The fork and the kinase: a DNA replication tale from a CHK1 perspective.

Science.gov (United States)

González Besteiro, Marina A; Gottifredi, Vanesa

2015-01-01

Replication fork progression is being continuously hampered by exogenously introduced and naturally occurring DNA lesions and other physical obstacles. Checkpoint kinase 1 (Chk1) is activated at replication forks that encounter damaged DNA. Subsequently, Chk1 inhibits the initiation of new replication factories and stimulates the firing of dormant origins (those in the vicinity of stalled forks). Chk1 also avoids fork collapse into DSBs (double strand breaks) and promotes fork elongation. At the molecular level, the current model considers stalled forks as the site of Chk1 activation and the nucleoplasm as the location where Chk1 phosphorylates target proteins. This model certainly serves to explain how Chk1 modulates origin firing, but how Chk1 controls the fate of stalled forks is less clear. Interestingly, recent reports demonstrating that Chk1 phosphorylates chromatin-bound proteins and even holds kinase-independent functions might shed light on how Chk1 contributes to the elongation of damaged DNA. Indeed, such findings have unveiled a puzzling connection between Chk1 and DNA lesion bypass, which might be central to promoting fork elongation and checkpoint attenuation. In summary, Chk1 is a multifaceted and versatile signaling factor that acts at ongoing forks and replication origins to determine the extent and quality of the cellular response to replication stress. Copyright © 2014 Elsevier B.V. All rights reserved.

Replication stress, a source of epigenetic aberrations in cancer?

DEFF Research Database (Denmark)

Jasencakova, Zusana; Groth, Anja

2010-01-01

. Chromatin organization is transiently disrupted during DNA replication and maintenance of epigenetic information thus relies on faithful restoration of chromatin on the new daughter strands. Acute replication stress challenges proper chromatin restoration by deregulating histone H3 lysine 9 mono......-methylation on new histones and impairing parental histone recycling. This could facilitate stochastic epigenetic silencing by laying down repressive histone marks at sites of fork stalling. Deregulation of replication in response to oncogenes and other tumor-promoting insults is recognized as a significant source...... of genome instability in cancer. We propose that replication stress not only presents a threat to genome stability, but also jeopardizes chromatin integrity and increases epigenetic plasticity during tumorigenesis....

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...

Replication Catastrophe

DEFF Research Database (Denmark)

Toledo, Luis; Neelsen, Kai John; Lukas, Jiri

2017-01-01

Proliferating cells rely on the so-called DNA replication checkpoint to ensure orderly completion of genome duplication, and its malfunction may lead to catastrophic genome disruption, including unscheduled firing of replication origins, stalling and collapse of replication forks, massive DNA...... breakage, and, ultimately, cell death. Despite many years of intensive research into the molecular underpinnings of the eukaryotic replication checkpoint, the mechanisms underlying the dismal consequences of its failure remain enigmatic. A recent development offers a unifying model in which the replication...... checkpoint guards against global exhaustion of rate-limiting replication regulators. Here we discuss how such a mechanism can prevent catastrophic genome disruption and suggest how to harness this knowledge to advance therapeutic strategies to eliminate cancer cells that inherently proliferate under...

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...... as a versatile, site-specific, heterologous DNA replication-perturbing system, with a variety of potential applications....

Functions of Ubiquitin and SUMO in DNA Replication and Replication Stress

Science.gov (United States)

García-Rodríguez, Néstor; Wong, Ronald P.; Ulrich, Helle D.

2016-01-01

Complete and faithful duplication of its entire genetic material is one of the essential prerequisites for a proliferating cell to maintain genome stability. Yet, during replication DNA is particularly vulnerable to insults. On the one hand, lesions in replicating DNA frequently cause a stalling of the replication machinery, as most DNA polymerases cannot cope with defective templates. This situation is aggravated by the fact that strand separation in preparation for DNA synthesis prevents common repair mechanisms relying on strand complementarity, such as base and nucleotide excision repair, from working properly. On the other hand, the replication process itself subjects the DNA to a series of hazardous transformations, ranging from the exposure of single-stranded DNA to topological contortions and the generation of nicks and fragments, which all bear the risk of inducing genomic instability. Dealing with these problems requires rapid and flexible responses, for which posttranslational protein modifications that act independently of protein synthesis are particularly well suited. Hence, it is not surprising that members of the ubiquitin family, particularly ubiquitin itself and SUMO, feature prominently in controlling many of the defensive and restorative measures involved in the protection of DNA during replication. In this review we will discuss the contributions of ubiquitin and SUMO to genome maintenance specifically as they relate to DNA replication. We will consider cases where the modifiers act during regular, i.e., unperturbed stages of replication, such as initiation, fork progression, and termination, but also give an account of their functions in dealing with lesions, replication stalling and fork collapse. PMID:27242895

Wider stall space affects behavior, lesion scores, and productivity of gestating sows.

Science.gov (United States)

Salak-Johnson, J L; DeDecker, A E; Levitin, H A; McGarry, B M

2015-10-01

Limited space allowance within the standard gestation stall is an important welfare concern because it restricts the ability of the sow to make postural adjustments and hinders her ability to perform natural behaviors. Therefore, we evaluated the impacts of increasing stall space and/or providing sows the freedom to access a small pen area on sow well-being using multiple welfare metrics. A total of 96 primi- and multiparous crossbred sows were randomly assigned in groups of 4 sows/treatment across 8 replicates to 1 of 3 stall treatments (TRT): standard stall (CTL; dimensions: 61 by 216 cm), width-adjustable stall (flex stall [FLX]; dimensions: adjustable width of 56 to 79 cm by 216 cm), or an individual walk-in/lock-in stall with access to a small communal open-pen area at the rear of the stall (free-access stall [FAS]; dimensions: 69 by 226 cm). Lesion scores, behavior, and immune and productivity traits were measured at various gestational days throughout the study. Total lesion scores were greatest for sows in FAS and least for sows in FLX ( pregnancy progressed, lesion scores increased among sows in CTL ( postural behaviors and sham chew behavior were affected by TRT ( changes in postural behaviors, lesion severity scores, and other sow traits. Moreover, compromised welfare measures found among sows in various stall environments may be partly attributed to the specific constraints of each stall system such as restricted stall space in CTL, insufficient floor space in the open-pen area of the FAS system, and gate design of the FLX (e.g., direction of bars and feeder space). These results also indicate that parity and gestational day are additional factors that may exacerbate the effects of restricted stall space or insufficient pen space, further compromising sow well-being.

Molecular basis for PrimPol recruitment to replication forks by RPA.

Science.gov (United States)

Guilliam, Thomas A; Brissett, Nigel C; Ehlinger, Aaron; Keen, Benjamin A; Kolesar, Peter; Taylor, Elaine M; Bailey, Laura J; Lindsay, Howard D; Chazin, Walter J; Doherty, Aidan J

2017-05-23

DNA damage and secondary structures can stall the replication machinery. Cells possess numerous tolerance mechanisms to complete genome duplication in the presence of such impediments. In addition to translesion synthesis (TLS) polymerases, most eukaryotic cells contain a multifunctional replicative enzyme called primase-polymerase (PrimPol) that is capable of directly bypassing DNA damage by TLS, as well as repriming replication downstream of impediments. Here, we report that PrimPol is recruited to reprime through its interaction with RPA. Using biophysical and crystallographic approaches, we identify that PrimPol possesses two RPA-binding motifs and ascertained the key residues required for these interactions. We demonstrate that one of these motifs is critical for PrimPol's recruitment to stalled replication forks in vivo. In addition, biochemical analysis reveals that RPA serves to stimulate the primase activity of PrimPol. Together, these findings provide significant molecular insights into PrimPol's mode of recruitment to stalled forks to facilitate repriming and restart.

Overcoming natural replication barriers: differential helicase requirements.

Science.gov (United States)

Anand, Ranjith P; Shah, Kartik A; Niu, Hengyao; Sung, Patrick; Mirkin, Sergei M; Freudenreich, Catherine H

2012-02-01

DNA sequences that form secondary structures or bind protein complexes are known barriers to replication and potential inducers of genome instability. In order to determine which helicases facilitate DNA replication across these barriers, we analyzed fork progression through them in wild-type and mutant yeast cells, using 2-dimensional gel-electrophoretic analysis of the replication intermediates. We show that the Srs2 protein facilitates replication of hairpin-forming CGG/CCG repeats and prevents chromosome fragility at the repeat, whereas it does not affect replication of G-quadruplex forming sequences or a protein-bound repeat. Srs2 helicase activity is required for hairpin unwinding and fork progression. Also, the PCNA binding domain of Srs2 is required for its in vivo role of replication through hairpins. In contrast, the absence of Sgs1 or Pif1 helicases did not inhibit replication through structural barriers, though Pif1 did facilitate replication of a telomeric protein barrier. Interestingly, replication through a protein barrier but not a DNA structure barrier was modulated by nucleotide pool levels, illuminating a different mechanism by which cells can regulate fork progression through protein-mediated stall sites. Our analyses reveal fundamental differences in the replication of DNA structural versus protein barriers, with Srs2 helicase activity exclusively required for fork progression through hairpin structures.

Development of tooling suitable for stall regulated blades

Energy Technology Data Exchange (ETDEWEB)

Hancock, M.

2001-07-01

The objectives of the project were to make significant improvements in the production of stall regulated blades in the areas of (a) the tip box, its housing, its mechanism and small GRP parts; (b) mould technology; (c) resins and glues and (d) root tooling. Although wood composite had been identified as a competitive technology for blades, compared with GRP blades, production volumes had been lower; reasons are given. The way in which the four areas identified for investigation were tackled are discussed. The study showed that the mould cycle time can be reduced to two days for a stall regulated blade and the blade quality can be improved by using the composite tip box and new resins. The time required for replication of moulds can be reduced by 40%.

Managing Single-Stranded DNA during Replication Stress in Fission Yeast

Directory of Open Access Journals (Sweden)

Sarah A. Sabatinos

2015-09-01

Full Text Available Replication fork stalling generates a variety of responses, most of which cause an increase in single-stranded DNA. ssDNA is a primary signal of replication distress that activates cellular checkpoints. It is also a potential source of genome instability and a substrate for mutation and recombination. Therefore, managing ssDNA levels is crucial to chromosome integrity. Limited ssDNA accumulation occurs in wild-type cells under stress. In contrast, cells lacking the replication checkpoint cannot arrest forks properly and accumulate large amounts of ssDNA. This likely occurs when the replication fork polymerase and helicase units are uncoupled. Some cells with mutations in the replication helicase (mcm-ts mimic checkpoint-deficient cells, and accumulate extensive areas of ssDNA to trigger the G2-checkpoint. Another category of helicase mutant (mcm4-degron causes fork stalling in early S-phase due to immediate loss of helicase function. Intriguingly, cells realize that ssDNA is present, but fail to detect that they accumulate ssDNA, and continue to divide. Thus, the cellular response to replication stalling depends on checkpoint activity and the time that replication stress occurs in S-phase. In this review we describe the signs, signals, and symptoms of replication arrest from an ssDNA perspective. We explore the possible mechanisms for these effects. We also advise the need for caution when detecting and interpreting data related to the accumulation of ssDNA.

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.

A Molecular Toolbox to Engineer Site-Specific DNA Replication Perturbation.

Science.gov (United States)

Larsen, Nicolai B; Hickson, Ian D; Mankouri, Hocine W

2018-01-01

Site-specific arrest of DNA replication is a useful tool for analyzing cellular responses to DNA replication perturbation. The E. coli Tus-Ter replication barrier can be reconstituted in eukaryotic cells as a system to engineer an unscheduled collision between a replication fork and an "alien" impediment to DNA replication. To further develop this system as a versatile tool, we describe a set of reagents and a detailed protocol that can be used to engineer Tus-Ter barriers into any locus in the budding yeast genome. Because the Tus-Ter complex is a bipartite system with intrinsic DNA replication-blocking activity, the reagents and protocols developed and validated in yeast could also be optimized to engineer site-specific replication fork barriers into other eukaryotic cell types.

A Molecular Toolbox to Engineer Site-Specific DNA Replication Perturbation

DEFF Research Database (Denmark)

Larsen, Nicolai B; Hickson, Ian D; Mankouri, Hocine W

2018-01-01

" impediment to DNA replication. To further develop this system as a versatile tool, we describe a set of reagents and a detailed protocol that can be used to engineer Tus-Ter barriers into any locus in the budding yeast genome. Because the Tus-Ter complex is a bipartite system with intrinsic DNA replication......Site-specific arrest of DNA replication is a useful tool for analyzing cellular responses to DNA replication perturbation. The E. coli Tus-Ter replication barrier can be reconstituted in eukaryotic cells as a system to engineer an unscheduled collision between a replication fork and an "alien......-blocking activity, the reagents and protocols developed and validated in yeast could also be optimized to engineer site-specific replication fork barriers into other eukaryotic cell types....

Anaphase onset before complete DNA replication with intact checkpoint responses

DEFF Research Database (Denmark)

Torres-Rosell, Jordi; De Piccoli, Giacomo; Cordon-Preciado, Violeta

2007-01-01

Cellular checkpoints prevent mitosis in the presence of stalled replication forks. Whether checkpoints also ensure the completion of DNA replication before mitosis is unknown. Here, we show that in yeast smc5-smc6 mutants, which are related to cohesin and condensin, replication is delayed, most...

Education stalls and subsequent stalls in African fertility: A descriptive overview

Directory of Open Access Journals (Sweden)

Anne Goujon

2015-12-01

Full Text Available Background: Recent stalls in fertility decline have been observed in a few countries in sub-Saharan Africa, and so far no plausible common reason has been identified in the literature. This paper develops the hypothesis that these fertility stalls could be associated with stalls in the progress of education among the women of the relevant cohorts, possibly resulting partly from the Structural Adjustment Programs (SAPs of the 1980s. Methods: We descriptively link the change in the education composition of successive cohorts of young women in sub-Saharan Africa and the recent fertility stalls. We use reconstructed data on population by age, gender, and level of education from www.wittgenstein centre.org/dataexplorer, and fertility rates from the United Nations. Results: In most sub-Saharan African countries, we observe that the same countries that had fertility stalls had a stall in the progress of education, particularly for young women who were of primary school age during the 1980s, when most of the countries were under structural adjustment. Conversely, stalls in fertility are less common in countries that did not have an education stall, possibly in relation to SAPs. Conclusions: The results point to the possibility of a link between the recent fertility stalls and discontinuities in the improvement of the education of the relevant cohorts, which in turn could be related to the SAPs in the 1980s. This descriptive finding now needs to be corroborated through more detailed cohort-specific fertility analysis. If the education-fertility link can be further established, it will have important implications for the projections of population growth in affected countries.

Genomic mapping of single-stranded DNA in hydroxyurea-challenged yeasts identifies origins of replication.

Science.gov (United States)

Feng, Wenyi; Collingwood, David; Boeck, Max E; Fox, Lindsay A; Alvino, Gina M; Fangman, Walton L; Raghuraman, Mosur K; Brewer, Bonita J

2006-02-01

During DNA replication one or both strands transiently become single stranded: first at the sites where initiation of DNA synthesis occurs (known as origins of replication) and subsequently on the lagging strands of replication forks as discontinuous Okazaki fragments are generated. We report a genome-wide analysis of single-stranded DNA (ssDNA) formation in the presence of hydroxyurea during DNA replication in wild-type and checkpoint-deficient rad53 Saccharomyces cerevisiae cells. In wild-type cells, ssDNA was first observed at a subset of replication origins and later 'migrated' bi-directionally, suggesting that ssDNA formation is associated with continuously moving replication forks. In rad53 cells, ssDNA was observed at virtually every known origin, but remained there over time, suggesting that replication forks stall. Telomeric regions seemed to be particularly sensitive to the loss of Rad53 checkpoint function. Replication origins in Schizosaccharomyces pombe were also mapped using our method.

Observations of the Growth and Decay of Stall Cells during Stall and Surge in an Axial Compressor

Directory of Open Access Journals (Sweden)

Adam R. Hickman

2017-01-01

Full Text Available This research investigated unsteady events such as stall inception, stall-cell development, and surge. Stall is characterized by a decrease in overall pressure rise and nonaxisymmetric throughflow. Compressor stall can lead to surge which is characterized by quasi-axisymmetric fluctuations in mass flow and pressure. Unsteady measurements of the flow field around the compressor rotor are examined. During the stall inception process, initial disturbances were found within the rotor passage near the tip region. As the stall cell develops, blade lift and pressure ratio decrease within the stall cell and increase ahead of the stall cell. The stall inception event, stall-cell development, and stall recovery event were found to be nearly identical for stable rotating stall and surge cases. As the stall cell grows, the leading edge of the cell will rotate at a higher rate than the trailing edge in the rotor frame. The opposite occurs during stall recovery. The trailing edge of the stall cell will rotate at the approximate speed as the fully developed stall cell, while the leading edge decreases in rotational speed in the rotor frame.

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

Science.gov (United States)

Larsen, Nicolai B; Sass, Ehud; Suski, Catherine; Mankouri, Hocine W; Hickson, Ian D

2014-04-07

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 Tof1-dependent nor counteracted by the Rrm3 helicase. Although the yeast replisome can overcome RF pausing at Tus-Ter modules, this event triggers site-specific homologous recombination that requires the RecQ helicase, Sgs1, for its timely resolution. We propose that Tus-Ter can be utilized as a versatile, site-specific, heterologous DNA replication-perturbing system, with a variety of potential applications.

DVC1 (C1orf124) is a DNA damage-targeting p97 adaptor that promotes ubiquitin-dependent responses to replication blocks

DEFF Research Database (Denmark)

Mosbech, Anna; Gibbs-Seymour, Ian; Kagias, Konstantinos

2012-01-01

Ubiquitin-mediated processes orchestrate critical DNA-damage signaling and repair pathways. We identify human DVC1 (C1orf124; Spartan) as a cell cycle-regulated anaphase-promoting complex (APC) substrate that accumulates at stalled replication forks. DVC1 recruitment to sites of replication stress...... synthesis (TLS) DNA polymerase η (Pol η) from monoubiquitylated PCNA. DVC1 knockdown enhances UV light-induced mutagenesis, and depletion of human DVC1 or the Caenorhabditis elegans ortholog DVC-1 causes hypersensitivity to replication stress-inducing agents. Our findings establish DVC1 as a DNA damage...

ATR prohibits replication catastrophe by preventing global exhaustion of RPA.

Science.gov (United States)

Toledo, Luis Ignacio; Altmeyer, Matthias; Rask, Maj-Britt; Lukas, Claudia; Larsen, Dorthe Helena; Povlsen, Lou Klitgaard; Bekker-Jensen, Simon; Mailand, Niels; Bartek, Jiri; Lukas, Jiri

2013-11-21

ATR, activated by replication stress, protects replication forks locally and suppresses origin firing globally. Here, we show that these functions of ATR are mechanistically coupled. Although initially stable, stalled forks in ATR-deficient cells undergo nucleus-wide breakage after unscheduled origin firing generates an excess of single-stranded DNA that exhausts the nuclear pool of RPA. Partial reduction of RPA accelerated fork breakage, and forced elevation of RPA was sufficient to delay such "replication catastrophe" even in the absence of ATR activity. Conversely, unscheduled origin firing induced breakage of stalled forks even in cells with active ATR. Thus, ATR-mediated suppression of dormant origins shields active forks against irreversible breakage via preventing exhaustion of nuclear RPA. This study elucidates how replicating genomes avoid destabilizing DNA damage. Because cancer cells commonly feature intrinsically high replication stress, this study also provides a molecular rationale for their hypersensitivity to ATR inhibitors. Copyright © 2013 Elsevier Inc. All rights reserved.

FANCJ couples replication past natural fork barriers with maintenance of chromatin structure.

Science.gov (United States)

Schwab, Rebekka A; Nieminuszczy, Jadwiga; Shin-ya, Kazuo; Niedzwiedz, Wojciech

2013-04-01

Defective DNA repair causes Fanconi anemia (FA), a rare childhood cancer-predisposing syndrome. At least 15 genes are known to be mutated in FA; however, their role in DNA repair remains unclear. Here, we show that the FANCJ helicase promotes DNA replication in trans by counteracting fork stalling on replication barriers, such as G4 quadruplex structures. Accordingly, stabilization of G4 quadruplexes in ΔFANCJ cells restricts fork movements, uncouples leading- and lagging-strand synthesis and generates small single-stranded DNA gaps behind the fork. Unexpectedly, we also discovered that FANCJ suppresses heterochromatin spreading by coupling fork movement through replication barriers with maintenance of chromatin structure. We propose that FANCJ plays an essential role in counteracting chromatin compaction associated with unscheduled replication fork stalling and restart, and suppresses tumorigenesis, at least partially, in this replication-specific manner.

Transition Process from Diffuser Stall to Stage Stall in a Centrifugal Compressor with a Vaned Diffuser

Directory of Open Access Journals (Sweden)

Nobumichi Fujisawa

2017-01-01

Full Text Available The transition process from a diffuser rotating stall to a stage stall in a centrifugal compressor with a vaned diffuser was investigated by experimental and numerical analyses. From the velocity measurements, it was found that the rotating stall existed on the shroud side of the diffuser passage in the off-design flow condition. The numerical results revealed the typical vortical structure of the diffuser stall. The diffuser stall cell was caused by the systematic vortical structure which consisted of the tornado-type vortex, the longitudinal vortex at the shroud/suction surface corner (i.e., leading edge vortex (LEV, and the vortex in the throat area of the diffuser passages. Furthermore, the stage stall, which rotated within both the impeller and diffuser passages, occurred instead of the diffuser stall as the mass flow rate was decreased. According to the velocity measurements at the diffuser inlet, the diffuser stall which rotated on the shroud side was shifted to the hub side. Then, the diffuser stall moved into the impeller passages and formed the stage stall. Therefore, the stage stall was caused by the development of the diffuser stall, which transferred from the shroud side to the hub side in the vaneless space and expanded to the impeller passages.

The relevance of the dynamic stall effect for transient fault operations of active-stall wind turbines

Energy Technology Data Exchange (ETDEWEB)

Jauch, Clemens; Soerensen, Poul; Jensen, Birgitte Bak

2005-06-15

This article describes a methodology to quantify the influence of dynamic stall on transient fault operations of active-stall turbines. The model of the dynamic stall effect is introduced briefly. The behaviour of the dynamic stall model during a transient fault operation is described mathematically, and from this its effect quantified. Two quantities are chosen to describe the influence of the dynamic stall effect: one is active power and the other is time delay. Subsequently a transient fault scenario is simulated with and without the dynamic stall effect and the differences discussed. From this comparison, the conclusion is drawn that the dynamic stall effect has some influence on the post-fault behaviour of the wind turbine, and it is hence suggested that the dynamic stall effect is considered if an active-stall wind turbine is to be modelled realistically. (Author)

Comfort zone-design free stalls: do they influence the stall use behavior of lame cows?

Science.gov (United States)

Cook, N B; Marin, M J; Mentink, R L; Bennett, T B; Schaefer, M J

2008-12-01

The behavior of 59 cows in 4 herds, each with Comfort Zone-design free stalls with dimensions suitable for 700-kg, mature Holstein dairy cows, was filmed for a 48-h period. Comparison was made between nonlame, slightly lame, and moderately lame cows on either rubber-crumb-filled mattress stall surfaces bedded with a small amount of sawdust (2 herds) or a Pack Mat design, which consisted of a rubber-crumb-filled mattress pad installed 5 cm below a raised rear curb, bedded with 5 to 8 cm of sand bedding (2 herds). All other stall design components were similar. Despite adequate resting space and freedom to perform normal rising and lying movements, lame cows on mattresses stood in the stall for >2 h longer than nonlame cows. Although a significant increase in stall standing behavior was observed in lame cows on Pack Mat stalls, the mean (95% confidence interval) standing time in the stall was only 0.7 (0 to 3.0) h/d for nonlame cows and 1.6 (0 to 4.2) h/d for moderately lame cows, which was less than the 2.1 (0 to 4.4), 4.3 (1.6 to 6.9), and 4.9 (2.5 to 7.3) h/d spent standing in the stall for nonlame, slightly lame, and moderately lame cows on mattresses, respectively. This observation supports the hypothesis that it is the nature of the stall surface that dictates changes in stall standing behavior observed in lame cows, rather than other components of stall design. The finding that only 5 to 8 cm of sand over a mattress pad provides most of the benefits of deep sand-bedded stalls, along with other advantages related to stall maintenance and manure handling, gives farmers another useful housing alternative with which to improve cow comfort and well-being.

Recovery of arrested replication forks by homologous recombination is error-prone.

Directory of Open Access Journals (Sweden)

Ismail Iraqui

Full Text Available Homologous recombination is a universal mechanism that allows repair of DNA and provides support for DNA replication. Homologous recombination is therefore a major pathway that suppresses non-homology-mediated genome instability. Here, we report that recovery of impeded replication forks by homologous recombination is error-prone. Using a fork-arrest-based assay in fission yeast, we demonstrate that a single collapsed fork can cause mutations and large-scale genomic changes, including deletions and translocations. Fork-arrest-induced gross chromosomal rearrangements are mediated by inappropriate ectopic recombination events at the site of collapsed forks. Inverted repeats near the site of fork collapse stimulate large-scale genomic changes up to 1,500 times over spontaneous events. We also show that the high accuracy of DNA replication during S-phase is impaired by impediments to fork progression, since fork-arrest-induced mutation is due to erroneous DNA synthesis during recovery of replication forks. The mutations caused are small insertions/duplications between short tandem repeats (micro-homology indicative of replication slippage. Our data establish that collapsed forks, but not stalled forks, recovered by homologous recombination are prone to replication slippage. The inaccuracy of DNA synthesis does not rely on PCNA ubiquitination or trans-lesion-synthesis DNA polymerases, and it is not counteracted by mismatch repair. We propose that deletions/insertions, mediated by micro-homology, leading to copy number variations during replication stress may arise by progression of error-prone replication forks restarted by homologous recombination.

The onset of dynamic stall revisited

Energy Technology Data Exchange (ETDEWEB)

Mulleners, Karen; Raffel, Markus [German Aerospace Center (DLR), Goettingen (Germany)

2012-03-15

Dynamic stall on a helicopter rotor blade comprises a series of complex aerodynamic phenomena in response to the unsteady change of the blade's angle of attack. It is accompanied by a lift overshoot and delayed massive flow separation with respect to static stall. The classical hallmark of the dynamic stall phenomenon is the dynamic stall vortex. The flow over an oscillating OA209 airfoil under dynamic stall conditions was investigated by means of unsteady surface pressure measurements and time-resolved particle image velocimetry. The characteristic features of the unsteady flow field were identified and analysed utilising different coherent structure identification methods. An Eulerian and a Lagrangian procedure were adopted to locate the axes of vortices and the edges of Lagrangian coherent structures, respectively; a proper orthogonal decomposition of the velocity field revealed the energetically dominant coherent flow patterns and their temporal evolution. Based on the complementary information obtained by these methods the dynamics and interaction of vortical structures were analysed within a single dynamic stall life cycle leading to a classification of the unsteady flow development into five successive stages: the attached flow stage; the stall development stage; stall onset; the stalled stage; and flow reattachment. The onset of dynamic stall was specified here based on a characteristic mode of the proper orthogonal decomposition of the velocity field. Variations in the flow field topology that accompany the stall onset were verified by the Lagrangian coherent structure analysis. The instantaneous effective unsteadiness was defined as a single representative parameter to describe the influence of the motion parameters. Dynamic stall onset was found to be promoted by increasing unsteadiness. The mechanism that results in the detachment of the dynamic stall vortex from the airfoil was identified as vortex-induced separation caused by strong viscous

Physical Interaction between Replication Protein A (RPA) and MRN: Involvement of RPA2 Phosphorylation and the N-terminus of RPA1

OpenAIRE

Oakley, Greg; Tillison, Kristin; Opiyo, Stephen; Glanzer, Jason; Horn, Jeffrey M.; Patrick, Steve M.

2009-01-01

Replication protein A (RPA) is a heterotrimeric protein consisting of RPA1, RPA2 and RPA3 subunits that binds to ssDNA with high affinity. The response to replication stress requires the recruitment of RPA and the MRE11/RAD50/NBS1 (MRN) complex. RPA bound to ssDNA stabilizes stalled replication forks by recruiting checkpoint proteins involved in fork stabilization. MRN can bind DNA structures encountered at stalled or collapsed replication forks, such as ssDNA-dsDNA junctions or breaks and pr...

Why do Cross-Flow Turbines Stall?

Science.gov (United States)

Cavagnaro, Robert; Strom, Benjamin; Polagye, Brian

2015-11-01

Hydrokinetic turbines are prone to instability and stall near their peak operating points under torque control. Understanding the physics of turbine stall may help to mitigate this undesirable occurrence and improve the robustness of torque controllers. A laboratory-scale two-bladed cross-flow turbine operating at a chord-based Reynolds number ~ 3 ×104 is shown to stall at a critical tip-speed ratio. Experiments are conducting bringing the turbine to this critical speed in a recirculating current flume by increasing resistive torque and allowing the rotor to rapidly decelerate while monitoring inflow velocity, torque, and drag. The turbine stalls probabilistically with a distribution generated from hundreds of such events. A machine learning algorithm identifies stall events and indicates the effectiveness of available measurements or combinations of measurements as predictors. Bubble flow visualization and PIV are utilized to observe fluid conditions during stall events including the formation, separation, and advection of leading-edge vortices involved in the stall process.

Factors affecting stall use for different freestall bases.

Science.gov (United States)

Wagner-Storch, A M; Palmer, R W; Kammel, D W

2003-06-01

The objective of this study was to compare stall use (stall occupancy and cow position) by barn side for factors affecting stall use. A closed circuit television system recorded stall use four times per day for a 9-mo period starting May 9, 2001. Six factors were analyzed: stall base, distance to water, stall location within stall base section, stall location within barn, inside barn temperature, and length of time cows were exposed to stall bases. Two barn sides with different stocking densities were analyzed: low (66%), with cows milked by robotic milker; and high (100%), with cows milked 2X in parlor. Six stall base types were tested: two mattresses, a waterbed, a rubber mat, concrete, and sand (high side only). The base types were grouped 3 to 7 stalls/section and randomly placed in each row. Cows spent more time in mattress-based stalls, but the highest percentage lying was in sand-based stalls. The following significant stall occupancy percentages were found: sand had the highest percentage of cows lying on the high stocking density side (69%), followed by mattress type 1 (65%) > mattress type 2 (57%) > waterbed (45%) > rubber mat (33%) > concrete (23%). Mattress type 1 had the highest percentage stalls occupied (88%), followed by mattress type 2 (84%) > sand (79%) > soft rubber mat (65%) > waterbed (62%) > concrete (39%). On the low stocking rate side, mattress type 1 had the highest percentage cows lying (45%) and occupied (59.6%), followed by mattress type 2 > waterbed > soft rubber mat > concrete. Cow lying and stalls occupied percentages were highest for stalls 1) not at the end of a section, and 2) on the outside row, and varied by base type for time cows exposed to stalls and inside barn temperature. Lying and occupied percentages were different for different mattress types. The percentage of stalls with cows standing was higher for mat and mattress-based stalls. Results show mattress type 1 and sand to be superior and rubber mats and concrete inferior

Systematic identification of fragile sites via genome-wide location analysis of γ-H2AX

Science.gov (United States)

Szilard, Rachel K.; Jacques, Pierre-Étienne; Laramée, Louise; Cheng, Benjamin; Galicia, Sarah; Bataille, Alain R.; Yeung, ManTek; Mendez, Megan; Bergeron, Maxime; Robert, François; Durocher, Daniel

2011-01-01

Phosphorylation of histone H2AX is an early response to DNA damage in eukaryotes. In Saccharomyces cerevisiae, DNA damage or replication fork stalling results in histone H2A phosphorylation to yield γ-H2A (yeast γ-H2AX) in a Mec1 (ATR)- and Tel1 (ATM)- dependent manner. Here, we describe the genome-wide location analysis of γ-H2A as a strategy to identify loci prone to engage the Mec1 and Tel1 pathways. Remarkably, γ-H2A enrichment overlaps with loci prone to replication fork stalling and is caused by the action of Mec1 and Tel1, indicating that these loci are prone to breakage. Moreover, about half the sites enriched for γ-H2A map to repressed protein-coding genes, and histone deacetylases are necessary for formation of γ-H2A at these loci. Finally, our work indicates that high resolution mapping of γ-H2AX is a fruitful route to map fragile sites in eukaryotic genomes. PMID:20139982

RPA-Binding Protein ETAA1 Is an ATR Activator Involved in DNA Replication Stress Response.

Science.gov (United States)

Lee, Yuan-Cho; Zhou, Qing; Chen, Junjie; Yuan, Jingsong

2016-12-19

ETAA1 (Ewing tumor-associated antigen 1), also known as ETAA16, was identified as a tumor-specific antigen in the Ewing family of tumors. However, the biological function of this protein remains unknown. Here, we report the identification of ETAA1 as a DNA replication stress response protein. ETAA1 specifically interacts with RPA (Replication protein A) via two conserved RPA-binding domains and is therefore recruited to stalled replication forks. Interestingly, further analysis of ETAA1 function revealed that ETAA1 participates in the activation of ATR signaling pathway via a conserved ATR-activating domain (AAD) located near its N terminus. Importantly, we demonstrate that both RPA binding and ATR activation are required for ETAA1 function at stalled replication forks to maintain genome stability. Therefore, our data suggest that ETAA1 is a new ATR activator involved in replication checkpoint control. Copyright © 2016 Elsevier Ltd. All rights reserved.

14 CFR 25.203 - Stall characteristics.

Science.gov (United States)

2010-01-01

... 14 Aeronautics and Space 1 2010-01-01 2010-01-01 false Stall characteristics. 25.203 Section 25.203 Aeronautics and Space FEDERAL AVIATION ADMINISTRATION, DEPARTMENT OF TRANSPORTATION AIRCRAFT AIRWORTHINESS STANDARDS: TRANSPORT CATEGORY AIRPLANES Flight Stalls § 25.203 Stall characteristics. (a) It must...

Effects of three types of free-stall surfaces on preferences and stall usage by dairy cows.

Science.gov (United States)

Tucker, C B; Weary, D M; Fraser, D

2003-02-01

One important criterion in choosing appropriate housing systems for dairy cattle is that the freestall provides a comfortable surface for the cow. This paper describes two experiments testing the effects of commonly used lying surfaces on stall preference and stall usage by Holstein cows. In both experiments, 12 cows were housed individually in separate pens. Each pen contained three free stalls with a different surface: deep-bedded sawdust, deep-bedded sand, and a geotextile mattress covered with 2 to 3 cm of sawdust. The animals were restricted to each surface in turn, in a random order for either 2 (Experiment 1) or 3 d (Experiment 2). Both before and after this restriction phase, the animals were allowed access to all three surfaces, and preference was determined, based on lying times. Of the 12 cows used in Experiment 1, 10 preferred sawdust before and nine after the restriction phase. During the restriction phase, average lying times and number of lying events during the restriction phase were significantly lower for the sand-bedded stalls (P sand bedded stalls. In this experiment, about half the cows preferred sand and half sawdust, after the restriction phase. During the restriction phase of experiment, lying times and number of lying events were lower, and standing times were higher when the animals were restricted to the mattresses compared to either sand or sawdust (P < or = 0.05). These results indicate that (1) free stall surface can affect both stall preferences and stall usage, and (2) mattresses are less preferred.

Stop Stalling: Mus81 Required for Efficient Replication | Center for Cancer Research

Science.gov (United States)

DNA replication is precisely controlled to ensure that daughter cells receive intact, accurate genetic information. Each segment of DNA must be copied only once, and the rate of replication coordinated genome-wide. Mild replication stress slows DNA synthesis and activates a pathway involving the Mus81 endonuclease, which generates a series of DNA breaks that are rapidly repaired, allowing the cell to avoid activating the S-phase checkpoint and its potentially damaging outcomes of apoptosis or error-prone repair. Mirit Aladjem, Ph.D., of CCR’s Developmental Therapeutics Branch, and her colleagues wondered whether Mus81 also plays a role in regulating the replication rate during growth in the absence of stress.

Distinct functions of human RecQ helicases during DNA replication.

Science.gov (United States)

Urban, Vaclav; Dobrovolna, Jana; Janscak, Pavel

2017-06-01

DNA replication is the most vulnerable process of DNA metabolism in proliferating cells and therefore it is tightly controlled and coordinated with processes that maintain genomic stability. Human RecQ helicases are among the most important factors involved in the maintenance of replication fork integrity, especially under conditions of replication stress. RecQ helicases promote recovery of replication forks being stalled due to different replication roadblocks of either exogenous or endogenous source. They prevent generation of aberrant replication fork structures and replication fork collapse, and are involved in proper checkpoint signaling. The essential role of human RecQ helicases in the genome maintenance during DNA replication is underlined by association of defects in their function with cancer predisposition. Copyright © 2016 Elsevier B.V. All rights reserved.

Simulator Studies of the Deep Stall

Science.gov (United States)

White, Maurice D.; Cooper, George E.

1965-01-01

Simulator studies of the deep-stall problem encountered with modern airplanes are discussed. The results indicate that the basic deep-stall tendencies produced by aerodynamic characteristics are augmented by operational considerations. Because of control difficulties to be anticipated in the deep stall, it is desirable that adequate safeguards be provided against inadvertent penetrations.

The analysis on centrifugal compressor rotating stall

International Nuclear Information System (INIS)

Kim, Ji Hwan; Kim, Kwang Ho; Shin, You Hwan

2003-01-01

In the present study, the performance characteristics and the number of stall cell during rotating stall of a centrifugal air compressor were experimentally investigated. Rotating stall in the vaneless diffuser were investigated by measuring unsteady pressure fluctuations at several different diffuser radius using a high frequency pressure transducer. The number of stall cell and their rotational speeds are distinctive features of the rotating stall phenomenon. The present study is mainly forced on the analysis for the stall cell number and its propagation speed unstable operating region of the compressor. The interpretation method of visualization is based on the pressure distribution in the circumference pressure fields while plotting the pressure and its harmonics variations in time in polar coordinates. To obtain the visualize the existence rotating stall, auto-correlation function and the frequency spectra of the pressure fluctuations were measured at r/r2=1.52. When the flow coefficient is lower than 0.150, the static pressure at impeller inlet is higher than that at inlet duct of the compressor. And the flow coefficient is lower than 0.086, several stall cell groups of discrete frequencies are observed

The quest for stall-free dynamic lift

Science.gov (United States)

Tung, C.; Mcalister, K. W.; Carr, Lawrence W.; Duque, E.; Zinner, R.

1992-01-01

During the past decade, numerous major effects have addressed the question of how to control or alleviate dynamic stall effects on helicopter rotors, but little concrete evidence of any significant reduction of the adverse characteristics of the dynamic stall phenomenon has been demonstrated. Nevertheless, it is important to remember that the control of dynamic stall is an achievable goal. Experiments performed at the US Army Aeroflight-dynamics Directorate more than a decade ago demonstrated that dynamic stall is not an unavoidable penalty of high amplitude motion, and that airfoils can indeed operate dynamically at angles far above the static-stall angle without necessarily forming a stall vortex. These experiments, one of them featuring a slat that was designed from static airfoil considerations, showed that unsteadiness can be a very beneficial factor in the development of high-lift devices for helicopter rotors. The experience drawn from these early experiments is now being focused on a program for the alleviation of dynamic-stall effects on helicopter rotors. The purpose of this effort is to demonstrate that rotor stall can be controlled through an improved understanding of the unsteady effects on airfoil stall and to document the role of specific means that lead to stall alleviation in the three dimensional unsteady environment of helicopter rotors in forward flight. The first concept to be addressed in this program will be a slatted airfoil. A two dimensional unsteady Navier-Stokes code has been modified to compute the flow around a two-element airfoil.

Rotating stall simulation for axial and centrifugal compressors

Science.gov (United States)

Halawa, Taher; Gadala, Mohamed S.

2017-05-01

This study presents a numerical simulation of the rotating stall phenomenon in axial and centrifugal compressors with detailed descriptions of stall precursors and its development with time. Results showed that the vaneless region of the centrifugal compressor is the most critical location affected by stall. It was found that the tip leakage flow and the back flow impingement are the main cause of the stall development at the impeller exit area for centrifugal compressors. The results of the axial compressor simulations indicated that the early separated flow combined with the tip leakage flow can block the impeller passages during stall.

Genome-wide identification and characterisation of human DNA replication origins by initiation site sequencing (ini-seq).

Science.gov (United States)

Langley, Alexander R; Gräf, Stefan; Smith, James C; Krude, Torsten

2016-12-01

Next-generation sequencing has enabled the genome-wide identification of human DNA replication origins. However, different approaches to mapping replication origins, namely (i) sequencing isolated small nascent DNA strands (SNS-seq); (ii) sequencing replication bubbles (bubble-seq) and (iii) sequencing Okazaki fragments (OK-seq), show only limited concordance. To address this controversy, we describe here an independent high-resolution origin mapping technique that we call initiation site sequencing (ini-seq). In this approach, newly replicated DNA is directly labelled with digoxigenin-dUTP near the sites of its initiation in a cell-free system. The labelled DNA is then immunoprecipitated and genomic locations are determined by DNA sequencing. Using this technique we identify >25,000 discrete origin sites at sub-kilobase resolution on the human genome, with high concordance between biological replicates. Most activated origins identified by ini-seq are found at transcriptional start sites and contain G-quadruplex (G4) motifs. They tend to cluster in early-replicating domains, providing a correlation between early replication timing and local density of activated origins. Origins identified by ini-seq show highest concordance with sites identified by SNS-seq, followed by OK-seq and bubble-seq. Furthermore, germline origins identified by positive nucleotide distribution skew jumps overlap with origins identified by ini-seq and OK-seq more frequently and more specifically than do sites identified by either SNS-seq or bubble-seq. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

RPA mediates recombination repair during replication stress and is displaced from DNA by checkpoint signalling in human cells

DEFF Research Database (Denmark)

Sleeth, Kate M; Sørensen, Claus Storgaard; Issaeva, Natalia

2007-01-01

The replication protein A (RPA) is involved in most, if not all, nuclear metabolism involving single-stranded DNA. Here, we show that RPA is involved in genome maintenance at stalled replication forks by the homologous recombination repair system in humans. Depletion of the RPA protein inhibited...... the formation of RAD51 nuclear foci after hydroxyurea-induced replication stalling leading to persistent unrepaired DNA double-strand breaks (DSBs). We demonstrate a direct role of RPA in homology directed recombination repair. We find that RPA is dispensable for checkpoint kinase 1 (Chk1) activation...... and that RPA directly binds RAD52 upon replication stress, suggesting a direct role in recombination repair. In addition we show that inhibition of Chk1 with UCN-01 decreases dissociation of RPA from the chromatin and inhibits association of RAD51 and RAD52 with DNA. Altogether, our data suggest a direct role...

Tus-Ter as a tool to study site-specific DNA replication perturbation in eukaryotes

DEFF Research Database (Denmark)

Larsen, Nicolai B; Hickson, Ian D; Mankouri, Hocine W

2014-01-01

The high-affinity binding of the Tus protein to specific 21-bp sequences, called Ter, causes site-specific, and polar, DNA replication fork arrest in E coli. The Tus-Ter complex serves to coordinate DNA replication with chromosome segregation in this organism. A number of recent and ongoing studies...... have demonstrated that Tus-Ter can be used as a heterologous tool to generate site-specific perturbation of DNA replication when reconstituted in eukaryotes. Here, we review these recent findings and explore the molecular mechanism by which Tus-Ter mediates replication fork (RF) arrest in the budding...... yeast, S. cerevisiae. We propose that Tus-Ter is a versatile, genetically tractable, and regulatable RF blocking system that can be utilized for disrupting DNA replication in a diverse range of host cells....

Tus-Ter as a tool to study site-specific DNA replication perturbation in eukaryotes.

Science.gov (United States)

Larsen, Nicolai B; Hickson, Ian D; Mankouri, Hocine W

2014-01-01

The high-affinity binding of the Tus protein to specific 21-bp sequences, called Ter, causes site-specific, and polar, DNA replication fork arrest in E coli. The Tus-Ter complex serves to coordinate DNA replication with chromosome segregation in this organism. A number of recent and ongoing studies have demonstrated that Tus-Ter can be used as a heterologous tool to generate site-specific perturbation of DNA replication when reconstituted in eukaryotes. Here, we review these recent findings and explore the molecular mechanism by which Tus-Ter mediates replication fork (RF) arrest in the budding yeast, S. cerevisiae. We propose that Tus-Ter is a versatile, genetically tractable, and regulatable RF blocking system that can be utilized for disrupting DNA replication in a diverse range of host cells.

16 CFR 1505.50 - Stalled motor testing.

Science.gov (United States)

2010-01-01

... 16 Commercial Practices 2 2010-01-01 2010-01-01 false Stalled motor testing. 1505.50 Section 1505... USE BY CHILDREN Policies and Interpretations § 1505.50 Stalled motor testing. (a) § 1505.6(e)(4)(ii) requires that a motor-operated toy be tested with the motor stalled if the construction of the toy is such...

The Relevance of the Dynamic Stall Effect for Transient

DEFF Research Database (Denmark)

Jauch, Clemens; Sørensen, Poul; Bak-Jensen, Birgitte

2005-01-01

This article describes a methodology to quantify the influence of dynamic stall on transient fault operations of active-stall turbines. The model of the dynamic stall effect is introduced briefly. The behaviour of the dynamic stall model during a transient fault operation is described mathematica...

Inclusion bodies are a site of ebolavirus replication.

Science.gov (United States)

Hoenen, Thomas; Shabman, Reed S; Groseth, Allison; Herwig, Astrid; Weber, Michaela; Schudt, Gordian; Dolnik, Olga; Basler, Christopher F; Becker, Stephan; Feldmann, Heinz

2012-11-01

Inclusion bodies are a characteristic feature of ebolavirus infections in cells. They contain large numbers of preformed nucleocapsids, but their biological significance has been debated, and they have been suggested to be aggregates of viral proteins without any further biological function. However, recent data for other viruses that produce similar structures have suggested that inclusion bodies might be involved in genome replication and transcription. In order to study filovirus inclusion bodies, we fused mCherry to the ebolavirus polymerase L, which is found in inclusion bodies. The resulting L-mCherry fusion protein was functional in minigenome assays and incorporated into virus-like particles. Importantly, L-mCherry fluorescence in transfected cells was readily detectable and distributed in a punctate pattern characteristic for inclusion bodies. A recombinant ebolavirus encoding L-mCherry instead of L was rescued and showed virtually identical growth kinetics and endpoint titers to those for wild-type virus. Using this virus, we showed that the onset of inclusion body formation corresponds to the onset of viral genome replication, but that viral transcription occurs prior to inclusion body formation. Live-cell imaging further showed that inclusion bodies are highly dynamic structures and that they can undergo dramatic reorganization during cell division. Finally, by labeling nascent RNAs using click technology we showed that inclusion bodies are indeed the site of viral RNA synthesis. Based on these data we conclude that, rather than being inert aggregates of nucleocapsids, ebolavirus inclusion bodies are in fact complex and dynamic structures and an important site at which viral RNA replication takes place.

Biomimetic Wind Turbine Design with Lift Enhancing Periodic Stall

NARCIS (Netherlands)

Stamhuis, Eize Jan

2017-01-01

A wind turbine includes a rotor; a blade; and a periodic stall system. The periodic stall system selectively moves at least part of the blade in an oscillating motion whereby an angle of incidence continuously varies to invoke periodic stall. The periodic stall system can move the entire blade or

The Location of the Bacterial Origin of Replication is Critical for Initial Ciproflaxcin Antibiotic Resistance

Science.gov (United States)

Bos, Julia; Nehring, Ralph; Cruz, Diane; Austin, Doug; Rosenberg, Susan; Austin, Robert

By using E. coli cells in which the unique origin of replication has been moved to a ectopic chromosome location distant from the native one, we probe how perturbation of gene order near the origin of replication impacts genome stability and survival under genomic attack. We find that when challenged with sub-inhibitory doses of ciprofloxacin, an antibiotic that generates replication fork stalling, cells with the ectopic origin show significant fitness loss. We show that genes functionally relevant to the cipro-induced stress response are largely located near the native origin, even in distantly related species. We show that while cipro induces increased copy number of genes proximal to the origin of replication as a direct consequence of replication fork stalling, gene copy number variation was reduced near the ectopic origin. Altered gene dosage in cells with an ectopic origin resulted in impaired replication fork repair and chromosome instability. We propose that gene distribution in the origin region acts as a fundamental first line of defense when the integrity of the genome is threatened and that genes proximal to the origin of replication serve as a mechanism of genetic innovation and a driving force of genome evolution in the presence of genotoxic antibiotics. Lewis Sigler Institute for Integrative Genomics and the Physics Department at Princeton University.

Close-loop Dynamic Stall Control on a Pitching Airfoil

Science.gov (United States)

Giles, Ian; Corke, Thomas

2017-11-01

A closed-loop control scheme utilizing a plasma actuator to control dynamic stall is presented. The plasma actuator is located at the leading-edge of a pitching airfoil. It initially pulses at an unsteady frequency that perturbs the boundary layer flow over the suction surface of the airfoil. As the airfoil approaches and enters stall, the amplification of the unsteady disturbance is detected by an onboard pressure sensor also located near the leading edge. Once detected, the actuator is switched to a higher voltage control state that in static airfoil experiments would reattach the flow. The threshold level of the detection is a parameter in the control scheme. Three stall regimes were examined: light, medium, and deep stall, that were defined by their stall penetration angles. The results showed that in general, the closed-loop control scheme was effective at controlling dynamic stall. The cycle-integrated lift improved in all cases, and increased by as much as 15% at the lowest stall penetration angle. As important, the cycle-integrated aerodynamic damping coefficient also increased in all cases, and was made to be positive at the light stall regime where it traditionally is negative. The latter is important in applications where negative damping can lead to stall flutter.

Stalling Tropical Cyclones over the Atlantic Basin

Science.gov (United States)

Nielsen-Gammon, J. W.; Emanuel, K.

2017-12-01

Hurricane Harvey produced massive amounts of rain over southeast Texas and southwest Louisiana. Average storm total rainfall amounts over a 10,000 square mile (26,000 square km) area exceeded 30 inches (750 mm). An important aspect of the storm that contributed to the large rainfall totals was its unusual motion. The storm stalled shortly after making landfall, then moved back offshore before once again making landfall five days later. This storm motion permitted heavy rainfall to occur in the same general area for an extended period of time. The unusual nature of this event motivates an investigation into the characteristics and potential climate change influences on stalled tropical cyclones in the Atlantic basin using the HURDAT 2 storm track database for 1866-2016 and downscaled tropical cyclones driven by simulations of present and future climate. The motion of cyclones is quantified as the size of a circle circumscribing all storm locations during a given length of time. For a three-day period, Harvey remained inside a circle with a radius of 123 km. This ranks within the top 0.6% of slowest-moving historical storm instances. Among the 2% of slowest-moving storm instances prior to Harvey, only 13 involved storms that stalled near the continental United States coast, where they may have produced substantial rainfall onshore while tapping into marine moisture. Only two such storms stalled in the month of September, in contrast to 20 September stalls out of the 36 storms that stalled over the nearby open Atlantic. Just four of the stalled coastal storms were hurricanes, implying a return frequency for such storms of much less than once per decade. The synoptic setting of these storms is examined for common features, and historical and projected trends in occurrences of stalled storms near the coast and farther offshore are investigated.

