cells, Resulting in Enhanced T-Cell Infection.

    Science.gov (United States)

    Ferreira, Victor H; Dizzell, Sara; Nazli, Aisha; Kafka, Jessica K; Mueller, Kristen; Nguyen, Philip V; Tremblay, Michel J; Cochrane, Alan; Kaushic, Charu

    2015-06-01

    Although clinical and experimental evidence indicates that female sex hormones and hormonal contraceptives regulate susceptibility to human immunodeficiency virus type 1 (HIV-1) infection, the underlying mechanism remains unknown. Genital epithelial cells (GECs) are the first cells to encounter HIV during sexual transmission and their interaction with HIV may determine the outcome of exposure. This is the first report that HIV uptake by GECs increased significantly in the presence of the hormonal contraceptive medroxyprogesterone acetate (MPA) and progesterone and that uptake occurred primarily via endocytosis. No productive infection was detected, but endocytosed virus was released into apical and basolateral compartments. Significantly higher viral transcytosis was observed in the presence of MPA. In GEC and T-cell cocultures, maximum viral replication in T cells was observed in the presence of MPA, which also broadly upregulated chemokine production by GECs. These results suggest that MPA may play a significant role in regulating susceptibility to HIV. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  13. Single molecule analysis of replicated DNA reveals the usage of multiple KSHV genome regions for latent replication.

    Directory of Open Access Journals (Sweden)

    Subhash C Verma

    2011-11-01

    Full Text Available Kaposi's sarcoma associated herpesvirus (KSHV, an etiologic agent of Kaposi's sarcoma, Body Cavity Based Lymphoma and Multicentric Castleman's Disease, establishes lifelong latency in infected cells. The KSHV genome tethers to the host chromosome with the help of a latency associated nuclear antigen (LANA. Additionally, LANA supports replication of the latent origins within the terminal repeats by recruiting cellular factors. Our previous studies identified and characterized another latent origin, which supported the replication of plasmids ex-vivo without LANA expression in trans. Therefore identification of an additional origin site prompted us to analyze the entire KSHV genome for replication initiation sites using single molecule analysis of replicated DNA (SMARD. Our results showed that replication of DNA can initiate throughout the KSHV genome and the usage of these regions is not conserved in two different KSHV strains investigated. SMARD also showed that the utilization of multiple replication initiation sites occurs across large regions of the genome rather than a specified sequence. The replication origin of the terminal repeats showed only a slight preference for their usage indicating that LANA dependent origin at the terminal repeats (TR plays only a limited role in genome duplication. Furthermore, we performed chromatin immunoprecipitation for ORC2 and MCM3, which are part of the pre-replication initiation complex to determine the genomic sites where these proteins accumulate, to provide further characterization of potential replication initiation sites on the KSHV genome. The ChIP data confirmed accumulation of these pre-RC proteins at multiple genomic sites in a cell cycle dependent manner. Our data also show that both the frequency and the sites of replication initiation vary within the two KSHV genomes studied here, suggesting that initiation of replication is likely to be affected by the genomic context rather than the DNA

  14. Serotonin transporter gene moderates childhood maltreatment’s effects on persistent but not single-episode depression: Replications and implications for resolving inconsistent results

    Science.gov (United States)

    Uher, Rudolf; Caspi, Avshalom; Houts, Renate; Sugden, Karen; Williams, Benjamin; Poulton, Richie; Moffitt, Terrie E.

    2013-01-01

    Background Genetic and environmental factors shape life-long vulnerability to depression, but most gene–environment interaction (G×E) research has focused on cross-sectional assessments rather than life-course phenotypes. This study tests the hypothesis that the G×E involving the length polymorphism in the serotonin-transporter-gene-linked-promoter-region (5-HTTLPR) and childhood maltreatment is specific to depression that runs a persistent course in adulthood. Methods The hypothesis is tested in two cohorts. Men and women in the Dunedin Study (N=847), New Zealand, followed to age 32 years with 96% retention and women in the E-Risk Study (N=930), England, followed to age 40 years with 96% retention. Diagnoses of past-year major depressive episode were established at four separate assessments. Depression diagnosed on two or more occasions was considered persistent. Results In both cohorts, statistical tests of gene–environment interactions showed positive results for persistent depression but not single-episode depression. Individuals with two short 5-HTTLPR alleles and childhood maltreatment had elevated risk of persistent but not single-episode depression. Limitations Some cases of recurrent depression may have been misclassified as single-episode due to non-contiguous assessment windows, but this would have a conservative effect on the findings. Chronic and recurrent depression could not be reliably distinguished due to non-contiguous periods of assessment. Therefore, the term persistent depression is used to describe either chronic or recurrent course. Conclusions The specific effect on persistent depression increases the significance of this G×E for public health. Research that does not distinguish persistent course may underestimate G×E effects and account for some replication failures in G×E research. PMID:21439648

  15. Columnar modelling of nucleation burst evolution in the convective boundary layer – first results from a feasibility study Part IV: A compilation of previous observations for valuation of simulation results from a columnar modelling study

    Directory of Open Access Journals (Sweden)

    O. Hellmuth

    2006-01-01

    Full Text Available In the preceding Papers I, II and III a revised columnar high-order modelling approach to model gas-aerosol-turbulence interactions in the convective boundary layer (CBL was proposed, and simulation results of two synthetic nucleation scenarios (binary vs. ternary on new particle formation (NPF in the anthropogenically influenced CBL were presented and discussed. The purpose of the present finishing Paper IV is twofold: Firstly, an attempt is made to compile previous observational findings on NPF bursts in the CBL, obtained from a number of field experiments. Secondly, the scenario simulations discussed in Paper III will be evaluated with respect to the role of CBL turbulence in NPF burst evolution. It was demonstrated, that completely different nucleation mechanisms can lead to the occurrence of NPF bursts in the surface layer, but the corresponding evolution patterns strongly differ with respect to the origin, amplitude and phase of the NPF burst as well as with respect to the time-height evolution of turbulent vertical fluxes and double correlation terms of physicochemical and aerosoldynamical variables. The large differences between the binary and ternary case scenario indicate, that ammonia (NH3 can not be considered as a time-independent tuning parameter in nucleation modelling. Its contribution to the evolution of the NPF burst pattern is much more complicated and reflects the influence of CBL turbulence as well as the strong non-linearity of the ternary nucleation rate. The impact of water (H2O vapour on the nucleation rate is quite varying depending on the considered nucleation mechanism. According to the classical theory of binary nucleation involving H2O and sulphuric acid (H2SO4, H2O vapour favours NPF, according to the classical theory of ternary nuncleation involving H2O, H2SO4 and NH3 and according to organic nucleation via chemical reactions involving stabilised Criegee intermediates (SCIs, H2O vapour disfavours nucleation, and

  16. The Problem of False Discovery: Many Scientific Results Can't Be Replicated, Leading to Serious Questions about What's True and False in the World of Research.

    Science.gov (United States)

    Foster, Kenneth R; Skufca, Joseph

    2016-01-01

    Is there a Cheshire Cat in science? One might believe so, given the many published scientific discoveries that cannot be independently reproduced. The ?replication crisis? in science has become a widely discussed issue among scientists and the lay media and even has its own entry in Wikipedia.

  17. Archaeal DNA replication.

    Science.gov (United States)

    Kelman, Lori M; Kelman, Zvi

    2014-01-01

    DNA replication is essential for all life forms. Although the process is fundamentally conserved in the three domains of life, bioinformatic, biochemical, structural, and genetic studies have demonstrated that the process and the proteins involved in archaeal DNA replication are more similar to those in eukaryal DNA replication than in bacterial DNA replication, but have some archaeal-specific features. The archaeal replication system, however, is not monolithic, and there are some differences in the replication process between different species. In this review, the current knowledge of the mechanisms governing DNA replication in Archaea is summarized. The general features of the replication process as well as some of the differences are discussed.

  18. Patterns of Human Immunodeficiency Virus type 1 recombination ex vivo provide evidence for coadaptation of distant sites, resulting in purifying selection for intersubtype recombinants during replication

    DEFF Research Database (Denmark)

    Galli, Andrea; Kearney, Mary; Nikolaitchik, Olga A

    2010-01-01

    High-frequency recombination is a hallmark of HIV-1 replication. Recombination can occur between two members of the same subtype or between viruses from two different subtypes, generating intra- or intersubtype recombinants, respectively. Many intersubtype recombinants have been shown to circulate....../B) and between viruses with pol genes from subtype B or F (B/F). Recombination events generated during a single cycle of infection without selection pressure on pol gene function were analyzed by single-genome sequencing. We found that recombination occurred slightly ( approximately 30%) less frequently in B...... subtypes; these sites may be segregated by recombination events, causing the newly generated intersubtype recombinants to undergo purifying selection. Therefore, the ability of the recombinants to replicate is the major barrier for many of these viruses....

  19. Efficacy and safety of olodaterol once daily delivered via Respimat® in patients with GOLD 2–4 COPD: results from two replicate 48-week studies

    Directory of Open Access Journals (Sweden)

    Ferguson GT

    2014-06-01

    Full Text Available Gary T Ferguson,1 Gregory J Feldman,2 Peter Hofbauer,3 Alan Hamilton,4 Lisa Allen,5 Lawrence Korducki,5 Paul Sachs6 1Pulmonary Research Institute of Southeast Michigan, Livonia, MI, 2S Carolina Pharmaceutical Research, Spartanburg, SC, USA; 3Pneumologie, Weinheim, Germany; 4Boehringer Ingelheim, Burlington, ON, Canada; 5Boehringer Ingelheim Pharmaceuticals Inc., Ridgefield, CT, 6Pulmonary Associates of Stamford, Stamford, CT, USA Background: Olodaterol is a long-acting β2-agonist with a 24-hour bronchodilator profile. Two replicate, randomized, double-blind, placebo-controlled, parallel-group, Phase III trials were performed as part of a comprehensive clinical program to investigate the long-term safety and efficacy of olodaterol in patients with moderate to very severe chronic obstructive pulmonary disease (COPD receiving usual-care background therapy. Methods: Patients received olodaterol 5 µg or 10 µg or placebo once daily for 48 weeks. Coprimary end points were forced expiratory volume in 1 second (FEV1 area under the curve from 0 to 3 hours (AUC0–3 response (change from baseline, and trough FEV1 response at 12 weeks. Secondary end points included additional lung function assessments, use of rescue medications, FEV1 AUC response from 0 to 12 hours, and Patient Global Rating over 48 weeks. Results: Overall, 624 and 642 patients were evaluated in studies 1222.11 and 1222.12, respectively. In both studies, olodaterol 5 µg and 10 µg significantly improved the FEV1 AUC0–3 response (P<0.0001 and trough FEV1 (study 1222.11, P<0.0001; study 1222.12, P<0.05, post hoc at week 12, with an incidence of adverse events comparable with that of placebo. Secondary end points supported the efficacy of olodaterol. Conclusion: These studies demonstrate the long-term efficacy and safety of once-daily olodaterol 5 µg and 10 µg in patients with moderate to very severe COPD continuing with usual-care maintenance therapy. Keywords: chronic obstructive

  20. DATABASE REPLICATION IN HETEROGENOUS PLATFORM

    Directory of Open Access Journals (Sweden)

    Hendro Nindito

    2014-01-01

    Full Text Available The application of diverse database technologies in enterprises today is increasingly a common practice. To provide high availability and survavibality of real-time information, a database replication technology that has capability to replicate databases under heterogenous platforms is required. The purpose of this research is to find the technology with such capability. In this research, the data source is stored in MSSQL database server running on Windows. The data will be replicated to MySQL running on Linux as the destination. The method applied in this research is prototyping in which the processes of development and testing can be done interactively and repeatedly. The key result of this research is that the replication technology applied, which is called Oracle GoldenGate, can successfully manage to do its task in replicating data in real-time and heterogeneous platforms.

  1. Dynamics of Escherichia coli chromosome segregation during multifork replication.

    Science.gov (United States)

    Nielsen, Henrik J; Youngren, Brenda; Hansen, Flemming G; Austin, Stuart

    2007-12-01

    Slowly growing Escherichia coli cells have a simple cell cycle, with replication and progressive segregation of the chromosome completed before cell division. In rapidly growing cells, initiation of replication occurs before the previous replication rounds are complete. At cell division, the chromosomes contain multiple replication forks and must be segregated while this complex pattern of replication is still ongoing. Here, we show that replication and segregation continue in step, starting at the origin and progressing to the replication terminus. Thus, early-replicated markers on the multiple-branched chromosomes continue to separate soon after replication to form separate protonucleoids, even though they are not segregated into different daughter cells until later generations. The segregation pattern follows the pattern of chromosome replication and does not follow the cell division cycle. No extensive cohesion of sister DNA regions was seen at any growth rate. We conclude that segregation is driven by the progression of the replication forks.

  2. A whole genome RNAi screen identifies replication stress response genes.

    Science.gov (United States)

    Kavanaugh, Gina; Ye, Fei; Mohni, Kareem N; Luzwick, Jessica W; Glick, Gloria; Cortez, David

    2015-11-01

    Proper DNA replication is critical to maintain genome stability. When the DNA replication machinery encounters obstacles to replication, replication forks stall and the replication stress response is activated. This response includes activation of cell cycle checkpoints, stabilization of the replication fork, and DNA damage repair and tolerance mechanisms. Defects in the replication stress response can result in alterations to the DNA sequence causing changes in protein function and expression, ultimately leading to disease states such as cancer. To identify additional genes that control the replication stress response, we performed a three-parameter, high content, whole genome siRNA screen measuring DNA replication before and after a challenge with replication stress as well as a marker of checkpoint kinase signalling. We identified over 200 replication stress response genes and subsequently analyzed how they influence cellular viability in response to replication stress. These data will serve as a useful resource for understanding the replication stress response.

  3. Biomarkers of replicative senescence revisited

    DEFF Research Database (Denmark)

    Nehlin, Jan

    2016-01-01

    Biomarkers of replicative senescence can be defined as those ultrastructural and physiological variations as well as molecules whose changes in expression, activity or function correlate with aging, as a result of the gradual exhaustion of replicative potential and a state of permanent cell cycle...... with their chronological age and present health status, help define their current rate of aging and contribute to establish personalized therapy plans to reduce, counteract or even avoid the appearance of aging biomarkers....

  4. Replication Restart in Bacteria.

    Science.gov (United States)

    Michel, Bénédicte; Sandler, Steven J

    2017-07-01

    In bacteria, replication forks assembled at a replication origin travel to the terminus, often a few megabases away. They may encounter obstacles that trigger replisome disassembly, rendering replication restart from abandoned forks crucial for cell viability. During the past 25 years, the genes that encode replication restart proteins have been identified and genetically characterized. In parallel, the enzymes were purified and analyzed in vitro, where they can catalyze replication initiation in a sequence-independent manner from fork-like DNA structures. This work also revealed a close link between replication and homologous recombination, as replication restart from recombination intermediates is an essential step of DNA double-strand break repair in bacteria and, conversely, arrested replication forks can be acted upon by recombination proteins and converted into various recombination substrates. In this review, we summarize this intense period of research that led to the characterization of the ubiquitous replication restart protein PriA and its partners, to the definition of several replication restart pathways in vivo, and to the description of tight links between replication and homologous recombination, responsible for the importance of replication restart in the maintenance of genome stability. Copyright © 2017 American Society for Microbiology.

  5. The Psychology of Replication and Replication in Psychology.

    Science.gov (United States)

    Francis, Gregory

    2012-11-01

    Like other scientists, psychologists believe experimental replication to be the final arbiter for determining the validity of an empirical finding. Reports in psychology journals often attempt to prove the validity of a hypothesis or theory with multiple experiments that replicate a finding. Unfortunately, these efforts are sometimes misguided because in a field like experimental psychology, ever more successful replication does not necessarily ensure the validity of an empirical finding. When psychological experiments are analyzed with statistics, the rules of probability dictate that random samples should sometimes be selected that do not reject the null hypothesis, even if an effect is real. As a result, it is possible for a set of experiments to have too many successful replications. When there are too many successful replications for a given set of experiments, a skeptical scientist should be suspicious that null or negative findings have been suppressed, the experiments were run improperly, or the experiments were analyzed improperly. This article describes the implications of this observation and demonstrates how to test for too much successful replication by using a set of experiments from a recent research paper.

  6. Ultrafine anaphase bridges, broken DNA and illegitimate recombination induced by a replication fork barrier

    Science.gov (United States)

    Sofueva, Sevil; Osman, Fekret; Lorenz, Alexander; Steinacher, Roland; Castagnetti, Stefania; Ledesma, Jennifer; Whitby, Matthew C.

    2011-01-01

    Most DNA double-strand breaks (DSBs) in S- and G2-phase cells are repaired accurately by Rad51-dependent sister chromatid recombination. However, a minority give rise to gross chromosome rearrangements (GCRs), which can result in disease/death. What determines whether a DSB is repaired accurately or inaccurately is currently unclear. We provide evidence that suggests that perturbing replication by a non-programmed protein–DNA replication fork barrier results in the persistence of replication intermediates (most likely regions of unreplicated DNA) into mitosis, which results in anaphase bridge formation and ultimately to DNA breakage. However, unlike previously characterised replication-associated DSBs, these breaks are repaired mainly by Rad51-independent processes such as single-strand annealing, and are therefore prone to generate GCRs. These data highlight how a replication-associated DSB can be predisposed to give rise to genome rearrangements in eukaryotes. PMID:21576223

  7. Dissection of the beta-globin replication-initiation region reveals specific requirements for replicator elements during gene amplification.

    Directory of Open Access Journals (Sweden)

    Naoya Okada

    Full Text Available Gene amplification plays a pivotal role in malignant transformation of human cells. A plasmid with both a mammalian replication-initiation region (IR/origin/replicator and a nuclear matrix-attachment region (MAR is spontaneously amplified in transfected cells by a mechanism that involves amplification at the extrachromosomal site, followed by amplification at the chromosomal arm, ultimately generating a long homogeneously staining region (HSR. Several observations suggest that replication initiation from IR sequences might mediate amplification. To test this idea, we previously dissected c-myc and DHFR IRs to identify the minimum sequence required to support amplification. In this study, we applied an improved analysis that discriminates between two amplification steps to the ß-globin RepP IR, which contains separate elements already known to be essential for initiation on the chromosome arm. The IR sequence was required at least for the extrachromosomal amplification step. In addition to the vector-encoded MAR, amplification also required an AT-rich region and a MAR-like element, consistent with the results regarding replicator activity on the chromosome. However, amplification did not require the AG-rich tract necessary for replicator activity, but instead required a novel sequence containing another AG-rich tract. The differential sequence requirement might be a consequence of extrachromosomal replication.

  8. Replication-Uncoupled Histone Deposition during Adenovirus DNA Replication

    OpenAIRE

    Komatsu, Tetsuro; Nagata, Kyosuke

    2012-01-01

    In infected cells, the chromatin structure of the adenovirus genome DNA plays critical roles in its genome functions. Previously, we reported that in early phases of infection, incoming viral DNA is associated with both viral core protein VII and cellular histones. Here we show that in late phases of infection, newly synthesized viral DNA is also associated with histones. We also found that the knockdown of CAF-1, a histone chaperone that functions in the replication-coupled deposition of his...

  9. Rif1 regulates initiation timing of late replication origins throughout the S. cerevisiae genome.

    Directory of Open Access Journals (Sweden)

    Jared M Peace

    Full Text Available Chromosomal DNA replication involves the coordinated activity of hundreds to thousands of replication origins. Individual replication origins are subject to epigenetic regulation of their activity during S-phase, resulting in differential efficiencies and timings of replication initiation during S-phase. This regulation is thought to involve chromatin structure and organization into timing domains with differential ability to recruit limiting replication factors. Rif1 has recently been identified as a genome-wide regulator of replication timing in fission yeast and in mammalian cells. However, previous studies in budding yeast have suggested that Rif1's role in controlling replication timing may be limited to subtelomeric domains and derives from its established role in telomere length regulation. We have analyzed replication timing by analyzing BrdU incorporation genome-wide, and report that Rif1 regulates the timing of late/dormant replication origins throughout the S. cerevisiae genome. Analysis of pfa4Δ cells, which are defective in palmitoylation and membrane association of Rif1, suggests that replication timing regulation by Rif1 is independent of its role in localizing telomeres to the nuclear periphery. Intra-S checkpoint signaling is intact in rif1Δ cells, and checkpoint-defective mec1Δ cells do not comparably deregulate replication timing, together indicating that Rif1 regulates replication timing through a mechanism independent of this checkpoint. Our results indicate that the Rif1 mechanism regulates origin timing irrespective of proximity to a chromosome end, and suggest instead that telomere sequences merely provide abundant binding sites for proteins that recruit Rif1. Still, the abundance of Rif1 binding in telomeric domains may facilitate Rif1-mediated repression of non-telomeric origins that are more distal from centromeres.

  10. The prevalence and correlates of lifetime psychiatric disorders and trauma exposures in urban and rural settings: results from the national comorbidity survey replication (NCS-R.

    Directory of Open Access Journals (Sweden)

    Jennifer S McCall-Hosenfeld

    Full Text Available Distinctions between rural and urban environments produce different frequencies of traumatic exposures and psychiatric disorders. We examine the prevalence of psychiatric disorders and frequency of trauma exposures by position on the rural-urban continuum.The National Comorbidity Survey Replication (NCS-R was used to evaluate psychiatric disorders among a nationally-representative sample of the U.S. population. Rurality was designated using the Department of Agriculture's 2003 rural-urban continuum codes (RUCC, which differentiate counties into levels of rurality by population density and adjacency to metropolitan areas. Lifetime psychiatric disorders included post-traumatic stress disorder (PTSD, anxiety disorders, major depressive disorder, mood disorders, impulse-control disorders, and substance abuse. Trauma exposures were classified as war-related, accident-related, disaster-related, interpersonal or other. Weighted logistic regression models examined the odds of psychiatric disorders and trauma exposures by position on the rural-urban continuum, adjusted for relevant covariates.75% of participants were metropolitan, 12.2% were suburban, and 12.8% were from rural counties. The most common disorder reported was any anxiety disorder (38.5%. Drug abuse was more common among metropolitan (8.7%, p = 0.018, compared to nonmetropolitan (5.1% suburban, 6.1% rural participants. A one-category increase in rurality was associated with decreased odds for war-related trauma (aOR = 0.86, 95%CI 0.78-0.95. Rurality was not associated with risk for any other lifetime psychiatric disorders or trauma exposure.Contrary to the expectation of some rural primary care providers, the frequencies of most psychiatric disorders and trauma exposures are similar across the rural-urban continuum, reinforcing calls to improve mental healthcare access in resource-poor rural communities.

  11. Replication Stress: A Lifetime of Epigenetic Change

    Directory of Open Access Journals (Sweden)

    Simran Khurana

    2015-09-01

    Full Text Available DNA replication is essential for cell division. Challenges to the progression of DNA polymerase can result in replication stress, promoting the stalling and ultimately collapse of replication forks. The latter involves the formation of DNA double-strand breaks (DSBs and has been linked to both genome instability and irreversible cell cycle arrest (senescence. Recent technological advances have elucidated many of the factors that contribute to the sensing and repair of stalled or broken replication forks. In addition to bona fide repair factors, these efforts highlight a range of chromatin-associated changes at and near sites of replication stress, suggesting defects in epigenome maintenance as a potential outcome of aberrant DNA replication. Here, we will summarize recent insight into replication stress-induced chromatin-reorganization and will speculate on possible adverse effects for gene expression, nuclear integrity and, ultimately, cell function.

  12. Unscheduled DNA replication origin activation at inserted HPV 18 sequences in a HPV-18/MYC amplicon.

    Science.gov (United States)

    Conti, Chiara; Herrick, John; Bensimon, Aaron

    2007-08-01

    Oncogene amplification is a critical step leading to tumorigenesis, but the underlying mechanisms are still poorly understood. Despite data suggesting that DNA replication is a major source of genomic instability, little is known about replication origin usage and replication fork progression in rearranged regions. Using a single DNA molecule approach, we provide here the first study of replication kinetics on a previously characterized MYC/papillomavirus (HPV18) amplicon in a cervical cancer. Using this amplicon as a model, we investigated the role DNA replication control plays in generating amplifications in human cancers. The data reveal severely perturbed DNA replication kinetics in the amplified region when compared with other regions of the same genome. It was found that DNA replication is initiated from both genomic and viral sequences, resulting in a higher median frequency of origin firings. In addition, it was found that the higher initiation frequency was associated with an equivalent increase in the number of stalled replication forks. These observations raise the intriguing possibility that unscheduled replication origin activation at inserted HPV-18 viral DNA sequences triggers DNA amplification in this cancer cell line and the subsequent overexpression of the MYC oncogene.

  13. Single molecule analysis of Trypanosoma brucei DNA replication dynamics.

    Science.gov (United States)

    Calderano, Simone Guedes; Drosopoulos, William C; Quaresma, Marina Mônaco; Marques, Catarina A; Kosiyatrakul, Settapong; McCulloch, Richard; Schildkraut, Carl L; Elias, Maria Carolina

    2015-03-11

    Eukaryotic genome duplication relies on origins of replication, distributed over multiple chromosomes, to initiate DNA replication. A recent genome-wide analysis of Trypanosoma brucei, the etiological agent of sleeping sickness, localized its replication origins to the boundaries of multigenic transcription units. To better understand genomic replication in this organism, we examined replication by single molecule analysis of replicated DNA. We determined the average speed of replication forks of procyclic and bloodstream form cells and we found that T. brucei DNA replication rate is similar to rates seen in other eukaryotes. We also analyzed the replication dynamics of a central region of chromosome 1 in procyclic forms. We present evidence for replication terminating within the central part of the chromosome and thus emanating from both sides, suggesting a previously unmapped origin toward the 5' extremity of chromosome 1. Also, termination is not at a fixed location in chromosome 1, but is rather variable. Importantly, we found a replication origin located near an ORC1/CDC6 binding site that is detected after replicative stress induced by hydroxyurea treatment, suggesting it may be a dormant origin activated in response to replicative stress. Collectively, our findings support the existence of more replication origins in T. brucei than previously appreciated.

  14. Effectiveness of Peer-Led Eating Disorders Prevention: A Replication Trial

    Science.gov (United States)

    Becker, Carolyn Black; Bull, Stephanie; Schaumberg, Katherine; Cauble, Adele; Franco, Amanda

    2008-01-01

    The aim of this study was to replicate and extend results of a previous trial that investigated the effectiveness of 2 peer-led eating disorders prevention interventions in reducing eating disorder risk factors in undergraduate women (C. B. Becker, L. M. Smith, & A. C. Ciao, 2006). To extend findings from the previous study by allowing for…

  15. Data from a pre-publication independent replication initiative examining ten moral judgement effects

    Science.gov (United States)

    Tierney, Warren; Schweinsberg, Martin; Jordan, Jennifer; Kennedy, Deanna M.; Qureshi, Israr; Sommer, S. Amy; Thornley, Nico; Madan, Nikhil; Vianello, Michelangelo; Awtrey, Eli; Zhu, Luke Lei; Diermeier, Daniel; Heinze, Justin E.; Srinivasan, Malavika; Tannenbaum, David; Bivolaru, Eliza; Dana, Jason; Davis-Stober, Clintin P.; du Plessis, Christilene; Gronau, Quentin F.; Hafenbrack, Andrew C.; Liao, Eko Yi; Ly, Alexander; Marsman, Maarten; Murase, Toshio; Schaerer, Michael; Tworek, Christina M.; Wagenmakers, Eric-Jan; Wong, Lynn; Anderson, Tabitha; Bauman, Christopher W.; Bedwell, Wendy L.; Brescoll, Victoria; Canavan, Andrew; Chandler, Jesse J.; Cheries, Erik; Cheryan, Sapna; Cheung, Felix; Cimpian, Andrei; Clark, Mark A.; Cordon, Diana; Cushman, Fiery; Ditto, Peter H.; Amell, Alice; Frick, Sarah E.; Gamez-Djokic, Monica; Grady, Rebecca Hofstein; Graham, Jesse; Gu, Jun; Hahn, Adam; Hanson, Brittany E.; Hartwich, Nicole J.; Hein, Kristie; Inbar, Yoel; Jiang, Lily; Kellogg, Tehlyr; Legate, Nicole; Luoma, Timo P.; Maibeucher, Heidi; Meindl, Peter; Miles, Jennifer; Mislin, Alexandra; Molden, Daniel C.; Motyl, Matt; Newman, George; Ngo, Hoai Huong; Packham, Harvey; Ramsay, P. Scott; Ray, Jennifer L.; Sackett, Aaron M.; Sellier, Anne-Laure; Sokolova, Tatiana; Sowden, Walter; Storage, Daniel; Sun, Xiaomin; Van Bavel, Jay J.; Washburn, Anthony N.; Wei, Cong; Wetter, Erik; Wilson, Carlos T.; Darroux, Sophie-Charlotte; Uhlmann, Eric Luis

    2016-01-01

    We present the data from a crowdsourced project seeking to replicate findings in independent laboratories before (rather than after) they are published. In this Pre-Publication Independent Replication (PPIR) initiative, 25 research groups attempted to replicate 10 moral judgment effects from a single laboratory’s research pipeline of unpublished findings. The 10 effects were investigated using online/lab surveys containing psychological manipulations (vignettes) followed by questionnaires. Results revealed a mix of reliable, unreliable, and culturally moderated findings. Unlike any previous replication project, this dataset includes the data from not only the replications but also from the original studies, creating a unique corpus that researchers can use to better understand reproducibility and irreproducibility in science. PMID:27727246

  16. Internal RNA Replication Elements are Prevalent in Tombusviridae

    Directory of Open Access Journals (Sweden)

    Beth L Nicholson

    2012-08-01

    Full Text Available Internal replication elements (IREs are RNA structures that are present at internal positions in the genomes of different types of plus-strand RNA viruses. Members of the genus Tombusvirus (family Tombusviridae contain an IRE within the polymerase coding region of their genomes and this RNA element participates in both genome targeting to sites of replication and replicase complex assembly. Here we propose that other members of the virus family Tombusviridae also possess comparable IREs. Through sequence and structural analyses, candidate IREs in several genera of this family were identified, including aureusviruses, necroviruses, carmoviruses and pelarspoviruses. The results from subsequent mutational analysis of selected proposed IREs were consistent with a critical role for these structures in viral genome accumulation during infections. Our study supports the existence of IREs in several genera in Tombusviridae and points to previously unappreciated similarities in genome replication strategies between members of this virus family.

  17. DNA replication and cancer

    DEFF Research Database (Denmark)

    Boyer, Anne-Sophie; Walter, David; Sørensen, Claus Storgaard

    2016-01-01

    A dividing cell has to duplicate its DNA precisely once during the cell cycle to preserve genome integrity avoiding the accumulation of genetic aberrations that promote diseases such as cancer. A large number of endogenous impacts can challenge DNA replication and cells harbor a battery of pathways...... causing DNA replication stress and genome instability. Further, we describe cellular and systemic responses to these insults with a focus on DNA replication restart pathways. Finally, we discuss the therapeutic potential of exploiting intrinsic replicative stress in cancer cells for targeted therapy....

  18. Role of Ribonucleic Acid Synthesis in Replication of Deoxyribonucleic Acid

    Science.gov (United States)

    Pato, Martin L.

    1975-01-01

    An experiment previously interpreted to show a ribonucleic acid requirement for propagation of deoxyribonucleic replication is reexamined and the earlier interpretation is shown to be incorrect. PMID:1090599

  19. Catalysis of Strand Annealing by Replication Protein A Derives from Its Strand Melting Properties*

    OpenAIRE

    Bartos, Jeremy D.; Willmott, Lyndsay J.; Binz, Sara K.; Wold, Marc S.; Bambara, Robert A.

    2008-01-01

    Eukaryotic DNA-binding protein replication protein A (RPA) has a strand melting property that assists polymerases and helicases in resolving DNA secondary structures. Curiously, previous results suggested that human RPA (hRPA) promotes undesirable recombination by facilitating annealing of flaps produced transiently during DNA replication; however, the mechanism was not understood. We designed a series of substrates, representing displaced DNA flaps generated during ma...

  20. Theoretical Analysis of a Self-Replicator With Reduced Template Inhibition Based on an Informational Leaving Group.

    Science.gov (United States)

    Bigan, Erwan; Mattelaer, Henri-Philippe; Herdewijn, Piet

    2016-03-01

    The first non-enzymatic self-replicating systems, as proposed by von Kiedrowski (Angew Chem Int Ed Engl 25(10):932-935, 1986) and Orgel (Nature 327(6120):346-347, 1987), gave rise to the analytical background still used today to describe artificial replicators. What separates a self-replicating from an autocatalytic system is the ability to pass on structural information (Orgel, Nature 358(6383):203-209, 1992). Utilising molecular information, nucleic acids were the first choice as prototypical examples. But early self-replicators showed parabolic over exponential growth due to the strongly bound template duplex after template-directed ligation of substrates. We propose a self-replicating scheme with a weakly bound template duplex, using an informational leaving group. Such a scheme is inspired by the role of tRNA as leaving group and information carrier during protein synthesis, and is based on our previous experience with nucleotide chemistry. We analyse theoretically this scheme and compare it to the classical minimal replicator model. We show that for an example hexanucleotide template mirroring that is used by von Kiedrowski (Bioorganic chemistry frontiers, 1993) for the analysis of the classical minimal replicator, the proposed scheme is expected to result in higher template self-replication rate. The proposed self-replicating scheme based on an informational leaving group is expected to outperform the classical minimal replicator because of a weaker template duplex bonding, resulting in reduced template inhibition.

  1. DNA Damage Reduces the Quality, but Not the Quantity of Human Papillomavirus 16 E1 and E2 DNA Replication

    Directory of Open Access Journals (Sweden)

    Molly L. Bristol

    2016-06-01

    Full Text Available Human papillomaviruses (HPVs are causative agents in almost all cervical carcinomas. HPVs are also causative agents in head and neck cancer, the cases of which are increasing rapidly. Viral replication activates the DNA damage response (DDR pathway; associated proteins are recruited to replication foci, and this pathway may serve to allow for viral genome amplification. Likewise, HPV genome double-strand breaks (DSBs could be produced during replication and could lead to linearization and viral integration. Many studies have shown that viral integration into the host genome results in unregulated expression of the viral oncogenes, E6 and E7, promoting HPV-induced carcinogenesis. Previously, we have demonstrated that DNA-damaging agents, such as etoposide, or knocking down viral replication partner proteins, such as topoisomerase II β binding protein I (TopBP1, does not reduce the level of DNA replication. Here, we investigated whether these treatments alter the quality of DNA replication by HPV16 E1 and E2. We confirm that knockdown of TopBP1 or treatment with etoposide does not reduce total levels of E1/E2-mediated DNA replication; however, the quality of replication is significantly reduced. The results demonstrate that E1 and E2 continue to replicate under genomically-stressed conditions and that this replication is mutagenic. This mutagenesis would promote the formation of substrates for integration of the viral genome into that of the host, a hallmark of cervical cancer.

  2. Direct visualization of replication dynamics in early zebrafish embryos.

    Science.gov (United States)

    Kuriya, Kenji; Higashiyama, Eriko; Avşar-Ban, Eriko; Okochi, Nanami; Hattori, Kaede; Ogata, Shin; Takebayashi, Shin-Ichiro; Ogata, Masato; Tamaru, Yutaka; Okumura, Katsuzumi

    2016-05-01

    We analyzed DNA replication in early zebrafish embryos. The replicating DNA of whole embryos was labeled with the thymidine analog 5-ethynyl-2'-deoxyuridine (EdU), and spatial regulation of replication sites was visualized in single embryo-derived cells. The results unveiled uncharacterized replication dynamics during zebrafish early embryogenesis.

  3. Excess Cdt1 inhibits nascent strand elongation by repressing the progression of replication forks in Xenopus egg extracts.

    Science.gov (United States)

    Nakazaki, Yuta; Tsuyama, Takashi; Seki, Masayuki; Takahashi, Mikiko; Enomoto, Takemi; Tada, Shusuke

    2016-02-01

    Cdt1 is a protein essential for initiation of DNA replication; it recruits MCM helicase, a core component of the replicative DNA helicase, onto replication origins. In our previous study, we showed that addition of excess Cdt1 inhibits nascent strand elongation during DNA replication in Xenopus egg extracts. In the present study, we investigated the mechanism behind the inhibitory effect of Cdt1. We found that addition of recombinant Cdt1 inhibited nascent DNA synthesis in a reinitiation-independent manner. To identify the mechanism by which Cdt1 inhibits nascent strand elongation, the effect of Cdt1 on loading of Mcm4 and Rpa70 onto chromatin was examined. The results showed that Cdt1 suppressed the excessive Rpa70 binding caused by extensive, aphidicolin-induced DNA unwinding; this unwinding occurs between stalled DNA polymerases and advancing replication forks. These findings suggested that excess Cdt1 suppressed the progression of replication forks.

  4. Chromosomal context and replication properties of ARS plasmids in Schizosaccharomyces pombe

    Indian Academy of Sciences (India)

    Aditya S Pratihar; Vishnu P Tripathi; Mukesh P Yadav; Dharani D Dubey

    2015-12-01

    Short, specific DNA sequences called as Autonomously Replicating Sequence (ARS) elements function as plasmid as well as chromosomal replication origins in yeasts. As compared to ARSs, different chromosomal origins vary greatly in their efficiency and timing of replication probably due to their wider chromosomal context. The two Schizosaccharomyces pombe ARS elements, ars727 and ars2OO4, represent two extremities in their chromosomal origin activity - ars727 is inactive and late replicating, while ars2OO4 is a highly active, early-firing origin. To determine the effect of chromosomal context on the activity of these ARS elements, we have cloned them with their extended chromosomal context as well as in the context of each other in both orientations and analysed their replication efficiency by ARS and plasmid stability assays. We found that these ARS elements retain their origin activity in their extended/altered context. However, deletion of a 133-bp region of the previously reported ars727-associated late replication enforcing element (LRE) caused advancement in replication timing of the resulting plasmid. These results confirm the role of LRE in directing plasmid replication timing and suggest that the plasmid origin efficiency of ars2OO4 or ars727 remains unaltered by the extended chromosomal context.

  5. Replicating animal mitochondrial DNA

    Directory of Open Access Journals (Sweden)

    Emily A. McKinney

    2013-01-01

    Full Text Available The field of mitochondrial DNA (mtDNA replication has been experiencing incredible progress in recent years, and yet little is certain about the mechanism(s used by animal cells to replicate this plasmid-like genome. The long-standing strand-displacement model of mammalian mtDNA replication (for which single-stranded DNA intermediates are a hallmark has been intensively challenged by a new set of data, which suggests that replication proceeds via coupled leading-and lagging-strand synthesis (resembling bacterial genome replication and/or via long stretches of RNA intermediates laid on the mtDNA lagging-strand (the so called RITOLS. The set of proteins required for mtDNA replication is small and includes the catalytic and accessory subunits of DNA polymerase y, the mtDNA helicase Twinkle, the mitochondrial single-stranded DNA-binding protein, and the mitochondrial RNA polymerase (which most likely functions as the mtDNA primase. Mutations in the genes coding for the first three proteins are associated with human diseases and premature aging, justifying the research interest in the genetic, biochemical and structural properties of the mtDNA replication machinery. Here we summarize these properties and discuss the current models of mtDNA replication in animal cells.

  6. Defining multiple, distinct, and shared spatiotemporal patterns of DNA replication and endoreduplication from 3D image analysis of developing maize (Zea mays L.) root tip nuclei.

    Science.gov (United States)

    Bass, Hank W; Hoffman, Gregg G; Lee, Tae-Jin; Wear, Emily E; Joseph, Stacey R; Allen, George C; Hanley-Bowdoin, Linda; Thompson, William F

    2015-11-01

    Spatiotemporal patterns of DNA replication have been described for yeast and many types of cultured animal cells, frequently after cell cycle arrest to aid in synchronization. However, patterns of DNA replication in nuclei from plants or naturally developing organs remain largely uncharacterized. Here we report findings from 3D quantitative analysis of DNA replication and endoreduplication in nuclei from pulse-labeled developing maize root tips. In both early and middle S phase nuclei, flow-sorted on the basis of DNA content, replicative labeling was widely distributed across euchromatic regions of the nucleoplasm. We did not observe the perinuclear or perinucleolar replicative labeling patterns characteristic of middle S phase in mammals. Instead, the early versus middle S phase patterns in maize could be distinguished cytologically by correlating two quantitative, continuous variables, replicative labeling and DAPI staining. Early S nuclei exhibited widely distributed euchromatic labeling preferentially localized to regions with weak DAPI signals. Middle S nuclei also exhibited widely distributed euchromatic labeling, but the label was preferentially localized to regions with strong DAPI signals. Highly condensed heterochromatin, including knobs, replicated during late S phase as previously reported. Similar spatiotemporal replication patterns were observed for both mitotic and endocycling maize nuclei. These results revealed that maize euchromatin exists as an intermingled mixture of two components distinguished by their condensation state and replication timing. These different patterns might reflect a previously described genome organization pattern, with "gene islands" mostly replicating during early S phase followed by most of the intergenic repetitive regions replicating during middle S phase.

  7. Semiconservative replication in the quasispecies model

    Science.gov (United States)

    Tannenbaum, Emmanuel; Deeds, Eric J.; Shakhnovich, Eugene I.

    2004-06-01

    This paper extends Eigen’s quasispecies equations to account for the semiconservative nature of DNA replication. We solve the equations in the limit of infinite sequence length for the simplest case of a static, sharply peaked fitness landscape. We show that the error catastrophe occurs when μ , the product of sequence length and per base pair mismatch probability, exceeds 2 ln [2/ ( 1+1/k ) ] , where k>1 is the first-order growth rate constant of the viable “master” sequence (with all other sequences having a first-order growth rate constant of 1 ). This is in contrast to the result of ln k for conservative replication. In particular, as k→∞ , the error catastrophe is never reached for conservative replication, while for semiconservative replication the critical μ approaches 2 ln 2 . Semiconservative replication is therefore considerably less robust than conservative replication to the effect of replication errors. We also show that the mean equilibrium fitness of a semiconservatively replicating system is given by k ( 2 e-μ/2 -1 ) below the error catastrophe, in contrast to the standard result of k e-μ for conservative replication (derived by Kimura and Maruyama in 1966). From this result it is readily shown that semiconservative replication is necessary to account for the observation that, at sufficiently high mutagen concentrations, faster replicating cells will die more quickly than more slowly replicating cells. Thus, in contrast to Eigen’s original model, the semiconservative quasispecies equations are able to provide a mathematical basis for explaining the efficacy of mutagens as chemotherapeutic agents.

  8. Data from Investigating Variation in Replicability: A “Many Labs” Replication Project

    Directory of Open Access Journals (Sweden)

    Richard A. Klein

    2014-04-01

    Full Text Available This dataset is from the Many Labs Replication Project in which 13 effects were replicated across 36 samples and over 6,000 participants. Data from the replications are included, along with demographic variables about the participants and contextual information about the environment in which the replication was conducted. Data were collected in-lab and online through a standardized procedure administered via an online link. The dataset is stored on the Open Science Framework website. These data could be used to further investigate the results of the included 13 effects or to study replication and generalizability more broadly.

  9. Involvement of the skin during bluetongue virus infection and replication in the ruminant host.

    Science.gov (United States)

    Darpel, Karin E; Monaghan, Paul; Simpson, Jennifer; Anthony, Simon J; Veronesi, Eva; Brooks, Harriet W; Elliott, Heather; Brownlie, Joe; Takamatsu, Haru-Hisa; Mellor, Philip S; Mertens, Peter Pc

    2012-04-30

    Bluetongue virus (BTV) is a double stranded (ds) RNA virus (genus Orbivirus; family Reoviridae), which is considered capable of infecting all species of domestic and wild ruminants, although clinical signs are seen mostly in sheep. BTV is arthropod-borne ("arbovirus") and able to productively infect and replicate in many different cell types of both insects and mammalian hosts. Although the organ and cellular tropism of BTV in ruminants has been the subject of several studies, many aspects of its pathogenesis are still poorly understood, partly because of inherent problems in distinguishing between "virus replication" and "virus presence".BTV replication and organ tropism were studied in a wide range of infected sheep tissues, by immuno-fluorescence-labeling of non-structural or structural proteins (NS2 or VP7 and core proteins, respectively) using confocal microscopy to distinguish between virus presence and replication. These results are compared to gross and microscopic pathological findings in selected organs from infected sheep. Replication was demonstrated in two major cell types: vascular endothelial cells, and agranular leukocytes which morphologically resemble lymphocytes, monocytes/macrophages and/or dendritic cells. Two organs (the skin and tonsils) were shown to support relatively high levels of BTV replication, although they have not previously been proposed as important replication sites during BTV infection. The high level of BTV replication in the skin is thought to be of major significance for the pathogenesis and transmission of BTV (via biting insects) and a refinement of our current model of BTV pathogenesis is discussed.

  10. Recruitment of wild-type and recombinant adeno-associated virus into adenovirus replication centers.

    Science.gov (United States)

    Weitzman, M D; Fisher, K J; Wilson, J M

    1996-03-01

    Replication of a human parvovirus, adeno-associated virus (AAV), is facilitated by coinfection with adeno-virus to provide essential helper functions. We have used the techniques of in situ hybridization and immunocytochemistry to characterize the localization of AAV replication within infected cells, Previous studies have shown that adenovirus establishes foci called replication centers within the nucleus, where adenoviral replication and transcription occur. Our studies indicate that AAV is colocalized with the adenovirus replication centers, where it may utilize adenovirus and cellular proteins for its own replication. Expression of the AAV Rep protein inhibits the normal maturation of the adenovirus centers. Similar experiments were performed with recombinant AAV (rAAV) to establish a relationship between intranuclear localization and rAAV transduction. rAAV efficiently entered the cell, and its genome was faintly detectable in a perinuclear distribution and was mobilized to replication centers when the cell was infected with adenovirus. The recruitment of the replication-defective genome into the intranuclear adenovirus domains resulted in enhanced transduction. These studies illustrate the importance of intracellular compartmentalization for such complex interactions as the relationship between AAV and adenovirus.

  11. Using autonomous replication to physically and genetically define human origins of replication

    Energy Technology Data Exchange (ETDEWEB)

    Krysan, P.J.

    1993-01-01

    The author previously developed a system for studying autonomous replication in human cells involving the use of sequences from the Epstein-Barr virus (EBV) genome to provide extrachromosomal plasmids with a nuclear retention function. Using this system, it was demonstrated that large fragments of human genomic DNA could be isolated which replicate autonomously in human cells. In this study the DNA sequences which function as origins of replication in human cells are defined physically and genetically. These experiments demonstrated that replication initiates at multiple locations distributed throughout the plasmid. Another line of experiments addressed the DNA sequence requirements for autonomous replication in human cells. These experiments demonstrated that human DNA fragments have a higher replication activity than bacterial fragments do. It was also found, however, that the bacterial DNA sequence could support efficient replication if enough copies of it were present on the plasmid. These findings suggested that autonomous replication in human cells does not depend on extensive, specific DNA sequences. The autonomous replication system which the author has employed for these experiments utilizes a cis-acting sequence from the EBV origin and the trans-acting EBNA-1 protein to provide plasmids with a nuclear retention function. It was therefore relevant to verify that the autonomous replication of human DNA fragments did not depend on the replication activity associated with the EBV sequences utilized for nuclear retention. To accomplish this goal, the author demonstrated that plasmids carrying the EBV sequences and large fragments of human DNA could support long-term autonomous replication in hamster cells, which are not permissive for EBV replication.

  12. Quality-of-life and performance status results from the phase III RAINBOW study of ramucirumab plus paclitaxel versus placebo plus paclitaxel in patients with previously treated gastric or gastroesophageal junction adenocarcinoma†

    Science.gov (United States)

    Al-Batran, S.-E.; Van Cutsem, E.; Oh, S. C.; Bodoky, G.; Shimada, Y.; Hironaka, S.; Sugimoto, N.; Lipatov, O. N.; Kim, T.-Y.; Cunningham, D.; Rougier, P.; Muro, K.; Liepa, A. M.; Chandrawansa, K.; Emig, M.; Ohtsu, A.; Wilke, H.

    2016-01-01

    Background The phase III RAINBOW trial demonstrated that the addition of ramucirumab to paclitaxel improved overall survival, progression-free survival, and tumor response rate in fluoropyrimidine–platinum previously treated patients with advanced gastric/gastroesophageal junction (GEJ) adenocarcinoma. Here, we present results from quality-of-life (QoL) and performance status (PS) analyses. Patients and methods Patients with Eastern Cooperative Oncology Group PS of 0/1 were randomized to receive ramucirumab (8 mg/kg i.v.) or placebo on days 1 and 15 of a 4-week cycle, with both arms receiving paclitaxel (80 mg/m2) on days 1, 8, and 15. Patient-reported outcomes were assessed with the QoL/health status questionnaires EORTC QLQ-C30 and EQ-5D at baseline and 6-week intervals. PS was assessed at baseline and day 1 of every cycle. Time to deterioration (TtD) in each QLQ-C30 scale was defined as randomization to first worsening of ≥10 points (on 100-point scale) and TtD in PS was defined as first worsening to ≥2. Hazard ratios (HRs) for treatment effect were estimated using stratified Cox proportional hazards models. Results Of the 665 patients randomized, 650 (98%) provided baseline QLQ-C30 and EQ-5D data, and 560 (84%) also provided data from ≥1 postbaseline time point. Baseline scores for both instruments were similar between arms. Of the 15 QLQ-C30 scales, 14 had HR < 1, indicating similar or longer TtD in QoL for ramucirumab + paclitaxel. Treatment with ramucirumab + paclitaxel was also associated with a delay in TtD in PS to ≥2 (HR = 0.798, P = 0.0941). Alternate definitions of PS deterioration yielded similar results: PS ≥ 3 (HR = 0.656, P = 0.0508), deterioration by ≥1 PS level (HR = 0.802, P = 0.0444), and deterioration by ≥2 PS levels (HR = 0.608, P = 0.0063). EQ-5D scores were comparable between treatment arms, stable during treatment, and worsened at discontinuation. Conclusion In patients with previously treated advanced gastric

  13. The Replication Recipe: What makes for a convincing replication?

    NARCIS (Netherlands)

    Brandt, M.J.; IJzerman, H.; Dijksterhuis, A.J.; Farach, F.J.; Geller, J.; Giner-Sorolla, R.; Grange, J.A.; Perugini, M.; Spies, J.R.; Veer, A. van 't

    2014-01-01

    Psychological scientists have recently started to reconsider the importance of close replications in building a cumulative knowledge base; however, there is no consensus about what constitutes a convincing close replication study. To facilitate convincing close replication attempts we have developed

  14. Comparison of three replication strategies in complex multicellular organisms: Asexual replication, sexual replication with identical gametes, and sexual replication with distinct sperm and egg gametes

    Science.gov (United States)

    Tannenbaum, Emmanuel

    2008-01-01

    This paper studies the mutation-selection balance in three simplified replication models. The first model considers a population of organisms replicating via the production of asexual spores. The second model considers a sexually replicating population that produces identical gametes. The third model considers a sexually replicating population that produces distinct sperm and egg gametes. All models assume diploid organisms whose genomes consist of two chromosomes, each of which is taken to be functional if equal to some master sequence, and defective otherwise. In the asexual population, the asexual diploid spores develop directly into adult organisms. In the sexual populations, the haploid gametes enter a haploid pool, where they may fuse with other haploids. The resulting immature diploid organisms then proceed to develop into mature organisms. Based on an analysis of all three models, we find that, as organism size increases, a sexually replicating population can only outcompete an asexually replicating population if the adult organisms produce distinct sperm and egg gametes. A sexual replication strategy that is based on the production of large numbers of sperm cells to fertilize a small number of eggs is found to be necessary in order to maintain a sufficiently low cost for sex for the strategy to be selected for over a purely asexual strategy. We discuss the usefulness of this model in understanding the evolution and maintenance of sexual replication as the preferred replication strategy in complex, multicellular organisms.

  15. Modeling DNA Replication.

    Science.gov (United States)

    Bennett, Joan

    1998-01-01

    Recommends the use of a model of DNA made out of Velcro to help students visualize the steps of DNA replication. Includes a materials list, construction directions, and details of the demonstration using the model parts. (DDR)

  16. Eukaryotic DNA Replication Fork.

    Science.gov (United States)

    Burgers, Peter M J; Kunkel, Thomas A

    2017-06-20

    This review focuses on the biogenesis and composition of the eukaryotic DNA replication fork, with an emphasis on the enzymes that synthesize DNA and repair discontinuities on the lagging strand of the replication fork. Physical and genetic methodologies aimed at understanding these processes are discussed. The preponderance of evidence supports a model in which DNA polymerase ε (Pol ε) carries out the bulk of leading strand DNA synthesis at an undisturbed replication fork. DNA polymerases α and δ carry out the initiation of Okazaki fragment synthesis and its elongation and maturation, respectively. This review also discusses alternative proposals, including cellular processes during which alternative forks may be utilized, and new biochemical studies with purified proteins that are aimed at reconstituting leading and lagging strand DNA synthesis separately and as an integrated replication fork.

  17. Chikungunya virus infection results in higher and persistent viral replication in aged rhesus macaques due to defects in anti-viral immunity.

    Directory of Open Access Journals (Sweden)

    Ilhem Messaoudi

    Full Text Available Chikungunya virus (CHIKV is a re-emerging mosquito-borne Alphavirus that causes a clinical disease involving fever, myalgia, nausea and rash. The distinguishing feature of CHIKV infection is the severe debilitating poly-arthralgia that may persist for several months after viral clearance. Since its re-emergence in 2004, CHIKV has spread from the Indian Ocean region to new locations including metropolitan Europe, Japan, and even the United States. The risk of importing CHIKV to new areas of the world is increasing due to high levels of viremia in infected individuals as well as the recent adaptation of the virus to the mosquito species Aedes albopictus. CHIKV re-emergence is also associated with new clinical complications including severe morbidity and, for the first time, mortality. In this study, we characterized disease progression and host immune responses in adult and aged Rhesus macaques infected with either the recent CHIKV outbreak strain La Reunion (LR or the West African strain 37997. Our results indicate that following intravenous infection and regardless of the virus used, Rhesus macaques become viremic between days 1-5 post infection. While adult animals are able to control viral infection, aged animals show persistent virus in the spleen. Virus-specific T cell responses in the aged animals were reduced compared to adult animals and the B cell responses were also delayed and reduced in aged animals. Interestingly, regardless of age, T cell and antibody responses were more robust in animals infected with LR compared to 37997 CHIKV strain. Taken together these data suggest that the reduced immune responses in the aged animals promotes long-term virus persistence in CHIKV-LR infected Rhesus monkeys.

  18. Adenovirus DNA Replication

    OpenAIRE

    Hoeben, Rob C.; Uil, Taco G.

    2013-01-01

    Adenoviruses have attracted much attention as probes to study biological processes such as DNA replication, transcription, splicing, and cellular transformation. More recently these viruses have been used as gene-transfer vectors and oncolytic agents. On the other hand, adenoviruses are notorious pathogens in people with compromised immune functions. This article will briefly summarize the basic replication strategy of adenoviruses and the key proteins involved and will deal with the new deve...

  19. APOBEC3G induces a hypermutation gradient: purifying selection at multiple steps during HIV-1 replication results in levels of G-to-A mutations that are high in DNA, intermediate in cellular viral RNA, and low in virion RNA

    Directory of Open Access Journals (Sweden)

    Pathak Vinay K

    2009-02-01

    Full Text Available Abstract Background Naturally occurring Vif variants that are unable to inhibit the host restriction factor APOBEC3G (A3G have been isolated from infected individuals. A3G can potentially induce G-to-A hypermutation in these viruses, and hypermutation could contribute to genetic variation in HIV-1 populations through recombination between hypermutant and wild-type genomes. Thus, hypermutation could contribute to the generation of immune escape and drug resistant variants, but the genetic contribution of hypermutation to the viral evolutionary potential is poorly understood. In addition, the mechanisms by which these viruses persist in the host despite the presence of A3G remain unknown. Results To address these questions, we generated a replication-competent HIV-1 Vif mutant in which the A3G-binding residues of Vif, Y40RHHY44, were substituted with five alanines. As expected, the mutant was severely defective in an A3G-expressing T cell line and exhibited a significant delay in replication kinetics. Analysis of viral DNA showed the expected high level of G-to-A hypermutation; however, we found substantially reduced levels of G-to-A hypermutation in intracellular viral RNA (cRNA, and the levels of G-to-A mutations in virion RNA (vRNA were even further reduced. The frequencies of hypermutation in DNA, cRNA, and vRNA were 0.73%, 0.12%, and 0.05% of the nucleotides sequenced, indicating a gradient of hypermutation. Additionally, genomes containing start codon mutations and early termination codons within gag were isolated from the vRNA. Conclusion These results suggest that sublethal levels of hypermutation coupled with purifying selection at multiple steps during the early phase of viral replication lead to the packaging of largely unmutated genomes, providing a mechanism by which mutant Vif variants can persist in infected individuals. The persistence of genomes containing mutated gag genes despite this selection pressure indicates that dual

  20. Hyperthermia stimulates HIV-1 replication.

    Directory of Open Access Journals (Sweden)

    Ferdinand Roesch

    Full Text Available HIV-infected individuals may experience fever episodes. Fever is an elevation of the body temperature accompanied by inflammation. It is usually beneficial for the host through enhancement of immunological defenses. In cultures, transient non-physiological heat shock (42-45°C and Heat Shock Proteins (HSPs modulate HIV-1 replication, through poorly defined mechanisms. The effect of physiological hyperthermia (38-40°C on HIV-1 infection has not been extensively investigated. Here, we show that culturing primary CD4+ T lymphocytes and cell lines at a fever-like temperature (39.5°C increased the efficiency of HIV-1 replication by 2 to 7 fold. Hyperthermia did not facilitate viral entry nor reverse transcription, but increased Tat transactivation of the LTR viral promoter. Hyperthermia also boosted HIV-1 reactivation in a model of latently-infected cells. By imaging HIV-1 transcription, we further show that Hsp90 co-localized with actively transcribing provirus, and this phenomenon was enhanced at 39.5°C. The Hsp90 inhibitor 17-AAG abrogated the increase of HIV-1 replication in hyperthermic cells. Altogether, our results indicate that fever may directly stimulate HIV-1 replication, in a process involving Hsp90 and facilitation of Tat-mediated LTR activity.