Dynamic Stall Control Using Plasma Actuators

Science.gov (United States)

Webb, Nathan; Singhal, Achal; Castaneda, David; Samimy, Mo

2017-11-01

Dynamic stall occurs in many applications, including sharp maneuvers of fixed wing aircraft, wind turbines, and rotorcraft and produces large unsteady aerodynamic loads that can lead to flutter and mechanical failure. This work uses flow control to reduce the unsteady loads by excitation of instabilities in the shear layer over the separated region using nanosecond pulse driven dielectric barrier discharge (NS-DBD) plasma actuators. These actuators have been shown to effectively delay or mitigate static stall. A wide range of flow parameters were explored in the current work: Reynolds number (Re = 167,000 to 500,000), reduced frequency (k = 0.025 to 0.075), and excitation Strouhal number (Ste = 0 to 10). Based on the results, three major conclusions were drawn: (a) Low Strouhal number excitation (Ste <0.5) results in oscillatory aerodynamic loads in the stalled stage of dynamic stall; (b) All excitation resulted in earlier flow reattachment; and (c) Excitation at progressively higher Ste weakened and eventually eliminated the dynamic stall vortex (DSV), thereby dramatically reducing the unsteady loading. The decrease in the strength of the DSV is achieved by the formation of shear layer coherent structures that bleed the leading-edge vorticity prior to the ejection of the DSV.

Observations of dynamic stall on Darrieus wind turbine blades

Energy Technology Data Exchange (ETDEWEB)

Fujisawa, N.; Shibuya, S. [Department of Mechanical and Production Engineering, Niigata University, 8050 Ikarashi 2, 950-2181 Niigata (Japan)

2001-02-01

Flow field around a Darrieus wind turbine blade in dynamic stall is studied by flow visualization and particle image velocimetry (PIV) measurement in stationary and rotating frames of reference. The experiment is carried out using the small-scale Darrieus wind turbine in a water tunnel. The unsteady nature of the dynamic stall observed by the flow visualization is quantitatively reproduced in the instantaneous velocity distributions by PIV measurement, which describes the successive shedding of two pairs of stall vortices from the blade moving upstream. The mechanism of dynamic stall is due to the successive generation of separation on the inner surface of the blade followed by the formation of roll-up vortices from the outer surface. Although the qualitative nature of the dynamic stall is independent of the tip-speed ratios, the blade angle for stall appearance and the growth rate of the stall vortices are influenced by the change in tip-speed ratios.

Simulation model of an active stall wind turbine controller

Energy Technology Data Exchange (ETDEWEB)

Jauch, C.; Hansen, A.D.; Soerensen, P. [Risoe National Lab., Wind Energy Dept., Rosilde (Denmark); Blaabjerg, F. [Aalborg Univ., Inst. of Energy Technology (Denmark)

2004-07-01

This paper describes an active stall wind turbine controller. The objective is to develop a general model of an active stall controller in order to simulate the operation of grid connected active stall wind turbines. The active stall turbine concept and its control strategies are presented and evaluated on the basis of simulations. The presented controller is described for continuous operation under all wind speeds from start-up wind speed to shut doven wind speed. Due to its parametric implementation it is general i.e. it can represent different active stall wind turbine controllers and can be implemented in different simulation tools. (au)

ATR Prohibits Replication Catastrophe by Preventing Global Exhaustion of RPA

DEFF Research Database (Denmark)

Toledo Lazaro, Luis Ignacio; Altmeyer, Matthias; Rask, Maj-Britt

2013-01-01

origin firing generates an excess of single-stranded DNA that exhausts the nuclear pool of RPA. Partial reduction of RPA accelerated fork breakage, and forced elevation of RPA was sufficient to delay such "replication catastrophe" even in the absence of ATR activity. Conversely, unscheduled origin firing...... induced breakage of stalled forks even in cells with active ATR. Thus, ATR-mediated suppression of dormant origins shields active forks against irreversible breakage via preventing exhaustion of nuclear RPA. This study elucidates how replicating genomes avoid destabilizing DNA damage. Because cancer cells...

Checkpoint-dependent RNR induction promotes fork restart after replicative stress.

Science.gov (United States)

Morafraile, Esther C; Diffley, John F X; Tercero, José Antonio; Segurado, Mónica

2015-01-20

The checkpoint kinase Rad53 is crucial to regulate DNA replication in the presence of replicative stress. Under conditions that interfere with the progression of replication forks, Rad53 prevents Exo1-dependent fork degradation. However, although EXO1 deletion avoids fork degradation in rad53 mutants, it does not suppress their sensitivity to the ribonucleotide reductase (RNR) inhibitor hydroxyurea (HU). In this case, the inability to restart stalled forks is likely to account for the lethality of rad53 mutant cells after replication blocks. Here we show that Rad53 regulates replication restart through the checkpoint-dependent transcriptional response, and more specifically, through RNR induction. Thus, in addition to preventing fork degradation, Rad53 prevents cell death in the presence of HU by regulating RNR-expression and localization. When RNR is induced in the absence of Exo1 and RNR negative regulators, cell viability of rad53 mutants treated with HU is increased and the ability of replication forks to restart after replicative stress is restored.

Acute Smc5/6 depletion reveals its primary role in rDNA replication by restraining recombination at fork pausing sites.

Directory of Open Access Journals (Sweden)

Xiao P Peng

2018-01-01

Full Text Available Smc5/6, a member of the conserved SMC family of complexes, is essential for growth in most organisms. Its exact functions in a mitotic cell cycle are controversial, as chronic Smc5/6 loss-of-function alleles produce varying phenotypes. To circumvent this issue, we acutely depleted Smc5/6 in budding yeast and determined the first cell cycle consequences of Smc5/6 removal. We found a striking primary defect in replication of the ribosomal DNA (rDNA array. Each rDNA repeat contains a programmed replication fork barrier (RFB established by the Fob1 protein. Fob1 removal improves rDNA replication in Smc5/6 depleted cells, implicating Smc5/6 in the management of programmed fork pausing. A similar improvement is achieved by removing the DNA helicase Mph1 whose recombinogenic activity can be inhibited by Smc5/6 under DNA damage conditions. DNA 2D gel analyses further show that Smc5/6 loss increases recombination structures at RFB regions; moreover, mph1∆ and fob1∆ similarly reduce this accumulation. These findings point to an important mitotic role for Smc5/6 in restraining recombination events when protein barriers in rDNA stall replication forks. As rDNA maintenance influences multiple essential cellular processes, Smc5/6 likely links rDNA stability to overall mitotic growth.

Escherichia coli DinB inhibits replication fork progression without significantly inducing the SOS response.

Science.gov (United States)

Mori, Tetsuya; Nakamura, Tatsuro; Okazaki, Naoto; Furukohri, Asako; Maki, Hisaji; Akiyama, Masahiro Tatsumi

2012-01-01

The SOS response is readily triggered by replication fork stalling caused by DNA damage or a dysfunctional replicative apparatus in Escherichia coli cells. E. coli dinB encodes DinB DNA polymerase and its expression is upregulated during the SOS response. DinB catalyzes translesion DNA synthesis in place of a replicative DNA polymerase III that is stalled at a DNA lesion. We showed previously that DNA replication was suppressed without exogenous DNA damage in cells overproducing DinB. In this report, we confirm that this was due to a dose-dependent inhibition of ongoing replication forks by DinB. Interestingly, the DinB-overproducing cells did not significantly induce the SOS response even though DNA replication was perturbed. RecA protein is activated by forming a nucleoprotein filament with single-stranded DNA, which leads to the onset of the SOS response. In the DinB-overproducing cells, RecA was not activated to induce the SOS response. However, the SOS response was observed after heat-inducible activation in strain recA441 (encoding a temperature-sensitive RecA) and after replication blockage in strain dnaE486 (encoding a temperature-sensitive catalytic subunit of the replicative DNA polymerase III) at a non-permissive temperature when DinB was overproduced in these cells. Furthermore, since catalytically inactive DinB could avoid the SOS response to a DinB-promoted fork block, it is unlikely that overproduced DinB takes control of primer extension and thus limits single-stranded DNA. These observations suggest that DinB possesses a feature that suppresses DNA replication but does not abolish the cell's capacity to induce the SOS response. We conclude that DinB impedes replication fork progression in a way that does not activate RecA, in contrast to obstructive DNA lesions and dysfunctional replication machinery.

Compressible dynamic stall control using high momentum microjets

Science.gov (United States)

Beahan, James J.; Shih, Chiang; Krothapalli, Anjaneyulu; Kumar, Rajan; Chandrasekhara, Muguru S.

2014-09-01

Control of the dynamic stall process of a NACA 0015 airfoil undergoing periodic pitching motion is investigated experimentally at the NASA Ames compressible dynamic stall facility. Multiple microjet nozzles distributed uniformly in the first 12 % chord from the airfoil's leading edge are used for the dynamic stall control. Point diffraction interferometry technique is used to characterize the control effectiveness, both qualitatively and quantitatively. The microjet control has been found to be very effective in suppressing both the emergence of the dynamic stall vortex and the associated massive flow separation at the entire operating range of angles of attack. At the high Mach number ( M = 0.4), the use of microjets appears to eliminate the shock structures that are responsible for triggering the shock-induced separation, establishing the fact that the use of microjets is effective in controlling dynamic stall with a strong compressibility effect. In general, microjet control has an overall positive effect in terms of maintaining leading edge suction pressure and preventing flow separation.

The Dynamics of SecM-Induced Translational Stalling

Directory of Open Access Journals (Sweden)

Albert Tsai

2014-06-01

Full Text Available SecM is an E. coli secretion monitor capable of stalling translation on the prokaryotic ribosome without cofactors. Biochemical and structural studies have demonstrated that the SecM nascent chain interacts with the 50S subunit exit tunnel to inhibit peptide bond formation. However, the timescales and pathways of stalling on an mRNA remain undefined. To provide a dynamic mechanism for stalling, we directly tracked the dynamics of elongation on ribosomes translating the SecM stall sequence (FSTPVWISQAQGIRAGP using single-molecule fluorescence techniques. Within 1 min, three peptide-ribosome interactions work cooperatively over the last five codons of the SecM sequence, leading to severely impaired elongation rates beginning from the terminal proline and lasting four codons. Our results suggest that stalling is tightly linked to the dynamics of elongation and underscore the roles that the exit tunnel and nascent chain play in controlling fundamental steps in translation.

Disaster recovery using VMware vSphere Replication and vCenter Site Recovery Manager

CERN Document Server

GB, Abhilash

2014-01-01

This is a step-by-step guide that will help you understand disaster recovery using VMware vSphere Replication 5.5 and VMware vCenter Site Recovery Manager (SRM) 5.5. The topics and configuration procedures are accompanied with relevant screenshots, flowcharts, and logical diagrams that makes grasping the concepts easier. This book is a guide for anyone who is keen on using vSphere Replication or vCenter Site Recovery Manager as a disaster recovery solution. This is an excellent handbook for solution architects, administrators, on-field engineers, and support professionals. Although the book as

Miscoding-induced stalling of substrate translocation on the bacterial ribosome.

Science.gov (United States)

Alejo, Jose L; Blanchard, Scott C

2017-10-10

Directional transit of the ribosome along the messenger RNA (mRNA) template is a key determinant of the rate and processivity of protein synthesis. Imaging of the multistep translocation mechanism using single-molecule FRET has led to the hypothesis that substrate movements relative to the ribosome resolve through relatively long-lived late intermediates wherein peptidyl-tRNA enters the P site of the small ribosomal subunit via reversible, swivel-like motions of the small subunit head domain within the elongation factor G (GDP)-bound ribosome complex. Consistent with translocation being rate-limited by recognition and productive engagement of peptidyl-tRNA within the P site, we now show that base-pairing mismatches between the peptidyl-tRNA anticodon and the mRNA codon dramatically delay this rate-limiting, intramolecular process. This unexpected relationship between aminoacyl-tRNA decoding and translocation suggests that miscoding antibiotics may impact protein synthesis by impairing the recognition of peptidyl-tRNA in the small subunit P site during EF-G-catalyzed translocation. Strikingly, we show that elongation factor P (EF-P), traditionally known to alleviate ribosome stalling at polyproline motifs, can efficiently rescue translocation defects arising from miscoding. These findings help reveal the nature and origin of the rate-limiting steps in substrate translocation on the bacterial ribosome and indicate that EF-P can aid in resuming translation elongation stalled by miscoding errors.

Progerin sequestration of PCNA promotes replication fork collapse and mislocalization of XPA in laminopathy-related progeroid syndromes.

Science.gov (United States)

Hilton, Benjamin A; Liu, Ji; Cartwright, Brian M; Liu, Yiyong; Breitman, Maya; Wang, Youjie; Jones, Rowdy; Tang, Hui; Rusinol, Antonio; Musich, Phillip R; Zou, Yue

2017-09-01

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder that is caused by a point mutation in the LMNA gene, resulting in production of a truncated farnesylated-prelamin A protein (progerin). We previously reported that XPA mislocalized to the progerin-induced DNA double-strand break (DSB) sites, blocking DSB repair, which led to DSB accumulation, DNA damage responses, and early replication arrest in HGPS. In this study, the XPA mislocalization to DSBs occurred at stalled or collapsed replication forks, concurrent with a significant loss of PCNA at the forks, whereas PCNA efficiently bound to progerin. This PCNA sequestration likely exposed ds-ssDNA junctions at replication forks for XPA binding. Depletion of XPA or progerin each significantly restored PCNA at replication forks. Our results suggest that although PCNA is much more competitive than XPA in binding replication forks, PCNA sequestration by progerin may shift the equilibrium to favor XPA binding. Furthermore, we demonstrated that progerin-induced apoptosis could be rescued by XPA, suggesting that XPA-replication fork binding may prevent apoptosis in HGPS cells. Our results propose a mechanism for progerin-induced genome instability and accelerated replicative senescence in HGPS.-Hilton, B. A., Liu, J., Cartwright, B. M., Liu, Y., Breitman, M., Wang, Y., Jones, R., Tang, H., Rusinol, A., Musich, P. R., Zou, Y. Progerin sequestration of PCNA promotes replication fork collapse and mislocalization of XPA in laminopathy-related progeroid syndromes. © FASEB.

A numerical strategy for modelling rotating stall in core compressors

Science.gov (United States)

Vahdati, M.

2007-03-01

The paper will focus on one specific core-compressor instability, rotating stall, because of the pressing industrial need to improve current design methods. The determination of the blade response during rotating stall is a difficult problem for which there is no reliable procedure. During rotating stall, the blades encounter the stall cells and the excitation depends on the number, size, exact shape and rotational speed of these cells. The long-term aim is to minimize the forced response due to rotating stall excitation by avoiding potential matches between the vibration modes and the rotating stall pattern characteristics. Accurate numerical simulations of core-compressor rotating stall phenomena require the modelling of a large number of bladerows using grids containing several tens of millions of points. The time-accurate unsteady-flow computations may need to be run for several engine revolutions for rotating stall to get initiated and many more before it is fully developed. The difficulty in rotating stall initiation arises from a lack of representation of the triggering disturbances which are inherently present in aeroengines. Since the numerical model represents a symmetric assembly, the only random mechanism for rotating stall initiation is provided by numerical round-off errors. In this work, rotating stall is initiated by introducing a small amount of geometric mistuning to the rotor blades. Another major obstacle in modelling flows near stall is the specification of appropriate upstream and downstream boundary conditions. Obtaining reliable boundary conditions for such flows can be very difficult. In the present study, the low-pressure compression (LPC) domain is placed upstream of the core compressor. With such an approach, only far field atmospheric boundary conditions are specified which are obtained from aircraft speed and altitude. A chocked variable-area nozzle, placed after the last compressor bladerow in the model, is used to impose boundary

Dynamic Stall Characteristics of Drooped Leading Edge Airfoils

Science.gov (United States)

Sankar, Lakshmi N.; Sahin, Mehmet; Gopal, Naveen

2000-01-01

Helicopters in high-speed forward flight usually experience large regions of dynamic stall over the retreating side of the rotor disk. The rapid variations in the lift and pitching moments associated with the stall process can result in vibratory loads, and can cause fatigue and failure of pitch links. In some instances, the large time lag between the aerodynamic forces and the blade motion can trigger stall flutter. A number of techniques for the alleviation of dynamic stall have been proposed and studied by researchers. Passive and active control techniques have both been explored. Passive techniques include the use of high solidity rotors that reduce the lift coefficients of individual blades, leading edge slots and leading edge slats. Active control techniques include steady and unsteady blowing, and dynamically deformable leading edge (DDLE) airfoils. Considerable amount of experimental and numerical data has been collected on the effectiveness of these concepts. One concept that has not received as much attention is the drooped-leading edge airfoil idea. It has been observed in wind tunnel studies and flight tests that drooped leading edge airfoils can have a milder dynamic stall, with a significantly milder load hysteresis. Drooped leading edge airfoils may not, however, be suitable at other conditions, e.g. in hover, or in transonic flow. Work needs to be done on the analysis and design of drooped leading edge airfoils for efficient operation in a variety of flight regimes (hover, dynamic stall, and transonic flow). One concept that is worthy of investigation is the dynamically drooping airfoil, where the leading edge shape is changed roughly once-per-rev to mitigate the dynamic stall.

Mutations in DONSON disrupt replication fork stability and cause microcephalic dwarfism.

Science.gov (United States)

Reynolds, John J; Bicknell, Louise S; Carroll, Paula; Higgs, Martin R; Shaheen, Ranad; Murray, Jennie E; Papadopoulos, Dimitrios K; Leitch, Andrea; Murina, Olga; Tarnauskaitė, Žygimantė; Wessel, Sarah R; Zlatanou, Anastasia; Vernet, Audrey; von Kriegsheim, Alex; Mottram, Rachel M A; Logan, Clare V; Bye, Hannah; Li, Yun; Brean, Alexander; Maddirevula, Sateesh; Challis, Rachel C; Skouloudaki, Kassiani; Almoisheer, Agaadir; Alsaif, Hessa S; Amar, Ariella; Prescott, Natalie J; Bober, Michael B; Duker, Angela; Faqeih, Eissa; Seidahmed, Mohammed Zain; Al Tala, Saeed; Alswaid, Abdulrahman; Ahmed, Saleem; Al-Aama, Jumana Yousuf; Altmüller, Janine; Al Balwi, Mohammed; Brady, Angela F; Chessa, Luciana; Cox, Helen; Fischetto, Rita; Heller, Raoul; Henderson, Bertram D; Hobson, Emma; Nürnberg, Peter; Percin, E Ferda; Peron, Angela; Spaccini, Luigina; Quigley, Alan J; Thakur, Seema; Wise, Carol A; Yoon, Grace; Alnemer, Maha; Tomancak, Pavel; Yigit, Gökhan; Taylor, A Malcolm R; Reijns, Martin A M; Simpson, Michael A; Cortez, David; Alkuraya, Fowzan S; Mathew, Christopher G; Jackson, Andrew P; Stewart, Grant S

2017-04-01

To ensure efficient genome duplication, cells have evolved numerous factors that promote unperturbed DNA replication and protect, repair and restart damaged forks. Here we identify downstream neighbor of SON (DONSON) as a novel fork protection factor and report biallelic DONSON mutations in 29 individuals with microcephalic dwarfism. We demonstrate that DONSON is a replisome component that stabilizes forks during genome replication. Loss of DONSON leads to severe replication-associated DNA damage arising from nucleolytic cleavage of stalled replication forks. Furthermore, ATM- and Rad3-related (ATR)-dependent signaling in response to replication stress is impaired in DONSON-deficient cells, resulting in decreased checkpoint activity and the potentiation of chromosomal instability. Hypomorphic mutations in DONSON substantially reduce DONSON protein levels and impair fork stability in cells from patients, consistent with defective DNA replication underlying the disease phenotype. In summary, we have identified mutations in DONSON as a common cause of microcephalic dwarfism and established DONSON as a critical replication fork protein required for mammalian DNA replication and genome stability.

Prediction of induced vibrations in stall

Energy Technology Data Exchange (ETDEWEB)

Thirstrup Petersen, J; Thomsen, K; Aagaard Madsen, H [Risoe National Lab., Wind Energy and Atmospheric Physics Dept., Roskilde (Denmark)

1999-03-01

The main results from recent research in stall induced vibrations are presented. The focus is on the edgewise blade vibrations, which during the last decade have turned out to be a potential threat against the stable operation of stall regulated wind turbines and a fact, which must be dealt with by the designer. The basic physical explanation for the phenomenon and examples of design precaution, which can be taken, are presented. (au)

Flow and Noise Characteristics of Centrifugal Fan under Different Stall Conditions

Directory of Open Access Journals (Sweden)

Lei Zhang

2014-01-01

Full Text Available An implicit, time-accurate 3D Reynolds-averaged Navier-Stokes (RANS solver is used to simulate the rotating stall phenomenon in a centrifugal fan. The goal of the present work is to shed light on the flow field and particularly the aerodynamic noise at different stall conditions. Aerodynamic characteristics, frequency domain characteristics, and the contours of sound power level under two different stall conditions are discussed in this paper. The results show that, with the decrease of valve opening, the amplitude of full pressure and flow fluctuations tends to be larger and the stall frequency remains the same. The flow field analysis indicates that the area occupied by stall cells expands with the decrease of flow rate. The noise calculation based on the simulation underlines the role of vortex noise after the occurrence of rotating stall, showing that the high noise area rotates along with the stall cell in the circumferential direction.

Simulation of Entropy Generation under Stall Conditions in a Centrifugal Fan

Directory of Open Access Journals (Sweden)

Lei Zhang

2014-06-01

Full Text Available Rotating stalls are generally the first instability met in turbomachinery, before surges. This 3D phenomenon is characterized by one or more stalled flow cells which rotate at a fraction of the impeller speed. The goal of the present work is to shed some light on the entropy generation in a centrifugal fan under rotating stall conditions. A numerical simulation of entropy generation is carried out with the ANSYS Fluent software which solves the Navier-Stokes equations and user defined function (UDF. The entropy generation characteristics in the centrifugal fan for five typical conditions are presented and discussed, involving the design condition, conditions on occurrence and development of stall inception, the rotating stall conditions with two throttle coefficients. The results show that the entropy generation increases after the occurrence of stall inception. The high entropy generation areas move along the circumferential and axial directions, and finally merge into one stall cell. The entropy generation rate during circumferential propagation of the stall cell is also discussed, showing that the entropy generation history is similar to sine curves in impeller and volute, and the volute tongue has a great influence on entropy generation in the centrifugal fan.

Economic evaluation of stall stocking density of lactating dairy cows

NARCIS (Netherlands)

Vries, De Albert; Dechassa, Hailegziabher; Hogeveen, Henk

2016-01-01

An increase in stall stocking density (SSD), as measured by the number of lactating cows per stall in a freestall barn, reduces cow performance, such as milk yield and fertility, but may increase farm profitability. Our objectives were to calculate effects of varying SSD on profit per stall for a

Simulation model of an active-stall fixed-speed wind turbine controller

Energy Technology Data Exchange (ETDEWEB)

Jauch, C.; Hansen, A.D.; Sorensen, P.; Blaabjerg, F.

2004-07-01

This paper describes an active-stall wind turbine controller. The objective is to develop a general model of an active stall controller in order to simulate the operation of grid connected active stall wind turbines. The active stall turbine concept and its control strategies are presented and evaluated by simulations. The presented controller is described for continuous operation under all wind speeds from start-up wind speed to shut down wind speed. Due to its parametric implementation it is general i. e. it can represent different active stall wind turbine controllers and can be implemented in different simulation tools. (author)

A CFD Database for Airfoils and Wings at Post-Stall Angles of Attack

Science.gov (United States)

Petrilli, Justin; Paul, Ryan; Gopalarathnam, Ashok; Frink, Neal T.

2013-01-01

This paper presents selected results from an ongoing effort to develop an aerodynamic database from Reynolds-Averaged Navier-Stokes (RANS) computational analysis of airfoils and wings at stall and post-stall angles of attack. The data obtained from this effort will be used for validation and refinement of a low-order post-stall prediction method developed at NCSU, and to fill existing gaps in high angle of attack data in the literature. Such data could have potential applications in post-stall flight dynamics, helicopter aerodynamics and wind turbine aerodynamics. An overview of the NASA TetrUSS CFD package used for the RANS computational approach is presented. Detailed results for three airfoils are presented to compare their stall and post-stall behavior. The results for finite wings at stall and post-stall conditions focus on the effects of taper-ratio and sweep angle, with particular attention to whether the sectional flows can be approximated using two-dimensional flow over a stalled airfoil. While this approximation seems reasonable for unswept wings even at post-stall conditions, significant spanwise flow on stalled swept wings preclude the use of two-dimensional data to model sectional flows on swept wings. Thus, further effort is needed in low-order aerodynamic modeling of swept wings at stalled conditions.

Conducting Classroom Observations : Stallings 'Classroom Snapshot' Observation System for an Electronic Tablet

OpenAIRE

World Bank Group

2017-01-01

The “Stallings Classroom Snapshot” instrument, technically called the “Stanford Research Institute Classroom Observation System”, was developed by Professor Jane Stallings for research on the efficiency and quality of basic education teachers in the United States in the 1970s. (Stallings, 1977; Stallings and Mohlman, 1988). The Stallings instrument generates robust quantitative data on the interaction of teachers and students in the classroom, with a high degree of inter-rater rel...

A role for the weak DnaA binding sites in bacterial replication origins

DEFF Research Database (Denmark)

Charbon, Godefroid; Løbner-Olesen, Anders

2011-01-01

DnaA initiates the chromosomal DNA replication in nearly all bacteria, and replication origins are characterized by binding sites for the DnaA protein (DnaA-boxes) along with an ‘AT-rich’ region. However, great variation in number, spatial organization and specificity of DnaA-boxes is observed...... between species. In the study by Taylor et al. (2011), new and unexpectedly weak DnaA-boxes were identified within the Caulobacter crescentus origin of replication (Cori). The position of weak and stronger DnaA-boxes follows a pattern seen in Escherichia coli oriC. This raises the possibility...... that bacterial origins might be more alike than previously thought....

Inception mechanism and suppression of rotating stall in an axial-flow fan

International Nuclear Information System (INIS)

Nishioka, T

2013-01-01

Inception patterns of rotating stall at two stagger-angle settings for the highly loaded rotor blades were experimentally investigated in a low-speed axial-flow fan. Rotor-tip flow fields were also numerically investigated to clarify the mechanism behind the rotating stall inception. The stall inception patterns depended on the rotor stagger-angle settings. The stall inception from a rotating instability was confirmed at the design stagger-angle settings. The stall inception from a short length-scale stall cell (spike) was also confirmed at the small stagger-angle setting. The spillage of tip-leakage flow and the tip-leakage vortex breakdown influence the rotating stall inception. An air-separator has been developed based on the clarified inception mechanism of rotating stall. The rotating stall was suppressed by the developed air-separator, and the operating range of fan was extended towards low flow rate. The effect of developed air-separator was also confirmed by application to a primary air fan used in a coal fired power plant. It is concluded from these results that the developed air-separator can provide a wide operating range for an axial-flow fan

Monitoring indices of cow comfort in free-stall-housed dairy herds.

Science.gov (United States)

Cook, N B; Bennett, T B; Nordlund, K V

2005-11-01

Indices of cow comfort are used widely by consultants in the dairy industry, with a general understanding that they are representative of lying behavior. This study examines the influence of stall base type (sand or a geotextile mattress filled with rubber crumbs) and time of measurement on 4 indices of comfort collected at hourly intervals in 12 herds, aligned by morning and afternoon milking. Stall base type significantly influenced all indices of comfort. For example, the least squares mean (SE) cow comfort index (proportion of cows touching a stall that are lying down) was 0.76 (0.015) in herds with mattresses compared with 0.86 (0.015) in herds with sand stalls. Significant hourly variation was also identified suggesting that timing of measurement is important. None of the indices of cow comfort derived from the high-yielding group pen was associated with the mean 24-h lying time of 10 sentinel cows whose time budgets were known in each herd. However, the cow comfort index was associated with the herd mean 24-h stall standing time, with the strongest relationships occurring 2 h before the morning and afternoon milking, when stall base type did not significantly influence the association. When measured at these times, we recommend use of the stall standing index (proportion of cows touching a stall that are standing), with values greater than 0.20 being associated with abnormally long herd mean stall standing times greater than 2 h/d.

14 CFR 33.65 - Surge and stall characteristics.

Science.gov (United States)

2010-01-01

... 14 Aeronautics and Space 1 2010-01-01 2010-01-01 false Surge and stall characteristics. 33.65 Section 33.65 Aeronautics and Space FEDERAL AVIATION ADMINISTRATION, DEPARTMENT OF TRANSPORTATION AIRCRAFT... stall characteristics. When the engine is operated in accordance with operating instructions required by...

Dedicated education unit: implementing an innovation in replication sites.

Science.gov (United States)

Moscato, Susan R; Nishioka, Vicki M; Coe, Michael T

2013-05-01

An important measure of an innovation is the ease of replication and achievement of the same positive outcomes. The dedicated education unit (DEU) clinical education model uses a collaborative academic-service partnership to develop an optimal learning environment for students. The University of Portland adapted this model from Flinders University, Australia, to increase the teaching capacity and quality of nursing education. This article identifies DEU implementation essentials and reports on the outcomes of two replication sites that received consultation support from the University of Portland. Program operation information, including education requirements for clinician instructors, types of patient care units, and clinical faculty-to-student ratios is presented. Case studies of the three programs suggest the DEU model is adaptable to a range of different clinical settings and continues to show promise as one strategy for addressing the nurse faculty shortage and strengthening academic-clinical collaborations while maintaining quality clinical education for students. Copyright 2013, SLACK Incorporated.

How MCM loading and spreading specify eukaryotic DNA replication initiation sites.

Science.gov (United States)

Hyrien, Olivier

2016-01-01

DNA replication origins strikingly differ between eukaryotic species and cell types. Origins are localized and can be highly efficient in budding yeast, are randomly located in early fly and frog embryos, which do not transcribe their genomes, and are clustered in broad (10-100 kb) non-transcribed zones, frequently abutting transcribed genes, in mammalian cells. Nonetheless, in all cases, origins are established during the G1-phase of the cell cycle by the loading of double hexamers of the Mcm 2-7 proteins (MCM DHs), the core of the replicative helicase. MCM DH activation in S-phase leads to origin unwinding, polymerase recruitment, and initiation of bidirectional DNA synthesis. Although MCM DHs are initially loaded at sites defined by the binding of the origin recognition complex (ORC), they ultimately bind chromatin in much greater numbers than ORC and only a fraction are activated in any one S-phase. Data suggest that the multiplicity and functional redundancy of MCM DHs provide robustness to the replication process and affect replication time and that MCM DHs can slide along the DNA and spread over large distances around the ORC. Recent studies further show that MCM DHs are displaced along the DNA by collision with transcription complexes but remain functional for initiation after displacement. Therefore, eukaryotic DNA replication relies on intrinsically mobile and flexible origins, a strategy fundamentally different from bacteria but conserved from yeast to human. These properties of MCM DHs likely contribute to the establishment of broad, intergenic replication initiation zones in higher eukaryotes.

Nuclear insulin-like growth factor 1 receptor phosphorylates proliferating cell nuclear antigen and rescues stalled replication forks after DNA damage.

Science.gov (United States)

Waraky, Ahmed; Lin, Yingbo; Warsito, Dudi; Haglund, Felix; Aleem, Eiman; Larsson, Olle

2017-11-03

We have previously shown that the insulin-like growth factor 1 receptor (IGF-1R) translocates to the cell nucleus, where it binds to enhancer-like regions and increases gene transcription. Further studies have demonstrated that nuclear IGF-1R (nIGF-1R) physically and functionally interacts with some nuclear proteins, i.e. the lymphoid enhancer-binding factor 1 (Lef1), histone H3, and Brahma-related gene-1 proteins. In this study, we identified the proliferating cell nuclear antigen (PCNA) as a nIGF-1R-binding partner. PCNA is a pivotal component of the replication fork machinery and a main regulator of the DNA damage tolerance (DDT) pathway. We found that IGF-1R interacts with and phosphorylates PCNA in human embryonic stem cells and other cell lines. In vitro MS analysis of PCNA co-incubated with the IGF-1R kinase indicated tyrosine residues 60, 133, and 250 in PCNA as IGF-1R targets, and PCNA phosphorylation was followed by mono- and polyubiquitination. Co-immunoprecipitation experiments suggested that these ubiquitination events may be mediated by DDT-dependent E2/E3 ligases ( e.g. RAD18 and SHPRH/HLTF). Absence of IGF-1R or mutation of Tyr-60, Tyr-133, or Tyr-250 in PCNA abrogated its ubiquitination. Unlike in cells expressing IGF-1R, externally induced DNA damage in IGF-1R-negative cells caused G 1 cell cycle arrest and S phase fork stalling. Taken together, our results suggest a role of IGF-1R in DDT. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Dairy Cows Produce Less Milk and Modify Their Behaviour during the Transition between Tie-Stall to Free-Stall

Science.gov (United States)

Broucek, Jan; Uhrincat, Michal; Mihina, Stefan; Soch, Miloslav; Mrekajova, Andrea; Hanus, Anton

2017-01-01

Simple Summary The purpose of this study was to evaluate the influence of moving cows from the barn with stanchion-stall housing to free-stall housing on their behaviour and production. Cows lay down up to ten hours after removing. The cows in their second lactation and open cows tended to lie sooner after removing than cows in their first lactation and pregnant cows. The times of total lying and rumination were increasing from the first day to the tenth day after removing. Cows produced 23.3% less milk at the first day following the transfer than at the last day prior to moving (23.76 ± 7.20 kg vs. 30.97 ± 7.26 kg, p cows achieved maximum production. The difference was found in milk losses due to the shift between cows in first and second lactation. Abstract Transfer of cattle to an unknown barn may result in a reduction in its welfare. Housing and management practices can result in signs of stress that include a long-term suppression of milk efficiency. The purpose of this study was to evaluate the influence of moving cows from the stanchion-stall housing to free-stall housing on their behaviour and production. The Holstein cows were moved into the new facility with free-stall housing from the old barn with stanchion-stall housing. Cows lay down up to ten hours (596.3 ± 282.7 min) after removing. The cows in their second lactation and open cows tended to lie sooner after removing than cows in their first lactation and pregnant cows. The times of total lying and rumination were increasing from the first day to the tenth day after removing (23.76 ± 7.20 kg vs. 30.97 ± 7.26 kg, p Cows produced 23.3% less milk at the first day following the transfer than at the last day prior to moving (p cows on the first and second lactation (p cows’ milk production. However, when the cows are moved to a better environment, they rapidly adapt to the change. PMID:28273810

Dynamic stall and 3D effects

Energy Technology Data Exchange (ETDEWEB)

Bjoerck, A.; Thor, S.E. [Aeronautical Research Inst. of Sweden, Bromma (Sweden)

1996-12-01

The JOULE II project `Dynamic stall and 3D effects` started in January 1994 and was completed in September 1995. The objective of the project has been to increase the understanding of the three-dimensional and unsteady aerodynamics of stall controlled HAWT`s. The objectives have also been to develop `engineering models` suitable for inclusion into aero-elastic codes. The project included the participation of 13 parties within Europe. This paper describes an overview of the work carried out within the project and key results. 3 refs, 4 figs

Airfoil stall interpreted through linear stability analysis

Science.gov (United States)

Busquet, Denis; Juniper, Matthew; Richez, Francois; Marquet, Olivier; Sipp, Denis

2017-11-01

Although airfoil stall has been widely investigated, the origin of this phenomenon, which manifests as a sudden drop of lift, is still not clearly understood. In the specific case of static stall, multiple steady solutions have been identified experimentally and numerically around the stall angle. We are interested here in investigating the stability of these steady solutions so as to first model and then control the dynamics. The study is performed on a 2D helicopter blade airfoil OA209 at low Mach number, M 0.2 and high Reynolds number, Re 1.8 ×106 . Steady RANS computation using a Spalart-Allmaras model is coupled with continuation methods (pseudo-arclength and Newton's method) to obtain steady states for several angles of incidence. The results show one upper branch (high lift), one lower branch (low lift) connected by a middle branch, characterizing an hysteresis phenomenon. A linear stability analysis performed around these equilibrium states highlights a mode responsible for stall, which starts with a low frequency oscillation. A bifurcation scenario is deduced from the behaviour of this mode. To shed light on the nonlinear behavior, a low order nonlinear model is created with the same linear stability behavior as that observed for that airfoil.

A comparison of free-stall barns used by modernized Wisconsin dairies.

Science.gov (United States)

Bewley, J; Palmer, R W; Jackson-Smith, D B

2001-02-01

A primary objective of the Wisconsin Dairy Modernization Survey was to compare features of free-stall barns available to dairy producers. This study used data from a large random sample of expanding dairy farms to determine whether the theoretical benefits of particular free-stall configurations bear out under on-farm conditions. Comparisons were made among herds using free-stall barns as their primary housing for new versus remodeled facilities, barn design, bedding used, feed-delivery design, manure removal strategies, animal restraint, maternity areas, overcrowding, and cooling methods. Producers who made the transition from tie-stall housing to free-stall housing were satisfied with this decision. New free-stall barns provided a more desirable environment for the herds than remodeled free-stall barns, although initial investments were higher. When new free-stall barns were compared, herds with four-row barns had higher production, lower somatic cell count, and higher stocking rates than herds with six-row barns. Respondents were more satisfied with four- and six-row barns than with two- and three-row barns. Respondents felt sand provided some advantages for cow comfort, while satisfaction with bedding cost and manure handling was higher with mattresses. Dairy Herd Improvement data showed no difference in milk production or somatic cell count for producers who chose sand or mattress-based free stalls. Respondents were more satisfied with the use of drive-through feeding than other feed-delivery designs. Most producers chose to use tractor scrapers to remove manure; however, producers who used automated systems were more satisfied with manure management. Few differences were observed when comparing self-locking head gates to palpation rails. Overcrowding did not have any adverse affect on production or user satisfaction with feed intake or cow comfort. Using supplemental cooling appeared to facilitate higher production.

Associations between cow hygiene, hock injuries, and free stall usage on US dairy farms.

Science.gov (United States)

Lombard, J E; Tucker, C B; von Keyserlingk, M A G; Kopral, C A; Weary, D M

2010-10-01

This cross-sectional study evaluated cow comfort measures in free stall dairies across the United States as part of the National Animal Health Monitoring System's Dairy 2007 study. The study was conducted in 17 states and evaluations were completed between March 5 and September 5, 2007. Assessors recorded hygiene and hock scores, number of cows housed in the pen, the number of cows standing with only the front feet in a stall, standing fully in a stall, and lying in a stall. Facility design measures included bedding type, bedding quantity, stall length and width, presence of a neck rail or brisket locator, and relevant distances from the rear and bed of the stall. Of the 491 operations that completed the cow comfort assessment, 297 had Holstein cows housed in free stalls and were included in this analysis. Negative binomial models were constructed to evaluate the following outcomes: the number of cows that were very dirty, had severe hock injuries, stood with front feet in the stall, stood with all feet in the stall, and were lying in the stall. Hygiene was better on farms that did not tail dock cows compared with those that did (5.7 vs. 8.8% were dirty) and on farms located in the study's west region compared with those located in the east region (5.2 vs. 9.7% were dirty). Severe hock injuries were less common on farms in the west than those in the east (0.5 vs. 4.1%). In addition, severe hock injuries were less common on farms that used dirt as a stall base or sand as bedding compared with farms that did not. A higher percentage of cows was standing with front feet in the stall at higher ambient temperatures (incidence rate ratio=1.016) and as time since feeding increased (incidence rate ratio=1.030). A lower percentage of cows were standing with front feet in the stall when the stalls were shorter and when there were fewer cows per stall. Standing fully in a stall was performed by a higher percentage of cows during the summer than during the spring (13.6 vs. 8

A Comparative Study of Three Methodologies for Modeling Dynamic Stall

Science.gov (United States)

Sankar, L.; Rhee, M.; Tung, C.; ZibiBailly, J.; LeBalleur, J. C.; Blaise, D.; Rouzaud, O.

2002-01-01

During the past two decades, there has been an increased reliance on the use of computational fluid dynamics methods for modeling rotors in high speed forward flight. Computational methods are being developed for modeling the shock induced loads on the advancing side, first-principles based modeling of the trailing wake evolution, and for retreating blade stall. The retreating blade dynamic stall problem has received particular attention, because the large variations in lift and pitching moments encountered in dynamic stall can lead to blade vibrations and pitch link fatigue. Restricting to aerodynamics, the numerical prediction of dynamic stall is still a complex and challenging CFD problem, that, even in two dimensions at low speed, gathers the major difficulties of aerodynamics, such as the grid resolution requirements for the viscous phenomena at leading-edge bubbles or in mixing-layers, the bias of the numerical viscosity, and the major difficulties of the physical modeling, such as the turbulence models, the transition models, whose both determinant influences, already present in static maximal-lift or stall computations, are emphasized by the dynamic aspect of the phenomena.

Dynamic stall characterization using modal analysis of phase-averaged pressure distributions

Science.gov (United States)

Harms, Tanner; Nikoueeyan, Pourya; Naughton, Jonathan

2017-11-01

Dynamic stall characterization by means of surface pressure measurements can simplify the time and cost associated with experimental investigation of unsteady airfoil aerodynamics. A unique test capability has been developed at University of Wyoming over the past few years that allows for time and cost efficient measurement of dynamic stall. A variety of rotorcraft and wind turbine airfoils have been tested under a variety of pitch oscillation conditions resulting in a range of dynamic stall behavior. Formation, development and separation of different flow structures are responsible for the complex aerodynamic loading behavior experienced during dynamic stall. These structures have unique signatures on the pressure distribution over the airfoil. This work investigates the statistical behavior of phase-averaged pressure distribution for different types of dynamic stall by means of modal analysis. The use of different modes to identify specific flow structures is being investigated. The use of these modes for different types of dynamic stall can provide a new approach for understanding and categorizing these flows. This work uses airfoil data acquired under Army contract W911W60160C-0021, DOE Grant DE-SC0001261, and a gift from BP Alternative Energy North America, Inc.

Precautions against axial fan stall in reactor building to Tianwan NPP

International Nuclear Information System (INIS)

Liu Chunlong; Pei Junmin

2011-01-01

The paper introduces the mechanism and harm of rotating stall of axial fans, analyzes the necessity for prevention against axial fan stall in reactor building of Tianwan NPP, introduces the precautions, and then makes an assessment on anti-stall effect of flow separators. It can provide reference for model-selection or reconstruction of similar fans in power stations, and for operation and maintenance of axial fans. (authors)

Simulation of Broadband Noise Sources of an Axial Fan under Rotating Stall Conditions

Directory of Open Access Journals (Sweden)

Lei Zhang

2014-11-01

Full Text Available Study on the influence of rotating stall on the aerodynamic noise of axial fan has important value to warn of the occurrence of stall through monitoring the noise variations. The present work is to analyze the aerodynamic noise before and after the phenomenon of rotating stall by solving Navier-Stokes equations, coupled with the throttle condition and the broadband noise sources model. The impeller exit rotational Mach number and rotational Reynolds number are separately 0.407 and 8.332 × 106. The results show that the aerodynamic noise source of the fan is mainly the rotation noise under the design condition. The vortex noise accounts for the major part of fan noise after the occurrence of stall, and the maximum acoustic power level of the fan appears in the rotor domains. In the evolution process from the stall inception to the stall cell, the high noise regions of the rotor develop along the radial, circumferential, and axial directions, and the area occupied by high noise regions increases from 33% to 46% impeller channels area. On rotating stall condition, the high noise regions occupying about 46% impeller channels area propagate with the stall cell along the circumferential direction at a half of rotor speed.

Replication stalling by catalytically impaired Twinkle induces mitochondrial DNA rearrangements in cultured cells

NARCIS (Netherlands)

Pohjoismaki, J.L.; Goffart, S.; Spelbrink, J.N.

2011-01-01

Pathological mitochondrial DNA (mtDNA) rearrangements have been proposed to result from repair of double-strand breaks caused by blockage of mitochondrial DNA (mtDNA) replication. As mtDNA deletions are seen only in post-mitotic tissues, it has been suggested that they are selected out in actively

Preferences of dairy cows for three stall surface materials with small amounts of bedding.

Science.gov (United States)

Norring, M; Manninen, E; de Passillé, A M; Rushen, J; Saloniemi, H

2010-01-01

Farmers' concerns about the economy, cost of labor, and hygiene have resulted in reduced use of organic bedding in stalls for dairy cows; however, the reduced use of organic bedding possibly impairs cow comfort. The effects of different stall surface materials were evaluated in an unheated building in which only a small amount of bedding was used. The lying time and preferences of 18 cows using 3 stall surface materials (concrete, soft rubber mat, and sand) were compared. All materials were lightly bedded with a small amount of straw, and the amount of straw added to each stall was measured. The cows only had access to stalls of one surface type while their lying time was observed. Lying times were longest on the rubber mats compared with other surfaces (rubber mat 768; concrete 727; sand 707+/-16 min/d). In a preference test, cows had access to 2 of the 3 types of stalls for 10 d and their stall preference was measured. Cows preferred stalls with rubber mats to stalls with a concrete floor (median 73 vs. 18 from a total of 160 observations per day; interquartile range was 27 and 12, respectively), but showed no preference for sand stalls compared with stalls with a concrete floor or with rubber mats. More straw was needed on sand stalls compared with concrete or mat (638+/-13 g/d on sand, 468+/-10 g/d on concrete, and 464+/-8 g/d on rubber mats). Lying times on bedded mats indicated that mats were comfortable for the cows. If availability or cost of bedding material requires limiting the amount of bedding used, rubber mats may help maintain cow comfort. Copyright 2010 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Recruitment of Mcm10 to Sites of Replication Initiation Requires Direct Binding to the Minichromosome Maintenance (MCM) Complex*

Science.gov (United States)

Douglas, Max E.

2016-01-01

Mcm10 is required for the initiation of eukaryotic DNA replication and contributes in some unknown way to the activation of the Cdc45-MCM-GINS (CMG) helicase. How Mcm10 is localized to sites of replication initiation is unclear, as current models indicate that direct binding to minichromosome maintenance (MCM) plays a role, but the details and functional importance of this interaction have not been determined. Here, we show that purified Mcm10 can bind both DNA-bound double hexamers and soluble single hexamers of MCM. The binding of Mcm10 to MCM requires the Mcm10 C terminus. Moreover, the binding site for Mcm10 on MCM includes the Mcm2 and Mcm6 subunits and overlaps that for the loading factor Cdt1. Whether Mcm10 recruitment to replication origins depends on CMG helicase assembly has been unclear. We show that Mcm10 recruitment occurs via two modes: low affinity recruitment in the absence of CMG assembly (“G1-like”) and high affinity recruitment when CMG assembly takes place (“S-phase-like”). Mcm10 that cannot bind directly to MCM is defective in both modes of recruitment and is unable to support DNA replication. These findings indicate that Mcm10 is localized to replication initiation sites by directly binding MCM through the Mcm10 C terminus. PMID:26719337

Recruitment of Mcm10 to Sites of Replication Initiation Requires Direct Binding to the Minichromosome Maintenance (MCM) Complex.

Science.gov (United States)

Douglas, Max E; Diffley, John F X

2016-03-11

Mcm10 is required for the initiation of eukaryotic DNA replication and contributes in some unknown way to the activation of the Cdc45-MCM-GINS (CMG) helicase. How Mcm10 is localized to sites of replication initiation is unclear, as current models indicate that direct binding to minichromosome maintenance (MCM) plays a role, but the details and functional importance of this interaction have not been determined. Here, we show that purified Mcm10 can bind both DNA-bound double hexamers and soluble single hexamers of MCM. The binding of Mcm10 to MCM requires the Mcm10 C terminus. Moreover, the binding site for Mcm10 on MCM includes the Mcm2 and Mcm6 subunits and overlaps that for the loading factor Cdt1. Whether Mcm10 recruitment to replication origins depends on CMG helicase assembly has been unclear. We show that Mcm10 recruitment occurs via two modes: low affinity recruitment in the absence of CMG assembly ("G1-like") and high affinity recruitment when CMG assembly takes place ("S-phase-like"). Mcm10 that cannot bind directly to MCM is defective in both modes of recruitment and is unable to support DNA replication. These findings indicate that Mcm10 is localized to replication initiation sites by directly binding MCM through the Mcm10 C terminus. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

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.

Bacillus cereus in free-stall bedding.

Science.gov (United States)

Magnusson, M; Svensson, B; Kolstrup, C; Christiansson, A

2007-12-01

To increase the understanding of how different factors affect the bacterial growth in deep sawdust beds for dairy cattle, the microbiological status of Bacillus cereus and coliforms in deep sawdust-bedded free stalls was investigated over two 14-d periods on one farm. High counts of B. cereus and coliforms were found in the entire beds. On average, 4.1 log(10) B. cereus spores, 5.5 log(10) B. cereus, and 6.7 log(10) coliforms per gram of bedding could be found in the upper layers of the sawdust likely to be in contact with the cows' udders. The highest counts of B. cereus spores, B. cereus, and coliforms were found in the bedding before fresh bedding was added, and the lowest immediately afterwards. Different factors of importance for the growth of B. cereus in the bedding material were explored in laboratory tests. These were found to be the type of bedding, pH, and the type and availability of nutrients. Alternative bedding material such as peat and mixtures of peat and sawdust inhibited the bacterial growth of B. cereus. The extent of growth of B. cereus in the sawdust was increased in a dose-dependent manner by the availability of feces. Urine added to different bedding material raised the pH and also led to bacterial growth of B. cereus in the peat. In sawdust, a dry matter content greater than 70% was needed to lower the water activity to 0.95, which is needed to inhibit the growth of B. cereus. In an attempt to reduce the bacterial growth of B. cereus and coliforms in deep sawdust beds on the farm, the effect of giving bedding daily or a full replacement of the beds was studied. The spore count of B. cereus in the back part of the free stalls before fresh bedding was added was 0.9 log units lower in stalls given daily bedding than in stalls given bedding twice weekly. No effect on coliform counts was found. Replacement of the entire sawdust bedding had an effect for a short period, but by 1 to 2 mo after replacement, the counts of B. cereus spores in the

Stall Recovery Guidance Algorithms Based on Constrained Control Approaches

Science.gov (United States)

Stepanyan, Vahram; Krishnakumar, Kalmanje; Kaneshige, John; Acosta, Diana

2016-01-01

Aircraft loss-of-control, in particular approach to stall or fully developed stall, is a major factor contributing to aircraft safety risks, which emphasizes the need to develop algorithms that are capable of assisting the pilots to identify the problem and providing guidance to recover the aircraft. In this paper we present several stall recovery guidance algorithms, which are implemented in the background without interfering with flight control system and altering the pilot's actions. They are using input and state constrained control methods to generate guidance signals, which are provided to the pilot in the form of visual cues. It is the pilot's decision to follow these signals. The algorithms are validated in the pilot-in-the loop medium fidelity simulation experiment.

Prediction of active control of subsonic centrifugal compressor rotating stall

Science.gov (United States)

Lawless, Patrick B.; Fleeter, Sanford

1993-01-01

A mathematical model is developed to predict the suppression of rotating stall in a centrifugal compressor with a vaned diffuser. This model is based on the employment of a control vortical waveform generated upstream of the impeller inlet to damp weak potential disturbances that are the early stages of rotating stall. The control system is analyzed by matching the perturbation pressure in the compressor inlet and exit flow fields with a model for the unsteady behavior of the compressor. The model was effective at predicting the stalling behavior of the Purdue Low Speed Centrifugal Compressor for two distinctly different stall patterns. Predictions made for the effect of a controlled inlet vorticity wave on the stability of the compressor show that for minimum control wave magnitudes, on the order of the total inlet disturbance magnitude, significant damping of the instability can be achieved. For control waves of sufficient amplitude, the control phase angle appears to be the most important factor in maintaining a stable condition in the compressor.

An archival analysis of stall warning system effectiveness during airborne icing encounters

Science.gov (United States)

Maris, John Michael

An archival study was conducted to determine the influence of stall warning system performance on aircrew decision-making outcomes during airborne icing encounters. A Conservative Icing Response Bias (CIRB) model was developed to explain the historical variability in aircrew performance in the face of airframe icing. The model combined Bayes' Theorem with Signal Detection Theory (SDT) concepts to yield testable predictions that were evaluated using a Binary Logistic Regression (BLR) multivariate technique applied to two archives: the NASA Aviation Safety Reporting System (ASRS) incident database, and the National Transportation Safety Board (NTSB) accident databases, both covering the period January 1, 1988 to October 2, 2015. The CIRB model predicted that aircrew would experience more incorrect response outcomes in the face of missed stall warnings than with stall warning False Alarms. These predicted outcomes were observed at high significance levels in the final sample of 132 NASA/NTSB cases. The CIRB model had high sensitivity and specificity, and explained 71.5% (Nagelkerke R2) of the variance of aircrew decision-making outcomes during the icing encounters. The reliability and validity metrics derived from this study suggest indicate that the findings are generalizable to the population of U.S. registered turbine-powered aircraft. These findings suggest that icing-related stall events could be reduced if the incidence of stall warning Misses could be minimized. Observed stall warning Misses stemmed from three principal causes: aerodynamic icing effects, which reduced the stall angle-of-attack (AoA) to below the stall warning calibration threshold; tail stalls, which are not monitored by contemporary protection systems; and icing-induced system issues (such as frozen pitot tubes), which compromised stall warning system effectiveness and airframe envelope protections. Each of these sources of missed stall warnings could be addressed by Aerodynamic Performance

Replication fork stability confers chemoresistance in BRCA-deficient cells

DEFF Research Database (Denmark)

Chaudhuri, Arnab Ray; Callen, Elsa; Ding, Xia

2016-01-01

/4 complex protein, PTIP, protects Brca1/2-deficient cells from DNA damage and rescues the lethality of Brca2-deficient embryonic stem cells. However, PTIP deficiency does not restore homologous recombination activity at double-strand breaks. Instead, its absence inhibits the recruitment of the MRE11......Cells deficient in the Brca1 and Brca2 genes have reduced capacity to repair DNA double-strand breaks by homologous recombination and consequently are hypersensitive to DNA-damaging agents, including cisplatin and poly(ADP-ribose) polymerase (PARP) inhibitors. Here we show that loss of the MLL3...... nuclease to stalled replication forks, which in turn protects nascent DNA strands from extensive degradation. More generally, acquisition of PARP inhibitors and cisplatin resistance is associated with replication fork protection in Brca2-deficient tumour cells that do not develop Brca2 reversion mutations...

Dynamic Characteristics of Rotating Stall in Mixed Flow Pump

Directory of Open Access Journals (Sweden)

Xiaojun Li

2013-01-01

Full Text Available Rotating stall, a phenomenon that causes flow instabilities and pressure hysteresis by propagating at some fraction of the impeller rotational speed, can occur in centrifugal impellers, mixed impellers, radial diffusers, or axial diffusers. Despite considerable efforts devoted to the study of rotating stall in pumps, the mechanics of this phenomenon are not sufficiently understood. The propagation mechanism and onset of rotating stall are not only affected by inlet flow but also by outlet flow as well as the pressure gradient in the flow passage. As such, the complexity of these concepts is not covered by the classical explanation. To bridge this research gap, the current study investigated prerotation generated at the upstream of the impeller, leakage flow at the tip clearance between the casing and the impeller, and strong reserve flow at the inlet of the diffuser. Understanding these areas will clarify the origin of the positive slope of the head-flow performance curve for a mixed flow pump. Nonuniform pressure distribution and adverse pressure gradient were also introduced to evaluate the onset and development of rotating stall within the diffuser.

Numerical Investigations of Dynamic Stall Control

Directory of Open Access Journals (Sweden)

Florin FRUNZULICA

2014-04-01

Full Text Available In this paper we investigated numerically the dynamic stall phenomenon and the possibilities to control it, with application to vertical axis wind turbines (for urban users. The Phenomenon appear at low tip speed ratio (TSR<4 and it has a great impact on structural integrity of the wind turbine and power performances. For this reason we performed a computational study of dynamic stall around NACA 0012 airfoil in pitching motion at relative low Reynolds number (105. Also, we performed the same analysis for four flow control methods: two passive (Gurney flap and slot and two active (blowing jet on the rounded trailing edge and synthetic jet periodically activated. The Results are compared to those of an existing experimental case test.

Shallow and deep dynamic stall for flapping low Reynolds number airfoils

Energy Technology Data Exchange (ETDEWEB)

Ol, Michael V. [Wright-Patterson AFB, Air Force Research Lab., Dayton, OH (United States); Bernal, Luis; Kang, Chang-Kwon; Shyy, Wei [University of Michigan, Department of Aerospace Engineering, Ann Arbor, MI (United States)

2009-05-15

We consider a combined experimental (based on flow visualization, direct force measurement and phase-averaged 2D particle image velocimetry in a water tunnel), computational (2D Reynolds-averaged Navier-Stokes) and theoretical (Theodorsen's formula) approach to study the fluid physics of rigid-airfoil pitch-plunge in nominally two-dimensional conditions. Shallow-stall (combined pitch-plunge) and deep-stall (pure-plunge) are compared at a reduced frequency commensurate with flapping-flight in cruise in nature. Objectives include assessment of how well attached-flow theory can predict lift coefficient even in the presence of significant separation, and how well 2D velocimetry and 2D computation can mutually validate one another. The shallow-stall case shows promising agreement between computation and experiment, while in the deep-stall case, the computation's prediction of flow separation lags that of the experiment, but eventually evinces qualitatively similar leading edge vortex size. Dye injection was found to give good qualitative match with particle image velocimetry in describing leading edge vortex formation and return to flow reattachment, and also gave evidence of strong spanwise growth of flow separation after leading-edge vortex formation. Reynolds number effects, in the range of 10,000-60,000, were found to influence the size of laminar separation in those phases of motion where instantaneous angle of attack was well below stall, but have limited effect on post-stall flowfield behavior. Discrepancy in lift coefficient time history between experiment, theory and computation was mutually comparable, with no clear failure of Theodorsen's formula. This is surprising and encouraging, especially for the deep-stall case, because the theory's assumptions are clearly violated, while its prediction of lift coefficient remains useful for capturing general trends. (orig.)

Prevalence of lameness among dairy cattle in Wisconsin as a function of housing type and stall surface.

Science.gov (United States)

Cook, Nigel B

2003-11-01

To determine the prevalence of lameness as a function of season (summer vs winter), housing type (free stalls vs tie stalls), and stall surface (sand vs any other surface) among lactating dairy cows in Wisconsin. Epidemiologic survey. 3,621 lactating dairy cows in 30 herds. Herds were visited once during the summer and once during the winter, and a locomotion score ranging from 1 (no gait abnormality) to 4 (severe lameness) was assigned to all lactating cows. Cows with a score of 3 or 4 were considered to be clinically lame. Mean +/- SD herd lameness prevalence was 21.1 +/- 10.5% during the summer and 23.9 +/- 10.7% during the winter; these values were significantly different. During the winter, mean prevalence of lameness in free-stall herds with non-sand stall surfaces (33.7%) was significantly higher than prevalences in free-stall herds with sand stall surfaces (21.2%), tie-stall herds with non-sand stall surfaces (21.7%), and tie-stall herds with sand stall surfaces (12.1%). Results suggest that the prevalence of lameness among dairy cattle in Wisconsin is higher than previously thought and that lameness prevalence is associated with season, housing type, and stall surface.

Effects of laminar separation bubbles and turbulent separation on airfoil stall

Energy Technology Data Exchange (ETDEWEB)

Dini, P. [Carleton College, Northfield, MN (United States); Coiro, D.P. [Universita di Napoli (Italy)

1997-12-31

An existing two-dimensional, interactive, stall prediction program is extended by improving its laminar separation bubble model. The program now accounts correctly for the effects of the bubble on airfoil performance characteristics when it forms at the mid-chord and on the leading edge. Furthermore, the model can now predict bubble bursting on very sharp leading edges at high angles of attack. The details of the model are discussed in depth. Comparisons of the predicted stall and post-stall pressure distributions show excellent agreement with experimental measurements for several different airfoils at different Reynolds numbers.

The SmpB C-terminal tail helps tmRNA to recognize and enter stalled ribosomes

Directory of Open Access Journals (Sweden)

Mickey R. Miller

2014-09-01

Full Text Available In bacteria, transfer-messenger RNA (tmRNA and SmpB comprise the most common and effective system for rescuing stalled ribosomes. Ribosomes stall on mRNA transcripts lacking stop codons and are rescued as the defective mRNA is swapped for the tmRNA template in a process known as trans-translation. The tmRNA–SmpB complex is recruited to the ribosome independent of a codon–anticodon interaction. Given that the ribosome uses robust discriminatory mechanisms to select against non-cognate tRNAs during canonical decoding, it has been hard to explain how this can happen. Recent structural and biochemical studies show that SmpB licenses tmRNA entry through its interactions with the decoding center and mRNA channel. In particular, the C-terminal tail of SmpB promotes both EFTu activation and accommodation of tmRNA, the former through interactions with 16S rRNA nucleotide G530 and the latter through interactions with the mRNA channel downstream of the A site. Here we present a detailed model of the earliest steps in trans-translation, and in light of these mechanistic considerations, revisit the question of how tmRNA preferentially reacts with stalled, non-translating ribosomes.

Visualization and PIV measurement of unsteady flow around a darrieus wind turbine in dynamic stall

Energy Technology Data Exchange (ETDEWEB)

Shibuya, Satoshi; Fujisawa, Nobuyuki; Takano, Tsuyoshi [Dept. of Mechanical and Production Engineering, Niigata Univ., Niigata (Japan)

1999-07-01

Flow around a Darrieus wind turbine in dynamic stall is studied by flow visualization and PIV (particle image velocimeter) measurement in a rotating frame of reference, which allows the successive observation of the dynamic stall over the blade. The qualitative features of the flow field in dynamic stall observed by the flow visualization, such as the formation and shedding of the stall vortices, are quantitatively reproduced in the instantaneous velocity distributions near the blade by using PIV. These results indicate that two pairs of stall vortices are generated from the blade during one rotation of the blade and that the size and the generating blade angle of the stall vortices are enlarged as the tip-speed ratio decreases. These stall vortices are produced by the in-flow motion from the outer surface to the inner surface through the trailing edge of the blade and the flow separation over the inner surface of the blade. (author)

Identify the Rotating Stall in Centrifugal Compressors by Fractal Dimension in Reconstructed Phase Space

Directory of Open Access Journals (Sweden)

Le Wang

2015-11-01

Full Text Available Based on phase space reconstruction and fractal dynamics in nonlinear dynamics, a method is proposed to extract and analyze the dynamics of the rotating stall in the impeller of centrifugal compressor, and some numerical examples are given to verify the results as well. First, the rotating stall of an existing low speed centrifugal compressor (LSCC is numerically simulated, and the time series of pressure in the rotating stall is obtained at various locations near the impeller outlet. Then, the phase space reconstruction is applied to these pressure time series, and a low-dimensional dynamical system, which the dynamics properties are included in, is reconstructed. In phase space reconstruction, C–C method is used to obtain the key parameters, such as time delay and the embedding dimension of the reconstructed phase space. Further, the fractal characteristics of the rotating stall are analyzed in detail, and the fractal dimensions are given for some examples to measure the complexity of the flow in the post-rotating stall. The results show that the fractal structures could reveal the intrinsic dynamics of the rotating stall flow and could be considered as a characteristic to identify the rotating stall.

An airloads theory for morphing airfoils in dynamic stall with experimental correlation

Science.gov (United States)

Ahaus, Loren A.

Helicopter rotor blades frequently encounter dynamic stall during normal flight conditions, limiting the applicability of classical thin-airfoil theory at large angles of attack. Also, it is evident that because of the largely different conditions on the advancing and retreating sides of the rotor, future rotorcraft may incorporate dynamically morphing airfoils (trailing-edge aps, dynamic camber, dynamic droop, etc.). Reduced-order aerodynamic models are needed for preliminary design and ight simulation. A unified model for predicting the airloads on a morphing airfoil in dynamic stall is presented, consisting of three components. First, a linear airloads theory allows for arbitrary airfoil deformations consistent with a morphing airfoil. Second, to capture the effects of the wake, the airloads theory is coupled to an induced ow model. Third, the overshoot and time delay associated with dynamic stall are modeled by a second-order dynamic filter, along the lines of the ONERA dynamic stall model. This paper presents a unified airloads model that allows arbitrary airfoil morphing with dynamic stall. Correlations with experimental data validate the theory.

Stall Margin Improvement in a Centrifugal Compressor through Inducer Casing Treatment

Directory of Open Access Journals (Sweden)

V. V. N. K. Satish Koyyalamudi

2016-01-01

Full Text Available The increasing trend of high stage pressure ratio with increased aerodynamic loading has led to reduction in stable operating range of centrifugal compressors with stall and surge initiating at relatively higher mass flow rates. The casing treatment technique of stall control is found to be effective in axial compressors, but very limited research work is published on the application of this technique in centrifugal compressors. Present research was aimed to investigate the effect of casing treatment on the performance and stall margin of a high speed, 4 : 1 pressure ratio centrifugal compressor through numerical simulations using ANSYS CFX software. Three casing treatment configurations were developed and incorporated in the shroud over the inducer of the impeller. The predicted performance of baseline compressor (without casing treatment was in good agreement with published experimental data. The compressor with different inducer casing treatment geometries showed varying levels of stall margin improvement, up to a maximum of 18%. While the peak efficiency of the compressor with casing treatment dropped by 0.8%–1% compared to the baseline compressor, the choke mass flow rate was improved by 9.5%, thus enhancing the total stable operating range. The inlet configuration of the casing treatment was found to play an important role in stall margin improvement.

Load prediction of stall regulated wind turbines

Energy Technology Data Exchange (ETDEWEB)

Bjoerck, A.; Dahlberg, J.Aa. [Aeronautical Research Inst. of Sweden, Bromma (Sweden); Carlen, I. [Chalmers Univ. of Technology, Goeteborg (Sweden). Div. of Marine Structural Engineering; Ganander, H. [Teknikgruppen AB, Sollentua (Sweden)

1996-12-01

Measurements of blade loads on a turbine situated in a small wind farm shows that the highest blade loads occur during operation close to the peak power i.e. when the turbine operates in the stall region. In this study the extensive experimental data base has been utilised to compare loads in selected campaigns with corresponding load predictions. The predictions are based on time domain simulations of the wind turbine structure, performed by the aeroelastic code VIDYN. In the calculations a model were adopted in order to include the effects of dynamic stall. This paper describes the work carried out so far within the project and key results. 5 refs, 10 figs

Alleviation of spike stall in axial compressors utilizing grooved casing treatment

Directory of Open Access Journals (Sweden)

Reza Taghavi-Zenouz

2015-06-01

Full Text Available This article deals with application of grooved type casing treatment for suppression of spike stall in an isolated axial compressor rotor blade row. The continuous grooved casing treatment covering the whole compressor circumference is of 1.8 mm in depth and located between 90% and 108% chord of the blade tip as measured from leading edge. The method of investigation is based on time-accurate three-dimensional full annulus numerical simulations for cases with and without casing treatment. Discretization of the Navier–Stokes equations has been carried out based on an upwind second-order scheme and k-ω-SST (Shear Stress Transport turbulence modeling has been used for estimation of eddy viscosity. Time-dependent flow structure results for the smooth casing reveal that there are two criteria for spike stall inception known as leading edge spillage and trailing edge backflow, which occur at specific mass flow rates in near-stall conditions. In this case, two dominant stall cells of different sizes could be observed. The larger one is caused by the spike stall covering roughly two blade passages in the circumferential direction and about 25% span in the radial direction. Spike stall disturbances are accompanied by lower frequencies and higher amplitudes of the pressure signals. Casing treatment causes flow blockages to reduce due to alleviation of backflow regions, which in turn reduces the total pressure loss and increases the axial velocity in the blade tip gap region, as well as tip leakage flow fluctuation at higher frequencies and lower amplitudes. Eventually, it can be concluded that the casing treatment of the stepped tip gap type could increase the stall margin of the compressor. This fact is basically due to retarding the movement of the interface region between incoming and tip leakage flows towards the rotor leading edge plane and suppressing the reversed flow around the blade trailing edge.

Development of High Speed Imaging and Analysis Techniques Compressible Dynamics Stall

Science.gov (United States)

Chandrasekhara, M. S.; Carr, L. W.; Wilder, M. C.; Davis, Sanford S. (Technical Monitor)

1996-01-01

Dynamic stall has limited the flight envelope of helicopters for many years. The problem has been studied in the laboratory as well as in flight, but most research, even in the laboratory, has been restricted to surface measurement techniques such as pressure transducers or skin friction gauges, except at low speed. From this research, it became apparent that flow visualization tests performed at Mach numbers representing actual flight conditions were needed if the complex physics associated with dynamic stall was to be properly understood. However, visualization of the flow field during compressible conditions required carefully aligned and meticulously reconstructed holographic interferometry. As part of a long-range effort focused on exposing of the physics of compressible dynamic stall, a research wind tunnel was developed at NASA Ames Research Center which permits visual access to the full flow field surrounding an oscillating airfoil during compressible dynamic stall. Initially, a stroboscopic schlieren technique was used for visualization of the stall process, but the primary research tool has been point diffraction interferometry(PDI), a technique carefully optimized for use in th is project. A review of the process of development of PDI will be presented in the full paper. One of the most valuable aspects of PDI is the fact that interferograms are produced in real time on a continuous basis. The use of a rapidly-pulsed laser makes this practical; a discussion of this approach will be presented in the full paper. This rapid pulsing(up to 40,000 pulses/sec) produces interferograms of the rapidly developing dynamic stall field in sufficient resolution(both in space and time) that the fluid physics of the compressible dynamic stall flowfield can be quantitatively determined, including the gradients of pressure in space and time. This permits analysis of the influence of the effect of pitch rate, Mach number, Reynolds number, amplitude of oscillation, and other

The role of HERC2 and RNF8 ubiquitin E3 ligases in the promotion of translesion DNA synthesis in the chicken DT40 cell line

DEFF Research Database (Denmark)

Mohiuddin, Mohammed; Kobayashi, Shunsuke; Keka, Islam Shamima

2016-01-01

immediately after exposure to UV while retaining proficient post-replicative gap filling. These mutants are both proficient in mono-ubiquitination of PCNA. Taken together, these results suggest that HERC2 and RNF8 promote TLS past abasic sites and UV-lesions at or very close to stalled replication forks....

Analysis of Low Speed Stall Aerodynamics of a Swept Wing with Laminar Flow Glove

Science.gov (United States)

Bui, Trong T.

2014-01-01

Reynolds-Averaged Navier-Stokes (RANS) computational fluid dynamics (CFD) analysis was conducted to study the low-speed stall aerodynamics of a GIII aircraft's swept wing modified with a laminar-flow wing glove. The stall aerodynamics of the gloved wing were analyzed and compared with the unmodified wing for the flight speed of 120 knots and altitude of 2300 ft above mean sea level (MSL). The Star-CCM+ polyhedral unstructured CFD code was first validated for wing stall predictions using the wing-body geometry from the First American Institute of Aeronautics and Astronautics (AIAA) CFD High-Lift Prediction Workshop. It was found that the Star-CCM+ CFD code can produce results that are within the scattering of other CFD codes considered at the workshop. In particular, the Star-CCM+ CFD code was able to predict wing stall for the AIAA wing-body geometry to within 1 degree of angle of attack as compared to benchmark wind-tunnel test data. Current results show that the addition of the laminar-flow wing glove causes the gloved wing to stall much earlier than the unmodified wing. Furthermore, the gloved wing has a different stall characteristic than the clean wing, with no sharp lift drop-off at stall for the gloved wing.

Analysis of Low-Speed Stall Aerodynamics of a Swept Wing with Laminar-Flow Glove

Science.gov (United States)

Bui, Trong T.

2014-01-01

Reynolds-Averaged Navier-Stokes (RANS) computational fluid dynamics (CFD) analysis was conducted to study the low-speed stall aerodynamics of a GIII aircraft's swept wing modified with a laminar-flow wing glove. The stall aerodynamics of the gloved wing were analyzed and compared with the unmodified wing for the flight speed of 120 knots and altitude of 2300 ft above mean sea level (MSL). The Star-CCM+ polyhedral unstructured CFD code was first validated for wing stall predictions using the wing-body geometry from the First American Institute of Aeronautics and Astronautics (AIAA) CFD High-Lift Prediction Workshop. It was found that the Star-CCM+ CFD code can produce results that are within the scattering of other CFD codes considered at the workshop. In particular, the Star-CCM+ CFD code was able to predict wing stall for the AIAA wing-body geometry to within 1 degree of angle of attack as compared to benchmark wind-tunnel test data. Current results show that the addition of the laminar-flow wing glove causes the gloved wing to stall much earlier than the unmodified wing. Furthermore, the gloved wing has a different stall characteristic than the clean wing, with no sharp lift drop-off at stall for the gloved wing.

Stall inception and warning in a single-stage transonic axial compressor with axial skewed slot casing treatment

International Nuclear Information System (INIS)

Lim, Byeung Jun; Kwon, Se Jin; Park, Tae Choon

2014-01-01

Characteristic changes in the stall inception in a single-stage transonic axial compressor with an axial skewed slot casing treatment were investigated experimentally. A rotating stall occurred intermittently in a compressor with an axial skewed slot, whereas spike-type rotating stalls occurred in the case of smooth casing. The axial skewed slot suppressed stall cell growth and increased the operating range. A mild surge, the frequency of which is the Helmholtz frequency of the compressor system, occurred with the rotating stall. The irregularity in the pressure signals at the slot bottom increased decreasing flow rate. An autocorrelation-based stall warning method was applied to the measured pressure signals. Results estimate and warn against the stall margin in a compressor with an axial skewed slot.

ATR-Chk1-APC/C-dependent stabilization of Cdc7-ASK (Dbf4) kinase is required for DNA lesion bypass under replication stress

DEFF Research Database (Denmark)

Yamada, M.; Watanabe, K.; Mistrik, M.

2013-01-01

replication. Stalled DNA replication evoked stabilization of the Cdc7-ASK (Dbf4) complex in a manner dependent on ATR-Chk1-mediated checkpoint signaling and its interplay with the anaphase-promoting complex/cyclosomeCdh1 (APC/C) ubiquitin ligase. Mechanistically, Chk1 kinase inactivates APC/C through...... degradation of Cdh1 upon replication block, thereby stabilizing APC/C substrates, including Cdc7-ASK (Dbf4). Furthermore, motif C of ASK (Dbf4) interacts with the N-terminal region of RAD18 ubiquitin ligase, and this interaction is required for chromatin binding of RAD18. Impaired interaction of ASK (Dbf4...

Stability Analysis for Rotating Stall Dynamics in Axial Flow Compressors

Science.gov (United States)

1999-01-01

modes determines collectively local stability of the compressor model. Explicit conditions are obtained for local stability of rotating stall which...critical modes determines the stability for rotating stall collectively . We point out that although in a special case our stability condition for...strict crossing assumption implies that the zero solution changes its stability as ~, crosses ~’c. For instance, odk (yc ) > 0 implies that the zero

Modeling dynamic stall on wind turbine blades under rotationally augmented flow fields

DEFF Research Database (Denmark)

Guntur, Srinivas; Sørensen, Niels N.; Schreck, Scott

2016-01-01

a reduced order dynamic stall model that uses rotationally augmented steady-state polars obtained from steady Phase VI experimental sequences, instead of the traditional two-dimensional, non-rotating data. The aim of this work is twofold. First, the blade loads estimated by the DDES simulations are compared...... Experiment Phase VI experimental data, including constant as well as continuously pitching blade conditions during axial operation; (2) data from unsteady delayed detached eddy simulations (DDES) carried out using the Technical University of Denmark’s in-house flow solver Ellipsys3D; and (3) data from...... with those from the dynamic stall model. This allowed the differences between the stall phenomenon on the inboard parts of harmonically pitching blades on a rotating wind turbine and the classic dynamic stall representation in two-dimensional flow to be investigated. Results indicated a good qualitative...

Experimental investigation on the effects of non-cyclical frequency and amplitude variation on dynamic stall

Science.gov (United States)

Heintz, Kyle C.

An experimental study of a cambered airfoil undergoing non-cyclical, transient pitch trajectories and the resulting effects on the dynamic stall phenomenon is presented. Surface pressure measurements and airfoil incidence angle are acquired simultaneously to resolve instantaneous aerodynamic load coefficients at Mach numbers ranging from 0.2 to 0.4. Derived from these coefficients are various formulations of the aerodynamic damping factor, referred to copiously throughout. Using a two-motor mechanism, each providing independent frequency and amplitude input to the airfoil, unique pitch motions can be implemented by actively controlling the phase between inputs. This work primarily focuses on three pitch motion schemas, the first of which is a "chirp" style trajectory featuring concurrent exponential frequency growth and amplitude decay. Second, these parameters are tested separately to determine their individual contributions. Lastly, a novel dual harmonic pitch motion is devised which rapidly traverses dynamic stall regimes on an inter-cycle basis by modulating the static-stall penetration angle. Throughout all results presented, there is evidence that for consecutive pitch-cycles, the process of dynamic stall is affected when prior oscillations prior have undergone deeper stall-penetration angles. In other words when stall-penetration is descending, retreating from a regime of light or deep stall, statistics of load coefficients, such as damping coefficient, maximum lift, minimum quarter-chord moment, and their phase relationships, do not match the values seen when stall-penetration was growing. The outcomes herein suggest that the airfoil retains some memory of previous flow separation which has the potential to change the influence of the dynamic stall vortex.

pUL34 binding near the human cytomegalovirus origin of lytic replication enhances DNA replication and viral growth.

Science.gov (United States)

Slayton, Mark; Hossain, Tanvir; Biegalke, Bonita J

2018-05-01

The human cytomegalovirus (HCMV) UL34 gene encodes sequence-specific DNA-binding proteins (pUL34) which are required for viral replication. Interactions of pUL34 with DNA binding sites represses transcription of two viral immune evasion genes, US3 and US9. 12 additional predicted pUL34-binding sites are present in the HCMV genome (strain AD169) with three binding sites concentrated near the HCMV origin of lytic replication (oriLyt). We used ChIP-seq analysis of pUL34-DNA interactions to confirm that pUL34 binds to the oriLyt region during infection. Mutagenesis of the UL34-binding sites in an oriLyt-containing plasmid significantly reduced viral-mediated oriLyt-dependent DNA replication. Mutagenesis of these sites in the HCMV genome reduced the replication efficiencies of the resulting viruses. Protein-protein interaction analyses demonstrated that pUL34 interacts with the viral proteins IE2, UL44, and UL84, that are essential for viral DNA replication, suggesting that pUL34-DNA interactions in the oriLyt region are involved in the DNA replication cascade. Copyright © 2018 Elsevier Inc. All rights reserved.

Human CST Facilitates Genome-wide RAD51 Recruitment to GC-Rich Repetitive Sequences in Response to Replication Stress.

Science.gov (United States)

Chastain, Megan; Zhou, Qing; Shiva, Olga; Fadri-Moskwik, Maria; Whitmore, Leanne; Jia, Pingping; Dai, Xueyu; Huang, Chenhui; Ye, Ping; Chai, Weihang

2016-08-02

The telomeric CTC1/STN1/TEN1 (CST) complex has been implicated in promoting replication recovery under replication stress at genomic regions, yet its precise role is unclear. Here, we report that STN1 is enriched at GC-rich repetitive sequences genome-wide in response to hydroxyurea (HU)-induced replication stress. STN1 deficiency exacerbates the fragility of these sequences under replication stress, resulting in chromosome fragmentation. We find that upon fork stalling, CST proteins form distinct nuclear foci that colocalize with RAD51. Furthermore, replication stress induces physical association of CST with RAD51 in an ATR-dependent manner. Strikingly, CST deficiency diminishes HU-induced RAD51 foci formation and reduces RAD51 recruitment to telomeres and non-telomeric GC-rich fragile sequences. Collectively, our findings establish that CST promotes RAD51 recruitment to GC-rich repetitive sequences in response to replication stress to facilitate replication restart, thereby providing insights into the mechanism underlying genome stability maintenance. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

Replication Protein A (RPA) Phosphorylation Prevents RPA Association with Replication Centers

OpenAIRE

Vassin, Vitaly M.; Wold, Marc S.; Borowiec, James A.

2004-01-01

Mammalian replication protein A (RPA) undergoes DNA damage-dependent phosphorylation at numerous sites on the N terminus of the RPA2 subunit. To understand the functional significance of RPA phosphorylation, we expressed RPA2 variants in which the phosphorylation sites were converted to aspartate (RPA2D) or alanine (RPA2A). Although RPA2D was incorporated into RPA heterotrimers and supported simian virus 40 DNA replication in vitro, the RPA2D mutant was selectively unable to associate with re...

Enhancing BEM simulations of a stalled wind turbine using a 3D correction model

Science.gov (United States)

Bangga, Galih; Hutomo, Go; Syawitri, Taurista; Kusumadewi, Tri; Oktavia, Winda; Sabila, Ahmad; Setiadi, Herlambang; Faisal, Muhamad; Hendranata, Yongki; Lastomo, Dwi; Putra, Louis; Kristiadi, Stefanus; Bumi, Ilmi

2018-03-01

Nowadays wind turbine rotors are usually employed with pitch control mechanisms to avoid deep stall conditions. Despite that, wind turbines often operate under pitch fault situation causing massive flow separation to occur. Pure Blade Element Momentum (BEM) approaches are not designed for this situation and inaccurate load predictions are already expected. In the present studies, BEM predictions are improved through the inclusion of a stall delay model for a wind turbine rotor operating under pitch fault situation of -2.3° towards stall. The accuracy of the stall delay model is assessed by comparing the results with available Computational Fluid Dynamics (CFD) simulations data.

Prediction of dynamic loads and induced vibrations in stall

Energy Technology Data Exchange (ETDEWEB)

Thirstrup Petersen, J.; Aagaard Madsen, H. [Risoe National Lab. (Denmark); Bjoerck, A. [Aeronautical Research Inst. of Sweden (Sweden); Enevoldsen, P. [Bonus Energy A/S (Denmark); Oeye, S. [The Technical Univ. of Denmark (Denmark); Ganander, H. [Teknikgruppen AB (Sweden); Winkelaar, D. [Netherlands Energy Research Foundation (Netherlands)

1998-05-01

Results from research in an EC Joule-III project and from national projects are presented. The objectives are improvement of design methods for stall regulated wind turbines with emphasis on stall induced vibrations and dynamic stall. The primary concern is limitation of the edgewise vibrations in the fundamental blade natural mode shape, which have caused trouble on modern wind turbines of approximate size 500 kW nominal power and 40 m rotor diameter. A theoretical study of quasi-steady aerodynamics confirms that the vibrations are driven basically by energy supplied from the aerodynamic forces during stalled operation. This energy exchange is equivalent to negative aerodynamic damping. The theoretical approach identifies the main parameters controlling the phenomenon. These parameters describe the steady and the dynamic airfoil characteristics, the overall aerodynamic layout of the blade, e.g. chord length and twist, the structural properties of the blade, e.g. structural damping and properties controlling the resulting vibration direction. Furthermore, full aeroelastic calculations and comparison with measurements show that the properties of the supporting structure, i.e. the main shaft, the nacelle and the tower, are important, as the global vibration of the rotor on its support may exchange energy with the blade vibration, when the blade natural frequency is close to one of the frequencies of the coupled rotor tilt-yaw mode shapes, usually denoted the global rotor whirl frequencies. It is confirmed that the influence of changing the primary design parameters can be determined by use of qualified aeroelastic calculations. Presented design guidelines therefore build on both the simple quasi-steady models, which can be used for the preliminary choice of the design variables mentioned above, and on full aeroelastic calculations. The aeroelastic calculations refine the design basis and should be used for choosing the final design variables and for final

A theory of post-stall transients in axial compression systems. I - Development of equations

Science.gov (United States)

Moore, F. K.; Greitzer, E. M.

1985-01-01

An approximate theory is presented for post-stall transients in multistage axial compression systems. The theory leads to a set of three simultaneous nonlinear third-order partial differential equations for pressure rise, and average and disturbed values of flow coefficient, as functions of time and angle around the compressor. By a Galerkin procedure, angular dependence is averaged, and the equations become first order in time. These final equations are capable of describing the growth and possible decay of a rotating-stall cell during a compressor mass-flow transient. It is shown how rotating-stall-like and surgelike motions are coupled through these equations, and also how the instantaneous compressor pumping characteristic changes during the transient stall process.

Evaluating Classroom Interaction with the iPad®: An Updated Stalling's Tool

Science.gov (United States)

MacKinnon, Gregory; Schep, Lourens; Borden, Lisa Lunney; Murray-Orr, Anne; Orr, Jeff; MacKinnon, Paula

2016-01-01

A large study of classrooms in the Caribbean context necessitated the use of a validated classroom observation tool. In practice, the paper-version Stalling's instrument (Stallings & Kaskowitz 1974) presented specific challenges with respect to (a) facile data collection and (b) qualitative observations of classrooms. In response to these…

Active Suppression of Rotating Stall Inception with Distributed Jet Actuation

Directory of Open Access Journals (Sweden)

Huu Duc Vo

2007-01-01

Full Text Available An analytical and experimental investigation of the effectiveness of full-span distributed jet actuation for active suppression of long length-scale rotating stall inception is carried out. Detailed modeling and experimental verification highlight the important effects of mass addition, discrete injectors, and feedback dynamics, which may be overlooked in preliminary theoretical studies of active control with jet injection. A model of the compression system incorporating nonideal injection and feedback dynamics is verified with forced response measurements to predict the right trends in the movement of the critical pole associated with the stall precursor. Active control experiments with proportional feedback control show that the predicted stall precursors are suppressed to give a 5.5% range extension in compressor flow coefficient. In addition, results suggest that the proposed model could be used to design a more sophisticated controller to further improve performance while reducing actuator bandwidth requirements.

Development and application of a dynamic stall model for rotating wind turbine blades

International Nuclear Information System (INIS)

Xu, B F; Yuan, Y; Wang, T G

2014-01-01

In unsteady conditions of wind turbines, both the dynamic stall phenomenon and the three-dimensional (3D) rotational effect affect the rotor aerodynamics. The dynamic stall mechanism for rotating wind turbine blades is first investigated. Through the comparison of the aerodynamic data between the rotating blade and the two-dimensional (2D) airfoil, the normal force slope in the attached flow and the separation point expression in the separated flow are modified in the Beddoes-Leishman (B-L) dynamic stall model for rotating NREL wind turbine blades. The modified model is validated by the comparison between the calculation results and the experimental results of the lift and drag coefficients at different radial positions. Both the hysteresis loop shapes and the calculation values are closer to the experiment than the 2D dynamic stall model. The present dynamic stall model is then coupled to a free vortex wake model. The coupled model is used to calculate the unsteady blade aerodynamic loads and the low speed shaft torque of the NREL wind turbine in a yawed condition. The accuracy is greatly improved by the corrections presented in the paper

Physical interaction between replication protein A (RPA) and MRN: involvement of RPA2 phosphorylation and the N-terminus of RPA1.

Science.gov (United States)

Oakley, Greg G; Tillison, Kristin; Opiyo, Stephen A; Glanzer, Jason G; Horn, Jeffrey M; Patrick, Steve M

2009-08-11

Replication protein A (RPA) is a heterotrimeric protein consisting of RPA1, RPA2, and RPA3 subunits that binds to single-stranded DNA (ssDNA) with high affinity. The response to replication stress requires the recruitment of RPA and the MRE11-RAD50-NBS1 (MRN) complex. RPA bound to ssDNA stabilizes stalled replication forks by recruiting checkpoint proteins involved in fork stabilization. MRN can bind DNA structures encountered at stalled or collapsed replication forks, such as ssDNA-double-stranded DNA (dsDNA) junctions or breaks, and promote the restart of DNA replication. Here, we demonstrate that RPA2 phosphorylation regulates the assembly of DNA damage-induced RPA and MRN foci. Using purified proteins, we observe a direct interaction between RPA with both NBS1 and MRE11. By utilizing RPA bound to ssDNA, we demonstrate that substituting RPA with phosphorylated RPA or a phosphomimetic weakens the interaction with the MRN complex. Also, the N-terminus of RPA1 is a critical component of the RPA-MRN protein-protein interaction. Deletion of the N-terminal oligonucleotide-oligosaccharide binding fold (OB-fold) of RPA1 abrogates interactions of RPA with MRN and individual proteins of the MRN complex. Further identification of residues critical for MRN binding in the N-terminus of RPA1 shows that substitution of Arg31 and Arg41 with alanines disrupts the RPA-MRN interaction and alters cell cycle progression in response to DNA damage. Thus, the N-terminus of RPA1 and phosphorylation of RPA2 regulate RPA-MRN interactions and are important in the response to DNA damage.

Comparison of driven and simulated "free" stall flutter in a wind tunnel

Science.gov (United States)

Culler, Ethan; Farnsworth, John; Fagley, Casey; Seidel, Jurgen

2016-11-01

Stall flutter and dynamic stall have received a significant amount of attention over the years. To experimentally study this problem, the body undergoing stall flutter is typically driven at a characteristic, single frequency sinusoid with a prescribed pitching amplitude and mean angle of attack offset. This approach allows for testing with repeatable kinematics, however it effectively decouples the structural motion from the aerodynamic forcing. Recent results suggest that this driven approach could misrepresent the forcing observed in a "free" stall flutter scenario. Specifically, a dynamically pitched rigid NACA 0018 wing section was tested in the wind tunnel under two modes of operation: (1) Cyber-Physical where "free" stall flutter was physically simulated through a custom motor-control system modeling a torsional spring and (2) Direct Motor-Driven Dynamic Pitch at a single frequency sinusoid representative of the cyber-physical motion. The time-resolved pitch angle and moment were directly measured and compared for each case. It was found that small deviations in the pitch angle trajectory between these two operational cases generate significantly different aerodynamic pitching moments on the wing section, with the pitching moments nearly 180o out of phase in some cases. This work is supported by the Air Force Office of Scientific Research through the Flow Interactions and Control Program and by the National Defense Science and Engineering Graduate Fellowship Program.

Simulasi Numerik Dynamic Stall Pada Airfoil Yang Berosilasi

Directory of Open Access Journals (Sweden)

Galih S.T.A. Bangga

2012-09-01

Full Text Available Kebutuhan analisa pada sudu helikopter, kompresor, kincir angin dan struktur streamline lainya yang beroperasi pada angle of attack yang tinggi dan melibatkan instationary effects yang disebut dynamic stall menjadi semakin penting. Fenomena ini ditandai dengan naiknya dynamic lift melewati static lift maksimum pada critical static stall angle, vortex yang terbentuk pada leading edge mengakibatkan naiknya suction contribution yang kemudian terkonveksi sepanjang permukaan hingga mencapai trailling edge diikuti terbentuknya trailling edge vortex yang menunjukkan terjadinya lift stall. Fenomena ini sangat berbahaya terhadap struktur airfoil itu sendiri. Secara umum, beban fatique yang ditimbulkan oleh adanya efek histerisis karena fluktuasi gaya lift akibat induksi vibrasi lebih besar dibandingkan kondisi statis. Simulasi numerik dilakukan secara 2D dengan menggunakan profil Boeing-Vertol V23010-1.58 pada α0 = 14.92°. Standard-kω dan SST-kω digunakan sebagai URANS turbulence modelling. Model osilasi dari airfoil disusun dalam suatu user defined function (UDF. Gerakan meshing beserta airfoil diakomodasi dengan menggunakan dynamic mesh approach. Simulasi numerik menunjukkan bahwa, model SST-kω menunjukkan performa yang lebih baik dibandingkan dengan Standard-kω. Fenomena travelling vortex yang terjadi mampu ditangkap dengan baik, meski pada angle of attack yang tinggi URANS turbulence model gagal memprediksikan fenomena yang terjadi karena dominasi efek 3D.

Effects of alley and stall surfaces on indices of claw and leg health in dairy cattle housed in a free-stall barn.

Science.gov (United States)

Vokey, F J; Guard, C L; Erb, H N; Galton, D M

2001-12-01

A 15-wk 2 x 3 factorial trial in a university dairy herd compared the effects of two alley surfaces and three free-stall beds on indices of lameness. Alley surfaces were grooved concrete (Ct) or 1.9-cm-thick interlocking rubber mats (R). Stalls were deep sand (S), rubber mattresses (M), or concrete (C). Mattress and concrete stalls were bedded with sawdust. At wk 1 and 15, the hind claws and hocks of 120 primi- (n = 69) and multiparous (n = 51) cows were scored for lesions and three claw measurements (dorsal wall length, heel depth, and toe angle) were recorded. Rates of lateral and medial claw growth and wear were calculated by measuring the migration of a reference mark away from the coronet. Digital photographs of claw surfaces were used to rescore claw lesions. Clinical lameness was evaluated by assigning a locomotion score from 1 to 4 to each cow during wk 1, 5, 10, and 14. Digital dermatitis (present/not present) and interdigital dermatitis (mild, moderate, or severe) were recorded at wk 15. The number of days that cows spent in a hospital barn was recorded. Before assignment, cows were professionally foot trimmed, sorted by initial claw lesion score, and then randomized in consecutive blocks of three to stall treatments. Photograph scores were highly repeatable. Nonparametric statistical techniques were used for analyses of rank data. Claw lesion score increased significantly for all treatment groups except RC and RS; however, when early lactation cows were excluded, no differences were found between treatment groups. Hock scores increased significantly more for cows in CtC than in CtS or RS. Significantly more animals from RC spent more than 10 d in the hospital pen compared with RM and RS. Groups did not significantly differ for clinical lameness. Cows in RS and RC had significantly lower rates for lateral claw net growth than those in CtM. Having moderate or severe interdigital dermatitis at wk 15 was associated with greater increases in claw lesion score

Cow comfort in tie-stalls: increased depth of shavings or straw bedding increases lying time.

Science.gov (United States)

Tucker, C B; Weary, D M; von Keyserlingk, M A G; Beauchemin, K A

2009-06-01

Over half of US dairy operations use tie-stalls, but these farming systems have received relatively little research attention in terms of stall design and management. The current study tested the effects of the amount of 2 bedding materials, straw and shavings, on dairy cattle lying behavior. The effects of 4 levels of shavings, 3, 9, 15, and 24 kg/stall (experiment 1, n = 12), and high and low levels of straw in 2 separate experiments: 1, 3, 5, and 7 kg/stall (experiment 2, n = 12) and 0.5, 1, 2, and 3 kg/stall (experiment 3, n = 12) were assessed. Treatments were compared using a crossover design with lactating cows housed in tie-stalls fitted with mattresses. Treatments were applied for 1 wk. Total lying time, number of lying bouts, and the length of each lying bout was recorded with data loggers. In experiment 1, cows spent 3 min more lying down for each additional kilogram of shavings (11.0, 11.7, 11.6, and 12.1 +/- 0.24 h/d for 3, 9, 15, and 24 kg/stall shavings, respectively). In experiment 2, cows increased lying time by 12 min for every additional kilogram of straw (11.2, 12.0, 11.8, and 12.4 +/- 0.24 h/d for 1, 3, 5, and 7 kg/stall of straw, respectively). There were no differences in lying behavior among the lower levels of straw tested in experiment 3 (11.7 +/- 0.32 h/d). These results indicated that additional bedding above a scant amount improves cow comfort, as measured by lying time, likely because a well-bedded surface is more compressible.

Combustion-Powered Actuation for Dynamic Stall Suppression - Simulations and Low-Mach Experiments

Science.gov (United States)

Matalanis, Claude G.; Min, Byung-Young; Bowles, Patrick O.; Jee, Solkeun; Wake, Brian E.; Crittenden, Tom; Woo, George; Glezer, Ari

2014-01-01

An investigation on dynamic-stall suppression capabilities of combustion-powered actuation (COMPACT) applied to a tabbed VR-12 airfoil is presented. In the first section, results from computational fluid dynamics (CFD) simulations carried out at Mach numbers from 0.3 to 0.5 are presented. Several geometric parameters are varied including the slot chordwise location and angle. Actuation pulse amplitude, frequency, and timing are also varied. The simulations suggest that cycle-averaged lift increases of approximately 4% and 8% with respect to the baseline airfoil are possible at Mach numbers of 0.4 and 0.3 for deep and near-deep dynamic-stall conditions. In the second section, static-stall results from low-speed wind-tunnel experiments are presented. Low-speed experiments and high-speed CFD suggest that slots oriented tangential to the airfoil surface produce stronger benefits than slots oriented normal to the chordline. Low-speed experiments confirm that chordwise slot locations suitable for Mach 0.3-0.4 stall suppression (based on CFD) will also be effective at lower Mach numbers.

Effect of artificial UV irradiation on spore content of stall air and fattening pig breeding

International Nuclear Information System (INIS)

Kalich, J.; Blendl, H.M.

1978-01-01

The influence of a continuous UV irradiation (emitter NN 33/89 original Hanau) during the fattening periods primarily in the bactericide region of 253.7 nm of various intensities on the spore content of air, on the state of health and on the fattening breeding of pigs was tested in two fattening procedures. The high spore number per m 3 air of over 700 000 upon occupying the stall in the first fattening procedure was reduced by 90.5% to about 70 000 after 1 week of UV irradiation, and in the second procedure, from 111 500 to 16 000, i.e. a reduction of 85.5%. The spore content of the stall air then exhibited large deviations reducing and increasing. The same deviations were recorded for dust content. There was no absolute correlation between dust and spore content of the air until the 11th week after starting UV irradiation in either test. The spore content sank in the reference stalls also without UV irradiation, by 29.9% in the first fattening procedure 1 week after occupying the stall and even by 75% in the second procedure. The spore content of the air in the reference stalls also then exhibited deviations sinking and rising as in the test stalls with UV irradiation. Here too, there was no correlation between dust and spore content of the air. The spore content in the air was 2 to 7 times higher in the reference stalls than in the test stalls. One may conclude from the tests that the promoting irradiation strength is between 15 and 20 μW/cm 2 and that short-term stool production in danish stalling, 60 μW/cm 2 are not harmful. Air disinfection with UV irradiation, can only be part of the total hygiene measures taken in veterinary medicine and may only be considered as an important link in the chain of the health promoting and increased efficient hygiene measures in the intensification of aggriculturally useful animals. (orig./AJ) [de

Enhancement of internal ribosome entry site-mediated translation and replication of hepatitis C virus by PD98059

International Nuclear Information System (INIS)

Murata, Takayuki; Hijikata, Makoto; Shimotohno, Kunitada

2005-01-01

Translation initiation of hepatitis C virus (HCV) occurs in an internal ribosome entry site (IRES)-dependent manner. We found that HCV IRES-dependent protein synthesis is enhanced by PD98059, an inhibitor of the extracellular signal-regulated kinase (ERK) signaling pathway, while cellular cap-dependent translation was relatively unaffected by the compound. Treatment of cells with PD98059 allowed for robust HCV replication following cellular incubation with HCV-positive serum. Though the molecular mechanism underlying IRES enhancement remains elusive, PD98059 is a potent accelerator of HCV RNA replication

Modeling dynamic stall on wind turbine blades under rotationally augmented flow fields

Energy Technology Data Exchange (ETDEWEB)

Guntur, S. [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Schreck, S. [National Renewable Energy Laboratory (NREL), Golden, CO (United States); Sorensen, N. N. [Technical Univ. of Denmark, Lyngby (Denmark); Bergami, L. [Technical Univ. of Denmark, Lyngby (Denmark)

2015-04-22

It is well known that airfoils under unsteady flow conditions with a periodically varying angle of attack exhibit aerodynamic characteristics different from those under steady flow conditions, a phenomenon commonly known as dynamic stall. It is also well known that the steady aerodynamic characteristics of airfoils in the inboard region of a rotating blade differ from those under steady two-dimensional (2D) flow conditions, a phenomenon commonly known as rotational augmentation. This paper presents an investigation of these two phenomena together in the inboard parts of wind turbine blades. This analysis is carried out using data from three sources: (1) the National Renewable Energy Laboratory’s Unsteady Aerodynamics Experiment Phase VI experimental data, including constant as well as continuously pitching blade conditions during axial operation, (2) data from unsteady Delayed Detached Eddy Simulations (DDES) carried out using the Technical University of Denmark’s in-house flow solver Ellipsys3D, and (3) data from a simplified model based on the blade element momentum method with a dynamic stall subroutine that uses rotationally augmented steady-state polars obtained from steady Phase VI experimental sequences, instead of the traditional 2D nonrotating data. The aim of this work is twofold. First, the blade loads estimated by the DDES simulations are compared to three select cases of the N sequence experimental data, which serves as a validation of the DDES method. Results show reasonable agreement between the two data in two out of three cases studied. Second, the dynamic time series of the lift and the moment polars obtained from the experiments are compared to those from the dynamic stall subroutine that uses the rotationally augmented steady polars. This allowed the differences between the stall phenomenon on the inboard parts of harmonically pitching blades on a rotating wind turbine and the classic dynamic stall representation in 2D flow to be

Study and Control of a Radial Vaned Diffuser Stall

Directory of Open Access Journals (Sweden)

Aurélien Marsan

2012-01-01

Full Text Available The aim of the present study is to evaluate the efficiency of a boundary layer suction technique in case of a centrifugal compressor stage in order to extend its stable operating range. First, an analysis of the flow pattern within the radial vaned diffuser is presented. It highlights the stall of the diffuser vanes when reaching a low massflow. A boundary layer separation in the hub-suction side corner grows when decreasing the massflow from the nominal operating point to the surge and finally leads to a massive stall. An aspiration strategy is investigated in order to control the stall. The suction slot is put in the vicinity of the saddle that originates the main separating skin-friction line, identified thanks to the analysis of the skin-friction pattern. Several aspiration massflow rates are tested, and two different modelings of the aspiration are evaluated. Finally, an efficient control is reached with a removal of only 0,1% of the global massflow and leads—from a steady-state calculations point of view—to an increase by 40% of the compressor operating range extent.