  1. Hyperthermia stimulates HIV-1 replication.

    Science.gov (United States)

    Roesch, Ferdinand; Meziane, Oussama; Kula, Anna; Nisole, Sébastien; Porrot, Françoise; Anderson, Ian; Mammano, Fabrizio; Fassati, Ariberto; Marcello, Alessandro; Benkirane, Monsef; Schwartz, Olivier

    2012-01-01

    HIV-infected individuals may experience fever episodes. Fever is an elevation of the body temperature accompanied by inflammation. It is usually beneficial for the host through enhancement of immunological defenses. In cultures, transient non-physiological heat shock (42-45°C) and Heat Shock Proteins (HSPs) modulate HIV-1 replication, through poorly defined mechanisms. The effect of physiological hyperthermia (38-40°C) on HIV-1 infection has not been extensively investigated. Here, we show that culturing primary CD4+ T lymphocytes and cell lines at a fever-like temperature (39.5°C) increased the efficiency of HIV-1 replication by 2 to 7 fold. Hyperthermia did not facilitate viral entry nor reverse transcription, but increased Tat transactivation of the LTR viral promoter. Hyperthermia also boosted HIV-1 reactivation in a model of latently-infected cells. By imaging HIV-1 transcription, we further show that Hsp90 co-localized with actively transcribing provirus, and this phenomenon was enhanced at 39.5°C. The Hsp90 inhibitor 17-AAG abrogated the increase of HIV-1 replication in hyperthermic cells. Altogether, our results indicate that fever may directly stimulate HIV-1 replication, in a process involving Hsp90 and facilitation of Tat-mediated LTR activity.

  2. Crinivirus replication and host interactions

    Directory of Open Access Journals (Sweden)

    Zsofia A Kiss

    2013-05-01

    Full Text Available Criniviruses comprise one of the genera within the family Closteroviridae. Members in this family are restricted to the phloem and rely on whitefly vectors of the genera Bemisia and/or Trialeurodes for plant-to-plant transmission. All criniviruses have bipartite, positive-sense ssRNA genomes, although there is an unconfirmed report of one having a tripartite genome. Lettuce infectious yellows virus (LIYV is the type species of the genus, the best studied so far of the criniviruses and the first for which a reverse genetics system was available. LIYV RNA 1 encodes for proteins predicted to be involved in replication, and alone is competent for replication in protoplasts. Replication results in accumulation of cytoplasmic vesiculated membranous structures which are characteristic of most studied members of the Closteroviridae. These membranous structures, often referred to as BYV-type vesicles, are likely sites of RNA replication. LIYV RNA 2 is replicated in trans when co-infecting cells with RNA 1, but is temporally delayed relative to RNA1. Efficient RNA 2 replication also is dependent on the RNA 1-encoded RNA binding protein, P34. No LIYV RNA 2-encoded proteins have been shown to affect RNA replication, but at least four, CP, CPm, Hsp70h, and p59 are virion structural components and CPm is a determinant of whitefly transmissibility. Roles of other LIYV RNA 2-encoded proteins are largely as yet unknown, but P26 is a non-virion protein that accumulates in cells as characteristic plasmalemma deposits which in plants are localized within phloem parenchyma and companion cells over plasmodesmata connections to sieve elements. The two remaining crinivirus-conserved RNA 2-encoded proteins are P5 and P9. P5 is 39 amino acid protein and is encoded at the 5’ end of RNA 2 as ORF1 and is part of the hallmark closterovirus gene array. The orthologous gene in BYV has been shown to play a role in cell-to-cell movement and indicated to be localized to the

  3. Replicative Intermediates of Human Papillomavirus Type 11 in Laryngeal Papillomas: Site of Replication Initiation and Direction of Replication

    Science.gov (United States)

    Auborn, K. J.; Little, R. D.; Platt, T. H. K.; Vaccariello, M. A.; Schildkraut, C. L.

    1994-07-01

    We have examined the structures of replication intermediates from the human papillomavirus type 11 genome in DNA extracted from papilloma lesions (laryngeal papillomas). The sites of replication initiation and termination utilized in vivo were mapped by using neutral/neutral and neutral/alkaline two-dimensional agarose gel electrophoresis methods. Initiation of replication was detected in or very close to the upstream regulatory region (URR; the noncoding, regulatory sequences upstream of the open reading frames in the papillomavirus genome). We also show that replication forks proceed bidirectionally from the origin and converge 180circ opposite the URR. These results demonstrate the feasibility of analysis of replication of viral genomes directly from infected tissue.

  4. Investigating variation in replicability: A "Many Labs" replication project

    NARCIS (Netherlands)

    Klein, R.A.; Ratliff, K.A.; Vianello, M.; Adams, R.B.; Bahnik, S.; Bernstein, M.J.; Bocian, K.; Brandt, M.J.; Brooks, B.; Brumbaugh, C.C.; Cemalcilar, Z.; Chandler, J.; Cheong, W.; Davis, W.E.; Devos, T.; Eisner, M.; Frankowska, N.; Furrow, D.; Galliani, E.M.; Hasselman, F.W.; Hicks, J.A.; Hovermale, J.F.; Hunt, S.J.; Huntsinger, J.R.; IJzerman, H.; John, M.S.; Joy-Gaba, J.A.; Kappes, H.B.; Krueger, L.E.; Kurtz, J.; Levitan, C.A.; Mallett, R.K.; Morris, W.L.; Nelson, A.J.; Nier, J.A.; Packard, G.; Pilati, R.; Rutchick, A.M.; Schmidt, K.; Skorinko, J.L.M.; Smith, R.; Steiner, T.G.; Storbeck, J.; Van Swol, L.M.; Thompson, D.; Veer, A.E. van 't; Vaughn, L.A.; Vranka, M.; Wichman, A.L.; Woodzicka, J.A.; Nosek, B.A.

    2014-01-01

    Although replication is a central tenet of science, direct replications are rare in psychology. This research tested variation in the replicability of 13 classic and contemporary effects across 36 independent samples totaling 6,344 participants. In the aggregate, 10 effects replicated consistently.

  5. Hepatitis B virus replication

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Hepadnaviruses, including human hepatitis B virus (HBV), replicate through reverse transcription of an RNA intermediate, the pregenomic RNA (pgRNA). Despite this kinship to retroviruses, there are fundamental differences beyond the fact that hepadnavirions contain DNA instead of RNA. Most peculiar is the initiation of reverse transcription: it occurs by protein-priming, is strictly committed to using an RNA hairpin on the pgRNA,ε, as template, and depends on cellular chaperones;moreover, proper replication can apparently occur only in the specialized environment of intact nucleocapsids.This complexity has hampered an in-depth mechanistic understanding. The recent successful reconstitution in the test tube of active replication initiation complexes from purified components, for duck HBV (DHBV),now allows for the analysis of the biochemistry of hepadnaviral replication at the molecular level. Here we review the current state of knowledge at all steps of the hepadnaviral genome replication cycle, with emphasis on new insights that turned up by the use of such cellfree systems. At this time, they can, unfortunately,not be complemented by three-dimensional structural information on the involved components. However, at least for the s RNA element such information is emerging,raising expectations that combining biophysics with biochemistry and genetics will soon provide a powerful integrated approach for solving the many outstanding questions. The ultimate, though most challenging goal,will be to visualize the hepadnaviral reverse transcriptase in the act of synthesizing DNA, which will also have strong implications for drug development.

  6. The Role of the Transcriptional Response to DNA Replication Stress.

    Science.gov (United States)

    Herlihy, Anna E; de Bruin, Robertus A M

    2017-03-02

    During DNA replication many factors can result in DNA replication stress. The DNA replication stress checkpoint prevents the accumulation of replication stress-induced DNA damage and the potential ensuing genome instability. A critical role for post-translational modifications, such as phosphorylation, in the replication stress checkpoint response has been well established. However, recent work has revealed an important role for transcription in the cellular response to DNA replication stress. In this review, we will provide an overview of current knowledge of the cellular response to DNA replication stress with a specific focus on the DNA replication stress checkpoint transcriptional response and its role in the prevention of replication stress-induced DNA damage.

  7. Completion of DNA replication in Escherichia coli.

    Science.gov (United States)

    Wendel, Brian M; Courcelle, Charmain T; Courcelle, Justin

    2014-11-18

    The mechanism by which cells recognize and complete replicated regions at their precise doubling point must be remarkably efficient, occurring thousands of times per cell division along the chromosomes of humans. However, this process remains poorly understood. Here we show that, in Escherichia coli, the completion of replication involves an enzymatic system that effectively counts pairs and limits cellular replication to its doubling point by allowing converging replication forks to transiently continue through the doubling point before the excess, over-replicated regions are incised, resected, and joined. Completion requires RecBCD and involves several proteins associated with repairing double-strand breaks including, ExoI, SbcDC, and RecG. However, unlike double-strand break repair, completion occurs independently of homologous recombination and RecA. In some bacterial viruses, the completion mechanism is specifically targeted for inactivation to allow over-replication to occur during lytic replication. The results suggest that a primary cause of genomic instabilities in many double-strand-break-repair mutants arises from an impaired ability to complete replication, independent from DNA damage.

  8. Psychology, replication & beyond.

    Science.gov (United States)

    Laws, Keith R

    2016-06-01

    Modern psychology is apparently in crisis and the prevailing view is that this partly reflects an inability to replicate past findings. If a crisis does exists, then it is some kind of 'chronic' crisis, as psychologists have been censuring themselves over replicability for decades. While the debate in psychology is not new, the lack of progress across the decades is disappointing. Recently though, we have seen a veritable surfeit of debate alongside multiple orchestrated and well-publicised replication initiatives. The spotlight is being shone on certain areas and although not everyone agrees on how we should interpret the outcomes, the debate is happening and impassioned. The issue of reproducibility occupies a central place in our whig history of psychology.

  9. Mechanism of chromosomal DNA replication initiation and replication fork stabilization in eukaryotes.

    Science.gov (United States)

    Wu, LiHong; Liu, Yang; Kong, DaoChun

    2014-05-01

    Chromosomal DNA replication is one of the central biological events occurring inside cells. Due to its large size, the replication of genomic DNA in eukaryotes initiates at hundreds to tens of thousands of sites called DNA origins so that the replication could be completed in a limited time. Further, eukaryotic DNA replication is sophisticatedly regulated, and this regulation guarantees that each origin fires once per S phase and each segment of DNA gets duplication also once per cell cycle. The first step of replication initiation is the assembly of pre-replication complex (pre-RC). Since 1973, four proteins, Cdc6/Cdc18, MCM, ORC and Cdt1, have been extensively studied and proved to be pre-RC components. Recently, a novel pre-RC component called Sap1/Girdin was identified. Sap1/Girdin is required for loading Cdc18/Cdc6 to origins for pre-RC assembly in the fission yeast and human cells, respectively. At the transition of G1 to S phase, pre-RC is activated by the two kinases, cyclindependent kinase (CDK) and Dbf4-dependent kinase (DDK), and subsequently, RPA, primase-polα, PCNA, topoisomerase, Cdc45, polδ, and polɛ are recruited to DNA origins for creating two bi-directional replication forks and initiating DNA replication. As replication forks move along chromatin DNA, they frequently stall due to the presence of a great number of replication barriers on chromatin DNA, such as secondary DNA structures, protein/DNA complexes, DNA lesions, gene transcription. Stalled forks must require checkpoint regulation for their stabilization. Otherwise, stalled forks will collapse, which results in incomplete DNA replication and genomic instability. This short review gives a concise introduction regarding the current understanding of replication initiation and replication fork stabilization.

  10. Therapeutic targeting of replicative immortality

    OpenAIRE

    Yaswen, Paul; MacKenzie, Karen L.; Keith, W. Nicol; Hentosh, Patricia; Rodier, Francis; Zhu, Jiyue; Firestone, Gary L.; Matheu, Ander; Carnero, Amancio; Bilsland, Alan; Sundin, Tabetha; Honoki, Kanya; Fujii, Hiromasa; Georgakilas, Alexandros G.; Amedei, Amedeo

    2015-01-01

    One of the hallmarks of malignant cell populations is the ability to undergo continuous proliferation. This property allows clonal lineages to acquire sequential aberrations that can fuel increasingly autonomous growth, invasiveness, and therapeutic resistance. Innate cellular mechanisms have evolved to regulate replicative potential as a hedge against malignant progression. When activated in the absence of normal terminal differentiation cues, these mechanisms can result in a state of persis...

  11. ASAR15, A cis-acting locus that controls chromosome-wide replication timing and stability of human chromosome 15.

    Directory of Open Access Journals (Sweden)

    Nathan Donley

    2015-01-01

    Full Text Available DNA replication initiates at multiple sites along each mammalian chromosome at different times during each S phase, following a temporal replication program. We have used a Cre/loxP-based strategy to identify cis-acting elements that control this replication-timing program on individual human chromosomes. In this report, we show that rearrangements at a complex locus at chromosome 15q24.3 result in delayed replication and structural instability of human chromosome 15. Characterization of this locus identified long, RNA transcripts that are retained in the nucleus and form a "cloud" on one homolog of chromosome 15. We also found that this locus displays asynchronous replication that is coordinated with other random monoallelic genes on chromosome 15. We have named this locus ASynchronous replication and Autosomal RNA on chromosome 15, or ASAR15. Previously, we found that disruption of the ASAR6 lincRNA gene results in delayed replication, delayed mitotic condensation and structural instability of human chromosome 6. Previous studies in the mouse found that deletion of the Xist gene, from the X chromosome in adult somatic cells, results in a delayed replication and instability phenotype that is indistinguishable from the phenotype caused by disruption of either ASAR6 or ASAR15. In addition, delayed replication and chromosome instability were detected following structural rearrangement of many different human or mouse chromosomes. These observations suggest that all mammalian chromosomes contain similar cis-acting loci. Thus, under this scenario, all mammalian chromosomes contain four distinct types of essential cis-acting elements: origins, telomeres, centromeres and "inactivation/stability centers", all functioning to promote proper replication, segregation and structural stability of each chromosome.

  12. Replication of a chronic hepatitis B virus genotype F1b construct.

    Science.gov (United States)

    Hernández, Sergio; Jiménez, Gustavo; Alarcón, Valentina; Prieto, Cristian; Muñoz, Francisca; Riquelme, Constanza; Venegas, Mauricio; Brahm, Javier; Loyola, Alejandra; Villanueva, Rodrigo A

    2016-03-01

    Genotype F is one of the less-studied genotypes of human hepatitis B virus, although it is widely distributed in regions of Central and South American. Our previous studies have shown that HBV genotype F is prevalent in Chile, and phylogenetic analysis of its full-length sequence amplified from the sera of chronically infected patients identified it as HBV subgenotype F1b. We have previously reported the full-length sequence of a HBV molecular clone obtained from a patient chronically infected with genotype F1b. In this report, we established a system to study HBV replication based on hepatoma cell lines transfected with full-length monomers of the HBV genome. Culture supernatants were analyzed after transfection and found to contain both HBsAg and HBeAg viral antigens. Consistently, fractionated cell extracts revealed the presence of viral replication, with both cytoplasmic and nuclear DNA intermediates. Analysis of HBV-transfected cells by indirect immunofluorescence or immunoelectron microscopy revealed the expression of viral antigens and cytoplasmic viral particles, respectively. To test the functionality of the ongoing viral replication further at the level of chromatinized cccDNA, transfected cells were treated with a histone deacetylase inhibitor, and this resulted in increased viral replication. This correlated with changes posttranslational modifications of histones at viral promoters. Thus, the development of this viral replication system for HBV genotype F will facilitate studies on the regulation of viral replication and the identification of new antiviral drugs.

  13. Metadata Control Agent approach for Replication in Grid Environments

    Directory of Open Access Journals (Sweden)

    P. SunilGavaskar

    2013-10-01

    Full Text Available since grid environment is dynamic, network latency and user requests may change. In order to provide better communication, access time and fault tolerant in decentralized systems, the replication is a technique to reduce access time, storage space. The objective of the work is to propose an agent control approach for Heterogeneous environments using the Agents for storing objects as replicas in decentralized environments. Our idea minimizes the more replicas (i.e. causes overhead on response time and update cost, therefore maintaining suitable number of replicas is important. Fixed replicas provides file access structure to identify the esteem files and gives optimal replication location, which minimize replication issues like access time and update cost by assuming a given traffic pattern. In this context we present the Agents as replicas to maintain a suitable scalable architecture. The solution uses fewer replicas, which lead to fewer agents as a result of that frequent updating is possible. Our tests show that the proposed strategy outperforms previous solutions in terms of replication issues.

  14. Replication intermediate analysis confirms that chromosomal replication origin initiates from an unusual intergenic region in Caulobacter crescentus.

    Science.gov (United States)

    Brassinga, A K; Marczynski, G T

    2001-11-01

    The alpha-proteobacterium Caulobacter crescentus possesses a developmental cell cycle that restricts chromosome replication to a stalked cell type. The proposed C.crescentus chromosome replication origin (Cori) lies between hemE and RP001, an unusual intergenic region not previously associated with bacterial replication origins, although a similar genomic arrangement is also present at the putative replication origin in the related bacterium Rickettsia prowazekii. The cloned Cori supports autonomous plasmid replication selectively in the stalked cell type implying that replication of the entire chromosome also initiates between hemE and RP001. To confirm this location, we applied the 2-D (N/N) agarose gel electrophoresis technique to resolve and identify chromosome replication intermediates throughout a 30 kb region spanning Cori. Replication initiation in Cori was uniquely characterized by an 'origin bubble and Y-arc' pattern and this observation was supported by simple replication fork 'Y-arc' patterns that characterized the regions flanking Cori. These replication forks originated bi-directionally from within Cori as determined by the fork direction assay. Therefore, chromosomal replication initiates from the unusual hemE/RP001 intergenic region that we propose represents a new class of replication origins.

  15. DNA replication origins in archaea

    OpenAIRE

    Zhenfang eWu; Jingfang eLiu; Haibo eYang; Hua eXiang

    2014-01-01

    DNA replication initiation, which starts at specific chromosomal site (known as replication origins), is the key regulatory stage of chromosome replication. Archaea, the third domain of life, use a single or multiple origin(s) to initiate replication of their circular chromosomes. The basic structure of replication origins is conserved among archaea, typically including an AT-rich unwinding region flanked by several conserved repeats (origin recognition box, ORB) that are located adjacent to ...

  16. Replicator dynamics in value chains

    DEFF Research Database (Denmark)

    Cantner, Uwe; Savin, Ivan; Vannuccini, Simone

    2016-01-01

    The pure model of replicator dynamics though providing important insights in the evolution of markets has not found much of empirical support. This paper extends the model to the case of firms vertically integrated in value chains. We show that i) by taking value chains into account, the replicator...... dynamics may revert its effect. In these regressive developments of market selection, firms with low fitness expand because of being integrated with highly fit partners, and the other way around; ii) allowing partner's switching within a value chain illustrates that periods of instability in the early...... stage of industry life-cycle may be the result of an 'optimization' of partners within a value chain providing a novel and simple explanation to the evidence discussed by Mazzucato (1998); iii) there are distinct differences in the contribution to market selection between the layers of a value chain...

  17. Replication-Fork Dynamics

    NARCIS (Netherlands)

    Duderstadt, Karl E.; Reyes-Lamothe, Rodrigo; van Oijen, Antoine M.; Sherratt, David J.

    2014-01-01

    The proliferation of all organisms depends on the coordination of enzymatic events within large multiprotein replisomes that duplicate chromosomes. Whereas the structure and function of many core replisome components have been clarified, the timing and order of molecular events during replication re

  18. Coronavirus Attachment and Replication

    Science.gov (United States)

    1988-03-28

    synthesis during RNA replication of vesicular stomatitis virus. J. Virol. 49:303-309. Pedersen, N.C. 1976a. Feline infectious peritonitis: Something old...receptors on intestinal brush border membranes from normal host species were developed for canine (CCV), feline (FIPV), porcine (TGEV), human (HCV...gastroenteritis receptor on pig BBMs ...... ................. ... 114 Feline infectious peritonitis virus receptor on cat BBMs ... .............. 117 Human

  19. Mcm2 phosphorylation and the response to replicative stress

    Directory of Open Access Journals (Sweden)

    Stead Brent E

    2012-05-01

    Full Text Available Abstract Background The replicative helicase in eukaryotic cells is comprised of minichromosome maintenance (Mcm proteins 2 through 7 (Mcm2-7 and is a key target for regulation of cell proliferation. In addition, it is regulated in response to replicative stress. One of the protein kinases that targets Mcm2-7 is the Dbf4-dependent kinase Cdc7 (DDK. In a previous study, we showed that alanine mutations of the DDK phosphorylation sites at S164 and S170 in Saccharomyces cerevisiae Mcm2 result in sensitivity to caffeine and methyl methanesulfonate (MMS leading us to suggest that DDK phosphorylation of Mcm2 is required in response to replicative stress. Results We show here that a strain with the mcm2 allele lacking DDK phosphorylation sites (mcm2AA is also sensitive to the ribonucleotide reductase inhibitor, hydroxyurea (HU and to the base analogue 5-fluorouracil (5-FU but not the radiomimetic drug, phleomycin. We screened the budding yeast non-essential deletion collection for synthetic lethal interactions with mcm2AA and isolated deletions that include genes involved in the control of genome integrity and oxidative stress. In addition, the spontaneous mutation rate, as measured by mutations in CAN1, was increased in the mcm2AA strain compared to wild type, whereas with a phosphomimetic allele (mcm2EE the mutation rate was decreased. These results led to the idea that the mcm2AA strain is unable to respond properly to DNA damage. We examined this by screening the deletion collection for suppressors of the caffeine sensitivity of mcm2AA. Deletions that decrease spontaneous DNA damage, increase homologous recombination or slow replication forks were isolated. Many of the suppressors of caffeine sensitivity suppressed other phenotypes of mcm2AA including sensitivity to genotoxic drugs, the increased frequency of cells with RPA foci and the increased mutation rate. Conclusions Together these observations point to a role for DDK-mediated phosphorylation

  20. On the scattering of DNA replication completion times

    Science.gov (United States)

    Meilikhov, E. Z.; Farzetdinova, R. M.

    2015-07-01

    Stochasticity of Eukaryotes' DNA replication should not lead to large fluctuations of replication times, which could result in mitotic catastrophes. Fundamental problem that cells face is how to be ensured that entire genome is replicated on time. We develop analytic approach of calculating DNA replication times, that being simplified and approximate, leads, nevertheless, to results practically coincident with those that were obtained by some sophisticated methods. In the framework of that model we consider replication times' scattering and discuss the influence of repair stopping on kinetics of DNA replication. Our main explicit formulae for DNA replication time t r ∝ ( N is the total number of DNA base pairs) is of general character and explains basic features of DNA replication kinetics.

  1. From the chromatin interaction network to the organization of the human genome into replication N/U-domains

    Science.gov (United States)

    Boulos, Rasha E.; Julienne, Hanna; Baker, Antoine; Chen, Chun-Long; Petryk, Nataliya; Kahli, Malik; dʼAubenton-Carafa, Yves; Goldar, Arach; Jensen, Pablo; Hyrien, Olivier; Thermes, Claude; Arneodo, Alain; Audit, Benjamin

    2014-11-01

    The three-dimensional (3D) architecture of the mammalian nucleus is now being unraveled thanks to the recent development of chromatin conformation capture (3C) technologies. Here we report the results of a combined multiscale analysis of genome-wide mean replication timing and chromatin conformation data that reveal some intimate relationships between chromatin folding and human DNA replication. We previously described megabase replication N/U-domains as mammalian multiorigin replication units, and showed that their borders are ‘master’ replication initiation zones that likely initiate cascades of origin firing responsible for the stereotypic replication of these domains. Here, we demonstrate that replication N/U-domains correspond to the structural domains of self-interacting chromatin, and that their borders act as insulating regions both in high-throughput 3C (Hi-C) data and high-resolution 3C (4C) experiments. Further analyses of Hi-C data using a graph-theoretical approach reveal that N/U-domain borders are long-distance, interconnected hubs of the chromatin interaction network. Overall, these results and the observation that a well-defined ordering of chromatin states exists from N/U-domain borders to centers suggest that ‘master’ replication initiation zones are at the heart of a high-order, epigenetically controlled 3D organization of the human genome.

  2. DNA moves sequentially towards the nuclear matrix during DNA replication in vivo

    Directory of Open Access Journals (Sweden)

    Aranda-Anzaldo Armando

    2011-01-01

    Full Text Available Abstract Background In the interphase nucleus of metazoan cells DNA is organized in supercoiled loops anchored to a nuclear matrix (NM. There is varied evidence indicating that DNA replication occurs in replication factories organized upon the NM and that DNA loops may correspond to the actual replicons in vivo. In normal rat liver the hepatocytes are arrested in G0 but they synchronously re-enter the cell cycle after partial-hepatectomy leading to liver regeneration in vivo. We have previously determined in quiescent rat hepatocytes that a 162 kbp genomic region containing members of the albumin gene family is organized into five structural DNA loops. Results In the present work we tracked down the movement relative to the NM of DNA sequences located at different points within such five structural DNA loops during the S phase and after the return to cellular quiescence during liver regeneration. Our results indicate that looped DNA moves sequentially towards the NM during replication and then returns to its original position in newly quiescent cells, once the liver regeneration has been achieved. Conclusions Looped DNA moves in a sequential fashion, as if reeled in, towards the NM during DNA replication in vivo thus supporting the notion that the DNA template is pulled progressively towards the replication factories on the NM so as to be replicated. These results provide further evidence that the structural DNA loops correspond to the actual replicons in vivo.

  3. Efficacy and Safety of Vorapaxar With and Without a Thienopyridine for Secondary Prevention in Patients With Previous Myocardial Infarction and No History of Stroke or Transient Ischemic Attack: Results from TRA 2°P-TIMI 50.

    Science.gov (United States)

    Bohula, Erin A; Aylward, Philip E; Bonaca, Marc P; Corbalan, Ramon L; Kiss, Robert G; Murphy, Sabina A; Scirica, Benjamin M; White, Harvey; Braunwald, Eugene; Morrow, David A

    2015-11-17

    Vorapaxar antagonizes protease-activated receptor 1, the primary receptor for thrombin on human platelets, and reduces recurrent thrombotic events in stable patients with a previous myocardial infarction (MI). We wished to determine whether the efficacy and safety of antiplatelet therapy with vorapaxar was modified by concurrent thienopyridine use. The Thrombin Receptor Antagonist in Secondary Prevention of Atherothrombotic Ischemic Events-Thrombolysis in Myocardial Infarction 50 (TRA 2°P-TIMI 50) was a randomized, double-blind, placebo-controlled trial of vorapaxar in 26,449 patients with previous atherothrombosis. This prespecified analysis included 16,897 patients who qualified with a MI in the preceding 2 weeks to 12 months and was restricted to patients without a history of stroke or transient ischemic attack given its contraindication in that population. Randomization was stratified on the basis of planned thienopyridine use. Thienopyridine was planned at randomization in 12,410 (73%). Vorapaxar significantly reduced the composite of cardiovascular death, MI, and stroke in comparison with placebo regardless of planned thienopyridine therapy (planned thienopyridine, hazard ratio, 0.80, 0.70-0.91, P<0.001; no planned thienopyridine, hazard ratio, 0.75; 0.60-0.94, P=0.011; P-interaction=0.67). Findings were similar when patients were stratified by actual thienopyridine use at baseline (P-interaction=0.82) and through 18 months (P-interaction=0.44). Global Use of Strategies to Open Occluded Coronary Arteries (GUSTO) moderate or severe bleeding risk was increased with vorapaxar and was not significantly altered by planned thienopyridine (planned, hazard ratio, 1.50; 1.18-1.89, P<0.001; no planned, hazard ratio, 1.90, 1.17-3.07, P=0.009; P-interaction=0.37) or actual thienopyridine use (P-interaction=0.24). Vorapaxar reduced cardiovascular death, MI, or stroke in stable patients with a history of previous MI, whether treated concomitantly with a thienopyridine or

  4. Multifork chromosome replication in slow-growing bacteria

    Science.gov (United States)

    Trojanowski, Damian; Hołówka, Joanna; Ginda, Katarzyna; Jakimowicz, Dagmara; Zakrzewska-Czerwińska, Jolanta

    2017-01-01

    The growth rates of bacteria must be coordinated with major cell cycle events, including chromosome replication. When the doubling time (Td) is shorter than the duration of chromosome replication (C period), a new round of replication begins before the previous round terminates. Thus, newborn cells inherit partially duplicated chromosomes. This phenomenon, which is termed multifork replication, occurs among fast-growing bacteria such as Escherichia coli and Bacillus subtilis. In contrast, it was historically believed that slow-growing bacteria (including mycobacteria) do not reinitiate chromosome replication until the previous round has been completed. Here, we use single-cell time-lapse analyses to reveal that mycobacterial cell populations exhibit heterogeneity in their DNA replication dynamics. In addition to cells with non-overlapping replication rounds, we observed cells in which the next replication round was initiated before completion of the previous replication round. We speculate that this heterogeneity may reflect a relaxation of cell cycle checkpoints, possibly increasing the ability of slow-growing mycobacteria to adapt to environmental conditions. PMID:28262767

  5. PCNA Modifications for Regulation of Post-Replication Repair Pathways

    OpenAIRE

    2008-01-01

    Stalled DNA replication forks activate specific DNA repair mechanism called post-replication repair (PRR) pathways that simply bypass DNA damage. The bypassing of DNA damage by PRR prevents prolonged stalling of DNA replication that could result in double strand breaks (DSBs). Proliferating cell nuclear antigen (PCNA) functions to initiate and choose different bypassing pathways of PRR. In yeast, DNA replication forks stalled by DNA damage induces monoubiquitination of PCNA at K164, which is ...

  6. Is the Previously Reported Increase in Second- and Higher-order Birth Rates in Norway and Sweden from the mid-1970s Real or a Result of Inadequate Estimation Methods?

    Directory of Open Access Journals (Sweden)

    2002-03-01

    Full Text Available According to models estimated separately for second-, third-, and fourth-birth rates in Norway, an increase took place from the mid-1970s to about 1990, given age and duration since last previous birth. A similar rise in the birth rates was seen in Sweden, except that the upturn at short durations was sharper. It is shown in this study, using Norwegian register data, that the increase partly reflects earlier changes in lower-order parity transitions. When models for each parity transition are estimated jointly, with a common unobserved factor included, there is no longer an upward trend in Norwegian second-birth rates, but a very weak decline, and the increase in the higher-order birth rates is strongly reduced compared to that found in the simpler approach.

  7. Immunogenicity, efficacy and safety of Nuwiq(®) (human-cl rhFVIII) in previously untreated patients with severe haemophilia A-Interim results from the NuProtect Study.

    Science.gov (United States)

    Liesner, R J; Abashidze, M; Aleinikova, O; Altisent, C; Belletrutti, M J; Borel-Derlon, A; Carcao, M; Chambost, H; Chan, A K C; Dubey, L; Ducore, J; Fouzia, N A; Gattens, M; Gruel, Y; Guillet, B; Kavardakova, N; El Khorassani, M; Klukowska, A; Lambert, T; Lohade, S; Sigaud, M; Turea, V; Wu, J K M; Vdovin, V; Pavlova, A; Jansen, M; Belyanskaya, L; Walter, O; Knaub, S; Neufeld, E J

    2017-08-16

    Nuwiq(®) (Human-cl rhFVIII) is a fourth generation recombinant FVIII, produced in a human cell line, without chemical modification or protein fusion. No inhibitors developed in studies with Nuwiq(®) in 201 previously treated patients with haemophilia A (HA). The immunogenicity, efficacy and safety of Nuwiq(®) in previously untreated patients (PUPs) with severe HA are being assessed in the ongoing NuProtect study. The study, conducted across 38 centres worldwide, is evaluating 110 true PUPs of all ages and ethnicities enrolled for study up to 100 exposure days (EDs) or 5 years maximum. The primary objective is to assess the immunogenicity of Nuwiq(®) (inhibitor activity ≥0.6 BU) using the Nijmegen-modified Bethesda assay at a central laboratory. Data for 66 PUPs with ≥20 EDs from a preplanned interim analysis were analysed. High-titre (HT) inhibitors developed in 8 of 66 patients after a median of 11.5 EDs (range 6-24). Five patients developed low-titre inhibitors (4 transient). The cumulative incidence (95% confidence interval) was 12.8% (4.5%, 21.2%) for HT inhibitors and 20.8% (10.7%, 31.0%) for all inhibitors. During inhibitor-free periods, median annualized bleeding rates during prophylaxis were 0 for spontaneous bleeds and 2.40 for all bleeds. Efficacy was rated as "excellent" or "good" in treating 91.8% of bleeds. Efficacy of surgical prophylaxis was "excellent" or "good" for 8 (89%) procedures and "moderate" for 1 (11%). No tolerability concerns were evident. These interim data show a cumulative incidence of 12.8% for HT inhibitors and convincing efficacy and tolerability in PUPs treated with Nuwiq(®) . © 2017 The Authors. Haemophilia published by John Wiley & Sons Ltd.

  8. Oncogene v-jun modulates DNA replication.

    Science.gov (United States)

    Wasylyk, C; Schneikert, J; Wasylyk, B

    1990-07-01

    Cell transformation leads to alterations in both transcription and DNA replication. Activation of transcription by the expression of a number of transforming oncogenes is mediated by the transcription factor AP1 (Herrlich & Ponta, 1989; Imler & Wasylyk, 1989). AP1 is a composite transcription factor, consisting of members of the jun and fos gene-families. c-jun and c-fos are progenitors of oncogenes, suggestion that an important transcriptional event in cell transformation is altered activity of AP1, which may arise either indirectly by oncogene expression or directly by structural modification of AP1. We report here that the v-jun oncogene and its progenitor c-jun, as fusion proteins with the lex-A-repressor DNA binding domain, can activate DNA replication from the Polyoma virus (Py) origin of replication, linked to the lex-A operator. The transcription-activation region of v-jun is required for activation of replication. When excess v-jun is expressed in the cell, replication is inhibited or 'squelched'. These results suggest that one consequence of deregulated jun activity could be altered DNA replication and that there are similarities in the way v-jun activates replication and transcription.

  9. Reversible Switching of Cooperating Replicators

    Science.gov (United States)

    Urtel, Georg C.; Rind, Thomas; Braun, Dieter

    2017-02-01

    How can molecules with short lifetimes preserve their information over millions of years? For evolution to occur, information-carrying molecules have to replicate before they degrade. Our experiments reveal a robust, reversible cooperation mechanism in oligonucleotide replication. Two inherently slow replicating hairpin molecules can transfer their information to fast crossbreed replicators that outgrow the hairpins. The reverse is also possible. When one replication initiation site is missing, single hairpins reemerge from the crossbreed. With this mechanism, interacting replicators can switch between the hairpin and crossbreed mode, revealing a flexible adaptation to different boundary conditions.

  10. Replication of M13 single—stranded DNA bearing a sitespecific ethenocytosine lesion by Escherichia coil cell extracts

    Institute of Scientific and Technical Information of China (English)

    WANGGE; PAULMDUNMAN; 等

    1997-01-01

    Previous investigation on the mutagenic effects of 3,N4-Ethenocytosine (εC),a nonpairing DNA lesion,revealed the existence of a novel SOS-independent inducible mutagenic mechanism in E.coli termed UVM for UV modulation of mutagenesis.To investigate whether UVM is mediated by an alteration of DNA replication,we have set up an in vitro replication system in which phage M13 viral single-stranded DNA bearing a single site-specific (εC) residue is replicated by soluble protein extracts from E.coli cells.Replication products were analyzed by agarose gel electrophoresis and the frequency of translesion synthesis was determined by restriction endonuclease analyses.Our data indicate that DNA replication is strongly inhibited by εC,but that translesion DNA synthesis does occur in about 14% of the replicated DNA molecules.These results are very similar to those observed previously in vivo,and suggest that this experimental system may be suitable for evaluating alterations in DNA replication in UVM-induced cells.

  11. The glyceraldehyde-3-phosphate dehydrogenase and the small GTPase Rab 2 are crucial for Brucella replication.

    Directory of Open Access Journals (Sweden)

    Emilie Fugier

    2009-06-01

    Full Text Available The intracellular pathogen Brucella abortus survives and replicates inside host cells within an endoplasmic reticulum (ER-derived replicative organelle named the "Brucella-containing vacuole" (BCV. Here, we developed a subcellular fractionation method to isolate BCVs and characterize for the first time the protein composition of its replicative niche. After identification of BCV membrane proteins by 2 dimensional (2D gel electrophoresis and mass spectrometry, we focused on two eukaryotic proteins: the glyceraldehyde-3-phosphate dehydrogenase (GAPDH and the small GTPase Rab 2 recruited to the vacuolar membrane of Brucella. These proteins were previously described to localize on vesicular and tubular clusters (VTC and to regulate the VTC membrane traffic between the endoplasmic reticulum (ER and the Golgi. Inhibition of either GAPDH or Rab 2 expression by small interfering RNA strongly inhibited B. abortus replication. Consistent with this result, inhibition of other partners of GAPDH and Rab 2, such as COPI and PKC iota, reduced B. abortus replication. Furthermore, blockage of Rab 2 GTPase in a GDP-locked form also inhibited B. abortus replication. Bacteria did not fuse with the ER and instead remained in lysosomal-associated membrane vacuoles. These results reveal an essential role for GAPDH and the small GTPase Rab 2 in B. abortus virulence within host cells.

  12. Characteristics of DNA replication in isolated nuclei initiated by an aprotinin-binding protein.

    Science.gov (United States)

    Coffman, F D; Fresa, K L; Hameed, M; Cohen, S

    1993-02-01

    Isolated cell nuclei were used as the source of template DNA to investigate the role of a cytosolic aprotinin-binding protein (ADR) in the initiation of eukaryotic DNA replication. Computerized image cytometry demonstrated that the DNA content of individual nuclei increased significantly following incubation with ADR-containing preparations, and the extent of DNA synthesis is consistent with that allowed by the limiting concentration of dTTP. Thus, dTTP incorporation into isolated nuclei represents DNA synthesis and not parent strand repair. We found that dTTP incorporation into the isolated nuclei is dependent on DNA polymerase alpha (a principal polymerase in DNA replication) but that DNA polymerase beta (a principal polymerase in DNA repair processes) does not play a significant role in this system. Finally, neither aprotinin nor a previously described cytosolic ADR inhibitor can block the replication of nuclease-treated calf thymus DNA, while both strongly inhibit replication of DNA in isolated nuclei. This result, coupled with the relative ineffectiveness of nuclease-treated DNA compared with nuclear DNA to serve as a replicative template in this assay, argues against a significant contribution from repair or synthesis which initiates at a site of DNA damage. These data indicate that ADR-mediated incorporation of 3H-dTTP into isolated nuclei results from DNA replicative processes that are directly relevant to in vivo S phase events.

  13. Replicator-dynamics models of sexual conflict.

    Science.gov (United States)

    Kimura, Mariko; Ihara, Yasuo

    2009-09-07

    Evolutionary conflict between the sexes has been studied in various taxa and in various contexts. When the sexes are in conflict over mating rates, natural selection favors both males that induce higher mating rates and females that are more successful at resisting mating attempts. Such sexual conflict may result in an escalating coevolutionary arms race between males and females. In this article, we develop simple replicator-dynamics models of sexual conflict in order to investigate its evolutionary dynamics. Two specific models of the dependence of a female's fitness on her number of matings are considered: in model 1, female fitness decreases linearly with increasing number of matings and in model 2, there is an optimal number of matings that maximizes female fitness. For each of these models, we obtain the conditions for a coevolutionary process to establish costly male and female traits and examine under what circumstances polymorphism is maintained at equilibrium. Then we discuss how assumptions in previous models of sexual conflict are translated to fit to our model framework and compare our results with those of the previous studies. The simplicity of our models allows us to consider sexual conflict in various contexts within a single framework. In addition, we find that our model 2 shows more complicated evolutionary dynamics than model 1. In particular, the population exhibits bistability, where the evolutionary outcome depends on the initial state, only in model 2.

  14. Characterization of ambient aerosols in Mexico City during the MCMA-2003 campaign with Aerosol Mass Spectrometry Part II: overview of the results at the CENICA supersite and comparison to previous studies

    Science.gov (United States)

    Salcedo, D.; Dzepina, K.; Onasch, T. B.; Canagaratna, M. R.; Jayne, J. T.; Worsnop, D. R.; Gaffney, J. S.; Marley, N. A.; Johnson, K. S.; Zuberi, B.; Molina, L. T.; Molina, M. J.; Shutthanandan, V.; Xie, Y.; Jimenez, J. L.

    2005-06-01

    An Aerodyne Aerosol Mass Spectrometer (AMS) was deployed at the CENICA Supersite during the Mexico City Metropolitan Area field study from 31 March-4 May 2003. The AMS provides real time information on mass concentration and composition of the non-refractory species in particulate matter less than 1 µm (NR-PM1) with high time and size-resolution. Measurements of Black Carbon (BC) using an aethalometer, and estimated soil concentrations from Proton-Induced X-Ray Emission (PIXE) analysis of impactor substrates are also presented and combined with the AMS in order to include refractory material and estimate the total PM2.5 mass concentration at CENICA during this campaign. In Mexico City, the organic fraction of the estimated PM2.5 at CENICA represents 54.6% of the mass, with the rest consisting of inorganic compounds (mainly ammonium nitrate and sulfate/ammonium salts), BC, and soil. Inorganic compounds represent 27.5% of PM2.5; BC mass concentration is about 11%; while soil represents about 6.9%. The NR species and BC have diurnal cycles that can be qualitatively interpreted as the interplay of direct emissions, photochemical production in the atmosphere followed by condensation and gas-to-particle partitioning, boundary layer dynamics, and/or advection. Bi- and trimodal size distributions are observed for the AMS species, with a small combustion (likely traffic) organic particle mode and an accumulation mode that contains mainly organic and secondary inorganic compounds. The AMS and BC mass concentrations, size distributions, and diurnal cycles are found to be qualitatively similar to those from most previous field measurements in Mexico City.

  15. Usefulness of indirect alcohol biomarkers for predicting recidivism of drunk-driving among previously convicted drunk-driving offenders: results from the recidivism of alcohol-impaired driving (ROAD) study.

    Science.gov (United States)

    Maenhout, Thomas M; Poll, Anneleen; Vermassen, Tijl; De Buyzere, Marc L; Delanghe, Joris R

    2014-01-01

    In several European countries, drivers under the influence (DUI), suspected of chronic alcohol abuse are referred for medical and psychological examination. This study (the ROAD study, or Recidivism Of Alcohol-impaired Driving) investigated the usefulness of indirect alcohol biomarkers for predicting drunk-driving recidivism in previously convicted drunk-driving offenders. The ROAD study is a prospective study (2009-13) that was performed on 517 randomly selected drivers in Belgium. They were convicted for drunk-driving for which their licence was confiscated. The initial post-arrest blood samples were collected and analysed for percentage carbohydrate-deficient transferrin (%CDT), transaminsase activities [alanine amino transferase (ALT), aspartate amino transferase (AST)], gamma-glutamyltransferase (γGT) and red cell mean corpuscular volume (MCV). The observation time for each driver was 3 years and dynamic. A logistic regression analysis revealed that ln(%CDT) (P drunk-driving. The ROAD index (which includes ln(%CDT), ln(γGT), -ln(ALT) and the sex of the driver) was calculated and had a significantly higher area under the receiver operator characteristic curve (0.71) than the individual biomarkers for drunk-driving recidivism. Drivers with a high risk of recidivating (ROAD index ≥ 25%; third tertile) could be distinguished from drivers with an intermediate risk (16% ≤ ROAD index drunk-driving. The association with gamma-glutamyltransferase, alanine amino transferase and the sex of the driver could have additional value for identifying drunk-drivers at intermediate risk of recidivism. Non-specific indirect alcohol markers, such as alanine amino transferase, gamma-glutamyltransferase, aspartate amino transferase and red cell mean corpuscular volume have minimal added value to % carbohydrate-deficient transferrin for distinguishing drunk drivers with a low or high risk of recidivism. © 2013 Society for the Study of Addiction.

  16. Chromatin replication and epigenome maintenance

    DEFF Research Database (Denmark)

    Alabert, Constance; Groth, Anja

    2012-01-01

    initiates, whereas the replication process itself disrupts chromatin and challenges established patterns of genome regulation. Specialized replication-coupled mechanisms assemble new DNA into chromatin, but epigenome maintenance is a continuous process taking place throughout the cell cycle. If DNA...

  17. Chromatin replication and epigenome maintenance

    DEFF Research Database (Denmark)

    Alabert, Constance; Groth, Anja

    2012-01-01

    initiates, whereas the replication process itself disrupts chromatin and challenges established patterns of genome regulation. Specialized replication-coupled mechanisms assemble new DNA into chromatin, but epigenome maintenance is a continuous process taking place throughout the cell cycle. If DNA...

  18. Replicating Cardiovascular Condition-Birth Month Associations

    Science.gov (United States)

    Li, Li; Boland, Mary Regina; Miotto, Riccardo; Tatonetti, Nicholas P.; Dudley, Joel T.

    2016-01-01

    Independent replication is vital for study findings drawn from Electronic Health Records (EHR). This replication study evaluates the relationship between seasonal effects at birth and lifetime cardiovascular condition risk. We performed a Season-wide Association Study on 1,169,599 patients from Mount Sinai Hospital (MSH) to compute phenome-wide associations between birth month and CVD. We then evaluated if seasonal patterns found at MSH matched those reported at Columbia University Medical Center. Coronary arteriosclerosis, essential hypertension, angina, and pre-infarction syndrome passed phenome-wide significance and their seasonal patterns matched those previously reported. Atrial fibrillation, cardiomyopathy, and chronic myocardial ischemia had consistent patterns but were not phenome-wide significant. We confirm that CVD risk peaks for those born in the late winter/early spring among the evaluated patient populations. The replication findings bolster evidence for a seasonal birth month effect in CVD. Further study is required to identify the environmental and developmental mechanisms. PMID:27624541

  19. Infidelity of SARS-CoV Nsp14-Exonuclease Mutant Virus Replication Is Revealed by Complete Genome Sequencing

    Science.gov (United States)

    Eckerle, Lance D.; Becker, Michelle M.; Halpin, Rebecca A.; Li, Kelvin; Venter, Eli; Lu, Xiaotao; Scherbakova, Sana; Graham, Rachel L.; Baric, Ralph S.; Stockwell, Timothy B.; Spiro, David J.; Denison, Mark R.

    2010-01-01

    Most RNA viruses lack the mechanisms to recognize and correct mutations that arise during genome replication, resulting in quasispecies diversity that is required for pathogenesis and adaptation. However, it is not known how viruses encoding large viral RNA genomes such as the Coronaviridae (26 to 32 kb) balance the requirements for genome stability and quasispecies diversity. Further, the limits of replication infidelity during replication of large RNA genomes and how decreased fidelity impacts virus fitness over time are not known. Our previous work demonstrated that genetic inactivation of the coronavirus exoribonuclease (ExoN) in nonstructural protein 14 (nsp14) of murine hepatitis virus results in a 15-fold decrease in replication fidelity. However, it is not known whether nsp14-ExoN is required for replication fidelity of all coronaviruses, nor the impact of decreased fidelity on genome diversity and fitness during replication and passage. We report here the engineering and recovery of nsp14-ExoN mutant viruses of severe acute respiratory syndrome coronavirus (SARS-CoV) that have stable growth defects and demonstrate a 21-fold increase in mutation frequency during replication in culture. Analysis of complete genome sequences from SARS-ExoN mutant viral clones revealed unique mutation sets in every genome examined from the same round of replication and a total of 100 unique mutations across the genome. Using novel bioinformatic tools and deep sequencing across the full-length genome following 10 population passages in vitro, we demonstrate retention of ExoN mutations and continued increased diversity and mutational load compared to wild-type SARS-CoV. The results define a novel genetic and bioinformatics model for introduction and identification of multi-allelic mutations in replication competent viruses that will be powerful tools for testing the effects of decreased fidelity and increased quasispecies diversity on viral replication, pathogenesis, and

  20. [Postoperative radiotherapy in patients with invasive uterine cervix cancer treated previously with simple hysterectomy. Results from the Hospital de Oncología, Centro Médico Nacional SXXI. ].

    Science.gov (United States)

    Huerta Bahena, Judith; Labastida Almendaro, Sonia; Cortez Arroyo, Héctor; Calva, Angel

    2003-06-01

    Invasive carcinoma found after simple hysterectomy for several causes is an increasing clinical condition. That situation results in the necessity for further therapy: re-operation or radiotherapy. The purpose of this review is to analyze our results with postoperative radiotherapy for these patients. In 1998, 59 patients were submitted to postoperative radiotherapy at the Oncology Hospital, Medical Cancer Center for invasive cervical cancer found after simple hysterectomy for several causes. All patients had postoperative radiotherapy, the dose depended of the magnitude of residual disease after surgery (range 10-80 Gy). After surgery 27 patients had gross residual disease; and no residual disease or microscopic disease was present in 32 patients. Three years overall survival rate was 59% for the whole group. With the results of the univariate analysis, prognostic factors found that significantly affect disease free survival were interval between hysterectomy and radiotherapy of more than 6 months, radiotherapy dose less than 50 Grays and histology. With multivariate analysis only presence of gross residual disease affect significantly disease free survival p = 0.0000. Postoperative radiotherapy results in patients with invasor cervical cancer incidentally found in hysterectomy specimens with minimal residual disease are excellent. However with the presence of more extensive residual disease the results in disease free survival are worse than those obtained in patients with similar stages who had not been treated with hysterectomy. A well established diagnostic procedure must be done in all patients candidates to simple hysterectomy.

  1. Initiation of adenovirus DNA replication.

    OpenAIRE

    Reiter, T; Fütterer, J; Weingärtner, B; Winnacker, E L

    1980-01-01

    In an attempt to study the mechanism of initiation of adenovirus DNA replication, an assay was developed to investigate the pattern of DNA synthesis in early replicative intermediates of adenovirus DNA. By using wild-type virus-infected cells, it was possible to place the origin of adenovirus type 2 DNA replication within the terminal 350 to 500 base pairs from either of the two molecular termini. In addition, a variety of parameters characteristic of adenovirus DNA replication were compared ...

  2. Chromatin replication and epigenome maintenance

    DEFF Research Database (Denmark)

    Alabert, Constance; Groth, Anja

    2012-01-01

    Stability and function of eukaryotic genomes are closely linked to chromatin structure and organization. During cell division the entire genome must be accurately replicated and the chromatin landscape reproduced on new DNA. Chromatin and nuclear structure influence where and when DNA replication...... initiates, whereas the replication process itself disrupts chromatin and challenges established patterns of genome regulation. Specialized replication-coupled mechanisms assemble new DNA into chromatin, but epigenome maintenance is a continuous process taking place throughout the cell cycle. If DNA...

  3. Checkpoint responses to replication stalling: inducing tolerance and preventing mutagenesis

    Energy Technology Data Exchange (ETDEWEB)

    Kai, Mihoko; Wang, Teresa S.-F

    2003-11-27

    Replication mutants often exhibit a mutator phenotype characterized by point mutations, single base frameshifts, and the deletion or duplication of sequences flanked by homologous repeats. Mutation in genes encoding checkpoint proteins can significantly affect the mutator phenotype. Here, we use fission yeast (Schizosaccharomyces pombe) as a model system to discuss the checkpoint responses to replication perturbations induced by replication mutants. Checkpoint activation induced by a DNA polymerase mutant, aside from delay of mitotic entry, up-regulates the translesion polymerase DinB (Pol{kappa}). Checkpoint Rad9-Rad1-Hus1 (9-1-1) complex, which is loaded onto chromatin by the Rad17-Rfc2-5 checkpoint complex in response to replication perturbation, recruits DinB onto chromatin to generate the point mutations and single nucleotide frameshifts in the replication mutator. This chain of events reveals a novel checkpoint-induced tolerance mechanism that allows cells to cope with replication perturbation, presumably to make possible restarting stalled replication forks. Fission yeast Cds1 kinase plays an essential role in maintaining DNA replication fork stability in the face of DNA damage and replication fork stalling. Cds1 kinase is known to regulate three proteins that are implicated in maintaining replication fork stability: Mus81-Eme1, a hetero-dimeric structure-specific endonuclease complex; Rqh1, a RecQ-family helicase involved in suppressing inappropriate recombination during replication; and Rad60, a protein required for recombinational repair during replication. These Cds1-regulated proteins are thought to cooperatively prevent mutagenesis and maintain replication fork stability in cells under replication stress. These checkpoint-regulated processes allow cells to survive replication perturbation by preventing stalled replication forks from degenerating into deleterious DNA structures resulting in genomic instability and cancer development.

  4. No Previous Public Services Required

    Science.gov (United States)

    Taylor, Kelley R.

    2009-01-01

    In 2007, the Supreme Court heard a case that involved the question of whether a school district could be required to reimburse parents who unilaterally placed their child in private school when the child had not previously received special education and related services in a public institution ("Board of Education v. Tom F."). The…

  5. Spacetime replication of continuous variable quantum information

    Science.gov (United States)

    Hayden, Patrick; Nezami, Sepehr; Salton, Grant; Sanders, Barry C.

    2016-08-01

    The theory of relativity requires that no information travel faster than light, whereas the unitarity of quantum mechanics ensures that quantum information cannot be cloned. These conditions provide the basic constraints that appear in information replication tasks, which formalize aspects of the behavior of information in relativistic quantum mechanics. In this article, we provide continuous variable (CV) strategies for spacetime quantum information replication that are directly amenable to optical or mechanical implementation. We use a new class of homologically constructed CV quantum error correcting codes to provide efficient solutions for the general case of information replication. As compared to schemes encoding qubits, our CV solution requires half as many shares per encoded system. We also provide an optimized five-mode strategy for replicating quantum information in a particular configuration of four spacetime regions designed not to be reducible to previously performed experiments. For this optimized strategy, we provide detailed encoding and decoding procedures using standard optical apparatus and calculate the recovery fidelity when finite squeezing is used. As such we provide a scheme for experimentally realizing quantum information replication using quantum optics.

  6. Adenovirus E1A/E1B Transformed Amniotic Fluid Cells Support Human Cytomegalovirus Replication

    Directory of Open Access Journals (Sweden)

    Natascha Krömmelbein

    2016-02-01

    Full Text Available The human cytomegalovirus (HCMV replicates to high titers in primary human fibroblast cell cultures. A variety of primary human cells and some tumor-derived cell lines do also support permissive HCMV replication, yet at low levels. Cell lines established by transfection of the transforming functions of adenoviruses have been notoriously resistant to HCMV replication and progeny production. Here, we provide first-time evidence that a permanent cell line immortalized by adenovirus type 5 E1A and E1B (CAP is supporting the full HCMV replication cycle and is releasing infectious progeny. The CAP cell line had previously been established from amniotic fluid cells which were likely derived from membranes of the developing fetus. These cells can be grown under serum-free conditions. HCMV efficiently penetrated CAP cells, expressed its immediate-early proteins and dispersed restrictive PML-bodies. Viral DNA replication was initiated and viral progeny became detectable by electron microscopy in CAP cells. Furthermore, infectious virus was released from CAP cells, yet to lower levels compared to fibroblasts. Subviral dense bodies were also secreted from CAP cells. The results show that E1A/E1B expression in transformed cells is not generally repressive to HCMV replication and that CAP cells may be a good substrate for dense body based vaccine production.

  7. HIV-1 Vpr increases HCV replication through VprBP in cell culture.

    Science.gov (United States)

    Yan, Yanling; Huang, Fang; Yuan, Ting; Sun, Binlian; Yang, Rongge

    2016-09-02

    Coinfection of human immunodeficiency virus (HIV) and hepatitis C virus (HCV) occurs at a high frequency, in which HIV shows a promotion of HCV-derived liver diseases. However, the mechanism of how this occurs is not well understood. Our previous work has demonstrated that the HIV-1 accessory protein Vpr enhances HCV RNA replication in cell culture. Because Vpr performs most of its functions through host protein VprBP (DCAF1), the role of VprBP in the regulation of HCV by Vpr was investigated in this study. We found that the Vpr mutant Q65R, which is deficient in VprBP binding, could not enhance HCV replication. Furthermore, Vpr-mediated enhancement of HCV replication was severely diminished in VprBP knockdown cells. In addition, an inhibitor of Cullin RING E3 ligases, MLN4924, impaired the function of Vpr during HCV replication. Together, these results suggest that Vpr promotes HCV replication in a VprBP-dependent manner, and that the activity of Cullin RING E3 ligases is essential to this process. In conclusion, our findings demonstrate that HIV-1 Vpr makes the cellular environment more suitable for HCV replication, which might relate with the host ubiquitination system.