Questionnaire-based study to assess the association between management practices and mastitis within tie-stall and free-stall dairy housing systems in Switzerland

Science.gov (United States)

2013-01-01

Background Prophylactic measures are key components of dairy herd mastitis control programs, but some are only relevant in specific housing systems. To assess the association between management practices and mastitis incidence, data collected in 2011 by a survey among 979 randomly selected Swiss dairy farms, and information from the regular test day recordings from 680 of these farms was analyzed. Results The median incidence of farmer-reported clinical mastitis (ICM) was 11.6 (mean 14.7) cases per 100 cows per year. The median annual proportion of milk samples with a composite somatic cell count (PSCC) above 200,000 cells/ml was 16.1 (mean 17.3) %. A multivariable negative binomial regression model was fitted for each of the mastitis indicators for farms with tie-stall and free-stall housing systems separately to study the effect of other (than housing system) management practices on the ICM and PSCC events (above 200,000 cells/ml). The results differed substantially by housing system and outcome. In tie-stall systems, clinical mastitis incidence was mainly affected by region (mountainous production zone; incidence rate ratio (IRR) = 0.73), the dairy herd replacement system (1.27) and farmers age (0.81). The proportion of high SCC was mainly associated with dry cow udder controls (IRR = 0.67), clean bedding material at calving (IRR = 1.72), using total merit values to select bulls (IRR = 1.57) and body condition scoring (IRR = 0.74). In free-stall systems, the IRR for clinical mastitis was mainly associated with stall climate/temperature (IRR = 1.65), comfort mats as resting surface (IRR = 0.75) and when no feed analysis was carried out (IRR = 1.18). The proportion of high SSC was only associated with hand and arm cleaning after calving (IRR = 0.81) and beef producing value to select bulls (IRR = 0.66). Conclusions There were substantial differences in identified risk factors in the four models. Some of the factors were in agreement with the reported literature

Human FAN1 promotes strand incision in 5'-flapped DNA complexed with RPA.

Science.gov (United States)

Takahashi, Daisuke; Sato, Koichi; Hirayama, Emiko; Takata, Minoru; Kurumizaka, Hitoshi

2015-09-01

Fanconi anaemia (FA) is a human infantile recessive disorder. Seventeen FA causal proteins cooperatively function in the DNA interstrand crosslink (ICL) repair pathway. Dual DNA strand incisions around the crosslink are critical steps in ICL repair. FA-associated nuclease 1 (FAN1) is a DNA structure-specific endonuclease that is considered to be involved in DNA incision at the stalled replication fork. Replication protein A (RPA) rapidly assembles on the single-stranded DNA region of the stalled fork. However, the effect of RPA on the FAN1-mediated DNA incision has not been determined. In this study, we purified human FAN1, as a bacterially expressed recombinant protein. FAN1 exhibited robust endonuclease activity with 5'-flapped DNA, which is formed at the stalled replication fork. We found that FAN1 efficiently promoted DNA incision at the proper site of RPA-coated 5'-flapped DNA. Therefore, FAN1 possesses the ability to promote the ICL repair of 5'-flapped DNA covered by RPA. © The Authors 2015. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved.

Analysis of Low-Speed Stall Aerodynamics of a Business Jets Wing Using STAR-CCM+

Science.gov (United States)

Bui, Trong

2016-01-01

Reynolds-Averaged Navier-Stokes (RANS) computational fluid dynamics (CFD) analysis was conducted: to study the low-speed stall aerodynamics of a GIII aircrafts swept wing modified with (1) a laminar-flow wing glove, or (2) a seamless flap. The stall aerodynamics of these two different wing configurations were analyzed and compared with the unmodified baseline wing for low-speed flight. The Star-CCM+ polyhedral unstructured CFD code was first validated for wing stall predictions using the wing-body geometry from the First AIAA CFD High-Lift Prediction Workshop.

Nonlinear control of rotating stall and surge with axisymmetric bleed and air injection on axial flow compressors

Science.gov (United States)

Yeung, Chung-Hei (Simon)

The study of compressor instabilities in gas turbine engines has received much attention in recent years. In particular, rotating stall and surge are major causes of problems ranging from component stress and lifespan reduction to engine explosion. In this thesis, modeling and control of rotating stall and surge using bleed valve and air injection is studied and validated on a low speed, single stage, axial compressor at Caltech. Bleed valve control of stall is achieved only when the compressor characteristic is actuated, due to the fast growth rate of the stall cell compared to the rate limit of the valve. Furthermore, experimental results show that the actuator rate requirement for stall control is reduced by a factor of fourteen via compressor characteristic actuation. Analytical expressions based on low order models (2--3 states) and a high fidelity simulation (37 states) tool are developed to estimate the minimum rate requirement of a bleed valve for control of stall. A comparison of the tools to experiments show a good qualitative agreement, with increasing quantitative accuracy as the complexity of the underlying model increases. Air injection control of stall and surge is also investigated. Simultaneous control of stall and surge is achieved using axisymmetric air injection. Three cases with different injector back pressure are studied. Surge control via binary air injection is achieved in all three cases. Simultaneous stall and surge control is achieved for two of the cases, but is not achieved for the lowest authority case. This is consistent with previous results for control of stall with axisymmetric air injection without a plenum attached. Non-axisymmetric air injection control of stall and surge is also studied. Three existing control algorithms found in literature are modeled and analyzed. A three-state model is obtained for each algorithm. For two cases, conditions for linear stability and bifurcation criticality on control of rotating stall are

How MCM loading and spreading specify eukaryotic DNA replication initiation sites [version 1; referees: 4 approved

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Olivier Hyrien

2016-08-01

Full Text Available DNA replication origins strikingly differ between eukaryotic species and cell types. Origins are localized and can be highly efficient in budding yeast, are randomly located in early fly and frog embryos, which do not transcribe their genomes, and are clustered in broad (10-100 kb non-transcribed zones, frequently abutting transcribed genes, in mammalian cells. Nonetheless, in all cases, origins are established during the G1-phase of the cell cycle by the loading of double hexamers of the Mcm 2-7 proteins (MCM DHs, the core of the replicative helicase. MCM DH activation in S-phase leads to origin unwinding, polymerase recruitment, and initiation of bidirectional DNA synthesis. Although MCM DHs are initially loaded at sites defined by the binding of the origin recognition complex (ORC, they ultimately bind chromatin in much greater numbers than ORC and only a fraction are activated in any one S-phase. Data suggest that the multiplicity and functional redundancy of MCM DHs provide robustness to the replication process and affect replication time and that MCM DHs can slide along the DNA and spread over large distances around the ORC. Recent studies further show that MCM DHs are displaced along the DNA by collision with transcription complexes but remain functional for initiation after displacement. Therefore, eukaryotic DNA replication relies on intrinsically mobile and flexible origins, a strategy fundamentally different from bacteria but conserved from yeast to human. These properties of MCM DHs likely contribute to the establishment of broad, intergenic replication initiation zones in higher eukaryotes.

Recent Insight into the Kinetic Mechanisms and Conformational Dynamics of Y-Family DNA Polymerases

OpenAIRE

Maxwell, Brian A.; Suo, Zucai

2014-01-01

The kinetic mechanisms by which DNA polymerases catalyze DNA replication and repair have long been areas of active research. Recently discovered Y-family DNA polymerases catalyze the bypass of damaged DNA bases that would otherwise block replicative DNA polymerases and stall replication forks. Unlike DNA polymerases from the five other families, the Y-family DNA polymerases have flexible, solvent-accessible active sites that are able to tolerate various types of damaged template bases and all...

Internal Flow of a High Specific-Speed Diagonal-Flow Fan (Rotor Outlet Flow Fields with Rotating Stall

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Norimasa Shiomi

2003-01-01

Full Text Available We carried out investigations for the purpose of clarifying the rotor outlet flow fields with rotating stall cell in a diagonal-flow fan. The test fan was a high–specific-speed (ns=1620 type of diagonal-flow fan that had 6 rotor blades and 11 stator blades. It has been shown that the number of the stall cell is 1, and its propagating speed is approximately 80% of its rotor speed, although little has been known about the behavior of the stall cell because a flow field with a rotating stall cell is essentially unsteady. In order to capture the behavior of the stall cell at the rotor outlet flow fields, hot-wire surveys were performed using a single-slant hotwire probe. The data obtained by these surveys were processed by means of a double phase-locked averaging technique, which enabled us to capture the flow field with the rotating stall cell in the reference coordinate system fixed to the rotor. As a result, time-dependent ensemble averages of the three-dimensional velocity components at the rotor outlet flow fields were obtained. The behavior of the stall cell was shown for each velocity component, and the flow patterns on the meridional planes were illustrated.

Conical Magnetic Bearings Developed for Active Stall Control in Gas Turbine Engines

Science.gov (United States)

Trudell, Jeffrey J.; Kascak, Albert F.; Provenza, Andrew J.; Buccieri, Carl J.

2004-01-01

Active stall control is a current research area at the NASA Glenn Research Center that offers a great benefit in specific fuel consumption by allowing the gas turbine to operate beyond the onset of stall. Magnetic bearings are being investigated as a new method to perform active stall control. This enabling global aviation safety technology would result in improved fuel efficiency and decreased carbon dioxide emissions, as well as improve safety and reliability by eliminating oil-related delays and failures of engine components, which account for 40 percent of the commercial aircraft departure delays. Active stall control works by perturbing the flow in front of the compressor stage such that it cancels the pressure wave, which causes the compressor to go into stall. Radial magnetic bearings are able to whirl the shaft so that variations in blade tip leakage would flow upstream causing a perturbation wave that could cancel the rotating stall cell. Axial or thrust magnetic bearings cannot be used to cancel the surge mode in the compressor because they have a very low bandwidth and thus cannot modulate at a high enough frequency. Frequency response is limited because the thrust runner cannot be laminated. To improve the bandwidth of magnetic thrust bearings, researchers must use laminations to suppress the eddy currents. A conical magnetic bearing can be laminated, resulting in increased bandwidth in the axial direction. In addition, this design can produce both radial and thrust force in a single bearing, simplifying the installation. The proposed solution combines the radial and thrust bearing into one design that can be laminated--a conical magnetic bearing. The new conical magnetic bearing test rig, funded by a Glenn fiscal year 2002 Director's Discretionary Fund, was needed because none of the existing rigs has an axial degree of freedom. The rotor bearing configuration will simulate that of the main shaft on a gas turbine engine. One conical magnetic bearing

A dynamic stall model for airfoils with deformable trailing edges

DEFF Research Database (Denmark)

Andersen, Peter Bjørn; Gaunaa, Mac; Bak, Dan Christian

2007-01-01

on an airfoil section undergoing arbitrary motion in heave, lead-lag, pitch, Trailing Edge (TE) flapping. In the linear region, the model reduces to the inviscid model of Gaunaa [4], which includes the aerodynamic effect of a thin airfoil with a deformable camberline in inviscid flow. Therefore, the proposed......The present work contains an extension of the Beddoes-Leishman (B-L) type dynamic stall model, as described by Hansen et al. [7]. In this work a Deformable Trailing Edge Geometry (DTEG) has been added to the dynamic stall model. The model predicts the unsteady aerodynamic forces and moments...

Model of OSBP-Mediated Cholesterol Supply to Aichi Virus RNA Replication Sites Involving Protein-Protein Interactions among Viral Proteins, ACBD3, OSBP, VAP-A/B, and SAC1.

Science.gov (United States)

Ishikawa-Sasaki, Kumiko; Nagashima, Shigeo; Taniguchi, Koki; Sasaki, Jun

2018-04-15

Positive-strand RNA viruses, including picornaviruses, utilize cellular machinery for genome replication. Previously, we reported that each of the 2B, 2BC, 2C, 3A, and 3AB proteins of Aichi virus (AiV), a picornavirus, forms a complex with the Golgi apparatus protein ACBD3 and phosphatidylinositol 4-kinase IIIβ (PI4KB) at viral RNA replication sites (replication organelles [ROs]), enhancing PI4KB-dependent phosphatidylinositol 4-phosphate (PI4P) production. Here, we demonstrate AiV hijacking of the cellular cholesterol transport system involving oxysterol-binding protein (OSBP), a PI4P-binding cholesterol transfer protein. AiV RNA replication was inhibited by silencing cellular proteins known to be components of this pathway, OSBP, the ER membrane proteins VAPA and VAPB (VAP-A/B), the PI4P-phosphatase SAC1, and PI-transfer protein β. OSBP, VAP-A/B, and SAC1 were present at RNA replication sites. We also found various previously unknown interactions among the AiV proteins (2B, 2BC, 2C, 3A, and 3AB), ACBD3, OSBP, VAP-A/B, and SAC1, and the interactions were suggested to be involved in recruiting the component proteins to AiV ROs. Importantly, the OSBP-2B interaction enabled PI4P-independent recruitment of OSBP to AiV ROs, indicating preferential recruitment of OSBP among PI4P-binding proteins. Protein-protein interaction-based OSBP recruitment has not been reported for other picornaviruses. Cholesterol was accumulated at AiV ROs, and inhibition of OSBP-mediated cholesterol transfer impaired cholesterol accumulation and AiV RNA replication. Electron microscopy showed that AiV-induced vesicle-like structures were close to ER membranes. Altogether, we conclude that AiV directly recruits the cholesterol transport machinery through protein-protein interactions, resulting in formation of membrane contact sites between the ER and AiV ROs and cholesterol supply to the ROs. IMPORTANCE Positive-strand RNA viruses utilize host pathways to modulate the lipid composition of

A time-varying subjective quality model for mobile streaming videos with stalling events

Science.gov (United States)

Ghadiyaram, Deepti; Pan, Janice; Bovik, Alan C.

2015-09-01

Over-the-top mobile video streaming is invariably influenced by volatile network conditions which cause playback interruptions (stalling events), thereby impairing users' quality of experience (QoE). Developing models that can accurately predict users' QoE could enable the more efficient design of quality-control protocols for video streaming networks that reduce network operational costs while still delivering high-quality video content to the customers. Existing objective models that predict QoE are based on global video features, such as the number of stall events and their lengths, and are trained and validated on a small pool of ad hoc video datasets, most of which are not publicly available. The model we propose in this work goes beyond previous models as it also accounts for the fundamental effect that a viewer's recent level of satisfaction or dissatisfaction has on their overall viewing experience. In other words, the proposed model accounts for and adapts to the recency, or hysteresis effect caused by a stall event in addition to accounting for the lengths, frequency of occurrence, and the positions of stall events - factors that interact in a complex way to affect a user's QoE. On the recently introduced LIVE-Avvasi Mobile Video Database, which consists of 180 distorted videos of varied content that are afflicted solely with over 25 unique realistic stalling events, we trained and validated our model to accurately predict the QoE, attaining standout QoE prediction performance.

DNA Replication Dynamics of the GGGGCC Repeat of the C9orf72 Gene.

Science.gov (United States)

Thys, Ryan Griffin; Wang, Yuh-Hwa

2015-11-27

DNA has the ability to form a variety of secondary structures in addition to the normal B-form DNA, including hairpins and quadruplexes. These structures are implicated in a number of neurological diseases and cancer. Expansion of a GGGGCC repeat located at C9orf72 is associated with familial amyotrophic lateral sclerosis and frontotemporal dementia. This repeat expands from two to 24 copies in normal individuals to several hundreds or thousands of repeats in individuals with the disease. Biochemical studies have demonstrated that as little as four repeats have the ability to form a stable DNA secondary structure known as a G-quadruplex. Quadruplex structures have the ability to disrupt normal DNA processes such as DNA replication and transcription. Here we examine the role of GGGGCC repeat length and orientation on DNA replication using an SV40 replication system in human cells. Replication through GGGGCC repeats leads to a decrease in overall replication efficiency and an increase in instability in a length-dependent manner. Both repeat expansions and contractions are observed, and replication orientation is found to influence the propensity for expansions or contractions. The presence of replication stress, such as low-dose aphidicolin, diminishes replication efficiency but has no effect on instability. Two-dimensional gel electrophoresis analysis demonstrates a replication stall with as few as 20 GGGGCC repeats. These results suggest that replication of the GGGGCC repeat at C9orf72 is perturbed by the presence of expanded repeats, which has the potential to result in further expansion, leading to disease. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Analysis of compressible light dynamic stall flow at transitional Reynolds numbers

DEFF Research Database (Denmark)

Dyken, R.D. Van; Ekaterinaris, John A.; Chandrasekhara, M.S.

1996-01-01

Numerical and experimental results of steady and light dynamic stall flow over an oscillating NACA 0012 airfoil at a freestream Mach number of 0.3 and Reynolds number of 0.54 x 10(6) are compared, The experimental observation that dynamic stall is induced from the bursting of a laminar separation...... point is specified suitably and a simple transition length model is incorporated to determine the extent of the laminar separation bubble. The thin-layer approximations of compressible, Reynolds-averaged, Navier-Stokes equations are used for the numerical solution, with an implicit, upwind-biased, third...

Telomere healing following DNA polymerase arrest-induced breakages is likely the main mechanism generating chromosome 4p terminal deletions.

Science.gov (United States)

Hannes, Femke; Van Houdt, Jeroen; Quarrell, Oliver W; Poot, Martin; Hochstenbach, Ron; Fryns, Jean-Pierre; Vermeesch, Joris R

2010-12-01

Constitutional developmental disorders are frequently caused by terminal chromosomal deletions. The mechanisms and/or architectural features that might underlie those chromosome breakages remain largely unexplored. Because telomeres are the vital DNA protein complexes stabilizing linear chromosomes against chromosome degradation, fusion, and incomplete replication, those terminal-deleted chromosomes acquired new telomeres either by telomere healing or by telomere capture. To unravel the mechanisms leading to chromosomal breakage and healing, we sequenced nine chromosome 4p terminal deletion boundaries. A computational analysis of the breakpoint flanking region, including 12 previously published pure terminal breakage sites, was performed in order to identify architectural features that might be involved in this process. All terminal 4p truncations were likely stabilized by telomerase-mediated telomere healing. In the majority of breakpoints multiple genetic elements have a potential to induce secondary structures and an enrichment in replication stalling site motifs were identified. These findings suggest DNA replication stalling-induced chromosome breakage during early development is the first mechanistic step leading toward terminal deletion syndromes. © 2010 Wiley-Liss, Inc.

RPA-Mediated Recruitment of the E3 Ligase RFWD3 Is Vital for Interstrand Crosslink Repair and Human Health.

Science.gov (United States)

Feeney, Laura; Muñoz, Ivan M; Lachaud, Christophe; Toth, Rachel; Appleton, Paul L; Schindler, Detlev; Rouse, John

2017-06-01

Defects in the repair of DNA interstrand crosslinks (ICLs) are associated with the genome instability syndrome Fanconi anemia (FA). Here we report that cells with mutations in RFWD3, an E3 ubiquitin ligase that interacts with and ubiquitylates replication protein A (RPA), show profound defects in ICL repair. An amino acid substitution in the WD40 repeats of RFWD3 (I639K) found in a new FA subtype abolishes interaction of RFWD3 with RPA, thereby preventing RFWD3 recruitment to sites of ICL-induced replication fork stalling. Moreover, single point mutations in the RPA32 subunit of RPA that abolish interaction with RFWD3 also inhibit ICL repair, demonstrating that RPA-mediated RFWD3 recruitment to stalled replication forks is important for ICL repair. We also report that unloading of RPA from sites of ICL induction is perturbed in RFWD3-deficient cells. These data reveal important roles for RFWD3 localization in protecting genome stability and preserving human health. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Intrinsic bent DNA sites in the chromosomal replication origin of Xylella fastidiosa 9a5c

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F. Gimenes

2008-04-01

Full Text Available The features of the nucleotide sequences in both replication and promoter regions have been investigated in many organisms. Intrinsically bent DNA sites associated with transcription have been described in several prokaryotic organisms. The aim of the present study was to investigate intrinsic bent DNA sites in the segment that holds the chromosomal replication origin, oriC, of Xylella fastidiosa 9a5c. Electrophoretic behavior analyses, as well as in silico analyses of both the 2-D projection and helical parameters, were performed. The chromosomal segment analyzed contains the initial sequence of the rpmH gene, an intergenic region, the dnaA gene, the oriC sequence, and the 5' partial sequence of the dnaN gene. The analysis revealed fragments with reduced electrophoretic mobility, which indicates the presence of curved DNA segments. The analysis of the helical parameter ENDS ratio revealed three bent DNA sites (b1, b2, and b3 located in the rpmH-dnaA intergenic region, the dnaA gene, and the oriC 5' end, respectively. The chromosomal segment of X. fastidiosa analyzed here is rich in phased AT tracts and in CAnT motifs. The 2-D projection indicated a segment whose structure was determined by the cumulative effect of all bent DNA sites. Further, the in silico analysis of the three different bacterial oriC sequences indicated similar negative roll and twist >34.00° values. The DnaA box sequences, and other motifs in them, may be associated with the intrinsic DNA curvature.

The computation of the post-stall behavior of a circulation controlled airfoil

Science.gov (United States)

Linton, Samuel W.

1993-01-01

The physics of the circulation controlled airfoil is complex and poorly understood, particularly with regards to jet stall, which is the eventual breakdown of lift augmentation by the jet at some sufficiently high blowing rate. The present paper describes the numerical simulation of stalled and unstalled flows over a two-dimensional circulation controlled airfoil using a fully implicit Navier-Stokes code, and the comparison with experimental results. Mach numbers of 0.3 and 0.5 and jet total to freestream pressure ratios of 1.4 and 1.8 are investigated. The Baldwin-Lomax and k-epsilon turbulence models are used, each modified to include the effect of strong streamline curvature. The numerical solutions of the post-stall circulation controlled airfoil show a highly regular unsteady periodic flowfield. This is the result of an alternation between adverse pressure gradient and shock induced separation of the boundary layer on the airfoil trailing edge.

Stalled RNAP-II molecules bound to non-coding rDNA spacers are required for normal nucleolus architecture.

Science.gov (United States)

Freire-Picos, M A; Landeira-Ameijeiras, V; Mayán, María D

2013-07-01

The correct distribution of nuclear domains is critical for the maintenance of normal cellular processes such as transcription and replication, which are regulated depending on their location and surroundings. The most well-characterized nuclear domain, the nucleolus, is essential for cell survival and metabolism. Alterations in nucleolar structure affect nuclear dynamics; however, how the nucleolus and the rest of the nuclear domains are interconnected is largely unknown. In this report, we demonstrate that RNAP-II is vital for the maintenance of the typical crescent-shaped structure of the nucleolar rDNA repeats and rRNA transcription. When stalled RNAP-II molecules are not bound to the chromatin, the nucleolus loses its typical crescent-shaped structure. However, the RNAP-II interaction with Seh1p, or cryptic transcription by RNAP-II, is not critical for morphological changes. Copyright © 2013 John Wiley & Sons, Ltd.

Parametric analyses on dynamic stall control of rotor airfoil via synthetic jet

Directory of Open Access Journals (Sweden)

Qijun ZHAO

2017-12-01

Full Text Available The effects of synthetic jet control on unsteady dynamic stall over rotor airfoil are investigated numerically. A moving-embedded grid method and an Unsteady Reynolds Averaged Navier-Stokes (URANS solver coupled with k-ω Shear Stress Transport (SST turbulence model are established for predicting the complex flowfields of oscillatory airfoil under jet control. Additionally, a velocity boundary condition modeled by sinusoidal function has been developed to fulfill the perturbation effect of periodic jet. The validity of present CFD method is evaluated by comparisons of the calculated results of baseline dynamic stall case for rotor airfoil and jet control case for VR-7B airfoil with experimental data. Then, parametric analyses are conducted emphatically for an OA212 rotor airfoil to investigate the effects of jet control parameters (jet location, dimensionless frequency, momentum coefficient, jet angle, jet type and dual-jet on dynamic stall characteristics of rotor airfoil. It is demonstrated by the calculated results that efficiency of jet control could be improved with specific momentum coefficient and jet angle when the jet is located near separation point of rotor airfoil. Furthermore, the dual-jet could improve control efficiency more obviously on dynamic stall of rotor airfoil with respect to the unique jet, and the influence laws of dual-jet’s angles and momentum coefficients on control effects are similar to those of the unique jet. Finally, unsteady aerodynamic characteristics of rotor via synthetic jet which is located on the upper surface of rotor blade in forward flight are calculated, and as a result, the aerodynamic characteristics of rotor are improved compared with the baseline. The results indicate that synthetic jet has the capability in improving aerodynamic characteristics of rotor. Keywords: Airfoil, Dynamic stall characteristics, Flow control, Moving-embedded grid methodology, Navier-Stokes equations, Parametric

Time Accurate Unsteady Simulation of the Stall Inception Process in the Compression System of a US Army Helicopter Gas Turbine Engine

National Research Council Canada - National Science Library

Hathaway, Michael D; Herrick, Greg; Chen, Jenping; Webster, Robert

2004-01-01

.... Improved understanding of the stall inception process and how stall control technologies mitigate such will provide compressors with increased tolerance to stall, thereby expanding the operational...

Aerodynamic shape optimization for alleviating dynamic stall characteristics of helicopter rotor airfoil

Directory of Open Access Journals (Sweden)

Wang Qing

2015-04-01

Full Text Available In order to alleviate the dynamic stall effects in helicopter rotor, the sequential quadratic programming (SQP method is employed to optimize the characteristics of airfoil under dynamic stall conditions based on the SC1095 airfoil. The geometry of airfoil is parameterized by the class-shape-transformation (CST method, and the C-topology body-fitted mesh is then automatically generated around the airfoil by solving the Poisson equations. Based on the grid generation technology, the unsteady Reynolds-averaged Navier-Stokes (RANS equations are chosen as the governing equations for predicting airfoil flow field and the highly-efficient implicit scheme of lower–upper symmetric Gauss–Seidel (LU-SGS is adopted for temporal discretization. To capture the dynamic stall phenomenon of the rotor more accurately, the Spalart–Allmaras turbulence model is employed to close the RANS equations. The optimized airfoil with a larger leading edge radius and camber is obtained. The leading edge vortex and trailing edge separation of the optimized airfoil under unsteady conditions are obviously weakened, and the dynamic stall characteristics of optimized airfoil at different Mach numbers, reduced frequencies and angles of attack are also obviously improved compared with the baseline SC1095 airfoil. It is demonstrated that the optimized method is effective and the optimized airfoil is suitable as the helicopter rotor airfoil.

Constitutive role of the Fanconi anemia D2 gene in the replication stress response.

Science.gov (United States)

Tian, Yanyan; Shen, Xi; Wang, Rui; Klages-Mundt, Naeh L; Lynn, Erica J; Martin, Sara K; Ye, Yin; Gao, Min; Chen, Junjie; Schlacher, Katharina; Li, Lei

2017-12-08

In response to DNA cross-linking damage, the Fanconi anemia (FA) core complex activates the FA pathway by monoubiquitinating Fanconi anemia complementation group D2 (FANCD2) for the initiation of the nucleolytic processing of the DNA cross-links and stabilization of stalled replication forks. Given that all the classic FA proteins coordinately monoubiquitinate FANCD2, it is unclear why losses of individual classic FA genes yield varying cellular sensitivities to cross-linking damage. To address this question, we generated cellular knock-out models of FA core complex components and FANCD2 and found that FANCD2-null mutants display higher levels of spontaneous chromosomal damage and hypersensitivity to replication-blocking lesions than Fanconi anemia complementation group L (FANCL)-null mutants, suggesting that FANCD2 provides a basal level of DNA protection countering endogenous lesions in the absence of monoubiquitination. FANCD2's ubiquitination-independent function is likely involved in optimized recruitment of nucleolytic activities for the processing and protection of stressed replication forks. Our results reveal that FANCD2 has a ubiquitination-independent role in countering endogenous levels of replication stress, a function that is critical for the maintenance of genomic stability. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

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. Copyright © 2015 Elsevier Inc. All rights reserved.

Dynamic stall - The case of the vertical axis wind turbine

Science.gov (United States)

Laneville, A.; Vittecoq, P.

1986-05-01

This paper presents the results of an experimental investigation on a driven Darrieus turbine rotating at different tip speed ratios. For a Reynolds number of 3.8 x 10 to the 4th, the results indicate the presence of dynamic stall at tip speed ratio less than 4, and that helicopter blade aerodynamics can be used in order to explain some aspects of the phenomenon. It was observed that in deep stall conditions, a vortex is formed at the leading edge; this vortex moves over the airfoil surface with 1/3 of the airfoil speed and then is shed at the trailing edge. After its shedding, the vortex can interact with the airfoil surface as the blade passes downstream.

DYNSTALL: Subroutine package with a dynamic stall model

Energy Technology Data Exchange (ETDEWEB)

Bjoerck, Anders [Aeronautical Research Inst. of Sweden, Bromma (Sweden)

2001-03-01

A subroutine package, called DYNSTALL, for the calculation of 2D unsteady airfoil aerodynamics is described. The subroutines are written in FORTRAN. DYNSTALL is basically an implementation of the Beddoes-Leishman dynamic stall model. This model is a semi-empirical model for dynamic stall. It includes, however, also models for attached flow unsteady aerodynamics. It is complete in the sense that it treats attached flow as well as separated flow. Semi-empirical means that the model relies on empirically determined constants. Semi because the constants are constants in equations with some physical interpretation. It requires the input of 2D airfoil aerodynamic data via tables as function of angle of attack. The method is intended for use in an aeroelastic code with the aerodynamics solved by blade/element method. DYNSTALL was written to work for any 2D angles of attack relative to the airfoil, e.g. flow from the rear of an airfoil.

Effect of summer grazing on welfare of dairy cows reared in mountain tie-stall barns

Directory of Open Access Journals (Sweden)

Simonetta Dovier

2010-09-01

Full Text Available Traditional mountain farms have an important economic, social and environmental role. The Alps management system for dairy cows consists of animals kept indoors from autumn to spring, mostly in tie-stalls, and moved to mountain pasture in summer. The aim of our study was to assess the effect of mountain summer grazing on the welfare of dairy cows housed in tie-stall barns. Twenty-four farms were considered. In twelve of them, animals were reared in tie-stalls and moved to mountain pasture for three months in summer; they were visited three times: (i four weeks before grazing during the indoor period in the stall; (ii about three weeks after the start of grazing; and (iii in the stall, in autumn, at least three weeks after returning from grazing. The other twelve farms kept the animals in tie-stalls all year; they were visited once in autumn. Data were collected following a protocol that considers animal-based measures and structure information on the basis of Quality Welfare Consortium® indications. Data allowed the calculation of both the Animal Needs Index score (ANI 35L and an overall assessment of the cows’ welfare obtained from three general aspects: housing, animal’s physical condition, and animal’s behaviour. Summer grazing had a significant positive effect on injuries, lameness and animal’s rising duration but a negative effect on faeces consistency. Moreover, a reduction of tongue playing was observed. The ANI 35L and the overall assessment did not show significant differences linked to summer grazing, which tended to have a positive but temporary effect on animal behaviour.

Dynamic Stall Vortex Formation of OA-209 Airfoil at Low Reynolds Number

OpenAIRE

Aung Myo Thu; Sang Eon Jeon; Yung Hwan Byun; Soo Hyung Park

2014-01-01

The unsteady flow field around oscillating OA-209 airfoil at a Reynolds number of 3.5×105 were investigated. Three different reduced frequencies were tested in order to see how it affects the hysteresis loop of an airfoil. At a reduced frequency of 0.05 the deep dynamic stall phenomenon was observed. Lift overshooting was observed as a result of dynamic stall vortex (DSV) shedding. Further investigation was carried out to find out the cause of DSV formation and shedding over airfoil. Particle...

Mechanisms and regulation of DNA replication initiation in eukaryotes.

Science.gov (United States)

Parker, Matthew W; Botchan, Michael R; Berger, James M

2017-04-01

Cellular DNA replication is initiated through the action of multiprotein complexes that recognize replication start sites in the chromosome (termed origins) and facilitate duplex DNA melting within these regions. In a typical cell cycle, initiation occurs only once per origin and each round of replication is tightly coupled to cell division. To avoid aberrant origin firing and re-replication, eukaryotes tightly regulate two events in the initiation process: loading of the replicative helicase, MCM2-7, onto chromatin by the origin recognition complex (ORC), and subsequent activation of the helicase by its incorporation into a complex known as the CMG. Recent work has begun to reveal the details of an orchestrated and sequential exchange of initiation factors on DNA that give rise to a replication-competent complex, the replisome. Here, we review the molecular mechanisms that underpin eukaryotic DNA replication initiation - from selecting replication start sites to replicative helicase loading and activation - and describe how these events are often distinctly regulated across different eukaryotic model organisms.

Flight Measurements of the Flying Qualities of a Lockheed P-80A Airplane (Army No. 44-85099) - Stalling Characteristics

Science.gov (United States)

Anderson, Seth B.; Cooper, George E.

1947-01-01

This report contains the flight-test results of the stalling characteristics measured during the flying-qualities investigation of the Lockheed P-8OA airplane (Army No. 44-85099). The tests were conducted in straight and turning flight with and without wing-tip tanks. These tests showed satisfactory stalling characteristics and adequate stall warning for all configurations and conditions tested.

Diagnosis of voltage collapse due to induction motor stalling using static analysis

International Nuclear Information System (INIS)

Karbalaei, F.; Kalantar, M.; Kazemi, A.

2008-01-01

Induction motor stalling is one of the important reasons for voltage collapse. This paper presents that, for induction motor stalling diagnosis, it is not necessary to use a third or first order dynamic model of induction motors. Instead, a method is presented based on algebraic calculations for which the steady state model of the induction motor considering different kinds of mechanical loads (constant and variable torque) is added to the power flow equations. Simulation results for a simple system confirm the correctness of the proposed method as compared to dynamic simulation results

Piloted Simulator Evaluation Results of Flight Physics Based Stall Recovery Guidance

Science.gov (United States)

Lombaerts, Thomas; Schuet, Stefan; Stepanyan, Vahram; Kaneshige, John; Hardy, Gordon; Shish, Kimberlee; Robinson, Peter

2018-01-01

In recent studies, it has been observed that loss of control in flight is the most frequent primary cause of accidents. A significant share of accidents in this category can be remedied by upset prevention if possible, and by upset recovery if necessary, in this order of priorities. One of the most important upsets to be recovered from is stall. Recent accidents have shown that a correct stall recovery maneuver remains a big challenge in civil aviation, partly due to a lack of pilot training. A possible strategy to support the flight crew in this demanding context is calculating a recovery guidance signal, and showing this signal in an intuitive way on one of the cockpit displays, for example by means of the flight director. Different methods for calculating the recovery signal, one based on fast model predictive control and another using an energy based approach, have been evaluated in four relevant operational scenarios by experienced commercial as well as test pilots in the Vertical Motion Simulator at NASA Ames Research Center. Evaluation results show that this approach could be able to assist the pilots in executing a correct stall recovery maneuver.

Plasticity of DNA replication initiation in Epstein-Barr virus episomes.

Directory of Open Access Journals (Sweden)

Paolo Norio

2004-06-01

Full Text Available In mammalian cells, the activity of the sites of initiation of DNA replication appears to be influenced epigenetically, but this regulation is not fully understood. Most studies of DNA replication have focused on the activity of individual initiation sites, making it difficult to evaluate the impact of changes in initiation activity on the replication of entire genomic loci. Here, we used single molecule analysis of replicated DNA (SMARD to study the latent duplication of Epstein-Barr virus (EBV episomes in human cell lines. We found that initiation sites are present throughout the EBV genome and that their utilization is not conserved in different EBV strains. In addition, SMARD shows that modifications in the utilization of multiple initiation sites occur across large genomic regions (tens of kilobases in size. These observations indicate that individual initiation sites play a limited role in determining the replication dynamics of the EBV genome. Long-range mechanisms and the genomic context appear to play much more important roles, affecting the frequency of utilization and the order of activation of multiple initiation sites. Finally, these results confirm that initiation sites are extremely redundant elements of the EBV genome. We propose that these conclusions also apply to mammalian chromosomes.

Replication dynamics of the yeast genome.

Science.gov (United States)

Raghuraman, M K; Winzeler, E A; Collingwood, D; Hunt, S; Wodicka, L; Conway, A; Lockhart, D J; Davis, R W; Brewer, B J; Fangman, W L

2001-10-05

Oligonucleotide microarrays were used to map the detailed topography of chromosome replication in the budding yeast Saccharomyces cerevisiae. The times of replication of thousands of sites across the genome were determined by hybridizing replicated and unreplicated DNAs, isolated at different times in S phase, to the microarrays. Origin activations take place continuously throughout S phase but with most firings near mid-S phase. Rates of replication fork movement vary greatly from region to region in the genome. The two ends of each of the 16 chromosomes are highly correlated in their times of replication. This microarray approach is readily applicable to other organisms, including humans.

An experimental description of the flow in a centrifugal compressor from alternate stall to surge

Science.gov (United States)

Moënne-Loccoz, V.; Trébinjac, I.; Benichou, E.; Goguey, S.; Paoletti, B.; Laucher, P.

2017-08-01

The present paper gives the experimental results obtained in a centrifugal compressor stage designed and built by SAFRAN Helicopter Engines. The compressor is composed of inlet guide vanes, a backswept splittered unshrouded impeller, a splittered vaned radial diffuser and axial outlet guide vanes. Previous numerical simulations revealed a particular S-shape pressure rise characteristic at partial rotation speed and predicted an alternate flow pattern in the vaned radial diffuser at low mass flow rate. This alternate flow pattern involves two adjacent vane passages. One passage exhibits very low momentum and a low pressure recovery, whereas the adjacent passage has very high momentum in the passage inlet and diffuses efficiently. Experimental measurements confirm the S-shape of the pressure rise characteristic even if the stability limit experimentally occurs at higher mass flow than numerically predicted. At low mass flow the alternate stall pattern is confirmed thanks to the data obtained by high-frequency pressure sensors. As the compressor is throttled the path to instability has been registered and a first scenario of the surge inception is given. The compressor first experiences a steady alternate stall in the diffuser. As the mass flow decreases, the alternate stall amplifies and triggers the mild surge in the vaned diffuser. An unsteady behavior results from the interaction of the alternate stall and the mild surge. Finally, when the pressure gradient becomes too strong, the alternate stall blows away and the compressor enters into deep surge.

The FFA dynamic stall model. The Beddoes-Leishman dynamic stall model modified for lead-lag oscillations

Energy Technology Data Exchange (ETDEWEB)

Bjoerck, A. [FFA, The Aeronautical Research Institute of Sweden, Bromma (Sweden)

1997-08-01

For calculations of the dynamics of wind turbines the inclusion of a dynamic stall model is necessary in order to obtain reliable results at high winds. For blade vibrations in the lead-lag motion the velocity relative to the blade will vary in time. In the present paper modifications to the Beddoes-Leishman model is presented in order to improve the model for calculations of cases with a varying relative velocity. Comparisons with measurement are also shown and the influence on the calculated aerodynamic damping by the modifications are investigated. (au)

A Dynamic Stall Model for Airfoils with Deformable Trailing Edges

International Nuclear Information System (INIS)

Andersen, Peter Bjoern; Gaunaa, Mac; Bak, Christian; Hansen, Morten Hartvig

2007-01-01

The present work contains an extension of the Beddoes-Leishman (B-L) type dynamic stall model, as described by Hansen et al. In this work a Deformable Trailing Edge Geometry (DTEG) has been added to the dynamic stall model. The model predicts the unsteady aerodynamic forces and moments on an airfoil section undergoing arbitrary motion in heave, lead-lag, pitch, Trailing Edge (TE) flapping. In the linear region, the model reduces to the inviscid model of Gaunaa, which includes the aerodynamic effect of a thin airfoil with a deformable camberline in inviscid flow. Therefore, the proposed model can be considered a crossover between the work of Gaunaa for the attached flow region and Hansen et al. The model will be compared to wind tunnel measurements from Velux described by Bak et al

Monitoring of the spatial and temporal dynamics of BER/SSBR pathway proteins, including MYH, UNG2, MPG, NTH1 and NEIL1-3, during DNA replication.

Science.gov (United States)

Bj Rås, Karine Ø; Sousa, Mirta M L; Sharma, Animesh; Fonseca, Davi M; S Gaard, Caroline K; Bj Rås, Magnar; Otterlei, Marit

2017-08-21

Base lesions in DNA can stall the replication machinery or induce mutations if bypassed. Consequently, lesions must be repaired before replication or in a post-replicative process to maintain genomic stability. Base excision repair (BER) is the main pathway for repair of base lesions and is known to be associated with DNA replication, but how BER is organized during replication is unclear. Here we coupled the iPOND (isolation of proteins on nascent DNA) technique with targeted mass-spectrometry analysis, which enabled us to detect all proteins required for BER on nascent DNA and to monitor their spatiotemporal orchestration at replication forks. We demonstrate that XRCC1 and other BER/single-strand break repair (SSBR) proteins are enriched in replisomes in unstressed cells, supporting a cellular capacity of post-replicative BER/SSBR. Importantly, we identify for the first time the DNA glycosylases MYH, UNG2, MPG, NTH1, NEIL1, 2 and 3 on nascent DNA. Our findings suggest that a broad spectrum of DNA base lesions are recognized and repaired by BER in a post-replicative process. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Intrinsically bent DNA in replication origins and gene promoters.

Science.gov (United States)

Gimenes, F; Takeda, K I; Fiorini, A; Gouveia, F S; Fernandez, M A

2008-06-24

Intrinsically bent DNA is an alternative conformation of the DNA molecule caused by the presence of dA/dT tracts, 2 to 6 bp long, in a helical turn phase DNA or with multiple intervals of 10 to 11 bp. Other than flexibility, intrinsic bending sites induce DNA curvature in particular chromosome regions such as replication origins and promoters. Intrinsically bent DNA sites are important in initiating DNA replication, and are sometimes found near to regions associated with the nuclear matrix. Many methods have been developed to localize bent sites, for example, circular permutation, computational analysis, and atomic force microscopy. This review discusses intrinsically bent DNA sites associated with replication origins and gene promoter regions in prokaryote and eukaryote cells. We also describe methods for identifying bent DNA sites for circular permutation and computational analysis.

An insight into the separate flow and stall delay for HAWT

Energy Technology Data Exchange (ETDEWEB)

Yu, Guohua; Shen, Xin; Zhu, Xiaocheng; Du, Zhaohui [School of Mechanical Engineering, Shanghai Jiao Tong University, Shanghai 200240 (China)

2011-01-15

The flow characteristics and the stall delay phenomenon of wind turbine rotor due to blade rotation in the steady state non-yawed conditions are investigated. An incompressible Reynolds-averaged Navier-Stokes solver is applied to carry out all the cases at different wind speeds from 5 m/s to 10 m/s with an interval of 1 m/s. CFD results turn out to agree well with experimental ones at incoming wind speeds below 10 m/s, though at 10 m/s some deviations exist due to the relative large flow separation and 3D spanwise flow over the suction surface of the blade. In the meanwhile, a lifting surface code with and without Du-Selig stall delay model is used to predict the power. A MATLAB code is developed to extract aerodynamic force coefficients from 3D CFD computations which are compared with the 2D airfoil wind tunnel experiment to demonstrate the stall delay and augmented lift phenomenon particularly at inboard span locations of the blade. The computational results are compared with the corrected value by the Du-Selig model and a lifting surface method derived data based on the measurements of the Unsteady Aerodynamic Experiment at the NASA Ames wind tunnel. (author)

Effect of stall design on dairy calf transition to voluntary feeding on an automatic milk feeder after introduction to group housing.

Science.gov (United States)

Wilson, Tanya R; LeBlanc, Stephen J; DeVries, Trevor J; Haley, Derek B

2018-06-01

Automatic milk feeders (AMF) for young dairy calves are widely used in the dairy industry. These feeders are thought to have benefits for calf health and welfare and may reduce labor required for feeding; however, little is known about how calves adapt to feeding with AMF. The objective of this study was to observe the effects of feeding stall design on calves learning to use the AMF. The hypothesis was that solid side stalls, compared with steel bar stalls, would result in a longer latency to approach and feed from the AMF without assistance. A total of 147 Holstein calves (80 male and 67 female) were enrolled at 4 d of age, introduced to a group pen, and, at the same time, trained on an AMF. For training, calves were allowed to suck on the trainer's fingers and guided to the teat. Calves were allocated to 1 of 2 stall designs at the pen level, depending on which treatment cohort they were born into, either with steel bar stall walls (n = 46 male, 34 female calves) or with solid side stall walls (n = 34 male, 33 female calves). For 72 h after introductory training on the AMF, data from the feeders were collected and calf behavior was monitored by video. Outcomes measured included latency to first voluntary visit to the feeder and to first feeding, time spent in the feeder, amount of milk consumed over 72 h, number of retraining sessions required (retrained if linear regression models or a Poisson model for the outcome of retraining. For certain outcomes the effects of stall design interacted with difficulty of training (willingness to enter feeder and drink); for the 38% of calves that were scored as moderately difficult to train on a scale of easy, moderate, or difficult, treatment (stall design) differences were detected. These calves took 2× longer to lick or bite toward the nipple, 2× longer to first voluntarily feeding, and consumed less milk over 72 h following training when trained on the steel bar stall design. These results suggest simple features of a

Replication stress-induced chromosome breakage is correlated with replication fork progression and is preceded by single-stranded DNA formation.

Science.gov (United States)

Feng, Wenyi; Di Rienzi, Sara C; Raghuraman, M K; Brewer, Bonita J

2011-10-01

Chromosome breakage as a result of replication stress has been hypothesized to be the direct consequence of defective replication fork progression, or "collapsed" replication forks. However, direct and genome-wide evidence that collapsed replication forks give rise to chromosome breakage is still lacking. Previously we showed that a yeast replication checkpoint mutant mec1-1, after transient exposure to replication impediment imposed by hydroxyurea (HU), failed to complete DNA replication, accumulated single-stranded DNA (ssDNA) at the replication forks, and fragmented its chromosomes. In this study, by following replication fork progression genome-wide via ssDNA detection and by direct mapping of chromosome breakage after HU exposure, we have tested the hypothesis that the chromosome breakage in mec1 cells occurs at collapsed replication forks. We demonstrate that sites of chromosome breakage indeed correlate with replication fork locations. Moreover, ssDNA can be detected prior to chromosome breakage, suggesting that ssDNA accumulation is the common precursor to double strand breaks at collapsed replication forks.

Effect of free stall surface on daily activity patterns in dairy cows with relevance to lameness prevalence.

Science.gov (United States)

Cook, N B; Bennett, T B; Nordlund, K V

2004-09-01

Differences in behavior of nonlame cows, slightly lame cows, and moderately lame cows in 6 free stall barns with sand bedding (SAND) vs. 6 free stall barns with rubber-crumb geotextile mattress surfaces (MAT) were documented in Wisconsin dairy herds. All lactating cows in the 12 herds were observed and given a locomotion score based on a 4-point scale: 1 = nonlame, 2 = slightly lame, 3 = moderately lame, and 4 = severely lame. Herd least square means +/-SE for prevalence of clinical lameness (locomotion scores = 3 and 4) were 11.1 vs. 24.0 +/- 1.7% for herds using SAND vs. MAT surfaces, respectively. Subsets of 10 cows per herd with locomotion scores of 1 to 3 were observed via video cameras for 24-h periods. Cows in MAT herds spent more time standing in free stalls per day than cows in SAND herds. Differences in standing times were 0.73 h/d for cows that were not lame, 2.32 h/d for cows that were slightly lame, and 4.31 h/d for cows that were moderately lame in MAT herds compared with equivalent cows in SAND herds. In MAT herds, the increase in time spent standing in the stall in moderately lame cows was associated with a significant reduction in stall use sessions per day, which impacted daily lying time. Although cause and effect are not clear, these findings have implications for housing, comfort, and care of cows in dairy herds with different types of free stall surfaces.

Prediction of rotating stall within an impeller of a centrifugal pump based on spectral analysis of pressure and velocity data

International Nuclear Information System (INIS)

Ullum, Ulrik; Wright, Jennifer; Dayi, Oguz; Ecder, Ali; Soulaimani, Azzeddine; Piche, Robert; Kamath, Hemant

2006-01-01

Experimental data, which was acquired in two centrifugal pumps and provided by Grundfos A/S, were analysed to determine if rotating stall could be detected from the velocity and pressure time series. The pressure data, which were uniformly acquired in time at high sample rates(10 kHz), were measured simultaneously in four adjacent di.user channels just downstream of the impeller outlet. The velocity data, which were non-uniformly sampled in time at fairly low rates(100 Hz to 3.5 kHz), were acquired either in or downstream of the impeller. Two di.erent methodologies were employed for detection of stall. The first method, which involved direct analysis of raw data, yielded qualitatively useful flow reversal information from the time series for the radial velocity. The second approach, which was based on power spectrum analysis of velocity and pressure data, could detect the onset and identify the frequency of rotating stall to a satisfactory extent in one of the two pumps. Nearly identical stall frequencies were observed in both velocity and pressure power spectra and this rotating stall phenomenon, which occurred at a very low frequency relative to the impeller speed, did not reveal any noticeable degree of sensitivity to the flow rate. In the other pump, where the available data was limited to velocity time series, the power spectrum analysis was successful in detecting stationary stall for a 6 bladed impeller but did not provide conclusive results for the existence of stall in the case of the 7 bladed impeller. Recommendations on the type of experimental data required for accurate detection of stall are provided based upon the present study

Airborne Microorganisms in Tie-stall Dairy Barns from Brasov County

Directory of Open Access Journals (Sweden)

Silvana Popescu

2011-05-01

Full Text Available The aim of this study was the assessment of the airborne microorganisms in tie-stall dairy cattle barns, through determination of the total number of bacteria (mesophilic bacteria, staphylococci, streptococci and gram-negative bacteria and fungi. We investigated 8 dairy cattle barns with tie-stalls in Brasov County during the winter of 2009. The mean numbers of bacteria and fungi in the morning and in the evening were: 1.02 °— 105 - 1.26 °— 105 CFU/m3 for mesophilic bacteria, 5.34 °— 104 - 5.91 °— 104 CFU/m3 for staphylococci, 2.93 °— 104 - 3.60 °— 104 CFU/m3 for streptococci, 2.17 °— 103 - 3.48 °— 103 CFU/m3 for gram-negative bacteria and 1.54 °— 104 - 2.75 °— 104 CFU/m3 for fungi. In the investigated cattle houses staphylococci represented 52.35 – 46.90%, streptococci were 28.73 – 28.57%, and the gram-negative bacteria were 2.13 – 2.76% within the overall number of mesophilic bacteria. The numbers of bacteria and fungi were slightly elevated in the evening comparative to the morning, but the differences were statistically insignificant (p>0.05. The great numbers of bacteria and fungi in the air of dairy cattle tie-stall barns indicate an elevated risk of disease for animals and human workers.

A dynamic stall model for airfoils with deformable trailing edges

DEFF Research Database (Denmark)

Andersen, Peter Bjørn; Gaunaa, Mac; Bak, Christian

2009-01-01

, lead-lag, pitch, trailing-edge flapping. In the linear region, the model reduces to the inviscid model, which includes the aerodynamic effect of a thin airfoil with a deformable camberline in inviscid flow. Therefore, the proposed model can be considered a crossover between the work of Gaunaa......The present work contains an extension of the Beddoes-Leishman-type dynamic stall model. In this work, a deformable trailing-edge flap has been added to the dynamic stall model. The model predicts the unsteady aerodynamic forces and moments on an airfoil section undergoing arbitrary motion in heave...... for the attached flow region and Hansen et al. The model is compared qualitatively to wind tunnel measurements of a Riso/ B1-18 blade section equipped with deformable trailing-edge flap devices in the form of piezoelectric devices. Copyright © 2009 John Wiley & Sons, Ltd....

Simulating Dynamic Stall Effects for Vertical Axis Wind Turbines Applying a Double Multiple Streamtube Model

Directory of Open Access Journals (Sweden)

Eduard Dyachuk

2015-02-01

Full Text Available The complex unsteady aerodynamics of vertical axis wind turbines (VAWT poses significant challenges to the simulation tools. Dynamic stall is one of the phenomena associated with the unsteady conditions for VAWTs, and it is in the focus of the study. Two dynamic stall models are compared: the widely-used Gormont model and a Leishman–Beddoes-type model. The models are included in a double multiple streamtube model. The effects of flow curvature and flow expansion are also considered. The model results are assessed against the measured data on a Darrieus turbine with curved blades. To study the dynamic stall effects, the comparison of force coefficients between the simulations and experiments is done at low tip speed ratios. Simulations show that the Leishman–Beddoes model outperforms the Gormont model for all tested conditions.

Strategic role of the ubiquitin-dependent segregase p97 (VCP or Cdc48) in DNA replication.

Science.gov (United States)

Ramadan, Kristijan; Halder, Swagata; Wiseman, Katherine; Vaz, Bruno

2017-02-01

Genome amplification (DNA synthesis) is one of the most demanding cellular processes in all proliferative cells. The DNA replication machinery (also known as the replisome) orchestrates genome amplification during S-phase of the cell cycle. Genetic material is particularly vulnerable to various events that can challenge the replisome during its assembly, activation (firing), progression (elongation) and disassembly from chromatin (termination). Any disturbance of the replisome leads to stalling of the DNA replication fork and firing of dormant replication origins, a process known as DNA replication stress. DNA replication stress is considered to be one of the main causes of sporadic cancers and other pathologies related to tissue degeneration and ageing. The mechanisms of replisome assembly and elongation during DNA synthesis are well understood. However, once DNA synthesis is complete, the process of replisome disassembly, and its removal from chromatin, remains unclear. In recent years, a growing body of evidence has alluded to a central role in replisome regulation for the ubiquitin-dependent protein segregase p97, also known as valosin-containing protein (VCP) in metazoans and Cdc48 in lower eukaryotes. By orchestrating the spatiotemporal turnover of the replisome, p97 plays an essential role in DNA replication. In this review, we will summarise our current knowledge about how p97 controls the replisome from replication initiation, to elongation and finally termination. We will also further examine the more recent findings concerning the role of p97 and how mutations in p97 cofactors, also known as adaptors, cause DNA replication stress induced genomic instability that leads to cancer and accelerated ageing. To our knowledge, this is the first comprehensive review concerning the mechanisms involved in the regulation of DNA replication by p97.

Compressible dynamic stall vorticity flux control using a dynamic ...

Indian Academy of Sciences (India)

systems, such as a wind turbine, are prevented from ever entering dynamic stall, essentially disregarding potential ... future generations of such systems, an overwhelming need has developed to avail this benefit safely. ... approach must diffuse the vorticity prior to its coalescence, but keep the vorticity over the airfoil up to ...

Identification of a divalent metal cation binding site in herpes simplex virus 1 (HSV-1) ICP8 required for HSV replication.

Science.gov (United States)

Bryant, Kevin F; Yan, Zhipeng; Dreyfus, David H; Knipe, David M

2012-06-01

Herpes simplex virus 1 (HSV-1) ICP8 is a single-stranded DNA-binding protein that is necessary for viral DNA replication and exhibits recombinase activity in vitro. Alignment of the HSV-1 ICP8 amino acid sequence with ICP8 homologs from other herpesviruses revealed conserved aspartic acid (D) and glutamic acid (E) residues. Amino acid residue D1087 was conserved in every ICP8 homolog analyzed, indicating that it is likely critical for ICP8 function. We took a genetic approach to investigate the functions of the conserved ICP8 D and E residues in HSV-1 replication. The E1086A D1087A mutant form of ICP8 failed to support the replication of an ICP8 mutant virus in a complementation assay. E1086A D1087A mutant ICP8 bound DNA, albeit with reduced affinity, demonstrating that the protein is not globally misfolded. This mutant form of ICP8 was also recognized by a conformation-specific antibody, further indicating that its overall structure was intact. A recombinant virus expressing E1086A D1087A mutant ICP8 was defective in viral replication, viral DNA synthesis, and late gene expression in Vero cells. A class of enzymes called DDE recombinases utilize conserved D and E residues to coordinate divalent metal cations in their active sites. We investigated whether the conserved D and E residues in ICP8 were also required for binding metal cations and found that the E1086A D1087A mutant form of ICP8 exhibited altered divalent metal binding in an in vitro iron-induced cleavage assay. These results identify a novel divalent metal cation-binding site in ICP8 that is required for ICP8 functions during viral replication.

Band-pass filtering algorithms for adaptive control of compressor pre-stall modes in aircraft gas-turbine engine

Science.gov (United States)

Kuznetsova, T. A.

2018-05-01

The methods for increasing gas-turbine aircraft engines' (GTE) adaptive properties to interference based on empowerment of automatic control systems (ACS) are analyzed. The flow pulsation in suction and a discharge line of the compressor, which may cause the stall, are considered as the interference. The algorithmic solution to the problem of GTE pre-stall modes’ control adapted to stability boundary is proposed. The aim of the study is to develop the band-pass filtering algorithms to provide the detection functions of the compressor pre-stall modes for ACS GTE. The characteristic feature of pre-stall effect is the increase of pressure pulsation amplitude over the impeller at the multiples of the rotor’ frequencies. The used method is based on a band-pass filter combining low-pass and high-pass digital filters. The impulse response of the high-pass filter is determined through a known low-pass filter impulse response by spectral inversion. The resulting transfer function of the second order band-pass filter (BPF) corresponds to a stable system. The two circuit implementations of BPF are synthesized. Designed band-pass filtering algorithms were tested in MATLAB environment. Comparative analysis of amplitude-frequency response of proposed implementation allows choosing the BPF scheme providing the best quality of filtration. The BPF reaction to the periodic sinusoidal signal, simulating the experimentally obtained pressure pulsation function in the pre-stall mode, was considered. The results of model experiment demonstrated the effectiveness of applying band-pass filtering algorithms as part of ACS to identify the pre-stall mode of the compressor for detection of pressure fluctuations’ peaks, characterizing the compressor’s approach to the stability boundary.

Inhibition of Lassa virus glycoprotein cleavage and multicycle replication by site 1 protease-adapted alpha(1-antitrypsin variants.

Directory of Open Access Journals (Sweden)

Anna Maisa

2009-06-01

Full Text Available Proteolytic processing of the Lassa virus envelope glycoprotein precursor GP-C by the host proprotein convertase site 1 protease (S1P is a prerequisite for the incorporation of the subunits GP-1 and GP-2 into viral particles and, hence, essential for infectivity and virus spread. Therefore, we tested in this study the concept of using S1P as a target to block efficient virus replication.We demonstrate that stable cell lines inducibly expressing S1P-adapted alpha(1-antitrypsin variants inhibit the proteolytic maturation of GP-C. Introduction of the S1P recognition motifs RRIL and RRLL into the reactive center loop of alpha(1-antitrypsin resulted in abrogation of GP-C processing by endogenous S1P to a similar level observed in S1P-deficient cells. Moreover, S1P-specific alpha(1-antitrypsins significantly inhibited replication and spread of a replication-competent recombinant vesicular stomatitis virus expressing the Lassa virus glycoprotein GP as well as authentic Lassa virus. Inhibition of viral replication correlated with the ability of the different alpha(1-antitrypsin variants to inhibit the processing of the Lassa virus glycoprotein precursor.Our data suggest that glycoprotein cleavage by S1P is a promising target for the development of novel anti-arenaviral strategies.

Rad51-Rad52 mediated maintenance of centromeric chromatin in Candida albicans.

Directory of Open Access Journals (Sweden)

Sreyoshi Mitra

2014-04-01

epigenetically maintain centromere functioning by regulating CENP-ACaCse4 levels at the programmed stall sites of early replicating centromeres.

Accounting for biases in riboprofiling data indicates a major role for proline in stalling translation.

Science.gov (United States)

Artieri, Carlo G; Fraser, Hunter B

2014-12-01

The recent advent of ribosome profiling-sequencing of short ribosome-bound fragments of mRNA-has offered an unprecedented opportunity to interrogate the sequence features responsible for modulating translational rates. Nevertheless, numerous analyses of the first riboprofiling data set have produced equivocal and often incompatible results. Here we analyze three independent yeast riboprofiling data sets, including two with much higher coverage than previously available, and find that all three show substantial technical sequence biases that confound interpretations of ribosomal occupancy. After accounting for these biases, we find no effect of previously implicated factors on ribosomal pausing. Rather, we find that incorporation of proline, whose unique side-chain stalls peptide synthesis in vitro, also slows the ribosome in vivo. We also reanalyze a method that implicated positively charged amino acids as the major determinant of ribosomal stalling and demonstrate that it produces false signals of stalling in low-coverage data. Our results suggest that any analysis of riboprofiling data should account for sequencing biases and sparse coverage. To this end, we establish a robust methodology that enables analysis of ribosome profiling data without prior assumptions regarding which positions spanned by the ribosome cause stalling. © 2014 Artieri and Fraser; Published by Cold Spring Harbor Laboratory Press.

Performance augmentation with vortex generators: Design and testing for stall-regulated AWT-26 turbine

Energy Technology Data Exchange (ETDEWEB)

Griffin, D.A. [Advanced Wind Turbines Inc., Seattle, WA (United States)

1996-12-31

A study investigated the use of vortex generators (VGs) for performance augmentation of the stall-regulated AWT-26 wind turbine. Based on wind-tunnel results and analysis, a VG array was designed for and tested on the AWT-26 prototype, designated Pt. Performance and loads data were measured for P1, both with and without VGs installed. The turbine performance with VGs met most of the design requirements; power output was increased at moderate wind speeds with a minimal effect on peak power. However, VG drag penalties caused a loss in power output for low wind speeds, such that performance with VGs resulted in a net decrease in AEP for wind speed sites up to 8.5 m/s. 8 refs., 8 figs., 3 tabs.

The influence of elevated feed stalls on feeding behaviour of lactating dairy cows

Directory of Open Access Journals (Sweden)

Barbara Benz

2014-10-01

Full Text Available The performance level of high yielding cows can only be guaranteed by high quality forage and high feed intake. An about 15–20 cm elevated and 160 cm long feed stall with rubber flooring doesn’t only offer undisturbed meals but also a yielding and dry standing surface. In a pilot stable with 130 dairy cows (German Simmental the feeding alley was subsequently equipped with elevated feed stalls. The results show that animals frequented the feeding barn less often while the duration of single meals prolonged. The specific behavioural changes differed depending on milk yield and number of lactation.

Hepatitis C Virus Replication Depends on Endosomal Cholesterol Homeostasis.

Science.gov (United States)

Stoeck, Ina Karen; Lee, Ji-Young; Tabata, Keisuke; Romero-Brey, Inés; Paul, David; Schult, Philipp; Lohmann, Volker; Kaderali, Lars; Bartenschlager, Ralf

2018-01-01

Similar to other positive-strand RNA viruses, hepatitis C virus (HCV) causes massive rearrangements of intracellular membranes, resulting in a membranous web (MW) composed of predominantly double-membrane vesicles (DMVs), the presumed sites of RNA replication. DMVs are enriched for cholesterol, but mechanistic details on the source and recruitment of cholesterol to the viral replication organelle are only partially known. Here we focused on selected lipid transfer proteins implicated in direct lipid transfer at various endoplasmic reticulum (ER)-membrane contact sites. RNA interference (RNAi)-mediated knockdown identified several hitherto unknown HCV dependency factors, such as steroidogenic acute regulatory protein-related lipid transfer domain protein 3 (STARD3), oxysterol-binding protein-related protein 1A and -B (OSBPL1A and -B), and Niemann-Pick-type C1 (NPC1), all residing at late endosome and lysosome membranes and required for efficient HCV RNA replication but not for replication of the closely related dengue virus. Focusing on NPC1, we found that knockdown or pharmacological inhibition caused cholesterol entrapment in lysosomal vesicles concomitant with decreased cholesterol abundance at sites containing the viral replicase factor NS5A. In untreated HCV-infected cells, unesterified cholesterol accumulated at the perinuclear region, partially colocalizing with NS5A at DMVs, arguing for NPC1-mediated endosomal cholesterol transport to the viral replication organelle. Consistent with cholesterol being an important structural component of DMVs, reducing NPC1-dependent endosomal cholesterol transport impaired MW integrity. This suggests that HCV usurps lipid transfer proteins, such as NPC1, at ER-late endosome/lysosome membrane contact sites to recruit cholesterol to the viral replication organelle, where it contributes to MW functionality. IMPORTANCE A key feature of the replication of positive-strand RNA viruses is the rearrangement of the host cell

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.

LHCb experience with LFC replication

International Nuclear Information System (INIS)

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

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

A kinesthetic-tactual display for stall deterrence

Science.gov (United States)

Gilson, R. D.; Ventola, R. W.; Fenton, R. E.

1975-01-01

A kinesthetic tactual display may be effectively used as a control aid per previous flight tests. Angle of attack information would be continuously presented to a pilot, via this display, during critical operational phases where stalls are probable. A two phase plan for evaluating this concept is presented. A first development phase would encompass: (1) display fabrication for a conventional control yoke; (2) its installation, together with other necessary instrumentation, in an experimental aircraft; and (3) preliminary flight testing by experienced pilots.

Design of advanced airfoil for stall-regulated wind turbines

Directory of Open Access Journals (Sweden)

F. Grasso

2017-07-01

Full Text Available Nowadays, all the modern megawatt-class wind turbines make use of pitch control to optimise the rotor performance and control the turbine. However, for kilowatt-range machines, stall-regulated solutions are still attractive and largely used for their simplicity and robustness. In the design phase, the aerodynamics plays a crucial role, especially concerning the selection/design of the necessary airfoils. This is because the airfoil performance is supposed to guarantee high wind turbine performance but also the necessary machine control capabilities. In the present work, the design of a new airfoil dedicated to stall machines is discussed. The design strategy makes use of a numerical optimisation scheme, where a gradient-based algorithm is coupled with the RFOIL code and an original Bezier-curves-based parameterisation to describe the airfoil shape. The performances of the new airfoil are compared in free- and fixed-transition conditions. In addition, the performance of the rotor is analysed, comparing the impact of the new geometry with alternative candidates. The results show that the new airfoil offers better performance and control than existing candidates do.

High-resolution LES of the rotating stall in a reduced scale model pump-turbine

International Nuclear Information System (INIS)

Pacot, Olivier; Avellan, François; Kato, Chisachi

2014-01-01

Extending the operating range of modern pump-turbines becomes increasingly important in the course of the integration of renewable energy sources in the existing power grid. However, at partial load condition in pumping mode, the occurrence of rotating stall is critical to the operational safety of the machine and on the grid stability. The understanding of the mechanisms behind this flow phenomenon yet remains vague and incomplete. Past numerical simulations using a RANS approach often led to inconclusive results concerning the physical background. For the first time, the rotating stall is investigated by performing a large scale LES calculation on the HYDRODYNA pump-turbine scale model featuring approximately 100 million elements. The computations were performed on the PRIMEHPC FX10 of the University of Tokyo using the overset Finite Element open source code FrontFlow/blue with the dynamic Smagorinsky turbulence model and the no-slip wall condition. The internal flow computed is the one when operating the pump-turbine at 76% of the best efficiency point in pumping mode, as previous experimental research showed the presence of four rotating cells. The rotating stall phenomenon is accurately reproduced for a reduced Reynolds number using the LES approach with acceptable computing resources. The results show an excellent agreement with available experimental data from the reduced scale model testing at the EPFL Laboratory for Hydraulic Machines. The number of stall cells as well as the propagation speed corroborates the experiment

High-resolution LES of the rotating stall in a reduced scale model pump-turbine

Science.gov (United States)

Pacot, Olivier; Kato, Chisachi; Avellan, François

2014-03-01

Extending the operating range of modern pump-turbines becomes increasingly important in the course of the integration of renewable energy sources in the existing power grid. However, at partial load condition in pumping mode, the occurrence of rotating stall is critical to the operational safety of the machine and on the grid stability. The understanding of the mechanisms behind this flow phenomenon yet remains vague and incomplete. Past numerical simulations using a RANS approach often led to inconclusive results concerning the physical background. For the first time, the rotating stall is investigated by performing a large scale LES calculation on the HYDRODYNA pump-turbine scale model featuring approximately 100 million elements. The computations were performed on the PRIMEHPC FX10 of the University of Tokyo using the overset Finite Element open source code FrontFlow/blue with the dynamic Smagorinsky turbulence model and the no-slip wall condition. The internal flow computed is the one when operating the pump-turbine at 76% of the best efficiency point in pumping mode, as previous experimental research showed the presence of four rotating cells. The rotating stall phenomenon is accurately reproduced for a reduced Reynolds number using the LES approach with acceptable computing resources. The results show an excellent agreement with available experimental data from the reduced scale model testing at the EPFL Laboratory for Hydraulic Machines. The number of stall cells as well as the propagation speed corroborates the experiment.

Modelling of multiple short-length-scale stall cells in an axial compressor using evolved GMDH neural networks

International Nuclear Information System (INIS)

Amanifard, N.; Nariman-Zadeh, N.; Farahani, M.H.; Khalkhali, A.

2008-01-01

Over the past 15 years there have been several research efforts to capture the stall inception nature in axial flow compressors. However previous analytical models could not explain the formation of short-length-scale stall cells. This paper provides a new model based on evolved GMDH neural network for transient evolution of multiple short-length-scale stall cells in an axial compressor. Genetic Algorithms (GAs) are also employed for optimal design of connectivity configuration of such GMDH-type neural networks. In this way, low-pass filter (LPF) pressure trace near the rotor leading edge is modelled with respect to the variation of pressure coefficient, flow rate coefficient, and number of rotor rotations which are defined as inputs

Towards observing the encounter of the T7 DNA replication fork with a lesion site at the Single molecule level

KAUST Repository

Shirbini, Afnan

2017-05-01

Single-molecule DNA flow-stretching assays have been a powerful approach to study various aspects on the mechanism of DNA replication for more than a decade. This technique depends on flow-induced force on a bead attached to a surface-tethered DNA. The difference in the elastic property between double-strand DNA (long) and single-strand DNA (short) at low regime force allows the observation of the beads motion when the dsDNA is converted to ssDNA by the replisome machinery during DNA replication. Here, I aim to develop an assay to track in real-time the encounter of the bacteriophage T7 replisome with abasic lesion site inserted on the leading strand template. I optimized methods to construct the DNA substrate that contains the abasic site and established the T7 leading strand synthesis at the single molecule level. I also optimized various control experiments to remove any interference from the nonspecific interactions of the DNA with the surface. My work established the foundation to image the encounter of the T7 replisome with abasic site and to characterize how the interactions between the helicase and the polymerase could influence the polymerase proofreading ability and its direct bypass of this highly common DNA damage type.

Dynamics and Control of Three-Dimensional Perching Maneuver under Dynamic Stall Influence

Science.gov (United States)

Feroskhan, Mir Alikhan Bin Mohammad

Perching is a type of aggressive maneuver performed by the class 'Aves' species to attain precision point landing with a generally short landing distance. Perching capability is desirable on unmanned aerial vehicles (UAVs) due to its efficient deceleration process that potentially expands the functionality and flight envelope of the aircraft. This dissertation extends the previous works on perching, which is mostly limited to two-dimensional (2D) cases, to its state-of-the-art threedimensional (3D) variety. This dissertation presents the aerodynamic modeling and optimization framework adopted to generate unprecedented variants of the 3D perching maneuver that include the sideslip perching trajectory, which ameliorates the existing 2D perching concept by eliminating the undesirable undershoot and reliance on gravity. The sideslip perching technique methodically utilizes the lateral and longitudinal drag mechanisms through consecutive phases of yawing and pitching-up motion. Since perching maneuver involves high rates of change in the angles of attack and large turn rates, introduction of three internal variables thus becomes necessary for addressing the influence of dynamic stall delay on the UAV's transient post-stall behavior. These variables are then integrated into a static nonlinear aerodynamic model, developed using empirical and analytical methods, and into an optimization framework that generates a trajectory of sideslip perching maneuver, acquiring over 70% velocity reduction. An impact study of the dynamic stall influence on the optimal perching trajectories suggests that consideration of dynamic stall delay is essential due to the significant discrepancies in the corresponding control inputs required. A comparative study between 2D and 3D perching is also conducted to examine the different drag mechanisms employed by 2D and 3D perching respectively. 3D perching is presented as a more efficient deceleration technique with respect to spatial costs and

A model for the selective amplification of spatially coherent waves in a centrifugal compressor on the verge of rotating stall

Science.gov (United States)

Lawless, Patrick B.; Fleeter, Sanford

1993-01-01

A simple model for the stability zones of a low speed centrifugal compressor is developed, with the goal of understanding the driving mechanism for the changes in stalling behavior predicted for, and observed in, the Purdue Low Speed Centrifugal Research Compressor Facility. To this end, earlier analyses of rotating stall suppression in centrifugal compressors are presented in a reduced form that preserves the essential parameters of the model that affect the stalling behavior of the compressor. The model is then used to illuminate the relationship between compressor geometry, expected mode shape, and regions of amplification for weak waves which are indicative of the susceptibility of the system to rotating stall. The results demonstrate that increasing the stagger angle of the diffuser vanes, and consequently the diffusion path length, results in the compressor moving towards a condition where higher-order spatial modes are excited during stall initiation. Similarly, flow acceleration in the diffuser section caused by an increase in the number of diffuser vanes also results in the excitation of higher modes.

Power reduction and the radial limit of stall delay in revolving wings of different aspect ratio.

Science.gov (United States)

Kruyt, Jan W; van Heijst, GertJan F; Altshuler, Douglas L; Lentink, David

2015-04-06

Airplanes and helicopters use high aspect ratio wings to reduce the power required to fly, but must operate at low angle of attack to prevent flow separation and stall. Animals capable of slow sustained flight, such as hummingbirds, have low aspect ratio wings and flap their wings at high angle of attack without stalling. Instead, they generate an attached vortex along the leading edge of the wing that elevates lift. Previous studies have demonstrated that this vortex and high lift can be reproduced by revolving the animal wing at the same angle of attack. How do flapping and revolving animal wings delay stall and reduce power? It has been hypothesized that stall delay derives from having a short radial distance between the shoulder joint and wing tip, measured in chord lengths. This non-dimensional measure of wing length represents the relative magnitude of inertial forces versus rotational accelerations operating in the boundary layer of revolving and flapping wings. Here we show for a suite of aspect ratios, which represent both animal and aircraft wings, that the attachment of the leading edge vortex on a revolving wing is determined by wing aspect ratio, defined with respect to the centre of revolution. At high angle of attack, the vortex remains attached when the local radius is shorter than four chord lengths and separates outboard on higher aspect ratio wings. This radial stall limit explains why revolving high aspect ratio wings (of helicopters) require less power compared with low aspect ratio wings (of hummingbirds) at low angle of attack and vice versa at high angle of attack. © 2015 The Author(s) Published by the Royal Society. All rights reserved.

Unsteady behavior of leading-edge vortex and diffuser stall in a centrifugal compressor with vaned diffuser

Science.gov (United States)

Fujisawa, Nobumichi; Hara, Shotaro; Ohta, Yutaka

2016-02-01

The characteristics of a rotating stall of an impeller and diffuser and the evolution of a vortex generated at the diffuser leading-edge (i.e., the leading-edge vortex (LEV)) in a centrifugal compressor were investigated by experiments and numerical analysis. The results of the experiments revealed that both the impeller and diffuser rotating stalls occurred at 55 and 25 Hz during off-design flow operation. For both, stall cells existed only on the shroud side of the flow passages, which is very close to the source location of the LEV. According to the CFD results, the LEV is made up of multiple vortices. The LEV is a combination of a separated vortex near the leading- edge and a longitudinal vortex generated by the extended tip-leakage flow from the impeller. Therefore, the LEV is generated by the accumulation of vorticity caused by the velocity gradient of the impeller discharge flow. In partial-flow operation, the spanwise extent and the position of the LEV origin are temporarily transmuted. The LEV develops with a drop in the velocity in the diffuser passage and forms a significant blockage within the diffuser passage. Therefore, the LEV may be regarded as being one of the causes of a diffuser stall in a centrifugal compressor.

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

Science.gov (United States)

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

2014-01-01

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

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

Directory of Open Access Journals (Sweden)

Gregory A Sowd

2014-12-01

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

Inhibition and recovery of the replication of depurinated parvovirus DNA in mouse fibroblasts

International Nuclear Information System (INIS)

Vos, J.M.; Avalosse, B.; Su, Z.Z.; Rommelaere, J.

1984-01-01

Apurinic sites were introduced in the single-stranded DNA of parvovirus minute-virus-of-mice (MVM) and their effect on viral DNA synthesis was measured in mouse fibroblasts. Approximately one apurinic site per viral genome, is sufficient to block its replication in untreated cells. The exposure of host cells to a sublethal dose of UV-light 15 hours prior to virus infection, enhances their ability to support the replication of depurinated MVM. Cell preirradiation induces the apparent overcome of 10-15% of viral DNA replication blocks. These results indicate that apurinic sites prevent mammalian cells from replicating single-stranded DNA unless a recovery process is activated by cell UV-irradiation

DNA replication origins—where do we begin?

Science.gov (United States)

Prioleau, Marie-Noëlle; MacAlpine, David M.

2016-01-01

For more than three decades, investigators have sought to identify the precise locations where DNA replication initiates in mammalian genomes. The development of molecular and biochemical approaches to identify start sites of DNA replication (origins) based on the presence of defining and characteristic replication intermediates at specific loci led to the identification of only a handful of mammalian replication origins. The limited number of identified origins prevented a comprehensive and exhaustive search for conserved genomic features that were capable of specifying origins of DNA replication. More recently, the adaptation of origin-mapping assays to genome-wide approaches has led to the identification of tens of thousands of replication origins throughout mammalian genomes, providing an unprecedented opportunity to identify both genetic and epigenetic features that define and regulate their distribution and utilization. Here we summarize recent advances in our understanding of how primary sequence, chromatin environment, and nuclear architecture contribute to the dynamic selection and activation of replication origins across diverse cell types and developmental stages. PMID:27542827

Replication termination and chromosome dimer resolution in the archaeon Sulfolobus solfataricus.

Science.gov (United States)

Duggin, Iain G; Dubarry, Nelly; Bell, Stephen D

2011-01-05

Archaea of the genus Sulfolobus have a single-circular chromosome with three replication origins. All three origins fire in every cell in every cell cycle. Thus, three pairs of replication forks converge and terminate in each replication cycle. Here, we report 2D gel analyses of the replication fork fusion zones located between origins. These indicate that replication termination involves stochastic fork collision. In bacteria, replication termination is linked to chromosome dimer resolution, a process that requires the XerC and D recombinases, FtsK and the chromosomal dif site. Sulfolobus encodes a single-Xer homologue and its deletion gave rise to cells with aberrant DNA contents and increased volumes. Identification of the chromosomal dif site that binds Xer in vivo, and biochemical characterization of Xer/dif recombination revealed that, in contrast to bacteria, dif is located outside the fork fusion zones. Therefore, it appears that replication termination and dimer resolution are temporally and spatially distinct processes in Sulfolobus.

Replication protein A and γ-H2AX foci assembly is triggered by cellular response to DNA double-strand breaks

International Nuclear Information System (INIS)

Balajee, Adayabalam S.; Geard, Charles R.

2004-01-01

Human replication protein A (RPA p34), a crucial component of diverse DNA excision repair pathways, is implicated in DNA double-strand break (DSB) repair. To evaluate its role in DSB repair, the intranuclear dynamics of RPA was investigated after DNA damage and replication blockage in human cells. Using two different agents [ionizing radiation (IR) and hydroxyurea (HU)] to generate DSBs, we found that RPA relocated into distinct nuclear foci and colocalized with a well-known DSB binding factor, γ-H2AX, at the sites of DNA damage in a time-dependent manner. Colocalization of RPA and γ-H2AX foci peaked at 2 h after IR treatment and subsequently declined with increasing postrecovery times. The time course of RPA and γ-H2AX foci association correlated well with the DSB repair activity detected by a neutral comet assay. A phosphatidylinositol-3 (PI-3) kinase inhibitor, wortmannin, completely abolished both RPA and γ-H2AX foci formation triggered by IR. Additionally, radiosensitive ataxia telangiectasia (AT) cells harboring mutations in ATM gene product were found to be deficient in RPA and γ-H2AX colocalization after IR. Transfection of AT cells with ATM cDNA fully restored the association of RPA foci with γ-H2AX illustrating the requirement of ATM gene product for this process. The exact coincidence of RPA and γ-H2AX in response to HU specifically in S-phase cells supports their role in DNA replication checkpoint control. Depletion of RPA by small interfering RNA (SiRNA) substantially elevated the frequencies of IR-induced micronuclei (MN) and apoptosis in human cells suggestive of a role for RPA in DSB repair. We propose that RPA in association with γ-H2AX contributes to both DNA damage checkpoint control and repair in response to strand breaks and stalled replication forks in human cells

Impaired TIP60-mediated H4K16 acetylation accounts for the aberrant chromatin accumulation of 53BP1 and RAP80 in Fanconi anemia pathway-deficient cells.

Science.gov (United States)

Renaud, Emilie; Barascu, Aurelia; Rosselli, Filippo

2016-01-29

To rescue collapsed replication forks cells utilize homologous recombination (HR)-mediated mechanisms to avoid the induction of gross chromosomal abnormalities that would be generated by non-homologous end joining (NHEJ). Using DNA interstrand crosslinks as a replication barrier, we investigated how the Fanconi anemia (FA) pathway promotes HR at stalled replication forks. FA pathway inactivation results in Fanconi anemia, which is associated with a predisposition to cancer. FANCD2 monoubiquitination and assembly in subnuclear foci appear to be involved in TIP60 relocalization to the chromatin to acetylates histone H4K16 and prevents the binding of 53BP1 to its docking site, H4K20Me2. Thus, FA pathway loss-of-function results in accumulation of 53BP1, RIF1 and RAP80 at damaged chromatin, which impair DNA resection at stalled replication fork-associated DNA breaks and impede HR. Consequently, DNA repair in FA cells proceeds through the NHEJ pathway, which is likely responsible for the accumulation of chromosome abnormalities. We demonstrate that the inhibition of NHEJ or deacetylase activity rescue HR in FA cells. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

A flight investigation of the ultra-deep-stall descent and spin recovery characteristics of a 1/6 scale radiocontrolled model of the Piper PA38 Tomahawk

Science.gov (United States)

Blanchard, W. S., Jr.

1981-01-01

Ultradeep stall descent and spin recovery characteristics of a 1/6 scale radio controlled model of the Piper PA38 Tomahawk aircraft was investigated. It was shown that the full scale PA38 is a suitable aircraft for conducting ultradeep stall research. Spin recovery was accomplished satisfactorily by entry to the ultradeep stall mode, followed by the exit from the ultradeep stall mode. It is concluded that since the PA38 has excellent spin recovery characteristics using normal recovery techniques (opposite rudder and forward control colum pressure), recovery using ultradeep stall would be beneficial only if the pilot suffered from disorientation.

Robust post-stall perching with a simple fixed-wing glider using LQR-Trees

International Nuclear Information System (INIS)

Moore, Joseph; Cory, Rick; Tedrake, Russ

2014-01-01

Birds routinely execute post-stall maneuvers with a speed and precision far beyond the capabilities of our best aircraft control systems. One remarkable example is a bird exploiting post-stall pressure drag in order to rapidly decelerate to land on a perch. Stall is typically associated with a loss of control authority, and it is tempting to attribute this agility of birds to the intricate morphology of the wings and tail, to their precision sensing apparatus, or their ability to perform thrust vectoring. Here we ask whether an extremely simple fixed-wing glider (no propeller) with only a single actuator in the tail is capable of landing precisely on a perch from a large range of initial conditions. To answer this question, we focus on the design of the flight control system; building upon previous work which used linear feedback control design based on quadratic regulators (LQR), we develop nonlinear feedback control based on nonlinear model-predictive control and ‘LQR-Trees’. Through simulation using a flat-plate model of the glider, we find that both nonlinear methods are capable of achieving an accurate bird-like perching maneuver from a large range of initial conditions; the ‘LQR-Trees’ algorithm is particularly useful due to its low computational burden at runtime and its inherent performance guarantees. With this in mind, we then implement the ‘LQR-Trees’ algorithm on real hardware and demonstrate a 95 percent perching success rate over 147 flights for a wide range of initial speeds. These results suggest that, at least in the absence of significant disturbances like wind gusts, complex wing morphology and sensing are not strictly required to achieve accurate and robust perching even in the post-stall flow regime. (papers)

Identification of phlebovirus and arenavirus RNA sequences that stall and repress the exoribonuclease XRN1.

Science.gov (United States)

Charley, Phillida A; Wilusz, Carol J; Wilusz, Jeffrey

2018-01-05

Regulated mRNA decay plays a vital role in determining both the level and quality of cellular gene expression. Viral RNAs must successfully evade this host RNA decay machinery to establish a productive infection. One way for RNA viruses to accomplish this is to target the cellular exoribonuclease XRN1, because this enzyme is accessible in the cytoplasm and plays a major role in mRNA decay. Members of the Flaviviridae use RNA structures in their 5'- or 3'-untranslated regions to stall and repress XRN1, effectively stabilizing viral RNAs while also causing significant dysregulation of host cell mRNA stability. Here, we use a series of biochemical assays to demonstrate that the 3'-terminal portion of the nucleocapsid (N) mRNA of Rift Valley fever virus, a phlebovirus of the Bunyaviridae family, also can effectively stall and repress XRN1. The region responsible for impeding XRN1 includes a G-rich portion that likely forms a G-quadruplex structure. The 3'-terminal portions of ambisense-derived transcripts of multiple arenaviruses also stalled XRN1. Therefore, we conclude that RNAs from two additional families of mammalian RNA viruses stall and repress XRN1. This observation. emphasizes the importance and commonality of this viral strategy to interfere with the 5'-to-3'-exoribonuclease component of the cytoplasmic RNA decay machinery. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Dynamic Stall Control on the Wind Turbine Airfoil via a Co-Flow Jet

Directory of Open Access Journals (Sweden)

He-Yong Xu

2016-06-01

Full Text Available Dynamic stall control of a S809 airfoil is numerically investigated by implementing a co-flow jet (CFJ. The numerical methods of the solver are validated by comparing results with the baseline experiment as well as a NACA 6415-based CFJ experiment, showing good agreement in both static and dynamic characteristics. The CFJ airfoil with inactive jet is simulated to study the impact that the jet channel imposes upon the dynamic characteristics. It is shown that the presence of a long jet channel could cause a negative effect of decreasing lift and increasing drag, leading to fluctuating extreme loads in terms of drag and moment. The main focus of the present research is the investigation of the dynamic characteristics of the CFJ airfoil with three different jet momentum coefficients, which are compared with the baseline, giving encouraging results. Dynamic stall can be greatly suppressed, showing a very good control performance of significantly increased lift and reduced drag and moment. Analysis of the amplitude of variation in the aerodynamic coefficients indicates that the fluctuating extreme aerodynamic loads are significantly alleviated, which is conducive to structural reliability and improved life cycle. The energy consumption analysis shows that the CFJ concept is applicable and economical in controlling dynamic stall.

Nonlinear Aeroelastic Study of Stall Induced Oscillation in a Symmetric Airfoil

NARCIS (Netherlands)

Sarkar, S.; Bijl, H.

2006-01-01

In this paper the aeroelastic stability of a wind turbine rotor in the dynamic stall regime is investigated. Increased flexibility of modern turbine blades makes them more susceptible to aeroelastic instabilities. Complex oscillation modes like flap/lead-lag are of particular concern, which give way

Pressure-based high-order TVD methodology for dynamic stall control

Science.gov (United States)

Yang, H. Q.; Przekwas, A. J.

1992-01-01

The quantitative prediction of the dynamics of separating unsteady flows, such as dynamic stall, is of crucial importance. This six-month SBIR Phase 1 study has developed several new pressure-based methodologies for solving 3D Navier-Stokes equations in both stationary and moving (body-comforting) coordinates. The present pressure-based algorithm is equally efficient for low speed incompressible flows and high speed compressible flows. The discretization of convective terms by the presently developed high-order TVD schemes requires no artificial dissipation and can properly resolve the concentrated vortices in the wing-body with minimum numerical diffusion. It is demonstrated that the proposed Newton's iteration technique not only increases the convergence rate but also strongly couples the iteration between pressure and velocities. The proposed hyperbolization of the pressure correction equation is shown to increase the solver's efficiency. The above proposed methodologies were implemented in an existing CFD code, REFLEQS. The modified code was used to simulate both static and dynamic stalls on two- and three-dimensional wing-body configurations. Three-dimensional effect and flow physics are discussed.

Ingestive behavior of lambs confined in individual and group stalls.

Science.gov (United States)

Filho, A Eustáquio; Carvalho, G G P; Pires, A J V; Silva, R R; Santos, P E F; Murta, R M; Pereira, F M

2014-02-01

The experiment was conducted to evaluate the ingestive behavior of lambs confined in individual and group stalls. We used thirty-four lambs in their growing phase, aged an average of three months, with mean initial live weight of 17.8±5.2 kg. They were allotted in a completely randomized design with 24 animals kept in individual stalls and 10 animals confined as a group. The experiment lasted for a total of 74 days, and the first 14 days were dedicated to the animals' adaption to the management, facilities and diets. The data collection period lasted 60 days, divided into three 20-d periods for the behavior evaluation. The animals were subjected to five days of visual observation during the experiment period, by the quantification of 24 h a day, with evaluations on the 15th day of each period and an interim evaluation consisting of two consecutive days on the 30th and 31st day of the experiment. The animals confined as a group consumed less (pbehavior.

A Method to Predict Compressor Stall in the TF34-100 Turbofan Engine Utilizing Real-Time Performance Data

Science.gov (United States)

2015-06-01

A METHOD TO PREDICT COMPRESSOR STALL IN THE TF34-100 TURBOFAN ENGINE UTILIZING REAL-TIME PERFORMANCE...THE TF34-100 TURBOFAN ENGINE UTILIZING REAL-TIME PERFORMANCE DATA THESIS Presented to the Faculty Department of Systems Engineering and...036 A METHOD TO PREDICT COMPRESSOR STALL IN THE TF34-100 TURBOFAN ENGINE UTILIZING REAL-TIME PERFORMANCE DATA Shuxiang ‘Albert’ Li, BS

Methods for sampling geographically mobile female traders in an East African market setting

Science.gov (United States)

Achiro, Lillian; Kwena, Zachary A.; McFarland, Willi; Neilands, Torsten B.; Cohen, Craig R.; Bukusi, Elizabeth A.; Camlin, Carol S.

2018-01-01

Background The role of migration in the spread of HIV in sub-Saharan Africa is well-documented. Yet migration and HIV research have often focused on HIV risks to male migrants and their partners, or migrants overall, often failing to measure the risks to women via their direct involvement in migration. Inconsistent measures of mobility, gender biases in those measures, and limited data sources for sex-specific population-based estimates of mobility have contributed to a paucity of research on the HIV prevention and care needs of migrant and highly mobile women. This study addresses an urgent need for novel methods for developing probability-based, systematic samples of highly mobile women, focusing on a population of female traders operating out of one of the largest open air markets in East Africa. Our method involves three stages: 1.) identification and mapping of all market stall locations using Global Positioning System (GPS) coordinates; 2.) using female market vendor stall GPS coordinates to build the sampling frame using replicates; and 3.) using maps and GPS data for recruitment of study participants. Results The location of 6,390 vendor stalls were mapped using GPS. Of these, 4,064 stalls occupied by women (63.6%) were used to draw four replicates of 128 stalls each, and a fifth replicate of 15 pre-selected random alternates for a total of 527 stalls assigned to one of five replicates. Staff visited 323 stalls from the first three replicates and from these successfully recruited 306 female vendors into the study for a participation rate of 94.7%. Mobilization strategies and involving traders association representatives in participant recruitment were critical to the study’s success. Conclusion The study’s high participation rate suggests that this geospatial sampling method holds promise for development of probability-based samples in other settings that serve as transport hubs for highly mobile populations. PMID:29324780

Flow Observations with Tufts and Lampblack of the Stalling of Four Typical Airfoil Sections in the NACA Variable-density Tunnel

Science.gov (United States)

Abbott, Ira H; Sherman, Albert

1938-01-01

A preliminary investigation of the stalling processes of four typical airfoil sections was made over the critical range of the Reynolds Number. Motion pictures were taken of the movements of small silk tufts on the airfoil surface as the angle of attack increased through a range of angles including the stall. The boundary-layer flow also at certain angles of attack was indicated by the patterns formed by a suspension of lampblack in oil brushed onto the airfoil surface. These observations were analyzed together with corresponding force-test measurements to derive a picture of the stalling processes of airfoils.

DNA replication origins-where do we begin?

Science.gov (United States)

Prioleau, Marie-Noëlle; MacAlpine, David M

2016-08-01

For more than three decades, investigators have sought to identify the precise locations where DNA replication initiates in mammalian genomes. The development of molecular and biochemical approaches to identify start sites of DNA replication (origins) based on the presence of defining and characteristic replication intermediates at specific loci led to the identification of only a handful of mammalian replication origins. The limited number of identified origins prevented a comprehensive and exhaustive search for conserved genomic features that were capable of specifying origins of DNA replication. More recently, the adaptation of origin-mapping assays to genome-wide approaches has led to the identification of tens of thousands of replication origins throughout mammalian genomes, providing an unprecedented opportunity to identify both genetic and epigenetic features that define and regulate their distribution and utilization. Here we summarize recent advances in our understanding of how primary sequence, chromatin environment, and nuclear architecture contribute to the dynamic selection and activation of replication origins across diverse cell types and developmental stages. © 2016 Prioleau and MacAlpine; Published by Cold Spring Harbor Laboratory Press.

Interactions and Localization of Escherichia coli Error-Prone DNA Polymerase IV after DNA Damage.

Science.gov (United States)

Mallik, Sarita; Popodi, Ellen M; Hanson, Andrew J; Foster, Patricia L

2015-09-01

Escherichia coli's DNA polymerase IV (Pol IV/DinB), a member of the Y family of error-prone polymerases, is induced during the SOS response to DNA damage and is responsible for translesion bypass and adaptive (stress-induced) mutation. In this study, the localization of Pol IV after DNA damage was followed using fluorescent fusions. After exposure of E. coli to DNA-damaging agents, fluorescently tagged Pol IV localized to the nucleoid as foci. Stepwise photobleaching indicated ∼60% of the foci consisted of three Pol IV molecules, while ∼40% consisted of six Pol IV molecules. Fluorescently tagged Rep, a replication accessory DNA helicase, was recruited to the Pol IV foci after DNA damage, suggesting that the in vitro interaction between Rep and Pol IV reported previously also occurs in vivo. Fluorescently tagged RecA also formed foci after DNA damage, and Pol IV localized to them. To investigate if Pol IV localizes to double-strand breaks (DSBs), an I-SceI endonuclease-mediated DSB was introduced close to a fluorescently labeled LacO array on the chromosome. After DSB induction, Pol IV localized to the DSB site in ∼70% of SOS-induced cells. RecA also formed foci at the DSB sites, and Pol IV localized to the RecA foci. These results suggest that Pol IV interacts with RecA in vivo and is recruited to sites of DSBs to aid in the restoration of DNA replication. DNA polymerase IV (Pol IV/DinB) is an error-prone DNA polymerase capable of bypassing DNA lesions and aiding in the restart of stalled replication forks. In this work, we demonstrate in vivo localization of fluorescently tagged Pol IV to the nucleoid after DNA damage and to DNA double-strand breaks. We show colocalization of Pol IV with two proteins: Rep DNA helicase, which participates in replication, and RecA, which catalyzes recombinational repair of stalled replication forks. Time course experiments suggest that Pol IV recruits Rep and that RecA recruits Pol IV. These findings provide in vivo evidence

Indoor and outdoor concentrations of RSP, NO2 and selected volatile organic compounds at 32 shoe stalls located near busy roadways in Seoul, Korea

International Nuclear Information System (INIS)

Bae, Hyunjoo; Chung, Moonho; Yang, Wonho

2004-01-01

It is suspected that persons who work in indoor environments near busy roadways are exposed to elevated levels of air pollutants during working hours. This study evaluated the potential exposure and source contribution associated with traffic-related air pollution for workers (polishers and repairmen) in shoe stalls from each of 32 districts during working hours in Seoul, Korea. The shoe stalls have been located at very close distances to the busy roadways. In this study, shoe stall workers could be exposed to high levels of respirable suspended particulate (RSP), nitrogen dioxide (NO 2 ) and volatile organic compounds (VOCs) from outdoor sources such as traffic exhaust, as well as indoor sources in the shoe stalls such as dust on the shoes, portable gas ranges, organic solvents, adhesives and shoe polish. Compounds of particular note included indoor mean concentrations of benzene, toluene, m/p-xylene and o-xylene were 0.732, 6.777, 4.080 and 1.302 mg/m 3 , respectively, in all shoe stalls. Mean indoor/outdoor ratios for toluene and m/p-xylene concentrations were 54.52 and 20.84, respectively. The contribution of vehicle exhaust emissions to indoor air quality of shoe stalls was identified by means of correlating the relationships between simultaneously measured air pollutant concentrations indoors and outdoors. Unlike RSP and NO 2 , indoor VOCs concentrations of shoe stalls mainly originated from indoor sources vs. outdoor sources

Short revolving wings enable hovering animals to avoid stall and reduce drag

Science.gov (United States)

Lentink, David; Kruyt, Jan W.; Heijst, Gertjan F.; Altshuler, Douglas L.

2014-11-01

Long and slender wings reduce the drag of airplanes, helicopters, and gliding animals, which operate at low angle of attack (incidence). Remarkably, there is no evidence for such influence of wing aspect ratio on the energetics of hovering animals that operate their wings at much higher incidence. High incidence causes aircraft wings to stall, hovering animals avoid stall by generating an attached vortex along the leading edge of their wings that elevates lift. Hypotheses that explain this capability include the necessity for a short radial distance between the shoulder joint and wing tip, measured in chord lengths, instead of the long tip-to-tip distance that elevates aircraft performance. This stems from how hovering animals revolve their wings around a joint, a condition for which the precise effect of aspect ratio on stall performance is unknown. Here we show that the attachment of the leading edge vortex is determined by wing aspect ratio with respect to the center of rotation-for a suite of aspect ratios that represent both animal and aircraft wings. The vortex remains attached when the local radius is shorter than 4 chord lengths, and separates outboard on more slender wings. Like most other hovering animals, hummingbirds have wing aspect ratios between 3 and 4, much stubbier than helicopters. Our results show this makes their wings robust against flow separation, which reduces drag below values obtained with more slender wings. This revises our understanding of how aspect ratio improves performance at low Reynolds numbers.

Fragile genomic sites are associated with origins of replication.

Science.gov (United States)

Di Rienzi, Sara C; Collingwood, David; Raghuraman, M K; Brewer, Bonita J

2009-09-09

Genome rearrangements are mediators of evolution and disease. Such rearrangements are frequently bounded by transfer RNAs (tRNAs), transposable elements, and other repeated elements, suggesting a functional role for these elements in creating or repairing breakpoints. Though not well explored, there is evidence that origins of replication also colocalize with breakpoints. To investigate a potential correlation between breakpoints and origins, we analyzed evolutionary breakpoints defined between Saccharomyces cerevisiae and Kluyveromyces waltii and S. cerevisiae and a hypothetical ancestor of both yeasts, as well as breakpoints reported in the experimental literature. We find that origins correlate strongly with both evolutionary breakpoints and those described in the literature. Specifically, we find that origins firing earlier in S phase are more strongly correlated with breakpoints than are later-firing origins. Despite origins being located in genomic regions also bearing tRNAs and Ty elements, the correlation we observe between origins and breakpoints appears to be independent of these genomic features. This study lays the groundwork for understanding the mechanisms by which origins of replication may impact genome architecture and disease.

Animal hygiene assessment of microclimate in semi open free-stall barns for dairy cows

Directory of Open Access Journals (Sweden)

D. Dimov

2017-03-01

Full Text Available Abstract. The study was conducted in three semi open free-stall barns (B1, B2, and B3 for dairy cows with capacities for 120, 120 and 500 cows, respectively, from three different dairy farms (F-1, F-2 and F-3, situated in Central Southern Bulgaria. The investigated farms had the same production system – loose housing in semi open free-stall dairy barn. For each of the farms the main microclimatic parameters – air temperature, relative humidity and speed of airflow were recorded twice a month at 10.00 h 12.00 h, 14.00 h, 16.00 h and 18.00 h of the day inside the barns in three main technological zones - above the stalls, above manure and feed alleys and outside the buildings. It was found that: a Microclimatic parameters (air temperature, air relative humidity and speed of airflow in technological zones (above the stalls, the manure and feed alleys of three semi open free-stall dairy barns meet the animal hygienic requirements for all seasons according to Regulation No. 44 (2006. Exceptions are some values of relative humidity in B1 and B2 in the spring, and in B1 in winter and summer, which are lower than the minimum humidity (50% according to the standard. b The investigated barns are characterized with poor insulation and do not provide enough isolation from the external ambient temperatures. With the exception of winter, the temperature of the air inside the buildings was lower than that outside, with minor differences for all seasons. The fans in the barns have no effect on the inside air temperature, especially in summer. There was a risk of higher temperatures mainly during the summer period. c There is no significant difference between the average temperatures, air humidity and speed of airflow in all technological zones of the investigated barns. d The largest and statistically significant is the difference between the relative air humidity outside and inside the building in Farm 3, followed by buildings in Farm 1 and 2, where the

RAD52 Facilitates Mitotic DNA Synthesis Following Replication Stress

DEFF Research Database (Denmark)

Bhowmick, Rahul; Minocherhomji, Sheroy; Hickson, Ian D

2016-01-01

Homologous recombination (HR) is necessary to counteract DNA replication stress. Common fragile site (CFS) loci are particularly sensitive to replication stress and undergo pathological rearrangements in tumors. At these loci, replication stress frequently activates DNA repair synthesis in mitosis...... replication stress at CFS loci during S-phase. In contrast, MiDAS is RAD52 dependent, and RAD52 is required for the timely recruitment of MUS81 and POLD3 to CFSs in early mitosis. Our results provide further mechanistic insight into MiDAS and define a specific function for human RAD52. Furthermore, selective...

Feline coronavirus replication is affected by both cyclophilin A and cyclophilin B.

Science.gov (United States)

Tanaka, Yoshikazu; Sato, Yuka; Sasaki, Takashi

2017-02-01

Feline coronavirus (FCoV) causes the fatal disease feline infectious peritonitis, which is currently incurable by drug treatment, and no effective vaccines are available. Cyclosporin A (CsA), a cyclophilin (Cyp) inhibitor, inhibits the replication of FCoV in vitro and in vivo as well as the replication of human and animal coronaviruses. However, the mechanism underlying the regulation of coronavirus replication by CsA is unknown. In this study, we analysed the role of Cyps in FCoV replication using knockdown and knockout cells specific to Cyps. Inhibition of CypA and CypB reduced FCoV replication, with replication in knockout cells being much less than that in knockdown cells. Furthermore, the proteins expressed by CypA and CypB harbouring mutations in their respective predicted peptidyl-prolyl cis-transisomerase active sites, which also alter the affinities between Cyps and CsA, inhibited FCoV replication. These findings indicate that the peptidyl-prolyl cis-transisomerase active sites of Cyps might be required for FCoV replication.