  8. Insights into the Determination of the Templating Nucleotide at the Initiation of φ29 DNA Replication.

    Science.gov (United States)

    del Prado, Alicia; Lázaro, José M; Longás, Elisa; Villar, Laurentino; de Vega, Miguel; Salas, Margarita

    2015-11-06

    Bacteriophage φ29 from Bacillus subtilis starts replication of its terminal protein (TP)-DNA by a protein-priming mechanism. To start replication, the DNA polymerase forms a heterodimer with a free TP that recognizes the replication origins, placed at both 5' ends of the linear chromosome, and initiates replication using as primer the OH-group of Ser-232 of the TP. The initiation of φ29 TP-DNA replication mainly occurs opposite the second nucleotide at the 3' end of the template. Earlier analyses of the template position that directs the initiation reaction were performed using single-stranded and double-stranded oligonucleotides containing the replication origin sequence without the parental TP. Here, we show that the parental TP has no influence in the determination of the nucleotide used as template in the initiation reaction. Previous studies showed that the priming domain of the primer TP determines the template position used for initiation. The results obtained here using mutant TPs at the priming loop where Ser-232 is located indicate that the aromatic residue Phe-230 is one of the determinants that allows the positioning of the penultimate nucleotide at the polymerization active site to direct insertion of the initiator dAMP during the initiation reaction. The role of Phe-230 in limiting the internalization of the template strand in the polymerization active site is discussed.

  9. Mycobacterium tuberculosis replicates within necrotic human macrophages

    Science.gov (United States)

    Lerner, Thomas R.; Repnik, Urska; Herbst, Susanne; Collinson, Lucy M.; Griffiths, Gareth

    2017-01-01

    Mycobacterium tuberculosis modulation of macrophage cell death is a well-documented phenomenon, but its role during bacterial replication is less characterized. In this study, we investigate the impact of plasma membrane (PM) integrity on bacterial replication in different functional populations of human primary macrophages. We discovered that IFN-γ enhanced bacterial replication in macrophage colony-stimulating factor–differentiated macrophages more than in granulocyte–macrophage colony-stimulating factor–differentiated macrophages. We show that permissiveness in the different populations of macrophages to bacterial growth is the result of a differential ability to preserve PM integrity. By combining live-cell imaging, correlative light electron microscopy, and single-cell analysis, we found that after infection, a population of macrophages became necrotic, providing a niche for M. tuberculosis replication before escaping into the extracellular milieu. Thus, in addition to bacterial dissemination, necrotic cells provide first a niche for bacterial replication. Our results are relevant to understanding the environment of M. tuberculosis replication in the host. PMID:28242744

  10. Correlação entre fratura por queda em idosos e uso prévio de medicamentos Correlation between fractures resulting from falls and previous drug use

    Directory of Open Access Journals (Sweden)

    Alberto Hamra

    2007-01-01

    Full Text Available O uso de medicamentos por idosos é freqüente e tem crescido a cada dia em virtude do aumento da expectativa de vida no Brasil. Algumas destas drogas quando administradas, podem provocar efeitos colaterais como tontura e diminuição dos reflexos podendo ocasionar quedas e conseqüentes fraturas. Este trabalho verificou se o uso de medicamentos pode ser considerado como fator de risco para fratura por queda no idoso. Para isso fez-se um estudo no ano de 2004 com 205 pacientes a partir de 60 anos de idade internados com fratura por queda e comparou-se estatisticamente com grupo-controle de 205 pacientes do mesmo grupo etário sem fratura. Verificou-se que o uso de medicamentos pode ser considerado como fator de risco para fratura por queda. Os resultados também podem servir como orientação para equipe médica, pacientes e seus familiares, no sentido de se tentar evitar quedas, principalmente quando o uso de medicamentos é necessário.The use of medication by elderly individuals is frequently and is constantly growing due to the increase of life expectance rates in Brazil. When some of these drugs are administered, they can cause collateral effects like dizziness and decreased reflex, potentially causing the occurrence of falls resulting in fractures. This study examined whether the use of medications by the elderly could be regarded as a risk factor to fractures resulting from falls. The study, conducted in 2004, assessed 205 hospitalized patients > 60 years old, with fractures resulting from falls. These patients were statistically compared to a control group (205 patients in the same age group and without fractures. We’ve found that the use of medication can be regarded as a risk factor to fractures resulting from falls. Our results could be useful to raise the awareness of doctors, patients and their families so that to avoid accidents when any drug therapy is required.

  11. Anatomy of Mammalian Replication Domains

    Science.gov (United States)

    Takebayashi, Shin-ichiro; Ogata, Masato; Okumura, Katsuzumi

    2017-01-01

    Genetic information is faithfully copied by DNA replication through many rounds of cell division. In mammals, DNA is replicated in Mb-sized chromosomal units called “replication domains.” While genome-wide maps in multiple cell types and disease states have uncovered both dynamic and static properties of replication domains, we are still in the process of understanding the mechanisms that give rise to these properties. A better understanding of the molecular basis of replication domain regulation will bring new insights into chromosome structure and function. PMID:28350365

  12. Effect of host cell lipid metabolism on alphavirus replication, virion morphogenesis, and infectivity.

    Science.gov (United States)

    Ng, Ching G; Coppens, Isabelle; Govindarajan, Dhanasekaran; Pisciotta, John; Shulaev, Vladimir; Griffin, Diane E

    2008-10-21

    The alphavirus Sindbis virus (SINV) causes encephalomyelitis in mice. Lipid-containing membranes, particularly cholesterol and sphingomyelin (SM), play important roles in virus entry, RNA replication, glycoprotein transport, and budding. Levels of SM are regulated by sphingomyelinases (SMases). Acid SMase (ASMase) deficiency results in the lipid storage disease type A Niemann-Pick disease (NPD-A), mimicked in mice by interruption of the ASMase gene. We previously demonstrated that ASMase-deficient mice are more susceptible to fatal SINV encephalomyelitis, with increased viral replication, spread, and neuronal death. To determine the mechanisms by which ASMase deficiency enhances SINV replication, we compared NPD-A fibroblasts (NPAF) to normal human fibroblasts (NHF). NPAF accumulated cholesterol- and sphingolipid-rich late endosomes/lysosomes in the perinuclear region. SINV replication was faster and reached higher titer in NPAF than in NHF, and NPAF died more quickly. SINV RNA and protein synthesis was greater in NHF than in NPAF, but virions budding from NPAF were 26 times more infectious and were regular dense particles whereas virions from NHF were larger particles containing substantial amounts of CD63. Cellular regulation of alphavirus morphogenesis is a previously unrecognized mechanism for control of virus replication and spread.

  13. Predicting Behavior Assessment System for Children-Second Edition Self-Report of Personality Child Form Results Using the Behavioral and Emotional Screening System Student Form: A Replication Study with an Urban, Predominantly Latino/a Sample

    Science.gov (United States)

    Kiperman, Sarah; Black, Mary S.; McGill, Tia M.; Harrell-Williams, Leigh M.; Kamphaus, Randy W.

    2014-01-01

    This study assesses the ability of a brief screening form, the Behavioral and Emotional Screening System-Student Form (BESS-SF), to predict scores on the much longer form from which it was derived: the Behavior Assessment System for Children-Second Edition Self-Report of Personality-Child Form (BASC-2-SRP-C). The present study replicates a former…

  14. Commercial Building Partnerships Replication and Diffusion

    Energy Technology Data Exchange (ETDEWEB)

    Antonopoulos, Chrissi A.; Dillon, Heather E.; Baechler, Michael C.

    2013-09-16

    This study presents findings from survey and interview data investigating replication efforts of Commercial Building Partnership (CBP) partners that worked directly with the Pacific Northwest National Laboratory (PNNL). PNNL partnered directly with 12 organizations on new and retrofit construction projects, which represented approximately 28 percent of the entire U.S. Department of Energy (DOE) CBP program. Through a feedback survey mechanism, along with personal interviews, PNNL gathered quantitative and qualitative data relating to replication efforts by each organization. These data were analyzed to provide insight into two primary research areas: 1) CBP partners’ replication efforts of technologies and approaches used in the CBP project to the rest of the organization’s building portfolio (including replication verification), and, 2) the market potential for technology diffusion into the total U.S. commercial building stock, as a direct result of the CBP program. The first area of this research focused specifically on replication efforts underway or planned by each CBP program participant. Factors that impact replication include motivation, organizational structure and objectives firms have for implementation of energy efficient technologies. Comparing these factors between different CBP partners revealed patterns in motivation for constructing energy efficient buildings, along with better insight into market trends for green building practices. The second area of this research develops a diffusion of innovations model to analyze potential broad market impacts of the CBP program on the commercial building industry in the United States.

  15. Systematic identification of novel, essential host genes affecting bromovirus RNA replication.

    Directory of Open Access Journals (Sweden)

    Brandi L Gancarz

    Full Text Available Positive-strand RNA virus replication involves viral proteins and cellular proteins at nearly every replication step. Brome mosaic virus (BMV is a well-established model for dissecting virus-host interactions and is one of very few viruses whose RNA replication, gene expression and encapsidation have been reproduced in the yeast Saccharomyces cerevisiae. Previously, our laboratory identified ∼100 non-essential host genes whose loss inhibited or enhanced BMV replication at least 3-fold. However, our isolation of additional BMV-modulating host genes by classical genetics and other results underscore that genes essential for cell growth also contribute to BMV RNA replication at a frequency that may be greater than that of non-essential genes. To systematically identify novel, essential host genes affecting BMV RNA replication, we tested a collection of ∼900 yeast strains, each with a single essential gene promoter replaced by a doxycycline-repressible promoter, allowing repression of gene expression by adding doxycycline to the growth medium. Using this strain array of ∼81% of essential yeast genes, we identified 24 essential host genes whose depleted expression reproducibly inhibited or enhanced BMV RNA replication. Relevant host genes are involved in ribosome biosynthesis, cell cycle regulation and protein homeostasis, among other cellular processes. BMV 2a(Pol levels were significantly increased in strains depleted for a heat shock protein (HSF1 or proteasome components (PRE1 and RPT6, suggesting these genes may affect BMV RNA replication by directly or indirectly modulating 2a(Pol localization, post-translational modification or interacting partners. Investigating the diverse functions of these newly identified essential host genes should advance our understanding of BMV-host interactions and normal cellular pathways, and suggest new modes of virus control.

  16. Systematic identification of novel, essential host genes affecting bromovirus RNA replication.

    Science.gov (United States)

    Gancarz, Brandi L; Hao, Linhui; He, Qiuling; Newton, Michael A; Ahlquist, Paul

    2011-01-01

    Positive-strand RNA virus replication involves viral proteins and cellular proteins at nearly every replication step. Brome mosaic virus (BMV) is a well-established model for dissecting virus-host interactions and is one of very few viruses whose RNA replication, gene expression and encapsidation have been reproduced in the yeast Saccharomyces cerevisiae. Previously, our laboratory identified ∼100 non-essential host genes whose loss inhibited or enhanced BMV replication at least 3-fold. However, our isolation of additional BMV-modulating host genes by classical genetics and other results underscore that genes essential for cell growth also contribute to BMV RNA replication at a frequency that may be greater than that of non-essential genes. To systematically identify novel, essential host genes affecting BMV RNA replication, we tested a collection of ∼900 yeast strains, each with a single essential gene promoter replaced by a doxycycline-repressible promoter, allowing repression of gene expression by adding doxycycline to the growth medium. Using this strain array of ∼81% of essential yeast genes, we identified 24 essential host genes whose depleted expression reproducibly inhibited or enhanced BMV RNA replication. Relevant host genes are involved in ribosome biosynthesis, cell cycle regulation and protein homeostasis, among other cellular processes. BMV 2a(Pol) levels were significantly increased in strains depleted for a heat shock protein (HSF1) or proteasome components (PRE1 and RPT6), suggesting these genes may affect BMV RNA replication by directly or indirectly modulating 2a(Pol) localization, post-translational modification or interacting partners. Investigating the diverse functions of these newly identified essential host genes should advance our understanding of BMV-host interactions and normal cellular pathways, and suggest new modes of virus control.

  17. A dynamic stochastic model for DNA replication initiation in early embryos.

    Directory of Open Access Journals (Sweden)

    Arach Goldar

    Full Text Available BACKGROUND: Eukaryotic cells seem unable to monitor replication completion during normal S phase, yet must ensure a reliable replication completion time. This is an acute problem in early Xenopus embryos since DNA replication origins are located and activated stochastically, leading to the random completion problem. DNA combing, kinetic modelling and other studies using Xenopus egg extracts have suggested that potential origins are much more abundant than actual initiation events and that the time-dependent rate of initiation, I(t, markedly increases through S phase to ensure the rapid completion of unreplicated gaps and a narrow distribution of completion times. However, the molecular mechanism that underlies this increase has remained obscure. METHODOLOGY/PRINCIPAL FINDINGS: Using both previous and novel DNA combing data we have confirmed that I(t increases through S phase but have also established that it progressively decreases before the end of S phase. To explore plausible biochemical scenarios that might explain these features, we have performed comparisons between numerical simulations and DNA combing data. Several simple models were tested: i recycling of a limiting replication fork component from completed replicons; ii time-dependent increase in origin efficiency; iii time-dependent increase in availability of an initially limiting factor, e.g. by nuclear import. None of these potential mechanisms could on its own account for the data. We propose a model that combines time-dependent changes in availability of a replication factor and a fork-density dependent affinity of this factor for potential origins. This novel model quantitatively and robustly accounted for the observed changes in initiation rate and fork density. CONCLUSIONS/SIGNIFICANCE: This work provides a refined temporal profile of replication initiation rates and a robust, dynamic model that quantitatively explains replication origin usage during early embryonic S phase

  18. Intensive versus Guideline Blood Pressure and Lipid Lowering in Patients with Previous Stroke: Main Results from the Pilot ‘Prevention of Decline in Cognition after Stroke Trial’ (PODCAST) Randomised Controlled Trial

    Science.gov (United States)

    Scutt, Polly; Blackburn, Daniel J.; Ankolekar, Sandeep; Krishnan, Kailash; Ballard, Clive; Burns, Alistair; Mant, Jonathan; Passmore, Peter; Pocock, Stuart; Reckless, John; Sprigg, Nikola; Stewart, Rob; Wardlaw, Joanna M.; Ford, Gary A.

    2017-01-01

    Background Stroke is associated with the development of cognitive impairment and dementia. We assessed the effect of intensive blood pressure (BP) and/or lipid lowering on cognitive outcomes in patients with recent stroke in a pilot trial. Methods In a multicentre, partial-factorial trial, patients with recent stroke, absence of dementia, and systolic BP (SBP) 125–170 mmHg were assigned randomly to at least 6 months of intensive (target SBP <125 mmHg) or guideline (target SBP <140 mmHg) BP lowering. The subset of patients with ischaemic stroke and total cholesterol 3.0–8.0 mmol/l were also assigned randomly to intensive (target LDL-cholesterol <1.3 mmol/l) or guideline (target LDL-c <3.0 mmol/l) lipid lowering. The primary outcome was the Addenbrooke’s Cognitive Examination-Revised (ACE-R). Results We enrolled 83 patients, mean age 74.0 (6.8) years, and median 4.5 months after stroke. The median follow-up was 24 months (range 1–48). Mean BP was significantly reduced with intensive compared to guideline treatment (difference –10·6/–5·5 mmHg; p<0·01), as was total/LDL-cholesterol with intensive lipid lowering compared to guideline (difference –0·54/–0·44 mmol/l; p<0·01). The ACE-R score during treatment did not differ for either treatment comparison; mean difference for BP lowering -3.6 (95% CI -9.7 to 2.4), and lipid lowering 4.4 (95% CI -2.1 to 10.9). However, intensive lipid lowering therapy was significantly associated with improved scores for ACE-R at 6 months, trail making A, modified Rankin Scale and Euro-Qol Visual Analogue Scale. There was no difference in rates of dementia or serious adverse events for either comparison. Conclusion In patients with recent stroke and normal cognition, intensive BP and lipid lowering were feasible and safe, but did not alter cognition over two years. The association between intensive lipid lowering and improved scores for some secondary outcomes suggests further trials are warranted. Trial Registration

  19. The Genomic Replication of the Crenarchaeal Virus SIRV2

    DEFF Research Database (Denmark)

    Martinez Alvarez, Laura

    of the crenarchaeal virus SIRV2, a model among archaeal viruses. SIRV2 was found to employ multiple replication mechanisms, with DNA synthesis starting by a strand-displacement mode that later derived in a rolling-circle replication from a circular intermediate. Interestingly, evidence for a secondary, bidirectional...... reinitiation events may partially explain the branched topology of the viral replication intermediates. We also analyzed the intracellular location of viral replication, showing the formation of viral peripheral replication centers in SIRV2-infected cells, where viral DNA synthesis and replication......-related proteins are concentrated. Our data indicates that the host DNA polymerase Dpo1 is also the viral replicative polymerase. Moreover, additional data suggests that other processes, such as translation, are also reorganized after infection. Based on our results, a model for the organization of viral...

  20. Replication, falsification, and the crisis of confidence in social psychology

    Directory of Open Access Journals (Sweden)

    Brian D. Earp

    2015-05-01

    Full Text Available The (latest crisis in confidence in social psychology has generated much heated discussion about the importance of replication, including how such replication should be carried out as well as interpreted by scholars in the field. What does it mean if a replication attempt fails—does it mean that the original results, or the theory that predicted them, have been falsified? And how should failed replications affect our belief in the validity of the original research? In this paper, we consider the replication debate from a historical and philosophical perspective, and provide a conceptual analysis of both replication and falsification as they pertain to this important discussion. Along the way, we introduce a Bayesian framework for assessing failed replications in terms of how they should affect our confidence in purported findings.

  1. Replication, falsification, and the crisis of confidence in social psychology.

    Science.gov (United States)

    Earp, Brian D; Trafimow, David

    2015-01-01

    The (latest) crisis in confidence in social psychology has generated much heated discussion about the importance of replication, including how it should be carried out as well as interpreted by scholars in the field. For example, what does it mean if a replication attempt "fails"-does it mean that the original results, or the theory that predicted them, have been falsified? And how should "failed" replications affect our belief in the validity of the original research? In this paper, we consider the replication debate from a historical and philosophical perspective, and provide a conceptual analysis of both replication and falsification as they pertain to this important discussion. Along the way, we highlight the importance of auxiliary assumptions (for both testing theories and attempting replications), and introduce a Bayesian framework for assessing "failed" replications in terms of how they should affect our confidence in original findings.

  2. Replication, falsification, and the crisis of confidence in social psychology

    Science.gov (United States)

    Earp, Brian D.; Trafimow, David

    2015-01-01

    The (latest) crisis in confidence in social psychology has generated much heated discussion about the importance of replication, including how it should be carried out as well as interpreted by scholars in the field. For example, what does it mean if a replication attempt “fails”—does it mean that the original results, or the theory that predicted them, have been falsified? And how should “failed” replications affect our belief in the validity of the original research? In this paper, we consider the replication debate from a historical and philosophical perspective, and provide a conceptual analysis of both replication and falsification as they pertain to this important discussion. Along the way, we highlight the importance of auxiliary assumptions (for both testing theories and attempting replications), and introduce a Bayesian framework for assessing “failed” replications in terms of how they should affect our confidence in original findings. PMID:26042061

  3. Novel Mutant AAV2 Rep Proteins Support AAV2 Replication without Blocking HSV-1 Helpervirus Replication

    Science.gov (United States)

    Seyffert, Michael; Glauser, Daniel L.; Schraner, Elisabeth M.; de Oliveira, Anna-Paula; Mansilla-Soto, Jorge; Vogt, Bernd; Büning, Hildegard; Linden, R. Michael; Ackermann, Mathias; Fraefel, Cornel

    2017-01-01

    As their names imply, parvoviruses of the genus Dependovirus rely for their efficient replication on the concurrent presence of a helpervirus, such as herpesvirus, adenovirus, or papilloma virus. Adeno-associated virus 2 (AAV2) is such an example, which in turn can efficiently inhibit the replication of each helpervirus by distinct mechanisms. In a previous study we have shown that expression of the AAV2 rep gene is not compatible with efficient replication of herpes simplex virus 1 (HSV-1). In particular, the combined DNA-binding and ATPase/helicase activities of the Rep68/78 proteins have been shown to exert opposite effects on the replication of AAV2 and HSV-1. While essential for AAV2 DNA replication these protein activities account for the Rep-mediated inhibition of HSV-1 replication. Here, we describe a novel Rep mutant (Rep-D371Y), which displayed an unexpected phenotype. Rep-D371Y did not block HSV-1 replication, but still supported efficient AAV2 replication, at least when a double-stranded AAV2 genome template was used. We also found that the capacity of Rep-D371Y to induce apoptosis and a Rep-specific DNA damage response was significantly reduced compared to wild-type Rep. These findings suggest that AAV2 Rep-helicase subdomains exert diverging activities, which contribute to distinct steps of the AAV2 life cycle. More important, the novel AAV2 mutant Rep-D371Y may allow deciphering yet unsolved activities of the AAV2 Rep proteins such as DNA second-strand synthesis, genomic integration or packaging, which all involve the Rep-helicase activity. PMID:28125695

  4. Replicated Spectrographs in Astronomy

    CERN Document Server

    Hill, Gary J

    2014-01-01

    As telescope apertures increase, the challenge of scaling spectrographic astronomical instruments becomes acute. The next generation of extremely large telescopes (ELTs) strain the availability of glass blanks for optics and engineering to provide sufficient mechanical stability. While breaking the relationship between telescope diameter and instrument pupil size by adaptive optics is a clear path for small fields of view, survey instruments exploiting multiplex advantages will be pressed to find cost-effective solutions. In this review we argue that exploiting the full potential of ELTs will require the barrier of the cost and engineering difficulty of monolithic instruments to be broken by the use of large-scale replication of spectrographs. The first steps in this direction have already been taken with the soon to be commissioned MUSE and VIRUS instruments for the Very Large Telescope and the Hobby-Eberly Telescope, respectively. MUSE employs 24 spectrograph channels, while VIRUS has 150 channels. We compa...

  5. SUMO and KSHV Replication

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Pei-Ching [Institute of Microbiology and Immunology, National Yang-Ming University, Taipei 112, Taiwan (China); Kung, Hsing-Jien, E-mail: hkung@nhri.org.tw [Institute for Translational Medicine, College of Medical Science and Technology, Taipei Medical University, Taipei 110, Taiwan (China); Department of Biochemistry and Molecular Medicine, University of California, Davis, CA 95616 (United States); UC Davis Cancer Center, University of California, Davis, CA 95616 (United States); Division of Molecular and Genomic Medicine, National Health Research Institutes, 35 Keyan Road, Zhunan, Miaoli County 35053, Taiwan (China)

    2014-09-29

    Small Ubiquitin-related MOdifier (SUMO) modification was initially identified as a reversible post-translational modification that affects the regulation of diverse cellular processes, including signal transduction, protein trafficking, chromosome segregation, and DNA repair. Increasing evidence suggests that the SUMO system also plays an important role in regulating chromatin organization and transcription. It is thus not surprising that double-stranded DNA viruses, such as Kaposi’s sarcoma-associated herpesvirus (KSHV), have exploited SUMO modification as a means of modulating viral chromatin remodeling during the latent-lytic switch. In addition, SUMO regulation allows the disassembly and assembly of promyelocytic leukemia protein-nuclear bodies (PML-NBs), an intrinsic antiviral host defense, during the viral replication cycle. Overcoming PML-NB-mediated cellular intrinsic immunity is essential to allow the initial transcription and replication of the herpesvirus genome after de novo infection. As a consequence, KSHV has evolved a way as to produce multiple SUMO regulatory viral proteins to modulate the cellular SUMO environment in a dynamic way during its life cycle. Remarkably, KSHV encodes one gene product (K-bZIP) with SUMO-ligase activities and one gene product (K-Rta) that exhibits SUMO-targeting ubiquitin ligase (STUbL) activity. In addition, at least two viral products are sumoylated that have functional importance. Furthermore, sumoylation can be modulated by other viral gene products, such as the viral protein kinase Orf36. Interference with the sumoylation of specific viral targets represents a potential therapeutic strategy when treating KSHV, as well as other oncogenic herpesviruses. Here, we summarize the different ways KSHV exploits and manipulates the cellular SUMO system and explore the multi-faceted functions of SUMO during KSHV’s life cycle and pathogenesis.

  6. The N-Terminal of Aquareovirus NS80 Is Required for Interacting with Viral Proteins and Viral Replication.

    Directory of Open Access Journals (Sweden)

    Jie Zhang

    Full Text Available Reovirus replication and assembly occurs within viral inclusion bodies that formed in specific intracellular compartments of cytoplasm in infected cells. Previous study indicated that aquareovirus NS80 is able to form inclusion bodies, and also can retain viral proteins within its inclusions. To better understand how NS80 performed in viral replication and assembly, the functional regions of NS80 associated with other viral proteins in aquareovirus replication were investigated in this study. Deletion mutational analysis and rotavirus NSP5-based protein association platform were used to detect association regions. Immunofluorescence images indicated that different N-terminal regions of NS80 could associate with viral proteins VP1, VP4, VP6 and NS38. Further co-immunoprecipitation analysis confirmed the interaction between VP1, VP4, VP6 or NS38 with different regions covering the N-terminal amino acid (aa, 1-471 of NS80, respectively. Moreover, removal of NS80 N-terminal sequences required for interaction with proteins VP1, VP4, VP6 or NS38 not only prevented the capacity of NS80 to support viral replication in NS80 shRNA-based replication complementation assays, but also inhibited the expression of aquareovirus proteins, suggesting that N-terminal regions of NS80 are necessary for viral replication. These results provided a foundational basis for further understanding the role of NS80 in viral replication and assembly during aquareovirus infection.

  7. The Deacetylase SIRT1 Regulates the Replication Properties of Human Papillomavirus 16 E1 and E2.

    Science.gov (United States)

    Das, Dipon; Smith, Nathan; Wang, Xu; Morgan, Iain M

    2017-05-15

    Human papillomaviruses (HPV) replicate their genomes in differentiating epithelium using the viral proteins E1 and E2 in association with host proteins. While the roles of E1 and E2 in this process are understood, the host factors involved and how they interact with and regulate E1-E2 are not. Our previous work identified the host replication and repair factor TopBP1 as an E2 partner protein essential for optimal E1-E2 replication and for the viral life cycle. The role of TopBP1 in host DNA replication is regulated by the class III deacetylase SIRT1; activation of the DNA damage response prevents SIRT1 deacetylation of TopBP1, resulting in a switch from DNA replication to repair functions for this protein and cell cycle arrest. Others have demonstrated an essential role for SIRT1 in regulation of the HPV31 life cycle; here, we report that SIRT1 can directly regulate HPV16 E1-E2-mediated DNA replication. SIRT1 is part of the E1-E2 DNA replication complex and is recruited to the viral origin of replication in an E1-E2-dependent manner. CRISPR/Cas9 was used to generate C33a clones with undetectable SIRT1 expression and lack of SIRT1 elevated E1-E2 DNA replication, in part due to increased acetylation and stabilization of the E2 protein in the absence of SIRT1. The results demonstrate that SIRT1 is a member of, and can regulate, the HPV16 replication complex. We discuss the potential role of this protein in the viral life cycle.IMPORTANCE HPV are causative agents in a number of human diseases, and currently only the symptoms of these diseases are treated. To identify novel therapeutic approaches for combating these diseases, the viral life cycle must be understood in more detail. This report demonstrates that a cellular enzyme, SIRT1, is part of the HPV16 DNA replication complex and is brought to the viral genome by the viral proteins E1 and E2. Using gene editing technology (CRISPR/Cas9), the SIRT1 gene was removed from cervical cancer cells. The consequence of this

  8. Replication study of 34 common SNPs associated with prostate cancer in the Romanian population.

    Science.gov (United States)

    Jinga, Viorel; Csiki, Irma Eva; Manolescu, Andrei; Iordache, Paul; Mates, Ioan Nicolae; Radavoi, Daniel; Rascu, Stefan; Badescu, Daniel; Badea, Paula; Mates, Dana

    2016-04-01

    Prostate cancer is the third-most common form of cancer in men in Romania. The Romanian unscreened population represents a good sample to study common genetic risk variants. However, a comprehensive analysis has not been conducted yet. Here, we report our replication efforts in a Romanian population of 979 cases and 1027 controls, for potential association of 34 literature-reported single nucleotide polymorphisms (SNPs) with prostate cancer. We also examined whether any SNP was differentially associated with tumour grade or stage at diagnosis, with disease aggressiveness, and with the levels of PSA (prostate specific antigen). In the allelic analysis, we replicated the previously reported risk for 19 loci on 4q24, 6q25.3, 7p15.2, 8q24.21, 10q11.23, 10q26.13, 11p15.5, 11q13.2, 11q13.3. Statistically significant associations were replicated for other six SNPs only with a particular disease phenotype: low-grade tumour and low PSA levels (rs1512268), high PSA levels (rs401681 and rs11649743), less aggressive cancers (rs1465618, rs721048, rs17021918). The strongest association of our tested SNP's with PSA in controls was for rs2735839, with 29% increase for each copy of the major allele G, consistent with previous results. Our results suggest that rs4962416, previously associated only with prostate cancer, is also associated with PSA levels, with 12% increase for each copy of the minor allele C. The study enabled the replication of the effect for the majority of previously reported genetic variants in a set of clinically relevant prostate cancers. This is the first replication study on these loci, known to associate with prostate cancer, in a Romanian population.

  9. Replication of Prostate Cancer Risk Variants in a Danish Case-Control Association Study

    DEFF Research Database (Denmark)

    Bentzon, Diem Nguyen; Nyegaard, Mette; Børglum, Anders

    2012-01-01

    assays and associations between SNPs, prostate cancer risk, and clinico-pathological variables were assessed. Results: Seventeen SNPs were successfully replicated in our case-control study and the association estimates were consistent with previous reports. Four markers were excluded from further...... (P = 0.045). In addition, variants rs6983267 (GG) and rs5759167 (GG/GT) were significantly associated with negative family history (P = 0.04 and P = 0.02, respectively). Conclusion: We replicated 17 previously identified prostate cancer-associated risk SNPs in a Danish case-control study and found...... developed to predict prostate cancer risk. The association between genetic markers and clinico-pathological tumor variables has, however, been inconsistent. Methods and Materials: A total of 32 previously identified prostate cancer-associated risk SNPs were genotyped in 648 prostate cancer cases and 526 age...

  10. Cellular peptidyl-prolyl cis/trans isomerase Pin1 facilitates replication of feline coronavirus.

    Science.gov (United States)

    Tanaka, Yoshikazu; Amano, Arisa; Morisaki, Masateru; Sato, Yuka; Sasaki, Takashi

    2016-02-01

    Although feline coronavirus (FCoV) causes feline infectious peritonitis (FIP), which is a fatal infectious disease, there are no effective therapeutic medicines or vaccines. Previously, in vitro studies have shown that cyclosporin (CsA) and FK506 inhibit virus replication in diverse coronaviruses. CsA and FK506 are targets of clinically relevant immunosuppressive drugs and bind to cellular cyclophilins (Cyps) or FK506 binding proteins (FKBPs), respectively. Both Cyp and FKBP have peptidyl-prolyl cis-trans isomerase (PPIase) activity. However, protein interacting with NIMA (Pin1), a member of the parvulin subfamily of PPIases that differs from Cyps and FKBPs, is essential for various signaling pathways. Here we demonstrated that genetic silencing or knockout of Pin1 resulted in decreased FCoV replication in vitro. Dipentamethylene thiuram monosulfide, a specific inhibitor of Pin1, inhibited FCoV replication. These data indicate that Pin1 modulates FCoV propagation.

  11. PAR-4/LKB1 regulates DNA replication during asynchronous division of the early C. elegans embryo

    Science.gov (United States)

    Descoteaux, Catherine; Chartier, Nicolas T.; Pintard, Lionel; Labbé, Jean-Claude

    2014-01-01

    Regulation of cell cycle duration is critical during development, yet the underlying molecular mechanisms are still poorly understood. The two-cell stage Caenorhabditis elegans embryo divides asynchronously and thus provides a powerful context in which to study regulation of cell cycle timing during development. Using genetic analysis and high-resolution imaging, we found that deoxyribonucleic acid (DNA) replication is asymmetrically regulated in the two-cell stage embryo and that the PAR-4 and PAR-1 polarity proteins dampen DNA replication dynamics specifically in the posterior blastomere, independently of regulators previously implicated in the control of cell cycle timing. Our results demonstrate that accurate control of DNA replication is crucial during C. elegans early embryonic development and further provide a novel mechanism by which PAR proteins control cell cycle progression during asynchronous cell division. PMID:24841566

  12. Efficient usage of Adabas replication

    CERN Document Server

    Storr, Dieter W

    2011-01-01

    In today's IT organization replication becomes more and more an essential technology. This makes Software AG's Event Replicator for Adabas an important part of your data processing. Setting the right parameters and establishing the best network communication, as well as selecting efficient target components, is essential for successfully implementing replication. This book provides comprehensive information and unique best-practice experience in the field of Event Replicator for Adabas. It also includes sample codes and configurations making your start very easy. It describes all components ne

  13. Solving the Telomere Replication Problem

    Science.gov (United States)

    Maestroni, Laetitia; Matmati, Samah; Coulon, Stéphane

    2017-01-01

    Telomeres are complex nucleoprotein structures that protect the extremities of linear chromosomes. Telomere replication is a major challenge because many obstacles to the progression of the replication fork are concentrated at the ends of the chromosomes. This is known as the telomere replication problem. In this article, different and new aspects of telomere replication, that can threaten the integrity of telomeres, will be reviewed. In particular, we will focus on the functions of shelterin and the replisome for the preservation of telomere integrity. PMID:28146113

  14. 3D Spatially Resolved Models of the Intracellular Dynamics of the Hepatitis C Genome Replication Cycle

    KAUST Repository

    Knodel, Markus

    2017-10-02

    Mathematical models of virus dynamics have not previously acknowledged spatial resolution at the intracellular level despite substantial arguments that favor the consideration of intracellular spatial dependence. The replication of the hepatitis C virus (HCV) viral RNA (vRNA) occurs within special replication complexes formed from membranes derived from endoplasmatic reticulum (ER). These regions, termed membranous webs, are generated primarily through specific interactions between nonstructural virus-encoded proteins (NSPs) and host cellular factors. The NSPs are responsible for the replication of the vRNA and their movement is restricted to the ER surface. Therefore, in this study we developed fully spatio-temporal resolved models of the vRNA replication cycle of HCV. Our simulations are performed upon realistic reconstructed cell structures-namely the ER surface and the membranous webs-based on data derived from immunostained cells replicating HCV vRNA. We visualized 3D simulations that reproduced dynamics resulting from interplay of the different components of our models (vRNA, NSPs, and a host factor), and we present an evaluation of the concentrations for the components within different regions of the cell. Thus far, our model is restricted to an internal portion of a hepatocyte and is qualitative more than quantitative. For a quantitative adaption to complete cells, various additional parameters will have to be determined through further in vitro cell biology experiments, which can be stimulated by the results deccribed in the present study.

  15. ReplicationDomain: a visualization tool and comparative database for genome-wide replication timing data

    Directory of Open Access Journals (Sweden)

    Yokochi Tomoki

    2008-12-01

    Full Text Available Abstract Background Eukaryotic DNA replication is regulated at the level of large chromosomal domains (0.5–5 megabases in mammals within which replicons are activated relatively synchronously. These domains replicate in a specific temporal order during S-phase and our genome-wide analyses of replication timing have demonstrated that this temporal order of domain replication is a stable property of specific cell types. Results We have developed ReplicationDomain http://www.replicationdomain.org as a web-based database for analysis of genome-wide replication timing maps (replication profiles from various cell lines and species. This database also provides comparative information of transcriptional expression and is configured to display any genome-wide property (for instance, ChIP-Chip or ChIP-Seq data via an interactive web interface. Our published microarray data sets are publicly available. Users may graphically display these data sets for a selected genomic region and download the data displayed as text files, or alternatively, download complete genome-wide data sets. Furthermore, we have implemented a user registration system that allows registered users to upload their own data sets. Upon uploading, registered users may choose to: (1 view their data sets privately without sharing; (2 share with other registered users; or (3 make their published or "in press" data sets publicly available, which can fulfill journal and funding agencies' requirements for data sharing. Conclusion ReplicationDomain is a novel and powerful tool to facilitate the comparative visualization of replication timing in various cell types as well as other genome-wide chromatin features and is considerably faster and more convenient than existing browsers when viewing multi-megabase segments of chromosomes. Furthermore, the data upload function with the option of private viewing or sharing of data sets between registered users should be a valuable resource for the

  16. USP7 is a SUMO deubiquitinase essential for DNA replication

    DEFF Research Database (Denmark)

    Lecona, Emilio; Rodriguez-Acebes, Sara; Specks, Julia

    2016-01-01

    Post-translational modification of proteins by ubiquitin (Ub) and Ub-like modifiers regulates DNA replication. We have previously shown that chromatin around replisomes is rich in SUMO and poor in Ub, whereas mature chromatin exhibits an opposite pattern. How this SUMO-rich, Ub-poor environment...... is maintained at sites of DNA replication in mammalian cells remains unexplored. Here we identify USP7 as a replisome-enriched SUMO deubiquitinase that is essential for DNA replication. By acting on SUMO and SUMOylated proteins, USP7 counteracts their ubiquitination. Inhibition or genetic deletion of USP7 leads...... to the accumulation of Ub on SUMOylated proteins, which are displaced away from replisomes. Our findings provide a model explaining the differential accumulation of SUMO and Ub at replication forks and identify an essential role of USP7 in DNA replication that should be considered in the development of USP7...

  17. USP7 is a SUMO deubiquitinase essential for DNA replication.

    Science.gov (United States)

    Lecona, Emilio; Rodriguez-Acebes, Sara; Specks, Julia; Lopez-Contreras, Andres J; Ruppen, Isabel; Murga, Matilde; Muñoz, Javier; Mendez, Juan; Fernandez-Capetillo, Oscar

    2016-04-01

    Post-translational modification of proteins by ubiquitin (Ub) and Ub-like modifiers regulates DNA replication. We have previously shown that chromatin around replisomes is rich in SUMO and poor in Ub, whereas mature chromatin exhibits an opposite pattern. How this SUMO-rich, Ub-poor environment is maintained at sites of DNA replication in mammalian cells remains unexplored. Here we identify USP7 as a replisome-enriched SUMO deubiquitinase that is essential for DNA replication. By acting on SUMO and SUMOylated proteins, USP7 counteracts their ubiquitination. Inhibition or genetic deletion of USP7 leads to the accumulation of Ub on SUMOylated proteins, which are displaced away from replisomes. Our findings provide a model explaining the differential accumulation of SUMO and Ub at replication forks and identify an essential role of USP7 in DNA replication that should be considered in the development of USP7 inhibitors as anticancer agents.

  18. An optimal replication strategy for data grid systems

    Institute of Scientific and Technical Information of China (English)

    JIANG Jianjin; YANG Guangwen

    2007-01-01

    Data access latency is an important metric of system performance in data grid.By means of efficient replication strategy,the amount of data transferred in a wide area network will decrease,and the average access latency of data will decrease ultimately.The motivation of our research is to solve the optimized replica distribution problem in a data grid;that is,the system should utilize many replicas for every data with storage constraints to minimize the average access latency of data.This paper proposes a model of replication strategy in federated data grid and gives the optimized solution.The analysis results and simulation results show that the optimized replication strategy proposed in this paper is superior to LRU caching strategy,uniform replication strategy,proportional replication strategy and square root replication strategy in terms of wide area network bandwidth requirement and in the average access latency of data.

  19. Caspase-3-mediated cleavage of p65/RelA results in a carboxy-terminal fragment that inhibits IκBα and enhances HIV-1 replication in human T lymphocytes

    Directory of Open Access Journals (Sweden)

    Alcamí José

    2008-12-01

    Full Text Available Abstract Background Degradation of p65/RelA has been involved in both the inhibition of NF-κB-dependent activity and the onset of apoptosis. However, the mechanisms of NF-κB degradation are unclear and can vary depending on the cell type. Cleavage of p65/RelA can produce an amino-terminal fragment that was shown to act as a dominant-negative inhibitor of NF-κB, thereby promoting apoptosis. However, the opposite situation has also been described and the production of a carboxy-terminal fragment that contains two potent transactivation domains has also been related to the onset of apoptosis. In this context, a carboxy-terminal fragment of p65/RelA (ΔNH2p65, detected in non-apoptotic human T lymphocytes upon activation, has been studied. T cells constitute one of the long-lived cellular reservoirs of the human immunodeficiency virus type 1 (HIV-1. Because NF-κB is the most important inducible element involved in initiation of HIV-1 transcription, an adequate control of NF-κB response is of paramount importance for both T cell survival and viral spread. Its major inhibitor IκBα constitutes a master terminator of NF-κB response that is complemented by degradation of p65/RelA. Results and conclusions In this study, the function of a caspase-3-mediated carboxy-terminal fragment of p65/RelA, which was detected in activated human peripheral blood lymphocytes (PBLs, was analyzed. Cells producing this truncated p65/RelA did not undergo apoptosis but showed a high viability, in spite of caspase-3 activation. ΔNH2p65 lacked most of DNA-binding domain but retained the dimerization domain, NLS and transactivation domains. Consequently, it could translocate to the nucleus, associate with NF-κB1/p50 and IκBα, but could not bind -κB consensus sites. However, although ΔNH2p65 lacked transcriptional activity by itself, it could increase NF-κB activity in a dose-dependent manner by hijacking IκBα. Thus, its expression resulted in a persistent

  20. Charter School Replication. Policy Guide

    Science.gov (United States)

    Rhim, Lauren Morando

    2009-01-01

    "Replication" is the practice of a single charter school board or management organization opening several more schools that are each based on the same school model. The most rapid strategy to increase the number of new high-quality charter schools available to children is to encourage the replication of existing quality schools. This policy guide…

  1. Control of helicase loading in the coupled DNA replication and recombination systems of bacteriophage T4.

    Science.gov (United States)

    Branagan, Amy M; Klein, Jenny A; Jordan, Christian S; Morrical, Scott W

    2014-01-31

    The Gp59 protein of bacteriophage T4 promotes DNA replication by loading the replicative helicase, Gp41, onto replication forks and recombination intermediates. Gp59 also blocks DNA synthesis by Gp43 polymerase until Gp41 is loaded, ensuring that synthesis is tightly coupled to unwinding. The distinct polymerase blocking and helicase loading activities of Gp59 likely involve different binding interactions with DNA and protein partners. Here, we investigate how interactions of Gp59 with DNA and Gp32, the T4 single-stranded DNA (ssDNA)-binding protein, are related to these activities. A previously characterized mutant, Gp59-I87A, exhibits markedly reduced affinity for ssDNA and pseudo-fork DNA substrates. We demonstrate that on Gp32-covered ssDNA, the DNA binding defect of Gp59-I87A is not detrimental to helicase loading and translocation. In contrast, on pseudo-fork DNA the I87A mutation is detrimental to helicase loading and unwinding in the presence or absence of Gp32. Other results indicate that Gp32 binding to lagging strand ssDNA relieves the blockage of Gp43 polymerase activity by Gp59, whereas the inhibition of Gp43 exonuclease activity is maintained. Our findings suggest that Gp59-Gp32 and Gp59-DNA interactions perform separate but complementary roles in T4 DNA metabolism; Gp59-Gp32 interactions are needed to load Gp41 onto D-loops, and other nucleoprotein structures containing clusters of Gp32. Gp59-DNA interactions are needed to load Gp41 onto nascent or collapsed replication forks lacking clusters of Gp32 and to coordinate bidirectional replication from T4 origins. The dual functionalities of Gp59 allow it to promote the initiation or re-start of DNA replication from a wide variety of recombination and replication intermediates.

  2. Enhanced Replication of Hepatitis E Virus Strain 47832c in an A549-Derived Subclonal Cell Line

    Science.gov (United States)

    Schemmerer, Mathias; Apelt, Silke; Trojnar, Eva; Ulrich, Rainer G.; Wenzel, Jürgen J.; Johne, Reimar

    2016-01-01

    Hepatitis E virus (HEV) is a human pathogen with increasing importance. The lack of efficient cell culture systems hampers systematic studies on its replication cycle, virus neutralization and inactivation. Here, several cell lines were inoculated with the HEV genotype 3c strain 47832c, previously isolated from a chronically infected transplant patient. At 14 days after inoculation the highest HEV genome copy numbers were found in A549 cells, followed by PLC/PRF/5 cells, whereas HepG2/C3A, Huh-7 Lunet BLR and MRC-5 cells only weakly supported virus replication. Inoculation of A549-derived subclone cell lines resulted in most cases in reduced HEV replication. However, the subclone A549/D3 was susceptible to lower virus concentrations and resulted in higher virus yields as compared to parental A549 cells. Transcriptome analysis indicated a downregulation of genes for carcinoembryonic antigen-related cell adhesion molecules (CEACAM) 5 and 6, and an upregulation of the syndecan 2 (SDC2) gene in A549/D3 cells compared to A549 cells. However, treatment of A549/D3 cells or A549 cells with CEACAM- or syndecan 2-specific antisera did not influence HEV replication. The results show that cells supporting more efficient HEV replication can be selected from the A549 cell line. The specific mechanisms responsible for the enhanced replication remain unknown. PMID:27690085

  3. Binding Affinities among DNA Helicase-Primase, DNA Polymerase, and Replication Intermediates in the Replisome of Bacteriophage T7.

    Science.gov (United States)

    Zhang, Huidong; Tang, Yong; Lee, Seung-Joo; Wei, Zeliang; Cao, Jia; Richardson, Charles C

    2016-01-15

    The formation of a replication loop on the lagging strand facilitates coordinated synthesis of the leading- and lagging-DNA strands and provides a mechanism for recycling of the lagging-strand DNA polymerase. As an Okazaki fragment is completed, the loop is released, and a new loop is formed as the synthesis of a new Okazaki fragment is initiated. Loop release requires the dissociation of the complex formed by the interactions among helicase, DNA polymerase, and DNA. The completion of the Okazaki fragment may result in either a nick or a single-stranded DNA region. In the replication system of bacteriophage T7, the dissociation of the polymerase from either DNA region is faster than that observed for the dissociation of the helicase from DNA polymerase, implying that the replication loop is released more likely through the dissociation of the lagging-strand DNA from polymerase, retaining the polymerase at replication fork. Both dissociation of DNA polymerase from DNA and that of helicase from a DNA polymerase · DNA complex are much faster at a nick DNA region than the release from a ssDNA region. These results suggest that the replication loop is released as a result of the nick formed when the lagging-strand DNA polymerase encounters the previously synthesized Okazaki fragment, releasing lagging-strand DNA and retaining DNA polymerase at the replication fork for the synthesis of next Okazaki fragment.

  4. Synchronization of DNA array replication kinetics

    Science.gov (United States)

    Manturov, Alexey O.; Grigoryev, Anton V.

    2016-04-01

    In the present work we discuss the features of the DNA replication kinetics at the case of multiplicity of simultaneously elongated DNA fragments. The interaction between replicated DNA fragments is carried out by free protons that appears at the every nucleotide attachment at the free end of elongated DNA fragment. So there is feedback between free protons concentration and DNA-polymerase activity that appears as elongation rate dependence. We develop the numerical model based on a cellular automaton, which can simulate the elongation stage (growth of DNA strands) for DNA elongation process with conditions pointed above and we study the possibility of the DNA polymerases movement synchronization. The results obtained numerically can be useful for DNA polymerase movement detection and visualization of the elongation process in the case of massive DNA replication, eg, under PCR condition or for DNA "sequencing by synthesis" sequencing devices evaluation.

  5. GFLV replication in electroporated grapevine protoplasts.

    Science.gov (United States)

    Valat; Toutain; Courtois; Gaire; Decout; Pinck; Mauro; Burrus

    2000-06-29

    Grapevine fanleaf virus (GFLV), responsible for the economically important court-noué disease, is exclusively transmitted to its natural host in the vineyards through Xiphinema nematodes. We have developed direct inoculation of GFLV into grapevine through protoplast electroporation. Protoplasts were isolated from mesophyll of in vitro-grown plants and from embryogenic cell suspensions. Permeation conditions were determined by monitoring calcein uptake. Low salt poration medium was selected. Electrical conditions leading to strong transient gene expression were also tested for GFLV inoculation (isolate F13). GFLV replication was detected with either virus particles (2 µg) or viral RNA (10 ng) in both protoplast populations, as shown by anti-P38 Western blotting. Direct inoculation and replication were also observed with Arabis mosaic virus (ArMV), a closely related nepovirus, as well as with another GFLV isolate. These results will be valuable in grapevine biotechnology, for GFLV replication studies, transgenic plant screening for GFLV resistance, and biorisk evaluation.

  6. Optical tweezers reveal how proteins alter replication

    Science.gov (United States)

    Chaurasiya, Kathy

    acids. We use single molecule DNA stretching to show that the nucleocapsid protein (NC) of the yeast retrotransposon Ty3, which is likely to be an ancestor of HIV NC, has optimal nucleic acid chaperone activity with only a single zinc finger. We also show that the chaperone activity of the ORF1 protein is responsible for successful replication of the mouse LINE-1 retrotransposon. LINE-1 is also 17% of the human genome, where it generates insertion mutations and alters gene expression. Retrotransposons such as LINE-1 and Ty3 are likely to be ancestors of retroviruses such as HIV. Human APOBEC3G (A3G) inhibits HIV-1 replication via cytidine deamination of the viral ssDNA genome, as well as via a distinct deamination-independent mechanism. Efficient deamination requires rapid on-off binding kinetics, but a slow dissociation rate is required for the proposed deaminase-independent mechanism. We resolve this apparent contradiction with a new quantitative single molecule method, which shows that A3G initially binds ssDNA with fast on-off rates and subsequently converts to a slow binding mode. This suggests that oligomerization transforms A3G from a fast enzyme to a slow binding protein, which is the biophysical mechanism that allows A3G to inhibit HIV replication. A complete understanding of the mechanism of A3G-mediated antiviral activity is required to design drugs that disrupt the viral response to A3G, enhance A3G packaging inside the viral core, and other potential strategies for long-term treatment of HIV infection. We use single molecule biophysics to explore the function of proteins involved in bacterial DNA replication, endogenous retrotransposition of retroelements in eukaryotic hosts such yeast and mice, and HIV replication in human cells. Our quantitative results provide insight into protein function in a range of complex biological systems and have wide-ranging implications for human health.

  7. Epigenetic control of DNA replication dynamics in mammals

    OpenAIRE

    Casas Delucchi, Corella Susana

    2011-01-01

    One of the most critically important processes in any living organism, essential for development and reproduction, is that of the accurate replication of its genome before each cell division. The process of DNA replication can take place millions of times in a single organism and any mistake, if left unrepaired, is potentially transmitted into the next generation. Errors during replication can result in genetic mutations or karyotype aberrations, both of which can lead to disease or death. ...

  8. LHCb experience with LFC replication

    CERN Document Server

    Bonifazi, F; Perez, E D; D'Apice, A; dell'Agnello, L; Düllmann, D; Girone, M; Re, G L; Martelli, B; Peco, G; Ricci, P P; Sapunenko, V; Vagnoni, V; Vitlacil, D

    2008-01-01

    Database replication is a key topic in the framework of the LHC Computing Grid to allow processing of data in a distributed environment. In particular, the LHCb computing model relies on the LHC File Catalog, i.e. a database which stores information about files spread across the GRID, their logical names and the physical locations of all the replicas. The LHCb computing model requires the LFC to be replicated at Tier-1s. The LCG 3D project deals with the database replication issue and provides a replication service based on Oracle Streams technology. This paper describes the deployment of the LHC File Catalog replication to the INFN National Center for Telematics and Informatics (CNAF) and to other LHCb Tier-1 sites. We performed stress tests designed to evaluate any delay in the propagation of the streams and the scalability of the system. The tests show the robustness of the replica implementation with performance going much beyond the LHCb requirements.

  9. LHCb experience with LFC replication

    CERN Document Server

    Carbone, Angelo; Dafonte Perez, Eva; D'Apice, Antimo; dell'Agnello, Luca; Duellmann, Dirk; Girone, Maria; Lo Re, Giuseppe; Martelli, Barbara; Peco, Gianluca; Ricci, Pier Paolo; Sapunenko, Vladimir; Vagnoni, Vincenzo; Vitlacil, Dejan

    2007-01-01

    Database replication is a key topic in the framework of the LHC Computing Grid to allow processing of data in a distributed environment. In particular, the LHCb computing model relies on the LHC File Catalog, i.e. database which stores information about files spread across the GRID, their logical names and the physical locations of all the replicas. The LHCb computing model requires the LFC to be replicated at Tier-1s. The LCG 3D project deals with the database replication issue and provides a replication service based on Oracle Streams technology. This paper describes the deployment of the LHC File Catalog replication to the INFN National Center for Telematics and Informations (CNAF) and to other LHCb Tier-1 sites. We performed stress tests designed to evaluate any delay in the propagation of the streams and the scalability of the system. The tests show the robustness of the replica implementation with performance going much beyond the LHCb requirements.

  10. Autophagy facilitates Salmonella replication in HeLa cells.

    Science.gov (United States)

    Yu, Hong B; Croxen, Matthew A; Marchiando, Amanda M; Ferreira, Rosana B R; Cadwell, Ken; Foster, Leonard J; Finlay, B Brett

    2014-03-11

    Autophagy is a process whereby a double-membrane structure (autophagosome) engulfs unnecessary cytosolic proteins, organelles, and invading pathogens and delivers them to the lysosome for degradation. We examined the fate of cytosolic Salmonella targeted by autophagy and found that autophagy-targeted Salmonella present in the cytosol of HeLa cells correlates with intracellular bacterial replication. Real-time analyses revealed that a subset of cytosolic Salmonella extensively associates with autophagy components p62 and/or LC3 and replicates quickly, whereas intravacuolar Salmonella shows no or very limited association with p62 or LC3 and replicates much more slowly. Replication of cytosolic Salmonella in HeLa cells is significantly decreased when autophagy components are depleted. Eventually, hyperreplication of cytosolic Salmonella potentiates cell detachment, facilitating the dissemination of Salmonella to neighboring cells. We propose that Salmonella benefits from autophagy for its cytosolic replication in HeLa cells. IMPORTANCE As a host defense system, autophagy is known to target a population of Salmonella for degradation and hence restricting Salmonella replication. In contrast to this concept, a recent report showed that knockdown of Rab1, a GTPase required for autophagy of Salmonella, decreases Salmonella replication in HeLa cells. Here, we have reexamined the fate of Salmonella targeted by autophagy by various cell biology-based assays. We found that the association of autophagy components with cytosolic Salmonella increases shortly after initiation of intracellular bacterial replication. Furthermore, through a live-cell imaging method, a subset of cytosolic Salmonella was found to be extensively associated with autophagy components p62 and/or LC3, and they replicated quickly. Most importantly, depletion of autophagy components significantly reduced the replication of cytosolic Salmonella in HeLa cells. Hence, in contrast to previous reports, we propose

  11. Therapeutic targeting of replicative immortality.

    Science.gov (United States)

    Yaswen, Paul; MacKenzie, Karen L; Keith, W Nicol; Hentosh, Patricia; Rodier, Francis; Zhu, Jiyue; Firestone, Gary L; Matheu, Ander; Carnero, Amancio; Bilsland, Alan; Sundin, Tabetha; Honoki, Kanya; Fujii, Hiromasa; Georgakilas, Alexandros G; Amedei, Amedeo; Amin, Amr; Helferich, Bill; Boosani, Chandra S; Guha, Gunjan; Ciriolo, Maria Rosa; Chen, Sophie; Mohammed, Sulma I; Azmi, Asfar S; Bhakta, Dipita; Halicka, Dorota; Niccolai, Elena; Aquilano, Katia; Ashraf, S Salman; Nowsheen, Somaira; Yang, Xujuan

    2015-12-01

    One of the hallmarks of malignant cell populations is the ability to undergo continuous proliferation. This property allows clonal lineages to acquire sequential aberrations that can fuel increasingly autonomous growth, invasiveness, and therapeutic resistance. Innate cellular mechanisms have evolved to regulate replicative potential as a hedge against malignant progression. When activated in the absence of normal terminal differentiation cues, these mechanisms can result in a state of persistent cytostasis. This state, termed "senescence," can be triggered by intrinsic cellular processes such as telomere dysfunction and oncogene expression, and by exogenous factors such as DNA damaging agents or oxidative environments. Despite differences in upstream signaling, senescence often involves convergent interdependent activation of tumor suppressors p53 and p16/pRB, but can be induced, albeit with reduced sensitivity, when these suppressors are compromised. Doses of conventional genotoxic drugs required to achieve cancer cell senescence are often much lower than doses required to achieve outright cell death. Additional therapies, such as those targeting cyclin dependent kinases or components of the PI3K signaling pathway, may induce senescence specifically in cancer cells by circumventing defects in tumor suppressor pathways or exploiting cancer cells' heightened requirements for telomerase. Such treatments sufficient to induce cancer cell senescence could provide increased patient survival with fewer and less severe side effects than conventional cytotoxic regimens. This positive aspect is countered by important caveats regarding senescence reversibility, genomic instability, and paracrine effects that may increase heterogeneity and adaptive resistance of surviving cancer cells. Nevertheless, agents that effectively disrupt replicative immortality will likely be valuable components of new combinatorial approaches to cancer therapy. Copyright © 2015 The Authors

  12. Pyrimidine Pool Disequilibrium Induced by a Cytidine Deaminase Deficiency Inhibits PARP-1 Activity, Leading to the Under Replication of DNA.