Power control of a wind farm with active stall wind turbines and AC grid connection

DEFF Research Database (Denmark)

Hansen, Anca Daniela; Sørensen, Poul; Iov, Florin

both the control on wind turbine level as well as the central control on the wind farm level. The ability of active stall wind farms with AC grid connection to regulate the power production to the reference power ordered by the operators is assessed and discussed by means of simulations.......This paper describes the design of a centralised wind farm controller for a wind farm made-up exclusively of active stall wind turbines with AC grid connection. The overall aim of such controller is to enable the wind farms to provide the best grid support. The designed wind farm control involves...

Opposite replication polarities of transcribed and nontranscribed histone H5 genes

International Nuclear Information System (INIS)

Trempe, J.P.; Lindstrom, Y.I.; Leffak, M.

1988-01-01

The authors used an in vitro nuclear runoff replication assay to analyze the direction of replication of the active and inactive histone H5 genes in avian cells. In embryonic erythrocytes the transcribed histone H5 gene displayed sensitivity to endogenous nuclease cleavage. In contrast, this gene was insensitive to endogenous nuclease digestion under the same conditions in nuclei of the lymphoblastoid cell line MSB-1, and histone H5 gene transcripts were not detectable by dot-blot analysis of MSB-1 cell RNA. When nuclei were isolated from embryonic erythrocyctes and incubated with bromodeoxyuridine triphosphate, runoff replication from endogenous nuclease cleavage sites led to a relative enrichment for fragments near the 3' end of the histone H5 gene in the density-labeled DNA. In nuclei of MSB-1 cells or chicken embryo fibroblasts, however, runoff replication from restriction enzyme-cut sites (or induced endogenous nuclease-cut sites in MSB-1 nuclei) led to a relative enrichment for fragments near the 5' end of the H5 gene in dense DNA. Based on the enhanced incorporation of bromodeoxyuridine into origin-distal regions of DNA during the in vitro runoff replication assay, the authors conclude that the active histone H5 gene in embryonic erythrocytes is preferentially replicated in the transcriptional direction from an origin in the 5'-flanking DNA, whereas its inactive counterparts in MSB-1 cells and chicken embryo fibroblasts are preferentially replicated in the opposite direction

Physics of Prestall Propagating Disturbances in Axial Compressors and Their Potential as a Stall Warning Indicator

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Mario Eck

2017-03-01

Full Text Available Axial compressors in aero engines are prone to suffering a breakdown of orderly flow when operating at the peak of the pressure rise characteristic. The damaging potential of separated flows is why a safe distance has to be left between every possible operating point and an operating point at which stall occurs. During earlier investigations of stall inception mechanisms, a new type of prestall instability has been found. In this study, it could be demonstrated that the prestall instability characterised by discrete flow disturbances can be clearly assigned to the subject of “Rotating Instabilities”. Propagating disturbances are responsible for the rise in blade passing irregularity. If the mass flow is reduced successively, the level of irregularity increases until the prestall condition devolves into rotating stall. The primary objective of the current work is to highlight the basic physics behind these prestall disturbances by complementary experimental and numerical investigations. Before reaching the peak of the pressure rise characteristic flow, disturbances appear as small vortex tubes with one end attached to the casing and the other attached to the suction surface of the rotor blade. These vortex structures arise when the entire tip region is affected by blockage and at the same time the critical rotor incidence is not exceeded in this flow regime. Furthermore, a new stall indicator was developed by applying statistical methods to the unsteady pressure signal measured over the rotor blade tips, thus granting a better control of the safety margin.

Herpes simplex virus replication compartments can form by coalescence of smaller compartments

International Nuclear Information System (INIS)

Taylor, Travis J; McNamee, Elizabeth E.; Day, Cheryl; Knipe, David M.

2003-01-01

Herpes simplex virus (HSV) uses intranuclear compartmentalization to concentrate the viral and cellular factors required for the progression of the viral life cycle. Processes as varied as viral DNA replication, late gene expression, and capsid assembly take place within discrete structures within the nucleus called replication compartments. Replication compartments are hypothesized to mature from a few distinct structures, called prereplicative sites, that form adjacent to cellular nuclear matrix-associated ND10 sites. During productive infection, the HSV single-stranded DNA-binding protein ICP8 localizes to replication compartments. To further the understanding of replication compartment maturation, we have constructed and characterized a recombinant HSV-1 strain that expresses an ICP8 molecule with green fluorescent protein (GFP) fused to its C terminus. In transfected Vero cells that were infected with HSV, the ICP8-GFP protein localized to prereplicative sites in the presence of the viral DNA synthesis inhibitor phosphonoacetic acid (PAA) or to replication compartments in the absence of PAA. A recombinant HSV-1 strain expressing the ICP8-GFP virus replicated in Vero cells, but the yield was increased by 150-fold in an ICP8-complementing cell line. Using the ICP8-GFP protein as a marker for replication compartments, we show here that these structures start as punctate structures early in infection and grow into large, globular structures that eventually fill the nucleus. Large replication compartments were formed by small structures that either moved through the nucleus to merge with adjacent compartments or remained relatively stationary within the nucleus and grew by accretion and fused with neighboring structures

Low-Order Modeling of Dynamic Stall on Airfoils in Incompressible Flow

Science.gov (United States)

Narsipur, Shreyas

Unsteady aerodynamics has been a topic of research since the late 1930's and has increased in popularity among researchers studying dynamic stall in helicopters, insect/bird flight, micro air vehicles, wind-turbine aerodynamics, and ow-energy harvesting devices. Several experimental and computational studies have helped researchers gain a good understanding of the unsteady ow phenomena, but have proved to be expensive and time-intensive for rapid design and analysis purposes. Since the early 1970's, the push to develop low-order models to solve unsteady ow problems has resulted in several semi-empirical models capable of effectively analyzing unsteady aerodynamics in a fraction of the time required by high-order methods. However, due to the various complexities associated with time-dependent flows, several empirical constants and curve fits derived from existing experimental and computational results are required by the semi-empirical models to be an effective analysis tool. The aim of the current work is to develop a low-order model capable of simulating incompressible dynamic-stall type ow problems with a focus on accurately modeling the unsteady ow physics with the aim of reducing empirical dependencies. The lumped-vortex-element (LVE) algorithm is used as the baseline unsteady inviscid model to which augmentations are applied to model unsteady viscous effects. The current research is divided into two phases. The first phase focused on augmentations aimed at modeling pure unsteady trailing-edge boundary-layer separation and stall without leading-edge vortex (LEV) formation. The second phase is targeted at including LEV shedding capabilities to the LVE algorithm and combining with the trailing-edge separation model from phase one to realize a holistic, optimized, and robust low-order dynamic stall model. In phase one, initial augmentations to theory were focused on modeling the effects of steady trailing-edge separation by implementing a non-linear decambering

Replication of kinetoplast minicircle DNA

International Nuclear Information System (INIS)

Sheline, C.T.

1989-01-01

These studies describe the isolation and characterization of early minicircle replication intermediates from Crithidia fasciculata, and Leishmania tarentolae, the mitochondrial localization of a type II topoisomerase (TIImt) in C. fasciculata, and the implication of the aforementioned TIImt in minicircle replication in L. tarentolae. Early minicircle replication intermediates from C. fasciculata were identified and characterized using isolated kinetoplasts to incorporate radiolabeled nucleotides into its DNA. The pulse-label in an apparent theta-type intermediate chase into two daughter molecules. A uniquely gapped, ribonucleotide primed, knotted molecule represents the leading strand in the model proposed, and a highly gapped molecule represents the lagging strand. This theta intermediate is repaired in vitro to a doubly nicked catenated dimer which was shown to result from the replication of a single parental molecule. Very similar intermediates were found in the heterogeneous population of minicircles of L. tarentolae. The sites of the Leishmania specific discontinuities were mapped and shown to lie within the universally conserved sequence blocks in identical positions as compared to C. fasciculata and Trypanosoma equiperdum

Manual of Cupule Replication Technology

Directory of Open Access Journals (Sweden)

Giriraj Kumar

2015-09-01

Full Text Available Throughout the world, iconic rock art is preceded by non-iconic rock art. Cupules (manmade, roughly semi-hemispherical depressions on rocks form the major bulk of the early non-iconic rock art globally. The antiquity of cupules extends back to the Lower Paleolithic in Asia and Africa, hundreds of thousand years ago. When one observes these cupules, the inquisitive mind poses so many questions with regard to understanding their technology, reasons for selecting the site, which rocks were used to make the hammer stones used, the skill and cognitive abilities employed to create the different types of cupules, the objective of their creation, their age, and so on. Replication of the cupules can provide satisfactory answers to some of these questions. Comparison of the hammer stones and cupules produced by the replication process with those obtained from excavation can provide support to observations. This paper presents a manual of cupule replication technology based on our experience of cupule replication on hard quartzite rock near Daraki-Chattan in the Chambal Basin, India.

Non‐Canonical Replication Initiation: You’re Fired!

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Bazilė Ravoitytė

2017-01-01

Full Text Available The division of prokaryotic and eukaryotic cells produces two cells that inherit a perfect copy of the genetic material originally derived from the mother cell. The initiation of canonical DNA replication must be coordinated to the cell cycle to ensure the accuracy of genome duplication. Controlled replication initiation depends on a complex interplay of cis‐acting DNA sequences, the so‐called origins of replication (ori, with trans‐acting factors involved in the onset of DNA synthesis. The interplay of cis‐acting elements and trans‐acting factors ensures that cells initiate replication at sequence‐specific sites only once, and in a timely order, to avoid chromosomal endoreplication. However, chromosome breakage and excessive RNA:DNA hybrid formation can cause breakinduced (BIR or transcription‐initiated replication (TIR, respectively. These non‐canonical replication events are expected to affect eukaryotic genome function and maintenance, and could be important for genome evolution and disease development. In this review, we describe the difference between canonical and non‐canonical DNA replication, and focus on mechanistic differences and common features between BIR and TIR. Finally, we discuss open issues on the factors and molecular mechanisms involved in TIR.

Dairy cows welfare quality in tie-stall housing system with or without access to exercise

Science.gov (United States)

2013-01-01

Background Tie-stall housing of dairy cows is used extensively worldwide, despite of the welfare concerns regarding the restriction of voluntary movement and limitation of expression of the cows’ natural behaviour. The aim of this study was to compare the welfare quality of dairy cows kept in two types of tie-stall housing systems: with regular outdoor exercise and without access to exercise. In addition, the study investigated the relationship between different welfare measures of dairy cows kept in tie-stalls. Methods 3,192 lactating cows were assessed using the Welfare Quality® assessment protocol for cattle in 80 commercial dairy farms, half of the farms providing outdoor access for the animals to exercise. The descriptive statistical indicators were determined for the assessed measures and for the welfare criteria and principle scores. The data obtained in the two housing types were compared and the correlation coefficients were calculated between the different welfare measures. Results The significant differences found between the two housing systems for the majority of the animal based measures indicate the positive effect of exercise on the welfare of tethered cows. Many of the animal welfare parameters correlated with each other. For the farms allowing the cows’ turnout in a paddock, pasture or both, the mean scores for the welfare criteria and principles were higher than for the farms with permanent tethering of the cows, except the criteria absence of prolonged hunger and expression of social behaviours. The lowest scores were obtained for the criterion positive emotional state, in both housing systems. With regard to the overall classification, none of the farms were considered excellent. In the not classified category were only farms with all-year-round tethering of the animals and in the enhanced category only farms where the cows had outdoor access. Conclusions The welfare quality of the investigated dairy cows was significantly better in the

Lift hysteresis at stall as an unsteady boundary-layer phenomenon

Science.gov (United States)

Moore, Franklin K

1956-01-01

Analysis of rotating stall of compressor blade rows requires specification of a dynamic lift curve for the airfoil section at or near stall, presumably including the effect of lift hysteresis. Consideration of the magnus lift of a rotating cylinder suggests performing an unsteady boundary-layer calculation to find the movement of the separation points of an airfoil fixed in a stream of variable incidence. The consideration of the shedding of vorticity into the wake should yield an estimate of lift increment proportional to time rate of change of angle of attack. This increment is the amplitude of the hysteresis loop. An approximate analysis is carried out according to the foregoing ideas for a 6:1 elliptic airfoil at the angle of attack for maximum lift. The assumptions of small perturbations from maximum lift are made, permitting neglect of distributed vorticity in the wake. The calculated hysteresis loop is counterclockwise. Finally, a discussion of the forms of hysteresis loops is presented; and, for small reduced frequency of oscillation, it is concluded that the concept of a viscous "time lag" is appropriate only for harmonic variations of angle of attack with time at mean conditions other than maximum lift.

Hydroxyurea-Mediated Cytotoxicity Without Inhibition of Ribonucleotide Reductase.

Science.gov (United States)

Liew, Li Phing; Lim, Zun Yi; Cohen, Matan; Kong, Ziqing; Marjavaara, Lisette; Chabes, Andrei; Bell, Stephen D

2016-11-01

In many organisms, hydroxyurea (HU) inhibits class I ribonucleotide reductase, leading to lowered cellular pools of deoxyribonucleoside triphosphates. The reduced levels for DNA precursors is believed to cause replication fork stalling. Upon treatment of the hyperthermophilic archaeon Sulfolobus solfataricus with HU, we observe dose-dependent cell cycle arrest, accumulation of DNA double-strand breaks, stalled replication forks, and elevated levels of recombination structures. However, Sulfolobus has a HU-insensitive class II ribonucleotide reductase, and we reveal that HU treatment does not significantly impact cellular DNA precursor pools. Profiling of protein and transcript levels reveals modulation of a specific subset of replication initiation and cell division genes. Notably, the selective loss of the regulatory subunit of the primase correlates with cessation of replication initiation and stalling of replication forks. Furthermore, we find evidence for a detoxification response induced by HU treatment. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

MICROBIOLOGICAL STUDY ON ICE FROM A FISH STALL

Directory of Open Access Journals (Sweden)

E. Tirloni

2012-08-01

Full Text Available The ice used for exposure of fish products could be a source of secondary contamination due to ice machine, due to not respected good manufacturing practices, particularly when ice is left on the fish stall and the next day the new layer is deposited over the old one. Aim of this study was the verification of the hygienic risk of this procedure through analyses of the liquid produced by the zones “thawed cephalopods” and “fresh whole fish”. Almost the microorganisms found were Gram negative (in particular Pseudomonadaceae.

An automatic system for the detection of dairy cows lying behaviour in free-stall barns

Directory of Open Access Journals (Sweden)

Simona M.C. Porto

2013-09-01

Full Text Available In this paper, a method for the automatic detection of dairy cow lying behaviour in free-stall barns is proposed. A computer visionbased system (CVBS composed of a video-recording system and a cow lying behaviour detector based on the Viola Jones algorithm was developed. The CVBS performance was tested in a head-to-head free stall barn. Two classifiers were implemented in the software component of the CVBS to obtain the cow lying behaviour detector. The CVBS was validated by comparing its detection results with those generated from visual recognition. This comparison allowed the following accuracy indices to be calculated: the branching factor (BF, the miss factor (MF, the sensitivity, and the quality percentage (QP. The MF value of approximately 0.09 showed that the CVBS missed one cow every 11 well detected cows. Conversely, the BF value of approximately 0.08 indicated that one false positive was detected every 13 well detected cows. The high value of approximately 0.92 obtained for the sensitivity index and that obtained for QP of about 0.85 revealed the ability of the proposed system to detect cows lying in the stalls.

Inviscid double wake model for stalled airfoils

International Nuclear Information System (INIS)

Marion, L; Ramos-García, N; Sørensen, J N

2014-01-01

An inviscid double wake model based on a steady two-dimensional panel method has been developed to predict aerodynamic loads of wind turbine airfoils in the deep stall region. The separated flow is modelled using two constant vorticity sheets which are released at the trailing edge and at the separation point. A calibration of the code through comparison with experiments has been performed using one set of airfoils. A second set of airfoils has been used for the validation of the calibrated model. Predicted aerodynamic forces for a wide range of angles of attack (0 to 90 deg) are in overall good agreement with wind tunnel measurements

Zonal RANS/LES coupling simulation of a transitional and separated flow around an airfoil near stall

Energy Technology Data Exchange (ETDEWEB)

Richez, F.; Mary, I.; Gleize, V. [ONERA, Department of Computational Fluid Dynamics and Aeroacoustics, 29 Avenue de la Division Leclerc, BP 72, Chatillon (France); Basdevant, C. [Universite Paris-Nord, Laboratoire d' Analyse, Geometrie et Applications, CNRS, Villetaneuse (France)

2008-05-15

The objective of the current study is to examine the course of events leading to stall just before its occurrence. The stall mechanisms are very sensitive to the transition that the boundary layer undergoes near the leading edge of the profile by a so-called laminar separation bubble (LSB). In order to provide helpful insights into this complex flow, a zonal Reynolds-averaged Navier-Stokes (RANS)/large-eddy simulation (LES) simulation of the flow around an airfoil near stall has been achieved and its results are presented and analyzed in this paper. LSB has already been numerically studied by direct numerical simulation (DNS) or LES, but for a flat plate with an adverse pressure gradient only. We intend, in this paper, to achieve a detailed analysis of the transition process by a LSB in more realistic conditions. The comparison with a linear instability analysis has shown that the numerical instability mechanism in the LSB provides the expected frequency of the perturbations. Furthermore, the right order of magnitude for the turbulence intensities at the reattachment point is found. (orig.)

Suppression of inducer stall based on inlet recirculation in a centrifugal impeller. 1st Report. Improvement in stall limit by ring groove arrangement; Enshin haneguruma iriguchi junkanryu ni yoru inducer shissoku no yokusei. 1. kanjoko ni yoru shissoku genkai no kaizen

Energy Technology Data Exchange (ETDEWEB)

Ueki, H.; Ishida, M.; Sakaguchi, D.; Sun, Z. [Nagasaki University, Nagasaki (Japan). Faculty of Engineering

2000-07-25

A ring groove arrangement is proposed to suppress unstable flow or surge in a centrifugal blower. The ring groove arrangement connects the upstream position of impeller inlet and the inducer throat tip through a bypass. The recirculation flow 'vas formed by the pressure difference between the two positions, and the recirculation flow rate was changed by increasing the ring groove widths. The inlet recirculation results in a decrease in the flow rate of unstable flow inception, and an up to 800 improvement in stall limit was obtained by the ring groove arrangement at a small expense of the delivery pressure drop. The improvement of stall limit in the present experiment seems to be mainly due to decrease in flow incidence based on the inlet recirculation flow. Tre flow incidence decreases more as the recirculation flow rate increases, thus resulting in a larger improvement in stall limit. (author)

Identifying sites of replication initiation in yeast chromosomes: looking for origins in all the right places.

Science.gov (United States)

van Brabant, A J; Hunt, S Y; Fangman, W L; Brewer, B J

1998-06-01

DNA fragments that contain an active origin of replication generate bubble-shaped replication intermediates with diverging forks. We describe two methods that use two-dimensional (2-D) agarose gel electrophoresis along with DNA sequence information to identify replication origins in natural and artificial Saccharomyces cerevisiae chromosomes. The first method uses 2-D gels of overlapping DNA fragments to locate an active chromosomal replication origin within a region known to confer autonomous replication on a plasmid. A variant form of 2-D gels can be used to determine the direction of fork movement, and the second method uses this technique to find restriction fragments that are replicated by diverging forks, indicating that a bidirectional replication origin is located between the two fragments. Either of these two methods can be applied to the analysis of any genomic region for which there is DNA sequence information or an adequate restriction map.

Mapping replication origins in yeast chromosomes.

Science.gov (United States)

Brewer, B J; Fangman, W L

1991-07-01

The replicon hypothesis, first proposed in 1963 by Jacob and Brenner, states that DNA replication is controlled at sites called origins. Replication origins have been well studied in prokaryotes. However, the study of eukaryotic chromosomal origins has lagged behind, because until recently there has been no method for reliably determining the identity and location of origins from eukaryotic chromosomes. Here, we review a technique we developed with the yeast Saccharomyces cerevisiae that allows both the mapping of replication origins and an assessment of their activity. Two-dimensional agarose gel electrophoresis and Southern hybridization with total genomic DNA are used to determine whether a particular restriction fragment acquires the branched structure diagnostic of replication initiation. The technique has been used to localize origins in yeast chromosomes and assess their initiation efficiency. In some cases, origin activation is dependent upon the surrounding context. The technique is also being applied to a variety of eukaryotic organisms.

Transcription-replication conflicts at chromosomal fragile sites—consequences in M phase and beyond

DEFF Research Database (Denmark)

Østergaard, Vibe Hallundbæk; Lisby, Michael

2017-01-01

transcription and replication patterns. At the same time, these chromosomal fragile sites engage in aberrant DNA structures in mitosis. Here, we discuss the mechanistic details of transcription–replication conflicts including putative scenarios for R-loop-induced replication inhibition to understand how...... transcription–replication conflicts transition from S phase into various aberrant DNA structures in mitosis....

Monitoring Replication Protein A (RPA) dynamics in homologous recombination through site-specific incorporation of non-canonical amino acids.

Science.gov (United States)

Pokhrel, Nilisha; Origanti, Sofia; Davenport, Eric Parker; Gandhi, Disha; Kaniecki, Kyle; Mehl, Ryan A; Greene, Eric C; Dockendorff, Chris; Antony, Edwin

2017-09-19

An essential coordinator of all DNA metabolic processes is Replication Protein A (RPA). RPA orchestrates these processes by binding to single-stranded DNA (ssDNA) and interacting with several other DNA binding proteins. Determining the real-time kinetics of single players such as RPA in the presence of multiple DNA processors to better understand the associated mechanistic events is technically challenging. To overcome this hurdle, we utilized non-canonical amino acids and bio-orthogonal chemistry to site-specifically incorporate a chemical fluorophore onto a single subunit of heterotrimeric RPA. Upon binding to ssDNA, this fluorescent RPA (RPAf) generates a quantifiable change in fluorescence, thus serving as a reporter of its dynamics on DNA in the presence of multiple other DNA binding proteins. Using RPAf, we describe the kinetics of facilitated self-exchange and exchange by Rad51 and mediator proteins during various stages in homologous recombination. RPAf is widely applicable to investigate its mechanism of action in processes such as DNA replication, repair and telomere maintenance. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Autonomous replication of plasmids bearing monkey DNA origin-enriched sequences

International Nuclear Information System (INIS)

Frappier, L.; Zannis-Hadjopoulos, M.

1987-01-01

Twelve clones of origin-enriched sequences (ORS) isolated from early replicating monkey (CV-1) DNA were examined for transient episomal replication in transfected CV-1, COS-7, and HeLa cells. Plasmid DNA was isolated at time intervals after transfection and screened by the Dpn I resistance assay or by the bromodeoxyuridine substitution assay to differentiate between input and replicated DNA. The authors have identified four monkey ORS (ORS3, -8, -9, and -12) that can support plasmid replication in mammalian cells. This replication is carried out in a controlled and semiconservative manner characteristic of mammalian replicons. ORS replication was most efficient in HeLa cells. Electron microscopy showed ORS8 and ORS12 plasmids of the correct size with replication bubbles. Using a unique restriction site in ORS12, we have mapped the replication bubble within the monkey DNA sequence

Broken silence restored-remodeling primes for deacetylation at replication forks

DEFF Research Database (Denmark)

Jasencakova, Zuzana; Groth, Anja

2011-01-01

Faithful propagation of chromatin structures requires assimilation of new histones to the modification profile of individual loci. In this issue of Molecular Cell, Rowbotham and colleagues identify a remodeler, SMARCAD1, acting at replication sites to facilitate histone deacetylation and restorat......Faithful propagation of chromatin structures requires assimilation of new histones to the modification profile of individual loci. In this issue of Molecular Cell, Rowbotham and colleagues identify a remodeler, SMARCAD1, acting at replication sites to facilitate histone deacetylation...

Characterization of a Non-Canonical Signal Peptidase Cleavage Site in a Replication Protein from Tomato Ringspot Virus.

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Ting Wei

Full Text Available The NTB-VPg polyprotein from tomato ringspot virus is an integral membrane replication protein associated with endoplasmic reticulum membranes. A signal peptidase (SPase cleavage was previously detected in the C-terminal region of NTB-VPg downstream of a 14 amino acid (aa-long hydrophobic region (termed TM2. However, the exact location of the cleavage site was not determined. Using in vitro translation assays, we show that the SPase cleavage site is conserved in the NTB-VPg protein from various ToRSV isolates, although the rate of cleavage varies from one isolate to another. Systematic site-directed mutagenesis of the NTB-VPg SPase cleavage sites of two ToRSV isolates allowed the identification of sequences that affect cleavage efficiency. We also present evidence that SPase cleavage in the ToRSV-Rasp2 isolate occurs within a GAAGG sequence likely after the AAG (GAAG/G. Mutation of a downstream MAAV sequence to AAAV resulted in SPase cleavage at both the natural GAAG/G and the mutated AAA/V sequences. Given that there is a distance of seven aa between the two cleavage sites, this indicates that there is flexibility in the positioning of the cleavage sites relative to the inner surface of the membrane and the SPase active site. SPase cleavage sites are typically located 3-7 aa downstream of the hydrophobic region. However, the NTB-VPg GAAG/G cleavage site is located 17 aa downstream of the TM2 hydrophobic region, highlighting unusual features of the NTB-VPg SPase cleavage site. A putative 11 aa-long amphipathic helix was identified immediately downstream of the TM2 region and five aa upstream of the GAAG/G cleavage site. Based on these results, we present an updated topology model in which the hydrophobic and amphipathic domains form a long tilted helix or a bent helix in the membrane lipid bilayer, with the downstream cleavage site(s oriented parallel to the membrane inner surface.

A Beddoes-Leishman type dynamic stall model in state-space and indicial formulations

DEFF Research Database (Denmark)

Hansen, M.H.; Gaunaa, Mac; Aagaard Madsen, Helge

2004-01-01

This report contains a description of a Beddoes-Leishman type dynamic stall model in both a state-space and an indicial function formulation. The model predicts the unsteady aerodynamic forces and moment on an airfoil section undergoing arbitrary motionin heave, lead-lag, and pitch. The model...... features, such as overshoot of the lift, in the stall region. The linearized model is shown to give identicalresults to the full model for small amplitude oscillations. Furthermore, it is shown that the response of finite thichkness airfoils can be reproduced to a high accuracy by the use of specific...... is carried out by comparing the response of the model with inviscid solutions and observing the general behavior of the model using known airfoil data as input. Theproposed dynamic model gives results identical to inviscid solutions within the attached-flow region; and it exhibits the expected dynamic...

Prediction of RNA Polymerase II recruitment, elongation and stalling from histone modification data

DEFF Research Database (Denmark)

Chen, Yun; Jørgensen, Mette; Kolde, Raivo

2011-01-01

of RNAPII stalling. CONCLUSIONS: In this study we introduce a general framework to accurately predict the level of RNAPII recruitment, elongation, stalling and mRNA expression from chromatin signals. The versatility of the method also makes it ideally suited to investigate other genomic data....... strategies are needed to progress from descriptive annotation of data to quantitative, predictive models. RESULTS: Here, we describe a computational framework which with high accuracy can predict the locations of core promoters, the amount of recruited RNAPII at the promoter, the amount of elongating RNAPII...... of these four marks are found to be necessary for recruitment of RNAPII but not sufficient for the elongation. We also show that the spatial distributions of histone marks are almost as predictive as the signal strength and that a set of histone marks immediately downstream of the TSS is highly predictive...

Numerical simulation of the RISOe1-airfoil dynamic stall

Energy Technology Data Exchange (ETDEWEB)

Bertagnolio, F.; Soerensen, N. [Risoe National Lab., Wind Energy and Atmospheric Physics Dept., Roskilde (Denmark)

1997-12-31

In this paper we are concerned with the numerical computation of the dynamic stall that occur in the viscous flowfield over an airfoil. These results are compared to experimental data that were obtained with the new designed RISOe1-airfoil, both for a motionless airfoil and for a pitching motion. Moreover, we present some numerical computations of the plunging and lead-lag motions. We also investigate the possibility of using the pitching motion to simulate the plunging and lead-lag situations. (au)

Editorial: 3Rs tightly intertwined to maintain genome stability

DEFF Research Database (Denmark)

Lisby, Michael; Mortensen, Uffe H.

2017-01-01

, replication of damaged DNA results in stalled replication forks that await DNA damage repair before replication can be resumed. In turn, the repair of most lesions depends on processes involving DNA synthesis. At the same time, the stalled forks may engage in recombination, either as part of a controlled...... repair process or by accident, just because it can, with the risk of producing genome rearrangements and loss of heterozygosity. The set of reviews presented in this thematic issue (https://academic-oup-com.proxy.findit.dtu.dk/femsyr/pages/replication_recombination_and_repair) of FEMSYR has been selected...

HuR and Ago2 Bind the Internal Ribosome Entry Site of Enterovirus 71 and Promote Virus Translation and Replication.

Directory of Open Access Journals (Sweden)

Jing-Yi Lin

Full Text Available EV71 (enterovirus 71 RNA contains an internal ribosomal entry site (IRES that directs cap-independent initiation of translation. IRES-dependent translation requires the host's translation initiation factors and IRES-associated trans-acting factors (ITAFs. We reported recently that mRNA decay factor AUF1 is a negative-acting ITAF that binds IRES stem-loop II. We also reported that the small RNA-processing enzyme Dicer produces at least four small RNAs (vsRNAs from the EV71 IRES. One of these, vsRNA1, derived from IRES stem-loop II, reduces IRES activity and virus replication. Since its mechanism of action is unknown, we hypothesized that it might control association of ITAFs with the IRES. Here, we identified the mRNA stability factor HuR and the RISC subunit Argonaute 2 (Ago2 as two ITAFs that bind stem-loop II. In contrast to AUF1, HuR and Ago2 promote EV71 IRES activity and virus replication. In vitro RNA-binding assays revealed that vsRNA1 can alter association of Ago2, HuR, and AUF1 with stem-loop II. This presents a possible mechanism by which vsRNA1 could control viral translation and replication.

Visualization and measurement of ATP levels in living cells replicating hepatitis C virus genome RNA.

Directory of Open Access Journals (Sweden)

Tomomi Ando

Full Text Available Adenosine 5'-triphosphate (ATP is the primary energy currency of all living organisms and participates in a variety of cellular processes. Although ATP requirements during viral lifecycles have been examined in a number of studies, a method by which ATP production can be monitored in real-time, and by which ATP can be quantified in individual cells and subcellular compartments, is lacking, thereby hindering studies aimed at elucidating the precise mechanisms by which viral replication energized by ATP is controlled. In this study, we investigated the fluctuation and distribution of ATP in cells during RNA replication of the hepatitis C virus (HCV, a member of the Flaviviridae family. We demonstrated that cells involved in viral RNA replication actively consumed ATP, thereby reducing cytoplasmic ATP levels. Subsequently, a method to measure ATP levels at putative subcellular sites of HCV RNA replication in living cells was developed by introducing a recently-established Förster resonance energy transfer (FRET-based ATP indicator, called ATeam, into the NS5A coding region of the HCV replicon. Using this method, we were able to observe the formation of ATP-enriched dot-like structures, which co-localize with non-structural viral proteins, within the cytoplasm of HCV-replicating cells but not in non-replicating cells. The obtained FRET signals allowed us to estimate ATP concentrations within HCV replicating cells as ∼5 mM at possible replicating sites and ∼1 mM at peripheral sites that did not appear to be involved in HCV replication. In contrast, cytoplasmic ATP levels in non-replicating Huh-7 cells were estimated as ∼2 mM. To our knowledge, this is the first study to demonstrate changes in ATP concentration within cells during replication of the HCV genome and increased ATP levels at distinct sites within replicating cells. ATeam may be a powerful tool for the study of energy metabolism during replication of the viral genome.

Histone Modification Associated with Initiation of DNA Replication | Center for Cancer Research

Science.gov (United States)

Before cells are able to divide, they must first duplicate their chromosomes accurately. DNA replication and packaging of DNA into chromosomes by histone proteins need to be coordinated by the cell to ensure proper transmission of genetic and epigenetic information to the next generation. Mammalian DNA replication begins at specific chromosomal sites, called replication

Preliminary Results of the Determination of Inlet-Pressure Distortion Effects on Compressor Stall and Altitude Operating Limits of the J57-P-1 Turbojet Engine

Science.gov (United States)

Wallner, L. E.; Lubick, R. J.; Chelko, L. J.

1955-01-01

During an investigation of the J57-P-1 turbojet engine in the Lewis altitude wind tunnel, effects of inlet-flow distortion on engine stall characteristics and operating limits were determined. In addition to a uniform inlet-flow profile, the inlet-pressure distortions imposed included two radial, two circumferential, and one combined radial-circumferential profile. Data were obtained over a range of compressor speeds at an altitude of 50,000 and a flight Mach number of 0.8; in addition, the high- and low-speed engine operating limits were investigated up to the maximum operable altitude. The effect of changing the compressor bleed position on the stall and operating limits was determined for one of the inlet distortions. The circumferential distortions lowered the compressor stall pressure ratios; this resulted in less fuel-flow margin between steady-state operation and compressor stall. Consequently, the altitude operating Limits with circumferential distortions were reduced compared with the uniform inlet profile. Radial inlet-pressure distortions increased the pressure ratio required for compressor stall over that obtained with uniform inlet flow; this resulted in higher altitude operating limits. Likewise, the stall-limit fuel flows required with the radial inlet-pressure distortions were considerably higher than those obtained with the uniform inlet-pressure profile. A combined radial-circumferential inlet distortion had effects on the engine similar to the circumferential distortion. Bleeding air between the two compressors eliminated the low-speed stall limit and thus permitted higher altitude operation than was possible without compressor bleed.

Visualization by PIV of dynamic stall on a vertical axis wind turbine

NARCIS (Netherlands)

Ferreira, C.J.S.; Kuik, van G.A.M.; Bussel, van G.J.W.; Scarano, F.

2009-01-01

The aerodynamic behavior of a vertical axis wind turbine (VAWT) is analyzed by means of 2D particle image velocimetry (PIV), focusing on the development of dynamic stall at different tip speed ratios. The VAWT has an unsteady aerodynamic behavior due to the variation with the azimuth angle ¿ of the

Genome-wide mapping reveals single-origin chromosome replication in Leishmania, a eukaryotic microbe.

Science.gov (United States)

Marques, Catarina A; Dickens, Nicholas J; Paape, Daniel; Campbell, Samantha J; McCulloch, Richard

2015-10-19

DNA replication initiates on defined genome sites, termed origins. Origin usage appears to follow common rules in the eukaryotic organisms examined to date: all chromosomes are replicated from multiple origins, which display variations in firing efficiency and are selected from a larger pool of potential origins. To ask if these features of DNA replication are true of all eukaryotes, we describe genome-wide origin mapping in the parasite Leishmania. Origin mapping in Leishmania suggests a striking divergence in origin usage relative to characterized eukaryotes, since each chromosome appears to be replicated from a single origin. By comparing two species of Leishmania, we find evidence that such origin singularity is maintained in the face of chromosome fusion or fission events during evolution. Mapping Leishmania origins suggests that all origins fire with equal efficiency, and that the genomic sites occupied by origins differ from related non-origins sites. Finally, we provide evidence that origin location in Leishmania displays striking conservation with Trypanosoma brucei, despite the latter parasite replicating its chromosomes from multiple, variable strength origins. The demonstration of chromosome replication for a single origin in Leishmania, a microbial eukaryote, has implications for the evolution of origin multiplicity and associated controls, and may explain the pervasive aneuploidy that characterizes Leishmania chromosome architecture.

Ethnic Dimensions of Guatemala's Stalled Transition: A Parity-Specific Analysis of Ladino and Indigenous Fertility Regimes.

Science.gov (United States)

Grace, Kathryn; Sweeney, Stuart

2016-02-01

In some contemporary populations, fertility levels appear to plateau, with women maintaining a consistently high level of fertility for a relatively extended period. Because this plateau does not reflect the historical patterns observed in Europe, the focus of most studies on fertility patterns, mechanisms underlying the plateau and the reinstatement of a decline have not been fully explored and are not fully understood. Through the construction of fertility histories of 25,000 women using multiple years of health survey data, we analyze some of the components of stalled fertility as they pertain to Guatemala, the only Central American country to have experienced a stalled fertility decline.

Pyrimidine dimers block simian virus 40 replication forks

International Nuclear Information System (INIS)

Berger, C.A.; Edenberg, H.J.

1986-01-01

UV light produces lesions, predominantly pyrimidine dimers, which inhibit DNA replication in mammalian cells. The mechanism of inhibition is controversial: is synthesis of a daughter strand halted at a lesion while the replication fork moves on and reinitiates downstream, or is fork progression itself blocked for some time at the site of a lesion? We directly addressed this question by using electron microscopy to examine the distances of replication forks from the origin in unirradiated and UV-irradiated simian virus 40 chromosomes. If UV lesions block replication fork progression, the forks should be asymmetrically located in a large fraction of the irradiated molecules; if replication forks move rapidly past lesions, the forks should be symmetrically located. A large fraction of the simian virus 40 replication forks in irradiated molecules were asymmetrically located, demonstrating that UV lesions present at the frequency of pyrimidine dimers block replication forks. As a mechanism for this fork blockage, we propose that polymerization of the leading strand makes a significant contribution to the energetics of fork movement, so any lesion in the template for the leading strand which blocks polymerization should also block fork movement

Associations of soft flooring materials in free stalls with milk yield, clinical mastitis, teat lesions, and removal of dairy cows.

Science.gov (United States)

Ruud, L E; Bøe, K E; Osterås, O

2010-04-01

The objective was to test if there was an association between free-stall base softness and milk yield, incidence of clinical mastitis (CM), teat lesions, and removal of cows. In a questionnaire sent to 1,923 dairy farms presumed to be using free-stall housing, farmers were asked for information regarding housing and stall base; for example, the year of installation and the product name or brand of their mats or mattresses. This information was merged with data for milk yield, CM, teat lesions, and removal of cows extracted from the Norwegian Dairy Herd Recording System for the years after installation of mats or mattresses. After exclusion of invalid contributions, the data set consisted of 29,326 lactations for milk yield distributed over 363 free-stalled herds in Norway. The farms were stratified into 5 categories according to the softness of the stall surface measured as millimeter impact of a sphere with a diameter of 120 mm at 2-kN load: 1=concrete, softness of 0mm; 2=rubber, softness of 1 to 8mm; 3=soft mats, softness of 9 to 16 mm; 4=multilayer mats, softness of 17 to 24 mm; and 5=mattresses, softness over 24 mm. Lactation curves were estimated as modified Wood's lactation curves using test-day data and mixed models with repeated measurements, adjusting for days in milk, parity, and softness of free-stall flooring. Herds on concrete free-stall bases yielded 6,727+/-146 kg of milk from 5 to 305 days in milk. In comparison, herds showed a decrease of 0.3% on rubber, an increase of 2.4% on soft mats, an increase of 4.5% on multilayer mats, and an increase of 3.9% on mattresses. Compared with concrete, the hazard ratio (HR) of CM was less on rubber, multilayer mats, and mattresses [HR=0.89 (0.79-0.99), 0.85 (0.73-0.996), and 0.80 (0.73-0.88), respectively]. Compared with concrete, the HR of teat lesions was less on rubber, soft mats, multilayer mats, and mattresses [HR=0.41 (0.26-0.65), 0.33 (0.24-0.44), 0.12 (0.04-0.38), and 0.47 (0.33-0.67), respectively]. The

Replication stress activates DNA repair synthesis in mitosis

DEFF Research Database (Denmark)

Minocherhomji, Sheroy; Ying, Songmin; Bjerregaard, Victoria A

2015-01-01

Oncogene-induced DNA replication stress has been implicated as a driver of tumorigenesis. Many chromosomal rearrangements characteristic of human cancers originate from specific regions of the genome called common fragile sites (CFSs). CFSs are difficult-to-replicate loci that manifest as gaps...... into mitotic prophase triggers the recruitment of MUS81 to CFSs. The nuclease activity of MUS81 then promotes POLD3-dependent DNA synthesis at CFSs, which serves to minimize chromosome mis-segregation and non-disjunction. We propose that the attempted condensation of incompletely duplicated loci in early...... mitosis serves as the trigger for completion of DNA replication at CFS loci in human cells. Given that this POLD3-dependent mitotic DNA synthesis is enhanced in aneuploid cancer cells that exhibit intrinsically high levels of chromosomal instability (CIN(+)) and replicative stress, we suggest...

Stall Recovery in a Centrifuge-Based Flight Simulator With an Extended Aerodynamic Model

NARCIS (Netherlands)

Ledegang, W.D.; Groen, E.L.

2015-01-01

We investigated the performance of 12 airline pilots in recovering from an asymmetrical stall in a flight simulator featuring an extended aerodynamic model of a transport-category aircraft, and a centrifuge-based motion platform capable of generating enhanced buffet motion and g-cueing. All pilots

DNA breaks early in replication in B cell cancers

Science.gov (United States)

Research by scientists at the NCI has identified a new class of DNA sites in cells that break early in the replication process. They found that these break sites correlate with damage often seen in B cell cancers, such as diffuse large B cell lymphoma.

Critical 23S rRNA interactions for macrolide-dependent ribosome stalling on the ErmCL nascent peptide chain.

Science.gov (United States)

Koch, Miriam; Willi, Jessica; Pradère, Ugo; Hall, Jonathan; Polacek, Norbert

2017-06-20

The nascent peptide exit tunnel has recently been identified as a functional region of ribosomes contributing to translation regulation and co-translational protein folding. Inducible expression of the erm resistance genes depends on ribosome stalling at specific codons of an upstream open reading frame in the presence of an exit tunnel-bound macrolide antibiotic. The molecular basis for this translation arrest is still not fully understood. Here, we used a nucleotide analog interference approach to unravel important functional groups on 23S rRNA residues in the ribosomal exit tunnel for ribosome stalling on the ErmC leader peptide. By replacing single nucleobase functional groups or even single atoms we were able to demonstrate the importance of A2062, A2503 and U2586 for drug-dependent ribosome stalling. Our data show that the universally conserved A2062 and A2503 are capable of forming a non-Watson-Crick base pair that is critical for sensing and transmitting the stalling signal from the exit tunnel back to the peptidyl transferase center of the ribosome. The nucleobases of A2062, A2503 as well as U2586 do not contribute significantly to the overall mechanism of protein biosynthesis, yet their elaborate role for co-translational monitoring of nascent peptide chains inside the exit tunnel can explain their evolutionary conservation. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

Microbiological quality of air in free-range and box-stall stable horse keeping systems.

Science.gov (United States)

Wolny-Koładka, Katarzyna

2018-04-07

The aim of this study was to assess the microbiological quality of air in three horse riding centers differing in the horse keeping systems. The air samples were collected in one facility with free-range horse keeping system and two with box stalls of different sizes. The samples were collected over a period of 3 years (2015-2017), four times per year (spring, summer, autumn, winter) to assess the effect of seasonal changes. The prevalence of aerobic mesophilic bacteria, mold fungi, actinomycetes, Staphylococcus spp., and Escherichia coli was determined by the air collision method on Petri dishes with appropriate microbiological media. At the same time, air temperature, relative humidity, and particulate matter concentration (PM 10 , PM 2.5 ) were measured. It was found that the horse keeping system affects the occurrence of the examined airborne microorganisms. Over the 3-year period of study, higher temperature and humidity, as well as particulate matter concentration-which notoriously exceeded limit values-were observed in the facilities with the box-stall system. The air sampled from the largest horse riding center, with the largest number of horses and the box-stall system of horse keeping, was also characterized by the heaviest microbiological contamination. Among others, bacteria from the following genera: Staphylococcus spp., Streptococcus spp., Bacillus spp., and E. coli and fungi from the genera Aspergillus, Fusarium, Mucor, Rhizopus, Penicillium, Trichothecium, Cladosporium, and Alternaria were identified in the analyzed samples.

Structural properties of replication origins in yeast DNA sequences

International Nuclear Information System (INIS)

Cao Xiaoqin; Zeng Jia; Yan Hong

2008-01-01

Sequence-dependent DNA flexibility is an important structural property originating from the DNA 3D structure. In this paper, we investigate the DNA flexibility of the budding yeast (S. Cerevisiae) replication origins on a genome-wide scale using flexibility parameters from two different models, the trinucleotide and the tetranucleotide models. Based on analyzing average flexibility profiles of 270 replication origins, we find that yeast replication origins are significantly rigid compared with their surrounding genomic regions. To further understand the highly distinctive property of replication origins, we compare the flexibility patterns between yeast replication origins and promoters, and find that they both contain significantly rigid DNAs. Our results suggest that DNA flexibility is an important factor that helps proteins recognize and bind the target sites in order to initiate DNA replication. Inspired by the role of the rigid region in promoters, we speculate that the rigid replication origins may facilitate binding of proteins, including the origin recognition complex (ORC), Cdc6, Cdt1 and the MCM2-7 complex

Why Do Promising Therapies Stall in Development and How Can We Move Them Forward?

Science.gov (United States)

Wegner, Craig D; Goodwin, Andrew; Cook, Jon C; Allamneni, Krishna; Sohn, Jane; McVean, Maralee

There are many reasons that molecules fail to progress to market and various principles of risk-benefit decisions that can help drive the molecule through development. This symposium included discussions on global strategies involved in pushing promising molecules to market, what to do when a molecule stalls in its progress to market, and options for rescuing the molecule and pushing it forward again. Innovative partnerships that bring stalled drugs back into clinical development were also addressed. A regulatory perspective on common reasons for a molecule to fail in its forward progress was presented. In addition, situations arise when a third-party advisory committee can provide input to help overcome issues identified by a regulatory agency. Using examples from the private and public domain, presentations centered on how to repurpose a molecule and when more science is needed.

Cooling systems of the resting area in free stall dairy barn

Science.gov (United States)

Calegari, F.; Calamari, L.; Frazzi, E.

2016-04-01

A study during the summer season evaluated the effect of different cooling systems on behavioral and productive responses of Italian Friesian dairy cows kept in an experimental-free stall barn located in the Po Valley in Italy. The study involved 30 lactating dairy cows subdivided into two groups kept in two pens with external hard court paddock in each free stall. The same cooling system was applied in the feeding area in both pens. A different cooling system in the resting area was applied to the two pens: in the pen SW, the resting area was equipped with fans and misters; in the other, there was simple ventilation (SV). Breathing rate, rectal temperature, milk yield, and milk characteristics (fat, protein, and somatic cell count) were measured. Behavioral activities (standing and lying cows in the different areas, as well as the animals in the feed bunk) were recorded. Mild to moderate heat waves during the trial were observed. On average, the breathing rate was numerically greater in SV compared with SW cows (60.2 and 55.8 breath/min, respectively), and mean rectal temperature remained below 39 °C in both groups during the trial (on average 38.7 and 38.8 °C in SV and SW, respectively. During the hotter periods of the trial, the time spent lying indoor in the free stall was greater in SW (11.8 h/day) than SV (10.7 h/day). Conversely, the time spent standing indoor without feeding was greater in SV (4.3 h/day) than SW (3.8 h/day). Milk yield was slightly better maintained during hotter period in SW compared with SV and somatic cell count was also slightly greater in the former. In conclusion, the adoption of the cooling system by means of evaporative cooling also in the resting area reduces the alteration of time budget caused by heat stress.

P-body proteins regulate transcriptional rewiring to promote DNA replication stress resistance.

Science.gov (United States)

Loll-Krippleber, Raphael; Brown, Grant W

2017-09-15

mRNA-processing (P-) bodies are cytoplasmic granules that form in eukaryotic cells in response to numerous stresses to serve as sites of degradation and storage of mRNAs. Functional P-bodies are critical for the DNA replication stress response in yeast, yet the repertoire of P-body targets and the mechanisms by which P-bodies promote replication stress resistance are unknown. In this study we identify the complete complement of mRNA targets of P-bodies during replication stress induced by hydroxyurea treatment. The key P-body protein Lsm1 controls the abundance of HHT1, ACF4, ARL3, TMA16, RRS1 and YOX1 mRNAs to prevent their toxic accumulation during replication stress. Accumulation of YOX1 mRNA causes aberrant downregulation of a network of genes critical for DNA replication stress resistance and leads to toxic acetaldehyde accumulation. Our data reveal the scope and the targets of regulation by P-body proteins during the DNA replication stress response.P-bodies form in response to stress and act as sites of mRNA storage and degradation. Here the authors identify the mRNA targets of P-bodies during DNA replication stress, and show that P-body proteins act to prevent toxic accumulation of these target transcripts.

Analysis of the Unsteady Flow Field in a Centrifugal Compressor from Peak Efficiency to Near Stall with Full-Annulus Simulations

Directory of Open Access Journals (Sweden)

Yannick Bousquet

2014-01-01

Full Text Available This study concerns a 2.5 pressure ratio centrifugal compressor stage consisting of a splittered unshrouded impeller and a vaned diffuser. The aim of this paper is to investigate the modifications of the flow structure when the operating point moves from peak efficiency to near stall. The investigations are based on the results of unsteady three-dimensional simulations, in a calculation domain comprising all the blade. A detailed analysis is given in the impeller inducer and in the vaned diffuser entry region through time-averaged and unsteady flow field. In the impeller inducer, this study demonstrates that the mass flow reduction from peak efficiency to near stall leads to intensification of the secondary flow effects. The low momentum fluid accumulated near the shroud interacts with the main flow through a shear layer zone. At near stall condition, the interface between the two flow structures becomes unstable leading to vortices development. In the diffuser entry region, by reducing the mass flow, the high incidence angle from the impeller exit induces a separation on the diffuser vane suction side. At near stall operating point, vorticity from the separation is shed into vortex cores which are periodically formed and convected downstream along the suction side.

Activation of a yeast replication origin near a double-stranded DNA break.

Science.gov (United States)

Raghuraman, M K; Brewer, B J; Fangman, W L

1994-03-01

Irradiation in the G1 phase of the cell cycle delays the onset of DNA synthesis and transiently inhibits the activation of replication origins in mammalian cells. It has been suggested that this inhibition is the result of the loss of torsional tension in the DNA after it has been damaged. Because irradiation causes DNA damage at an undefined number of nonspecific sites in the genome, it is not known how cells respond to limited DNA damage, and how replication origins in the immediate vicinity of a damage site would behave. Using the sequence-specific HO endonuclease, we have created a defined double-stranded DNA break in a centromeric plasmid in G1-arrested cells of the yeast Saccharomyces cerevisiae. We show that replication does initiate at the origin on the cut plasmid, and that the plasmid replicates early in the S phase after linearization in vivo. These observations suggest that relaxation of a supercoiled DNA domain in yeast need not inactivate replication origins within that domain. Furthermore, these observations rule out the possibility that the late replication context associated with chromosomal termini is a consequence of DNA ends.

Recombination at DNA replication fork barriers is not universal and is differentially regulated by Swi1.

Science.gov (United States)

Pryce, David W; Ramayah, Soshila; Jaendling, Alessa; McFarlane, Ramsay J

2009-03-24

DNA replication stress has been implicated in the etiology of genetic diseases, including cancers. It has been proposed that genomic sites that inhibit or slow DNA replication fork progression possess recombination hotspot activity and can form potential fragile sites. Here we used the fission yeast, Schizosaccharomyces pombe, to demonstrate that hotspot activity is not a universal feature of replication fork barriers (RFBs), and we propose that most sites within the genome that form RFBs do not have recombination hotspot activity under nonstressed conditions. We further demonstrate that Swi1, the TIMELESS homologue, differentially controls the recombination potential of RFBs, switching between being a suppressor and an activator of recombination in a site-specific fashion.

ATAD2 is an epigenetic reader of newly synthesized histone marks during DNA replication.

Science.gov (United States)

Koo, Seong Joo; Fernández-Montalván, Amaury E; Badock, Volker; Ott, Christopher J; Holton, Simon J; von Ahsen, Oliver; Toedling, Joern; Vittori, Sarah; Bradner, James E; Gorjánácz, Mátyás

2016-10-25

ATAD2 (ATPase family AAA domain-containing protein 2) is a chromatin regulator harboring an AAA+ ATPase domain and a bromodomain, previously proposed to function as an oncogenic transcription co-factor. Here we suggest that ATAD2 is also required for DNA replication. ATAD2 is co-expressed with genes involved in DNA replication in various cancer types and predominantly expressed in S phase cells where it localized on nascent chromatin (replication sites). Our extensive biochemical and cellular analyses revealed that ATAD2 is recruited to replication sites through a direct interaction with di-acetylated histone H4 at K5 and K12, indicative of newly synthesized histones during replication-coupled chromatin reassembly. Similar to ATAD2-depletion, ectopic expression of ATAD2 mutants that are deficient in binding to these di-acetylation marks resulted in reduced DNA replication and impaired loading of PCNA onto chromatin, suggesting relevance of ATAD2 in DNA replication. Taken together, our data show a novel function of ATAD2 in cancer and for the first time identify a reader of newly synthesized histone di-acetylation-marks during replication.

Unsteady performance of a cavitating hydrofoil in stall conditions. Shissoku jotai ni okeru yokukei no hiteijo tokusei

Energy Technology Data Exchange (ETDEWEB)

Ogata, H. (Tohoku University, Sendai (Japan)); Ito, Y. (Hachinoe Institutea of Technology, Aomori (Japan)); Oba, R. (Tohoku University, Sendai (Japan). Institute of Fluid Science); Sunayama, Y.; Abe, J. (Suzuki Co. Ltd., Shizuoka (Japan))

1991-11-25

To elucidate the unsteady performance of cavitating hydrofoils in a stall condition, this paper describes a survey on unsteady conditions without cavitations and stall conditions as to their characteristics from a cavitation to a supercavitation, lift and drag. Flows with cavitations were also analyzed. As a result of comparing also data for the subcavitation regions, it was found that a large-scale vortex generation on the hydrofoil back-pressure plane in near stall condition has a close relation with the changes in lifts and drags or the cavitation breakdown. The experiment used a testing water tank of circulation flow type having a rectangular measuring cross section (70 mm in width and 190 mm in height), and the hydrofoil specimens of two-dimensional symmetric type with a chord length of 70 mm and an aspect ratio of 1.0. The test condition used a cavitation coefficient of 0.18-6.33 (from a supercavitation to non-cavitation). A numerical analysis proved that the power spectra around the hydrofoils having no cavitations agreed with the experimental results, and verified the reasonability of the application. 18 refs., 8 figs., 2 tabs.

Replisome speed determines the efficiency of the Tus−Ter replication termination barrier

KAUST Repository

Elshenawy, Mohamed; Jergic, Slobodan; Xu, Zhi Qiang; Sobhy, Mohamed Abdelmaboud; Takahashi, Masateru; Oakley, Aaron J.; Dixon, Nicholas E.; Hamdan, Samir

2015-01-01

In all domains of life, DNA synthesis occurs bidirectionally from replication origins. Despite variable rates of replication fork progression, fork convergence often occurs at specific sites. Escherichia coli sets a 'replication fork trap' that allows the first arriving fork to enter but not to leave the terminus region. The trap is set by oppositely oriented Tus-bound Ter sites that block forks on approach from only one direction. However, the efficiency of fork blockage by Tus-Ter does not exceed 50% in vivo despite its apparent ability to almost permanently arrest replication forks in vitro. Here we use data from single-molecule DNA replication assays and structural studies to show that both polarity and fork-arrest efficiency are determined by a competition between rates of Tus displacement and rearrangement of Tus-Ter interactions that leads to blockage of slower moving replisomes by two distinct mechanisms. To our knowledge this is the first example where intrinsic differences in rates of individual replisomes have different biological outcomes. ©2015 Macmillan Publishers Limited. All rights reserved.

Replisome speed determines the efficiency of the Tus−Ter replication termination barrier

KAUST Repository

Elshenawy, Mohamed

2015-08-31

In all domains of life, DNA synthesis occurs bidirectionally from replication origins. Despite variable rates of replication fork progression, fork convergence often occurs at specific sites. Escherichia coli sets a \\'replication fork trap\\' that allows the first arriving fork to enter but not to leave the terminus region. The trap is set by oppositely oriented Tus-bound Ter sites that block forks on approach from only one direction. However, the efficiency of fork blockage by Tus-Ter does not exceed 50% in vivo despite its apparent ability to almost permanently arrest replication forks in vitro. Here we use data from single-molecule DNA replication assays and structural studies to show that both polarity and fork-arrest efficiency are determined by a competition between rates of Tus displacement and rearrangement of Tus-Ter interactions that leads to blockage of slower moving replisomes by two distinct mechanisms. To our knowledge this is the first example where intrinsic differences in rates of individual replisomes have different biological outcomes. ©2015 Macmillan Publishers Limited. All rights reserved.

Phosphorylation of NS5A Serine-235 is essential to hepatitis C virus RNA replication and normal replication compartment formation

Energy Technology Data Exchange (ETDEWEB)

Eyre, Nicholas S., E-mail: nicholas.eyre@adelaide.edu.au [School of Biological Sciences and Research Centre for Infectious Diseases, University of Adelaide, Adelaide (Australia); Centre for Cancer Biology, SA Pathology, Adelaide (Australia); Hampton-Smith, Rachel J.; Aloia, Amanda L. [School of Biological Sciences and Research Centre for Infectious Diseases, University of Adelaide, Adelaide (Australia); Centre for Cancer Biology, SA Pathology, Adelaide (Australia); Eddes, James S. [Adelaide Proteomics Centre, School of Biological Sciences, University of Adelaide, Adelaide (Australia); Simpson, Kaylene J. [Victorian Centre for Functional Genomics, Peter MacCallum Cancer Centre, East Melbourne (Australia); The Sir Peter MacCallum Department of Oncology, University of Melbourne, Parkville (Australia); Hoffmann, Peter [Adelaide Proteomics Centre, School of Biological Sciences, University of Adelaide, Adelaide (Australia); Institute for Photonics and Advanced Sensing (IPAS), University of Adelaide, Adelaide (Australia); Beard, Michael R. [School of Biological Sciences and Research Centre for Infectious Diseases, University of Adelaide, Adelaide (Australia); Centre for Cancer Biology, SA Pathology, Adelaide (Australia)

2016-04-15

Hepatitis C virus (HCV) NS5A protein is essential for HCV RNA replication and virus assembly. Here we report the identification of NS5A phosphorylation sites Ser-222, Ser-235 and Thr-348 during an infectious HCV replication cycle and demonstrate that Ser-235 phosphorylation is essential for HCV RNA replication. Confocal microscopy revealed that both phosphoablatant (S235A) and phosphomimetic (S235D) mutants redistribute NS5A to large juxta-nuclear foci that display altered colocalization with known replication complex components. Using electron microscopy (EM) we found that S235D alters virus-induced membrane rearrangements while EM using ‘APEX2’-tagged viruses demonstrated S235D-mediated enrichment of NS5A in irregular membranous foci. Finally, using a customized siRNA screen of candidate NS5A kinases and subsequent analysis using a phospho-specific antibody, we show that phosphatidylinositol-4 kinase III alpha (PI4KIIIα) is important for Ser-235 phosphorylation. We conclude that Ser-235 phosphorylation of NS5A is essential for HCV RNA replication and normal replication complex formation and is regulated by PI4KIIIα. - Highlights: • NS5A residues Ser-222, Ser-235 and Thr-348 are phosphorylated during HCV infection. • Phosphorylation of Ser-235 is essential to HCV RNA replication. • Mutation of Ser-235 alters replication compartment localization and morphology. • Phosphatidylinositol-4 kinase III alpha is important for Ser-235 phosphorylation.

Wave energy plants: Control strategies for avoiding the stalling behaviour in the Wells turbine

Energy Technology Data Exchange (ETDEWEB)

Amundarain, Modesto; Alberdi, Mikel; Garrido, Aitor J.; Garrido, Izaskun; Maseda, Javier [Dept. of Automatic Control and Systems Engineering, EUITI Bilbao, University of the Basque Country, Plaza de la Casilla 3, 48012 Bilbao (Spain)

2010-12-15

This study analyzes the problem of the stalling behaviour in Wells turbines, one of the most widely used turbines in wave energy plants. For this purpose two different control strategies are presented and compared. In the first one, a rotational speed control system is employed to appropriately adapt the speed of the double-fed induction generator coupling to the turbine, according to the pressure drop entry. In the second control strategy, an airflow control regulates the power generated by the turbine generator module by means of the modulation valve avoiding the stalling behaviour. It is demonstrated that the proposed rotational speed control design adequately matches the desired relationship between the slip of the double-fed induction generator and the pressure drop input, whilst the valve control using a traditional PID controller successfully governs the flow that modulates the pressure drop across the turbine. (author)

A Detailed Study of the Rotational Augmentation and Dynamic Stall Phenomena for Wind Turbines

DEFF Research Database (Denmark)

Guntur, Srinivas

This thesis presents investigations into the aerodynamics of wind turbine rotors, with a focus on the in-board sections of the rotor. Two important aerodynamic phenomena that have challenged scientists over nearly the last half a century are the so-called rotational augmentation and dynamic stall....... This thesis presents an investigation into these two phenomena, using data from the MEXICO and the NREL UAE Phase VI experiments, as well as data obtained from full rotor CFD computations carried out using the in-house flow solver Ellipsys3D. The experimental data, CFD data and that from some of the existing...... on wind turbine blades using the N-sequence data of the NREL UAE Phase VI experiment. The experimental data is compared with the results from unsteady Delayed Detached Eddy Simulations (DDES). The same conditions are also modelled using a Beddoes-Leishman type dynamic stall model by Hansen et al. (2004...

Computer modeling of the stalled flow of a rotating cylinder and the reverse magnus effect

Science.gov (United States)

Belotserkovskii, S. M.; Kotovskii, V. N.; Nisht, M. I.; Fedorov, R. M.

1985-02-01

Unsteady stalled flow around a rotating cylinder is investigated in a numerical experiment. Attention is mostly given to the reverse Magnus effect which was discovered in tube experiments at some critical rotational speed of the cylinder.

Plum Pox Virus 6K1 Protein Is Required for Viral Replication and Targets the Viral Replication Complex at the Early Stage of Infection.

Science.gov (United States)

Cui, Hongguang; Wang, Aiming

2016-05-15

The potyviral RNA genome encodes two polyproteins that are proteolytically processed by three viral protease domains into 11 mature proteins. Extensive molecular studies have identified functions for the majority of the viral proteins. For example, 6K2, one of the two smallest potyviral proteins, is an integral membrane protein and induces the endoplasmic reticulum (ER)-originated replication vesicles that target the chloroplast for robust viral replication. However, the functional role of 6K1, the other smallest protein, remains uncharacterized. In this study, we developed a series of recombinant full-length viral cDNA clones derived from a Canadian Plum pox virus (PPV) isolate. We found that deletion of any of the short motifs of 6K1 (each of which ranged from 5 to 13 amino acids), most of the 6K1 sequence (but with the conserved sequence of the cleavage sites being retained), or all of the 6K1 sequence in the PPV infectious clone abolished viral replication. The trans expression of 6K1 or the cis expression of a dislocated 6K1 failed to rescue the loss-of-replication phenotype, suggesting the temporal and spatial requirement of 6K1 for viral replication. Disruption of the N- or C-terminal cleavage site of 6K1, which prevented the release of 6K1 from the polyprotein, either partially or completely inhibited viral replication, suggesting the functional importance of the mature 6K1. We further found that green fluorescent protein-tagged 6K1 formed punctate inclusions at the viral early infection stage and colocalized with chloroplast-bound viral replicase elements 6K2 and NIb. Taken together, our results suggest that 6K1 is required for viral replication and is an important viral element of the viral replication complex at the early infection stage. Potyviruses account for more than 30% of known plant viruses and consist of many agriculturally important viruses. The genomes of potyviruses encode two polyproteins that are proteolytically processed into 11 mature

GC-rich DNA elements enable replication origin activity in the methylotrophic yeast Pichia pastoris.

Science.gov (United States)

Liachko, Ivan; Youngblood, Rachel A; Tsui, Kyle; Bubb, Kerry L; Queitsch, Christine; Raghuraman, M K; Nislow, Corey; Brewer, Bonita J; Dunham, Maitreya J

2014-03-01

The well-studied DNA replication origins of the model budding and fission yeasts are A/T-rich elements. However, unlike their yeast counterparts, both plant and metazoan origins are G/C-rich and are associated with transcription start sites. Here we show that an industrially important methylotrophic budding yeast, Pichia pastoris, simultaneously employs at least two types of replication origins--a G/C-rich type associated with transcription start sites and an A/T-rich type more reminiscent of typical budding and fission yeast origins. We used a suite of massively parallel sequencing tools to map and dissect P. pastoris origins comprehensively, to measure their replication dynamics, and to assay the global positioning of nucleosomes across the genome. Our results suggest that some functional overlap exists between promoter sequences and G/C-rich replication origins in P. pastoris and imply an evolutionary bifurcation of the modes of replication initiation.

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

Directory of Open Access Journals (Sweden)

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.

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

Association between stall surface and some animal welfare measurements in freestall dairy herds using recycled manure solids for bedding.

Science.gov (United States)

Husfeldt, A W; Endres, M I

2012-10-01

The objective of this cross-sectional study was to investigate the association between stall surface and some animal welfare measurements in upper Midwest US dairy operations using recycled manure solids as bedding material. The study included 34 dairy operations with herd sizes ranging from 130 to 3,700 lactating cows. Forty-five percent of the herds had mattresses and 55% had deep-bedded stalls. Farms were visited once between July and October 2009. At the time of visit, at least 50% of the cows in each lactating pen were scored for locomotion, hygiene, and hock lesions. On-farm herd records were collected for the entire year and used to investigate mortality, culling, milk production, and mastitis incidence. Stall surface was associated with lameness and hock lesion prevalence. Lameness prevalence (locomotion score ≥ 3 on a 1 to 5 scale) was lower in deep-bedded freestalls (14.4%) than freestalls with mattresses (19.8%). Severe lameness prevalence (locomotion score ≥ 4) was also lower for cows housed in deep-bedded freestalls (3.6%) than for cows housed in freestalls with mattresses (5.9%). In addition, the prevalence of hock lesions (hock lesion scores ≥ 2 on a 1 to 3 scale, with 1=no lesion, 2=hair loss or mild lesion, and 3=swelling or severe lesion) and severe hock lesions (hock lesion score=3) was lower in herds with deep-bedded freestalls (49.4%; 6.4%) than in herds with mattresses (67.3%; 13.2%). Herd turnover rates were not associated with stall surface; however, the percentage of removals due to voluntary (low milk production, disposition, and dairy) and involuntary (death, illness, injury, and reproductive) reasons was different between deep-bedded and mattress-based freestalls. Voluntary removals averaged 16% of all herd removals in deep-bedded herds, whereas in mattress herds, these removals were 8%. Other welfare measurements such as cow hygiene, mortality rate, mastitis incidence, and milk production were not associated with stall surface

Experimental study of dynamic stall on Darrieus wind turbine blades

Science.gov (United States)

Brochier, G.; Fraunie, P.; Beguier, C.; Paraschivoiu, I.

1985-12-01

An experimental study of periodic vortex phenomena was performed on a model of a two straight-bladed Darrieus wind turbine under controlled-rotation conditions in the IMST water tunnel. The main focus of interest was the tip-speed ratios at which dynamic stall appears. Observations of this phenomenon from dye emission and the formation of hydrogen bubbles were made in the form of photographs, film and video recordings. Velocity measurements were obtained using the Laser-Doppler Velocimeter and components of velocity fluctuations could be determined quantitatively.

The Relationships between Selection and Processing Food with Escherichia coli Contaminant on Food Stall Serving

Directory of Open Access Journals (Sweden)

Tris Eryando

2014-04-01

Full Text Available Escherichia coli in food stalls surrounding the X Campuss in Depok, year 2012. The research conducted to examine food safety, which were served in surrounding the campus X in Depok. Escherichia coli (E. coli existence was used to indicate the quality of hygiene and sanitation of the food that was served. Using the cross sectional method, the research examined the persons who served the food to be sold in the food stalls in the campus. There were 173 food servers chosen as the respondents from 10 different food stalls around the university. The existence of E. coli examined in the microbiology laboratory in the Faculty of Public Health. Using the most probable number (MPN method found that 59.54% of the food served in the campus were contaminated E. coli. Factors affecting the existence of E. coli were the raw materials (vegetables treated and the length of cooking of the materials (rice/beens. The improper treatment such as washing with no running water or even unwashed vegetables had 5 times risk of the E. coli contamination. Cooking less than 15 minutes was also more risky than cooking more than 15 minutes. As a result, this is very important to find a method to improve knowledge and to increase practical skills in food safety. Furthermore, in this research area may give contribution to avoid E. coli contamination which will prevent unnecessary illness among students in the campus.

Reliability analysis of a replication with limited number of journaling files

International Nuclear Information System (INIS)

Kimura, Mitsutaka; Imaizumi, Mitsuhiro; Nakagawa, Toshio

2013-01-01

Recently, replication mechanisms using journaling files have been widely used for the server systems. We have already discussed the model of asynchronous replication system using journaling files [8]. This paper formulates a stochastic model of a server system with replication considering the number of transmitting journaling files. The server updates the storage database and transmits the journaling file when a client requests the data update. The server transmits the database content to a backup site either at a constant time or after a constant number of transmitting journaling files. We derive the expected number of the replication and of transmitting journaling files. Further, we calculate the expected cost and discuss optimal replication interval to minimize it. Finally, numerical examples are given

Intragenic origins due to short G1 phases underlie oncogene-induced DNA replication stress.

Science.gov (United States)

Macheret, Morgane; Halazonetis, Thanos D

2018-03-01

Oncogene-induced DNA replication stress contributes critically to the genomic instability that is present in cancer. However, elucidating how oncogenes deregulate DNA replication has been impeded by difficulty in mapping replication initiation sites on the human genome. Here, using a sensitive assay to monitor nascent DNA synthesis in early S phase, we identified thousands of replication initiation sites in cells before and after induction of the oncogenes CCNE1 and MYC. Remarkably, both oncogenes induced firing of a novel set of DNA replication origins that mapped within highly transcribed genes. These ectopic origins were normally suppressed by transcription during G1, but precocious entry into S phase, before all genic regions had been transcribed, allowed firing of origins within genes in cells with activated oncogenes. Forks from oncogene-induced origins were prone to collapse, as a result of conflicts between replication and transcription, and were associated with DNA double-stranded break formation and chromosomal rearrangement breakpoints both in our experimental system and in a large cohort of human cancers. Thus, firing of intragenic origins caused by premature S phase entry represents a mechanism of oncogene-induced DNA replication stress that is relevant for genomic instability in human cancer.

The stress granule component TIA-1 binds tick-borne encephalitis virus RNA and is recruited to perinuclear sites of viral replication to inhibit viral translation.

Science.gov (United States)

Albornoz, Amelina; Carletti, Tea; Corazza, Gianmarco; Marcello, Alessandro

2014-06-01

Flaviviruses are a major cause of disease in humans and animals worldwide. Tick-borne encephalitis virus (TBEV) is the most important arthropod-borne flavivirus endemic in Europe and is the etiological agent of tick-borne encephalitis, a potentially fatal infection of the central nervous system. However, the contributions of host proteins during TBEV infection are poorly understood. In this work, we investigate the cellular protein TIA-1 and its cognate factor TIAR, which are stress-induced RNA-binding proteins involved in the repression of initiation of translation of cellular mRNAs and in the formation of stress granules. We show that TIA-1 and TIAR interact with viral RNA in TBEV-infected cells. During TBEV infection, cytoplasmic TIA-1 and TIAR are recruited at sites of viral replication with concomitant depletion from stress granules. This effect is specific, since G3BP1, another component of these cytoplasmic structures, remains localized to stress granules. Moreover, heat shock induction of stress granules containing TIA-1, but not G3BP1, is inhibited in TBEV-infected cells. Infection of cells depleted of TIA-1 or TIAR by small interfering RNA (siRNA) or TIA-1(-/-) mouse fibroblasts, leads to a significant increase in TBEV extracellular infectivity. Interestingly, TIAR(-/-) fibroblasts show the opposite effect on TBEV infection, and this phenotype appears to be related to an excess of TIA-1 in these cells. Taking advantage of a TBE-luciferase replicon system, we also observed increased luciferase activity in TIA-1(-/-) mouse fibroblasts at early time points, consistent with TIA-1-mediated inhibition at the level of the first round of viral translation. These results indicate that, in response to TBEV infection, TIA-1 is recruited to sites of virus replication to bind TBEV RNA and modulate viral translation independently of stress granule (SG) formation. This study (i) extends previous work that showed TIA-1/TIAR recruitment at sites of flavivirus replication

An active site aromatic triad in Escherichia coli DNA Pol IV coordinates cell survival and mutagenesis in different DNA damaging agents.

Directory of Open Access Journals (Sweden)

Ryan W Benson

Full Text Available DinB (DNA Pol IV is a translesion (TLS DNA polymerase, which inserts a nucleotide opposite an otherwise replication-stalling N(2-dG lesion in vitro, and confers resistance to nitrofurazone (NFZ, a compound that forms these lesions in vivo. DinB is also known to be part of the cellular response to alkylation DNA damage. Yet it is not known if DinB active site residues, in addition to aminoacids involved in DNA synthesis, are critical in alkylation lesion bypass. It is also unclear which active site aminoacids, if any, might modulate DinB's bypass fidelity of distinct lesions. Here we report that along with the classical catalytic residues, an active site "aromatic triad", namely residues F12, F13, and Y79, is critical for cell survival in the presence of the alkylating agent methyl methanesulfonate (MMS. Strains expressing dinB alleles with single point mutations in the aromatic triad survive poorly in MMS. Remarkably, these strains show fewer MMS- than NFZ-induced mutants, suggesting that the aromatic triad, in addition to its role in TLS, modulates DinB's accuracy in bypassing distinct lesions. The high bypass fidelity of prevalent alkylation lesions is evident even when the DinB active site performs error-prone NFZ-induced lesion bypass. The analyses carried out with the active site aromatic triad suggest that the DinB active site residues are poised to proficiently bypass distinctive DNA lesions, yet they are also malleable so that the accuracy of the bypass is lesion-dependent.

Direct numerical simulation of a NACA0012 in full stall

International Nuclear Information System (INIS)

Rodríguez, I.; Lehmkuhl, O.; Borrell, R.; Oliva, A.

2013-01-01

Highlights: • Coherent structures at transitional and supercritical wake modes are presented. • Vortex shedding is detected in both wake modes. • KH instabilities and vortex shedding frequencies are identified. • Low-frequency flapping of the shear-layer is also detected after stall. • Local pressure distribution at both AOA is coherent with experimental observations. -- Abstract: This work aims at investigating the mechanisms of separation and the transition to turbulence in the separated shear-layer of aerodynamic profiles, while at the same time to gain insight into coherent structures formed in the separated zone at low-to-moderate Reynolds numbers. To do this, direct numerical simulations of the flow past a NACA0012 airfoil at Reynolds numbers Re = 50,000 (based on the free-stream velocity and the airfoil chord) and angles of attack AOA = 9.25° and AOA = 12° have been carried out. At low-to-moderate Reynolds numbers, NACA0012 exhibits a combination of leading-edge/trailing-edge stall which causes the massive separation of the flow on the suction side of the airfoil. The initially laminar shear layer undergoes transition to turbulence and vortices formed are shed forming a von Kármán like vortex street in the airfoil wake. The main characteristics of this flow together with its main features, including power spectra of a set of selected monitoring probes at different positions on the suction side and in the wake of the airfoil are provided and discussed in detail

GC-rich DNA elements enable replication origin activity in the methylotrophic yeast Pichia pastoris.

Directory of Open Access Journals (Sweden)

Ivan Liachko

2014-03-01

Full Text Available The well-studied DNA replication origins of the model budding and fission yeasts are A/T-rich elements. However, unlike their yeast counterparts, both plant and metazoan origins are G/C-rich and are associated with transcription start sites. Here we show that an industrially important methylotrophic budding yeast, Pichia pastoris, simultaneously employs at least two types of replication origins--a G/C-rich type associated with transcription start sites and an A/T-rich type more reminiscent of typical budding and fission yeast origins. We used a suite of massively parallel sequencing tools to map and dissect P. pastoris origins comprehensively, to measure their replication dynamics, and to assay the global positioning of nucleosomes across the genome. Our results suggest that some functional overlap exists between promoter sequences and G/C-rich replication origins in P. pastoris and imply an evolutionary bifurcation of the modes of replication initiation.

Avaliação de diferentes materiais para recobrimento de camas em baias de galpão modelo free-stall

OpenAIRE

Cecchin, Daiane; Campos, Alessandro T.; Pires, Maria de F. A.; Lima, Renato R. de; Yanagi Junior, Tadayuki; Souza, Myriam C. M.

2014-01-01

Objetivou-se, com o presente trabalho, comparar o uso de cama de colchão de borracha e de areia no recobrimento da superfície de baias para confinamento tipo free-stall. Foram monitoradas 18 vacas holandesas confinadas em galpão modelo free-stall cujas baias foram recobertas com camas de areia e colchão de borracha (tratamentos). O delineamento experimental foi em blocos casualizados. Os parâmetros comportamentais estudados foram os tempos despendidos nas atividades: deitada em ócio e deitada...

Compartmental HBV evolution and replication in liver and extrahepatic sites after nucleos/tide analogue therapy in chronic hepatitis B carriers.

Science.gov (United States)

Gao, Shan; Duan, Zhong-Ping; Chen, Yu; van der Meer, Frank; Lee, Samuel S; Osiowy, Carla; van Marle, Guido; Coffin, Carla S

2017-09-01

Hepatitis B virus (HBV) variants are associated with nucleos/tide analogue (NA) response and liver disease but it is unknown whether NA influences extrahepatic HBV persistence. To investigate HBV replication and genetic evolution in hepatic and extrahepatic sites of chronic hepatitis B (CHB) before and after NA therapy. A total of 13 paired plasma, peripheral blood mononuclear cells (PBMC), were collected from chronic HBV carriers at baseline and after a median 53 weeks NA therapy as well as liver biopsy (N=7 baseline, N=5 follow-up). HBV covalently closed circular DNA (cccDNA) and messenger (m) RNA in liver and PBMC were analyzed. HBV polymerase (P)/surface (S), basal core promoter (BCP)/pre-core (PC)/C gene clonal sequencing was done in plasma, peripheral blood mononuclear cells (PBMC), and liver. Compare to baseline, at ∼53 weeks follow-up, there was no significant change in HBV cccDNA levels in liver (0.2-0.08 copies/hepatocyte, p>0.05) or in PBMC 0.003-0.02 copies/PBMC, p>0.05), and HBV mRNA remained detectable in both sites. At baseline, BCP variants were higher in PBMC vs. liver and plasma. After therapy, drug resistant (DR) and immune escape (IE) variants increased in liver but IE and PC variants were more frequent in PBMC. HBV P/S diversity was significantly higher in PBMC compared to plasma. Continuous HBV replication occurs in liver and PBMC and shows compartmentalized evolution under selective pressure of potent NA therapy. Copyright © 2017 Elsevier B.V. All rights reserved.

Exploratory study of the effects of wing-leading-edge modifications on the stall/spin behavior of a light general aviation airplane

Science.gov (United States)

1979-01-01

Configurations with full-span and segmented leading-edge flaps and full-span and segmented leading-edge droop were tested. Studies were conducted with wind-tunnel models, with an outdoor radio-controlled model, and with a full-scale airplane. Results show that wing-leading-edge modifications can produce large effects on stall/spin characteristics, particularly on spin resistance. One outboard wing-leading-edge modification tested significantly improved lateral stability at stall, spin resistance, and developed spin characteristics.

Distributional Replication

OpenAIRE

Beare, Brendan K.

2009-01-01

Suppose that X and Y are random variables. We define a replicating function to be a function f such that f(X) and Y have the same distribution. In general, the set of replicating functions for a given pair of random variables may be infinite. Suppose we have some objective function, or cost function, defined over the set of replicating functions, and we seek to estimate the replicating function with the lowest cost. We develop an approach to estimating the cheapest replicating function that i...

Fluid mechanics of dynamic stall. II - Prediction of full scale characteristics

Science.gov (United States)

Ericsson, L. E.; Reding, J. P.

1988-01-01

Analytical extrapolations are made from experimental subscale dynamics to predict full scale characteristics of dynamic stall. The method proceeds by establishing analytic relationships between dynamic and static aerodynamic characteristics induced by viscous flow effects. The method is then validated by predicting dynamic test results on the basis of corresponding static test data obtained at the same subscale flow conditions, and the effect of Reynolds number on the static aerodynamic characteristics are determined from subscale to full scale flow conditions.

Prediction of H.A.W.T. blade stall and performance

Energy Technology Data Exchange (ETDEWEB)

Giannakidis, G.; Graham, J.M.R. [Imperial College, Dept. of Aeronautics, London (United Kingdom)

1996-09-01

A model is being developed for the prediction of Horizontal Axis Wind Turbine blade stall and performance coupled with a simple aeroelastic analysis model. For the aerodynamic calculation a two dimensional unsteady Navier-Stokes solver on a sectional basis on the blade is coupled with a three dimensional vortex lattice wake. Pressure coefficient distributions are calculated from the two dimensional viscous flow in each blade section. The aerodynamic computations are coupled with a vibrating beam model in order to incorporate flapwise deformations of the blade. (au) 17 refs.

Chromatin Immunoprecipitation of Replication Factors Moving with the Replication Fork

OpenAIRE

Rapp, Jordan B.; Ansbach, Alison B.; Noguchi, Chiaki; Noguchi, Eishi

2009-01-01

Replication of chromosomes involves a variety of replication proteins including DNA polymerases, DNA helicases, and other accessory factors. Many of these proteins are known to localize at replication forks and travel with them as components of the replisome complex. Other proteins do not move with replication forks but still play an essential role in DNA replication. Therefore, in order to understand the mechanisms of DNA replication and its controls, it is important to examine localization ...

Hepatitis C virus translation preferentially depends on active RNA replication.

Directory of Open Access Journals (Sweden)

Helene Minyi Liu

Full Text Available Hepatitis C virus (HCV RNA initiates its replication on a detergent-resistant membrane structure derived from the endoplasmic reticulum (ER in the HCV replicon cells. By performing a pulse-chase study of BrU-labeled HCV RNA, we found that the newly-synthesized HCV RNA traveled along the anterograde-membrane traffic and moved away from the ER. Presumably, the RNA moved to the site of translation or virion assembly in the later steps of viral life cycle. In this study, we further addressed how HCV RNA translation was regulated by HCV RNA trafficking. When the movement of HCV RNA from the site of RNA synthesis to the Golgi complex was blocked by nocodazole, an inhibitor of ER-Golgi transport, HCV protein translation was surprisingly enhanced, suggesting that the translation of viral proteins occurred near the site of RNA synthesis. We also found that the translation of HCV proteins was dependent on active RNA synthesis: inhibition of viral RNA synthesis by an NS5B inhibitor resulted in decreased HCV viral protein synthesis even when the total amount of intracellular HCV RNA remained unchanged. Furthermore, the translation activity of the replication-defective HCV replicons or viral RNA with an NS5B mutation was greatly reduced as compared to that of the corresponding wildtype RNA. By performing live cell labeling of newly synthesized HCV RNA and proteins, we further showed that the newly synthesized HCV proteins colocalized with the newly synthesized viral RNA, suggesting that HCV RNA replication and protein translation take place at or near the same site. Our findings together indicate that the translation of HCV RNA is coupled to RNA replication and that the both processes may occur at the same subcellular membrane compartments, which we term the replicasome.

Stall margin improvement of an axial flow fan with end wall injection and suction; Hekimen fukidashi suidashi ni yoru han'yo jikuryu sofuki no shissoku kaizen

Energy Technology Data Exchange (ETDEWEB)

Nishioka, K.; Kuroda, H.; Obata, S.; Chimura, O. [National Defense Academy, Kanagawa (Japan)

1999-06-25

The experimental studies are conducted to reveal the mechanism of stall margin improvement of an axial flow fan by injection or suction from the end wall. In case of injection, the largest improvement is obtained by the injection at about 0. 14 {approx} 0 .21 times axial chord length downstream from leading edge. The reason for large improvement is that stall vortex, shed intermittent separation vortex and tip leakage vortex are dissipated by this injection, and also that this blowing suppresses the separation of boundary layer. In case of suction, the largest improvement is found for the suction from the end wall near leading edge. The amplitude of periodic static pressure after stall inception becomes smaller in comparison with injection cases. These effects are increased with the increase of suction flow rate, because the discharge of the vortex occurs more easily. On the other hand, the suction from the upstream of leading edge reduces the axial velocity near rotor tip, and then it induces stall. Also we tried to visualize the tip region flow, The suppression mechanism is discussed based on the visualization. The suppression of stall is successfully photographed. (author)

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...

Visualization and image analysis of dynamic stall phenomenon for a Darrieus wind turbine; Darrieus fusha ni okeru doteki shissoku gensho no kashika to gazo kaisekini kansuru kenkyu

Energy Technology Data Exchange (ETDEWEB)

Fujisawa, N.; Shibuya, S.; Takano, T. [Niigata University, Niigata (Japan)

1999-10-25

The flow field around a Darrieus wind turbine is studied by flow visualization and PIV measurement in a rotating frame of reference to understand the unsteady nature of dynamic stall appearing at low tip-speed ratios. The qualitative nature of the dynamic stall observed by the flow visualization using dye injection technique is quantitatively reproduced in the instantaneous velocity distributions around the blade measured by PIV technique. These results indicate that two pairs of stall vortices are generated in one cycle of the turbine rotation and they grow in size as the tip-speed ratio decreases. The mechanism of the dynamic stall is found to be due to the flow separation over the suction side of the blade followed by the generation of in-flow motion from the pressure side to the suction side of the blade through the trailing edge. (author)

Signals Involved in Regulation of Hepatitis C Virus RNA Genome Translation and Replication

Directory of Open Access Journals (Sweden)

Michael Niepmann

2018-03-01

Full Text Available Hepatitis C virus (HCV preferentially replicates in the human liver and frequently causes chronic infection, often leading to cirrhosis and liver cancer. HCV is an enveloped virus classified in the genus Hepacivirus in the family Flaviviridae and has a single-stranded RNA genome of positive orientation. The HCV RNA genome is translated and replicated in the cytoplasm. Translation is controlled by the Internal Ribosome Entry Site (IRES in the 5′ untranslated region (5′ UTR, while also downstream elements like the cis-replication element (CRE in the coding region and the 3′ UTR are involved in translation regulation. The cis-elements controlling replication of the viral RNA genome are located mainly in the 5′- and 3′-UTRs at the genome ends but also in the protein coding region, and in part these signals overlap with the signals controlling RNA translation. Many long-range RNA–RNA interactions (LRIs are predicted between different regions of the HCV RNA genome, and several such LRIs are actually involved in HCV translation and replication regulation. A number of RNA cis-elements recruit cellular RNA-binding proteins that are involved in the regulation of HCV translation and replication. In addition, the liver-specific microRNA-122 (miR-122 binds to two target sites at the 5′ end of the viral RNA genome as well as to at least three additional target sites in the coding region and the 3′ UTR. It is involved in the regulation of HCV RNA stability, translation and replication, thereby largely contributing to the hepatotropism of HCV. However, we are still far from completely understanding all interactions that regulate HCV RNA genome translation, stability, replication and encapsidation. In particular, many conclusions on the function of cis-elements in HCV replication have been obtained using full-length HCV genomes or near-full-length replicon systems. These include both genome ends, making it difficult to decide if a cis-element in

Signals Involved in Regulation of Hepatitis C Virus RNA Genome Translation and Replication.

Science.gov (United States)

Niepmann, Michael; Shalamova, Lyudmila A; Gerresheim, Gesche K; Rossbach, Oliver

2018-01-01

Hepatitis C virus (HCV) preferentially replicates in the human liver and frequently causes chronic infection, often leading to cirrhosis and liver cancer. HCV is an enveloped virus classified in the genus Hepacivirus in the family Flaviviridae and has a single-stranded RNA genome of positive orientation. The HCV RNA genome is translated and replicated in the cytoplasm. Translation is controlled by the Internal Ribosome Entry Site (IRES) in the 5' untranslated region (5' UTR), while also downstream elements like the cis -replication element (CRE) in the coding region and the 3' UTR are involved in translation regulation. The cis -elements controlling replication of the viral RNA genome are located mainly in the 5'- and 3'-UTRs at the genome ends but also in the protein coding region, and in part these signals overlap with the signals controlling RNA translation. Many long-range RNA-RNA interactions (LRIs) are predicted between different regions of the HCV RNA genome, and several such LRIs are actually involved in HCV translation and replication regulation. A number of RNA cis -elements recruit cellular RNA-binding proteins that are involved in the regulation of HCV translation and replication. In addition, the liver-specific microRNA-122 (miR-122) binds to two target sites at the 5' end of the viral RNA genome as well as to at least three additional target sites in the coding region and the 3' UTR. It is involved in the regulation of HCV RNA stability, translation and replication, thereby largely contributing to the hepatotropism of HCV. However, we are still far from completely understanding all interactions that regulate HCV RNA genome translation, stability, replication and encapsidation. In particular, many conclusions on the function of cis -elements in HCV replication have been obtained using full-length HCV genomes or near-full-length replicon systems. These include both genome ends, making it difficult to decide if a cis -element in question acts on HCV

msCentipede: Modeling Heterogeneity across Genomic Sites and Replicates Improves Accuracy in the Inference of Transcription Factor Binding.

Directory of Open Access Journals (Sweden)

Anil Raj

Full Text Available Understanding global gene regulation depends critically on accurate annotation of regulatory elements that are functional in a given cell type. CENTIPEDE, a powerful, probabilistic framework for identifying transcription factor binding sites from tissue-specific DNase I cleavage patterns and genomic sequence content, leverages the hypersensitivity of factor-bound chromatin and the information in the DNase I spatial cleavage profile characteristic of each DNA binding protein to accurately infer functional factor binding sites. However, the model for the spatial profile in this framework fails to account for the substantial variation in the DNase I cleavage profiles across different binding sites. Neither does it account for variation in the profiles at the same binding site across multiple replicate DNase I experiments, which are increasingly available. In this work, we introduce new methods, based on multi-scale models for inhomogeneous Poisson processes, to account for such variation in DNase I cleavage patterns both within and across binding sites. These models account for the spatial structure in the heterogeneity in DNase I cleavage patterns for each factor. Using DNase-seq measurements assayed in a lymphoblastoid cell line, we demonstrate the improved performance of this model for several transcription factors by comparing against the Chip-seq peaks for those factors. Finally, we explore the effects of DNase I sequence bias on inference of factor binding using a simple extension to our framework that allows for a more flexible background model. The proposed model can also be easily applied to paired-end ATAC-seq and DNase-seq data. msCentipede, a Python implementation of our algorithm, is available at http://rajanil.github.io/msCentipede.

msCentipede: Modeling Heterogeneity across Genomic Sites and Replicates Improves Accuracy in the Inference of Transcription Factor Binding.

Science.gov (United States)

Raj, Anil; Shim, Heejung; Gilad, Yoav; Pritchard, Jonathan K; Stephens, Matthew

2015-01-01

Understanding global gene regulation depends critically on accurate annotation of regulatory elements that are functional in a given cell type. CENTIPEDE, a powerful, probabilistic framework for identifying transcription factor binding sites from tissue-specific DNase I cleavage patterns and genomic sequence content, leverages the hypersensitivity of factor-bound chromatin and the information in the DNase I spatial cleavage profile characteristic of each DNA binding protein to accurately infer functional factor binding sites. However, the model for the spatial profile in this framework fails to account for the substantial variation in the DNase I cleavage profiles across different binding sites. Neither does it account for variation in the profiles at the same binding site across multiple replicate DNase I experiments, which are increasingly available. In this work, we introduce new methods, based on multi-scale models for inhomogeneous Poisson processes, to account for such variation in DNase I cleavage patterns both within and across binding sites. These models account for the spatial structure in the heterogeneity in DNase I cleavage patterns for each factor. Using DNase-seq measurements assayed in a lymphoblastoid cell line, we demonstrate the improved performance of this model for several transcription factors by comparing against the Chip-seq peaks for those factors. Finally, we explore the effects of DNase I sequence bias on inference of factor binding using a simple extension to our framework that allows for a more flexible background model. The proposed model can also be easily applied to paired-end ATAC-seq and DNase-seq data. msCentipede, a Python implementation of our algorithm, is available at http://rajanil.github.io/msCentipede.

FBH1 Catalyzes Regression of Stalled Replication Forks

DEFF Research Database (Denmark)

Fugger, Kasper; Mistrik, Martin; Neelsen, Kai J

2015-01-01

, is required for early phosphorylation of ATM substrates such as CHK2 and CtIP as well as hyperphosphorylation of RPA. These phosphorylations occur prior to apparent DNA double-strand break formation. Furthermore, FBH1-dependent signaling promotes checkpoint control and preserves genome integrity. We propose...

Database Replication Prototype

OpenAIRE

Vandewall, R.

2000-01-01

This report describes the design of a Replication Framework that facilitates the implementation and com-parison of database replication techniques. Furthermore, it discusses the implementation of a Database Replication Prototype and compares the performance measurements of two replication techniques based on the Atomic Broadcast communication primitive: pessimistic active replication and optimistic active replication. The main contributions of this report can be split into four parts....

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

2016-01-01

Post-translational modification of proteins by ubiquitin (Ub) and Ub-like modifiers regulates various aspects of DNA replication. We previously showed that the chromatin around replisomes is rich in SUMO and depleted in Ub, whereas an opposite pattern is observed in mature chromatin. How this SUMO-rich/Ub-low environment is maintained at sites of DNA replication is not known. 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. Chemical inhibition or genetic deletion of USP7 leads to the accumulation of Ub on SUMOylated proteins, which are displaced to chromatin away from replisomes. Our findings provide a model to explain the differential accumulation of SUMO and Ub at replication forks, and identify an essential role of USP7 in DNA replication that should be taken into account for the use of USP7 inhibitors as anticancer agents. PMID:26950370

Asynchronous DNA replication within the human β-globin gene locus

International Nuclear Information System (INIS)

Epner, E.; Forrester, W.C.; Groudine, M.

1988-01-01

The timing of DNA replication of the human β-globin gene locus has been studied by blot hybridization of newly synthesized BrdUrd-substituted DNA from cells in different stages of the S phase. Using probes that span >120 kilobases across the human β-globin gene locus, the authors show that the majority of this domain replicates in early S phase in the human erythroleukemia cell line K562 and in middle-to-late S phase in the lymphoid cell line Manca. However, in K562 cells three small regions display a strikingly different replication pattern than adjacent sequences. These islands, located in the inter-γ-globin gene region and approximately 20 kilobases 5' to the ε-globin gene and 20 kilobases 3' to the β-globin gene, replicate later and throughout S phase. A similar area is also present in the α-globin gene region in K562 cells. They suggest that these regions may represent sites of termination of replication forks

RANS study of unsteady flow around a profile blade : application to stall of horizontal axis wind turbine

Energy Technology Data Exchange (ETDEWEB)

Belkheir, N. [Khemis Miliana Univ., Ain Defla (Algeria); Dizene, R. [Univ. des Sciences et de la Technologie Houari Boumediene, Algiers (Algeria). Laboratoire de Mecanique Avancee; Khelladi, S.; Massouh, F.; Dobrev, I. [Arts et Metiers Paris Tech., Paris (France)

2010-07-01

The shape of an airfoil is designed to achieve the best aerodynamic performance. An aerofoil section undergoes dynamic stall when subjected to any form of unsteady angle of pitch. The study of a horizontal-axis wind turbine (HAWT) under wind operating conditions is complex because it is subject to instantaneous speed and wind direction variation. When turbine blades are driven into a dynamic stall, the lift coefficient drops suddenly resulting in a degradation in aerodynamic performance. This study presented steady and unsteady wind load predictions over an oscillating S809 airfoil tested in a subsonic wind tunnel. A model of sinusoidal pitch oscillations was used. The values for the angles of attack in steady state ranged from -20 to +40 degrees. The model considered 3 frequencies and 2 amplitudes. The two-dimensional numerical model simulated the instantaneous change of wind direction with respect to the turbine blade. Results were compared with data measurements of S809 aerofoil. Reasonable deviations were obtained between the predicted and experimental results for pitch oscillations. The URANS approach was used to predict the stall while the software FLUENT was used for the numerical solution. It was concluded that the behaviour of the unsteady flow in the wind farm must be considered in order to obtain an accurate estimate of the wind turbine aerodynamic load. 12 refs., 5 figs.

Nuclear safety aspects of exported replicate nuclear power plants and associated problems

International Nuclear Information System (INIS)

Kern, H.G.

1978-01-01

The standardization of the export nuclear power plant is being pursued with the concept of replication. This concept entails using another exported Nuclear Power Plant (NPP) as the base design and adapting it to a new site. The general ground rule applied to this concept is upgrading the design where necessary and duplicating the design where it is superior. Such continuous improvement will result in a standard export NPP that incorporates design features which will make it essentially acceptable for any suitable site. The advantages of replication are, therefore, boundless. However, the replication mode requires superior design control by the engineer to assure that only improvements alter the base design. With this concept, the replicating engineer is essentially assigned the responsiblity of safeguarding the standard export plant design. He is delegated the task of filtering the design such that only the conservative aspects prevail. Tight control of design changes via properly administered procedures is necessary to assure that no unforeseen compromises are made in designs which have already achieved optimization. Techniques to accomplish successful replication include, among others, the use of PCNs, system cognizant engineers, design verfication review, and the participation of all engineering disciplines in the development of the project schedule. (author)

Influence of DNA Lesions on Polymerase-Mediated DNA Replication at Single-Molecule Resolution.

Science.gov (United States)

Gahlon, Hailey L; Romano, Louis J; Rueda, David

2017-11-20

Faithful replication of DNA is a critical aspect in maintaining genome integrity. DNA polymerases are responsible for replicating DNA, and high-fidelity polymerases do this rapidly and at low error rates. Upon exposure to exogenous or endogenous substances, DNA can become damaged and this can alter the speed and fidelity of a DNA polymerase. In this instance, DNA polymerases are confronted with an obstacle that can result in genomic instability during replication, for example, by nucleotide misinsertion or replication fork collapse. It is important to know how DNA polymerases respond to damaged DNA substrates to understand the mechanism of mutagenesis and chemical carcinogenesis. Single-molecule techniques have helped to improve our current understanding of DNA polymerase-mediated DNA replication, as they enable the dissection of mechanistic details that can otherwise be lost in ensemble-averaged experiments. These techniques have also been used to gain a deeper understanding of how single DNA polymerases behave at the site of the damage in a DNA substrate. In this review, we evaluate single-molecule studies that have examined the interaction between DNA polymerases and damaged sites on a DNA template.

Numerical study on a single bladed vertical axis wind turbine under dynamic stall

Energy Technology Data Exchange (ETDEWEB)

Bangga, Galih [Institute of Aerodynamics and Gas Dynamics, University of Stuttgart, Stuttgart (Germany); Hutomo, Go; Sasongko, Herman [Dept. of Mechanical Engineering, Institut Teknologi Sepuluh Nopember, Surabaya (Indonesia); Wiranegara, Raditya [School of Mechanical Aerospace and Civil Engineering, University of Manchester, Manchester (United Kingdom)

2017-01-15

The aim of this study is to investigate the flow development of a single bladed vertical axis wind turbine using Computational fluid dynamics (CFD) methods. The blade is constructed using the NACA 0012 profile and is operating under stalled conditions at tip speed ratio of 2. Two dimensional simulations are performed using a commercial CFD package, ANSYS Fluent 15.0, employing the Menter-SST turbulence model. For the preliminary study, simulations of the NACA 0012 airfoil under static conditions are carried out and compared with available measurement data and calculations using the boundary layer code XFOIL. The CFD results under the dynamic case are presented and the resulting aerodynamic forces are evaluated. The turbine is observed to generate negative power at certain azimuth angles which can be divided into three main zones. The blade vortex interaction is observed to strongly influence the flow behavior near the blade and contributes to the power production loss. However, the impact is considered small since it covers only 6.4 % of the azimuth angle range where the power is negative compared to the dynamic stall impact which covers almost 22 % of the azimuth angle range.

Blade tip, finite aspect ratio, and dynamic stall effects on the Darrieus rotor

Science.gov (United States)

Paraschivoiu, I.; Desy, P.; Masson, C.

1988-02-01

The objective of the work described in this paper was to apply the Boeing-Vertol dynamic stall model in an asymmetric manner to account for the asymmetry of the flow between the left and right sides of the rotor. This phenomenon has been observed by the flow visualization of a two-straight-bladed Darrieus rotor in the IMST water tunnel. Also introduced into the aerodynamic model are the effects of the blade tip and finite aspect ratio on the aerodynamic performance of the Darrieus wind turbine. These improvements are compatible with the double-multiple-streamtube model and have been included in the CARDAAV computer code for predicting the aerodynamic performance. Very good agreement has been observed between the test data (Sandia 17 m) and theoretical predictions; a significant improvement over the previous dynamic stall model was obtained for the rotor power at low tip speed ratios, while the inclusion of the finite aspect ratio effects enhances the prediction of the rotor power for high tip speed ratios. The tip losses and finite aspect ratio effects were also calculated for a small-scale vertical-axis wind turbine, with a two-straight-bladed (NACA 0015) rotor.

Grid support of a wind farm with active stall wind turbines and AC grid connection

DEFF Research Database (Denmark)

Hansen, Anca Daniela; Sørensen, Poul Ejnar; Iov, F.

2006-01-01

grid connection. The designed control system has the task of enabling such a wind farm to provide the best grid support. It is based on two control levels: a supervisory control level, which controls the power production of the whole farm by sending out reference signals to each individual wind turbine......One of the main concerns in the grid integration of large wind farms is their ability to behave as active controllable components in the power system. This article presents the design of a new integrated power control system for a wind farm made up exclusively of active stall wind turbines with AC......, and a local control level, which ensures that the reference power signals at the wind turbine level are reached. The ability of active stall wind farms with AC grid connection to control the power production to the reference power ordered by the operators is assessed and discussed by means of simulations....

Homologous regions of Fen1 and p21Cip1 compete for binding to the same site on PCNA: a potential mechanism to co-ordinate DNA replication and repair.