    Directory of Open Access Journals (Sweden)

    Simon Gemble

    2015-07-01

    Full Text Available Genome stability is jeopardized by imbalances of the dNTP pool; such imbalances affect the rate of fork progression. For example, cytidine deaminase (CDA deficiency leads to an excess of dCTP, slowing the replication fork. We describe here a novel mechanism by which pyrimidine pool disequilibrium compromises the completion of replication and chromosome segregation: the intracellular accumulation of dCTP inhibits PARP-1 activity. CDA deficiency results in incomplete DNA replication when cells enter mitosis, leading to the formation of ultrafine anaphase bridges between sister-chromatids at "difficult-to-replicate" sites such as centromeres and fragile sites. Using molecular combing, electron microscopy and a sensitive assay involving cell imaging to quantify steady-state PAR levels, we found that DNA replication was unsuccessful due to the partial inhibition of basal PARP-1 activity, rather than slower fork speed. The stimulation of PARP-1 activity in CDA-deficient cells restores replication and, thus, chromosome segregation. Moreover, increasing intracellular dCTP levels generates under-replication-induced sister-chromatid bridges as efficiently as PARP-1 knockdown. These results have direct implications for Bloom syndrome (BS, a rare genetic disease combining susceptibility to cancer and genomic instability. BS results from mutation of the BLM gene, encoding BLM, a RecQ 3'-5' DNA helicase, a deficiency of which leads to CDA downregulation. BS cells thus have a CDA defect, resulting in a high frequency of ultrafine anaphase bridges due entirely to dCTP-dependent PARP-1 inhibition and independent of BLM status. Our study describes previously unknown pathological consequences of the distortion of dNTP pools and reveals an unexpected role for PARP-1 in preventing DNA under-replication and chromosome segregation defects.

  13. Pyrimidine Pool Disequilibrium Induced by a Cytidine Deaminase Deficiency Inhibits PARP-1 Activity, Leading to the Under Replication of DNA.

    Directory of Open Access Journals (Sweden)

    Simon Gemble

    2015-07-01

    Full Text Available Genome stability is jeopardized by imbalances of the dNTP pool; such imbalances affect the rate of fork progression. For example, cytidine deaminase (CDA deficiency leads to an excess of dCTP, slowing the replication fork. We describe here a novel mechanism by which pyrimidine pool disequilibrium compromises the completion of replication and chromosome segregation: the intracellular accumulation of dCTP inhibits PARP-1 activity. CDA deficiency results in incomplete DNA replication when cells enter mitosis, leading to the formation of ultrafine anaphase bridges between sister-chromatids at "difficult-to-replicate" sites such as centromeres and fragile sites. Using molecular combing, electron microscopy and a sensitive assay involving cell imaging to quantify steady-state PAR levels, we found that DNA replication was unsuccessful due to the partial inhibition of basal PARP-1 activity, rather than slower fork speed. The stimulation of PARP-1 activity in CDA-deficient cells restores replication and, thus, chromosome segregation. Moreover, increasing intracellular dCTP levels generates under-replication-induced sister-chromatid bridges as efficiently as PARP-1 knockdown. These results have direct implications for Bloom syndrome (BS, a rare genetic disease combining susceptibility to cancer and genomic instability. BS results from mutation of the BLM gene, encoding BLM, a RecQ 3'-5' DNA helicase, a deficiency of which leads to CDA downregulation. BS cells thus have a CDA defect, resulting in a high frequency of ultrafine anaphase bridges due entirely to dCTP-dependent PARP-1 inhibition and independent of BLM status. Our study describes previously unknown pathological consequences of the distortion of dNTP pools and reveals an unexpected role for PARP-1 in preventing DNA under-replication and chromosome segregation defects.

  14. Replication of Avocado Sunblotch Viroid in the Yeast Saccharomyces cerevisiae▿

    Science.gov (United States)

    Delan-Forino, Clémentine; Maurel, Marie-Christine; Torchet, Claire

    2011-01-01

    Viroids are the smallest known pathogenic agents. They are noncoding, single-stranded, closed-circular, “naked” RNAs, which replicate through RNA-RNA transcription. Viroids of the Avsunviroidae family possess a hammerhead ribozyme in their sequence, allowing self-cleavage during their replication. To date, viroids have only been detected in plant cells. Here, we investigate the replication of Avocado sunblotch viroid (ASBVd) of the Avsunviroidae family in a nonconventional host, the yeast Saccharomyces cerevisiae. We demonstrate that ASBVd RNA strands of both polarities are able to self-cleave and to replicate in a unicellular eukaryote cell. We show that the viroid monomeric RNA is destabilized by the nuclear 3′ and the cytoplasmic 5′ RNA degradation pathways. For the first time, our results provide evidence that viroids can replicate in other organisms than plants and that yeast contains all of the essential cellular elements for the replication of ASBVd. PMID:21270165

  15. NACSA Charter School Replication Guide: The Spectrum of Replication Options. Authorizing Matters. Replication Brief 1

    Science.gov (United States)

    O'Neill, Paul

    2010-01-01

    One of the most important and high-profile issues in public education reform today is the replication of successful public charter school programs. With more than 5,000 failing public schools in the United States, there is a tremendous need for strong alternatives for parents and students. Replicating successful charter school models is an…

  16. Evidence for sequential and increasing activation of replication origins along replication timing gradients in the human genome.

    Science.gov (United States)

    Guilbaud, Guillaume; Rappailles, Aurélien; Baker, Antoine; Chen, Chun-Long; Arneodo, Alain; Goldar, Arach; d'Aubenton-Carafa, Yves; Thermes, Claude; Audit, Benjamin; Hyrien, Olivier

    2011-12-01

    Genome-wide replication timing studies have suggested that mammalian chromosomes consist of megabase-scale domains of coordinated origin firing separated by large originless transition regions. Here, we report a quantitative genome-wide analysis of DNA replication kinetics in several human cell types that contradicts this view. DNA combing in HeLa cells sorted into four temporal compartments of S phase shows that replication origins are spaced at 40 kb intervals and fire as small clusters whose synchrony increases during S phase and that replication fork velocity (mean 0.7 kb/min, maximum 2.0 kb/min) remains constant and narrowly distributed through S phase. However, multi-scale analysis of a genome-wide replication timing profile shows a broad distribution of replication timing gradients with practically no regions larger than 100 kb replicating at less than 2 kb/min. Therefore, HeLa cells lack large regions of unidirectional fork progression. Temporal transition regions are replicated by sequential activation of origins at a rate that increases during S phase and replication timing gradients are set by the delay and the spacing between successive origin firings rather than by the velocity of single forks. Activation of internal origins in a specific temporal transition region is directly demonstrated by DNA combing of the IGH locus in HeLa cells. Analysis of published origin maps in HeLa cells and published replication timing and DNA combing data in several other cell types corroborate these findings, with the interesting exception of embryonic stem cells where regions of unidirectional fork progression seem more abundant. These results can be explained if origins fire independently of each other but under the control of long-range chromatin structure, or if replication forks progressing from early origins stimulate initiation in nearby unreplicated DNA. These findings shed a new light on the replication timing program of mammalian genomes and provide a general

  17. Rif1 Regulates Initiation Timing of Late Replication Origins throughout the S. cerevisiae Genome

    OpenAIRE

    Peace, Jared M.; Anna Ter-Zakarian; Aparicio, Oscar M

    2014-01-01

    Chromosomal DNA replication involves the coordinated activity of hundreds to thousands of replication origins. Individual replication origins are subject to epigenetic regulation of their activity during S-phase, resulting in differential efficiencies and timings of replication initiation during S-phase. This regulation is thought to involve chromatin structure and organization into timing domains with differential ability to recruit limiting replication factors. Rif1 has recently been identi...

  18. Measurement of replication structures at the nanometer scale using super-resolution light microscopy.

    Science.gov (United States)

    Baddeley, D; Chagin, V O; Schermelleh, L; Martin, S; Pombo, A; Carlton, P M; Gahl, A; Domaing, P; Birk, U; Leonhardt, H; Cremer, C; Cardoso, M C

    2010-01-01

    DNA replication, similar to other cellular processes, occurs within dynamic macromolecular structures. Any comprehensive understanding ultimately requires quantitative data to establish and test models of genome duplication. We used two different super-resolution light microscopy techniques to directly measure and compare the size and numbers of replication foci in mammalian cells. This analysis showed that replication foci vary in size from 210 nm down to 40 nm. Remarkably, spatially modulated illumination (SMI) and 3D-structured illumination microscopy (3D-SIM) both showed an average size of 125 nm that was conserved throughout S-phase and independent of the labeling method, suggesting a basic unit of genome duplication. Interestingly, the improved optical 3D resolution identified 3- to 5-fold more distinct replication foci than previously reported. These results show that optical nanoscopy techniques enable accurate measurements of cellular structures at a level previously achieved only by electron microscopy and highlight the possibility of high-throughput, multispectral 3D analyses.

  19. Aquareovirus NS80 Initiates Efficient Viral Replication by Retaining Core Proteins within Replication-Associated Viral Inclusion Bodies

    OpenAIRE

    Liming Yan; Jie Zhang; Hong Guo; Shicui Yan; Qingxiu Chen; Fuxian Zhang; Qin Fang

    2015-01-01

    Viral inclusion bodies (VIBs) are specific intracellular compartments for reoviruses replication and assembly. Aquareovirus nonstructural protein NS80 has been identified to be the major constituent for forming globular VIBs in our previous study. In this study, we investigated the role of NS80 in viral structural proteins expression and viral replication. Immunofluorescence assays showed that NS80 could retain five core proteins or inner-capsid proteins (VP1-VP4 and VP6), but not outer-capsi...

  20. International Expansion through Flexible Replication

    DEFF Research Database (Denmark)

    Jonsson, Anna; Foss, Nicolai Juul

    2011-01-01

    to local environments and under the impact of new learning. To illuminate these issues, we draw on a longitudinal in-depth study of Swedish home furnishing giant IKEA, involving more than 70 interviews. We find that IKEA has developed organizational mechanisms that support an ongoing learning process aimed......, etc.) are replicated in a uniform manner across stores, and change only very slowly (if at all) in response to learning (“flexible replication”). We conclude by discussing the factors that influence the approach to replication adopted by an international replicator....

  1. Regulation of Replication Recovery and Genome Integrity

    DEFF Research Database (Denmark)

    Colding, Camilla Skettrup

    Preserving genome integrity is essential for cell survival. To this end, mechanisms that supervise DNA replication and respond to replication perturbations have evolved. One such mechanism is the replication checkpoint, which responds to DNA replication stress and acts to ensure replication pausing...

  2. Requirement of cellular DDX3 for hepatitis C virus replication is unrelated to its interaction with the viral core protein.

    Science.gov (United States)

    Angus, Allan G N; Dalrymple, David; Boulant, Steeve; McGivern, David R; Clayton, Reginald F; Scott, Martin J; Adair, Richard; Graham, Susan; Owsianka, Ania M; Targett-Adams, Paul; Li, Kui; Wakita, Takaji; McLauchlan, John; Lemon, Stanley M; Patel, Arvind H

    2010-01-01

    The cellular DEAD-box protein DDX3 was recently shown to be essential for hepatitis C virus (HCV) replication. Prior to that, we had reported that HCV core binds to DDX3 in yeast-two hybrid and transient transfection assays. Here, we confirm by co-immunoprecipitation that this interaction occurs in cells replicating the JFH1 virus. Consistent with this result, immunofluorescence staining of infected cells revealed a dramatic redistribution of cytoplasmic DDX3 by core protein to the virus assembly sites around lipid droplets. Given this close association of DDX3 with core and lipid droplets, and its involvement in virus replication, we investigated the importance of this host factor in the virus life cycle. Mutagenesis studies located a single amino acid in the N-terminal domain of JFH1 core that when changed to alanine significantly abrogated this interaction. Surprisingly, this mutation did not alter infectious virus production and RNA replication, indicating that the core-DDX3 interaction is dispensable in the HCV life cycle. Consistent with previous studies, siRNA-led knockdown of DDX3 lowered virus production and RNA replication levels of both WT JFH1 and the mutant virus unable to bind DDX3. Thus, our study shows for the first time that the requirement of DDX3 for HCV replication is unrelated to its interaction with the viral core protein.

  3. Nucleotide Metabolism and DNA Replication.

    Science.gov (United States)

    Warner, Digby F; Evans, Joanna C; Mizrahi, Valerie

    2014-10-01

    The development and application of a highly versatile suite of tools for mycobacterial genetics, coupled with widespread use of "omics" approaches to elucidate the structure, function, and regulation of mycobacterial proteins, has led to spectacular advances in our understanding of the metabolism and physiology of mycobacteria. In this article, we provide an update on nucleotide metabolism and DNA replication in mycobacteria, highlighting key findings from the past 10 to 15 years. In the first section, we focus on nucleotide metabolism, ranging from the biosynthesis, salvage, and interconversion of purine and pyrimidine ribonucleotides to the formation of deoxyribonucleotides. The second part of the article is devoted to DNA replication, with a focus on replication initiation and elongation, as well as DNA unwinding. We provide an overview of replication fidelity and mutation rates in mycobacteria and summarize evidence suggesting that DNA replication occurs during states of low metabolic activity, and conclude by suggesting directions for future research to address key outstanding questions. Although this article focuses primarily on observations from Mycobacterium tuberculosis, it is interspersed, where appropriate, with insights from, and comparisons with, other mycobacterial species as well as better characterized bacterial models such as Escherichia coli. Finally, a common theme underlying almost all studies of mycobacterial metabolism is the potential to identify and validate functions or pathways that can be exploited for tuberculosis drug discovery. In this context, we have specifically highlighted those processes in mycobacterial DNA replication that might satisfy this critical requirement.

  4. Plasmid Rolling-Circle Replication.

    Science.gov (United States)

    Ruiz-Masó, J A; MachóN, C; Bordanaba-Ruiseco, L; Espinosa, M; Coll, M; Del Solar, G

    2015-02-01

    Plasmids are DNA entities that undergo controlled replication independent of the chromosomal DNA, a crucial step that guarantees the prevalence of the plasmid in its host. DNA replication has to cope with the incapacity of the DNA polymerases to start de novo DNA synthesis, and different replication mechanisms offer diverse solutions to this problem. Rolling-circle replication (RCR) is a mechanism adopted by certain plasmids, among other genetic elements, that represents one of the simplest initiation strategies, that is, the nicking by a replication initiator protein on one parental strand to generate the primer for leading-strand initiation and a single priming site for lagging-strand synthesis. All RCR plasmid genomes consist of a number of basic elements: leading strand initiation and control, lagging strand origin, phenotypic determinants, and mobilization, generally in that order of frequency. RCR has been mainly characterized in Gram-positive bacterial plasmids, although it has also been described in Gram-negative bacterial or archaeal plasmids. Here we aim to provide an overview of the RCR plasmids' lifestyle, with emphasis on their characteristic traits, promiscuity, stability, utility as vectors, etc. While RCR is one of the best-characterized plasmid replication mechanisms, there are still many questions left unanswered, which will be pointed out along the way in this review.

  5. Multiseason occupancy models for correlated replicate surveys

    Science.gov (United States)

    Hines, James; Nichols, James; Collazo, Jaime

    2014-01-01

    Occupancy surveys collecting data from adjacent (sometimes correlated) spatial replicates have become relatively popular for logistical reasons. Hines et al. (2010) presented one approach to modelling such data for single-season occupancy surveys. Here, we present a multiseason analogue of this model (with corresponding software) for inferences about occupancy dynamics. We include a new parameter to deal with the uncertainty associated with the first spatial replicate for both single-season and multiseason models. We use a case study, based on the brown-headed nuthatch, to assess the need for these models when analysing data from the North American Breeding Bird Survey (BBS), and we test various hypotheses about occupancy dynamics for this species in the south-eastern United States. The new model permits inference about local probabilities of extinction, colonization and occupancy for sampling conducted over multiple seasons. The model performs adequately, based on a small simulation study and on results of the case study analysis. The new model incorporating correlated replicates was strongly favoured by model selection for the BBS data for brown-headed nuthatch (Sitta pusilla). Latitude was found to be an important source of variation in local colonization and occupancy probabilities for brown-headed nuthatch, with both probabilities being higher near the centre of the species range, as opposed to more northern and southern areas. We recommend this new occupancy model for detection–nondetection studies that use potentially correlated replicates.

  6. Resurgence of derived stimulus relations: replication and extensions.

    Science.gov (United States)

    Doughty, Adam H; Kastner, Rebecca M; Bismark, Bryan D

    2011-01-01

    Resurgence typically refers to the recovery of a previously reinforced response when a more recently reinforced response is extinguished. Under conditions of punishment, Wilson and Hayes (1996) observed the recovery of derived stimulus relations that never had been correlated with differential consequences. The present study systematically replicated the findings of Wilson and Hayes by observing the recovery of derived stimulus relations under extinction conditions and with an additional equivalence class. College students received arbitrary-matching-to-sample training in Phase 1 that resulted in four 4-member stimulus-equivalence classes. These derived relations were not correlated with differential consequences. In Phase 2, with the same stimuli, the students received training that resulted in four different 4-member stimulus-equivalence classes. After the emergence and reinforcement of the latter derived relations, their extinction generated the recurrence of the derived relations from Phase 1. The implications of these findings for conceptualizing resurgence are discussed.

  7. 5-Benzothiazole substituted pyrimidine derivatives as HCV replication (replicase) inhibitors.

    Science.gov (United States)

    Arasappan, Ashok; Bennett, Frank; Girijavallabhan, Vinay; Huang, Yuhua; Huelgas, Regina; Alvarez, Carmen; Chen, Lei; Gavalas, Stephen; Kim, Seong-Heon; Kosinski, Aneta; Pinto, Patrick; Rizvi, Razia; Rossman, Randall; Shankar, Bandarpalle; Tong, Ling; Velazquez, Francisco; Venkatraman, Srikanth; Verma, Vishal A; Kozlowski, Joseph; Shih, Neng-Yang; Piwinski, John J; MacCoss, Malcolm; Kwong, Cecil D; Clark, Jeremy L; Fowler, Anita T; Geng, Feng; Kezar, Hollis S; Roychowdhury, Abhijit; Reynolds, Robert C; Maddry, Joseph A; Ananthan, Subramaniam; Secrist, John A; Li, Cheng; Chase, Robert; Curry, Stephanie; Huang, Hsueh-Cheng; Tong, Xiao; Njoroge, F George

    2012-05-01

    Based on a previously identified HCV replication (replicase) inhibitor 1, SAR efforts were conducted around the pyrimidine core to improve the potency and pharmacokinetic profile of the inhibitors. A benzothiazole moiety was found to be the optimal substituent at the pyrimidine 5-position. Due to potential reactivity concern, the 4-chloro residue was replaced by a methyl group with some loss in potency and enhanced rat in vivo profile. Extensive investigations at the C-2 position resulted in identification of compound 16 that demonstrated very good replicon potency, selectivity and rodent plasma/target organ concentration. Inhibitor 16 also demonstrated good plasma levels and oral bioavailability in dogs, while monkey exposure was rather low. Chemistry optimization towards a practical route to install the benzothiazole moiety resulted in an efficient direct C-H arylation protocol.

  8. Replicability and heterogeneity of awake unrestrained canine FMRI responses.

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    Gregory S Berns

    Full Text Available Previously, we demonstrated the possibility of fMRI in two awake and unrestrained dogs. Here, we determined the replicability and heterogeneity of these results in an additional 11 dogs for a total of 13 subjects. Based on an anatomically placed region-of-interest, we compared the caudate response to a hand signal indicating the imminent availability of a food reward to a hand signal indicating no reward. 8 of 13 dogs had a positive differential caudate response to the signal indicating reward. The mean differential caudate response was 0.09%, which was similar to a comparable human study. These results show that canine fMRI is reliable and can be done with minimal stress to the dogs.

  9. Replicability and heterogeneity of awake unrestrained canine FMRI responses.

    Science.gov (United States)

    Berns, Gregory S; Brooks, Andrew; Spivak, Mark

    2013-01-01

    Previously, we demonstrated the possibility of fMRI in two awake and unrestrained dogs. Here, we determined the replicability and heterogeneity of these results in an additional 11 dogs for a total of 13 subjects. Based on an anatomically placed region-of-interest, we compared the caudate response to a hand signal indicating the imminent availability of a food reward to a hand signal indicating no reward. 8 of 13 dogs had a positive differential caudate response to the signal indicating reward. The mean differential caudate response was 0.09%, which was similar to a comparable human study. These results show that canine fMRI is reliable and can be done with minimal stress to the dogs.

  10. Termination of DNA replication forks: "Breaking up is hard to do".

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    Bailey, Rachael; Priego Moreno, Sara; Gambus, Agnieszka

    2015-01-01

    To ensure duplication of the entire genome, eukaryotic DNA replication initiates from thousands of replication origins. The replication forks move through the chromatin until they encounter forks from neighboring origins. During replication fork termination forks converge, the replisomes disassemble and topoisomerase II resolves the daughter DNA molecules. If not resolved efficiently, terminating forks result in genomic instability through the formation of pathogenic structures. Our recent findings shed light onto the mechanism of replisome disassembly upon replication fork termination. We have shown that termination-specific polyubiquitylation of the replicative helicase component - Mcm7, leads to dissolution of the active helicase in a process dependent on the p97/VCP/Cdc48 segregase. The inhibition of terminating helicase disassembly resulted in a replication termination defect. In this extended view we present hypothetical models of replication fork termination and discuss remaining and emerging questions in the DNA replication termination field.

  11. Pharmacological manipulation of the akt signaling pathway regulates myxoma virus replication and tropism in human cancer cells.

    Science.gov (United States)

    Werden, Steven J; McFadden, Grant

    2010-04-01

    Viruses have evolved an assortment of mechanisms for regulating the Akt signaling pathway to establish a cellular environment more favorable for viral replication. Myxoma virus (MYXV) is a rabbit-specific poxvirus that encodes many immunomodulatory factors, including an ankyrin repeat-containing host range protein termed M-T5 that functions to regulate tropism of MYXV for rabbit lymphocytes and certain human cancer cells. MYXV permissiveness in these human cancer cells is dependent upon the direct interaction between M-T5 and Akt, which has been shown to induce the kinase activity of Akt. In this study, an array of compounds that selectively manipulate Akt signaling was screened and we show that only a subset of Akt inhibitors significantly decreased the ability of MYXV to replicate in previously permissive human cancer cells. Furthermore, reduced viral replication efficiency was correlated with lower levels of phosphorylated Akt. In contrast, the PP2A-specific phosphatase inhibitor okadaic acid promoted increased Akt kinase activation and rescued MYXV replication in human cancer cells that did not previously support viral replication. Finally, phosphorylation of Akt at residue Thr308 was shown to dictate the physical interaction between Akt and M-T5, which then leads to phosphorylation of Ser473 and permits productive MYXV replication in these human cancer cells. The results of this study further characterize the mechanism by which M-T5 exploits the Akt signaling cascade and affirms this interaction as a major tropism determinant that regulates the replication efficiency of MYXV in human cancer cells.

  12. Antisense oligonucleotide inhibition of hepatitis C virus genotype 4 replication in HepG2 cells

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    Omran Moataza H

    2006-06-01

    Full Text Available Abstract Background Hepatitis C (HCV viral infection is a serious medical problem in Egypt and it has a devastating impact on the Egyptian economy. It is estimated that over 15% of Egyptians are infected by the virus and thus finding a cure for this disease is of utmost importance. Current therapies for hepatitis C virus (HCV genotype 4 with interferon/ribavirin have not been successful and thus the development of alternative therapy for this genotype is disparately needed. Results Although previous studies utilizing viral subgenomic or full cDNA fragments linked to reporter genes transfected into adhered cells or in a cell free system showed promise, demonstration of efficient viral replication was lacking. Thus, we utilized HepG2 cells infected with native HCV RNA genomes in a replication competent system and used antisense phosphorothioate Oligonucleotides (S-ODN against stem loop IIId and the AUG translation start site of the viral polyprotein precursor to monitor viral replication. We were able to show complete arrest of intracellular replication of HCV-4 at 1 uM S-ODN, thus providing a proof of concept for the potential antiviral activity of S-ODN on native genomic replication of HCV genotype 4. Conclusion We have successfully demonstrated that by using two S-ODNs [(S-ODN1 (nt 326–348 and S-ODN-2 (nt 264–282], we were able to completely inhibit viral replication in culture, thus confirming earlier reports on subgenomic constructs and suggesting a potential therapeutic value in HCV type 4.

  13. Murine leukemia virus (MLV replication monitored with fluorescent proteins

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    Bittner Alexandra

    2004-12-01

    Full Text Available Abstract Background Cancer gene therapy will benefit from vectors that are able to replicate in tumor tissue and cause a bystander effect. Replication-competent murine leukemia virus (MLV has been described to have potential as cancer therapeutics, however, MLV infection does not cause a cytopathic effect in the infected cell and viral replication can only be studied by immunostaining or measurement of reverse transcriptase activity. Results We inserted the coding sequences for green fluorescent protein (GFP into the proline-rich region (PRR of the ecotropic envelope protein (Env and were able to fluorescently label MLV. This allowed us to directly monitor viral replication and attachment to target cells by flow cytometry. We used this method to study viral replication of recombinant MLVs and split viral genomes, which were generated by replacement of the MLV env gene with the red fluorescent protein (RFP and separately cloning GFP-Env into a retroviral vector. Co-transfection of both plasmids into target cells resulted in the generation of semi-replicative vectors, and the two color labeling allowed to determine the distribution of the individual genomes in the target cells and was indicative for the occurrence of recombination events. Conclusions Fluorescently labeled MLVs are excellent tools for the study of factors that influence viral replication and can be used to optimize MLV-based replication-competent viruses or vectors for gene therapy.

  14. Systematic determination of replication activity type highlights interconnections between replication, chromatin structure and nuclear localization.

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    Shlomit Farkash-Amar

    Full Text Available DNA replication is a highly regulated process, with each genomic locus replicating at a distinct time of replication (ToR. Advances in ToR measurement technology enabled several genome-wide profiling studies that revealed tight associations between ToR and general genomic features and a remarkable ToR conservation in mammals. Genome wide studies further showed that at the hundreds kb-to-megabase scale the genome can be divided into constant ToR regions (CTRs in which the replication process propagates at a faster pace due to the activation of multiple origins and temporal transition regions (TTRs in which the replication process propagates at a slower pace. We developed a computational tool that assigns a ToR to every measured locus and determines its replication activity type (CTR versus TTR. Our algorithm, ARTO (Analysis of Replication Timing and Organization, uses signal processing methods to fit a constant piece-wise linear curve to the measured raw data. We tested our algorithm and provide performance and usability results. A Matlab implementation of ARTO is available at http://bioinfo.cs.technion.ac.il/people/zohar/ARTO/. Applying our algorithm to ToR data measured in multiple mouse and human samples allowed precise genome-wide ToR determination and replication activity type characterization. Analysis of the results highlighted the plasticity of the replication program. For example, we observed significant ToR differences in 10-25% of the genome when comparing different tissue types. Our analyses also provide evidence for activity type differences in up to 30% of the probes. Integration of the ToR data with multiple aspects of chromosome organization characteristics suggests that ToR plays a role in shaping the regional chromatin structure. Namely, repressive chromatin marks, are associated with late ToR both in TTRs and CTRs. Finally, characterization of the differences between TTRs and CTRs, with matching ToR, revealed that TTRs are

  15. Failed Replication of Oxytocin Effects on Trust: The Envelope Task Case.

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    Anthony Lane

    Full Text Available The neurohormone Oxytocin (OT has been one of the most studied peptides in behavioral sciences over the past two decades. Many studies have suggested that OT could increase trusting behaviors. A previous study, based on the "Envelope Task" paradigm, where trust is assessed by the degree of openness of an envelope containing participant's confidential information, showed that OT increases trusting behavior and reported one of the most powerful effects of OT on a behavioral variable. In this paper we present two failed replications of this effect, despite sufficient power to replicate the original large effect. The non-significant results of these two failed replications clearly exclude a large effect of OT on trust in this paradigm but are compatible with either a null effect of OT on trust, or a small effect, undetectable with small sample size (N = 95 and 61 in Study 1 and 2, respectively. Taken together, our results question the purported size of OT's effect on trust and emphasize the need for replications.

  16. Ciprofloxacin is an inhibitor of the Mcm2-7 replicative helicase

    Science.gov (United States)

    Simon, Nicholas; Bochman, Matthew L.; Seguin, Sandlin; Brodsky, Jeffrey L.; Seibel, William L.; Schwacha, Anthony

    2013-01-01

    Most currently available small molecule inhibitors of DNA replication lack enzymatic specificity, resulting in deleterious side effects during use in cancer chemotherapy and limited experimental usefulness as mechanistic tools to study DNA replication. Towards development of targeted replication inhibitors, we have focused on Mcm2-7 (minichromosome maintenance protein 2–7), a highly conserved helicase and key regulatory component of eukaryotic DNA replication. Unexpectedly we found that the fluoroquinolone antibiotic ciprofloxacin preferentially inhibits Mcm2-7. Ciprofloxacin blocks the DNA helicase activity of Mcm2-7 at concentrations that have little effect on other tested helicases and prevents the proliferation of both yeast and human cells at concentrations similar to those that inhibit DNA unwinding. Moreover, a previously characterized mcm mutant (mcm4chaos3) exhibits increased ciprofloxacin resistance. To identify more potent Mcm2-7 inhibitors, we screened molecules that are structurally related to ciprofloxacin and identified several that compromise the Mcm2-7 helicase activity at lower concentrations. Our results indicate that ciprofloxacin targets Mcm2-7 in vitro, and support the feasibility of developing specific quinolone-based inhibitors of Mcm2-7 for therapeutic and experimental applications. PMID:24001138

  17. Biochemical analysis of DNA polymerase η fidelity in the presence of replication protein A.

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    Samuel C Suarez

    Full Text Available DNA polymerase η (pol η synthesizes across from damaged DNA templates in order to prevent deleterious consequences like replication fork collapse and double-strand breaks. This process, termed translesion synthesis (TLS, is an overall positive for the cell, as cells deficient in pol η display higher mutation rates. This outcome occurs despite the fact that the in vitro fidelity of bypass by pol η alone is moderate to low, depending on the lesion being copied. One possible means of increasing the fidelity of pol η is interaction with replication accessory proteins present at the replication fork. We have previously utilized a bacteriophage based screening system to measure the fidelity of bypass using purified proteins. Here we report on the fidelity effects of a single stranded binding protein, replication protein A (RPA, when copying the oxidative lesion 7,8-dihydro-8-oxo-guanine(8-oxoG and the UV-induced cis-syn thymine-thymine cyclobutane pyrimidine dimer (T-T CPD. We observed no change in fidelity dependent on RPA when copying these damaged templates. This result is consistent in multiple position contexts. We previously identified single amino acid substitution mutants of pol η that have specific effects on fidelity when copying both damaged and undamaged templates. In order to confirm our results, we examined the Q38A and Y52E mutants in the same full-length construct. We again observed no difference when RPA was added to the bypass reaction, with the mutant forms of pol η displaying similar fidelity regardless of RPA status. We do, however, observe some slight effects when copying undamaged DNA, similar to those we have described previously. Our results indicate that RPA by itself does not affect pol η dependent lesion bypass fidelity when copying either 8-oxoG or T-T CPD lesions.

  18. Replication timing: a fingerprint for cell identity and pluripotency.

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    Tyrone Ryba

    2011-10-01

    Full Text Available Many types of epigenetic profiling have been used to classify stem cells, stages of cellular differentiation, and cancer subtypes. Existing methods focus on local chromatin features such as DNA methylation and histone modifications that require extensive analysis for genome-wide coverage. Replication timing has emerged as a highly stable cell type-specific epigenetic feature that is regulated at the megabase-level and is easily and comprehensively analyzed genome-wide. Here, we describe a cell classification method using 67 individual replication profiles from 34 mouse and human cell lines and stem cell-derived tissues, including new data for mesendoderm, definitive endoderm, mesoderm and smooth muscle. Using a Monte-Carlo approach for selecting features of replication profiles conserved in each cell type, we identify "replication timing fingerprints" unique to each cell type and apply a k nearest neighbor approach to predict known and unknown cell types. Our method correctly classifies 67/67 independent replication-timing profiles, including those derived from closely related intermediate stages. We also apply this method to derive fingerprints for pluripotency in human and mouse cells. Interestingly, the mouse pluripotency fingerprint overlaps almost completely with previously identified genomic segments that switch from early to late replication as pluripotency is lost. Thereafter, replication timing and transcription within these regions become difficult to reprogram back to pluripotency, suggesting these regions highlight an epigenetic barrier to reprogramming. In addition, the major histone cluster Hist1 consistently becomes later replicating in committed cell types, and several histone H1 genes in this cluster are downregulated during differentiation, suggesting a possible instrument for the chromatin compaction observed during differentiation. Finally, we demonstrate that unknown samples can be classified independently using site

  19. Two replications of "Hierarchical encoding makes individuals in a group seem more attractive (2014; Experiment 4".

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    Yuko Ojiro

    2015-06-01

    Full Text Available The cheerleader effect implies that a person in a group look like more attractive than in isolation. Walker and Vul (2014 reported results supporting the existence of the cheerleader effect. We replicated Walker and Vul’s Experiment 4, which manipulated group size. Their participants were asked to rate attractiveness of each female face image in a group (one of 4, 9, or 16 members and in isolation and revealed that attractiveness ratings significantly increased in all the group conditions. We performed two direct replications of this experiment using Japanese participants. As a result, at least one experiment yielded a pattern of results similar to those of the previous study, although the effect was not significant and the effect size was small.

  20. Integrative functional genomics of hepatitis C virus infection identifies host dependencies in complete viral replication cycle.

    Science.gov (United States)

    Li, Qisheng; Zhang, Yong-Yuan; Chiu, Stephan; Hu, Zongyi; Lan, Keng-Hsin; Cha, Helen; Sodroski, Catherine; Zhang, Fang; Hsu, Ching-Sheng; Thomas, Emmanuel; Liang, T Jake

    2014-05-01

    Recent functional genomics studies including genome-wide small interfering RNA (siRNA) screens demonstrated that hepatitis C virus (HCV) exploits an extensive network of host factors for productive infection and propagation. How these co-opted host functions interact with various steps of HCV replication cycle and exert pro- or antiviral effects on HCV infection remains largely undefined. Here we present an unbiased and systematic strategy to functionally interrogate HCV host dependencies uncovered from our previous infectious HCV (HCVcc) siRNA screen. Applying functional genomics approaches and various in vitro HCV model systems, including HCV pseudoparticles (HCVpp), single-cycle infectious particles (HCVsc), subgenomic replicons, and HCV cell culture systems (HCVcc), we identified and characterized novel host factors or pathways required for each individual step of the HCV replication cycle. Particularly, we uncovered multiple HCV entry factors, including E-cadherin, choline kinase α, NADPH oxidase CYBA, Rho GTPase RAC1 and SMAD family member 6. We also demonstrated that guanine nucleotide binding protein GNB2L1, E2 ubiquitin-conjugating enzyme UBE2J1, and 39 other host factors are required for HCV RNA replication, while the deubiquitinating enzyme USP11 and multiple other cellular genes are specifically involved in HCV IRES-mediated translation. Families of antiviral factors that target HCV replication or translation were also identified. In addition, various virologic assays validated that 66 host factors are involved in HCV assembly or secretion. These genes included insulin-degrading enzyme (IDE), a proviral factor, and N-Myc down regulated Gene 1 (NDRG1), an antiviral factor. Bioinformatics meta-analyses of our results integrated with literature mining of previously published HCV host factors allows the construction of an extensive roadmap of cellular networks and pathways involved in the complete HCV replication cycle. This comprehensive study of HCV host

  1. Integrative functional genomics of hepatitis C virus infection identifies host dependencies in complete viral replication cycle.

    Directory of Open Access Journals (Sweden)

    Qisheng Li

    2014-05-01

    Full Text Available Recent functional genomics studies including genome-wide small interfering RNA (siRNA screens demonstrated that hepatitis C virus (HCV exploits an extensive network of host factors for productive infection and propagation. How these co-opted host functions interact with various steps of HCV replication cycle and exert pro- or antiviral effects on HCV infection remains largely undefined. Here we present an unbiased and systematic strategy to functionally interrogate HCV host dependencies uncovered from our previous infectious HCV (HCVcc siRNA screen. Applying functional genomics approaches and various in vitro HCV model systems, including HCV pseudoparticles (HCVpp, single-cycle infectious particles (HCVsc, subgenomic replicons, and HCV cell culture systems (HCVcc, we identified and characterized novel host factors or pathways required for each individual step of the HCV replication cycle. Particularly, we uncovered multiple HCV entry factors, including E-cadherin, choline kinase α, NADPH oxidase CYBA, Rho GTPase RAC1 and SMAD family member 6. We also demonstrated that guanine nucleotide binding protein GNB2L1, E2 ubiquitin-conjugating enzyme UBE2J1, and 39 other host factors are required for HCV RNA replication, while the deubiquitinating enzyme USP11 and multiple other cellular genes are specifically involved in HCV IRES-mediated translation. Families of antiviral factors that target HCV replication or translation were also identified. In addition, various virologic assays validated that 66 host factors are involved in HCV assembly or secretion. These genes included insulin-degrading enzyme (IDE, a proviral factor, and N-Myc down regulated Gene 1 (NDRG1, an antiviral factor. Bioinformatics meta-analyses of our results integrated with literature mining of previously published HCV host factors allows the construction of an extensive roadmap of cellular networks and pathways involved in the complete HCV replication cycle. This comprehensive study

  2. NCOA4 transcriptional coactivator inhibits activation of DNA replication origins.

    Science.gov (United States)

    Bellelli, Roberto; Castellone, Maria Domenica; Guida, Teresa; Limongello, Roberto; Dathan, Nina Alayne; Merolla, Francesco; Cirafici, Anna Maria; Affuso, Andrea; Masai, Hisao; Costanzo, Vincenzo; Grieco, Domenico; Fusco, Alfredo; Santoro, Massimo; Carlomagno, Francesca

    2014-07-01

    NCOA4 is a transcriptional coactivator of nuclear hormone receptors that undergoes gene rearrangement in human cancer. By combining studies in Xenopus laevis egg extracts and mouse embryonic fibroblasts (MEFs), we show here that NCOA4 is a minichromosome maintenance 7 (MCM7)-interacting protein that is able to control DNA replication. Depletion-reconstitution experiments in Xenopus laevis egg extracts indicate that NCOA4 acts as an inhibitor of DNA replication origin activation by regulating CMG (CDC45/MCM2-7/GINS) helicase. NCOA4(-/-) MEFs display unscheduled origin activation and reduced interorigin distance; this results in replication stress, as shown by the presence of fork stalling, reduction of fork speed, and premature senescence. Together, our findings indicate that NCOA4 acts as a regulator of DNA replication origins that helps prevent inappropriate DNA synthesis and replication stress.

  3. Stress responses and replication of plasmids in bacterial cells

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    Wegrzyn Alicja

    2002-05-01

    Full Text Available Abstract Plasmids, DNA (or rarely RNA molecules which replicate in cells autonomously (independently of chromosomes as non-essential genetic elements, play important roles for microbes grown under specific environmental conditions as well as in scientific laboratories and in biotechnology. For example, bacterial plasmids are excellent models in studies on regulation of DNA replication, and their derivatives are the most commonly used vectors in genetic engineering. Detailed mechanisms of replication initiation, which is the crucial process for efficient maintenance of plasmids in cells, have been elucidated for several plasmids. However, to understand plasmid biology, it is necessary to understand regulation of plasmid DNA replication in response to different environmental conditions in which host cells exist. Knowledge of such regulatory processes is also very important for those who use plasmids as expression vectors to produce large amounts of recombinant proteins. Variable conditions in large-scale fermentations must influence replication of plasmid DNA in cells, thus affecting the efficiency of recombinant gene expression significantly. Contrary to extensively investigated biochemistry of plasmid replication, molecular mechanisms of regulation of plasmid DNA replication in response to various environmental stress conditions are relatively poorly understood. There are, however, recently published studies that add significant data to our knowledge on relations between cellular stress responses and control of plasmid DNA replication. In this review we focus on plasmids derived from bacteriophage λ that are among the best investigated replicons. Nevertheless, recent results of studies on other plasmids are also discussed shortly.

  4. Publication bias and the failure of replication in experimental psychology.

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    Francis, Gregory

    2012-12-01

    Replication of empirical findings plays a fundamental role in science. Among experimental psychologists, successful replication enhances belief in a finding, while a failure to replicate is often interpreted to mean that one of the experiments is flawed. This view is wrong. Because experimental psychology uses statistics, empirical findings should appear with predictable probabilities. In a misguided effort to demonstrate successful replication of empirical findings and avoid failures to replicate, experimental psychologists sometimes report too many positive results. Rather than strengthen confidence in an effect, too much successful replication actually indicates publication bias, which invalidates entire sets of experimental findings. Researchers cannot judge the validity of a set of biased experiments because the experiment set may consist entirely of type I errors. This article shows how an investigation of the effect sizes from reported experiments can test for publication bias by looking for too much successful replication. Simulated experiments demonstrate that the publication bias test is able to discriminate biased experiment sets from unbiased experiment sets, but it is conservative about reporting bias. The test is then applied to several studies of prominent phenomena that highlight how publication bias contaminates some findings in experimental psychology. Additional simulated experiments demonstrate that using Bayesian methods of data analysis can reduce (and in some cases, eliminate) the occurrence of publication bias. Such methods should be part of a systematic process to remove publication bias from experimental psychology and reinstate the important role of replication as a final arbiter of scientific findings.

  5. Genome-wide association of bipolar disorder suggests an enrichment of replicable associations in regions near genes.

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    Erin N Smith

    2011-06-01

    Full Text Available Although a highly heritable and disabling disease, bipolar disorder's (BD genetic variants have been challenging to identify. We present new genotype data for 1,190 cases and 401 controls and perform a genome-wide association study including additional samples for a total of 2,191 cases and 1,434 controls. We do not detect genome-wide significant associations for individual loci; however, across all SNPs, we show an association between the power to detect effects calculated from a previous genome-wide association study and evidence for replication (P = 1.5×10(-7. To demonstrate that this result is not likely to be a false positive, we analyze replication rates in a large meta-analysis of height and show that, in a large enough study, associations replicate as a function of power, approaching a linear relationship. Within BD, SNPs near exons exhibit a greater probability of replication, supporting an enrichment of reproducible associations near functional regions of genes. These results indicate that there is likely common genetic variation associated with BD near exons (±10 kb that could be identified in larger studies and, further, provide a framework for assessing the potential for replication when combining results from multiple studies.

  6. Genome-wide association of bipolar disorder suggests an enrichment of replicable associations in regions near genes.

    Directory of Open Access Journals (Sweden)

    Erin N Smith

    2011-06-01

    Full Text Available Although a highly heritable and disabling disease, bipolar disorder's (BD genetic variants have been challenging to identify. We present new genotype data for 1,190 cases and 401 controls and perform a genome-wide association study including additional samples for a total of 2,191 cases and 1,434 controls. We do not detect genome-wide significant associations for individual loci; however, across all SNPs, we show an association between the power to detect effects calculated from a previous genome-wide association study and evidence for replication (P = 1.5×10(-7. To demonstrate that this result is not likely to be a false positive, we analyze replication rates in a large meta-analysis of height and show that, in a large enough study, associations replicate as a function of power, approaching a linear relationship. Within BD, SNPs near exons exhibit a greater probability of replication, supporting an enrichment of reproducible associations near functional regions of genes. These results indicate that there is likely common genetic variation associated with BD near exons (±10 kb that could be identified in larger studies and, further, provide a framework for assessing the potential for replication when combining results from multiple studies.

  7. Defects of mitochondrial DNA replication.

    Science.gov (United States)

    Copeland, William C

    2014-09-01

    Mitochondrial DNA is replicated by DNA polymerase γ in concert with accessory proteins such as the mitochondrial DNA helicase, single-stranded DNA binding protein, topoisomerase, and initiating factors. Defects in mitochondrial DNA replication or nucleotide metabolism can cause mitochondrial genetic diseases due to mitochondrial DNA deletions, point mutations, or depletion, which ultimately cause loss of oxidative phosphorylation. These genetic diseases include mitochondrial DNA depletion syndromes such as Alpers or early infantile hepatocerebral syndromes, and mitochondrial DNA deletion disorders, such as progressive external ophthalmoplegia, ataxia-neuropathy, or mitochondrial neurogastrointestinal encephalomyopathy. This review focuses on our current knowledge of genetic defects of mitochondrial DNA replication (POLG, POLG2, C10orf2, and MGME1) that cause instability of mitochondrial DNA and mitochondrial disease.

  8. Regulation of beta cell replication

    DEFF Research Database (Denmark)

    Lee, Ying C; Nielsen, Jens Høiriis

    2008-01-01

    Beta cell mass, at any given time, is governed by cell differentiation, neogenesis, increased or decreased cell size (cell hypertrophy or atrophy), cell death (apoptosis), and beta cell proliferation. Nutrients, hormones and growth factors coupled with their signalling intermediates have been...... suggested to play a role in beta cell mass regulation. In addition, genetic mouse model studies have indicated that cyclins and cyclin-dependent kinases that determine cell cycle progression are involved in beta cell replication, and more recently, menin in association with cyclin-dependent kinase...... inhibitors has been demonstrated to be important in beta cell growth. In this review, we consider and highlight some aspects of cell cycle regulation in relation to beta cell replication. The role of cell cycle regulation in beta cell replication is mostly from studies in rodent models, but whether...

  9. Shell Separation for Mirror Replication

    Science.gov (United States)

    1999-01-01

    NASA's Space Optics Manufacturing Center has been working to expand our view of the universe via sophisticated new telescopes. The Optics Center's goal is to develop low-cost, advanced space optics technologies for the NASA program in the 21st century - including the long-term goal of imaging Earth-like planets in distant solar systems. To reduce the cost of mirror fabrication, Marshall Space Flight Center (MSFC) has developed replication techniques, the machinery, and materials to replicate electro-formed nickel mirrors. Optics replication uses reusable forms, called mandrels, to make telescope mirrors ready for final finishing. MSFC optical physicist Bill Jones monitors a device used to chill a mandrel, causing it to shrink and separate from the telescope mirror without deforming the mirror's precisely curved surface.

  10. Mutagen-mediated enhancement of HIV-1 replication in persistently infected cells.

    Science.gov (United States)

    Sánchez-Jiménez, Carmen; Olivares, Isabel; de Ávila Lucas, Ana Isabel; Toledano, Víctor; Gutiérrez-Rivas, Mónica; Lorenzo-Redondo, Ramón; Grande-Pérez, Ana; Domingo, Esteban; López-Galíndez, Cecilio

    2012-03-15

    Lethal mutagenesis, a new antiviral strategy to extinguish virus through elevated mutation rates, was explored in H61-D cells an HIV-1 persistently infected lymphoid cell line. Three mutagenic agents: 5-hydroxy-2(')-deoxycytidine (5-OHdC), 5-fluorouracil (5-FU) and 2,2(')-difluoro-2(')-deoxycytidine (gemcitabine) were used. After 54 passages, treatments with 5-FU and gemcitabine reduced virus infectivity, p24 and RT activity. Treatment with the pyrimidine analog 5-OHdC resulted in increases of p24 production, RT activity and infectivity. Rise in viral replication by 5-OHdC during HIV-1 persistence is in contrast with its inhibitory effect in acute infections. Viral replication enhancement by 5-OHdC was associated with an increase in intracellular HIV-1 RNA mutations. Mechanisms of HIV-1 replication enhancement by 5-OHdC are unknown but some potential factors are discussed. Increase of HIV-1 replication by 5-OHdC cautions against the use, without previous analyses, of mutagenic nucleoside analogs for AIDS treatment.

  11. Lack of a unique termination site for the first round of bacteriophage lambda DNA replication

    Energy Technology Data Exchange (ETDEWEB)

    Valenzuela, M.S. (Brandeis Univ., Waltham, MA); Freifelder, D.; Inman, R.B.

    1976-01-01

    From previous data on the first round of bacteriophage lambdacIIcIII DNA replication (Schnos and Inman, 1970) it is possible to estimate, by extrapolation, the position on circular lambda DNA where bidirectional growing points meet. In the present study we have investigated whether this position occurs at a genetically defined site. To this end, replicative intermediates of lambda mutants containing either deletions to the left of the replication origin, or one deletion plus a duplication to the right, were analyzed in the electron microscope. Our results indicate that (i) leftward growing points can traverse the extrapolated termination point calculated from the lambdacIIcIII data, (ii) no discontinuity of either right or leftward growing fork position is observed, and (iii) the extrapolated termination points for these mutants are well removed from those calculated for lambdacIIcIII DNA. From these data we conclude that there is probably no unique termination site for the first round of lambda DNA replication and that termination occurs simply by collision of the growing forks.

  12. Amiloride inhibits the initiation of Coxsackievirus and poliovirus RNA replication by inhibiting VPg uridylylation.

    Science.gov (United States)

    Ogram, Sushma A; Boone, Christopher D; McKenna, Robert; Flanegan, James B

    2014-09-01

    The mechanism of amiloride inhibition of Coxsackievirus B3 (CVB3) and poliovirus type 1 (PV1) RNA replication was investigated using membrane-associated RNA replication complexes. Amiloride was shown to inhibit viral RNA replication and VPgpUpU synthesis. However, the drug had no effect on polymerase elongation activity during either (-) strand or (+) strand synthesis. These findings indicated that amiloride inhibited the initiation of RNA synthesis by inhibiting VPg uridylylation. In addition, in silico binding studies showed that amiloride docks in the VPg binding site on the back of the viral RNA polymerase, 3D(pol). Since VPg binding at this site on PV1 3D(pol) was previously shown to be required for VPg uridylylation, our results suggest that amiloride inhibits VPg binding to 3D(pol). In summary, our findings are consistent with a model in which amiloride inhibits VPgpUpU synthesis and viral RNA replication by competing with VPg for binding to 3D(pol).

  13. Replication Banding Patterns in Human Chromosomes Detected Using 5-ethynyl-2'-deoxyuridine Incorporation.

    Science.gov (United States)

    Hoshi, Osamu; Ushiki, Tatsuo

    2011-10-26

    A novel technique using the incorporation of 5-ethynyl-2'-deoxyuridine (EdU) into replicating DNA is described for the analysis of replicating banding patterns of human metaphase chromosomes. Human lymphocytes were synchronized with excess thymidine and treated with EdU during the late S phase of the cell cycle. The incorporated EdU was then detected in metaphase chromosomes using Alexa Fluor® 488 azides, through the 1,3-dipolar cycloaddition reaction of organic azides with the terminal acetylene group of EdU. Chromosomes with incorporated EdU showed a banding pattern similar to G-banding of normal human chromosomes. Imaging by atomic force microscopy (AFM) in liquid conditions showed that the structure of the chromosomes was well preserved even after EdU treatment. Comparison between fluorescence microscopy and AFM images of the same chromosome 1 indicated the presence of ridges and grooves in the chromatid arm, features that have been previously reported in relation to G-banding. These results suggest an intimate relationship between EdU-induced replication bands and G- or R-bands in human chromosomes. This technique is thus useful for analyzing the structure of chromosomes in relation to their banding patterns following DNA replication in the S phase.

  14. Lung function efficacy and symptomatic benefit of olodaterol once daily delivered via Respimat® versus placebo and formoterol twice daily in patients with GOLD 2–4 COPD: results from two replicate 48-week studies

    Directory of Open Access Journals (Sweden)

    Koch A

    2014-07-01

    Full Text Available Andrea Koch,1 Emilio Pizzichini,2 Alan Hamilton,3 Lorna Hart,3 Lawrence Korducki,4 Maria Cristina De Salvo,5 Pierluigi Paggiaro6 1Medical Clinic III for Pneumology, Allergology, Sleep and Respiratory Medicine, University Hospital Bochum-Bergmannsheil, Bochum, Germany; 2NUPAIVA (Asthma Research Center, Universidade Federal de Santa Catarina, Santa Catarina, Brazil; 3Boehringer Ingelheim, Burlington, Ontario, Canada; 4Boehringer Ingelheim Pharmaceuticals Inc., Ridgefield, CT, USA; 5Centro Médico Dra. De Salvo, Fundación Respirar, Buenos Aires, Argentina; 6Cardio-Thoracic and Vascular Department, University of Pisa, Pisa, Italy Abstract: Two replicate, multicenter, randomized, double-blind, placebo-controlled, parallel-group, Phase III studies investigated the long-term efficacy and safety of once-daily olodaterol via Respimat® versus placebo and formoterol over 48 weeks in patients with moderate to very severe chronic obstructive pulmonary disease receiving usual-care background therapy. Patients received once-daily olodaterol 5 or 10 µg, twice-daily formoterol 12 µg, or placebo. Co-primary end points were forced expiratory volume in 1 second (FEV1 area under the curve from 0–3 hours response, FEV1 trough response, and Mahler transition dyspnea index total score after 24 weeks; secondary end points included St George's Respiratory Questionnaire. Overall, 904 (Study 1222.13 and 934 (Study 1222.14 patients received treatment. Olodaterol significantly improved FEV1 area under the curve from 0–3 hours versus placebo in both studies (with olodaterol 5 µg, 0.151 L and 0.129 L; with olodaterol 10 µg, 0.165 L and 0.154 L; for all comparisons P<0.0001 and FEV1 trough responses versus placebo (0.053–0.085 L; P<0.01, as did formoterol. Primary analysis revealed no significant difference in transition dyspnea index focal score for any active treatment versus placebo. Post hoc analysis using pattern mixture modeling (accounting for

  15. Role of a cdk5-associated protein, p35, in herpes simplex virus type 1 replication in vivo.

    Science.gov (United States)

    Haenchen, Steve D; Utter, Jeff A; Bayless, Adam M; Dobrowsky, Rick T; Davido, David J

    2010-10-01

    Previous studies have shown that herpes simplex virus type 1 (HSV-1) replication is inhibited by the cyclin-dependent kinase (cdk) inhibitor roscovitine. One roscovitine-sensitive cdk that functions in neurons is cdk5, which is activated in part by its binding partner, p35. Because HSV establishes latent infections in sensory neurons, we sought to determine the role p35 plays in HSV-1 replication in vivo. For these studies, wild-type (wt) and p35−/− mice were infected with HSV-1 using the mouse ocular model of HSV latency and reactivation. The current results indicate that p35 is an important determinant of viral replication in vivo.

  16. Cataract surgery in previously vitrectomized eyes.

    Science.gov (United States)

    Akinci, A; Batman, C; Zilelioglu, O

    2008-05-01

    To evaluate the results of extracapsular cataract extraction (ECCE) and phacoemulsification (PHACO) performed in previously vitrectomized eyes. In this retrospective study, 56 vitrectomized eyes that had ECCE and 60 vitrectomized eyes that had PHACO were included in the study group while 65 eyes that had PHACO in the control group. The evaluated parameters were the incidence of intra-operative and postoperative complications (IPC) and visual outcomes. Chi-squared, independent samples and paired samples tests were used for comparing the results. Deep anterior chamber (AC) was significantly more common in the PHACO group of vitrectomized eyes (PGVE) and observed in eyes that had undergone extensive vitreous removal (p ECCE group and the PGVE (p > 0.05). Some of the intra-operative conditions such as posterior synechiae, primary posterior capsular opacification (PCO) and postoperative complications such as retinal detachment (RD), PCO were significantly more common in vitrectomized eyes than the controls (p ECCE group and the PGVE (p > 0.05). Deep AC is more common in eyes with extensive vitreous removal during PHACO than ECCE. Decreasing the bottle height is advised in this case. Except for this, the results of ECCE and PHACO are similar in previously vitrectomized eyes. Posterior synechiaes, primary and postoperative PCO and RD are more common in vitrectomized eyes than the controls.

  17. Regulation of Replication Recovery and Genome Integrity

    DEFF Research Database (Denmark)

    Colding, Camilla Skettrup

    facilitate replication recovery after MMS-induced replication stress. Our data reveal that control of Mrc1 turnover through the interplay between posttranslational modifications and INQ localization adds another layer of regulation to the replication checkpoint. We also add replication recovery to the list...... is mediated by Mrc1, which ensures Mec1 presence at the stalled replication fork thus facilitating Rad53 phosphorylation. When replication can be resumed safely, the replication checkpoint is deactivated and replication forks restart. One mechanism for checkpoint deactivation is the ubiquitin......-targeted proteasomal degradation of Mrc1. In this study, we describe a novel nuclear structure, the intranuclear quality control compartment (INQ), which regulates protein turnover and is important for recovery after replication stress. We find that upon methyl methanesulfonate (MMS)-induced replication stress, INQ...

  18. USP7/HAUSP: A SUMO deubiquitinase at the heart of DNA replication.

    Science.gov (United States)

    Smits, Veronique A J; Freire, Raimundo

    2016-09-01

    DNA replication is both highly conserved and controlled. Problematic DNA replication can lead to genomic instability and therefore carcinogenesis. Numerous mechanisms work together to achieve this tight control and increasing evidence suggests that post-translational modifications (phosphorylation, ubiquitination, SUMOylation) of DNA replication proteins play a pivotal role in this process. Here we discuss such modifications in the light of a recent article that describes a novel role for the deubiquitinase (DUB) USP7/HAUSP in the control of DNA replication. USP7 achieves this function by an unusual and novel mechanism, namely deubiquitination of SUMOylated proteins at the replication fork, making USP7 also a SUMO DUB (SDUB). This work extends previous observations of increased levels of SUMO and low levels of ubiquitin at the on-going replication fork. Here, we discuss this novel study, its contribution to the DNA replication and genomic stability field and what questions arise from this work.

  19. Physically Embedded Minimal Self-Replicating Systems

    DEFF Research Database (Denmark)

    Fellermann, Harold

    Self-replication is a fundamental property of all living organisms, yet has only been accomplished to limited extend in manmade systems. This thesis is part of the ongoing research endeavor to bridge the two sides of this gap. In particular, we present simulation results of a minimal life......-like, artificial, molecular aggregate (i.e. protocell) that has been proposed by Steen Rasussen and coworkers and is currently pursued both experimentally and computationally in interdisciplinary international research projects. We develop a space-time continuous physically motivated simulation framework based...... computational models. This allows us to address key issues of the replicating subsystems – container, genome, and metabolism – both individually and in mutual coupling. We analyze each step in the life-cycle of the molecular aggregate, and a final integrated simulation of the entire life-cycle is prepared. Our...

  20. Experimental Replication of an Aeroengine Combustion Instability

    Science.gov (United States)

    Cohen, J. M.; Hibshman, J. R.; Proscia, W.; Rosfjord, T. J.; Wake, B. E.; McVey, J. B.; Lovett, J.; Ondas, M.; DeLaat, J.; Breisacher, K.

    2000-01-01

    Combustion instabilities in gas turbine engines are most frequently encountered during the late phases of engine development, at which point they are difficult and expensive to fix. The ability to replicate an engine-traceable combustion instability in a laboratory-scale experiment offers the opportunity to economically diagnose the problem (to determine the root cause), and to investigate solutions to the problem, such as active control. The development and validation of active combustion instability control requires that the causal dynamic processes be reproduced in experimental test facilities which can be used as a test bed for control system evaluation. This paper discusses the process through which a laboratory-scale experiment was designed to replicate an instability observed in a developmental engine. The scaling process used physically-based analyses to preserve the relevant geometric, acoustic and thermo-fluid features. The process increases the probability that results achieved in the single-nozzle experiment will be scalable to the engine.

  1. Cellular Responses to Replication Problems

    NARCIS (Netherlands)

    M. Budzowska (Magdalena)

    2008-01-01

    textabstractDuring every S-phase cells need to duplicate their genomes so that both daughter cells inherit complete copies of genetic information. It is a tremendous task, given the large sizes of mammalian genomes and the required precision of DNA replication. A major threat to the accuracy and eff

  2. Covert Reinforcement: A Partial Replication.

    Science.gov (United States)

    Ripstra, Constance C.; And Others

    A partial replication of an investigation of the effect of covert reinforcement on a perceptual estimation task is described. The study was extended to include an extinction phase. There were five treatment groups: covert reinforcement, neutral scene reinforcement, noncontingent covert reinforcement, and two control groups. Each subject estimated…

  3. Break-Induced Replication and Genome Stability

    Directory of Open Access Journals (Sweden)

    Anna Malkova

    2012-10-01

    Full Text Available Genetic instabilities, including mutations and chromosomal rearrangements, lead to cancer and other diseases in humans and play an important role in evolution. A frequent cause of genetic instabilities is double-strand DNA breaks (DSBs, which may arise from a wide range of exogeneous and endogeneous cellular factors. Although the repair of DSBs is required, some repair pathways are dangerous because they may destabilize the genome. One such pathway, break-induced replication (BIR, is the mechanism for repairing DSBs that possesses only one repairable end. This situation commonly arises as a result of eroded telomeres or collapsed replication forks. Although BIR plays a positive role in repairing DSBs, it can alternatively be a dangerous source of several types of genetic instabilities, including loss of heterozygosity, telomere maintenance in the absence of telomerase, and non-reciprocal translocations. Also, mutation rates in BIR are about 1000 times higher as compared to normal DNA replication. In addition, micro-homology-mediated BIR (MMBIR, which is a mechanism related to BIR, can generate copy-number variations (CNVs as well as various complex chromosomal rearrangements. Overall, activation of BIR may contribute to genomic destabilization resulting in substantial biological consequences including those affecting human health.

  4. Continuously Cumulating Meta-Analysis and Replicability.

    Science.gov (United States)

    Braver, Sanford L; Thoemmes, Felix J; Rosenthal, Robert

    2014-05-01

    The current crisis in scientific psychology about whether our findings are irreproducible was presaged years ago by Tversky and Kahneman (1971), who noted that even sophisticated researchers believe in the fallacious Law of Small Numbers-erroneous intuitions about how imprecisely sample data reflect population phenomena. Combined with the low power of most current work, this often leads to the use of misleading criteria about whether an effect has replicated. Rosenthal (1990) suggested more appropriate criteria, here labeled the continuously cumulating meta-analytic (CCMA) approach. For example, a CCMA analysis on a replication attempt that does not reach significance might nonetheless provide more, not less, evidence that the effect is real. Alternatively, measures of heterogeneity might show that two studies that differ in whether they are significant might have only trivially different effect sizes. We present a nontechnical introduction to the CCMA framework (referencing relevant software), and then explain how it can be used to address aspects of replicability or more generally to assess quantitative evidence from numerous studies. We then present some examples and simulation results using the CCMA approach that show how the combination of evidence can yield improved results over the consideration of single studies.

  5. Reproducible and replicable CFD: it's harder than you think

    CERN Document Server

    Mesnard, Olivier

    2016-01-01

    Completing a full replication study of our previously published findings on bluff-body aerodynamics was harder than we thought. Despite the fact that we have good reproducible-research practices, sharing our code and data openly. Here's what we learned from three years, four CFD codes and hundreds of runs.

  6. Two failures to replicate high-performance-goal priming effects

    National Research Council Canada - National Science Library

    Harris, Christine R; Coburn, Noriko; Rohrer, Doug; Pashler, Harold

    2013-01-01

    .... Two direct replication attempts were performed. Results from the first experiment (n = 98) found no effect of priming, and the means were in the opposite direction from those reported by Bargh and colleagues...

  7. Replication of SNP associations with keratoconus in a Czech cohort

    Science.gov (United States)

    Krepelova, Anna; Klema, Jiri; Hysi, Pirro G.

    2017-01-01

    Introduction Keratoconus is a relatively frequent disease leading to severe visual impairment. Existing therapies are imperfect and clinical management may benefit from improved understanding of mechanisms leading to this disease. We aim to investigate the replication of 11 single nucleotide polymorphisms (SNPs) with keratoconus. Methods SNPs from loci previously found in association with keratoconus were genotyped in 165 keratoconus cases of Caucasian Czech origin (108 males and 57 females) and 193 population and gender-matched controls. They included rs1536482 (COL5A1), rs4839200 (KCND3), rs757219 and rs214884 (IMMP2L), rs1328083 and rs1328089 (DAOA), rs2721051 (FOXO1), rs4894535 (FNDC3B), rs4954218 (MAP3K19, RAB3GAP1), rs9938149 (ZNF469) and rs1324183 (MPDZ). A case-control association analysis was assessed using Fisher’s exact tests. Results The strongest association was found for rs1324183 (allelic test OR = 1.58; 95% CI, 1.10–2.24, p = 0.01). Statistically significant values were also obtained for rs2721051 (allelic test OR = 1.72; 95% CI, 1.07–2.77, p = 0.025) and rs4954218 (allelic test OR = 1.53; 95% CI, 1.01–2.34; p = 0.047) which showed an opposite effect direction compared to previously reported one. Conclusion Independent replication of association between two SNPs and keratoconus supports the association of these loci with the risks for the disease development, while the effect of rs4954218 warrants further investigation. Understanding the role of the genetic factors involved in keratoconus etiopathogenesis may facilitate development of novel therapies and an early detection. PMID:28207827

  8. Books average previous decade of economic misery.

    Directory of Open Access Journals (Sweden)

    R Alexander Bentley

    Full Text Available For the 20(th century since the Depression, we find a strong correlation between a 'literary misery index' derived from English language books and a moving average of the previous decade of the annual U.S. economic misery index, which is the sum of inflation and unemployment rates. We find a peak in the goodness of fit at 11 years for the moving average. The fit between the two misery indices holds when using different techniques to measure the literary misery index, and this fit is significantly better than other possible correlations with different emotion indices. To check the robustness of the results, we also analysed books written in German language and obtained very similar correlations with the German economic misery index. The results suggest that millions of books published every year average the authors' shared economic experiences over the past decade.

  9. Books average previous decade of economic misery.

    Science.gov (United States)

    Bentley, R Alexander; Acerbi, Alberto; Ormerod, Paul; Lampos, Vasileios

    2014-01-01

    For the 20(th) century since the Depression, we find a strong correlation between a 'literary misery index' derived from English language books and a moving average of the previous decade of the annual U.S. economic misery index, which is the sum of inflation and unemployment rates. We find a peak in the goodness of fit at 11 years for the moving average. The fit between the two misery indices holds when using different techniques to measure the literary misery index, and this fit is significantly better than other possible correlations with different emotion indices. To check the robustness of the results, we also analysed books written in German language and obtained very similar correlations with the German economic misery index. The results suggest that millions of books published every year average the authors' shared economic experiences over the past decade.

  10. 77 FR 70176 - Previous Participation Certification

    Science.gov (United States)

    2012-11-23

    ... URBAN DEVELOPMENT Previous Participation Certification AGENCY: Office of the Chief Information Officer... digital submission of all data and certifications is available via HUD's secure Internet systems. However...: Previous Participation Certification. OMB Approval Number: 2502-0118. Form Numbers: HUD-2530 ....

  11. REPLICATION TOOL AND METHOD OF PROVIDING A REPLICATION TOOL

    DEFF Research Database (Denmark)

    2016-01-01

    structured master surface (3a, 3b, 3c, 3d) having a lateral master pattern and a vertical master profile. The microscale structured master surface (3a, 3b, 3c, 3d) has been provided by localized pulsed laser treatment to generate microscale phase explosions. A method for producing a part with microscale......The invention relates to a replication tool (1, 1a, 1b) for producing a part (4) with a microscale textured replica surface (5a, 5b, 5c, 5d). The replication tool (1, 1a, 1b) comprises a tool surface (2a, 2b) defining a general shape of the item. The tool surface (2a, 2b) comprises a microscale...... energy directors on flange portions thereof uses the replication tool (1, 1a, 1b) to form an item (4) with a general shape as defined by the tool surface (2a, 2b). The formed item (4) comprises a microscale textured replica surface (5a, 5b, 5c, 5d) with a lateral arrangement of polydisperse microscale...

  12. A Transactional Asynchronous Replication Scheme for Mobile Database Systems

    Institute of Scientific and Technical Information of China (English)

    丁治明; 孟小峰; 王珊

    2002-01-01

    In mobile database systems, mobility of users has a significant impact on data replication. As a result, the various replica control protocols that exist today in traditional distributed and multidatabase environments are no longer suitable. To solve this problem, a new mobile database replication scheme, the Transaction-Level Result-Set Propagation (TLRSP)model, is put forward in this paper. The conflict detection and resolution strategy based on TLRSP is discussed in detail, and the implementation algorithm is proposed. In order to compare the performance of the TLRSP model with that of other mobile replication schemes, we have developed a detailed simulation model. Experimental results show that the TLRSP model provides an efficient support for replicated mobile database systems by reducing reprocessing overhead and maintaining database consistency.

  13. Molecular basis for viral selective replication in cancer cells: activation of CDK2 by adenovirus-induced cyclin E.

    Directory of Open Access Journals (Sweden)

    Pei-Hsin Cheng

    Full Text Available Adenoviruses (Ads with deletion of E1b55K preferentially replicate in cancer cells and have been used in cancer therapies. We have previously shown that Ad E1B55K protein is involved in induction of cyclin E for Ad replication, but this E1B55K function is not required in cancer cells in which deregulation of cyclin E is frequently observed. In this study, we investigated the interaction of cyclin E and CDK2 in Ad-infected cells. Ad infection significantly increased the large form of cyclin E (cyclin EL, promoted cyclin E/CDK2 complex formation and increased CDK2 phosphorylation at the T160 site. Activated CDK2 caused pRb phosphorylation at the S612 site. Repression of CDK2 activity with the chemical inhibitor roscovitine or with specific small interfering RNAs significantly decreased pRb phosphorylation, with concomitant repression of viral replication. Our results suggest that Ad-induced cyclin E activates CDK2 that targets the transcriptional repressor pRb to generate a cellular environment for viral productive replication. This study reveals a new molecular basis for oncolytic replication of E1b-deleted Ads and will aid in the development of new strategies for Ad oncolytic virotherapies.

  14. New Methods to Address Old Challenges: The Use of Administrative Data for Longitudinal Replication Studies of Child Maltreatment.

    Science.gov (United States)

    Hurren, Emily; Stewart, Anna; Dennison, Susan

    2017-09-15

    Administrative data are crucial to the "big data" revolution of social science and have played an important role in the development of child maltreatment research. These data are also of value to administrators, policy makers, and clinicians. The focus of this paper is the use of administrative data to produce and replicate longitudinal studies of child maltreatment. Child protection administrative data have several advantages. They are often population-based, and allow longitudinal examination of child maltreatment and complex multi-level analyses. They also allow comparison across subgroups and minority groups, remove burden from individuals to disclose traumatic experiences, and can be less biased than retrospective recall. Finally, they can be linked to data from other agencies to explore comorbidity and outcomes, and are comparatively cost and time effective. The benefits and challenges associated with the use of administrative data for longitudinal child maltreatment research become magnified when these data are used to produce replications. Techniques to address challenges and support future replication efforts include developing a biographical understanding of the systems from which the data are drawn, using multiple data sources to contextualize the data and research results, recognizing and adopting various approaches to replication, and documenting all data coding and manipulation processes. These techniques are illustrated in this paper via a case study of previous replication work.

  15. Chromatin dynamics at the replication fork: there's more to life than histones.

    Science.gov (United States)

    Whitehouse, Iestyn; Smith, Duncan J

    2013-04-01

    Before each division, eukaryotic cells face the daunting task of completely and accurately replicating a heterogeneous, chromatinized genome and repackaging both resulting daughters. Because replication requires strand separation, interactions between the DNA and its many associated proteins--including histones--must be transiently broken to allow the passage of the replication fork. Here, we will discuss the disruption and re-establishment of chromatin structure during replication, and the consequences of these processes for epigenetic inheritance. Published by Elsevier Ltd.

  16. Involvement of proliferating cell nuclear antigen (Cyclin) in DNA replication in living cells

    Energy Technology Data Exchange (ETDEWEB)

    Zuber, M.; Tan, E.M.; Ryoji, M.

    1989-01-01

    Proliferating cell nuclear antigen (PCNA) (also called cyclin) is known to stimulate the activity of DNA polymerase /delta/ but not the other DNA polymerases in vitro. The authors injected a human autoimmune antibody against PCNA into unfertilized eggs of Xenopus laevis and examined the effects of this antibody on the replication of injected plasmid DNA as well as egg chromosomes. The anti-PCNA antibody inhibited plasmid replication by up to 67%, demonstrating that PCNA is involved in plasmid replication in living cells. This result further implies that DNA polymerase /delta/ is necessary for plasmid replication in vivo, Anti-PCNA antibody alone did not block plasmid replication completely, but the residual replication was abolished by coinjection of a monoclonal antibody against DNA polymerase /alpha/. Anti-DNA polymerase /alpha/ alone inhibited plasmid replication by 63%. Thus, DNA ploymerase /alpha/ is also required for plasmid replication in this system. In similar studies on the replication of egg chromosomes, the inhibition by anti-PCNA antibody was only 30%, while anti-DNA polymerase /alpha/ antibody blocked 73% of replication. They concluded that the replication machineries of chromosomes and plasmid differ in their relative content of DNA polymerase /delta/. In addition, they obtained evidence through the use of phenylbutyl deoxyguanosine, an inhibitor of DNA polymearse /alpha/, that the structure of DNA polymerase /alpha/ holoenzyme for chromosome replication is significantly different from that for plasmid replication.

  17. Replication of Mu prophages lacking the central strong gyrase site.

    Science.gov (United States)

    Pato, Martin L

    2004-09-01

    Replication of Mu prophages lacking the central strong gyrase site (SGS) is severely slowed. To study details of the replication of these prophages, an assay was developed for determining the rate and extent of introduction of nicks at the 3'-ends of a Mu prophage, an early step in Mu replicative transposition. The maximal level of end-nicking of a prophage with the SGS, about 70-90% depending upon the host strain, was achieved within about 15 min after induction, whereas at that time less than 5% nicking was observed with a prophage lacking the SGS. The amount of nicking at the end of the SGS(-) prophage increased with time, and approx. 30% nicking of the SGS(-) prophage was achieved by 60 min post-induction. Nicking kinetics were identical at each end of the prophages, and no nicking was observed at the 5'-ends of the prophages, verifying in vivo the earlier results with in vitro systems. To determine if prophage location affects the kinetics of replication, we examined prophages at numerous chromosomal locations. SGS(+) prophages at different chromosomal locations showed essentially identical replication kinetics. SGS(-) prophages showed a range of delays in replication and host lysis. A gradient of delays was apparent, with prophages further from the chromosomal origin of replication, oriC, showing longer delays than ones nearer to oriC. However, there were also exceptions to this overall gradient. Possible explanations for the differences in the delays observed with SGS(-) prophages are discussed.

  18. SMARCAL1 maintains telomere integrity during DNA replication.

    Science.gov (United States)

    Poole, Lisa A; Zhao, Runxiang; Glick, Gloria G; Lovejoy, Courtney A; Eischen, Christine M; Cortez, David

    2015-12-01

    The SMARCAL1 (SWI/SNF related, matrix-associated, actin-dependent, regulator of chromatin, subfamily A-like 1) DNA translocase is one of several related enzymes, including ZRANB3 (zinc finger, RAN-binding domain containing 3) and HLTF (helicase-like transcription factor), that are recruited to stalled replication forks to promote repair and restart replication. These enzymes can perform similar biochemical reactions such as fork reversal; however, genetic studies indicate they must have unique cellular activities. Here, we present data showing that SMARCAL1 has an important function at telomeres, which present an endogenous source of replication stress. SMARCAL1-deficient cells accumulate telomere-associated DNA damage and have greatly elevated levels of extrachromosomal telomere DNA (C-circles). Although these telomere phenotypes are often found in tumor cells using the alternative lengthening of telomeres (ALT) pathway for telomere elongation, SMARCAL1 deficiency does not yield other ALT phenotypes such as elevated telomere recombination. The activity of SMARCAL1 at telomeres can be separated from its genome-maintenance activity in bulk chromosomal replication because it does not require interaction with replication protein A. Finally, this telomere-maintenance function is not shared by ZRANB3 or HLTF. Our results provide the first identification, to our knowledge, of an endogenous source of replication stress that requires SMARCAL1 for resolution and define differences between members of this class of replication fork-repair enzymes.

  19. Obinutuzumab for previously untreated chronic lymphocytic leukemia.

    Science.gov (United States)

    Abraham, Jame; Stegner, Mark

    2014-04-01

    Obinutuzumab was approved by the Food and Drug Administration in late 2013 for use in combination with chlorambucil for the treatment of patients with previously untreated chronic lymphocytic leukemia (CLL). The approval was based on results of an open-label phase 3 trial that showed improved progression-free survival (PFS) with the combination of obinutuzumab plus chlorambucil compared with chlorambucil alone. Obinutuzumab is a monoclonal antibody that targets CD20 antigen expressed on the surface of pre B- and mature B-lymphocytes. After binding to CD20, obinutuzumab mediates B-cell lysis by engaging immune effector cells, directly activating intracellular death signaling pathways, and activating the complement cascade. Immune effector cell activities include antibody-dependent cellular cytotoxicity and antibody-dependent cellular phagocytosis.

  20. Induced vaginal birth after previous caesarean section

    Directory of Open Access Journals (Sweden)

    Akylbek Tussupkaliyev

    2016-11-01

    Full Text Available Introduction The rate of operative birth by Caesarean section is constantly rising. In Kazakhstan, it reaches 27 per cent. Research data confirm that the percentage of successful vaginal births after previous Caesarean section is 50–70 per cent. How safe the induction of vaginal birth after Caesarean (VBAC remains unclear. Methodology The studied techniques of labour induction were amniotomy of the foetal bladder with the vulsellum ramus, intravaginal administration of E1 prostaglandin (Misoprostol, and intravenous infusion of Oxytocin-Richter. The assessment of rediness of parturient canals was conducted by Bishop’s score; the labour course was assessed by a partogram. The effectiveness of labour induction techniques was assessed by the number of administered doses, the time of onset of regular labour, the course of labour and the postpartum period and the presence of complications, and the course of the early neonatal period, which implied the assessment of the child’s condition, described in the newborn development record. The foetus was assessed by medical ultrasound and antenatal and intranatal cardiotocography (CTG. Obtained results were analysed with SAS statistical processing software. Results The overall percentage of successful births with intravaginal administration of Misoprostol was 93 per cent (83 of cases. This percentage was higher than in the amniotomy group (relative risk (RR 11.7 and was similar to the oxytocin group (RR 0.83. Amniotomy was effective in 54 per cent (39 of cases, when it induced regular labour. Intravenous oxytocin infusion was effective in 94 per cent (89 of cases. This percentage was higher than that with amniotomy (RR 12.5. Conclusions The success of vaginal delivery after previous Caesarean section can be achieved in almost 70 per cent of cases. At that, labour induction does not decrease this indicator and remains within population boundaries.

  1. Alphavirus polymerase and RNA replication.

    Science.gov (United States)

    Pietilä, Maija K; Hellström, Kirsi; Ahola, Tero

    2017-01-16

    Alphaviruses are typically arthropod-borne, and many are important pathogens such as chikungunya virus. Alphaviruses encode four nonstructural proteins (nsP1-4), initially produced as a polyprotein P1234. nsP4 is the core RNA-dependent RNA polymerase but all four nsPs are required for RNA synthesis. The early replication complex (RC) formed by the polyprotein P123 and nsP4 synthesizes minus RNA strands, and the late RC composed of fully processed nsP1-nsP4 is responsible for the production of genomic and subgenomic plus strands. Different parts of nsP4 recognize the promoters for minus and plus strands but the binding also requires the other nsPs. The alphavirus polymerase has been purified and is capable of de novo RNA synthesis only in the presence of the other nsPs. The purified nsP4 also has terminal adenylyltransferase activity, which may generate the poly(A) tail at the 3' end of the genome. Membrane association of the nsPs is vital for replication, and alphaviruses induce membrane invaginations called spherules, which form a microenvironment for RNA synthesis by concentrating replication components and protecting double-stranded RNA intermediates. The RCs isolated as crude membrane preparations are active in RNA synthesis in vitro, but high-resolution structure of the RC has not been achieved, and thus the arrangement of viral and possible host components remains unknown. For some alphaviruses, Ras-GTPase-activating protein (Src-homology 3 (SH3) domain)-binding proteins (G3BPs) and amphiphysins have been shown to be essential for RNA replication and are present in the RCs. Host factors offer an additional target for antivirals, as only few alphavirus polymerase inhibitors have been described.

  2. PDTC inhibits picornavirus polyprotein processing and RNA replication by transporting zinc ions into cells.

    NARCIS (Netherlands)

    Lanke, K.H.W.; Krenn, B.M.; Melchers, W.J.G.; Seipelt, J.; Kuppeveld, F.J.M. van

    2007-01-01

    Previously, it was shown that pyrrolidine dithiocarbamate (PDTC) inhibits proteolytic polyprotein processing and replication of human rhinovirus by transporting metal ions into cells. Here, it is shown that PDTC also inhibits replication of two other picornaviruses: coxsackievirus B3 (CVB3), a close

  3. Genes and sequences involved in the replication of cowpea mosaic virus RNAs

    NARCIS (Netherlands)

    Eggen, R.

    1989-01-01

    The aim of the studies described in this thesis was to gain more insight in the complex molecular mechanisms underlying the RNA replication of the cowpea mosaic virus genome. Previously the replication of CPMV RNA has been examined extensively with crude membrane fractions prepared from CP

  4. Genes and sequences involved in the replication of cowpea mosaic virus RNAs.

    NARCIS (Netherlands)

    Eggen, R.

    1989-01-01

    The aim of the studies described in this thesis was to gain more insight in the complex molecular mechanisms underlying the RNA replication of the cowpea mosaic virus genome. Previously the replication of CPMV RNA has been examined extensively with crude membrane fractions prepared from CPMV inf

  5. New criteria for selecting the origin of DNA replication in Wolbachia and closely related bacteria

    Directory of Open Access Journals (Sweden)

    Baldo Laura

    2007-06-01

    Full Text Available Abstract Background The annotated genomes of two closely related strains of the intracellular bacterium Wolbachia pipientis have been reported without the identifications of the putative origin of replication (ori. Identifying the ori of these bacteria and related alpha-Proteobacteria as well as their patterns of sequence evolution will aid studies of cell replication and cell density, as well as the potential genetic manipulation of these widespread intracellular bacteria. Results Using features that have been previously experimentally verified in the alpha-Proteobacterium Caulobacter crescentus, the origin of DNA replication (ori regions were identified in silico for Wolbachia strains and eleven other related bacteria belonging to Ehrlichia, Anaplasma, and Rickettsia genera. These features include DnaA-, CtrA- and IHF-binding sites as well as the flanking genes in C. crescentus. The Wolbachia ori boundary genes were found to be hemE and COG1253 protein (CBS domain protein. Comparisons of the putative ori region among related Wolbachia strains showed higher conservation of bases within binding sites. Conclusion The sequences of the ori regions described here are only similar among closely related bacteria while fundamental characteristics like presence of DnaA and IHF binding sites as well as the boundary genes are more widely conserved. The relative paucity of CtrA binding sites in the ori regions, as well as the absence of key enzymes associated with DNA replication in the respective genomes, suggest that several of these obligate intracellular bacteria may have altered replication mechanisms. Based on these analyses, criteria are set forth for identifying the ori region in genome sequencing projects.

  6. The three faces of riboviral spontaneous mutation: spectrum, mode of genome replication, and mutation rate.

    Directory of Open Access Journals (Sweden)

    Libertad García-Villada

    Full Text Available Riboviruses (RNA viruses without DNA replication intermediates are the most abundant pathogens infecting animals and plants. Only a few riboviral infections can be controlled with antiviral drugs, mainly because of the rapid appearance of resistance mutations. Little reliable information is available concerning i kinds and relative frequencies of mutations (the mutational spectrum, ii mode of genome replication and mutation accumulation, and iii rates of spontaneous mutation. To illuminate these issues, we developed a model in vivo system based on phage Qß infecting its natural host, Escherichia coli. The Qß RT gene encoding the Read-Through protein was used as a mutation reporter. To reduce uncertainties in mutation frequencies due to selection, the experimental Qß populations were established after a single cycle of infection and selection against RT(- mutants during phage growth was ameliorated by plasmid-based RT complementation in trans. The dynamics of Qß genome replication were confirmed to reflect the linear process of iterative copying (the stamping-machine mode. A total of 32 RT mutants were detected among 7,517 Qß isolates. Sequencing analysis of 45 RT mutations revealed a spectrum dominated by 39 transitions, plus 4 transversions and 2 indels. A clear template•primer mismatch bias was observed: A•C>C•A>U•G>G•U> transversion mismatches. The average mutation rate per base replication was ≈9.1×10(-6 for base substitutions and ≈2.3×10(-7 for indels. The estimated mutation rate per genome replication, μ(g, was ≈0.04 (or, per phage generation, ≈0.08, although secondary RT mutations arose during the growth of some RT mutants at a rate about 7-fold higher, signaling the possible impact of transitory bouts of hypermutation. These results are contrasted with those previously reported for other riboviruses to depict the current state of the art in riboviral mutagenesis.

  7. Dynamic replication of Web contents

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    The phenomenal growth of the World Wide Web has brought huge increase in the traffic to the popular web sites.Long delays and denial of service experienced by the end-users,especially during the peak hours,continues to be the common problem while accessing popular sites.Replicating some of the objects at multiple sites in a distributed web-server environment is one of the possible solutions to improve the response time/Iatency. The decision of what and where to replicate requires solving a constraint optimization problem,which is NP-complete in general.In this paper, we consider the problem of placing copies of objects in a distributed web server system to minimize the cost of serving read and write requests when the web servers have Iimited storage capacity.We formulate the problem as a 0-1 optimization problem and present a polynomial time greedy algorithm with backtracking to dynamically replicate objects at the appropriate sites to minimize a cost function.To reduce the solution search space,we present necessary condi tions for a site to have a replica of an object jn order to minimize the cost function We present simulation resuIts for a variety of problems to illustrate the accuracy and efficiency of the proposed algorithms and compare them with those of some well-known algorithms.The simulation resuIts demonstrate the superiority of the proposed algorithms.

  8. Conserved amino acids within the N-terminus of the West Nile virus NS4A protein contribute to virus replication, protein stability and membrane proliferation

    Energy Technology Data Exchange (ETDEWEB)

    Ambrose, R.L.; Mackenzie, J.M., E-mail: jason.mackenzie@unimelb.edu.au

    2015-07-15

    The West Nile virus strain Kunjin virus (WNV{sub KUN}) NS4A protein is a multifunctional protein involved in many aspects of the virus life-cycle and is a major component of the WNV{sub KUN} replication complex (RC). Previously we identified a conserved region in the C-terminus of NS4A regulating proteolytic processing and RC assembly, and now investigate key conserved residues in the N-terminus of NS4A and their contribution to WNV{sub KUN} replication. Mutation of P13 completely ablated replication, whereas, mutation of P48 and D49, near the first transmembrane helix, and G66 within the helix, showed variable defects in replication, virion secretion and membrane proliferation. Intriguingly, the P48 and G66 NS4A mutants resulted in specific proteasome depletion of NS4A that could in part be rescued with a proteasome inhibitor. Our results suggest that the N-terminus of NS4A contributes to correct folding and stability, essential for facilitating the essential roles of NS4A during replication. - Highlights: • Mutation of Proline13 of the WNV NS4A protein is lethal to replication. • 1st TMB helix of NS4A contributes to protein stability and membrane remodelling. • Unstable mutants of NS4A can be rescued with a proteasome inhibitor. • This study (and of others) contributes to a functional mapping of the NS4A protein.

  9. 22 CFR 40.91 - Certain aliens previously removed.

    Science.gov (United States)

    2010-04-01

    ... 22 Foreign Relations 1 2010-04-01 2010-04-01 false Certain aliens previously removed. 40.91... IMMIGRANTS UNDER THE IMMIGRATION AND NATIONALITY ACT, AS AMENDED Aliens Previously Removed § 40.91 Certain aliens previously removed. (a) 5-year bar. An alien who has been found inadmissible, whether as a result...

  10. Protein kinase C-delta regulates HIV-1 replication at an early post-entry step in macrophages

    Directory of Open Access Journals (Sweden)

    Contreras Xavier

    2012-05-01

    Full Text Available Abstract Background Macrophages, which are CD4 and CCR5 positive, can sustain HIV-1 replication for long periods of time. Thus, these cells play critical roles in the transmission, dissemination and persistence of viral infection. Of note, current antiviral therapies do not target macrophages efficiently. Previously, it was demonstrated that interactions between CCR5 and gp120 stimulate PKC. However, the PKC isozymes involved were not identified. Results In this study, we identified PKC-delta as a major cellular cofactor for HIV-1 replication in macrophages. Indeed, PKC-delta was stimulated following the interaction between the virus and its target cell. Moreover, inhibition of PKC-delta blocked the replication of R5-tropic viruses in primary human macrophages. However, this inhibition did not have significant effects on receptor and co-receptor expression or fusion. Additionally, it did not affect the formation of the early reverse transcription product containing R/U5 sequences, but did inhibit the synthesis of subsequent cDNAs. Importantly, the inhibition of PKC-delta altered the redistribution of actin, a cellular cofactor whose requirement for the completion of reverse transcription was previously established. It also prevented the association of the reverse transcription complex with the cytoskeleton. Conclusion This work highlights the importance of PKC-delta during early steps of the replicative cycle of HIV-1 in human macrophages.

  11. The Escherichia coli Tus-Ter replication fork barrier causes site-specific DNA replication perturbation in yeast

    DEFF Research Database (Denmark)

    Larsen, Nicolai B; Sass, Ehud; Suski, Catherine

    2014-01-01

    Replication fork (RF) pausing occurs at both 'programmed' sites and non-physiological barriers (for example, DNA adducts). Programmed RF pausing is required for site-specific DNA replication termination in Escherichia coli, and this process requires the binding of the polar terminator protein, Tus......, to specific DNA sequences called Ter. Here, we demonstrate that Tus-Ter modules also induce polar RF pausing when engineered into the Saccharomyces cerevisiae genome. This heterologous RF barrier is distinct from a number of previously characterized, protein-mediated, RF pause sites in yeast, as it is neither...

  12. Evaluating replicability of laboratory experiments in economics.

    Science.gov (United States)

    Camerer, Colin F; Dreber, Anna; Forsell, Eskil; Ho, Teck-Hua; Huber, Jürgen; Johannesson, Magnus; Kirchler, Michael; Almenberg, Johan; Altmejd, Adam; Chan, Taizan; Heikensten, Emma; Holzmeister, Felix; Imai, Taisuke; Isaksson, Siri; Nave, Gideon; Pfeiffer, Thomas; Razen, Michael; Wu, Hang

    2016-03-25

    The replicability of some scientific findings has recently been called into question. To contribute data about replicability in economics, we replicated 18 studies published in the American Economic Review and the Quarterly Journal of Economics between 2011 and 2014. All of these replications followed predefined analysis plans that were made publicly available beforehand, and they all have a statistical power of at least 90% to detect the original effect size at the 5% significance level. We found a significant effect in the same direction as in the original study for 11 replications (61%); on average, the replicated effect size is 66% of the original. The replicability rate varies between 67% and 78% for four additional replicability indicators, including a prediction market measure of peer beliefs.

  13. DNA polymerase beta can substitute for DNA polymerase I in the initiation of plasmid DNA replication.

    OpenAIRE

    1995-01-01

    We previously demonstrated that mammalian DNA polymerase beta can substitute for DNA polymerase I of Escherichia coli in DNA replication and in base excision repair. We have now obtained genetic evidence suggesting that DNA polymerase beta can substitute for E. coli DNA polymerase I in the initiation of replication of a plasmid containing a pMB1 origin of DNA replication. Specifically, we demonstrate that a plasmid with a pMB1 origin of replication can be maintained in an E. coli polA mutant ...

  14. The Werner and Bloom syndrome proteins help resolve replication blockage by converting (regressed) holliday junctions to functional replication forks.

    Science.gov (United States)

    Machwe, Amrita; Karale, Rajashree; Xu, Xioahua; Liu, Yilun; Orren, David K

    2011-08-16

    Cells cope with blockage of replication fork progression in a manner that allows DNA synthesis to be completed and genomic instability minimized. Models for resolution of blocked replication involve fork regression to form Holliday junction structures. The human RecQ helicases WRN and BLM (deficient in Werner and Bloom syndromes, respectively) are critical for maintaining genomic stability and thought to function in accurate resolution of replication blockage. Consistent with this notion, WRN and BLM localize to sites of blocked replication after certain DNA-damaging treatments and exhibit enhanced activity on replication and recombination intermediates. Here we examine the actions of WRN and BLM on a special Holliday junction substrate reflective of a regressed replication fork. Our results demonstrate that, in reactions requiring ATP hydrolysis, both WRN and BLM convert this Holliday junction substrate primarily to a four-stranded replication fork structure, suggesting they target the Holliday junction to initiate branch migration. In agreement, the Holliday junction binding protein RuvA inhibits the WRN- and BLM-mediated conversion reactions. Importantly, this conversion product is suitable for replication with its leading daughter strand readily extended by DNA polymerases. Furthermore, binding to and conversion of this Holliday junction are optimal at low MgCl(2) concentrations, suggesting that WRN and BLM preferentially act on the square planar (open) conformation of Holliday junctions. Our findings suggest that, subsequent to fork regression events, WRN and/or BLM could re-establish functional replication forks to help overcome fork blockage. Such a function is highly consistent with phenotypes associated with WRN- and BLM-deficient cells.

  15. Adenovirus sequences required for replication in vivo.

    OpenAIRE

    Wang, K.; Pearson, G D

    1985-01-01

    We have studied the in vivo replication properties of plasmids carrying deletion mutations within cloned adenovirus terminal sequences. Deletion mapping located the adenovirus DNA replication origin entirely within the first 67 bp of the adenovirus inverted terminal repeat. This region could be further subdivided into two functional domains: a minimal replication origin and an adjacent auxillary region which boosted the efficiency of replication by more than 100-fold. The minimal origin occup...

  16. Symmetry of interactions rules in incompletely connected random replicator ecosystems.

    Science.gov (United States)

    Kärenlampi, Petri P

    2014-06-01

    The evolution of an incompletely connected system of species with speciation and extinction is investigated in terms of random replicators. It is found that evolving random replicator systems with speciation do become large and complex, depending on speciation parameters. Antisymmetric interactions result in large systems, whereas systems with symmetric interactions remain small. A co-dominating feature is within-species interaction pressure: large within-species interaction increases species diversity. Average fitness evolves in all systems, however symmetry and connectivity evolve in small systems only. Newcomers get extinct almost immediately in symmetric systems. The distribution in species lifetimes is determined for antisymmetric systems. The replicator systems investigated do not show any sign of self-organized criticality. The generalized Lotka-Volterra system is shown to be a tedious way of implementing the replicator system.

  17. Replication Origin Specification Gets a Push.

    Science.gov (United States)

    Plosky, Brian S

    2015-12-03

    During the gap between G1 and S phases when replication origins are licensed and fired, it is possible that DNA translocases could disrupt pre-replicative complexes (pre-RCs). In this issue of Molecular Cell, Gros et al. (2015) find that pre-RCs can be pushed along DNA and retain the ability to support replication.

  18. Exploiting replicative stress to treat cancer

    DEFF Research Database (Denmark)

    Dobbelstein, Matthias; Sørensen, Claus Storgaard

    2015-01-01

    DNA replication in cancer cells is accompanied by stalling and collapse of the replication fork and signalling in response to DNA damage and/or premature mitosis; these processes are collectively known as 'replicative stress'. Progress is being made to increase our understanding of the mechanisms...

  19. Roles of conserved residues within the pre-NH2-terminal domain of herpes simplex virus 1 DNA polymerase in replication and latency in mice.

    Science.gov (United States)

    Terrell, Shariya L; Pesola, Jean M; Coen, Donald M

    2014-04-01

    The catalytic subunit of the herpes simplex virus 1 DNA polymerase (HSV-1 Pol) is essential for viral DNA synthesis and production of infectious virus in cell culture. While mutations that affect 5'-3' polymerase activity have been evaluated in animal models of HSV-1 infection, mutations that affect other functions of HSV-1 Pol have not. In a previous report, we utilized bacterial artificial chromosome technology to generate defined HSV-1 pol mutants with lesions in the previously uncharacterized pre-NH2-terminal domain. We found that the extreme N-terminal 42 residues (deletion mutant polΔN43) were dispensable for replication in cell culture, while residues 44-49 (alanine-substitution mutant polA6) were required for efficient viral DNA synthesis and production of infectious virus. In this study, we sought to address the importance of these conserved elements in viral replication in a mouse corneal infection model. Mutant virus polΔN43 exhibited no meaningful defect in acute or latent infection despite strong conservation of residues 1-42 with HSV-2 Pol. The polA6 mutation caused a modest defect in replication at the site of inoculation, and was severely impaired for ganglionic replication, even at high inocula that permitted efficient corneal replication. Additionally, the polA6 mutation resulted in reduced latency establishment and subsequent reactivation. Moreover, we found that the polA6 replication defect in cultured cells was exacerbated in resting cells as compared to dividing cells. These results reveal an important role for the conserved motif at residues 44-49 of HSV-1 Pol for ganglionic viral replication.

  20. Genome-wide alterations of the DNA replication program during tumor progression

    Science.gov (United States)

    Arneodo, A.; Goldar, A.; Argoul, F.; Hyrien, O.; Audit, B.

    2016-08-01

    Oncogenic stress is a major driving force in the early stages of cancer development. Recent experimental findings reveal that, in precancerous lesions and cancers, activated oncogenes may induce stalling and dissociation of DNA replication forks resulting in DNA damage. Replication timing is emerging as an important epigenetic feature that recapitulates several genomic, epigenetic and functional specificities of even closely related cell types. There is increasing evidence that chromosome rearrangements, the hallmark of many cancer genomes, are intimately associated with the DNA replication program and that epigenetic replication timing changes often precede chromosomic rearrangements. The recent development of a novel methodology to map replication fork polarity using deep sequencing of Okazaki fragments has provided new and complementary genome-wide replication profiling data. We review the results of a wavelet-based multi-scale analysis of genomic and epigenetic data including replication profiles along human chromosomes. These results provide new insight into the spatio-temporal replication program and its dynamics during differentiation. Here our goal is to bring to cancer research, the experimental protocols and computational methodologies for replication program profiling, and also the modeling of the spatio-temporal replication program. To illustrate our purpose, we report very preliminary results obtained for the chronic myelogeneous leukemia, the archetype model of cancer. Finally, we discuss promising perspectives on using genome-wide DNA replication profiling as a novel efficient tool for cancer diagnosis, prognosis and personalized treatment.

  1. The IFITMs Inhibit Zika Virus Replication

    Directory of Open Access Journals (Sweden)

    George Savidis

    2016-06-01

    Full Text Available Zika virus has emerged as a severe health threat with a rapidly expanding range. The IFITM family of restriction factors inhibits the replication of a broad range of viruses, including the closely related flaviruses West Nile virus and dengue virus. Here, we show that IFITM1 and IFITM3 inhibit Zika virus infection early in the viral life cycle. Moreover, IFITM3 can prevent Zika-virus-induced cell death. These results suggest that strategies to boost the actions and/or levels of the IFITMs might be useful for inhibiting a broad range of emerging viruses.

  2. A Comprehensive Analysis of the Dynamic Response to Aphidicolin-Mediated Replication Stress Uncovers Targets for ATM and ATMIN.

    Science.gov (United States)

    Mazouzi, Abdelghani; Stukalov, Alexey; Müller, André C; Chen, Doris; Wiedner, Marc; Prochazkova, Jana; Chiang, Shih-Chieh; Schuster, Michael; Breitwieser, Florian P; Pichlmair, Andreas; El-Khamisy, Sherif F; Bock, Christoph; Kralovics, Robert; Colinge, Jacques; Bennett, Keiryn L; Loizou, Joanna I

    2016-04-14

    The cellular response to replication stress requires the DNA-damage-responsive kinase ATM and its cofactor ATMIN; however, the roles of this signaling pathway following replication stress are unclear. To identify the functions of ATM and ATMIN in response to replication stress, we utilized both transcriptomics and quantitative mass-spectrometry-based phosphoproteomics. We found that replication stress induced by aphidicolin triggered widespread changes in both gene expression and protein phosphorylation patterns. These changes gave rise to distinct early and late replication stress responses. Furthermore, our analysis revealed previously unknown targets of ATM and ATMIN downstream of replication stress. We demonstrate ATMIN-dependent phosphorylation of H2AX and of CRMP2, a protein previously implicated in Alzheimer's disease but not in the DNA damage response. Overall, our dataset provides a comprehensive resource for discovering the cellular responses to replication stress and, potentially, associated pathologies. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  3. A Comprehensive Analysis of the Dynamic Response to Aphidicolin-Mediated Replication Stress Uncovers Targets for ATM and ATMIN

    Directory of Open Access Journals (Sweden)

    Abdelghani Mazouzi

    2016-04-01

    Full Text Available The cellular response to replication stress requires the DNA-damage-responsive kinase ATM and its cofactor ATMIN; however, the roles of this signaling pathway following replication stress are unclear. To identify the functions of ATM and ATMIN in response to replication stress, we utilized both transcriptomics and quantitative mass-spectrometry-based phosphoproteomics. We found that replication stress induced by aphidicolin triggered widespread changes in both gene expression and protein phosphorylation patterns. These changes gave rise to distinct early and late replication stress responses. Furthermore, our analysis revealed previously unknown targets of ATM and ATMIN downstream of replication stress. We demonstrate ATMIN-dependent phosphorylation of H2AX and of CRMP2, a protein previously implicated in Alzheimer’s disease but not in the DNA damage response. Overall, our dataset provides a comprehensive resource for discovering the cellular responses to replication stress and, potentially, associated pathologies.

  4. RNase H confers specificity in the dnaA-dependent initiation of replication at the unique origin of the Escherichia coli chromosome in vivo and in vitro.

    Science.gov (United States)

    Ogawa, T; Pickett, G G; Kogoma, T; Kornberg, A

    1984-02-01

    Escherichia coli rnh mutants defective in RNase H activity display the features of previously described sdrA (stable DNA replication) and dasF (dnaA suppressor) mutants: (i) sustained DNA replication in the absence of protein synthesis, (ii) lack of requirement for dnaA protein and the origin of replication (oriC), and (iii) sensitivity of growth to a rich medium. Both the sdrA mutants (selected for continued DNA replication in the absence of protein synthesis) and the dasF mutants (selected as dnaA suppressors) are defective in RNase H activity, measured in vitro. Furthermore, a 760-base-pair fragment containing the rnh+ structural gene complements the phenotype of each of the rnh, sdrA, and dasF mutants, indicative of a single gene. One function of RNase H in vivo is in the initiation of a cycle of DNA replication at oriC dependent on dnaA+. In keeping with these results, RNase H contributes to the specificity of dnaA protein-dependent replication initiated at oriC in a partially purified enzyme system.

  5. Replication, Communication, and the Population Dynamics of Scientific Discovery.

    Directory of Open Access Journals (Sweden)

    Richard McElreath

    Full Text Available Many published research results are false (Ioannidis, 2005, and controversy continues over the roles of replication and publication policy in improving the reliability of research. Addressing these problems is frustrated by the lack of a formal framework that jointly represents hypothesis formation, replication, publication bias, and variation in research quality. We develop a mathematical model of scientific discovery that combines all of these elements. This model provides both a dynamic model of research as well as a formal framework for reasoning about the normative structure of science. We show that replication may serve as a ratchet that gradually separates true hypotheses from false, but the same factors that make initial findings unreliable also make replications unreliable. The most important factors in improving the reliability of research are the rate of false positives and the base rate of true hypotheses, and we offer suggestions for addressing each. Our results also bring clarity to verbal debates about the communication of research. Surprisingly, publication bias is not always an obstacle, but instead may have positive impacts-suppression of negative novel findings is often beneficial. We also find that communication of negative replications may aid true discovery even when attempts to replicate have diminished power. The model speaks constructively to ongoing debates about the design and conduct of science, focusing analysis and discussion on precise, internally consistent models, as well as highlighting the importance of population dynamics.

  6. Replication, Communication, and the Population Dynamics of Scientific Discovery.

    Science.gov (United States)

    McElreath, Richard; Smaldino, Paul E

    2015-01-01

    Many published research results are false (Ioannidis, 2005), and controversy continues over the roles of replication and publication policy in improving the reliability of research. Addressing these problems is frustrated by the lack of a formal framework that jointly represents hypothesis formation, replication, publication bias, and variation in research quality. We develop a mathematical model of scientific discovery that combines all of these elements. This model provides both a dynamic model of research as well as a formal framework for reasoning about the normative structure of science. We show that replication may serve as a ratchet that gradually separates true hypotheses from false, but the same factors that make initial findings unreliable also make replications unreliable. The most important factors in improving the reliability of research are the rate of false positives and the base rate of true hypotheses, and we offer suggestions for addressing each. Our results also bring clarity to verbal debates about the communication of research. Surprisingly, publication bias is not always an obstacle, but instead may have positive impacts-suppression of negative novel findings is often beneficial. We also find that communication of negative replications may aid true discovery even when attempts to replicate have diminished power. The model speaks constructively to ongoing debates about the design and conduct of science, focusing analysis and discussion on precise, internally consistent models, as well as highlighting the importance of population dynamics.

  7. A Blm-Recql5 partnership in replication stress response

    Institute of Scientific and Technical Information of China (English)

    Xincheng Lu; Hua Lou; Guangbin Luo

    2011-01-01

    Deficiencies in DNA damage response and repair not only can result in genome instability and cancer predisposition, but also can render the cancer cells intrinsically more vulnerable to certain types of DNA damage insults. Particularly, replication stress is both a hallmark of human cancers and a common instigator for genome instability and cell death. Here, we review our work based on the genetic knockout studies on Blm and Recql5, two members of the mammalian RecQ helicase family. These studies have uncovered a unique partnership between these two helicases in the implementation of proper mitigation strategies under different circumstances to promote DNA replication and cell survival and suppress genome instability and cancer. In particular, current studies have revealed the presence of a novel Recql5/RECQL5-dependent mechanism for suppressing replication fork collapse in response to global replication fork stalling following exposure to camptothecin (CPT), a topoisomerase I inhibitor, and a potent inhibitor of DNA replication. The unique partnership between Blm and Recql5 in coping with the challenge imposed by replication stress is discussed. In addition, given that irinotecan and topotecan, two CPT derivatives, are currently used in clinic for treating human cancer patients with very promising results, the potential implication of the new findings from these studies in anticancer treatments is also discussed.

  8. P body-associated protein Mov10 inhibits HIV-1 replication at multiple stages.

    Science.gov (United States)

    Burdick, Ryan; Smith, Jessica L; Chaipan, Chawaree; Friew, Yeshitila; Chen, Jianbo; Venkatachari, Narasimhan J; Delviks-Frankenberry, Krista A; Hu, Wei-Shau; Pathak, Vinay K

    2010-10-01

    Recent studies have shown that APOBEC3G (A3G), a potent inhibitor of human immunodeficiency virus type 1 (HIV-1) replication, is localized to cytoplasmic mRNA-processing bodies (P bodies). However, the functional relevance of A3G colocalization with P body marker proteins has not been established. To explore the relationship between HIV-1, A3G, and P bodies, we analyzed the effects of overexpression of P body marker proteins Mov10, DCP1a, and DCP2 on HIV-1 replication. Our results show that overexpression of Mov10, a putative RNA helicase that was previously reported to belong to the DExD superfamily and was recently reported to belong to the Upf1-like group of helicases, but not the decapping enzymes DCP1a and DCP2, leads to potent inhibition of HIV-1 replication at multiple stages. Mov10 overexpression in the virus producer cells resulted in reductions in the steady-state levels of the HIV-1 Gag protein and virus production; Mov10 was efficiently incorporated into virions and reduced virus infectivity, in part by inhibiting reverse transcription. In addition, A3G and Mov10 overexpression reduced proteolytic processing of HIV-1 Gag. The inhibitory effects of A3G and Mov10 were additive, implying a lack of functional interaction between the two inhibitors. Small interfering RNA (siRNA)-mediated knockdown of endogenous Mov10 by 80% resulted in a 2-fold reduction in virus production but no discernible impact on the infectivity of the viruses after normalization for the p24 input, suggesting that endogenous Mov10 was not required for viral infectivity. Overall, these results show that Mov10 can potently inhibit HIV-1 replication at multiple stages.

  9. Short hairpin-loop-structured oligodeoxynucleotides reduce HSV-1 replication

    Directory of Open Access Journals (Sweden)

    Heinrich Jochen

    2009-04-01

    Full Text Available Abstract The Herpes simplex virus (HSV is known as an infectious agent and widespread in the human population. The symptoms of HSV infections can range from mild to life threatening, especially in immune-compromised individuals. HSV infections are commonly treated with the guanosine analogue Aciclovir, but reports of resistance are increasing. Efforts are made to establish single-stranded antisense oligodeoxynucleotides (as and small interfering ribonucleic acids (siRNAs for antiviral treatment. Recently, another class of short interfering nucleic acids, partially double-stranded hairpin loop-structured 54 mer oligodeoxynucleotides (ODNs, was shown to allow hydrolysis of HIV RNA by binding to the viral RNA. This leads to a substrate for the viral RNase H. To assess the potential of such ODNs for inhibition of HSV-1 replication, five partially double-stranded ODNs were designed based on the sequences of known siRNAs against HSV-1 with antiviral activity. Three of them are directed against early and two against leaky late genes. Primary human lung fibroblasts, MRC-5, and African green monkey kidney cells, Vero, were transfected with ODNs and subsequently infected. The effect on HSV-1 replication was determined by analyzing the virus titer in cell culture supernatants by quantitative PCR and plaque assays. An inhibitory effect was observed with all five selected ODNs, with two cases showing statistical significance in both cell types. The observed effect was sequence-specific and dose dependent. In one case the ODN was more efficient than a previously described siRNA directed against the same target site in the mRNA of UL5, a component of the helicase/primase complex. HSV-1 virions and ODNs can be applied simultaneously without transfection reagent, but at a 50-fold higher concentration to Vero cells with similar efficiencies. The results underline the potential of partially double-stranded hairpin loop-structured ODNs as antiviral agents.

  10. Replication of micro and nano surface geometries

    DEFF Research Database (Denmark)

    Hansen, Hans Nørgaard; Hocken, R.J.; Tosello, Guido

    2011-01-01

    : manufacture of net-shape micro/nano surfaces, tooling (i.e. master making), and surface quality control (metrology, inspection). Replication processes and methods as well as the metrology of surfaces to determine the degree of replication are presented and classified. Examples from various application areas...... are given including replication for surface texture measurements, surface roughness standards, manufacture of micro and nano structured functional surfaces, replicated surfaces for optical applications (e.g. optical gratings), and process chains based on combinations of repeated surface replication steps....

  11. Replication of prions in differentiated muscle cells.

    Science.gov (United States)

    Herbst, Allen; Aiken, Judd M; McKenzie, Debbie

    2014-01-01

    We have demonstrated that prions accumulate to high levels in non-proliferative C2C12 myotubes. C2C12 cells replicate as myoblasts but can be differentiated into myotubes. Earlier studies indicated that C2C12 myoblasts are not competent for prion replication. (1) We confirmed that observation and demonstrated, for the first time, that while replicative myoblasts do not accumulate PrP(Sc), differentiated post-mitotic myotube cultures replicate prions robustly. Here we extend our observations and describe the implication and utility of this system for replicating prions.

  12. Inhibition of Bim enhances replication of varicella-zoster virus and delays plaque formation in virus-infected cells.

    Science.gov (United States)

    Liu, Xueqiao; Cohen, Jeffrey I

    2014-01-01

    Programmed cell death (apoptosis) is an important host defense mechanism against intracellular pathogens, such as viruses. Accordingly, viruses have evolved multiple mechanisms to modulate apoptosis to enhance replication. Varicella-zoster virus (VZV) induces apoptosis in human fibroblasts and melanoma cells. We found that VZV triggered the phosphorylation of the proapoptotic proteins Bim and BAD but had little or no effect on other Bcl-2 family members. Since phosphorylation of Bim and BAD reduces their proapoptotic activity, this may prevent or delay apoptosis in VZV-infected cells. Phosphorylation of Bim but not BAD in VZV-infected cells was dependent on activation of the MEK/extracellular signal-regulated kinase (ERK) pathway. Cells knocked down for Bim showed delayed VZV plaque formation, resulting in longer survival of VZV-infected cells and increased replication of virus, compared with wild-type cells infected with virus. Conversely, overexpression of Bim resulted in earlier plaque formation, smaller plaques, reduced virus replication, and increased caspase 3 activity. Inhibition of caspase activity in VZV-infected cells overexpressing Bim restored levels of virus production similar to those seen with virus-infected wild-type cells. Previously we showed that VZV ORF12 activates ERK and inhibits apoptosis in virus-infected cells. Here we found that VZV ORF12 contributes to Bim and BAD phosphorylation. In summary, VZV triggers Bim phosphorylation; reduction of Bim levels results in longer survival of VZV-infected cells and increased VZV replication.

  13. Replication and encapsidation of the viroid-like satellite RNA of lucerne transient streak virus are supported in divergent hosts by cocksfoot mottle virus and turnip rosette virus.

    Science.gov (United States)

    Sehgal, O P; Sinha, R C; Gellatly, D L; Ivanov, I; AbouHaidar, M G

    1993-04-01

    Cocksfoot mottle sobemovirus supports replication and encapsidation of the viroid-like satellite RNA (sat-RNA) of lucerne transient streak virus (LTSV) in two monocotyledonous species, Triticum aestivum and Dactylis glomerata. Additionally, LTSV sat-RNA replicates effectively in the presence of turnip rosette sobemovirus in Brassica rapa, Raphanus raphanistrum and Sinapsis arvensis, but not in Thlaspi arvense or Nicotiana bigelovii, indicating that host species markedly influence this interaction. Previous reports of the association between LTSV sat-RNA and helper sobemoviruses were limited to dicotyledonous hosts. Our results demonstrate that the biological interaction between these two entities spans divergent dicotyledonous and monocotyledonous species.

  14. Preventing DNA over-replication: a Cdk perspective

    Directory of Open Access Journals (Sweden)

    Porter Andrew CG

    2008-01-01

    Full Text Available Abstract The cell cycle is tightly controlled to ensure that replication origins fire only once per cycle and that consecutive S-phases are separated by mitosis. When controls fail, DNA over-replication ensues: individual origins fire more than once per S-phase (re-replication or consecutive S-phases occur without intervening mitoses (endoreduplication. In yeast the cell cycle is controlled by a single cyclin dependent kinase (Cdk that prevents origin licensing at times when it promotes origin firing, and that is inactivated, via proteolysis of its partner cyclin, as cells undergo mitosis. A quantitative model describes three levels of Cdk activity: low activity allows licensing, intermediate activity allows firing but prevents licensing, and high activity promotes mitosis. In higher eukaryotes the situation is complicated by the existence of additional proteins (geminin, Cul4-Ddb1Cdt2, and Emi1 that control licensing. A current challenge is to understand how these various control mechanisms are co-ordinated and why the degree of redundancy between them is so variable. Here the experimental induction of DNA over-replication is reviewed in the context of the quantitative model of Cdk action. Endoreduplication is viewed as a consequence of procedures that cause Cdk activity to fall below the threshold required to prevent licensing, and re-replication as the result of procedures that increase that threshold value. This may help to explain why over-replication does not necessarily require reduced Cdk activity and how different mechanisms conspire to prevent over-replication. Further work is nevertheless required to determine exactly how losing just one licensing control mechanism often causes over-replication, and why this varies between cell systems.

  15. Verifying likelihoods for low template DNA profiles using multiple replicates

    Science.gov (United States)

    Steele, Christopher D.; Greenhalgh, Matthew; Balding, David J.

    2014-01-01

    To date there is no generally accepted method to test the validity of algorithms used to compute likelihood ratios (LR) evaluating forensic DNA profiles from low-template and/or degraded samples. An upper bound on the LR is provided by the inverse of the match probability, which is the usual measure of weight of evidence for standard DNA profiles not subject to the stochastic effects that are the hallmark of low-template profiles. However, even for low-template profiles the LR in favour of a true prosecution hypothesis should approach this bound as the number of profiling replicates increases, provided that the queried contributor is the major contributor. Moreover, for sufficiently many replicates the standard LR for mixtures is often surpassed by the low-template LR. It follows that multiple LTDNA replicates can provide stronger evidence for a contributor to a mixture than a standard analysis of a good-quality profile. Here, we examine the performance of the likeLTD software for up to eight replicate profiling runs. We consider simulated and laboratory-generated replicates as well as resampling replicates from a real crime case. We show that LRs generated by likeLTD usually do exceed the mixture LR given sufficient replicates, are bounded above by the inverse match probability and do approach this bound closely when this is expected. We also show good performance of likeLTD even when a large majority of alleles are designated as uncertain, and suggest that there can be advantages to using different profiling sensitivities for different replicates. Overall, our results support both the validity of the underlying mathematical model and its correct implementation in the likeLTD software. PMID:25082140

  16. Human ribonuclease H1 resolves R-loops and thereby enables progression of the DNA replication fork.

    Science.gov (United States)

    Parajuli, Shankar; Teasley, Daniel C; Murali, Bhavna; Jackson, Jessica; Vindigni, Alessandro; Stewart, Sheila A

    2017-09-15

    Faithful DNA replication is essential for genome stability. To ensure accurate replication, numerous complex and redundant replication and repair mechanisms function in tandem with the core replication proteins to ensure DNA replication continues even when replication challenges are present that could impede progression of the replication fork. A unique topological challenge to the replication machinery is posed by RNA-DNA hybrids, commonly referred to as R-loops. Although R-loops play important roles in gene expression and recombination at immunoglobulin sites, their persistence is thought to interfere with DNA replication by slowing or impeding replication fork progression. Therefore, it is of interest to identify DNA-associated enzymes that help resolve replication-impeding R-loops. Here, using DNA fiber analysis, we demonstrate that human ribonuclease H1 (RNH1) plays an important role in replication fork movement in the mammalian nucleus by resolving R-loops. We found that RNH1 depletion results in accumulation of RNA-DNA hybrids, slowing of replication forks, and increased DNA damage. Our data uncovered a role for RNH1 in global DNA replication in the mammalian nucleus. Because accumulation of RNA-DNA hybrids is linked to various human cancers and neurodegenerative disorders, our study raises the possibility that replication fork progression might be impeded, adding to increased genomic instability and contributing to disease. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. DNA replication stress: causes, resolution and disease.

    Science.gov (United States)

    Mazouzi, Abdelghani; Velimezi, Georgia; Loizou, Joanna I

    2014-11-15

    DNA replication is a fundamental process of the cell that ensures accurate duplication of the genetic information and subsequent transfer to daughter cells. Various pertubations, originating from endogenous or exogenous sources, can interfere with proper progression and completion of the replication process, thus threatening genome integrity. Coordinated regulation of replication and the DNA damage response is therefore fundamental to counteract these challenges and ensure accurate synthesis of the genetic material under conditions of replication stress. In this review, we summarize the main sources of replication stress and the DNA damage signaling pathways that are activated in order to preserve genome integrity during DNA replication. We also discuss the association of replication stress and DNA damage in human disease and future perspectives in the field. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Evidence of a conserved role for Chlamydia HtrA in the replication phase of the chlamydial developmental cycle.

    Science.gov (United States)

    Patel, Pooja; De Boer, Leonore; Timms, Peter; Huston, Wilhelmina May

    2014-08-01

    Identification of the HtrA inhibitor JO146 previously enabled us to demonstrate an essential function for HtrA during the mid-replicative phase of the Chlamydia trachomatis developmental cycle. Here we extend our investigations to other members of the Chlamydia genus. C. trachomatis isolates with distinct replicative phase growth kinetics showed significant loss of viable infectious progeny after HtrA was inhibited during the replicative phase. Mid-replicative phase addition of JO146 was also significantly detrimental to Chlamydia pecorum, Chlamydia suis and Chlamydia cavie. These data combined indicate that HtrA has a conserved critical role during the replicative phase of the chlamydial developmental cycle.

  19. Replication-associated gene dosage effects shape the genomes of fast-growing bacteria but only for transcription and translation genes.

    Science.gov (United States)

    Couturier, Etienne; Rocha, Eduardo P C

    2006-03-01

    The bidirectional replication of bacterial genomes leads to transient gene dosage effects. Here, we show that such effects shape the chromosome organisation of fast-growing bacteria and that they correlate strongly with maximal growth rate. Surprisingly the predicted maximal number of replication rounds shows little if any phylogenetic inertia, suggesting that it is a very labile trait. Yet, a combination of theoretical and statistical analyses predicts that dozens of replication forks may be simultaneously present in the cells of certain species. This suggests a strikingly efficient management of the replication apparatus, of replication fork arrests and of chromosome segregation in such cells. Gene dosage effects strongly constrain the position of genes involved in translation and transcription, but not other highly expressed genes. The relative proximity of the former genes to the origin of replication follows the regulatory dependencies observed under exponential growth, as the bias is stronger for RNA polymerase, then rDNA, then ribosomal proteins and tDNA. Within tDNAs we find that only the positions of the previously proposed 'ubiquitous' tRNA, which translate the most frequent codons in highly expressed genes, show strong signs of selection for gene dosage effects. Finally, we provide evidence for selection acting upon genome organisation to take advantage of gene dosage effects by identifying a positive correlation between genome stability and the number of simultaneous replication rounds. We also show that gene dosage effects can explain the over-representation of highly expressed genes in the largest replichore of genomes containing more than one chromosome. Together, these results demonstrate that replication-associated gene dosage is an important determinant of chromosome organisation and dynamics, especially among fast-growing bacteria.

  20. Assembling semiconductor nanocomposites using DNA replication technologies.

    Energy Technology Data Exchange (ETDEWEB)

    Heimer, Brandon W.; Crown, Kevin K.; Bachand, George David

    2005-11-01

    Deoxyribonucleic acid (DNA) molecules represent Nature's genetic database, encoding the information necessary for all cellular processes. From a materials engineering perspective, DNA represents a nanoscale scaffold with highly refined structure, stability across a wide range of environmental conditions, and the ability to interact with a range of biomolecules. The ability to mass-manufacture functionalized DNA strands with Angstrom-level resolution through DNA replication technology, however, has not been explored. The long-term goal of the work presented in this report is focused on exploiting DNA and in vitro DNA replication processes to mass-manufacture nanocomposite materials. The specific objectives of this project were to: (1) develop methods for replicating DNA strands that incorporate nucleotides with ''chemical handles'', and (2) demonstrate attachment of nanocrystal quantum dots (nQDs) to functionalized DNA strands. Polymerase chain reaction (PCR) and primer extension methodologies were used to successfully synthesize amine-, thiol-, and biotin-functionalized DNA molecules. Significant variability in the efficiency of modified nucleotide incorporation was observed, and attributed to the intrinsic properties of the modified nucleotides. Noncovalent attachment of streptavidin-coated nQDs to biotin-modified DNA synthesized using the primer extension method was observed by epifluorescence microscopy. Data regarding covalent attachment of nQDs to amine- and thiol-functionalized DNA was generally inconclusive; alternative characterization tools are necessary to fully evaluate these attachment methods. Full realization of this technology may facilitate new approaches to manufacturing materials at the nanoscale. In addition, composite nQD-DNA materials may serve as novel recognition elements in sensor devices, or be used as diagnostic tools for forensic analyses. This report summarizes the results obtained over the course of this 1-year

  1. Metabolically Coupled Replicator Systems: Overview of an RNA-world model concept of prebiotic evolution on mineral surfaces.

    Science.gov (United States)

    Czárán, Tamás; Könnyű, Balázs; Szathmáry, Eörs

    2015-09-21

    Metabolically Coupled Replicator Systems (MCRS) are a family of models implementing a simple, physico-chemically and ecologically feasible scenario for the first steps of chemical evolution towards life. Evolution in an abiotically produced RNA-population sets in as soon as any one of the RNA molecules become autocatalytic by engaging in template directed self-replication from activated monomers, and starts increasing exponentially. Competition for the finite external supply of monomers ignites selection favouring RNA molecules with catalytic activity helping self-replication by any possible means. One way of providing such autocatalytic help is to become a replicase ribozyme. An additional way is through increasing monomer supply by contributing to monomer synthesis from external resources, i.e., by evolving metabolic enzyme activity. Retroevolution may build up an increasingly autotrophic, cooperating community of metabolic ribozymes running an increasingly complicated and ever more efficient metabolism. Maintaining such a cooperating community of metabolic replicators raises two serious ecological problems: one is keeping the system coexistent in spite of the different replicabilities of the cooperating replicators; the other is constraining parasitism, i.e., keeping "cheaters" in check. Surface-bound MCRS provide an automatic solution to both problems: coexistence and parasite resistance are the consequences of assuming the local nature of metabolic interactions. In this review we present an overview of results published in previous articles, showing that these effects are, indeed, robust in different MCRS implementations, by considering different environmental setups and realistic chemical details in a few different models. We argue that the MCRS model framework naturally offers a suitable starting point for the future modelling of membrane evolution and extending the theory to cover the emergence of the first protocell in a self-consistent manner. The

  2. Rapid transient production in plants by replicating and non-replicating vectors yields high quality functional anti-HIV antibody.

    Directory of Open Access Journals (Sweden)

    Frank Sainsbury

    Full Text Available The capacity of plants and plant cells to produce large amounts of recombinant protein has been well established. Due to advantages in terms of speed and yield, attention has recently turned towards the use of transient expression systems, including viral vectors, to produce proteins of pharmaceutical interest in plants. However, the effects of such high level expression from viral vectors and concomitant effects on host cells may affect the quality of the recombinant product.To assess the quality of antibodies transiently expressed to high levels in plants, we have expressed and characterised the human anti-HIV monoclonal antibody, 2G12, using both replicating and non-replicating systems based on deleted versions of Cowpea mosaic virus (CPMV RNA-2. The highest yield (approximately 100 mg/kg wet weight leaf tissue of affinity purified 2G12 was obtained when the non-replicating CPMV-HT system was used and the antibody was retained in the endoplasmic reticulum (ER. Glycan analysis by mass-spectrometry showed that the glycosylation pattern was determined exclusively by whether the antibody was retained in the ER and did not depend on whether a replicating or non-replicating system was used. Characterisation of the binding and neutralisation properties of all the purified 2G12 variants from plants showed that these were generally similar to those of the Chinese hamster ovary (CHO cell-produced 2G12.Overall, the results demonstrate that replicating and non-replicating CPMV-based vectors are able to direct the production of a recombinant IgG similar in activity to the CHO-produced control. Thus, a complex recombinant protein was produced with no apparent effect on its biochemical properties using either high-level expression or viral replication. The speed with which a recombinant pharmaceutical with excellent biochemical characteristics can be produced transiently in plants makes CPMV-based expression vectors an attractive option for

  3. High rate of mismatch extension during reverse transcription in a single round of retrovirus replication.

    Science.gov (United States)

    Pulsinelli, G A; Temin, H M

    1994-09-27

    We made spleen necrosis virus-based retroviral vectors with mutations at the 3' end of the primer binding site region to observe the effects of terminal mismatches on retroviral replication. These vectors, when compared to a vector with the wild-type primer binding sequence, allowed us to assay the effects of the mutations on the viral titer during a single cycle of replication. The mutant vectors had titers that were comparable to the wild-type vector, indicating that reverse transcriptase has no trouble extending mismatches of as many as 3 bases under normal in vivo conditions. These results confirm and extend previous in vitro studies [Yu, H. & Goodman, M. (1992) J. Biol. Chem. 15, 10888-10896] that showed that such mismatch extension could occur in a cell-free system at high concentrations of incorrect nucleotides and in the absence of correct nucleotides. We now show that mismatch extension can occur during normal retroviral replication in cells and at normal physiological nucleotide concentrations.

  4. Slingshot Dynamics for Self Replicating Probes and the Effect on Exploration Timescales

    CERN Document Server

    Nicholson, Arwen

    2013-01-01

    Interstellar probes can carry out slingshot manoeuvres around the stars they visit, gaining a boost in velocity by extracting energy from the star's motion around the Galactic Centre. These maneouvres carry little to no extra energy cost, and in previous work it has been shown that a single Voyager-like probe exploring the galaxy does so 100 times faster when carrying out these slingshots than when navigating purely by powered flight (Forgan et al. 2012). We expand on these results by repeating the experiment with self-replicating probes. The probes explore a box of stars representative of the local Solar neighbourhood, to investigate how self-replication affects exploration timescales when compared with a single non-replicating probe. We explore three different scenarios of probe behaviour: i) standard powered flight to the nearest unvisited star (no slingshot techniques used), ii) flight to the nearest unvisited star using slingshot techniques, and iii) flight to the next unvisited star that will give the m...

  5. Varicella-zoster virus (VZV) origin of DNA replication oriS influences origin-dependent DNA replication and flanking gene transcription.

    Science.gov (United States)

    Khalil, Mohamed I; Sommer, Marvin H; Hay, John; Ruyechan, William T; Arvin, Ann M

    2015-07-01

    The VZV genome has two origins of DNA replication (oriS), each of which consists of an AT-rich sequence and three origin binding protein (OBP) sites called Box A, C and B. In these experiments, the mutation in the core sequence CGC of the Box A and C not only inhibited DNA replication but also inhibited both ORF62 and ORF63 expression in reporter gene assays. In contrast the Box B mutation did not influence DNA replication or flanking gene transcription. These results suggest that efficient DNA replication enhances ORF62 and ORF63 transcription. Recombinant viruses carrying these mutations in both sites and one with a deletion of the whole oriS were constructed. Surprisingly, the recombinant virus lacking both copies of oriS retained the capacity to replicate in melanoma and HELF cells suggesting that VZV has another origin of DNA replication.

  6. Self-replication of DNA rings

    Science.gov (United States)

    Kim, Junghoon; Lee, Junwye; Hamada, Shogo; Murata, Satoshi; Ha Park, Sung

    2015-06-01

    Biology provides numerous examples of self-replicating machines, but artificially engineering such complex systems remains a formidable challenge. In particular, although simple artificial self-replicating systems including wooden blocks, magnetic systems, modular robots and synthetic molecular systems have been devised, such kinematic self-replicators are rare compared with examples of theoretical cellular self-replication. One of the principal reasons for this is the amount of complexity that arises when you try to incorporate self-replication into a physical medium. In this regard, DNA is a prime candidate material for constructing self-replicating systems due to its ability to self-assemble through molecular recognition. Here, we show that DNA T-motifs, which self-assemble into ring structures, can be designed to self-replicate through toehold-mediated strand displacement reactions. The inherent design of these rings allows the population dynamics of the systems to be controlled. We also analyse the replication scheme within a universal framework of self-replication and derive a quantitative metric of the self-replicability of the rings.

  7. Modulation of vacuolar pH is required for replication of Edwardsiella ictaluri in channel catfish macrophages.

    Science.gov (United States)

    Baumgartner, Wes A; Dubytska, Lidiya; Rogge, Matthew L; Mottram, Peter J; Thune, Ronald L

    2014-06-01

    Previous in vitro work demonstrated that Edwardsiella ictaluri produces an acid-activated urease that can modulate environmental pH through the production of ammonia from urea. Additional work revealed that expression of the E. ictaluri type III secretion system (T3SS) is upregulated by acidic pH. Both the urease and the T3SS were previously shown to be essential to intracellular replication. In this work, fluorescence microscopy with LysoTracker Red DND-99 (LTR) indicated that E. ictaluri-containing vacuoles (ECV) became acidified following ingestion by head kidney-derived macrophages (HKDM). In vivo ratiometric imaging demonstrated a lowered ECV pH, which fell to as low as pH 4 but subsequently increased to pH 6 or greater. Inhibition of vacuolar H(+)-ATPases by use of the specific inhibitor bafilomycin A1 abrogated both ECV acidification and intracellular replication in HKDM. Failure of an E. ictaluri urease knockout mutant to increase the ECV pH in the in vivo ratiometric assay suggests that ammonia produced by the urease reaction mediates the pH increase. Additionally, when the specific arginase inhibitor l-norvaline was used to treat E. ictaluri-infected HKDM, the ECV failed to neutralize and E. ictaluri was unable to replicate. This indicates that the HKDM-encoded arginase enzyme produces the urea used by the E. ictaluri urease enzyme. Failure of the ECV to acidify would prevent both upregulation of the T3SS and activation of the urease enzyme, either of which would prevent E. ictaluri from replicating in HKDM. Failure of the ECV to neutralize would result in a vacuolar pH too low to support E. ictaluri replication.

  8. Adenovirus replication and transcription sites are spatially separated in the nucleus of infected cells.

    Science.gov (United States)

    Pombo, A; Ferreira, J; Bridge, E; Carmo-Fonseca, M

    1994-11-01

    We have visualized the intranuclear topography of adenovirus replication and transcription in infected HeLa cells. The results show that viral DNA replication occurs in multiple foci that are highly organized in the nucleoplasm. Pulse-chase experiments indicate that newly synthesized viral double-stranded DNA molecules are displaced from the replication foci and spread throughout the nucleoplasm, while the single-stranded DNA replication intermediates accumulate in adjacent sites. Double-labelling experiments and confocal microscopy show that replication occurs in foci localized at the periphery of the sites where single-stranded DNA accumulates. The simultaneous visualization of viral replication and transcription reveals that the sites of transcription are predominantly separated from the sites of replication. Transcription is detected adjacent to the replication foci and extends around the sites of single-stranded DNA accumulation. These data indicate that newly synthesized double-stranded DNA molecules are displaced from the replication foci and spread in the surrounding nucleoplasm, where they are used as templates for transcription. Splicing snRNPs are shown to co-localize with the sites of transcription and to be excluded from the sites of replication. This provides evidence that splicing of viral RNAs occurs co-transcriptionally and that the sites of viral DNA replication are spatially distinct from the sites of RNA transcription and processing.

  9. The Dual Nature of Nek9 in Adenovirus Replication

    OpenAIRE

    Jung, Richard; Radko, Sandi; Pelka, Peter

    2016-01-01

    To successfully replicate in an infected host cell, a virus must overcome sophisticated host defense mechanisms. Viruses, therefore, have evolved a multitude of devices designed to circumvent cellular defenses that would lead to abortive infection. Previous studies have identified Nek9, a cellular kinase, as a binding partner of adenovirus E1A, but the biology behind this association remains a mystery. Here we show that Nek9 is a transcriptional repressor that functions together with E1A to s...

  10. DNA Replication via Entanglement Swapping

    CERN Document Server

    Pusuluk, Onur

    2010-01-01

    Quantum effects are mainly used for the determination of molecular shapes in molecular biology, but quantum information theory may be a more useful tool to understand the physics of life. Molecular biology assumes that function is explained by structure, the complementary geometries of molecules and weak intermolecular hydrogen bonds. However, both this assumption and its converse are possible if organic molecules and quantum circuits/protocols are considered as hardware and software of living systems that are co-optimized during evolution. In this paper, we try to model DNA replication as a multiparticle entanglement swapping with a reliable qubit representation of nucleotides. In the model, molecular recognition of a nucleotide triggers an intrabase entanglement corresponding to a superposition state of different tautomer forms. Then, base pairing occurs by swapping intrabase entanglements with interbase entanglements.

  11. Le Chatelier's principle in replicator dynamics

    Science.gov (United States)

    Allahverdyan, Armen E.; Galstyan, Aram

    2011-10-01

    The Le Chatelier principle states that physical equilibria are not only stable, but they also resist external perturbations via short-time negative-feedback mechanisms: a perturbation induces processes tending to diminish its results. The principle has deep roots, e.g., in thermodynamics it is closely related to the second law and the positivity of the entropy production. Here we study the applicability of the Le Chatelier principle to evolutionary game theory, i.e., to perturbations of a Nash equilibrium within the replicator dynamics. We show that the principle can be reformulated as a majorization relation. This defines a stability notion that generalizes the concept of evolutionary stability. We determine criteria for a Nash equilibrium to satisfy the Le Chatelier principle and relate them to mutualistic interactions (game-theoretical anticoordination) showing in which sense mutualistic replicators can be more stable than (say) competing ones. There are globally stable Nash equilibria, where the Le Chatelier principle is violated even locally: in contrast to the thermodynamic equilibrium a Nash equilibrium can amplify small perturbations, though both types of equilibria satisfy the detailed balance condition.

  12. DNA instability in replicating Huntington's disease lymphoblasts

    Directory of Open Access Journals (Sweden)

    Frati Luigi

    2009-02-01

    Full Text Available Abstract Background The expanded CAG repeat in the Huntington's disease (HD gene may display tissue-specific variability (e.g. triplet mosaicism in repeat length, the longest mutations involving mitotic (germ and glial cells and postmitotic (neurons cells. What contributes to the triplet mutability underlying the development of HD nevertheless remains unknown. We investigated whether, besides the increased DNA instability documented in postmitotic neurons, possible environmental and genetic mechanisms, related to cell replication, may concur to determine CAG repeat mutability. To test this hypothesis we used, as a model, cultured HD patients' lymphoblasts with various CAG repeat lengths. Results Although most lymphoblastoid cell lines (88% showed little or no repeat instability even after six or more months culture, in lymphoblasts with large expansion repeats beyond 60 CAG repeats the mutation size and triplet mosaicism always increased during replication, implying that the repeat mutability for highly expanded mutations may quantitatively depend on the triplet expansion size. None of the investigated genetic factors, potentially acting in cis to the mutation, significantly influence the repeat changes. Finally, in our experiments certain drugs controlled triplet expansion in two prone-to-expand HD cell lines carrying large CAG mutations. Conclusion Our data support quantitative evidence that the inherited CAG length of expanded alleles has a major influence on somatic repeat variation. The longest triplet expansions show wide somatic variations and may offer a mechanistic model to study triplet drug-controlled instability and genetic factors influencing it.

  13. The origin of replication, oriC, and the dnaA protein are dispensable in stable DNA replication (sdrA) mutants of Escherichia coli K-12.

    Science.gov (United States)

    Kogoma, T; von Meyenburg, K

    1983-01-01

    The sdrA224 mutants of Escherichia coli K-12, capable of continued DNA replication in the absence of protein synthesis (stable DNA replication), tolerate inactivation of the dnaA gene by insertion of transposon Tn10. Furthermore, oriC, the origin of E. coli chromosome replication, can be deleted from the chromosome of sdrA mutants without loss of viability. The results suggest the presence of a second, normally repressed, initiation system for chromosome replication alternative to the 'normal' dnaA+ oriC+-dependent initiation mechanism.

  14. Mismatch repair balances leading and lagging strand DNA replication fidelity.

    Directory of Open Access Journals (Sweden)

    Scott A Lujan

    Full Text Available The two DNA strands of the nuclear genome are replicated asymmetrically using three DNA polymerases, α, δ, and ε. Current evidence suggests that DNA polymerase ε (Pol ε is the primary leading strand replicase, whereas Pols α and δ primarily perform lagging strand replication. The fact that these polymerases differ in fidelity and error specificity is interesting in light of the fact that the stability of the nuclear genome depends in part on the ability of mismatch repair (MMR to correct different mismatches generated in different contexts during replication. Here we provide the first comparison, to our knowledge, of the efficiency of MMR of leading and lagging strand replication errors. We first use the strand-biased ribonucleotide incorporation propensity of a Pol ε mutator variant to confirm that Pol ε is the primary leading strand replicase in Saccharomyces cerevisiae. We then use polymerase-specific error signatures to show that MMR efficiency in vivo strongly depends on the polymerase, the mismatch composition, and the location of the mismatch. An extreme case of variation by location is a T-T mismatch that is refractory to MMR. This mismatch is flanked by an AT-rich triplet repeat sequence that, when interrupted, restores MMR to > 95% efficiency. Thus this natural DNA sequence suppresses MMR, placing a nearby base pair at high risk of mutation due to leading strand replication infidelity. We find that, overall, MMR most efficiently corrects the most potentially deleterious errors (indels and then the most common substitution mismatches. In combination with earlier studies, the results suggest that significant differences exist in the generation and repair of Pol α, δ, and ε replication errors, but in a generally complementary manner that results in high-fidelity replication of both DNA strands of the yeast nuclear genome.

  15. Subsequent pregnancy outcome after previous foetal death

    NARCIS (Netherlands)

    Nijkamp, J. W.; Korteweg, F. J.; Holm, J. P.; Timmer, A.; Erwich, J. J. H. M.; van Pampus, M. G.

    2013-01-01

    Objective: A history of foetal death is a risk factor for complications and foetal death in subsequent pregnancies as most previous risk factors remain present and an underlying cause of death may recur. The purpose of this study was to evaluate subsequent pregnancy outcome after foetal death and to

  16. Replication of genetic associations as pseudoreplication due to shared genealogy.

    Science.gov (United States)

    Rosenberg, Noah A; Vanliere, Jenna M

    2009-09-01

    The genotypes of individuals in replicate genetic association studies have some level of correlation due to shared descent in the complete pedigree of all living humans. As a result of this genealogical sharing, replicate studies that search for genotype-phenotype associations using linkage disequilibrium between marker loci and disease-susceptibility loci can be considered as "pseudoreplicates" rather than true replicates. We examine the size of the pseudoreplication effect in association studies simulated from evolutionary models of the history of a population, evaluating the excess probability that both of a pair of studies detect a disease association compared to the probability expected under the assumption that the two studies are independent. Each of nine combinations of a demographic model and a penetrance model leads to a detectable pseudoreplication effect, suggesting that the degree of support that can be attributed to a replicated genetic association result is less than that which can be attributed to a replicated result in a context of true independence.

  17. Phenotype switching is a natural consequence of Staphylococcus aureus replication.

    Science.gov (United States)

    Edwards, Andrew M

    2012-10-01

    The pathogen Staphylococcus aureus undergoes phenotype switching in vivo from its normal colony phenotype (NCP) to a slow-growing, antibiotic-resistant small-colony-variant (SCV) phenotype that is associated with persistence in host cells and tissues. However, it is not clear whether phenotype switching is the result of a constitutive process that is selected for under certain conditions or is triggered by particular environmental stimuli. Examination of cultures of diverse S. aureus strains in the absence of selective pressure consistently revealed a small gentamicin-resistant SCV subpopulation that emerged during exponential-phase NCP growth and increased in number until NCP stationary phase. Treatment of replicating bacteria with the antibiotic gentamicin, which inhibited NCP but not SCV replication, resulted in an initial decrease in SCV numbers, demonstrating that SCVs arise as a consequence of NCP replication. However, SCV population expansion in the presence of gentamicin was reestablished by selection of phenotype-stable SCVs and subsequent SCV replication. In the absence of selective pressure, however, phenotype switching was bidirectional and occurred at a high frequency during NCP replication, resulting in SCV turnover. In summary, these data demonstrate that S. aureus phenotype switching occurs via a constitutive mechanism that generates a dynamic, antibiotic-resistant subpopulation of bacteria that can revert to the parental phenotype. The emergence of SCVs can therefore be considered a normal part of the S. aureus life cycle and provides an insurance policy against exposure to antibiotics that would otherwise eliminate the entire population.

  18. Regulation of Unperturbed DNA Replication by Ubiquitylation

    Directory of Open Access Journals (Sweden)

    Sara Priego Moreno

    2015-06-01

    Full Text Available Posttranslational modification of proteins by means of attachment of a small globular protein ubiquitin (i.e., ubiquitylation represents one of the most abundant and versatile mechanisms of protein regulation employed by eukaryotic cells. Ubiquitylation influences almost every cellular process and its key role in coordination of the DNA damage response is well established. In this review we focus, however, on the ways ubiquitylation controls the process of unperturbed DNA replication. We summarise the accumulated knowledge showing the leading role of ubiquitin driven protein degradation in setting up conditions favourable for replication origin licensing and S-phase entry. Importantly, we also present the emerging major role of ubiquitylation in coordination of the active DNA replication process: preventing re-replication, regulating the progression of DNA replication forks, chromatin re-establishment and disassembly of the replisome at the termination of replication forks.

  19. Chromosome replication and segregation in bacteria.

    Science.gov (United States)

    Reyes-Lamothe, Rodrigo; Nicolas, Emilien; Sherratt, David J

    2012-01-01

    In dividing cells, chromosome duplication once per generation must be coordinated with faithful segregation of newly replicated chromosomes and with cell growth and division. Many of the mechanistic details of bacterial replication elongation are well established. However, an understanding of the complexities of how replication initiation is controlled and coordinated with other cellular processes is emerging only slowly. In contrast to eukaryotes, in which replication and segregation are separate in time, the segregation of most newly replicated bacterial genetic loci occurs sequentially soon after replication. We compare the strategies used by chromosomes and plasmids to ensure their accurate duplication and segregation and discuss how these processes are coordinated spatially and temporally with growth and cell division. We also describe what is known about the three conserved families of ATP-binding proteins that contribute to chromosome segregation and discuss their inter-relationships in a range of disparate bacteria.

  20. On Scalability and Replicability of Smart Grid Projects—A Case Study

    Directory of Open Access Journals (Sweden)

    Lukas Sigrist

    2016-03-01

    Full Text Available This paper studies the scalability and replicability of smart grid projects. Currently, most smart grid projects are still in the R&D or demonstration phases. The full roll-out of the tested solutions requires a suitable degree of scalability and replicability to prevent project demonstrators from remaining local experimental exercises. Scalability and replicability are the preliminary requisites to perform scaling-up and replication successfully; therefore, scalability and replicability allow for or at least reduce barriers for the growth and reuse of the results of project demonstrators. The paper proposes factors that influence and condition a project’s scalability and replicability. These factors involve technical, economic, regulatory and stakeholder acceptance related aspects, and they describe requirements for scalability and replicability. In order to assess and evaluate the identified scalability and replicability factors, data has been collected from European and national smart grid projects by means of a survey, reflecting the projects’ view and results. The evaluation of the factors allows quantifying the status quo of on-going projects with respect to the scalability and replicability, i.e., they provide a feedback on to what extent projects take into account these factors and on whether the projects’ results and solutions are actually scalable and replicable.

  1. Error-prone replication bypass of the primary aflatoxin B1 DNA adduct, AFB1-N7-Gua.

    Science.gov (United States)

    Lin, Ying-Chih; Li, Liang; Makarova, Alena V; Burgers, Peter M; Stone, Michael P; Lloyd, R Stephen

    2014-06-27

    Hepatocellular carcinomas (HCCs) are the third leading cause of cancer deaths worldwide. The highest rates of early onset HCCs occur in geographical regions with high aflatoxin B1 (AFB1) exposure, concomitant with hepatitis B infection. Although the carcinogenic basis of AFB1 has been ascribed to its mutagenic effects, the mutagenic property of the primary AFB1-DNA adduct, AFB1-N7-Gua, in mammalian cells has not been studied extensively. Taking advantage of the ability to create vectors containing a site-specific DNA adduct, the mutagenic potential was determined in primate cells. This adduct was highly mutagenic following replication in COS-7 cells, with a mutation frequency of 45%. The spectrum of mutations was predominantly G to T base substitutions, a result that is consistent with previous mutation data derived from aflatoxin-associated HCCs. To assess which DNA polymerases (pol) might contribute to the mutational outcome, in vitro replication studies were performed. Unexpectedly, replicative pol δ and the error-prone translesion synthesis pol ζ were able to accurately bypass AFB1-N7-Gua. In contrast, replication bypass using pol κ was shown to occur with low fidelity and could account for the commonly detected G to T transversions.

  2. Self-replicating alphavirus RNA vaccines.

    Science.gov (United States)

    Ljungberg, Karl; Liljeström, Peter

    2015-02-01

    Recombinant nucleic acids are considered as promising next-generation vaccines. These vaccines express the native antigen upon delivery into tissue, thus mimicking live attenuated vaccines without having the risk of reversion to pathogenicity. They also stimulate the innate immune system, thus potentiating responses. Nucleic acid vaccines are easy to produce at reasonable cost and are stable. During the past years, focus has been on the use of plasmid DNA for vaccination. Now mRNA and replicon vaccines have come into focus as promising technology platforms for vaccine development. This review discusses self-replicating RNA vaccines developed from alphavirus expression vectors. These replicon vaccines can be delivered as RNA, DNA or as recombinant virus particles. All three platforms have been pre-clinically evaluated as vaccines against a number of infectious diseases and cancer. Results have been very encouraging and propelled the first human clinical trials, the results of which have been promising.

  3. Regulation of chromosomal replication in Caulobacter crescentus.

    Science.gov (United States)

    Collier, Justine

    2012-03-01

    The alpha-proteobacterium Caulobacter crescentus is characterized by its asymmetric cell division, which gives rise to a replicating stalked cell and a non-replicating swarmer cell. Thus, the initiation of chromosomal replication is tightly regulated, temporally and spatially, to ensure that it is coordinated with cell differentiation and cell cycle progression. Waves of DnaA and CtrA activities control when and where the initiation of DNA replication will take place in C. crescentus cells. The conserved DnaA protein initiates chromosomal replication by directly binding to sites within the chromosomal origin (Cori), ensuring that DNA replication starts once and only once per cell cycle. The CtrA response regulator represses the initiation of DNA replication in swarmer cells and in the swarmer compartment of pre-divisional cells, probably by competing with DnaA for binding to Cori. CtrA and DnaA are controlled by multiple redundant regulatory pathways that include DNA methylation-dependent transcriptional regulation, temporally regulated proteolysis and the targeting of regulators to specific locations within the cell. Besides being critical regulators of chromosomal replication, CtrA and DnaA are also master transcriptional regulators that control the expression of many genes, thus connecting DNA replication with other events of the C. crescentus cell cycle. Copyright © 2012 Elsevier Inc. All rights reserved.

  4. Global profiling of DNA replication timing and efficiency reveals that efficient replication/firing occurs late during S-phase in S. pombe.

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    Majid Eshaghi

    Full Text Available BACKGROUND: During S. pombe S-phase, initiation of DNA replication occurs at multiple sites (origins that are enriched with AT-rich sequences, at various times. Current studies of genome-wide DNA replication profiles have focused on the DNA replication timing and origin location. However, the replication and/or firing efficiency of the individual origins on the genomic scale remain unclear. METHODOLOGY/PRINCIPAL FINDINGS: Using the genome-wide ORF-specific DNA microarray analysis, we show that in S. pombe, individual origins fire with varying efficiencies and at different times during S-phase. The increase in DNA copy number plotted as a function of time is approximated to the near-sigmoidal model, when considering the replication start and end timings at individual loci in cells released from HU-arrest. Replication efficiencies differ from origin to origin, depending on the origin's firing efficiency. We have found that DNA replication is inefficient early in S-phase, due to inefficient firing at origins. Efficient replication occurs later, attributed to efficient but late-firing origins. Furthermore, profiles of replication timing in cds1Delta cells are abnormal, due to the failure in resuming replication at the collapsed forks. The majority of the inefficient origins, but not the efficient ones, are found to fire in cds1Delta cells after HU removal, owing to the firing at the remaining unused (inefficient origins during HU treatment. CONCLUSIONS/SIGNIFICANCE: Taken together, our results indicate that efficient DNA replication/firing occurs late in S-phase progression in cells after HU removal, due to efficient late-firing origins. Additionally, checkpoint kinase Cds1p is required for maintaining the efficient replication/firing late in S-phase. We further propose that efficient late-firing origins are essential for ensuring completion of DNA duplication by the end of S-phase.

  5. Depletion of acidic phospholipids influences chromosomal replication in Escherichia coli.

    Science.gov (United States)

    Fingland, Nicholas; Flåtten, Ingvild; Downey, Christopher D; Fossum-Raunehaug, Solveig; Skarstad, Kirsten; Crooke, Elliott

    2012-12-01

    In Escherichia coli, coordinated activation and deactivation of DnaA allows for proper timing of the initiation of chromosomal synthesis at the origin of replication (oriC) and assures initiation occurs once per cell cycle. In vitro, acidic phospholipids reactivate DnaA, and in vivo depletion of acidic phospholipids, results in growth arrest. Growth can be restored by the expression of a mutant form of DnaA, DnaA(L366K), or by oriC-independent DNA synthesis, suggesting acidic phospholipids are required for DnaA- and oriC-dependent replication. We observe here that when acidic phospholipids were depleted, replication was inhibited with a concomitant reduction of chromosomal content and cell mass prior to growth arrest. This global shutdown of biosynthetic activity was independent of the stringent response. Restoration of acidic phospholipid synthesis resulted in a resumption of DNA replication prior to restored growth, indicating a possible cell-cycle-specific growth arrest had occurred with the earlier loss of acidic phospholipids. Flow cytometry, thymidine uptake, and quantitative polymerase chain reaction data suggest that a deficiency in acidic phospholipids prolonged the time required to replicate the chromosome. We also observed that regardless of the cellular content of acidic phospholipids, expression of mutant DnaA(L366K) altered the DNA content-to-cell mass ratio.

  6. Transcription-replication collision increases recombination efficiency between plasmids.

    Science.gov (United States)

    Jialiang, Li; Feng, Chen; Zhen, Xu; Jibing, Chen; Xiang, Lv; Lingling, Zhang; Depei, Liu

    2013-11-01

    It has been proposed that the stalling of the replication forks can induce homologous recombination in several organisms, and that arrested replication forks may offer nuclease targets, thereby providing a substrate for proteins involved in double-strand repair. In this article, we constructed a plasmid with the potential for transcription-replication collision (TRC), in which DNA replication and RNA transcription occur on the same DNA template simultaneously. Theoretically, transcription will impede DNA replication and increase homologous recombination. To validate this hypothesis, another plasmid was constructed that contained a homologous sequence with the exception of some mutated sites. Co-transfection of these two plasmids into 293T cells resulted in increased recombination frequency. The ratio of these two plasmids also affected the recombination frequency. Moreover, we found high expression levels of RAD51, which indicated that the increase in the recombination rate was probably via the homologous recombination pathway. These results indicate that mutant genes in plasmids can be repaired by TRC-induced recombination.

  7. Hydrogen peroxide thermochemical oscillator as driver for primordial RNA replication.

    Science.gov (United States)

    Ball, Rowena; Brindley, John

    2014-06-06

    This paper presents and tests a previously unrecognized mechanism for driving a replicating molecular system on the prebiotic earth. It is proposed that cell-free RNA replication in the primordial soup may have been driven by self-sustained oscillatory thermochemical reactions. To test this hypothesis, a well-characterized hydrogen peroxide oscillator was chosen as the driver and complementary RNA strands with known association and melting kinetics were used as the substrate. An open flow system model for the self-consistent, coupled evolution of the temperature and concentrations in a simple autocatalytic scheme is solved numerically, and it is shown that thermochemical cycling drives replication of the RNA strands. For the (justifiably realistic) values of parameters chosen for the simulated example system, the mean amount of replicant produced at steady state is 6.56 times the input amount, given a constant supply of substrate species. The spontaneous onset of sustained thermochemical oscillations via slowly drifting parameters is demonstrated, and a scheme is given for prebiotic production of complementary RNA strands on rock surfaces.

  8. Effects of DNA replication on mRNA noise.

    Science.gov (United States)

    Peterson, Joseph R; Cole, John A; Fei, Jingyi; Ha, Taekjip; Luthey-Schulten, Zaida A

    2015-12-29

    There are several sources of fluctuations in gene expression. Here we study the effects of time-dependent DNA replication, itself a tightly controlled process, on noise in mRNA levels. Stochastic simulations of constitutive and regulated gene expression are used to analyze the time-averaged mean and variation in each case. The simulations demonstrate that to capture mRNA distributions correctly, chromosome replication must be realistically modeled. Slow relaxation of mRNA from the low copy number steady state before gene replication to the high steady state after replication is set by the transcript's half-life and contributes significantly to the shape of the mRNA distribution. Consequently both the intrinsic kinetics and the gene location play an important role in accounting for the mRNA average and variance. Exact analytic expressions for moments of the mRNA distributions that depend on the DNA copy number, gene location, cell doubling time, and the rates of transcription and degradation are derived for the case of constitutive expression and subsequently extended to provide approximate corrections for regulated expression and RNA polymerase variability. Comparisons of the simulated models and analytical expressions to experimentally measured mRNA distributions show that they better capture the physics of the system than previous theories.

  9. Inhibition of simian virus 40 DNA replication by ultraviolet light

    Energy Technology Data Exchange (ETDEWEB)

    Edenberg, H.J.

    1983-07-30

    The effects of ultraviolet light (uv) upon SV40 DNA synthesis in monkey cells were examined to determine whether replication forks were halted upon encountering lesions in the DNA, or alternatively whether lesions were rapidly bypassed. Ultraviolet light inhibits elongation of nascent DNA strands; the extent of incorporation of (/sup 3/H)deoxythymidine ((/sup 3/H)dT) into DNA decreases with increasing uv fluence. Inhibition begins within minutes of irradiation, and becomes more pronounced with increasing time after irradiation. The synthesis of form I (covalently closed) molecules is inhibited even more severely than is total incorporation: post-uv incorporation is predominantly into replication intermediates. In contrast to previous reports, we find that replication intermediates labeled after uv resemble those in unirradiated cells, and contain covalently closed parental strands. DNA strands made after uv are approximately the size of parental DNA which has been cleaved at pyrimidine dimers by a uv endonuclease, indicating that they do not extend past dimers. The hypothesis that replication forks are halted upon encountering pyrimidine dimers in the template strand is consistent with these data.

  10. The replication timing of the amplified dihydrofolate reductase genes in the Chinese hamster ovary cell line CHOC 400.

    Science.gov (United States)

    Caddle, M S; Heintz, N H

    1990-07-16

    We have examined the timing of replication of the amplified dihydrofolate reductase genes in the methotrexate-resistant Chinese hamster ovary cell line CHOC 400 using two synchronization procedures. DNA replicated in the presence of 5-bromodeoxyuridine was collected from cells of various times during the DNA synthesis phase and the extent of replication for defined sequences was determined by Southern blotting analysis of CsCl density gradient fractions. We report that under these conditions the DHFR gene replicates throughout the course of S phase in a mode similar to the bulk of the replicated genomic DNA. This contrasts with previous data that shows the non-amplified DHFR gene replicates during the first quarter of S phase. Therefore, we conclude that gene amplification alters the replication timing of the DHFR gene in CHOC 400 cells.

  11. Induced vaginal birth after previous caesarean section

    OpenAIRE

    Akylbek Tussupkaliyev; Andrey Gayday; Bibigul Karimsakova; Saule Bermagambetova; Lunara Uteniyazova; Guldana Iztleuova; Gulkhanym Kusherbayeva; Meruyert Konakbayeva; Assylzada Merekeyeva; Zamira Imangaliyeva

    2016-01-01

    Introduction The rate of operative birth by Caesarean section is constantly rising. In Kazakhstan, it reaches 27 per cent. Research data confirm that the percentage of successful vaginal births after previous Caesarean section is 50–70 per cent. How safe the induction of vaginal birth after Caesarean (VBAC) remains unclear. Methodology The studied techniques of labour induction were amniotomy of the foetal bladder with the vulsellum ramus, intravaginal administra...

  12. Replication of an empirical approach to delineate the heterogeneity of chronic unexplained fatigue

    Directory of Open Access Journals (Sweden)

    White Peter D

    2009-10-01

    Full Text Available Abstract Background Chronic fatigue syndrome (CFS is defined by self-reported symptoms. There are no diagnostic signs or laboratory markers, and the pathophysiology remains inchoate. In part, difficulties identifying and replicating biomarkers and elucidating the pathophysiology reflect the heterogeneous nature of the syndromic illness CFS. We conducted this analysis of people from defined metropolitan, urban, and rural populations to replicate our earlier empirical delineation of medically unexplained chronic fatigue and CFS into discrete endophenotypes. Both the earlier and current analyses utilized quantitative measures of functional impairment and symptoms as well as laboratory data. This study and the earlier one enrolled participants from defined populations and measured the internal milieu, which differentiates them from studies of clinic referrals that examine only clinical phenotypes. Methods This analysis evaluated 386 women identified in a population-based survey of chronic fatigue and unwellness in metropolitan, urban, and rural populations of the state of Georgia, USA. We used variables previously demonstrated to effectively delineate endophenotypes in an attempt to replicate identification of these endophenotypes. Latent class analyses were used to derive the classes, and these were compared and contrasted to those described in the previous study based in Wichita, Kansas. Results We identified five classes in the best fit analysis. Participants in Class 1 (25% were polysymptomatic, with sleep problems and depressed mood. Class 2 (24% was also polysymptomatic, with insomnia and depression, but participants were also obese with associated metabolic strain. Class 3 (20% had more selective symptoms but was equally obese with metabolic strain. Class 4 (20% and Class 5 (11% consisted of nonfatigued, less symptomatic individuals, Class 4 being older and Class 5 younger. The classes were generally validated by independent variables. People

  13. Owner reports of attention, activity, and impulsivity in dogs: a replication study

    Directory of Open Access Journals (Sweden)

    Iosif Ana-Maria

    2010-01-01

    Full Text Available Abstract Background When developing behaviour measurement tools that use third party assessments, such as parent report, it is important to demonstrate reliability of resulting scales through replication using novel cohorts. The domestic dog has been suggested as a model to investigate normal variation in attention, hyperactivity, and impulsive behaviours impaired in Attention Deficit Hyperactive Disorder (ADHD. The human ADHD Rating Scale, modified for dogs and using owner-directed surveys, was applied in a European sample. We asked whether findings would be replicated utilizing an Internet survey in a novel sample, where unassisted survey completion, participant attitudes and breeds might affect previous findings. Methods Using a slightly modified version of the prior survey, we collected responses (n = 1030, 118 breeds representing 7 breed groups primarily in the United States and Canada. This study was conducted using an Internet survey mechanism. Results Reliability analyses confirmed two scales previously identified for dogs (inattention [IA], hyperactivity-impulsivity [HA-IM]. Models including age, training status, and breed group accounted for very little variance in subscales, with no effect of gender. Conclusions The factor invariance demonstrated in these findings confirms that owner report, using this modified human questionnaire, provides dog scores according to "inattention" and "hyperactivity-impulsivity" axes. Further characterization of naturally occurring variability of attention, activity, and impulsivity in domestic dogs may provide insight into genetic backgrounds underlying behaviours impaired in attention and associated disorders.

  14. Approximating the maximum weight clique using replicator dynamics.

    Science.gov (United States)

    Bomze, I R; Pelillo, M; Stix, V

    2000-01-01

    Given an undirected graph with weights on the vertices, the maximum weight clique problem (MWCP) is to find a subset of mutually adjacent vertices (i.e., a clique) having the largest total weight. This is a generalization of the classical problem of finding the maximum cardinality clique of an unweighted graph, which arises as a special case of the MWCP when all the weights associated to the vertices are equal. The problem is known to be NP-hard for arbitrary graphs and, according to recent theoretical results, so is the problem of approximating it within a constant factor. Although there has recently been much interest around neural-network algorithms for the unweighted maximum clique problem, no effort has been directed so far toward its weighted counterpart. In this paper, we present a parallel, distributed heuristic for approximating the MWCP based on dynamics principles developed and studied in various branches of mathematical biology. The proposed framework centers around a recently introduced continuous characterization of the MWCP which generalizes an earlier remarkable result by Motzkin and Straus. This allows us to formulate the MWCP (a purely combinatorial problem) in terms of a continuous quadratic programming problem. One drawback associated with this formulation, however, is the presence of "spurious" solutions, and we present characterizations of these solutions. To avoid them we introduce a new regularized continuous formulation of the MWCP inspired by previous works on the unweighted problem, and show how this approach completely solves the problem. The continuous formulation of the MWCP naturally maps onto a parallel, distributed computational network whose dynamical behavior is governed by the so-called replicator equations. These are dynamical systems introduced in evolutionary game theory and population genetics to model evolutionary processes on a macroscopic scale.We present theoretical results which guarantee that the solutions provided by

  15. A new light on DNA replication from the inactive X chromosome.

    Science.gov (United States)

    Aladjem, Mirit I; Fu, Haiqing

    2014-06-01

    While large portions of the mammalian genome are known to replicate sequentially in a distinct, tissue-specific order, recent studies suggest that the inactive X chromosome is duplicated rapidly via random, synchronous DNA synthesis at numerous adjacent regions. The rapid duplication of the inactive X chromosome was observed in high-resolution studies visualizing DNA replication patterns in the nucleus, and by allele-specific DNA sequencing studies measuring the extent of DNA synthesis. These studies conclude that inactive X chromosomes complete replication earlier than previously thought and suggest that the strict order of DNA replication detected in the majority of genomic regions is not preserved in non-transcribed, "silent" chromatin. These observations alter current concepts about the regulation of DNA replication in non-transcribed portions of the genome in general and in the inactive X-chromosome in particular.

  16. Genome-wide studies highlight indirect links between human replication origins and gene regulation.

    Science.gov (United States)

    Cadoret, Jean-Charles; Meisch, Françoise; Hassan-Zadeh, Vahideh; Luyten, Isabelle; Guillet, Claire; Duret, Laurent; Quesneville, Hadi; Prioleau, Marie-Noëlle

    2008-10-14

    To get insights into the regulation of replication initiation, we systematically mapped replication origins along 1% of the human genome in HeLa cells. We identified 283 origins, 10 times more than previously known. Origin density is strongly correlated with genomic landscapes, with clusters of closely spaced origins in GC-rich regions and no origins in large GC-poor regions. Origin sequences are evolutionarily conserved, and half of them map within or near CpG islands. Most of the origins overlap transcriptional regulatory elements, providing further evidence of a connection with gene regulation. Moreover, we identify c-JUN and c-FOS as important regulators of origin selection. Half of the identified replication initiation sites do not have an open chromatin configuration, showing the absence of a direct link with gene regulation. Replication timing analyses coupled with our origin mapping suggest that a relatively strict origin-timing program regulates the replication of the human genome.

  17. Electron microscopic analysis of rotavirus assembly-replication intermediates

    Energy Technology Data Exchange (ETDEWEB)

    Boudreaux, Crystal E.; Kelly, Deborah F. [Virginia Tech Carilion School of Medicine and Research Institute, Roanoke, VA (United States); McDonald, Sarah M., E-mail: mcdonaldsa@vtc.vt.edu [Virginia Tech Carilion School of Medicine and Research Institute, Roanoke, VA (United States); Department of Biomedical Sciences and Pathobiology, Virginia—Maryland Regional College of Veterinary Medicine, Blacksburg, VA (United States)

    2015-03-15

    Rotaviruses (RVs) replicate their segmented, double-stranded RNA genomes in tandem with early virion assembly. In this study, we sought to gain insight into the ultrastructure of RV assembly-replication intermediates (RIs) using transmission electron microscopy (EM). Specifically, we examined a replicase-competent, subcellular fraction that contains all known RV RIs. Three never-before-seen complexes were visualized in this fraction. Using in vitro reconstitution, we showed that ~15-nm doughnut-shaped proteins in strings were nonstructural protein 2 (NSP2) bound to viral RNA transcripts. Moreover, using immunoaffinity-capture EM, we revealed that ~20-nm pebble-shaped complexes contain the viral RNA polymerase (VP1) and RNA capping enzyme (VP3). Finally, using a gel purification method, we demonstrated that ~30–70-nm electron-dense, particle-shaped complexes represent replicase-competent core RIs, containing VP1, VP3, and NSP2 as well as capsid proteins VP2 and VP6. The results of this study raise new questions about the interactions among viral proteins and RNA during the concerted assembly–replicase process. - Highlights: • Rotaviruses replicate their genomes in tandem with early virion assembly. • Little is known about rotavirus assembly-replication intermediates. • Assembly-replication intermediates were imaged using electron microscopy.

  18. Replicator dynamics for optional public good games

    DEFF Research Database (Denmark)

    Hauert, C.; De Monte, Silvia; Hofbauer, J.;

    2002-01-01

    The public goods game represents a straightforward generalization of the prisoner's dilemma to an arbitrary number of players. Since the dominant strategy is to defect, both classical and evolutionary game theory predict the asocial outcome that no player contributes to the public goods....... In contrast to the compulsory public goods game, optional participation provides a natural way to avoid deadlocks in the state of mutual defection. The three resulting strategies-collaboration or defection in the public goods game, as well as not joining at all-are studied by means of a replicator dynamics...... participation makes cooperation feasible. But for each strategy, the average payoff value remains equal to the earnings of those not participating in the public goods game....

  19. Suggestibility and negative priming: two replication studies.

    Science.gov (United States)

    David, Daniel; Brown, Richard J

    2002-07-01

    Research suggests that inhibiting the effect of irrelevant stimuli on subsequent thought and action (cognitive inhibition) may be an important component of suggestibility. Two small correlation studies were conducted to address the relationship between different aspects of suggestibility and individual differences in cognitive inhibition, operationalized as the degree of negative priming generated by to-be-ignored stimuli in a semantic categorization task. The first study found significant positive correlations between negative priming, hypnotic suggestibility, and creative imagination; a significant negative correlation was obtained between negative priming and interrogative suggestibility, demonstrating the discriminant validity of the study results. The second study replicated the correlation between negative priming and hypnotic suggestibility, using a different suggestibility measurement procedure that assessed subjective experience and hypnotic involuntariness as well as objective responses to suggestions. These studies support the notion that the ability to engage in cognitive inhibition may be an important component of hypnotic responsivity and maybe of other forms of suggestibility.

  20. Temperamental profiles and language development: a replication and an extension.

    Science.gov (United States)

    Garello, Valentina; Viterbori, Paola; Usai, M Carmen

    2012-02-01

    Individual differences in child temperament are associated with individual differences in language development. The present study examined the relationship between temperament and language ability in 109 twenty-four- to 30-month-old children. Parents and day-care teachers completed two questionnaires: the Primo Vocabolario del Bambino (Caselli & Casadio, 1995) and the Questionari Italiani del Temperamento (Axia, 2002). Researchers administered the First Language Test (Axia, 1993) to assess productive and receptive language in each child. Replicating previous research (Usai, Garello, & Viterbori, 2009), day-care teachers identified three temperamental profiles: most of the children fit into the first profile, typical of the Italian population; another profile was made up of easily distractible and not very persistent children, with a poor capacity to modulate motor activity; and the third profile of children were inhibited in new situations. A relationship was found between temperament assessed by day-care teachers and different levels of linguistic competence. In particular, the groups of "inattentive" and "inhibited" children showed poorer lexical and morphological abilities and a more immature vocabulary, characterised by the presence of more primitive components of the lexical repertory compared to the group of "typical" children. Unlike the results from day-care teachers, temperament questionnaires completed by parents revealed a 4-cluster-solution. Also, for parents, the "typical" profile is characterised by the largest vocabulary (productive and receptive) and the most mature semantic production.

  1. DNA methyltransferases are required to induce heterochromatic re-replication in Arabidopsis.

    Directory of Open Access Journals (Sweden)

    Hume Stroud

    2012-07-01

    Full Text Available The relationship between epigenetic marks on chromatin and the regulation of DNA replication is poorly understood. Mutations of the H3K27 methyltransferase genes, Arabidopsis trithorax-related protein5 (ATXR5 and ATXR6, result in re-replication (repeated origin firing within the same cell cycle. Here we show that mutations that reduce DNA methylation act to suppress the re-replication phenotype of atxr5 atxr6 mutants. This suggests that DNA methylation, a mark enriched at the same heterochromatic regions that re-replicate in atxr5/6 mutants, is required for aberrant re-replication. In contrast, RNA sequencing analyses suggest that ATXR5/6 and DNA methylation cooperatively transcriptionally silence transposable elements (TEs. Hence our results suggest a complex relationship between ATXR5/6 and DNA methylation in the regulation of DNA replication and transcription of TEs.

  2. Choosing a suitable method for the identification of replication origins in microbial genomes

    Directory of Open Access Journals (Sweden)

    Chengcheng eSong

    2015-09-01

    Full Text Available As the replication of genomic DNA is arguably the most important task performed by a cell and given that it is controlled at the initiation stage, the events that occur at the replication origin play a central role in the cell cycle. Making sense of DNA replication origins is important for improving our capacity to study cellular processes and functions in the regulation of gene expression, genome integrity in much finer detail. Thus, clearly comprehending the positions and sequences of replication origins which are fundamental to chromosome organization and duplication is the first priority of all. In view of such important roles of replication origins, tremendous work has been aimed at identifying and testing the specificity of replication origins. A number of computational tools based on various skew types have been developed to predict replication origins. Using various in silico approaches such as Ori-Finder 2, and databases such as DoriC and oriDB, researchers have predicted the locations of replication origins sites for thousands bacterial chromosomes and archaeal genomes. Based on the predicted results, we should choose an effective method for identifying and confirming the interactions at origins of replication. Here we describe the main existing experimental methods that aimed to determine the replication origin regions and list some of the many the practical applications of these methods.

  3. Initiation of DNA replication from non-canonical sites on an origin-depleted chromosome.

    Directory of Open Access Journals (Sweden)

    Naomi L Bogenschutz

    Full Text Available Eukaryotic DNA replication initiates from multiple sites on each chromosome called replication origins (origins. In the budding yeast Saccharomyces cerevisiae, origins are defined at discrete sites. Regular spacing and diverse firing characteristics of origins are thought to be required for efficient completion of replication, especially in the presence of replication stress. However, a S. cerevisiae chromosome III harboring multiple origin deletions has been reported to replicate relatively normally, and yet how an origin-deficient chromosome could accomplish successful replication remains unknown. To address this issue, we deleted seven well-characterized origins from chromosome VI, and found that these deletions do not cause gross growth defects even in the presence of replication inhibitors. We demonstrated that the origin deletions do cause a strong decrease in the binding of the origin recognition complex. Unexpectedly, replication profiling of this chromosome showed that DNA replication initiates from non-canonical loci around deleted origins in yeast. These results suggest that replication initiation can be unexpectedly flexible in this organism.

  4. Proficient Replication of the Yeast Genome by a Viral DNA Polymerase.

    Science.gov (United States)

    Stodola, Joseph L; Stith, Carrie M; Burgers, Peter M

    2016-05-27

    DNA replication in eukaryotic cells requires minimally three B-family DNA polymerases: Pol α, Pol δ, and Pol ϵ. Pol δ replicates and matures Okazaki fragments on the lagging strand of the replication fork. Saccharomyces cerevisiae Pol δ is a three-subunit enzyme (Pol3-Pol31-Pol32). A small C-terminal domain of the catalytic subunit Pol3 carries both iron-sulfur cluster and zinc-binding motifs, which mediate interactions with Pol31, and processive replication with the replication clamp proliferating cell nuclear antigen (PCNA), respectively. We show that the entire N-terminal domain of Pol3, containing polymerase and proofreading activities, could be effectively replaced by those from bacteriophage RB69, and could carry out chromosomal DNA replication in yeast with remarkable high fidelity, provided that adaptive mutations in the replication clamp PCNA were introduced. This result is consistent with the model that all essential interactions for DNA replication in yeast are mediated through the small C-terminal domain of Pol3. The chimeric polymerase carries out processive replication with PCNA in vitro; however, in yeast, it requires an increased involvement of the mutagenic translesion DNA polymerase ζ during DNA replication.

  5. Surface micro topography replication in injection moulding

    DEFF Research Database (Denmark)

    Arlø, Uffe Rolf

    of the mechanisms controlling topography replication. Surface micro topography replication in injection moulding depends on the main elements of  Process conditions  Plastic material  Mould topography In this work, the process conditions is the main factor considered, but the impact of plastic material...

  6. Replication and Robustness in Developmental Research

    Science.gov (United States)

    Duncan, Greg J.; Engel, Mimi; Claessens, Amy; Dowsett, Chantelle J.

    2014-01-01

    Replications and robustness checks are key elements of the scientific method and a staple in many disciplines. However, leading journals in developmental psychology rarely include explicit replications of prior research conducted by different investigators, and few require authors to establish in their articles or online appendices that their key…

  7. Replication and Robustness in Developmental Research

    Science.gov (United States)

    Duncan, Greg J.; Engel, Mimi; Claessens, Amy; Dowsett, Chantelle J.

    2014-01-01

    Replications and robustness checks are key elements of the scientific method and a staple in many disciplines. However, leading journals in developmental psychology rarely include explicit replications of prior research conducted by different investigators, and few require authors to establish in their articles or online appendices that their key…

  8. Using Replication Projects in Teaching Research Methods

    Science.gov (United States)

    Standing, Lionel G.; Grenier, Manuel; Lane, Erica A.; Roberts, Meigan S.; Sykes, Sarah J.

    2014-01-01

    It is suggested that replication projects may be valuable in teaching research methods, and also address the current need in psychology for more independent verification of published studies. Their use in an undergraduate methods course is described, involving student teams who performed direct replications of four well-known experiments, yielding…

  9. How frog embryos replicate their DNA reliably

    Science.gov (United States)

    Bechhoefer, John; Marshall, Brandon

    2007-03-01

    Frog embryos contain three billion base pairs of DNA. In early embryos (cycles 2-12), DNA replication is extremely rapid, about 20 min., and the entire cell cycle lasts only 25 min., meaning that mitosis (cell division) takes place in about 5 min. In this stripped-down cell cycle, there are no efficient checkpoints to prevent the cell from dividing before its DNA has finished replication - a disastrous scenario. Even worse, the many origins of replication are laid down stochastically and are also initiated stochastically throughout the replication process. Despite the very tight time constraints and despite the randomness introduced by origin stochasticity, replication is extremely reliable, with cell division failing no more than once in 10,000 tries. We discuss a recent model of DNA replication that is drawn from condensed-matter theories of 1d nucleation and growth. Using our model, we discuss different strategies of replication: should one initiate all origins as early as possible, or is it better to hold back and initiate some later on? Using concepts from extreme-value statistics, we derive the distribution of replication times given a particular scenario for the initiation of origins. We show that the experimentally observed initiation strategy for frog embryos meets the reliability constraint and is close to the one that requires the fewest resources of a cell.

  10. Mammalian RAD52 Functions in Break-Induced Replication Repair of Collapsed DNA Replication Forks

    DEFF Research Database (Denmark)

    Sotiriou, Sotirios K; Kamileri, Irene; Lugli, Natalia

    2016-01-01

    Human cancers are characterized by the presence of oncogene-induced DNA replication stress (DRS), making them dependent on repair pathways such as break-induced replication (BIR) for damaged DNA replication forks. To better understand BIR, we performed a targeted siRNA screen for genes whose depl...

  11. A Replication Protocol for Real Time database System

    Directory of Open Access Journals (Sweden)

    Ashish Srivastava

    2012-06-01

    Full Text Available Database replication protocols for real time system based on a certification approach are usually the best ones for achieving good performance. The weak voting approach achieves a slightly longer transaction completion time, but with a lower abortion rate. So, both techniques can be considered as the best ones for replication when performance is a must, and both of them take advantage of the properties provided by atomic broadcast. We propose a new database replication strategy that shares many characteristics with such previous strategies. It is also based on totally ordering the application of writesets, using only an unordered reliable broadcast, instead of an atomic broadcast. Additionally, the writesets of transactions that are aborted in the final validation phase along with verification phase incorporated in the new system are not broadcast in our strategy rather than only validation phase. Thus, this new approach certainly reducesc the communication traffic and also achieves a good transaction response time (even shorter than those previous strategies associated with only validation phase in some system configurations.

  12. Delay Scheduling Based Replication Scheme for Hadoop Distributed File System

    Directory of Open Access Journals (Sweden)

    S. Suresh

    2015-03-01

    Full Text Available The data generated and processed by modern computing systems burgeon rapidly. MapReduce is an important programming model for large scale data intensive applications. Hadoop is a popular open source implementation of MapReduce and Google File System (GFS. The scalability and fault-tolerance feature of Hadoop makes it as a standard for BigData processing. Hadoop uses Hadoop Distributed File System (HDFS for storing data. Data reliability and faulttolerance is achieved through replication in HDFS. In this paper, a new technique called Delay Scheduling Based Replication Algorithm (DSBRA is proposed to identify and replicate (dereplicate the popular (unpopular files/blocks in HDFS based on the information collected from the scheduler. Experimental results show that, the proposed method achieves 13% and 7% improvements in response time and locality over existing algorithms respectively.

  13. Replication of Psycholinguistic Experiments and the Resolution of Inconsistencies.

    Science.gov (United States)

    Rákosi, Csilla

    2017-05-05

    Non-exact replications are regarded as effective tools of problem solving in psycholinguistic research because they lead to more plausible experimental results; however, they are also ineffective tools of problem solving because they trigger cumulative contradictions among different replications of an experiment. This paper intends to resolve this paradox by putting forward a metatheoretical model that clarifies the criteria with the help of which various aspects of the effectiveness of the problem solving process can be differentiated and evaluated. The key point is the reconstruction of the relationship between original experiments and their non-exact replications by introducing the concept of 'experimental complex' and analysing the problem solving strategies that the researchers apply. The workability of the proposed metatheoretical model is illustrated with the help of three case studies.

  14. Phosphorylation of hepatitis C virus RNA polymerases ser29 and ser42 by protein kinase C-related kinase 2 regulates viral RNA replication.

    Science.gov (United States)

    Han, Song-Hee; Kim, Seong-Jun; Kim, Eun-Jung; Kim, Tae-Eun; Moon, Jae-Su; Kim, Geon-Woo; Lee, Seung-Hoon; Cho, Kun; Yoo, Jong Shin; Son, Woo Sung; Rhee, Jin-Kyu; Han, Seung Hyun; Oh, Jong-Won

    2014-10-01

    Hepatitis C virus (HCV) nonstructural protein 5B (NS5B), an RNA-dependent RNA polymerase (RdRp), is the key enzyme for HCV RNA replication. We previously showed that HCV RdRp is phosphorylated by protein kinase C-related kinase 2 (PRK2). In the present study, we used biochemical and reverse-genetics approaches to demonstrate that HCV NS5B phosphorylation is crucial for viral RNA replication in cell culture. Two-dimensional phosphoamino acid analysis revealed that PRK2 phosphorylates NS5B exclusively at its serine residues in vitro and in vivo. Using in vitro kinase assays and mass spectrometry, we identified two phosphorylation sites, Ser29 and Ser42, in the Δ1 finger loop region that interacts with the thumb subdomain of NS5B. Colony-forming assays using drug-selectable HCV subgenomic RNA replicons revealed that preventing phosphorylation by Ala substitution at either Ser29 or Ser42 impairs HCV RNA replication. Furthermore, reverse-genetics studies using HCV infectious clones encoding phosphorylation-defective NS5B confirmed the crucial role of these PRK2 phosphorylation sites in viral RNA replication. Molecular-modeling studies predicted that the phosphorylation of NS5B stabilizes the interactions between its Δ1 loop and thumb subdomain, which are required for the formation of the closed conformation of NS5B known to be important for de novo RNA synthesis. Collectively, our results provide evidence that HCV NS5B phosphorylation has a positive regulatory role in HCV RNA replication. While the role of RNA-dependent RNA polymerases (RdRps) in viral RNA replication is clear, little is known about their functional regulation by phosphorylation. In this study, we addressed several important questions about the function and structure of phosphorylated hepatitis C virus (HCV) nonstructural protein 5B (NS5B). Reverse-genetics studies with HCV replicons encoding phosphorylation-defective NS5B mutants and analysis of their RdRp activities revealed previously unidentified

  15. Rescue from replication stress during mitosis.

    Science.gov (United States)

    Fragkos, Michalis; Naim, Valeria

    2017-04-03

    Genomic instability is a hallmark of cancer and a common feature of human disorders, characterized by growth defects, neurodegeneration, cancer predisposition, and aging. Recent evidence has shown that DNA replication stress is a major driver of genomic instability and tumorigenesis. Cells can undergo mitosis with under-replicated DNA or unresolved DNA structures, and specific pathways are dedicated to resolving these structures during mitosis, suggesting that mitotic rescue from replication stress (MRRS) is a key process influencing genome stability and cellular homeostasis. Deregulation of MRRS following oncogene activation or loss-of-function of caretaker genes may be the cause of chromosomal aberrations that promote cancer initiation and progression. In this review, we discuss the causes and consequences of replication stress, focusing on its persistence in mitosis as well as the mechanisms and factors involved in its resolution, and the potential impact of incomplete replication or aberrant MRRS on tumorigenesis, aging and disease.

  16. Power and precision of replicated helicopter surveys in mixed bushveld

    Directory of Open Access Journals (Sweden)

    B.K. Reilly

    1998-01-01

    Full Text Available It is well known that aerial game counts in South Africa are often applied in a non-standardised, unreplicated fashion. They contribute to poor management decisions based on their results as they may be subject to large statistical Type I and II errors. Replicate counts of large herbivores were conducted in a 8 500 ha sample site in the Loskop Dam Nature Reserve in July 1991. These data were used to estimate precision of the counts and estimate statistical power to detect population changes for different combinations of replications and significance levels.

  17. Initiation of Replication in Escherichia coli

    DEFF Research Database (Denmark)

    Frimodt-Møller, Jakob

    of initiation, which leads to hyperinitiation, results in double-strand breaks when replication forks encounters single-stranded DNA lesions generated while removing oxidized bases, primarily 8-oxoG, from the DNA. Thus, the number of replication forks can only increase when ROS formation is reduced or when...... that the cell needs a copy of both DARS1 and DARS2 for proper regulation of initiation; i.e. DARS1 is a poor replacement for DARS2 and vice versa. Last we suggest that transcription has a negative effect of the activity of the non-coding regions....

  18. Inhibition of hepatitis B virus replication by helper dependent adenoviral vectors expressing artificial anti-HBV pri-miRs from a liver-specific promoter.

    Science.gov (United States)

    Mowa, Mohube Betty; Crowther, Carol; Ely, Abdullah; Arbuthnot, Patrick

    2014-01-01

    Research on applying RNA interference (RNAi) to counter HBV replication has led to identification of potential therapeutic sequences. However, before clinical application liver-specific expression and efficient delivery of these sequences remain an important objective. We recently reported short-term inhibition of HBV replication in vivo by using helper dependent adenoviral vectors (HD Ads) expressing anti-HBV sequences from a constitutively active cytomegalovirus (CMV) promoter. To develop the use of liver-specific transcription regulatory elements we investigated the utility of the murine transthyretin (MTTR) promoter for expression of anti-HBV primary microRNAs (pri-miRs). HD Ads containing MTTR promoter effected superior expression of anti-HBV pri-miRs in mice compared to HD Ads containing the CMV promoter. MTTR-containing HD Ads resulted in HBV replication knockdown of up to 94% in mice. HD Ads expressing trimeric anti-HBV pri-miRs silenced HBV replication for 5 weeks. We previously showed that the product of the codelivered lacZ gene induces an immune response, and the duration of HBV silencing in vivo is likely to be attenuated by this effect. Nevertheless, expression of anti-HBV pri-miRs from MTTR promoter is well suited to countering HBV replication and development of HD Ads through attenuation of their immunostimulatory effects should advance their clinical utility.

  19. Transforming growth factor-β1 suppresses hepatitis B virus replication by the reduction of hepatocyte nuclear factor-4α expression.

    Directory of Open Access Journals (Sweden)

    Ming-Hsiang Hong

    Full Text Available Several studies have demonstrated that cytokine-mediated noncytopathic suppression of hepatitis B virus (HBV replication may provide an alternative therapeutic strategy for the treatment of chronic hepatitis B infection. In our previous study, we showed that transforming growth factor-beta1 (TGF-β1 could effectively suppress HBV replication at physiological concentrations. Here, we provide more evidence that TGF-β1 specifically diminishes HBV core promoter activity, which subsequently results in a reduction in the level of viral pregenomic RNA (pgRNA, core protein (HBc, nucleocapsid, and consequently suppresses HBV replication. The hepatocyte nuclear factor 4alpha (HNF-4α binding element(s within the HBV core promoter region was characterized to be responsive for the inhibitory effect of TGF-β1 on HBV regulation. Furthermore, we found that TGF-β1 treatment significantly repressed HNF-4α expression at both mRNA and protein levels. We demonstrated that RNAi-mediated depletion of HNF-4α was sufficient to reduce HBc synthesis as TGF-β1 did. Prevention of HNF-4α degradation by treating with proteasome inhibitor MG132 also prevented the inhibitory effect of TGF-β1. Finally, we confirmed that HBV replication could be rescued by ectopic expression of HNF-4α in TGF-β1-treated cells. Our data clarify the mechanism by which TGF-β1 suppresses HBV replication, primarily through modulating the expression of HNF-4α gene.

  20. Implications of “too good to be true” for replication, theoretical claims, and experimental design: An example using prominent studies of racial bias

    Directory of Open Access Journals (Sweden)

    Greg Francis

    2016-09-01

    Full Text Available In response to concerns about the validity of empirical findings in psychology, some scientists use replication studies as a way to validate good science and to identify poor science. Such efforts are resource intensive and are sometimes controversial (with accusations of researcher incompetence when a replication fails to show a previous result. An alternative approach is to examine the statistical properties of the reported literature to identify some cases of poor science. This review discusses some details of this process for prominent findings about racial bias, where a set of studies seems too good to be true. This kind of analysis is based on the original studies, so it avoids criticism from the original authors about the validity of replication studies. The analysis is also much easier to perform than a new empirical study. A variation of the analysis can also be used to explore whether it makes sense to run a replication study. As demonstrated here, there are situations where the existing data suggest that a direct replication of a set of studies is not worth the effort. Such a conclusion should motivate scientists to generate alternative experimental designs that better test theoretical ideas.

  1. Human immunodeficiency virus type 1-mediated syncytium formation is compatible with adenovirus replication and facilitates efficient dispersion of viral gene products and de novo-synthesized virus particles.

    Science.gov (United States)

    Li, H; Haviv, Y S; Derdeyn, C A; Lam, J; Coolidge, C; Hunter, E; Curiel, D T; Blackwell, J L

    2001-12-10

    Conditionally replicative adenovirus (CRAd) vectors are designed for specific oncolytic replication in tumor tissues with concomitant sparing of normal cells. As such, CRAds offer an unprecedented level of anticancer potential for malignancies that have been refractory to previous cancer gene therapy interventions. CRAd efficacy may, however, be compromised by inefficient dispersion of the replicating vector within the tumor tissue. To address this issue, we evaluated the utility of a fusogenic membrane glycoprotein (FMG), which induces the fusion of neighboring cellular membranes to form multinucleated syncytia. We hypothesized that the FMG-mediated syncytia would facilitate dispersion of the adenovirus (Ad) gene products and viral progeny. To test this, human immunodeficiency virus type 1 (HIV-1) envelope glycoproteins, which induce syncytia in the presence of CD4+ target cells, were expressed by an Ad (Ad5HIVenv) in permissive (CD4-positive) and nonpermissive (CD4-negative) cell lines. After validating this Ad-FMG model, the efficiency of Ad replication in the presence or absence of syncytia was evaluated. The results demonstrated that syncytium formation was compatible with Ad replication and dramatically increased the dispersion of virus gene products within the cytoplasm of the syncytia as well as viral particles in the nuclei of the syncytial mass. Moreover, progeny virions were released more efficiently from syncytia compared with nonsyncytial cells. These data demonstrate the utility of FMGs as a dispersion agent and suggest that FMGs can improve the efficacy of CRAd gene therapy.

  2. Inhibition of Hepatitis B Virus Replication by Helper Dependent Adenoviral Vectors Expressing Artificial Anti-HBV Pri-miRs from a Liver-Specific Promoter

    Directory of Open Access Journals (Sweden)

    Mohube Betty Mowa

    2014-01-01

    Full Text Available Research on applying RNA interference (RNAi to counter HBV replication has led to identification of potential therapeutic sequences. However, before clinical application liver-specific expression and efficient delivery of these sequences remain an important objective. We recently reported short-term inhibition of HBV replication in vivo by using helper dependent adenoviral vectors (HD Ads expressing anti-HBV sequences from a constitutively active cytomegalovirus (CMV promoter. To develop the use of liver-specific transcription regulatory elements we investigated the utility of the murine transthyretin (MTTR promoter for expression of anti-HBV primary microRNAs (pri-miRs. HD Ads containing MTTR promoter effected superior expression of anti-HBV pri-miRs in mice compared to HD Ads containing the CMV promoter. MTTR-containing HD Ads resulted in HBV replication knockdown of up to 94% in mice. HD Ads expressing trimeric anti-HBV pri-miRs silenced HBV replication for 5 weeks. We previously showed that the product of the codelivered lacZ gene induces an immune response, and the duration of HBV silencing in vivo is likely to be attenuated by this effect. Nevertheless, expression of anti-HBV pri-miRs from MTTR promoter is well suited to countering HBV replication and development of HD Ads through attenuation of their immunostimulatory effects should advance their clinical utility.

  3. Role of preterminal protein processing in adenovirus replication.

    Science.gov (United States)

    Webster, A; Leith, I R; Nicholson, J; Hounsell, J; Hay, R T

    1997-09-01

    Preterminal protein (pTP), the protein primer for adenovirus DNA replication, is processed at two sites by the virus-encoded protease to yield mature terminal protein (TP). Here we demonstrate that processing to TP, via an intermediate (iTP), is conserved in all serotypes sequenced to date; and in determining the sites cleaved in Ad4 pTP, we extend the previously published substrate specificity of human adenovirus proteases to include a glutamine residue at P4. Furthermore, using monoclonal antibodies raised against pTP, we show that processing to iTP and TP are temporally separated in the infectious cycle, with processing to iTP taking place outside the virus particles. In vitro and in vivo studies of viral DNA replication reveal that iTP can act as a template for initiation and elongation and argue against a role for virus-encoded protease in switching off DNA replication. Virus DNA with TP attached to its 5' end (TP-DNA) has been studied extensively in in vitro DNA replication assays. Given that in vivo pTP-DNA, not TP-DNA, is the template for all but the first round of replication, the two templates were compared in vitro and shown to have different properties. Immunofluorescence studies suggest that a region spanning the TP cleavage site is involved in defining the subnuclear localization of pTP. Therefore, a likely role for the processing of pTP-DNA is to create a distinct template for early transcription (TP-DNA), while the terminal protein moiety, be it TP or pTP, serves to guide the template to the appropriate subcellular location through the course of infection.

  4. Investigation of previously implicated genetic variants in chronic tic disorders

    DEFF Research Database (Denmark)

    Abdulkadir, Mohamed; Londono, Douglas; Gordon, Derek

    2017-01-01

    Genetic studies in Tourette syndrome (TS) are characterized by scattered and poorly replicated findings. We aimed to replicate findings from candidate gene and genome-wide association studies (GWAS). Our cohort included 465 probands with chronic tic disorder (93% TS) and both parents from 412 fam...

  5. Replication of urban innovations - prioritization of strategies for the replication of Dhaka's community-based decentralized composting model.

    Science.gov (United States)

    Yedla, Sudhakar

    2012-01-01

    Dhaka's community-based decentralized composting (DCDC) is a successful demonstration of solid waste management by adopting low-cost technology, local resources community participation and partnerships among the various actors involved. This paper attempts to understand the model, necessary conditions, strategies and their priorities to replicate DCDC in the other developing cities of Asia. Thirteen strategies required for its replication are identified and assessed based on various criteria, namely transferability, longevity, economic viability, adaptation and also overall replication. Priority setting by multi-criteria analysis by applying analytic hierarchy process revealed that immediate transferability without long-term and economic viability consideration is not advisable as this would result in unsustainable replication of DCDC. Based on the analysis, measures to ensure the product quality control; partnership among stakeholders (public-private-community); strategies to achieve better involvement of the private sector in solid waste management (entrepreneurship in approach); simple and low-cost technology; and strategies to provide an effective interface among the complementing sectors are identified as important strategies for its replication.

  6. A New Replication Norm for Psychology

    Directory of Open Access Journals (Sweden)

    Etienne P LeBel

    2015-10-01

    Full Text Available In recent years, there has been a growing concern regarding the replicability of findings in psychology, including a mounting number of prominent findings that have failed to replicate via high-powered independent replication attempts. In the face of this replicability “crisis of confidence”, several initiatives have been implemented to increase the reliability of empirical findings. In the current article, I propose a new replication norm that aims to further boost the dependability of findings in psychology. Paralleling the extant social norm that researchers should peer review about three times as many articles that they themselves publish per year, the new replication norm states that researchers should aim to independently replicate important findings in their own research areas in proportion to the number of original studies they themselves publish per year (e.g., a 4:1 original-to-replication studies ratio. I argue this simple approach could significantly advance our science by increasing the reliability and cumulative nature of our empirical knowledge base, accelerating our theoretical understanding of psychological phenomena, instilling a focus on quality rather than quantity, and by facilitating our transformation toward a research culture where executing and reporting independent direct replications is viewed as an ordinary part of the research process. To help promote the new norm, I delineate (1 how each of the major constituencies of the research process (i.e., funders, journals, professional societies, departments, and individual researchers can incentivize replications and promote the new norm and (2 any obstacles each constituency faces in supporting the new norm.

  7. Can previous learning alter future plasticity mechanisms?

    Science.gov (United States)

    Crestani, Ana Paula; Quillfeldt, Jorge Alberto

    2016-02-01

    The dynamic processes related to mnemonic plasticity have been extensively researched in the last decades. More recently, studies have attracted attention because they show an unusual plasticity mechanism that is independent of the receptor most usually related to first-time learning--that is, memory acquisition-the NMDA receptor. An interesting feature of this type of learning is that a previous experience may cause modifications in the plasticity mechanism of a subsequent learning, suggesting that prior experience in a very similar task triggers a memory acquisition process that does not depend on NMDARs. The intracellular molecular cascades necessary to assist the learning process seem to depend on the activation of hippocampal CP-AMPARs. Moreover, most of these studies were performed on hippocampus-dependent tasks, even though other brain areas, such as the basolateral amygdala, also display NMDAR-independent learning.

  8. Vocabulary Development in European Portuguese: A Replication Study Using the Language Development Survey.

    Science.gov (United States)

    Rescorla, Leslie; Nyame, Josephine; Dias, Pedro

    2016-12-01

    Our objective was to replicate previous cross-linguistic findings by comparing Portuguese and U.S. children with respect to (a) effects of language, gender, and age on vocabulary size; (b) lexical composition; and (c) late talking. We used the Language Development Survey (LDS; Rescorla, 1989) with children (18-35 months) learning European Portuguese (n = 181) and English (n = 206). In both languages, girls had higher vocabulary scores than boys and vocabulary scores increased with age. Portuguese LDS scores were significantly lower than English scores, but the effect size was small. Cross-linguistic concordance of percentage use scores yielded a Q correlation of .50, with 64 of the "top 100" words being exact matches. Cross-linguistic concordance was highest for the youngest age group. In both languages, vocabulary composition in late talkers (children ≥ 24 months with < 50 words) was highly correlated with composition in vocabulary size-matched younger children. Results replicated previous Greek, Korean, and Italian LDS studies. The early lexicons of typical talkers and late talkers contained many of the same words, indicating considerable universality and suggesting good targets for clinical intervention.

  9. Hope and the interpersonal-psychological theory of suicidal behavior: replication and extension of prior findings.

    Science.gov (United States)

    Anestis, Michael D; Moberg, Fallon B; Arnau, Randolph C

    2014-04-01

    The interpersonal-psychological theory of suicidal behavior (IPTS; Joiner, 2005) posits that suicidal behavior occurs when an individual has a desire for death (due to the combination of perceived burdensomeness and thwarted belongingness) in addition to an acquired capacity for suicide, which is present when the individual has a low fear of death and high pain tolerance. Previous research has demonstrated an expected negative relation between trait hope and perceived burdensomeness and thwarted belongingness, as well as a more perplexing finding that hope is positively associated with the acquired capability. In a sample of 230 college students, measures of the three components of the IPTS were administered, along with measures of hope, depression, and painful and/or provocative events. Hierarchical regression analyses replicated the previously found associations between hope and burdensomeness and belongingness while controlling for depression and demographic variables. The positive association between hope and acquired capacity was also replicated, but a mediation analysis demonstrated that the effect was statistically accounted for by distress tolerance. The results further support the incremental validity of hope as a consideration in suicide risk assessments and suggest that hope may serve as a protective factor with respect to suicidal desire.

  10. Inhibitory effect of interferon-gamma on adenovirus replication and late transcription.

    Science.gov (United States)

    Mistchenko, A S; Diez, R A; Falcoff, R

    1989-06-15

    We have previously shown that human interferon-gamma inhibited adenovirus multiplication in vitro in a dose-dependent fashion. This action was previous to capsid proteins synthesis and did not involve virus adsorption nor penetration. In this report we have analysed viral mRNA levels at early (7 hr post infection (p.i.)) or late (20 hr p.i.) times, as well as DNA replication in Wish cells pretreated with interferon-gamma and infected with adenovirus 5. Controls included untreated cells as well as cells treated with interferon-alpha, to which adenovirus are reported to be resistant. Transcription of adenovirus regions E1, E4, L1 and L2 has been analysed by Northern blot. Adenovirus DNA replication was determined by DNA-DNA hybridization with total adenovirus 2 DNA. We have also searched for adenovirus E1A proteins by immunoblot with a specific monoclonal antibody. Although pretreatment of cells with either interferon-alpha or interferon-gamma resulted in reduced amounts of E1 and E4 mRNA in the early phase of infection (7 hr p.i.), the near complete inhibition of viral DNA and late transcription was only achieved by interferon-gamma. Immunoblot has shown the absence of the 48-kD E1A protein in cells pretreated with interferon-gamma. The lack of this regulatory adenovirus protein may be involved in the inhibitory mechanism of interferon-gamma on adenovirus.

  11. A bridging model for persistence of a polycomb group protein complex through DNA replication in vitro.

    Science.gov (United States)

    Lo, Stanley M; Follmer, Nicole E; Lengsfeld, Bettina M; Madamba, Egbert V; Seong, Samuel; Grau, Daniel J; Francis, Nicole J

    2012-06-29

    Epigenetic regulation may involve heritable chromatin states, but how chromatin features can be inherited through DNA replication is incompletely understood. We address this question using cell-free replication of chromatin. Previously, we showed that a Polycomb group complex, PRC1, remains continuously associated with chromatin through DNA replication. Here we investigate the mechanism of persistence. We find that a single PRC1 subunit, Posterior sex combs (PSC), can reconstitute persistence through DNA replication. PSC binds nucleosomes and self-interacts, bridging nucleosomes into a stable, oligomeric structure. Within these structures, individual PSC-chromatin contacts are dynamic. Stable association of PSC with chromatin, including through DNA replication, depends on PSC-PSC interactions. Our data suggest that labile individual PSC-chromatin contacts allow passage of the DNA replication machinery while PSC-PSC interactions prevent PSC from dissociating, allowing it to rebind to replicated chromatin. This mechanism may allow inheritance of chromatin proteins including PRC1 through DNA replication to maintain chromatin states.

  12. Low doses of ultraviolet radiation and oxidative damage induce dramatic accumulation of mitochondrial DNA replication intermediates, fork regression, and replication initiation shift.

    Science.gov (United States)

    Torregrosa-Muñumer, Rubén; Goffart, Steffi; Haikonen, Juha A; Pohjoismäki, Jaakko L O

    2015-11-15

    Mitochondrial DNA is prone to damage by various intrinsic as well as environmental stressors. DNA damage can in turn cause problems for replication, resulting in replication stalling and double-strand breaks, which are suspected to be the leading cause of pathological mtDNA rearrangements. In this study, we exposed cells to subtle levels of oxidative stress or UV radiation and followed their effects on mtDNA maintenance. Although the damage did not influence mtDNA copy number, we detected a massive accumulation of RNA:DNA hybrid-containing replication intermediates, followed by an increase in cruciform DNA molecules, as well as in bidirectional replication initiation outside of the main replication origin, OH. Our results suggest that mitochondria maintain two different types of replication as an adaptation to different cellular environments; the RNA:DNA hybrid-involving replication mode maintains mtDNA integrity in tissues with low oxidative stress, and the potentially more error tolerant conventional strand-coupled replication operates when stress is high.

  13. MCNP5 CALCULATIONS REPLICATING ARH-600 NITRATE DATA

    Energy Technology Data Exchange (ETDEWEB)

    FINFROCK SH

    2011-10-25

    This report serves to extend the previous document: 'MCNP Calculations Replicating ARH-600 Data' by replicating the nitrate curves found in ARH-600. This report includes the MCNP models used, the calculated critical dimension for each analyzed parameter set, and the resulting data libraries for use with the CritView code. As with the ARH-600 data, this report is not meant to replace the analysis of the fissile systems by qualified criticality personnel. The M CNP data is presented without accounting for the statistical uncertainty (although this is typically less than 0.001) or bias and, as such, the application of a reasonable safety margin is required. The data that follows pertains to the uranyl nitrate and plutonium nitrate spheres, infinite cylinders, and infinite slabs of varying isotopic composition, reflector thickness, and molarity. Each of the cases was modeled in MCNP (version 5.1.40), using the ENDF/B-VI cross section set. Given a molarity, isotopic composition, and reflector thickness, the fissile concentration and diameter (or thicknesses in the case of the slab geometries) were varied. The diameter for which k-effective equals 1.00 for a given concentration could then be calculated and graphed. These graphs are included in this report. The pages that follow describe the regions modeled, formulas for calculating the various parameters, a list of cross-sections used in the calculations, a description of the automation routine and data, and finally the data output. The data of most interest are the critical dimensions of the various systems analyzed. This is presented graphically, and in table format, in Appendix B. Appendix C provides a text listing of the same data in a format that is compatible with the CritView code. Appendices D and E provide listing of example Template files and MCNP input files (these are discussed further in Section 4). Appendix F is a complete listing of all of the output data (i.e., all of the analyzed dimensions and

  14. Dengue virus binding and replication by platelets.

    Science.gov (United States)

    Simon, Ayo Y; Sutherland, Michael R; Pryzdial, Edward L G

    2015-07-16

    Dengue virus (DENV) infection causes ∼200 million cases of severe flulike illness annually, escalating to life-threatening hemorrhagic fever or shock syndrome in ∼500,000. Although thrombocytopenia is typical of both mild and severe diseases, the mechanism triggering platelet reduction is incompletely understood. As a probable initiating event, direct purified DENV-platelet binding was followed in the current study by quantitative reverse transcription-polymerase chain reaction and confirmed antigenically. Approximately 800 viruses specifically bound per platelet at 37°C. Fewer sites were observed at 25°C, the blood bank storage temperature (∼350 sites), or 4°C, known to attenuate virus cell entry (∼200 sites). Dendritic cell-specific intercellular adhesion molecule-3-grabbing nonintegrin (DC-SIGN) and heparan sulfate proteoglycan were implicated as coreceptors because only the combination of anti-DC-SIGN and low-molecular-weight heparin prevented binding. Interestingly, at 37°C and 25°C, platelets replicated the positive sense single-stranded RNA genome of DENV by up to ∼4-fold over 7 days. Further time course experiments demonstrated production of viral NS1 protein, which is known to be highly antigenic in patient serum. The infectivity of DENV intrinsically decayed in vitro, which was moderated by platelet-mediated generation of viable progeny. This was shown using a transcription inhibitor and confirmed by freeze-denatured platelets being incapable of replicating the DENV genome. For the first time, these data demonstrate that platelets directly bind DENV saturably and produce infectious virus. Thus, expression of antigen encoded by DENV is a novel consideration in the pathogen-induced thrombocytopenia mechanism. These results furthermore draw attention to the possibility that platelets may produce permissive RNA viruses in addition to DENV.

  15. Both Chromosome Decondensation and Condensation Are Dependent on DNA Replication in C. elegans Embryos.

    Science.gov (United States)

    Sonneville, Remi; Craig, Gillian; Labib, Karim; Gartner, Anton; Blow, J Julian

    2015-07-21

    During cell division, chromatin alternates between a condensed state to facilitate chromosome segregation and a decondensed form when DNA replicates. In most tissues, S phase and mitosis are separated by defined G1 and G2 gap phases, but early embryogenesis involves rapid oscillations between replication and mitosis. Using Caenorhabditis elegans embryos as a model system, we show that chromosome condensation and condensin II concentration on chromosomal axes require replicated DNA. In addition, we found that, during late telophase, replication initiates on condensed chromosomes and promotes the rapid decondensation of the chromatin. Upon replication initiation, the CDC-45-MCM-GINS (CMG) DNA helicase drives the release of condensin I complexes from chromatin and the activation or displacement of inactive MCM-2-7 complexes, which together with the nucleoporin MEL-28/ELYS tethers condensed chromatin to the nuclear envelope, thereby promoting chromatin decondensation. Our results show how, in an early embryo, the chromosome-condensation cycle is functionally linked with DNA replication.

  16. Replication Requires Psychological Rather than Statistical Hypotheses : The Case of Eye Movements Enhancing Word Recollection

    NARCIS (Netherlands)

    Phaf, R.H.

    2016-01-01

    Can an experiment be replicated in a mechanical fashion without considering the processes underlying the initial results? Here I will consider a non-replication of Saccade Induced Retrieval Enhancement (SIRE) and argue that it results from focusing on statistical instead of on substantive process

  17. Do gender and personality traits (BFI-10) influence trust? A replication

    DEFF Research Database (Denmark)

    Sudzina, Frantisek

    2016-01-01

    with adding personality traits into the equation. This article is a replication of a previous study. This study uses 1-5 Likert scales while the previous used 1-7 Likert scales, while all the questions/statements stayed the same. The difference is that both measures (not only the first measure) of trust were...

  18. Does Literacy Skill Level Predict Performance in Community College Courses: A Replication and Extension

    Science.gov (United States)

    Allen, Nancy J.; DeLauro, Kimberly A.; Perry, Julia K.; Carman, Carol A.

    2017-01-01

    Previous research has found a positive relationship between students who had completed a sequence of developmental reading and writing courses and success in a reading-intensive college-level course. This study replicates and expands upon the previous research of Goldstein and Perin (2008) by utilizing a differently diverse sample and an…

  19. Does Literacy Skill Level Predict Performance in Community College Courses: A Replication and Extension

    Science.gov (United States)

    Allen, Nancy J.; DeLauro, Kimberly A.; Perry, Julia K.; Carman, Carol A.

    2017-01-01

    Previous research has found a positive relationship between students who had completed a sequence of developmental reading and writing courses and success in a reading-intensive college-level course. This study replicates and expands upon the previous research of Goldstein and Perin (2008) by utilizing a differently diverse sample and an…

  20. Links between DNA Replication, Stem Cells and Cancer

    Directory of Open Access Journals (Sweden)

    Alex Vassilev

    2017-01-01

    Full Text Available Cancers can be categorized into two groups: those whose frequency increases with age, and those resulting from errors during mammalian development. The first group is linked to DNA replication through the accumulation of genetic mutations that occur during proliferation of developmentally acquired stem cells that give rise to and maintain tissues and organs. These mutations, which result from DNA replication errors as well as environmental insults, fall into two categories; cancer driver mutations that initiate carcinogenesis and genome destabilizing mutations that promote aneuploidy through excess genome duplication and chromatid missegregation. Increased genome instability results in accelerated clonal evolution leading to the appearance of more aggressive clones with increased drug resistance. The second group of cancers, termed germ cell neoplasia, results from the mislocation of pluripotent stem cells during early development. During normal development, pluripotent stem cells that originate in early embryos give rise to all of the cell lineages in the embryo and adult, but when they mislocate to ectopic sites, they produce tumors. Remarkably, pluripotent stem cells, like many cancer cells, depend on the Geminin protein to prevent excess DNA replication from triggering DNA damage-dependent apoptosis. This link between the control of DNA replication during early development and germ cell neoplasia reveals Geminin as a potential chemotherapeutic target in the eradication of cancer progenitor cells.

  1. Links between DNA Replication, Stem Cells and Cancer.

    Science.gov (United States)

    Vassilev, Alex; DePamphilis, Melvin L

    2017-01-25

    Cancers can be categorized into two groups: those whose frequency increases with age, and those resulting from errors during mammalian development. The first group is linked to DNA replication through the accumulation of genetic mutations that occur during proliferation of developmentally acquired stem cells that give rise to and maintain tissues and organs. These mutations, which result from DNA replication errors as well as environmental insults, fall into two categories; cancer driver mutations that initiate carcinogenesis and genome destabilizing mutations that promote aneuploidy through excess genome duplication and chromatid missegregation. Increased genome instability results in accelerated clonal evolution leading to the appearance of more aggressive clones with increased drug resistance. The second group of cancers, termed germ cell neoplasia, results from the mislocation of pluripotent stem cells during early development. During normal development, pluripotent stem cells that originate in early embryos give rise to all of the cell lineages in the embryo and adult, but when they mislocate to ectopic sites, they produce tumors. Remarkably, pluripotent stem cells, like many cancer cells, depend on the Geminin protein to prevent excess DNA replication from triggering DNA damage-dependent apoptosis. This link between the control of DNA replication during early development and germ cell neoplasia reveals Geminin as a potential chemotherapeutic target in the eradication of cancer progenitor cells.

  2. Love Matches and Arranged Marriages: A Chinese Replication.

    Science.gov (United States)

    Xiaohe, Xu; Whyte, Martin King

    1990-01-01

    Partially replicated Japanese study by examining transition from arranged to free-choice marriages in Chinese ever-married women (n=586). The results revealed that the role of parents had declined sharply, whereas young people more and more dominated the process of mate selection. However there was still little sign of a "dating culture" emerging.…

  3. Replicating Small Group Research Using the Functional Theory.

    Science.gov (United States)

    Cragan, John F.; Wright, David W.

    A replication study tested functional theory utilizing untrained full-fledged groups. One hundred forty undergraduate students who were enrolled in a small group communication course at a large midwestern university participated in small group discussions analyzing a plagiarism case used in an original study by R. Y. Hirokawa. Results indicated…

  4. Replication-based Inference Algorithms for Hard Computational Problems

    OpenAIRE

    Alamino, Roberto C.; Neirotti, Juan P.; Saad, David

    2013-01-01

    Inference algorithms based on evolving interactions between replicated solutions are introduced and analyzed on a prototypical NP-hard problem - the capacity of the binary Ising perceptron. The efficiency of the algorithm is examined numerically against that of the parallel tempering algorithm, showing improved performance in terms of the results obtained, computing requirements and simplicity of implementation.

  5. Viral replication is enhanced by an HIV-1 intersubtype recombination-derived Vpu protein

    Directory of Open Access Journals (Sweden)

    Salomón Horacio

    2010-10-01

    Full Text Available Abstract Background Multiple HIV-1 intersubtype recombinants have been identified in human populations. Previous studies from our lab group have shown that the epidemic in Argentina is characterized by the high prevalence of a circulating recombinant form, CRF12_BF, and many related BF recombinant forms. In these genomic structures a recombination breakpoint frequently involved the vpu coding region. Due to the scarce knowledge of Vpu participation in the virion release process and its impact on pathogenesis and of the functional capacities of intersubtype recombinant Vpu proteins, the aim of this work was to perform a comparative analysis on virion release capacity and relative replication capacity among viral variants harboring either a BF recombinant Vpu or a subtype B Vpu. Results Our results showed that BF recombinant Vpu was associated to an increased viral particles production when compared to WT B variant in tetherin-expressing cell lines. This observation was tested in the context of a competition assay between the above mentioned variants. The results showed that the replication of the BF Vpu-harboring variant was more efficient in cell cultures than subtype B, reaching a higher frequency in the viral population in a short period of time. Conclusion This study showed that as a result of intersubtype recombination, a structurally re-organized HIV-1 Vpu has an improved in vitro capacity of enhancing viral replication, and provides evidence of the changes occurring in this protein function that could play an important role in the successful spread of intersubtype recombinant variants.

  6. Replication forks reverse at high frequency upon replication stress in Physarum polycephalum.

    Science.gov (United States)

    Maric, Chrystelle; Bénard, Marianne

    2014-12-01

    The addition of hydroxyurea after the onset of S phase allows replication to start and permits the successive detecting of replication-dependent joint DNA molecules and chicken foot structures in the synchronous nuclei of Physarum polycephalum. We find evidence for a very high frequency of reversed replication forks upon replication stress. The formation of these reversed forks is dependent on the presence of joint DNA molecules, the impediment of the replication fork progression by hydroxyurea, and likely on the propensity of some replication origins to reinitiate replication to counteract the action of this compound. As hydroxyurea treatment enables us to successively detect the appearance of joint DNA molecules and then of reversed replication forks, we propose that chicken foot structures are formed both from the regression of hydroxyurea-frozen joint DNA molecules and from hydroxyurea-stalled replication forks. These experiments underscore the transient nature of replication fork regression, which becomes detectable due to the hydroxyurea-induced slowing down of replication fork progression.

  7. Replication initiator DnaA binds at the Caulobacter centromere and enables chromosome segregation.

    Science.gov (United States)

    Mera, Paola E; Kalogeraki, Virginia S; Shapiro, Lucy

    2014-11-11

    During cell division, multiple processes are highly coordinated to faithfully generate genetically equivalent daughter cells. In bacteria, the mechanisms that underlie the coordination of chromosome replication and segregation are poorly understood. Here, we report that the conserved replication initiator, DnaA, can mediate chromosome segregation independent of replication initiation. It does so by binding directly to the parS centromere region of the chromosome, and mutations that alter this interaction result in cells that display aberrant centromere translocation and cell division. We propose that DnaA serves to coordinate bacterial DNA replication with the onset of chromosome segregation.

  8. A quantitative model of DNA replication in Xenopus embryos: reliable replication despite stochasticity

    Science.gov (United States)

    Cheng-Hsin Yang, Scott; Bechhoefer, John

    2008-03-01

    DNA synthesis in Xenopus frog embryos initiates stochastically in time at many sites (origins) along the chromosome. Stochastic initiation implies fluctuations in the replication time and may lead to cell death if replication takes longer than the cell cycle time (˜ 25 min.). Surprisingly, although the typical replication time is about 20 min., in vivo experiments show that replication fails to complete only about 1 in 250 times. How is replication timing accurately controlled despite the stochasticity? Biologists have proposed two mechanisms: the first uses a regular spatial distribution of origins, while the second uses randomly located origins but increases their probability of initiation as the cell cycle proceeds. Here, we show that both mechanisms yield similar end-time distributions, implying that regular origin spacing is not needed for control of replication time. Moreover, we show that the experimentally inferred time-dependent initiation rate satisfies the observed low failure probability and nearly optimizes the use of replicative proteins.

  9. Inhibition of Monkeypox virus replication by RNA interference

    Directory of Open Access Journals (Sweden)

    Jahrling Peter B

    2009-11-01

    Full Text Available Abstract The Orthopoxvirus genus of Poxviridae family is comprised of several human pathogens, including cowpox (CPXV, Vaccinia (VACV, monkeypox (MPV and Variola (VARV viruses. Species of this virus genus cause human diseases with various severities and outcome ranging from mild conditions to death in fulminating cases. Currently, vaccination is the only protective measure against infection with these viruses and no licensed antiviral drug therapy is available. In this study, we investigated the potential of RNA interference pathway (RNAi as a therapeutic approach for orthopox virus infections using MPV as a model. Based on genome-wide expression studies and bioinformatic analysis, we selected 12 viral genes and targeted them by small interference RNA (siRNA. Forty-eight siRNA constructs were developed and evaluated in vitro for their ability to inhibit viral replication. Two genes, each targeted with four different siRNA constructs in one pool, were limiting to viral replication. Seven siRNA constructs from these two pools, targeting either an essential gene for viral replication (A6R or an important gene in viral entry (E8L, inhibited viral replication in cell culture by 65-95% with no apparent cytotoxicity. Further analysis with wild-type and recombinant MPV expressing green fluorescence protein demonstrated that one of these constructs, siA6-a, was the most potent and inhibited viral replication for up to 7 days at a concentration of 10 nM. These results emphasis the essential role of A6R gene in viral replication, and demonstrate the potential of RNAi as a therapeutic approach for developing oligonucleotide-based drug therapy for MPV and other orthopox viruses.

  10. APOBEC3A damages the cellular genome during DNA replication.

    Science.gov (United States)

    Green, Abby M; Landry, Sébastien; Budagyan, Konstantin; Avgousti, Daphne C; Shalhout, Sophia; Bhagwat, Ashok S; Weitzman, Matthew D

    2016-01-01

    The human APOBEC3 family of DNA-cytosine deaminases comprises 7 members (A3A-A3H) that act on single-stranded DNA (ssDNA). The APOBEC3 proteins function within the innate immune system by mutating DNA of viral genomes and retroelements to restrict infection and retrotransposition. Recent evidence suggests that APOBEC3 enzymes can also cause damage to the cellular genome. Mutational patterns consistent with APOBEC3 activity have been identified by bioinformatic analysis of tumor genome sequences. These mutational signatures include clusters of base substitutions that are proposed to occur due to APOBEC3 deamination. It has been suggested that transiently exposed ssDNA segments provide substrate for APOBEC3 deamination leading to mutation signatures within the genome. However, the mechanisms that produce single-stranded substrates for APOBEC3 deamination in mammalian cells have not been demonstrated. We investigated ssDNA at replication forks as a substrate for APOBEC3 deamination. We found that APOBEC3A (A3A) expression leads to DNA damage in replicating cells but this is reduced in quiescent cells. Upon A3A expression, cycling cells activate the DNA replication checkpoint and undergo cell cycle arrest. Additionally, we find that replication stress leaves cells vulnerable to A3A-induced DNA damage. We propose a model to explain A3A-induced damage to the cellular genome in which cytosine deamination at replication forks and other ssDNA substrates results in mutations and DNA breaks. This model highlights the risk of mutagenesis by A3A expression in replicating progenitor cells, and supports the emerging hypothesis that APOBEC3 enzymes contribute to genome instability in human tumors.

  11. A new fuzzy optimal data replication method for data grid

    Directory of Open Access Journals (Sweden)

    Zeinab Ghilavizadeh

    2013-03-01

    Full Text Available These days, There are several applications where we face with large data set and it has become an important part of common resources in different scientific areas. In fact, there are many applications where there are literally huge amount of information handled either in terabyte or in petabyte. Many scientists apply huge amount of data distributed geographically around the world through advanced computing systems. The huge volume data and calculations have created new problems in accessing, processing and distribution of data. The challenges of data management infrastructure have become very difficult under a large amount of data, different geographical spaces, and complicated involved calculations. Data Grid is a remedy to all mentioned problems. In this paper, a new method of dynamic optimal data replication in data grid is introduced where it reduces the total job execution time and increases the locality in accessibilities by detecting and impacting the factors influencing the data replication. Proposed method is composed of two main phases. During the first phase is the phase of file application and replication operation. In this phase, we evaluate three factors influencing the data replication and determine whether the requested file can be replicated or it can be used from distance. In the second phase or the replacement phase, the proposed method investigates whether there is enough space in the destination to store the requested file or not. In this phase, the proposed method also chooses a replica with the lowest value for deletion by considering three replica factors to increase the performance of system. The results of simulation also indicate the improved performance of our proposed method compared with other replication methods represented in the simulator Optorsim.

  12. Previous gastric bypass surgery complicating total thyroidectomy.

    Science.gov (United States)

    Alfonso, Bianca; Jacobson, Adam S; Alon, Eran E; Via, Michael A

    2015-03-01

    Hypocalcemia is a well-known complication of total thyroidectomy. Patients who have previously undergone gastric bypass surgery may be at increased risk of hypocalcemia due to gastrointestinal malabsorption, secondary hyperparathyroidism, and an underlying vitamin D deficiency. We present the case of a 58-year-old woman who underwent a total thyroidectomy for the follicular variant of papillary thyroid carcinoma. Her history included Roux-en-Y gastric bypass surgery. Following the thyroid surgery, she developed postoperative hypocalcemia that required large doses of oral calcium carbonate (7.5 g/day), oral calcitriol (up to 4 μg/day), intravenous calcium gluconate (2.0 g/day), calcium citrate (2.0 g/day), and ergocalciferol (50,000 IU/day). Her serum calcium levels remained normal on this regimen after hospital discharge despite persistent hypoparathyroidism. Bariatric surgery patients who undergo thyroid surgery require aggressive supplementation to maintain normal serum calcium levels. Preoperative supplementation with calcium and vitamin D is strongly recommended.

  13. Sebacinales everywhere: previously overlooked ubiquitous fungal endophytes.

    Science.gov (United States)

    Weiss, Michael; Sýkorová, Zuzana; Garnica, Sigisfredo; Riess, Kai; Martos, Florent; Krause, Cornelia; Oberwinkler, Franz; Bauer, Robert; Redecker, Dirk

    2011-02-15

    Inconspicuous basidiomycetes from the order Sebacinales are known to be involved in a puzzling variety of mutualistic plant-fungal symbioses (mycorrhizae), which presumably involve transport of mineral nutrients. Recently a few members of this fungal order not fitting this definition and commonly referred to as 'endophytes' have raised considerable interest by their ability to enhance plant growth and to increase resistance of their host plants against abiotic stress factors and fungal pathogens. Using DNA-based detection and electron microscopy, we show that Sebacinales are not only extremely versatile in their mycorrhizal associations, but are also almost universally present as symptomless endophytes. They occurred in field specimens of bryophytes, pteridophytes and all families of herbaceous angiosperms we investigated, including liverworts, wheat, maize, and the non-mycorrhizal model plant Arabidopsis thaliana. They were present in all habitats we studied on four continents. We even detected these fungi in herbarium specimens originating from pioneering field trips to North Africa in the 1830s/40s. No geographical or host patterns were detected. Our data suggest that the multitude of mycorrhizal interactions in Sebacinales may have arisen from an ancestral endophytic habit by specialization. Considering their proven beneficial influence on plant growth and their ubiquity, endophytic Sebacinales may be a previously unrecognized universal hidden force in plant ecosystems.

  14. Surgery of intracranial aneurysms previously treated endovascularly.

    Science.gov (United States)

    Tirakotai, Wuttipong; Sure, Ulrich; Yin, Yuhua; Benes, Ludwig; Schulte, Dirk Michael; Bien, Siegfried; Bertalanffy, Helmut

    2007-11-01

    To perform a retrospective study on the patients who underwent aneurysmal surgery following endovascular treatment. We performed a retrospective study on eight patients who underwent aneurysmal surgery following endovascular treatment (-attempts) with gugliemi detachable coils (GDCs). The indications for surgery, surgical techniques and clinical outcomes were analyzed. The indications for surgical treatment after GDC coiling of aneurysm were classified into three groups. First group: surgery of incompletely coiled aneurysms (n=4). Second group: surgery of mass effect on the neural structures due to coil compaction or rebleeding (n=2). Third group: surgery of vascular complications after endovascular procedure due to parent artery occlusion or thrombus propagation from aneurysm (n=2). Aneurysm obliterations could be performed in all cases confirmed by postoperative angiography. Six patients had an excellent outcome and returned to their profession. Patient's visual acuity was improved. One individual experienced right hemiparesis (grade IV/V) and hemihypesthesia. Microsurgical clipping is rarely necessary for previously coiled aneurysms. Surgical treatment is uncommonly required when an acute complication arises during endovascular treatment, or when there is a dynamic change of a residual aneurysm configuration over time that is considered to be insecure.

  15. [Electronic cigarettes - effects on health. Previous reports].

    Science.gov (United States)

    Napierała, Marta; Kulza, Maksymilian; Wachowiak, Anna; Jabłecka, Katarzyna; Florek, Ewa

    2014-01-01

    Currently very popular in the market of tobacco products have gained electronic cigarettes (ang. E-cigarettes). These products are considered to be potentially less harmful in compared to traditional tobacco products. However, current reports indicate that the statements of the producers regarding to the composition of the e- liquids not always are sufficient, and consumers often do not have reliable information on the quality of the product used by them. This paper contain a review of previous reports on the composition of e-cigarettes and their impact on health. Most of the observed health effects was related to symptoms of the respiratory tract, mouth, throat, neurological complications and sensory organs. Particularly hazardous effects of the e-cigarettes were: pneumonia, congestive heart failure, confusion, convulsions, hypotension, aspiration pneumonia, face second-degree burns, blindness, chest pain and rapid heartbeat. In the literature there is no information relating to passive exposure by the aerosols released during e-cigarette smoking. Furthermore, the information regarding to the use of these products in the long term are not also available.

  16. Sebacinales everywhere: previously overlooked ubiquitous fungal endophytes.

    Directory of Open Access Journals (Sweden)

    Michael Weiss

    Full Text Available Inconspicuous basidiomycetes from the order Sebacinales are known to be involved in a puzzling variety of mutualistic plant-fungal symbioses (mycorrhizae, which presumably involve transport of mineral nutrients. Recently a few members of this fungal order not fitting this definition and commonly referred to as 'endophytes' have raised considerable interest by their ability to enhance plant growth and to increase resistance of their host plants against abiotic stress factors and fungal pathogens. Using DNA-based detection and electron microscopy, we show that Sebacinales are not only extremely versatile in their mycorrhizal associations, but are also almost universally present as symptomless endophytes. They occurred in field specimens of bryophytes, pteridophytes and all families of herbaceous angiosperms we investigated, including liverworts, wheat, maize, and the non-mycorrhizal model plant Arabidopsis thaliana. They were present in all habitats we studied on four continents. We even detected these fungi in herbarium specimens originating from pioneering field trips to North Africa in the 1830s/40s. No geographical or host patterns were detected. Our data suggest that the multitude of mycorrhizal interactions in Sebacinales may have arisen from an ancestral endophytic habit by specialization. Considering their proven beneficial influence on plant growth and their ubiquity, endophytic Sebacinales may be a previously unrecognized universal hidden force in plant ecosystems.

  17. A previously undescribed pathway for pyrimidine catabolism.

    Science.gov (United States)

    Loh, Kevin D; Gyaneshwar, Prasad; Markenscoff Papadimitriou, Eirene; Fong, Rebecca; Kim, Kwang-Seo; Parales, Rebecca; Zhou, Zhongrui; Inwood, William; Kustu, Sydney

    2006-03-28

    The b1012 operon of Escherichia coli K-12, which is composed of seven unidentified ORFs, is one of the most highly expressed operons under control of nitrogen regulatory protein C. Examination of strains with lesions in this operon on Biolog Phenotype MicroArray (PM3) plates and subsequent growth tests indicated that they failed to use uridine or uracil as the sole nitrogen source and that the parental strain could use them at room temperature but not at 37 degrees C. A strain carrying an ntrB(Con) mutation, which elevates transcription of genes under nitrogen regulatory protein C control, could also grow on thymidine as the sole nitrogen source, whereas strains with lesions in the b1012 operon could not. Growth-yield experiments indicated that both nitrogens of uridine and thymidine were available. Studies with [(14)C]uridine indicated that a three-carbon waste product from the pyrimidine ring was excreted. After trimethylsilylation and gas chromatography, the waste product was identified by mass spectrometry as 3-hydroxypropionic acid. In agreement with this finding, 2-methyl-3-hydroxypropionic acid was released from thymidine. Both the number of available nitrogens and the waste products distinguished the pathway encoded by the b1012 operon from pyrimidine catabolic pathways described previously. We propose that the genes of this operon be named rutA-G for pyrimidine utilization. The product of the divergently transcribed gene, b1013, is a tetracycline repressor family regulator that controls transcription of the b1012 operon negatively.

  18. Targeting DNA Replication Stress for Cancer Therapy

    Science.gov (United States)

    Zhang, Jun; Dai, Qun; Park, Dongkyoo; Deng, Xingming

    2016-01-01

    The human cellular genome is under constant stress from extrinsic and intrinsic factors, which can lead to DNA damage and defective replication. In normal cells, DNA damage response (DDR) mediated by various checkpoints will either activate the DNA repair system or induce cellular apoptosis/senescence, therefore maintaining overall genomic integrity. Cancer cells, however, due to constitutive growth signaling and defective DDR, may exhibit “replication stress” —a phenomenon unique to cancer cells that is described as the perturbation of error-free DNA replication and slow-down of DNA synthesis. Although replication stress has been proven to induce genomic instability and tumorigenesis, recent studies have counterintuitively shown that enhancing replicative stress through further loosening of the remaining checkpoints in cancer cells to induce their catastrophic failure of proliferation may provide an alternative therapeutic approach. In this review, we discuss the rationale to enhance replicative stress in cancer cells, past approaches using traditional radiation and chemotherapy, and emerging approaches targeting the signaling cascades induced by DNA damage. We also summarize current clinical trials exploring these strategies and propose future research directions including the use of combination therapies, and the identification of potential new targets and biomarkers to track and predict treatment responses to targeting DNA replication stress. PMID:27548226

  19. Targeting DNA Replication Stress for Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Jun Zhang

    2016-08-01

    Full Text Available The human cellular genome is under constant stress from extrinsic and intrinsic factors, which can lead to DNA damage and defective replication. In normal cells, DNA damage response (DDR mediated by various checkpoints will either activate the DNA repair system or induce cellular apoptosis/senescence, therefore maintaining overall genomic integrity. Cancer cells, however, due to constitutive growth signaling and defective DDR, may exhibit “replication stress” —a phenomenon unique to cancer cells that is described as the perturbation of error-free DNA replication and slow-down of DNA synthesis. Although replication stress has been proven to induce genomic instability and tumorigenesis, recent studies have counterintuitively shown that enhancing replicative stress through further loosening of the remaining checkpoints in cancer cells to induce their catastrophic failure of proliferation may provide an alternative therapeutic approach. In this review, we discuss the rationale to enhance replicative stress in cancer cells, past approaches using traditional radiation and chemotherapy, and emerging approaches targeting the signaling cascades induced by DNA damage. We also summarize current clinical trials exploring these strategies and propose future research directions including the use of combination therapies, and the identification of potential new targets and biomarkers to track and predict treatment responses to targeting DNA replication stress.

  20. Identification of imprinted genes using a novel screening method based on asynchronous DNA replication

    Energy Technology Data Exchange (ETDEWEB)

    Kawame, H.; Hansen, R.S.; Gartler, S.M. [Univ. of Washington, Seattle, WA (United States)

    1994-09-01

    Genomic imprinting refers to the process of epigenetic change that occurs during germ cell development that results in either maternal- or paternal-specific gene expression. Identification of imprinted genes is of primary importance to the understanding of imprinting mechanisms and the role of specific imprinted genes in human disease. Recently, it has been established that chromosomal regions known to contain imprinted genes replicate asynchronously. We propose a novel screening method to identify imprinted genes based on replication asynchrony as a marker for imprinted domains. Dividing human cells were pulse-labeled with BrdU and separated into different fractions of S-phase by flow cytometry. A library of late-replicating inter-Alu sequences should be enriched in gene-associated sequences that replicate early on one chromosome and late on the other homologue. Clones were analyzed for replication timing by hybridization to inter-Alu replication profiles. Candidates for replication asynchrony exhibited broad or biphasic replication timing, and these were analyzed for chromosomal location by hybridizations to inter-Alu products from a hybrid mapping panel. Initial screening of 123 clones resulted in 3 asynchronously-replicating clones that localized to single chromosomes. Chromosome 17 and chromosome 19 candidates might be located in regions thought to be imprinted by synteny with mouse chromosomes. A chromosome 15 clone was further characterized because of its possible localization to the Prader-Willi/Angelman locus. This sequence was localized outside the region deleted in Prader-Willi patients, and was found to be expressed in human cell lines. Replication asynchrony for this sequence appears to be polymorphic because cells derived from some individuals indicated synchronous replication. This appears to be the first example of a polymorphism in replication asynchrony.

  1. Identification of putative DnaN-binding motifs in plasmid replication initiation proteins.

    Science.gov (United States)

    Dalrymple, Brian P; Kongsuwan, Kritaya; Wijffels, Gene

    2007-01-01

    Recently the plasmid RK2 replication initiation protein, TrfA, has been shown to bind to the beta subunit of DNA Polymerase III (DnaN) via a short pentapeptide with the consensus QL[S/D]LF. A second consensus peptide, the hexapeptide QLxLxL, has also been demonstrated to mediate binding to DnaN. Here we describe the results of a comprehensive survey of replication initiation proteins encoded by bacterial plasmids to identify putative DnaN-binding sites. Both pentapeptide and hexapeptide motifs have been identified in a number of families of replication initiation proteins. The distribution of sites is sporadic and closely related families of proteins may differ in the presence, location, or type of putative DnaN-binding motif. Neither motif has been identified in replication initiation proteins encoded by plasmids that replicate via rolling circles or strand displacement. The results suggest that the recruitment of DnaN to the origin of replication of a replisome by plasmid replication initiation proteins is not generally required for plasmid replication, but that in some cases it may be beneficial for efficiency of replication initiation.

  2. Host ESCRT proteins are required for bromovirus RNA replication compartment assembly and function.

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    Arturo Diaz

    2015-03-01

    Full Text Available Positive-strand RNA viruses genome replication invariably is associated with vesicles or other rearranged cellular membranes. Brome mosaic virus (BMV RNA replication occurs on perinuclear endoplasmic reticulum (ER membranes in ~70 nm vesicular invaginations (spherules. BMV RNA replication vesicles show multiple parallels with membrane-enveloped, budding retrovirus virions, whose envelopment and release depend on the host ESCRT (endosomal sorting complexes required for transport membrane-remodeling machinery. We now find that deleting components of the ESCRT pathway results in at least two distinct BMV phenotypes. One group of genes regulate RNA replication and the frequency of viral replication complex formation, but had no effect on spherule size, while a second group of genes regulate RNA replication in a way or ways independent of spherule formation. In particular, deleting SNF7 inhibits BMV RNA replication > 25-fold and abolishes detectable BMV spherule formation, even though the BMV RNA replication proteins accumulate and localize normally on perinuclear ER membranes. Moreover, BMV ESCRT recruitment and spherule assembly depend on different sets of protein-protein interactions from those used by multivesicular body vesicles, HIV-1 virion budding, or tomato bushy stunt virus (TBSV spherule formation. These and other data demonstrate that BMV requires cellular ESCRT components for proper formation and function of its vesicular RNA replication compartments. The results highlight growing but diverse interactions of ESCRT factors with many viruses and viral processes, and potential value of the ESCRT pathway as a target for broad-spectrum antiviral resistance.

  3. Host ESCRT proteins are required for bromovirus RNA replication compartment assembly and function.

    Science.gov (United States)

    Diaz, Arturo; Zhang, Jiantao; Ollwerther, Abigail; Wang, Xiaofeng; Ahlquist, Paul

    2015-03-01

    Positive-strand RNA viruses genome replication invariably is associated with vesicles or other rearranged cellular membranes. Brome mosaic virus (BMV) RNA replication occurs on perinuclear endoplasmic reticulum (ER) membranes in ~70 nm vesicular invaginations (spherules). BMV RNA replication vesicles show multiple parallels with membrane-enveloped, budding retrovirus virions, whose envelopment and release depend on the host ESCRT (endosomal sorting complexes required for transport) membrane-remodeling machinery. We now find that deleting components of the ESCRT pathway results in at least two distinct BMV phenotypes. One group of genes regulate RNA replication and the frequency of viral replication complex formation, but had no effect on spherule size, while a second group of genes regulate RNA replication in a way or ways independent of spherule formation. In particular, deleting SNF7 inhibits BMV RNA replication > 25-fold and abolishes detectable BMV spherule formation, even though the BMV RNA replication proteins accumulate and localize normally on perinuclear ER membranes. Moreover, BMV ESCRT recruitment and spherule assembly depend on different sets of protein-protein interactions from those used by multivesicular body vesicles, HIV-1 virion budding, or tomato bushy stunt virus (TBSV) spherule formation. These and other data demonstrate that BMV requires cellular ESCRT components for proper formation and function of its vesicular RNA replication compartments. The results highlight growing but diverse interactions of ESCRT factors with many viruses and viral processes, and potential value of the ESCRT pathway as a target for broad-spectrum antiviral resistance.

  4. Social learning and the replication process: an experimental investigation.

    Science.gov (United States)

    Derex, Maxime; Feron, Romain; Godelle, Bernard; Raymond, Michel

    2015-06-07

    Human cultural traits typically result from a gradual process that has been described as analogous to biological evolution. This observation has led pioneering scholars to draw inspiration from population genetics to develop a rigorous and successful theoretical framework of cultural evolution. Social learning, the mechanism allowing information to be transmitted between individuals, has thus been described as a simple replication mechanism. Although useful, the extent to which this idealization appropriately describes the actual social learning events has not been carefully assessed. Here, we used a specifically developed computer task to evaluate (i) the extent to which social learning leads to the replication of an observed behaviour and (ii) the consequences it has for fitness landscape exploration. Our results show that social learning does not lead to a dichotomous choice between disregarding and replicating social information. Rather, it appeared that individuals combine and transform information coming from multiple sources to produce new solutions. As a consequence, landscape exploration was promoted by the use of social information. These results invite us to rethink the way social learning is commonly modelled and could question the validity of predictions coming from models considering this process as replicative.

  5. Study on the micro-replication of shark skin

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Direct replication of creatural scarfskins to form biomimetic surfaces with relatively vivid morphology is a new attempt of the bio-replicated forming technology at animal body. Taking shark skins as the replication templates, and the micro-embossing and micro-molding as the material forming methods, the micro-replicating technology of the outward morphology on shark skins was demonstrated. The preliminary analysis on replication precision indicates that the bio-replicated forming technology can replicate the outward morphology of the shark scales with good precision, which validates the application of the bio-replicated forming technology in the direct morphology replication of the firm creatural scarfskins.

  6. Replicated Data Management for Mobile Computing

    CERN Document Server

    Douglas, Terry

    2008-01-01

    Managing data in a mobile computing environment invariably involves caching or replication. In many cases, a mobile device has access only to data that is stored locally, and much of that data arrives via replication from other devices, PCs, and services. Given portable devices with limited resources, weak or intermittent connectivity, and security vulnerabilities, data replication serves to increase availability, reduce communication costs, foster sharing, and enhance survivability of critical information. Mobile systems have employed a variety of distributed architectures from client-server

  7. IFN-Gamma Inhibits JC Virus Replication in Glial Cells by Suppressing T-Antigen Expression.

    Directory of Open Access Journals (Sweden)

    Francesca Isabella De-Simone

    Full Text Available Patients undergoing immune modulatory therapies for the treatment of autoimmune diseases such as multiple sclerosis, and individuals with an impaired-immune system, most notably AIDS patients, are in the high risk group of developing progressive multifocal leukoencephalopathy (PML, an often lethal disease of the brain characterized by lytic infection of oligodendrocytes in the central nervous system (CNS with JC virus (JCV. The immune system plays an important regulatory role in controlling JCV reactivation from latent sites by limiting viral gene expression and replication. However, little is known regarding the molecular mechanisms responsible for this regulation.Here, we investigated the impact of soluble immune mediators secreted by activated PBMCs on viral replication and gene expression by cell culture models and molecular virology techniques. Our data revealed that viral gene expression and viral replication were suppressed by soluble immune mediators. Further studies demonstrated that soluble immune mediators secreted by activated PBMCs inhibit viral replication induced by T-antigen, the major viral regulatory protein, by suppressing its expression in glial cells. This unexpected suppression of T-antigen was mainly associated with the suppression of translational initiation. Cytokine/chemokine array studies using conditioned media from activated PBMCs revealed several candidate cytokines with possible roles in this regulation. Among them, only IFN-γ showed a robust inhibition of T-antigen expression. While potential roles for IFN-β, and to a lesser extent IFN-α have been described for JCV, IFN-γ has not been previously implicated. Further analysis of IFN-γ signaling pathway revealed a novel role of Jak1 signaling in control of viral T-antigen expression. Furthermore, IFN-γ suppressed JCV replication and viral propagation in primary human fetal glial cells, and showed a strong anti-JCV activity.Our results suggest a novel role for

  8. The Saccharomyces cerevisiae Dna2 can function as a sole nuclease in the processing of Okazaki fragments in DNA replication.

    Science.gov (United States)

    Levikova, Maryna; Cejka, Petr

    2015-09-18

    During DNA replication, synthesis of the lagging strand occurs in stretches termed Okazaki fragments. Before adjacent fragments are ligated, any flaps resulting from the displacement of the 5' DNA end of the Okazaki fragment must be cleaved. Previously, Dna2 was implicated to function upstream of flap endonuclease 1 (Fen1 or Rad27) in the processing of long flaps bound by the replication protein A (RPA). Here we show that Dna2 efficiently cleaves long DNA flaps exactly at or directly adjacent to the base. A fraction of the flaps cleaved by Dna2 can be immediately ligated. When coupled with DNA replication, the flap processing activity of Dna2 leads to a nearly complete Okazaki fragment maturation at sub-nanomolar Dna2 concentrations. Our results indicate that a subsequent nucleolytic activity of Fen1 is not required in most cases. In contrast Dna2 is completely incapable to cleave short flaps. We show that also Dna2, like Fen1, interacts with proliferating cell nuclear antigen (PCNA). We propose a model where Dna2 alone is responsible for cleaving of RPA-bound long flaps, while Fen1 or exonuclease 1 (Exo1) cleave short flaps. Our results argue that Dna2 can function in a separate, rather than in a Fen1-dependent pathway.

  9. A conserved helicase processivity factor is needed for conjugation and replication of an integrative and conjugative element.

    Directory of Open Access Journals (Sweden)

    Jacob Thomas

    Full Text Available Integrative and conjugative elements (ICEs are agents of horizontal gene transfer and have major roles in evolution and acquisition of new traits, including antibiotic resistances. ICEs are found integrated in a host chromosome and can excise and transfer to recipient bacteria via conjugation. Conjugation involves nicking of the ICE origin of transfer (oriT by the ICE-encoded relaxase and transfer of the nicked single strand of ICE DNA. For ICEBs1 of Bacillus subtilis, nicking of oriT by the ICEBs1 relaxase NicK also initiates rolling circle replication. This autonomous replication of ICEBs1 is critical for stability of the excised element in growing cells. We found a conserved and previously uncharacterized ICE gene that is required for conjugation and replication of ICEBs1. Our results indicate that this gene, helP (formerly ydcP, encodes a helicase processivity factor that enables the host-encoded helicase PcrA to unwind the double-stranded ICEBs1 DNA. HelP was required for both conjugation and replication of ICEBs1, and HelP and NicK were the only ICEBs1 proteins needed for replication from ICEBs1 oriT. Using chromatin immunoprecipitation, we measured association of HelP, NicK, PcrA, and the host-encoded single-strand DNA binding protein Ssb with ICEBs1. We found that NicK was required for association of HelP and PcrA with ICEBs1 DNA. HelP was required for association of PcrA and Ssb with ICEBs1 regions distal, but not proximal, to oriT, indicating that PcrA needs HelP to progress beyond nicked oriT and unwind ICEBs1. In vitro, HelP directly stimulated the helicase activity of the PcrA homologue UvrD. Our findings demonstrate that HelP is a helicase processivity factor needed for efficient unwinding of ICEBs1 for conjugation and replication. Homologues of HelP and PcrA-type helicases are encoded on many known and putative ICEs. We propose that these factors are essential for ICE conjugation, replication, and genetic stability.

  10. A quantitative and high-throughput assay of human papillomavirus DNA replication.

    Science.gov (United States)

    Gagnon, David; Fradet-Turcotte, Amélie; Archambault, Jacques

    2015-01-01

    Replication of the human papillomavirus (HPV) double-stranded DNA genome is accomplished by the two viral proteins E1 and E2 in concert with host DNA replication factors. HPV DNA replication is an established model of eukaryotic DNA replication and a potential target for antiviral therapy. Assays to measure the transient replication of HPV DNA in transfected cells have been developed, which rely on a plasmid carrying the viral origin of DNA replication (ori) together with expression vectors for E1 and E2. Replication of the ori-plasmid is typically measured by Southern blotting or PCR analysis of newly replicated DNA (i.e., DpnI digested DNA) several days post-transfection. Although extremely valuable, these assays have been difficult to perform in a high-throughput and quantitative manner. Here, we describe a modified version of the transient DNA replication assay that circumvents these limitations by incorporating a firefly luciferase expression cassette in cis of the ori. Replication of this ori-plasmid by E1 and E2 results in increased levels of firefly luciferase activity that can be accurately quantified and normalized to those of Renilla luciferase expressed from a control plasmid, thus obviating the need for DNA extraction, digestion, and analysis. We provide a detailed protocol for performing the HPV type 31 DNA replication assay in a 96-well plate format suitable for small-molecule screening and EC50 determinations. The quantitative and high-throughput nature of the assay should greatly facilitate the study of HPV DNA replication and the identification of inhibitors thereof.

  11. NIPBL rearrangements in Cornelia de Lange syndrome: evidence for replicative mechanism and genotype–phenotype correlation

    Science.gov (United States)

    Pehlivan, Davut; Hullings, Melanie; Carvalho, Claudia M.B.; Gonzaga-Jauregui, Claudia G.; Loy, Elizabeth; Jackson, Laird G.; Krantz, Ian D.; Deardorff, Matthew A.; Lupski, James R.

    2013-01-01

    Purpose Cornelia de Lange syndrome (CdLS) is a multisystem congenital anomaly disorder characterized by mental retardation, limb abnormalities, distinctive facial features, and hirsutism. Mutations in three genes involved in sister chromatid cohesion, NIPBL, SMC1A, and SMC3, account for ~55% of CdLS cases. The molecular etiology of a significant fraction of CdLS cases remains unknown. We hypothesized that large genomic rearrangements of cohesin complex subunit genes may play a role in the molecular etiology of this disorder. Methods Custom high-resolution oligonucleotide array comparative genomic hybridization analyses interrogating candidate cohesin genes and breakpoint junction sequencing of identified genomic variants were performed. Results Of the 162 patients with CdLS, for whom mutations in known CdLS genes were previously negative by sequencing, deletions containing NIPBL exons were observed in 7 subjects (~5%). Breakpoint sequences in five patients implicated microhomology-mediated replicative mechanisms—such as serial replication slippage and fork stalling and template switching/microhomology-mediated break-induced replication—as a potential predominant contributor to these copy number variations. Most deletions are predicted to result in haploinsuflciency due to heterozygous loss-of-function mutations; such mutations may result in a more severe CdLS phenotype. Conclusion Our findings suggest a potential clinical utility to testing for copy number variations involving NIPBL when clinically diagnosed CdLS cases are mutation-negative by DNA-sequencing studies. PMID:22241092

  12. Small noncoding RNA modulates japanese encephalitis virus replication and translation in trans

    Directory of Open Access Journals (Sweden)

    Fan Yi-Hsin

    2011-11-01

    Full Text Available Abstract Background Sequence and structural elements in the 3'-untranslated region (UTR of Japanese encephalitis virus (JEV are known to regulate translation and replication. We previously reported an abundant accumulation of small subgenomic flaviviral RNA (sfRNA which is collinear with the highly conserved regions of the 3'-UTR in JEV-infected cells. However, function of the sfRNA in JEV life cycle remains unknown. Results Northern blot and real-time RT-PCR analyses indicated that the sfRNA becomes apparent at the time point at which minus-strand RNA (antigenome reaches a plateau suggesting a role for sfRNA in the regulation of antigenome synthesis. Transfection of minus-sense sfRNA into JEV-infected cells, in order to counter the effects of plus-sense sfRNA, resulted in higher levels of antigenome suggesting that the presence of the sfRNA inhibits antigenome synthesis. Trans-acting effect of sfRNA on JEV translation was studied using a reporter mRNA containing the luciferase gene fused to partial coding regions of JEV and flanked by the respective JEV UTRs. In vivo and in vitro translation revealed that sfRNA inhibited JEV translation. Conclusions Our results indicate that sfRNA modulates viral translation and replication in trans.

  13. Pairwise growth competition assay for determining the replication fitness of human immunodeficiency viruses.

    Science.gov (United States)

    Manocheewa, Siriphan; Lanxon-Cookson, Erinn C; Liu, Yi; Swain, J Victor; McClure, Jan; Rao, Ushnal; Maust, Brandon; Deng, Wenjie; Sunshine, Justine E; Kim, Moon; Rolland, Morgane; Mullins, James I

    2015-05-04

    In vitro fitness assays are essential tools for determining viral replication fitness for viruses such as HIV-1. Various measurements have been used to extrapolate viral replication fitness, ranging from the number of viral particles per infectious unit, growth rate in cell culture, and relative fitness derived from multiple-cycle growth competition assays. Growth competition assays provide a particularly sensitive measurement of fitness since the viruses are competing for cellular targets under identical growth conditions. There are several experimental factors to consider when conducting growth competition assays, including the multiplicity of infection (MOI), sampling times, and viral detection and fitness calculation methods. Each factor can affect the end result and hence must be considered carefully during the experimental design. The protocol presented here includes steps from constructing a new recombinant HIV-1 clone to performing growth competition assays and analyzing the experimental results. This protocol utilizes experimental parameter values previously shown to yield consistent and robust results. Alternatives are discussed, as some parameters need to be adjusted according to the cell type and viruses being studied. The protocol contains two alternative viral detection methods to provide flexibility as the availability of instruments, reagents and expertise varies between laboratories.

  14. Efficient Dynamic Replication Algorithm Using Agent for Data Grid

    Directory of Open Access Journals (Sweden)

    Priyanka Vashisht

    2014-01-01

    Full Text Available In data grids scientific and business applications produce huge volume of data which needs to be transferred among the distributed and heterogeneous nodes of data grids. Data replication provides a solution for managing data files efficiently in large grids. The data replication helps in enhancing the data availability which reduces the overall access time of the file. In this paper an algorithm, namely, EDRA using agents for data grid, has been proposed and implemented. EDRA consists of dynamic replication of hierarchical structure taken into account for the selection of best replica. Decision for selecting the best replica is based on scheduling parameters. The scheduling parameters are bandwidth, load gauge, and computing capacity of the node. The scheduling in data grid helps in reducing the data access time. The distribution of the load on the nodes of data grid is done evenly by considering scheduling parameters. EDRA is implemented using data grid simulator, namely, OptorSim. European Data Grid CMS test bed topology is used in this experiment. The simulation results are obtained by comparing BHR, LRU, No Replication, and EDRA. The result shows the efficiency of EDRA algorithm in terms of mean job execution time, network usage, and storage usage of node.

  15. A transcription and translation-coupled DNA replication system using rolling-circle replication.

    Science.gov (United States)

    Sakatani, Yoshihiro; Ichihashi, Norikazu; Kazuta, Yasuaki; Yomo, Tetsuya

    2015-05-27

    All living organisms have a genome replication system in which genomic DNA is replicated by a DNA polymerase translated from mRNA transcribed from the genome. The artificial reconstitution of this genome replication system is a great challenge in in vitro synthetic biology. In this study, we attempted to construct a transcription- and translation-coupled DNA replication (TTcDR) system using circular genomic DNA encoding phi29 DNA polymerase and a reconstituted transcription and translation system. In this system, phi29 DNA polymerase was translated from the genome and replicated the genome in a rolling-circle manner. When using a traditional translation system composition, almost no DNA replication was observed, because the tRNA and nucleoside triphosphates included in the translation system significantly inhibited DNA replication. To minimize these inhibitory effects, we optimized the composition of the TTcDR system and improved replication by approximately 100-fold. Using our system, genomic DNA was replicated up to 10 times in 12 hours at 30 °C. This system provides a step toward the in vitro construction of an artificial genome replication system, which is a prerequisite for the construction of an artificial cell.

  16. Replicating chromatin: a tale of histones

    DEFF Research Database (Denmark)

    Groth, Anja

    2009-01-01

    Chromatin serves structural and functional roles crucial for genome stability and correct gene expression. This organization must be reproduced on daughter strands during replication to maintain proper overlay of epigenetic fabric onto genetic sequence. Nucleosomes constitute the structural...

  17. Control of chromosome replication in caulobacter crescentus.

    Science.gov (United States)

    Marczynski, Gregory T; Shapiro, Lucy

    2002-01-01

    Caulobacter crescentus permits detailed analysis of chromosome replication control during a developmental cell cycle. Its chromosome replication origin (Cori) may be prototypical of the large and diverse class of alpha-proteobacteria. Cori has features that both affiliate and distinguish it from the Escherichia coli chromosome replication origin. For example, requirements for DnaA protein and RNA transcription affiliate both origins. However, Cori is distinguished by several features, and especially by five binding sites for the CtrA response regulator protein. To selectively repress and limit chromosome replication, CtrA receives both protein degradation and protein phosphorylation signals. The signal mediators, proteases, response regulators, and kinases, as well as Cori DNA and the replisome, all show distinct patterns of temporal and spatial organization during cell cycle progression. Future studies should integrate our knowledge of biochemical activities at Cori with our emerging understanding of cytological dynamics in C. crescentus and other bacteria.

  18. Initiation of Replication in Escherichia coli

    DEFF Research Database (Denmark)

    Frimodt-Møller, Jakob

    The circular chromosome of Escherichia coli is replicated by two replisomes assembled at the unique origin and moving in the opposite direction until they meet in the less well defined terminus. The key protein in initiation of replication, DnaA, facilitates the unwinding of double-stranded DNA...... to single-stranded DNA in oriC. Although DnaA is able to bind both ADP and ATP, DnaA is only active in initiation when bound to ATP. Although initiation of replication, and the regulation of this, is thoroughly investigated it is still not fully understood. The overall aim of the thesis was to investigate...... the regulation of initiation, the effect on the cell when regulation fails, and if regulation was interlinked to chromosomal organization. This thesis uncovers that there exists a subtle balance between chromosome replication and reactive oxygen species (ROS) inflicted DNA damage. Thus, failure in regulation...

  19. Surface Micro Topography Replication in Injection Moulding

    DEFF Research Database (Denmark)

    Arlø, Uffe Rolf; Hansen, Hans Nørgaard; Kjær, Erik Michael

    2005-01-01

    carried out with rough EDM (electrical discharge machining) mould surfaces, a PS grade, and by applying established three-dimensional topography parameters. Significant quantitative relationships between process parameters and topography parameters were established. It further appeared that replication...

  20. Eclipse period of R1 plasmids during downshift from elevated copy number: Nonrandom selection of copies for replication.

    Science.gov (United States)

    Olsson, Jan A; Berg, Otto; Nordström, Kurt; Dasgupta, Santanu

    2012-03-01

    The classical Meselson-Stahl density-shift method was used to study replication of pOU71, a runaway-replication derivative of plasmid R1 in Escherichia coli. The miniplasmid maintained the normal low copy number of R1 during steady growth at 30°C, but as growth temperatures were raised above 34°C, the copy number of the plasmid increased to higher levels, and at 42°C, it replicated without control in a runaway replication mode with lethal consequences for the host. The eclipse periods (minimum time between successive replication of the same DNA) of the plasmid shortened with rising copy numbers at increasing growth temperatures (Olsson et al., 2003). In this work, eclipse periods were measured during downshifts in copy number of pOU71 after it had replicated at 39 and 42°C, resulting in 7- and 50-fold higher than normal plasmid copy number per cell, respectively. Eclipse periods for plasmid replication, measured during copy number downshift, suggested that plasmid R1, normally selected randomly for replication, showed a bias such that a newly replicated DNA had a higher probability of replication compared to the bulk of the R1 population. However, even the unexpected nonrandom replication followed the copy number kinetics such that every generation, the plasmids underwent the normal inherited number of replication, n, independent of the actual number of plasmid copies in a newborn cell.

  1. A genetic screen for replication initiation defective (rid mutants in Schizosaccharomyces pombe

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    Locovei Alexandra M

    2010-08-01

    Full Text Available Abstract In fission yeast the intra-S phase and DNA damage checkpoints are activated in response to inhibition of DNA replication or DNA damage, respectively. The intra-S phase checkpoint responds to stalled replication forks leading to the activation of the Cds1 kinase that both delays cell cycle progression and stabilizes DNA replication forks. The DNA damage checkpoint, that operates during the G2 phase of the cell cycle delays mitotic progression through activation of the checkpoint kinase, Chk1. Delay of the cell cycle is believed to be essential to allow time for either replication restart (in S phase or DNA damage repair (in G2. Previously, our laboratory showed that fission yeast cells deleted for the N-terminal half of DNA polymerase ε (Cdc20 are delayed in S phase, but surprisingly require Chk1 rather than Cds1 to maintain cell viability. Several additional DNA replication mutants were then tested for their dependency on Chk1 or Cds1 when grown under semi-permissive temperatures. We discovered that mutants defective in DNA replication initiation are sensitive only to loss of Chk1, whilst mutations that inhibit DNA replication elongation are sensitive to loss of both Cds1 and Chk1. To confirm that the Chk1-sensitive, Cds1-insensitive phenotype (rid phenotype is specific to mutants defective in DNA replication initiation, we completed a genetic screen for cell cycle mutants that require Chk1, but not Cds1 to maintain cell viability when grown at semi-permissive temperatures. Our screen identified two mutants, rid1-1 and rid2-1, that are defective in Orc1 and Mcm4, respectively. Both mutants show defects in DNA replication initiation consistent with our hypothesis that the rid phenotype is replication initiation specific. In the case of Mcm4, the mutation has been mapped to a highly conserved region of the protein that appears to be required for DNA replication initiation, but not elongation. Therefore, we conclude that the cellular

  2. Regulation of the switch from early to late bacteriophage lambda DNA replication.

    Science.gov (United States)

    Baranska, S; Gabig, M; Wegrzyn, A; Konopa, G; Herman-Antosiewicz, A; Hernandez, P; Schvartzman, J B; Helinski, D R; Wegrzyn, G

    2001-03-01

    There are two modes of bacteriophage lambda DNA replication following infection of its host, Escherichia coli. Early after infection, replication occurs according to the theta (theta or circle-to-circle) mode, and is later switched to the sigma (sigma or rolling-circle) mode. It is not known how this switch, occurring at a specific time in the infection cycle, is regulated. Here it is demonstrated that in wild-type cells the replication starting from orilambda proceeds both bidirectionally and unidirectionally, whereas in bacteria devoid of a functional DnaA protein, replication from orilambda is predominantly unidirectional. The regulation of directionality of replication from orilambda is mediated by positive control of lambda p(R) promoter activity by DnaA, since the mode of replication of an artificial lambda replicon bearing the p(tet) promoter instead of p(R) was found to be independent of DnaA function. These findings and results of density-shift experiments suggest that in dnaA mutants infected with lambda, phage DNA replication proceeds predominantly according to the unidirectional theta mechanism and is switched early after infection to the sigma mode. It is proposed that in wild-type E. coli cells infected with lambda, phage DNA replication proceeds according to a bidirectional theta mechanism early after infection due to efficient transcriptional activation of orilambda, stimulated by the host DnaA protein. After a few rounds of this type of replication, the resulting increased copy number of lambda genomic DNA may cause a depletion of free DnaA protein because of its interaction with the multiple DnaA-binding sites in lambda DNA. It is proposed that this may lead to inefficient transcriptional activation of orilambda resulting in unidirectional theta replication followed by sigma type replication.

  3. Aggregate and Individual Replication Probability within an Explicit Model of the Research Process

    Science.gov (United States)

    Miller, Jeff; Schwarz, Wolf

    2011-01-01

    We study a model of the research process in which the true effect size, the replication jitter due to changes in experimental procedure, and the statistical error of effect size measurement are all normally distributed random variables. Within this model, we analyze the probability of successfully replicating an initial experimental result by…

  4. Aggregate and Individual Replication Probability within an Explicit Model of the Research Process

    Science.gov (United States)

    Miller, Jeff; Schwarz, Wolf

    2011-01-01

    We study a model of the research process in which the true effect size, the replication jitter due to changes in experimental procedure, and the statistical error of effect size measurement are all normally distributed random variables. Within this model, we analyze the probability of successfully replicating an initial experimental result by…

  5. Chikungunya triggers an autophagic process which promotes viral replication

    Directory of Open Access Journals (Sweden)

    Briant Laurence

    2011-09-01

    Full Text Available Abstract Background Chikungunya Virus (ChikV surprised by a massive re-emerging outbreak in Indian Ocean in 2006, reaching Europe in 2007 and exhibited exceptional severe physiopathology in infants and elderly patients. In this context, it is important to analyze the innate immune host responses triggered against ChikV. Autophagy has been shown to be an important component of the innate immune response and is involved in host defense elimination of different pathogens. However, the autophagic process was recently observed to be hijacked by virus for their own replication. Here we provide the first evidence that hallmarks of autophagy are specifically found in HEK.293 infected cells and are involved in ChikV replication. Methods To test the capacity of ChikV to mobilize the autophagic machinery, we performed fluorescence microscopy experiments on HEK.GFP.LC3 stable cells, and followed the LC3 distribution during the time course of ChikV infection. To confirm this, we performed electron microscopy on HEK.293 infected cells. To test the effect of ChikV-induced-autophagy on viral replication, we blocked the autophagic process, either by pharmacological (3-MA or genetic inhibition (siRNA against the transcript of Beclin 1, an autophagic protein, and analyzed the percentage of infected cells and the viral RNA load released in the supernatant. Moreover, the effect of induction of autophagy by Rapamycin on viral replication was tested. Results The increasing number of GFP-LC3 positive cells with a punctate staining together with the enhanced number of GFP-LC3 dots per cell showed that ChikV triggered an autophagic process in HEK.293 infected cells. Those results were confirmed by electron microscopy analysis since numerous membrane-bound vacuoles characteristic of autophagosomes were observed in infected cells. Moreover, we found that inhibition of autophagy, either by biochemical reagent and RNA interference, dramatically decreases ChikV replication

  6. Single molecular biology: coming of age in DNA replication.

    Science.gov (United States)

    Liu, Xiao-Jing; Lou, Hui-Qiang

    2017-09-20

    DNA replication is an essential process of the living organisms. To achieve precise and reliable replication, DNA polymerases play a central role in DNA synthesis. Previous investigations have shown that the average rates of DNA synthesis on the leading and lagging strands in a replisome must be similar to avoid the formation of significant gaps in the nascent strands. The underlying mechanism has been assumed to be coordination between leading- and lagging-strand polymerases. However, Kowalczykowski's lab members recently performed single molecule techniques in E. coli and showed the real-time behavior of a replisome. The leading- and lagging-strand polymerases function stochastically and independently. Furthermore, when a DNA polymerase is paused, the helicase slows down in a self-regulating fail-safe mechanism, akin to a ''dead-man's switch''. Based on the real-time single-molecular observation, the authors propose that leading- and lagging-strand polymerases synthesize DNA stochastically within a Gaussian distribution. Along with the development and application of single-molecule techniques, we will witness a new age of DNA replication and other biological researches.

  7. Lymphatic endothelial cells are a replicative niche for Mycobacterium tuberculosis

    Science.gov (United States)

    Lerner, Thomas R.; de Souza Carvalho-Wodarz, Cristiane; Repnik, Urska; Russell, Matthew R.G.; Borel, Sophie; Diedrich, Collin R.; Rohde, Manfred; Wainwright, Helen; Collinson, Lucy M.; Wilkinson, Robert J.; Griffiths, Gareth; Gutierrez, Maximiliano G.

    2016-01-01

    In extrapulmonary tuberculosis, the most common site of infection is within the lymphatic system, and there is growing recognition that lymphatic endothelial cells (LECs) are involved in immune function. Here, we identified LECs, which line the lymphatic vessels, as a niche for Mycobacterium tuberculosis in the lymph nodes of patients with tuberculosis. In cultured primary human LECs (hLECs), we determined that M. tuberculosis replicates both in the cytosol and within autophagosomes, but the bacteria failed to replicate when the virulence locus RD1 was deleted. Activation by IFN-γ induced a cell-autonomous response in hLECs via autophagy and NO production that restricted M. tuberculosis growth. Thus, depending on the activation status of LECs, autophagy can both promote and restrict replication. Together, these findings reveal a previously unrecognized role for hLECs and autophagy in tuberculosis pathogenesis and suggest that hLECs are a potential niche for M. tuberculosis that allows establishment of persistent infection in lymph nodes. PMID:26901813

  8. Defendants previous history and mock sentencing.

    Science.gov (United States)

    Wear, D A; Pasewark, R A

    1984-05-01

    Six hundred forty-four undergraduates served as mock judges in sentencing male or female defendants convicted of homicide, child molestation, embezzlement, fraudulent issuance of checks, heroin possession, and consensual homosexuality. Defendants had a reported history of psychiatric hospitalization, imprisonment, or neither hospitalization nor incarceration. Results indicated: (1) those defendants with a mental health history were more likely to be accorded a disposition that involved mandatory health treatment; (2) dispositions of persons with a mental health history tended to be more restrictive than those of defendants with neither a mental health nor criminal history; and (3) sex of defendant of mock judge influenced sentencing disposition only in child molestation cases.

  9. Elevated proteasome capacity extends replicative lifespan in Saccharomyces cerevisiae.

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    Undine Kruegel

    2011-09-01

    Full Text Available Aging is characterized by the accumulation of damaged cellular macromolecules caused by declining repair and elimination pathways. An integral component employed by cells to counter toxic protein aggregates is the conserved ubiquitin/proteasome system (UPS. Previous studies have described an age-dependent decline of proteasomal function and increased longevity correlates with sustained proteasome capacity in centenarians and in naked mole rats, a long-lived rodent. Proof for a direct impact of enhanced proteasome function on longevity, however, is still lacking. To determine the importance of proteasome function in yeast aging, we established a method to modulate UPS capacity by manipulating levels of the UPS-related transcription factor Rpn4. While cells lacking RPN4 exhibit a decreased non-adaptable proteasome pool, loss of UBR2, an ubiquitin ligase that regulates Rpn4 turnover, results in elevated Rpn4 levels, which upregulates UPS components. Increased UPS capacity significantly enhances replicative lifespan (RLS and resistance to proteotoxic stress, while reduced UPS capacity has opposing consequences. Despite tight transcriptional co-regulation of the UPS and oxidative detoxification systems, the impact of proteasome capacity on lifespan is independent of the latter, since elimination of Yap1, a key regulator of the oxidative stress response, does not affect lifespan extension of cells with higher proteasome capacity. Moreover, since elevated proteasome capacity results in improved clearance of toxic huntingtin fragments in a yeast model for neurodegenerative diseases, we speculate that the observed lifespan extension originates from prolonged elimination of damaged proteins in old mother cells. Epistasis analyses indicate that proteasome-mediated modulation of lifespan is at least partially distinct from dietary restriction, Tor1, and Sir2. These findings demonstrate that UPS capacity determines yeast RLS by a mechanism that is distinct

  10. Enrichment of Phosphatidylethanolamine in Viral Replication Compartments via Co-opting the Endosomal Rab5 Small GTPase by a Positive-Strand RNA Virus

    Science.gov (United States)

    Xu, Kai; Nagy, Peter D.

    2016-01-01

    Positive-strand RNA viruses build extensive membranous replication compartments to support replication and protect the virus from antiviral responses by the host. These viruses require host factors and various lipids to form viral replication complexes (VRCs). The VRCs built by Tomato bushy stunt virus (TBSV) are enriched with phosphatidylethanolamine (PE) through a previously unknown pathway. To unravel the mechanism of PE enrichment within the TBSV replication compartment, in this paper, the authors demonstrate that TBSV co-opts the guanosine triphosphate (GTP)-bound active form of the endosomal Rab5 small GTPase via direct interaction with the viral replication protein. Deletion of Rab5 orthologs in a yeast model host or expression of dominant negative mutants of plant Rab5 greatly decreases TBSV replication and prevents the redistribution of PE to the sites of viral replication. We also show that enrichment of PE in the viral replication compartment is assisted by actin filaments. Interestingly, the closely related Carnation Italian ringspot virus, which replicates on the boundary membrane of mitochondria, uses a similar strategy to the peroxisomal TBSV to hijack the Rab5-positive endosomes into the viral replication compartments. Altogether, usurping the GTP-Rab5–positive endosomes allows TBSV to build a PE-enriched viral replication compartment, which is needed to support peak-level replication. Thus, the Rab family of small GTPases includes critical host factors assisting VRC assembly and genesis of the viral replication compartment. PMID:27760128

  11. Identification, replication, and fine-mapping of Loci associated with adult height in individuals of african ancestry.

    Directory of Open Access Journals (Sweden)

    Amidou N'Diaye

    2011-10-01

    Full Text Available Adult height is a classic polygenic trait of high heritability (h(2 approximately 0.8. More than 180 single nucleotide polymorphisms (SNPs, identified mostly in populations of European descent, are associated with height. These variants convey modest effects and explain approximately10% of the variance in height. Discovery efforts in other populations, while limited, have revealed loci for height not previously implicated in individuals of European ancestry. Here, we performed a meta-analysis of genome-wide association (GWA results for adult height in 20,427 individuals of African ancestry with replication in up to 16,436 African Americans. We found two novel height loci (Xp22-rs12393627, P = 3.4×10(-12 and 2p14-rs4315565, P = 1.2×10(-8. As a group, height associations discovered in European-ancestry samples replicate in individuals of African ancestry (P = 1.7×10(-4 for overall replication. Fine-mapping of the European height loci in African-ancestry individuals showed an enrichment of SNPs that are associated with expression of nearby genes when compared to the index European height SNPs (P<0.01. Our results highlight the utility of genetic studies in non-European populations to understand the etiology of complex human diseases and traits.

  12. Identification, Replication, and Fine-Mapping of Loci Associated with Adult Height in Individuals of African Ancestry

    Science.gov (United States)

    Ge, Bing; Tayo, Bamidele; Mathias, Rasika A.; Ding, Jingzhong; Nalls, Michael A.; Adeyemo, Adebowale; Adoue, Véronique; Ambrosone, Christine B.; Atwood, Larry; Bandera, Elisa V.; Becker, Lewis C.; Berndt, Sonja I.; Bernstein, Leslie; Blot, William J.; Boerwinkle, Eric; Britton, Angela; Casey, Graham; Chanock, Stephen J.; Demerath, Ellen; Deming, Sandra L.; Diver, W. Ryan; Fox, Caroline; Harris, Tamara B.; Hernandez, Dena G.; Hu, Jennifer J.; Ingles, Sue A.; John, Esther M.; Johnson, Craig; Keating, Brendan; Kittles, Rick A.; Kolonel, Laurence N.; Kritchevsky, Stephen B.; Le Marchand, Loic; Lohman, Kurt; Liu, Jiankang; Millikan, Robert C.; Murphy, Adam; Musani, Solomon; Neslund-Dudas, Christine; North, Kari E.; Nyante, Sarah; Ogunniyi, Adesola; Ostrander, Elaine A.; Papanicolaou, George; Patel, Sanjay; Pettaway, Curtis A.; Press, Michael F.; Redline, Susan; Rodriguez-Gil, Jorge L.; Rotimi, Charles; Rybicki, Benjamin A.; Salako, Babatunde; Schreiner, Pamela J.; Signorello, Lisa B.; Singleton, Andrew B.; Stanford, Janet L.; Stram, Alex H.; Stram, Daniel O.; Strom, Sara S.; Suktitipat, Bhoom; Thun, Michael J.; Witte, John S.; Yanek, Lisa R.; Ziegler, Regina G.; Zheng, Wei; Zhu, Xiaofeng; Zmuda, Joseph M.; Zonderman, Alan B.; Evans, Michele K.; Liu, Yongmei; Becker, Diane M.; Cooper, Richard S.; Pastinen, Tomi; Henderson, Brian E.; Hirschhorn, Joel N.; Lettre, Guillaume; Haiman, Christopher A.

    2011-01-01

    Adult height is a classic polygenic trait of high heritability (h 2 ∼0.8). More than 180 single nucleotide polymorphisms (SNPs), identified mostly in populations of European descent, are associated with height. These variants convey modest effects and explain ∼10% of the variance in height. Discovery efforts in other populations, while limited, have revealed loci for height not previously implicated in individuals of European ancestry. Here, we performed a meta-analysis of genome-wide association (GWA) results for adult height in 20,427 individuals of African ancestry with replication in up to 16,436 African Americans. We found two novel height loci (Xp22-rs12393627, P = 3.4×10−12 and 2p14-rs4315565, P = 1.2×10−8). As a group, height associations discovered in European-ancestry samples replicate in individuals of African ancestry (P = 1.7×10−4 for overall replication). Fine-mapping of the European height loci in African-ancestry individuals showed an enrichment of SNPs that are associated with expression of nearby genes when compared to the index European height SNPs (P<0.01). Our results highlight the utility of genetic studies in non-European populations to understand the etiology of complex human diseases and traits. PMID:21998595

  13. Identification, replication, and fine-mapping of Loci associated with adult height in individuals of african ancestry.

    Science.gov (United States)

    N'Diaye, Amidou; Chen, Gary K; Palmer, Cameron D; Ge, Bing; Tayo, Bamidele; Mathias, Rasika A; Ding, Jingzhong; Nalls, Michael A; Adeyemo, Adebowale; Adoue, Véronique; Ambrosone, Christine B; Atwood, Larry; Bandera, Elisa V; Becker, Lewis C; Berndt, Sonja I; Bernstein, Leslie; Blot, William J; Boerwinkle, Eric; Britton, Angela; Casey, Graham; Chanock, Stephen J; Demerath, Ellen; Deming, Sandra L; Diver, W Ryan; Fox, Caroline; Harris, Tamara B; Hernandez, Dena G; Hu, Jennifer J; Ingles, Sue A; John, Esther M; Johnson, Craig; Keating, Brendan; Kittles, Rick A; Kolonel, Laurence N; Kritchevsky, Stephen B; Le Marchand, Loic; Lohman, Kurt; Liu, Jiankang; Millikan, Robert C; Murphy, Adam; Musani, Solomon; Neslund-Dudas, Christine; North, Kari E; Nyante, Sarah; Ogunniyi, Adesola; Ostrander, Elaine A; Papanicolaou, George; Patel, Sanjay; Pettaway, Curtis A; Press, Michael F; Redline, Susan; Rodriguez-Gil, Jorge L; Rotimi, Charles; Rybicki, Benjamin A; Salako, Babatunde; Schreiner, Pamela J; Signorello, Lisa B; Singleton, Andrew B; Stanford, Janet L; Stram, Alex H; Stram, Daniel O; Strom, Sara S; Suktitipat, Bhoom; Thun, Michael J; Witte, John S; Yanek, Lisa R; Ziegler, Regina G; Zheng, Wei; Zhu, Xiaofeng; Zmuda, Joseph M; Zonderman, Alan B; Evans, Michele K; Liu, Yongmei; Becker, Diane M; Cooper, Richard S; Pastinen, Tomi; Henderson, Brian E; Hirschhorn, Joel N; Lettre, Guillaume; Haiman, Christopher A

    2011-10-01

    Adult height is a classic polygenic trait of high heritability (h(2) approximately 0.8). More than 180 single nucleotide polymorphisms (SNPs), identified mostly in populations of European descent, are associated with height. These variants convey modest effects and explain approximately10% of the variance in height. Discovery efforts in other populations, while limited, have revealed loci for height not previously implicated in individuals of European ancestry. Here, we performed a meta-analysis of genome-wide association (GWA) results for adult height in 20,427 individuals of African ancestry with replication in up to 16,436 African Americans. We found two novel height loci (Xp22-rs12393627, P = 3.4×10(-12) and 2p14-rs4315565, P = 1.2×10(-8)). As a group, height associations discovered in European-ancestry samples replicate in individuals of African ancestry (P = 1.7×10(-4) for overall replication). Fine-mapping of the European height loci in African-ancestry individuals showed an enrichment of SNPs that are associated with expression of nearby genes when compared to the index European height SNPs (P<0.01). Our results highlight the utility of genetic studies in non-European populations to understand the etiology of complex human diseases and traits.

  14. Identification, replication, and fine-mapping of Loci associated with adult height in individuals of african ancestry.

    Directory of Open Access Journals (Sweden)

    Amidou N'Diaye

    2011-10-01

    Full Text Available Adult height is a classic polygenic trait of high heritability (h(2 approximately 0.8. More than 180 single nucleotide polymorphisms (SNPs, identified mostly in populations of European descent, are associated with height. These variants convey modest effects and explain approximately10% of the variance in height. Discovery efforts in other populations, while limited, have revealed loci for height not previously implicated in individuals of European ancestry. Here, we performed a meta-analysis of genome-wide association (GWA results for adult height in 20,427 individuals of African ancestry with replication in up to 16,436 African Americans. We found two novel height loci (Xp22-rs12393627, P = 3.4×10(-12 and 2p14-rs4315565, P = 1.2×10(-8. As a group, height associations discovered in European-ancestry samples replicate in individuals of African ancestry (P = 1.7×10(-4 for overall replication. Fine-mapping of the European height loci in African-ancestry individuals showed an enrichment of SNPs that are associated with expression of nearby genes when compared to the index European height SNPs (P<0.01. Our results highlight the utility of genetic studies in non-European populations to understand the etiology of complex human diseases and traits.

  15. On Node Replication Attack in Wireless Sensor Networks

    Directory of Open Access Journals (Sweden)

    Mumtaz Qabulio

    2016-04-01

    Full Text Available WSNs (Wireless Sensor Networks comprise a large number of small, inexpensive, low power and memory constrained sensing devices (called sensor nodes that are densely deployed to measure a given physical phenomenon. Since WSNs are commonly deployed in a hostile and unattended environment, it is easy for an adversary to physically capture one or more legitimate sensor nodes, re-program and redeploy them in the network. As a result, the adversary becomes able to deploy several identical copies of physically captured nodes in the network in order to perform illegitimate activities. This type of attack is referred to as Node Replication Attack or Clone Node Attack. By launching node replication attack, an adversary can easily get control on the network which consequently is the biggest threat to confidentiality, integrity and availability of data and services. Thus, detection and prevention of node replication attack in WSNs has become an active area of research and to date more than two dozen schemes have been proposed, which address this issue. In this paper, we present a comprehensive review, classification and comparative analysis of twenty five of these schemes which help to detect and/or prevent node replication attack in WSNs

  16. RNA interference-mediated inhibition of Hepatitis B Virus replication

    Institute of Scientific and Technical Information of China (English)

    TANG Ni; ZHANG Bingqiang; YAN Ge; PU Dan; GAO Xiaolin; Tong-Chuan He; HUANG Ailong

    2004-01-01

    Persistent and recurrent infection of hepatitis B virus (HBV) represents one of the most common and severe viral infections of humans, and has caused a formidable health problem in the affected countries. Currently used antiviral drugs have a very limited success on controlling HBV replication and infection. RNA interference (RNAi), a process by which double-stranded RNA (dsRNA) directs sequence-specific degradation of target mRNA in mammalian and plant cells, has recently been used to knockdown gene expression in various species. In this study, we sought to determine whether RNAi-mediated silencing of HBV viral gene expression could lead to the effective inhibition of HBV replication. We first developed RNAi vectors that expressed small interfering RNA (siRNA) and targeted the HBV core or surface gene sequence. Our results demonstrated that these specific siRNAs efficiently reduced the levels of corresponding viral RNAs and proteins, and thus suppressed viral replication. Treatment with siRNA gave the greatest reduction in the levels of HBsAg (92%) and in HBeAg (85%) respectively in the cultured cell medium. Our findings further demonstrated that the RNAi-mediated antiviral effect was sequence-specific and dose-dependent. Therefore, our findings strongly suggest that RNAi-mediated silencing of HBV viral genes could effectively inhibit the replication of HBV, hence RNAi-based strategy should be further explored as a more efficacious antiviral therapy of HBV infection.

  17. 18β-glycyrrhetinic acid inhibits rotavirus replication in culture

    Directory of Open Access Journals (Sweden)

    Hardy Michele E

    2012-05-01

    Full Text Available Abstract Background Glycyrrhizin (GA and primary metabolite 18β-glycyrrhetinic acid (GRA are pharmacologically active components of the medicinal licorice root, and both have been shown to have antiviral and immunomodulatory properties. Although these properties are well established, the mechanisms of action are not completely understood. In this study, GA and GRA were tested for the ability to inhibit rotavirus replication in cell culture, toward a long term goal of discovering natural compounds that may complement existing vaccines. Methods Epithelial cells were treated with GA or GRA various times pre- or post-infection and virus yields were measured by immunofluorescent focus assay. Levels of viral proteins VP2, VP6, and NSP2 in GRA treated cells were measured by immunoblot to determine if there was an effect of GRA treatment on the accumulation of viral protein. Results GRA treatment reduced rotavirus yields by 99% when added to infected cultures post-- virus adsorption, whereas virus yields in GA treated cultures were similar to mock treated controls. Time of addition experiments indicated that GRA-mediated replication inhibition likely occurs at a step or steps subsequent to virus entry. The amounts of VP2, VP6 and NSP2 were substantially reduced when GRA was added to cultures up to two hours post-entry. Conclusions GRA, but not GA, has significant antiviral activity against rotavirus replication in vitro, and studies to determine whether GRA attenuates rotavirus replication in vivo are underway.

  18. The LMO2 oncogene regulates DNA replication in hematopoietic cells.

    Science.gov (United States)

    Sincennes, Marie-Claude; Humbert, Magali; Grondin, Benoît; Lisi, Véronique; Veiga, Diogo F T; Haman, André; Cazaux, Christophe; Mashtalir, Nazar; Affar, El Bachir; Verreault, Alain; Hoang, Trang

    2016-02-02

    Oncogenic transcription factors are commonly activated in acute leukemias and subvert normal gene expression networks to reprogram hematopoietic progenitors into preleukemic stem cells, as exemplified by LIM-only 2 (LMO2) in T-cell acute lymphoblastic leukemia (T-ALL). Whether or not these oncoproteins interfere with other DNA-dependent processes is largely unexplored. Here, we show that LMO2 is recruited to DNA replication origins by interaction with three essential replication enzymes: DNA polymerase delta (POLD1), DNA primase (PRIM1), and minichromosome 6 (MCM6). Furthermore, tethering LMO2 to synthetic DNA sequences is sufficient to transform these sequences into origins of replication. We next addressed the importance of LMO2 in erythroid and thymocyte development, two lineages in which cell cycle and differentiation are tightly coordinated. Lowering LMO2 levels in erythroid progenitors delays G1-S progression and arrests erythropoietin-dependent cell growth while favoring terminal differentiation. Conversely, ectopic expression in thymocytes induces DNA replication and drives these cells into cell cycle, causing differentiation blockade. Our results define a novel role for LMO2 in directly promoting DNA synthesis and G1-S progression.

  19. DBR1 siRNA inhibition of HIV-1 replication

    Directory of Open Access Journals (Sweden)

    Naidu Yathi

    2005-10-01

    Full Text Available Abstract Background HIV-1 and all retroviruses are related to retroelements of simpler organisms such as the yeast Ty elements. Recent work has suggested that the yeast retroelement Ty1 replicates via an unexpected RNA lariat intermediate in cDNA synthesis. The putative genomic RNA lariat intermediate is formed by a 2'-5' phosphodiester bond, like that found in pre-mRNA intron lariats and it facilitates the minus-strand template switch during cDNA synthesis. We hypothesized that HIV-1 might also form a genomic RNA lariat and therefore that siRNA-mediated inhibition of expression of the human RNA lariat de-branching enzyme (DBR1 expression would specifically inhibit HIV-1 replication. Results We designed three short interfering RNA (siRNA molecules targeting DBR1, which were capable of reducing DBR1 mRNA expression by 80% and did not significantly affect cell viability. We assessed HIV-1 replication in the presence of DBR1 siRNA and found that DBR1 knockdown led to decreases in viral cDNA and protein production. These effects could be reversed by cotransfection of a DBR1 cDNA indicating that the inhibition of HIV-1 replication was a specific effect of DBR1 underexpression. Conclusion These data suggest that DBR1 function may be needed to debranch a putative HIV-1 genomic RNA lariat prior to completion of reverse transcription.

  20. Ultrashort pulsed laser treatment inactivates viruses by inhibiting viral replication and transcription in the host nucleus.

    Science.gov (United States)

    Tsen, Shaw-Wei D; Chapa, Travis; Beatty, Wandy; Xu, Baogang; Tsen, Kong-Thon; Achilefu, Samuel

    2014-10-01

    Ultrashort pulsed laser irradiation is a new method for virus reduction in pharmaceuticals and blood products. Current evidence suggests that ultrashort pulsed laser irradiation inactivates viruses through an impulsive stimulated Raman scattering process, resulting in aggregation of viral capsid proteins. However, the specific functional defect(s) in viruses inactivated in this manner have not been demonstrated. This information is critical for the optimization and the extension of this treatment platform to other applications. Toward this goal, we investigated whether viral internalization, replication, or gene expression in cells were altered by ultrashort pulsed laser irradiation. Murine Cytomegalovirus (MCMV), an enveloped DNA virus, was used as a model virus. Using electron and fluorescence microscopy, we found that laser-treated MCMV virions successfully internalized in cells, as evidenced by the detection of intracellular virions, which was confirmed by the detection of intracellular viral DNA via PCR. Although the viral DNA itself remained polymerase-amplifiable after laser treatment, no viral replication or gene expression was observed in cells infected with laser-treated virus. These results, along with evidence from previous studies, support a model whereby the laser treatment stabilizes the capsid, which inhibits capsid uncoating within cells. By targeting the mechanical properties of viral capsids, ultrashort pulsed laser treatment represents a unique potential strategy to overcome viral mutational escape, with implications for combatting emerging or drug-resistant pathogens.

  1. The replication origin of a repABC plasmid

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    Cevallos Miguel A

    2011-06-01

    Full Text Available Abstract Background repABC operons are present on large, low copy-number plasmids and on some secondary chromosomes in at least 19 α-proteobacterial genera, and are responsible for the replication and segregation properties of these replicons. These operons consist, with some variations, of three genes: repA, repB, and repC. RepA and RepB are involved in plasmid partitioning and in the negative regulatio