Science.gov (United States)

Warbrick, E; Lane, D P; Glover, D M; Cox, L S

1997-05-15

Following genomic damage, the cessation of DNA replication is co-ordinated with onset of DNA repair; this co-ordination is essential to avoid mutation and genomic instability. To investigate these phenomena, we have analysed proteins that interact with PCNA, which is required for both DNA replication and repair. One such protein is p21Cip1, which inhibits DNA replication through its interaction with PCNA, while allowing repair to continue. We have identified an interaction between PCNA and the structure specific nuclease, Fen1, which is involved in DNA replication. Deletion analysis suggests that p21Cip1 and Fen1 bind to the same region of PCNA. Within Fen1 and its homologues a small region (10 amino acids) is sufficient for PCNA binding, which contains an 8 amino acid conserved PCNA-binding motif. This motif shares critical residues with the PCNA-binding region of p21Cip1. A PCNA binding peptide from p21Cip1 competes with Fen1 peptides for binding to PCNA, disrupts the Fen1-PCNA complex in replicating cell extracts, and concomitantly inhibits DNA synthesis. Competition between homologous regions of Fen1 and p21Cip1 for binding to the same site on PCNA may provide a mechanism to co-ordinate the functions of PCNA in DNA replication and repair.

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

The replication recipe : What makes for a convincing replication?

NARCIS (Netherlands)

Brandt, M.J.; IJzerman, H.; Dijksterhuis, Ap; Farach, Frank J.; Geller, Jason; Giner-Sorolla, Roger; Grange, James A.; Perugini, Marco; Spies, Jeffrey R.; van 't Veer, Anna

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

Dynamic stall study of a multi-element airfoil

Science.gov (United States)

Tung, Chee; Mcalister, Kenneth W.; Wang, Clin M.

1992-01-01

Unsteady flow behavior and load characteristics of a VR-7 airfoil with and without a slat were studied in the water tunnel of the Aeroflightdynamics Directorate, NASA Ames Research Center. Both airfoils were oscillated sinusoidally between 5 and 25 degrees at a Reynolds number of 200,000 to obtain the unsteady lift, drag and pitching moment data. A fluorescing dye was released from an orifice located at the leading edge of the airfoil for the purpose of visualizing the boundary layer and wake flow. The flow field and load predictions of an incompressible Navier-Stokes code based on a velocity-vorticity formulation were compared with the test data. The test and predictions both confirm that the slatted VR-7 airfoil delays both static and dynamic stall as compared to the VR-7 airfoil alone.

Proposed Chevron Tengiz venture stalls amid Soviet political squabble

International Nuclear Information System (INIS)

Anon.

1991-01-01

This paper reports on the status of foreign investment in Soviet oil and gas joint ventures which has reached a critical juncture. Just as the U.S. is considering granting most favored nation trade status to the U.S.S.R., the joint venture petroleum deal seen as the litmus test for such deals-Chevron Corp.'s proposed addition of supergiant Tengiz oil field to its Caspian Sea joint venture-has stalled amid controversy. Unconfirmed reports from Soviet officials and other foreign joint venture participants in the U.S.S.R. have Chevron pulling out of the long negotiated, multibillion dollar project after the Soviets rejected the company's terms. Chevron, however, insists the project is still alive

Genetic variations in the DNA replication origins of human papillomavirus family correlate with their oncogenic potential.

Science.gov (United States)

Yilmaz, Gulden; Biswas-Fiss, Esther E; Biswas, Subhasis B

2018-04-01

Human papillomaviruses (HPVs) encompass a large family of viruses that range from benign to highly carcinogenic. The crucial differences between benign and carcinogenic types of HPV remain unknown, except that the two HPV types differ in the frequency of DNA replication. We have systematically analyzed the mechanism of HPV DNA replication initiation in low-risk and high-risk HPVs. Our results demonstrate that HPV-encoded E2 initiator protein and its four binding sites in the replication origin play pivotal roles in determining the destiny of the HPV-infected cell. We have identified strain-specific single nucleotide variations in E2 binding sites found only in the high-risk HPVs. We have demonstrated that these variations result in attenuated formation of the E2-DNA complex. E2 binding to these sites is linked to the activation of the DNA replication origin as well as initiation of DNA replication. Both electrophoretic mobility shift assay and atomic force microscopy studies demonstrated that binding of E2 from either low- or high-risk HPVs with variant binding sequences lacked multimeric E2-DNA complex formation in vitro. These results provided a molecular basis of differential DNA replication in the two types of HPVs and pointed to a correlation with the development of cancer. Copyright © 2017. Published by Elsevier B.V.

The cellular Mre11 protein interferes with adenovirus E4 mutant DNA replication

International Nuclear Information System (INIS)

Mathew, Shomita S.; Bridge, Eileen

2007-01-01

Adenovirus type 5 (Ad5) relocalizes and degrades the host DNA repair protein Mre11, and efficiently initiates viral DNA replication. Mre11 associates with Ad E4 mutant DNA replication centers and is important for concatenating viral genomes. We have investigated the role of Mre11 in the E4 mutant DNA replication defect. RNAi-mediated knockdown of Mre11 dramatically rescues E4 mutant DNA replication in cells that do or do not concatenate viral genomes, suggesting that Mre11 inhibits DNA replication independent of genome concatenation. The mediator of DNA damage checkpoint 1 (Mdc1) protein is involved in recruiting and sustaining Mre11 at sites of DNA damage following ionizing radiation. We observe foci formation by Mdc1 in response to viral infection, indicating that this damage response protein is activated. However, knockdown of Mdc1 does not prevent Mre11 from localizing at viral DNA replication foci or rescue E4 mutant DNA replication. Our results are consistent with a model in which Mre11 interferes with DNA replication when it is localized at viral DNA replication foci

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

Rev1, Rev3, or Rev7 siRNA Abolishes Ultraviolet Light-Induced Translesion Replication in HeLa Cells: A Comprehensive Study Using Alkaline Sucrose Density Gradient Sedimentation

Directory of Open Access Journals (Sweden)

Jun Takezawa

2010-01-01

Full Text Available When a replicative DNA polymerase stalls upon encountering a lesion on the template strand, it is relieved by other low-processivity polymerase(s, which insert nucleotide(s opposite the lesion, extend by a few nucleotides, and dissociate from the 3′-OH. The replicative polymerase then resumes DNA synthesis. This process, termed translesion replication (TLS or replicative bypass, may involve at least five different polymerases in mammals, although the participating polymerases and their roles have not been entirely characterized. Using siRNAs originally designed and an alkaline sucrose density gradient sedimentation technique, we verified the involvement of several polymerases in ultraviolet (UV light-induced TLS in HeLa cells. First, siRNAs to Rev3 or Rev7 largely abolished UV-TLS, suggesting that these 2 gene products, which comprise Polζ, play a main role in mutagenic TLS. Second, Rev1-targeted siRNA also abrogated UV-TLS, indicating that Rev1 is also indispensable to mutagenic TLS. Third, Polη-targeted siRNA also prevented TLS to a greater extent than our expectations. Forth, although siRNA to Polι had no detectable effect, that to Polκ delayed UV-TLS. To our knowledge, this is the first study reporting apparent evidence for the participation of Polκ in UV-TLS.

3D replicon distributions arise from stochastic initiation and domino-like DNA replication progression.

Science.gov (United States)

Löb, D; Lengert, N; Chagin, V O; Reinhart, M; Casas-Delucchi, C S; Cardoso, M C; Drossel, B

2016-04-07

DNA replication dynamics in cells from higher eukaryotes follows very complex but highly efficient mechanisms. However, the principles behind initiation of potential replication origins and emergence of typical patterns of nuclear replication sites remain unclear. Here, we propose a comprehensive model of DNA replication in human cells that is based on stochastic, proximity-induced replication initiation. Critical model features are: spontaneous stochastic firing of individual origins in euchromatin and facultative heterochromatin, inhibition of firing at distances below the size of chromatin loops and a domino-like effect by which replication forks induce firing of nearby origins. The model reproduces the empirical temporal and chromatin-related properties of DNA replication in human cells. We advance the one-dimensional DNA replication model to a spatial model by taking into account chromatin folding in the nucleus, and we are able to reproduce the spatial and temporal characteristics of the replication foci distribution throughout S-phase.

Site-directed mutagenesis of HIV-1 vpu gene demonstrates two clusters of replication-defective mutants with distinct ability to down-modulate cell surface CD4 and tetherin

Directory of Open Access Journals (Sweden)

Masako Nomaguchi

2010-11-01

Full Text Available HIV-1 Vpu acts positively on viral infectivity by mediating CD4 degradation in endoplasmic reticulum and enhances virion release by counteracting a virion release restriction factor, tetherin. In order to define the impact of Vpu activity on HIV-1 replication, we have generated a series of site-specific proviral vpu mutants. Of fifteen mutants examined, seven exhibited a replication-defect similar to that of a vpu-deletion mutant in a lymphocyte cell line H9. These mutations clustered in narrow regions within transmembrane domain (TMD and cytoplasmic domain (CTD. Replication-defective mutants displayed the reduced ability to enhance virion release from a monolayer cell line HEp2 without exception. Upon transfection with Vpu expression vectors, neither TMD mutants nor CTD mutants blocked CD4 expression at the cell surface in another monolayer cell line MAGI. While TMD mutants were unable to down-modulate cell surface tetherin in HEp2 cells, CTD mutants did quite efficiently. Confocal microscopy analysis revealed the difference of intracellular localization between TMD and CTD mutants. In total, replication capability of HIV-1 carrying vpu mutations correlates well with the ability of Vpu to enhance virion release and to impede the cell surface expression of CD4 but not with the ability to down-modulate cell surface tetherin. Our results here suggest that efficient viral replication requires not only down-regulation of cell surface tetherin but also its degradation.

Identification of Persistent RNA-DNA Hybrid Structures within the Origin of Replication of Human Cytomegalovirus

OpenAIRE

Prichard, Mark N.; Jairath, Sanju; Penfold, Mark E. T.; Jeor, Stephen St.; Bohlman, Marlene C.; Pari, Gregory S.

1998-01-01

Human cytomegalovirus (HCMV) lytic-phase DNA replication initiates at the cis-acting origin of replication, oriLyt. oriLyt is a structurally complex region containing repeat elements and transcription factor binding sites. We identified two site-specific alkali-labile regions within oriLyt which flank an alkali-resistant DNA segment. These alkali-sensitive regions were the result of the degradation of two RNA species embedded within oriLyt and covalently linked to viral DNA. The virus-associa...

Validation of an Actuator Line Model Coupled to a Dynamic Stall Model for Pitching Motions Characteristic to Vertical Axis Turbines

International Nuclear Information System (INIS)

Mendoza, Victor; Goude, Anders; Bachant, Peter; Wosnik, Martin

2016-01-01

Vertical axis wind turbines (VAWT) can be used to extract renewable energy from wind flows. A simpler design, low cost of maintenance, and the ability to accept flow from all directions perpendicular to the rotor axis are some of the most important advantages over conventional horizontal axis wind turbines (HAWT). However, VAWT encounter complex and unsteady fluid dynamics, which present significant modeling challenges. One of the most relevant phenomena is dynamic stall, which is caused by the unsteady variation of angle of attack throughout the blade rotation, and is the focus of the present study. Dynamic stall is usually used as a passive control for VAWT operating conditions, hence the importance of predicting its effects. In this study, a coupled model is implemented with the open-source CFD toolbox OpenFOAM for solving the Navier-Stokes equations, where an actuator line model and dynamic stall model are used to compute the blade loading and body force. Force coefficients obtained from the model are validated with experimental data of pitching airfoil in similar operating conditions as an H-rotor type VAWT. Numerical results show reasonable agreement with experimental data for pitching motion. (paper)

Validation of an Actuator Line Model Coupled to a Dynamic Stall Model for Pitching Motions Characteristic to Vertical Axis Turbines

Science.gov (United States)

Mendoza, Victor; Bachant, Peter; Wosnik, Martin; Goude, Anders

2016-09-01

Vertical axis wind turbines (VAWT) can be used to extract renewable energy from wind flows. A simpler design, low cost of maintenance, and the ability to accept flow from all directions perpendicular to the rotor axis are some of the most important advantages over conventional horizontal axis wind turbines (HAWT). However, VAWT encounter complex and unsteady fluid dynamics, which present significant modeling challenges. One of the most relevant phenomena is dynamic stall, which is caused by the unsteady variation of angle of attack throughout the blade rotation, and is the focus of the present study. Dynamic stall is usually used as a passive control for VAWT operating conditions, hence the importance of predicting its effects. In this study, a coupled model is implemented with the open-source CFD toolbox OpenFOAM for solving the Navier-Stokes equations, where an actuator line model and dynamic stall model are used to compute the blade loading and body force. Force coefficients obtained from the model are validated with experimental data of pitching airfoil in similar operating conditions as an H-rotor type VAWT. Numerical results show reasonable agreement with experimental data for pitching motion.

(1)H, (13)C, and (15)N backbone resonance assignments of the full-length 40 kDa S. acidocaldarius Y-family DNA polymerase, dinB homolog.

Science.gov (United States)

Moro, Sean L; Cocco, Melanie J

2015-10-01

The dinB homolog (Dbh) is a member of the Y-family of translesion DNA polymerases, which are specialized to accurately replicate DNA across from a wide variety of lesions in living cells. Lesioned bases block the progression of high-fidelity polymerases and cause detrimental replication fork stalling; Y-family polymerases can bypass these lesions. The active site of the translesion synthesis polymerase is more open than that of a replicative polymerase; consequently Dbh polymerizes with low fidelity. Bypass polymerases also have low processivity. Short extension past the lesion allows the high-fidelity polymerase to switch back onto the site of replication. Dbh and the other Y-family polymerases have been used as structural models to investigate the mechanisms of DNA polymerization and lesion bypass. Many high-resolution crystal structures of Y-family polymerases have been reported. NMR dynamics studies can complement these structures by providing a measure of protein motions. Here we report the (15)N, (1)H, and (13)C backbone resonance assignments at two temperatures (35 and 50 °C) for Sulfolobus acidocaldarius Dbh polymerase. Backbone resonance assignments have been obtained for 86 % of the residues. The polymerase active site is assigned as well as the majority of residues in each of the four domains.

Structures of minute virus of mice replication initiator protein N-terminal domain: Insights into DNA nicking and origin binding

International Nuclear Information System (INIS)

Tewary, Sunil K.; Liang, Lingfei; Lin, Zihan; Lynn, Annie; Cotmore, Susan F.; Tattersall, Peter; Zhao, Haiyan; Tang, Liang

2015-01-01

Members of the Parvoviridae family all encode a non-structural protein 1 (NS1) that directs replication of single-stranded viral DNA, packages viral DNA into capsid, and serves as a potent transcriptional activator. Here we report the X-ray structure of the minute virus of mice (MVM) NS1 N-terminal domain at 1.45 Å resolution, showing that sites for dsDNA binding, ssDNA binding and cleavage, nuclear localization, and other functions are integrated on a canonical fold of the histidine-hydrophobic-histidine superfamily of nucleases, including elements specific for this Protoparvovirus but distinct from its Bocaparvovirus or Dependoparvovirus orthologs. High resolution structural analysis reveals a nickase active site with an architecture that allows highly versatile metal ligand binding. The structures support a unified mechanism of replication origin recognition for homotelomeric and heterotelomeric parvoviruses, mediated by a basic-residue-rich hairpin and an adjacent helix in the initiator proteins and by tandem tetranucleotide motifs in the replication origins. - Highlights: • The structure of a parvovirus replication initiator protein has been determined; • The structure sheds light on mechanisms of ssDNA binding and cleavage; • The nickase active site is preconfigured for versatile metal ligand binding; • The binding site for the double-stranded replication origin DNA is identified; • A single domain integrates multiple functions in virus replication

Structures of minute virus of mice replication initiator protein N-terminal domain: Insights into DNA nicking and origin binding

Energy Technology Data Exchange (ETDEWEB)

Tewary, Sunil K.; Liang, Lingfei; Lin, Zihan; Lynn, Annie [Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045 (United States); Cotmore, Susan F. [Departments of Laboratory Medicine, Yale University Medical School, New Haven, CT 06510 (United States); Tattersall, Peter [Departments of Laboratory Medicine, Yale University Medical School, New Haven, CT 06510 (United States); Departments of Genetics, Yale University Medical School, New Haven, CT 06510 (United States); Zhao, Haiyan, E-mail: zhaohy@ku.edu [Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045 (United States); Tang, Liang, E-mail: tangl@ku.edu [Department of Molecular Biosciences, University of Kansas, Lawrence, KS 66045 (United States)

2015-02-15

Members of the Parvoviridae family all encode a non-structural protein 1 (NS1) that directs replication of single-stranded viral DNA, packages viral DNA into capsid, and serves as a potent transcriptional activator. Here we report the X-ray structure of the minute virus of mice (MVM) NS1 N-terminal domain at 1.45 Å resolution, showing that sites for dsDNA binding, ssDNA binding and cleavage, nuclear localization, and other functions are integrated on a canonical fold of the histidine-hydrophobic-histidine superfamily of nucleases, including elements specific for this Protoparvovirus but distinct from its Bocaparvovirus or Dependoparvovirus orthologs. High resolution structural analysis reveals a nickase active site with an architecture that allows highly versatile metal ligand binding. The structures support a unified mechanism of replication origin recognition for homotelomeric and heterotelomeric parvoviruses, mediated by a basic-residue-rich hairpin and an adjacent helix in the initiator proteins and by tandem tetranucleotide motifs in the replication origins. - Highlights: • The structure of a parvovirus replication initiator protein has been determined; • The structure sheds light on mechanisms of ssDNA binding and cleavage; • The nickase active site is preconfigured for versatile metal ligand binding; • The binding site for the double-stranded replication origin DNA is identified; • A single domain integrates multiple functions in virus replication.

Differential Tus-Ter binding and lock formation: implications for DNA replication termination in Escherichia coli.

Science.gov (United States)

Moreau, Morgane J J; Schaeffer, Patrick M

2012-10-01

In E. coli, DNA replication termination occurs at Ter sites and is mediated by Tus. Two clusters of five Ter sites are located on each side of the terminus region and constrain replication forks in a polar manner. The polarity is due to the formation of the Tus-Ter-lock intermediate. Recently, it has been shown that DnaB helicase which unwinds DNA at the replication fork is preferentially stopped at the non-permissive face of a Tus-Ter complex without formation of the Tus-Ter-lock and that fork pausing efficiency is sequence dependent, raising two essential questions: Does the affinity of Tus for the different Ter sites correlate with fork pausing efficiency? Is formation of the Tus-Ter-lock the key factor in fork pausing? The combined use of surface plasmon resonance and GFP-Basta showed that Tus binds strongly to TerA-E and G, moderately to TerH-J and weakly to TerF. Out of these ten Ter sites only two, TerF and H, were not able to form significant Tus-Ter-locks. Finally, Tus's resistance to dissociation from Ter sites and the strength of the Tus-Ter-locks correlate with the differences in fork pausing efficiency observed for the different Ter sites by Duggin and Bell (2009).

Fidelity of DNA Replication in Normal and Malignant Human Breast Cells.

Science.gov (United States)

1997-08-01

enzyme) into the multiple cloning site (MCS). This template will not only replicate inside a mammalian cell (utilizing the E-B virus origin), and...Maniatis, T. Commonly used techniques in molecular cloning . In: Molecular cloning : REFERENCES a laboratory manual, 2nd edition. Cold Spring Harbor...A vatit"Y Of DNA synthesis and the typt of DNA replica~tion Products " celular prca including DNA rsplicatlon. DNA repsair. R~NA formed in experiments

The DNA replication checkpoint directly regulates MBF-dependent G1/S transcription.

Science.gov (United States)

Dutta, Chaitali; Patel, Prasanta K; Rosebrock, Adam; Oliva, Anna; Leatherwood, Janet; Rhind, Nicholas

2008-10-01

The DNA replication checkpoint transcriptionally upregulates genes that allow cells to adapt to and survive replication stress. Our results show that, in the fission yeast Schizosaccharomyces pombe, the replication checkpoint regulates the entire G(1)/S transcriptional program by directly regulating MBF, the G(1)/S transcription factor. Instead of initiating a checkpoint-specific transcriptional program, the replication checkpoint targets MBF to maintain the normal G(1)/S transcriptional program during replication stress. We propose a mechanism for this regulation, based on in vitro phosphorylation of the Cdc10 subunit of MBF by the Cds1 replication-checkpoint kinase. Replacement of two potential phosphorylation sites with phosphomimetic amino acids suffices to promote the checkpoint transcriptional program, suggesting that Cds1 phosphorylation directly regulates MBF-dependent transcription. The conservation of MBF between fission and budding yeast, and recent results implicating MBF as a target of the budding yeast replication checkpoint, suggests that checkpoint regulation of the MBF transcription factor is a conserved strategy for coping with replication stress. Furthermore, the structural and regulatory similarity between MBF and E2F, the metazoan G(1)/S transcription factor, suggests that this checkpoint mechanism may be broadly conserved among eukaryotes.

The DNA Replication Checkpoint Directly Regulates MBF-Dependent G1/S Transcription▿

Science.gov (United States)

Dutta, Chaitali; Patel, Prasanta K.; Rosebrock, Adam; Oliva, Anna; Leatherwood, Janet; Rhind, Nicholas

2008-01-01

The DNA replication checkpoint transcriptionally upregulates genes that allow cells to adapt to and survive replication stress. Our results show that, in the fission yeast Schizosaccharomyces pombe, the replication checkpoint regulates the entire G1/S transcriptional program by directly regulating MBF, the G1/S transcription factor. Instead of initiating a checkpoint-specific transcriptional program, the replication checkpoint targets MBF to maintain the normal G1/S transcriptional program during replication stress. We propose a mechanism for this regulation, based on in vitro phosphorylation of the Cdc10 subunit of MBF by the Cds1 replication-checkpoint kinase. Replacement of two potential phosphorylation sites with phosphomimetic amino acids suffices to promote the checkpoint transcriptional program, suggesting that Cds1 phosphorylation directly regulates MBF-dependent transcription. The conservation of MBF between fission and budding yeast, and recent results implicating MBF as a target of the budding yeast replication checkpoint, suggests that checkpoint regulation of the MBF transcription factor is a conserved strategy for coping with replication stress. Furthermore, the structural and regulatory similarity between MBF and E2F, the metazoan G1/S transcription factor, suggests that this checkpoint mechanism may be broadly conserved among eukaryotes. PMID:18662996

New paradigms in the repair of oxidative damage in human genome: mechanisms ensuring repair of mutagenic base lesions during replication and involvement of accessory proteins.

Science.gov (United States)

Dutta, Arijit; Yang, Chunying; Sengupta, Shiladitya; Mitra, Sankar; Hegde, Muralidhar L

2015-05-01

Oxidized bases in the mammalian genome, which are invariably mutagenic due to their mispairing property, are continuously induced by endogenous reactive oxygen species and more abundantly after oxidative stress. Unlike bulky base adducts induced by UV and other environmental mutagens in the genome that block replicative DNA polymerases, oxidatively damaged bases such as 5-hydroxyuracil, produced by oxidative deamination of cytosine in the template strand, do not block replicative polymerases and thus need to be repaired prior to replication to prevent mutation. Following up our earlier studies, which showed that the Nei endonuclease VIII like 1 (NEIL1) DNA glycosylase, one of the five base excision repair (BER)-initiating enzymes in mammalian cells, has enhanced expression during the S-phase and higher affinity for replication fork-mimicking single-stranded (ss) DNA substrates, we recently provided direct experimental evidence for NEIL1's role in replicating template strand repair. The key requirement for this event, which we named as the 'cow-catcher' mechanism of pre-replicative BER, is NEIL1's non-productive binding (substrate binding without product formation) to the lesion base in ss DNA template to stall DNA synthesis, causing fork regression. Repair of the lesion in reannealed duplex is then carried out by NEIL1 in association with the DNA replication proteins. NEIL1 (and other BER-initiating enzymes) also interact with several accessory and non-canonical proteins including the heterogeneous nuclear ribonucleoprotein U and Y-box-binding protein 1 as well as high mobility group box 1 protein, whose precise roles in BER are still obscure. In this review, we have discussed the recent advances in our understanding of oxidative genome damage repair pathways with particular focus on the pre-replicative template strand repair and the role of scaffold factors like X-ray repairs cross-complementing protein 1 and poly (ADP-ribose) polymerase 1 and other accessory

Optimal parameters for the FFA-Beddoes dynamic stall model

Energy Technology Data Exchange (ETDEWEB)

Bjoerck, A; Mert, M [FFA, The Aeronautical Research Institute of Sweden, Bromma (Sweden); Madsen, H A [Risoe National Lab., Roskilde (Denmark)

1999-03-01

Unsteady aerodynamic effects, like dynamic stall, must be considered in calculation of dynamic forces for wind turbines. Models incorporated in aero-elastic programs are of semi-empirical nature. Resulting aerodynamic forces therefore depend on values used for the semi-empiricial parameters. In this paper a study of finding appropriate parameters to use with the Beddoes-Leishman model is discussed. Minimisation of the `tracking error` between results from 2D wind tunnel tests and simulation with the model is used to find optimum values for the parameters. The resulting optimum parameters show a large variation from case to case. Using these different sets of optimum parameters in the calculation of blade vibrations, give rise to quite different predictions of aerodynamic damping which is discussed. (au)

Deciphering DNA replication dynamics in eukaryotic cell populations in relation with their averaged chromatin conformations

Science.gov (United States)

Goldar, A.; Arneodo, A.; Audit, B.; Argoul, F.; Rappailles, A.; Guilbaud, G.; Petryk, N.; Kahli, M.; Hyrien, O.

2016-03-01

We propose a non-local model of DNA replication that takes into account the observed uncertainty on the position and time of replication initiation in eukaryote cell populations. By picturing replication initiation as a two-state system and considering all possible transition configurations, and by taking into account the chromatin’s fractal dimension, we derive an analytical expression for the rate of replication initiation. This model predicts with no free parameter the temporal profiles of initiation rate, replication fork density and fraction of replicated DNA, in quantitative agreement with corresponding experimental data from both S. cerevisiae and human cells and provides a quantitative estimate of initiation site redundancy. This study shows that, to a large extent, the program that regulates the dynamics of eukaryotic DNA replication is a collective phenomenon that emerges from the stochastic nature of replication origins initiation.

Cell lethality after selective irradiation of the DNA replication fork

International Nuclear Information System (INIS)

Hofer, K.G.; Warters, R.L.

1985-01-01

It has been suggested that nascent DNA located at the DNA replication fork may exhibit enhanced sensitivity to radiation damage. To evaluate this hypothesis, Chinese hamster ovary cells (CHO) were labeled with 125 I-iododeoxyuridine ( 125 IUdR) either in the presence or absence of aphidicolin. Aphidicolin (5 μg/ml) reduced cellular 125 IUdR incorporation to 3-5% of the control value. The residual 125 I incorporation appeared to be restricted to low molecular weight (sub-replicon sized) fragments of DNA which were more sensitive to micrococcal nuclease attack and less sensitive to high salt DNase I digestion than randomly labeled DNA. These findings suggest that DNA replicated in the presence of aphidicolin remains localized at the replication fork adjacent to the nuclear matrix. Based on these observations an attempt was made to compare the lethal consequences of 125 I decays at the replication fork to that of 125 I decays randomly distributed over the entire genome. Regardless of the distribution of decay events, all treatment groups exhibited identical dose-response curves (D 0 : 101 125 I decays/cell). Since differential irradiation of the replication complex did not result in enhanced cell lethality, it can be concluded that neither the nascent DNA nor the protein components (replicative enzymes, nuclear protein matrix) associated with the DNA replication site constitute key radiosensitive targets within the cellular genome. (orig.)

Soluble histone H2AX is induced by DNA replication stress and sensitizes cells to undergo apoptosis

Directory of Open Access Journals (Sweden)

Duensing Stefan

2008-07-01

Full Text Available Abstract Background Chromatin-associated histone H2AX is a key regulator of the cellular responses to DNA damage. However, non-nucleosomal functions of histone H2AX are poorly characterized. We have recently shown that soluble H2AX can trigger apoptosis but the mechanisms leading to non-chromatin-associated H2AX are unclear. Here, we tested whether stalling of DNA replication, a common event in cancer cells and the underlying mechanism of various chemotherapeutic agents, can trigger increased soluble H2AX. Results Transient overexpression of H2AX was found to lead to a detectable fraction of soluble H2AX and was associated with increased apoptosis. This effect was enhanced by the induction of DNA replication stress using the DNA polymerase α inhibitor aphidicolin. Cells manipulated to stably express H2AX did not contain soluble H2AX, however, short-term treatment with aphidicolin (1 h resulted in detectable amounts of H2AX in the soluble nuclear fraction and enhanced apoptosis. Similarly, soluble endogenous H2AX was detected under these conditions. We found that excessive soluble H2AX causes chromatin aggregation and inhibition of ongoing gene transcription as evidenced by the redistribution and/or loss of active RNA polymerase II as well as the transcriptional co-activators CBP and p300. Conclusion Taken together, these results show that DNA replication stress rapidly leads to increased soluble H2AX and that non-chromatin-associated H2AX can sensitize cells to undergo apoptosis. Our findings encourage further studies to explore H2AX and the cellular pathways that control its expression as anti-cancer drug targets.

The "enemies within": regions of the genome that are inherently difficult to replicate [version 1; referees: 2 approved

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Rahul Bhowmick

2017-05-01

Full Text Available An unusual feature of many eukaryotic genomes is the presence of regions that appear intrinsically difficult to copy during the process of DNA replication. Curiously, the location of these difficult-to-replicate regions is often conserved between species, implying a valuable role in some aspect of genome organization or maintenance. The most prominent class of these regions in mammalian cells is defined as chromosome fragile sites, which acquired their name because of a propensity to form visible gaps/breaks on otherwise-condensed chromosomes in mitosis. This fragility is particularly apparent following perturbation of DNA replication—a phenomenon often referred to as “replication stress”. Here, we review recent data on the molecular basis for chromosome fragility and the role of fragile sites in the etiology of cancer. In particular, we highlight how studies on fragile sites have provided unexpected insights into how the DNA repair machinery assists in the completion of DNA replication.

Roles for Dam methylation in bacterial chromosome replication

DEFF Research Database (Denmark)

Charbon, Godefroid; Koch, Birgit; Skovgaard, Ole

GATC sequences in the DNA of Escherichia coli and related species are methylated at the adenine residue by DNA adenine methyltransferase (DamMT). These methylated residues and/or the level of DamMT influence initiation of chromosome replication from the replication origin, oriC, which contain...... for about one third of the cell cycle. During sequestration at least three mechanisms operate to lower the activity of the initiator protein, DnaA. First, the dnaA promoter, which also contains an excess of GATC sequences, is sequestered for the same period of time as oriC to prevent de novo DnaA synthesis....... Second, new DnaA binding sites outside oriC are generated by replication which serve to titrate free DNA protein. Third, after initiation, DnaA-ATP is converted to inactive DnaA-ADP by a process called RIDA (regulatory inactivation of DnaA), which is dependent on the beta-clamp of DNA polymerase III...

The kissing-loop motif is a preferred site of 5' leader recombination during replication of SL3-3 murine leukemia viruses in mice

DEFF Research Database (Denmark)

Lund, Anders Henrik; Mikkelsen, J G; Schmidt, J

1999-01-01

, and the upstream part of the 5' untranslated region, enabled us to map recombination sites, guided by distinct scattered nucleotide differences. In 30 of 44 analyzed sequences, recombination was mapped to a 33-nucleotide similarity window coinciding with the kissing-loop stem-loop motif implicated in dimerization...... of the diploid genome. Interestingly, the recombination pattern preference found in replication-competent viruses from T-cell tumors is very similar to the pattern previously reported for retroviral vectors in cell culture experiments. The data therefore sustain the hypothesis that the kissing loop, presumably...

Choreography of the DNA damage response

DEFF Research Database (Denmark)

Lisby, Michael; Barlow, Jacqueline H; Burgess, Rebecca C

2004-01-01

DNA repair is an essential process for preserving genome integrity in all organisms. In eukaryotes, recombinational repair is choreographed by multiprotein complexes that are organized into centers (foci). Here, we analyze the cellular response to DNA double-strand breaks (DSBs) and replication...... stress in Saccharomyces cerevisiae. The Mre11 nuclease and the ATM-related Tel1 kinase are the first proteins detected at DSBs. Next, the Rfa1 single-strand DNA binding protein relocalizes to the break and recruits other key checkpoint proteins. Later and only in S and G2 phase, the homologous...... recombination machinery assembles at the site. Unlike the response to DSBs, Mre11 and recombination proteins are not recruited to hydroxyurea-stalled replication forks unless the forks collapse. The cellular response to DSBs and DNA replication stress is likely directed by the Mre11 complex detecting...

A Crystallographic Study of the Role of Sequence Context in Thymine Glycol Bypass by a Replicative DNA Polymerase Serendipitously Sheds Light on the Exonuclease Complex

Energy Technology Data Exchange (ETDEWEB)

Aller, Pierre; Duclos, Stéphanie; Wallace, Susan S.; Doublié, Sylvie (Vermont)

2012-06-27

Thymine glycol (Tg) is the most common oxidation product of thymine and is known to be a strong block to replicative DNA polymerases. A previously solved structure of the bacteriophage RB69 DNA polymerase (RB69 gp43) in complex with Tg in the sequence context 5'-G-Tg-G shed light on how Tg blocks primer elongation: The protruding methyl group of the oxidized thymine displaces the adjacent 5'-G, which can no longer serve as a template for primer elongation [Aller, P., Rould, M. A., Hogg, M, Wallace, S. S. and Doublie S. (2007). A structural rationale for stalling of a replicative DNA polymerase at the most common oxidative thymine lesion, thymine glycol. Proc. Natl. Acad. Sci. USA, 104, 814-818.]. Several studies showed that in the sequence context 5'-C-Tg-purine, Tg is more likely to be bypassed by Klenow fragment, an A-family DNA polymerase. We set out to investigate the role of sequence context in Tg bypass in a B-family polymerase and to solve the crystal structures of the bacteriophage RB69 DNA polymerase in complex with Tg-containing DNA in the three remaining sequence contexts: 5'-A-Tg-G, 5'-T-Tg-G, and 5'-C-Tg-G. A combination of several factors - including the associated exonuclease activity, the nature of the 3' and 5' bases surrounding Tg, and the cis-trans interconversion of Tg - influences Tg bypass. We also visualized for the first time the structure of a well-ordered exonuclease complex, allowing us to identify and confirm the role of key residues (Phe123, Met256, and Tyr257) in strand separation and in the stabilization of the primer strand in the exonuclease site.

Coordinating repair of oxidative DNA damage with transcription and replication

International Nuclear Information System (INIS)

Cooper, P.K.

2003-01-01

Transcription-coupled repair (TCR) preferentially removes DNA lesions from template strands of active genes. Defects in TCR, which acts both on lesions removed by nucleotide excision repair (NER) and on oxidative lesions removed by base excision repair (BER), underlie the fatal developmental disorder Cockayne syndrome. Although its detailed mechanism remains unknown, TCR involves recognition of a stalled RNA polymerase (RNAP), removal or remodeling of RNAP to allow access to the lesion, and recruitment of repair enzymes. At a minimum, these early steps require a non-enzymatic function of the multifunctional repair protein XPG, the CSB protein with ATP-dependent chromatin remodeling activity, and the TFIIH complex (including the XPB and XPD helicases) that is also required for basal transcription initiation and NER. XPG exists in the cell in a complex with TFIIH, and in vitro evidence has suggested that it interacts with CSB. To address the mechanism of TCR, we are characterizing protein-DNA and protein-protein interactions of XPG. We show that XPG preferentially binds to double-stranded DNA containing bubbles resembling in size the unpaired regions associated with transcription. Two distinct domains of XPG are required for the observed strong binding specificity and stability. XPG both interacts directly with CSB and synergistically binds with it to bubble DNA, and it strongly stimulates the bubble DNA-dependent ATPase activity of CSB. Significantly for TCR, XPG also interacts directly with RNAP II, binds both the protein and nucleic acid components (the R-loop) of a stalled RNA polymerase, and forms a ternary complex with CSB and the stalled RNAP. These results are consistent with the model that XPG and CSB jointly interact with the DNA/chromatin structure in the vicinity of the stalled transcriptional apparatus and with the transcriptional machinery itself to remodel the chromatin and either move or remodel the blocked RNA polymerase to expose the lesion

Stall/surge dynamics of a multi-stage air compressor in response to a load transient of a hybrid solid oxide fuel cell-gas turbine system

Science.gov (United States)

Azizi, Mohammad Ali; Brouwer, Jacob

2017-10-01

A better understanding of turbulent unsteady flows in gas turbine systems is necessary to design and control compressors for hybrid fuel cell-gas turbine systems. Compressor stall/surge analysis for a 4 MW hybrid solid oxide fuel cell-gas turbine system for locomotive applications is performed based upon a 1.7 MW multi-stage air compressor. Control strategies are applied to prevent operation of the hybrid SOFC-GT beyond the stall/surge lines of the compressor. Computational fluid dynamics tools are used to simulate the flow distribution and instabilities near the stall/surge line. The results show that a 1.7 MW system compressor like that of a Kawasaki gas turbine is an appropriate choice among the industrial compressors to be used in a 4 MW locomotive SOFC-GT with topping cycle design. The multi-stage radial design of the compressor enhances the ability of the compressor to maintain air flow rate during transient step-load changes. These transient step-load changes are exhibited in many potential applications for SOFC/GT systems. The compressor provides sustained air flow rate during the mild stall/surge event that occurs due to the transient step-load change that is applied, indicating that this type of compressor is well-suited for this hybrid application.

Diversification of DnaA dependency for DNA replication in cyanobacterial evolution.

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Ohbayashi, Ryudo; Watanabe, Satoru; Ehira, Shigeki; Kanesaki, Yu; Chibazakura, Taku; Yoshikawa, Hirofumi

2016-05-01

Regulating DNA replication is essential for all living cells. The DNA replication initiation factor DnaA is highly conserved in prokaryotes and is required for accurate initiation of chromosomal replication at oriC. DnaA-independent free-living bacteria have not been identified. The dnaA gene is absent in plastids and some symbiotic bacteria, although it is not known when or how DnaA-independent mechanisms were acquired. Here, we show that the degree of dependency of DNA replication on DnaA varies among cyanobacterial species. Deletion of the dnaA gene in Synechococcus elongatus PCC 7942 shifted DNA replication from oriC to a different site as a result of the integration of an episomal plasmid. Moreover, viability during the stationary phase was higher in dnaA disruptants than in wild-type cells. Deletion of dnaA did not affect DNA replication or cell growth in Synechocystis sp. PCC 6803 or Anabaena sp. PCC 7120, indicating that functional dependency on DnaA was already lost in some nonsymbiotic cyanobacterial lineages during diversification. Therefore, we proposed that cyanobacteria acquired DnaA-independent replication mechanisms before symbiosis and such an ancestral cyanobacterium was the sole primary endosymbiont to form a plastid precursor.

DNA replication and cancer: From dysfunctional replication origin activities to therapeutic opportunities.

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

2016-06-01

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 to promote genome integrity during DNA replication. This includes suppressing new replication origin firing, stabilization of replicating forks, and the safe restart of forks to prevent any loss of genetic information. Here, we describe mechanisms by which oncogenes can interfere with DNA replication thereby 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. Copyright © 2016 Elsevier Ltd. All rights reserved.

An unusual internal ribosomal entry site of inverted symmetry directs expression of a potato leafroll polerovirus replication-associated protein

Science.gov (United States)

Jaag, Hannah Miriam; Kawchuk, Lawrence; Rohde, Wolfgang; Fischer, Rainer; Emans, Neil; Prüfer, Dirk

2003-01-01

Potato leafroll polerovirus (PLRV) genomic RNA acts as a polycistronic mRNA for the production of proteins P0, P1, and P2 translated from the 5′-proximal half of the genome. Within the P1 coding region we identified a 5-kDa replication-associated protein 1 (Rap1) essential for viral multiplication. An internal ribosome entry site (IRES) with unusual structure and location was identified that regulates Rap1 translation. Core structural elements for internal ribosome entry include a conserved AUG codon and a downstream GGAGAGAGAGG motif with inverted symmetry. Reporter gene expression in potato protoplasts confirmed the internal ribosome entry function. Unlike known IRES motifs, the PLRV IRES is located completely within the coding region of Rap1 at the center of the PLRV genome. PMID:12835413

Power and precision of replicated helicopter surveys in mixed bushveld

Directory of Open Access Journals (Sweden)

B.K. Reilly

1998-07-01

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.

Replication origins oriGNAI3 and oriB of the mammalian AMPD2 locus nested in a region of straight DNA flanked by intrinsically bent DNA sites.

Science.gov (United States)

Balani, Valério Américo; de Lima Neto, Quirino Alves; Takeda, Karen Izumi; Gimenes, Fabrícia; Fiorini, Adriana; Debatisse, Michelle; Fernandez, Maria Aparecida

2010-11-01

The aim of this work was to determine whether intrinsically bent DNA sites are present at, or close to, the mammalian replication origins oriGNAI3 and oriB in the Chinese hamster AMPD2 locus. Using an electrophoretic mobility shift assay and in silico analysis, we located four intrinsically bent DNA sites (b1 to b4) in a fragment that contains the oriGNAI3 and one site (b5) proximal to oriB. The helical parameters show that each bent DNA site is curved in a left-handed superhelical writhe. A 2D projection of 3D fragment trajectories revealed that oriGNAI3 is located in a relatively straight segment flanked by bent sites b1 and b2, which map in previously identified Scaffold/Matrix Attachment Region. Sites b3 and b4 are located approximately 2 kb downstream and force the fragment into a strong closed loop structure. The b5 site is also located in an S/MAR that is found just downstream of oriB.

Crystallization and preliminary X-ray characterization of the eukaryotic replication terminator Reb1-Ter DNA complex.

Science.gov (United States)

Jaiswal, Rahul; Singh, Samarendra K; Bastia, Deepak; Escalante, Carlos R

2015-04-01

The Reb1 protein from Schizosaccharomyces pombe is a member of a family of proteins that control programmed replication termination and/or transcription termination in eukaryotic cells. These events occur at naturally occurring replication fork barriers (RFBs), where Reb1 binds to termination (Ter) DNA sites and coordinates the polar arrest of replication forks and transcription approaching in opposite directions. The Reb1 DNA-binding and replication-termination domain was expressed in Escherichia coli, purified and crystallized in complex with a 26-mer DNA Ter site. Batch crystallization under oil was required to produce crystals of good quality for data collection. Crystals grew in space group P2₁, with unit-cell parameters a = 68.9, b = 162.9, c = 71.1 Å, β = 94.7°. The crystals diffracted to a resolution of 3.0 Å. The crystals were mosaic and required two or three cycles of annealing. This study is the first to yield structural information about this important family of proteins and will provide insights into the mechanism of replication and transcription termination.

Power reduction and the radial limit of stall delay in revolving wings of different aspect ratio

NARCIS (Netherlands)

Kruyt, J.W.; Heijst, Van G.F.; Altshuler, D.L.; Lentink, David

2015-01-01

Airplanes and helicopters use high aspect ratio wings to reduce the power required to fly, but must operate at low angle of attack to prevent flow separation and stall. Animals capable of slow sustained flight, such as hummingbirds, have low aspect ratio wings and flap their wings at high angle

Aerodynamic loads calculation and analysis for large scale wind turbine based on combining BEM modified theory with dynamic stall model

Energy Technology Data Exchange (ETDEWEB)

Dai, J.C. [College of Mechanical and Electrical Engineering, Central South University, Changsha (China); School of Electromechanical Engineering, Hunan University of Science and Technology, Xiangtan (China); Hu, Y.P.; Liu, D.S. [School of Electromechanical Engineering, Hunan University of Science and Technology, Xiangtan (China); Long, X. [Hara XEMC Windpower Co., Ltd., Xiangtan (China)

2011-03-15

The aerodynamic loads for MW scale horizontal-axis wind turbines are calculated and analyzed in the established coordinate systems which are used to describe the wind turbine. In this paper, the blade element momentum (BEM) theory is employed and some corrections, such as Prandtl and Buhl models, are carried out. Based on the B-L semi-empirical dynamic stall (DS) model, a new modified DS model for NACA63-4xx airfoil is adopted. Then, by combing BEM modified theory with DS model, a set of calculation method of aerodynamic loads for large scale wind turbines is proposed, in which some influence factors such as wind shear, tower, tower and blade vibration are considered. The research results show that the presented dynamic stall model is good enough for engineering purpose; the aerodynamic loads are influenced by many factors such as tower shadow, wind shear, dynamic stall, tower and blade vibration, etc, with different degree; the single blade endures periodical changing loads but the variations of the rotor shaft power caused by the total aerodynamic torque in edgewise direction are very small. The presented study approach of aerodynamic loads calculation and analysis is of the university, and helpful for thorough research of loads reduction on large scale wind turbines. (author)

Fork rotation and DNA precatenation are restricted during DNA replication to prevent chromosomal instability.

Science.gov (United States)

Schalbetter, Stephanie A; Mansoubi, Sahar; Chambers, Anna L; Downs, Jessica A; Baxter, Jonathan

2015-08-18

Faithful genome duplication and inheritance require the complete resolution of all intertwines within the parental DNA duplex. This is achieved by topoisomerase action ahead of the replication fork or by fork rotation and subsequent resolution of the DNA precatenation formed. Although fork rotation predominates at replication termination, in vitro studies have suggested that it also occurs frequently during elongation. However, the factors that influence fork rotation and how rotation and precatenation may influence other replication-associated processes are unknown. Here we analyze the causes and consequences of fork rotation in budding yeast. We find that fork rotation and precatenation preferentially occur in contexts that inhibit topoisomerase action ahead of the fork, including stable protein-DNA fragile sites and termination. However, generally, fork rotation and precatenation are actively inhibited by Timeless/Tof1 and Tipin/Csm3. In the absence of Tof1/Timeless, excessive fork rotation and precatenation cause extensive DNA damage following DNA replication. With Tof1, damage related to precatenation is focused on the fragile protein-DNA sites where fork rotation is induced. We conclude that although fork rotation and precatenation facilitate unwinding in hard-to-replicate contexts, they intrinsically disrupt normal chromosome duplication and are therefore restricted by Timeless/Tipin.

Analysis of Vaneless Diffuser Stall Instability in a Centrifugal Compressor

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Elias Sundström

2017-11-01

Full Text Available Numerical simulations based on the large eddy simulation approach were conducted with the aim to explore vaneless diffuser rotating stall instability in a centrifugal compressor. The effect of the impeller blade passage was included as an inlet boundary condition with sufficiently low flow angle relative to the tangent to provoke the instability and cause circulation in the diffuser core flow. Flow quantities, velocity and pressure, were extracted to accumulate statistics for calculating mean velocity and mean Reynolds stresses in the wall-to-wall direction. The paper focuses on the assessment of the complex response of the system to the velocity perturbations imposed, the resulting pressure gradient and flow curvature effects.

Knockdown of RMI1 impairs DNA repair under DNA replication stress.

Science.gov (United States)

Xu, Chang; Fang, Lianying; Kong, Yangyang; Xiao, Changyan; Yang, Mengmeng; Du, Li-Qing; Liu, Qiang

2017-12-09

RMI1 (RecQ-mediated genome instability protein 1) forms a conserved BTR complex with BLM, Topo IIIα, and RMI2, and its absence causes genome instability. It has been revealed that RMI1 localizes to nuclear foci with BLM and Topo IIIα in response to replication stress, and that RMI1 functions downstream of BLM in promoting replication elongation. However, the precise functions of RMI1 during replication stress are not completely understood. Here we report that RMI1 knockdown cells are hypersensitive to hydroxyurea (HU). Using comet assay, we show that RMI1 knockdown cells exhibit accumulation of broken DNAs after being released from HU treatment. Moreover, we demonstrate that RMI1 facilitates the recovery from activated checkpoint and resuming the cell cycle after replicative stress. Surprisingly, loss of RMI1 results in a failure of RAD51 loading onto DNA damage sites. These findings reveal the importance of RMI1 in response to replication stress, which could explain the molecular basis for its function in maintaining genome integrity. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Distribution of DNA replication proteins in Drosophila cells

Science.gov (United States)

Easwaran, Hariharan P; Leonhardt, Heinrich; Cardoso, M Cristina

2007-01-01

Background DNA replication in higher eukaryotic cells is organized in discrete subnuclear sites called replication foci (RF). During the S phase, most replication proteins assemble at the RF by interacting with PCNA via a PCNA binding domain (PBD). This has been shown to occur for many mammalian replication proteins, but it is not known whether this mechanism is conserved in evolution. Results Fluorescent fusions of mammalian replication proteins, Dnmt1, HsDNA Lig I and HsPCNA were analyzed for their ability to target to RF in Drosophila cells. Except for HsPCNA, none of the other proteins and their deletions showed any accumulation at RF in Drosophila cells. We hypothesized that in Drosophila cells there might be some other peptide sequence responsible for targeting proteins to RF. To test this, we identified the DmDNA Lig I and compared the protein sequence with HsDNA Lig I. The two orthologs shared the PBD suggesting a functionally conserved role for this domain in the Drosophila counterpart. A series of deletions of DmDNA Lig I were analyzed for their ability to accumulate at RF in Drosophila and mammalian cells. Surprisingly, no accumulation at RF was observed in Drosophila cells, while in mammalian cells DmDNA Lig I accumulated at RF via its PBD. Further, GFP fusions with the PBD domains from Dnmt1, HsDNA Lig I and DmDNA Lig I, were able to target to RF only in mammalian cells but not in Drosophila cells. Conclusion We show that S phase in Drosophila cells is characterized by formation of RF marked by PCNA like in mammalian cells. However, other than PCNA none of the replication proteins and their deletions tested here showed accumulation at RF in Drosophila cells while the same proteins and deletions are capable of accumulating at RF in mammalian cells. We hypothesize that unlike mammalian cells, in Drosophila cells, replication proteins do not form long-lasting interactions with the replication machinery, and rather perform their functions via very

DNA Sequences Proximal to Human Mitochondrial DNA Deletion Breakpoints Prevalent in Human Disease Form G-quadruplexes, a Class of DNA Structures Inefficiently Unwound by the Mitochondrial Replicative Twinkle Helicase*

Science.gov (United States)

Bharti, Sanjay Kumar; Sommers, Joshua A.; Zhou, Jun; Kaplan, Daniel L.; Spelbrink, Johannes N.; Mergny, Jean-Louis; Brosh, Robert M.

2014-01-01

Mitochondrial DNA deletions are prominent in human genetic disorders, cancer, and aging. It is thought that stalling of the mitochondrial replication machinery during DNA synthesis is a prominent source of mitochondrial genome instability; however, the precise molecular determinants of defective mitochondrial replication are not well understood. In this work, we performed a computational analysis of the human mitochondrial genome using the “Pattern Finder” G-quadruplex (G4) predictor algorithm to assess whether G4-forming sequences reside in close proximity (within 20 base pairs) to known mitochondrial DNA deletion breakpoints. We then used this information to map G4P sequences with deletions characteristic of representative mitochondrial genetic disorders and also those identified in various cancers and aging. Circular dichroism and UV spectral analysis demonstrated that mitochondrial G-rich sequences near deletion breakpoints prevalent in human disease form G-quadruplex DNA structures. A biochemical analysis of purified recombinant human Twinkle protein (gene product of c10orf2) showed that the mitochondrial replicative helicase inefficiently unwinds well characterized intermolecular and intramolecular G-quadruplex DNA substrates, as well as a unimolecular G4 substrate derived from a mitochondrial sequence that nests a deletion breakpoint described in human renal cell carcinoma. Although G4 has been implicated in the initiation of mitochondrial DNA replication, our current findings suggest that mitochondrial G-quadruplexes are also likely to be a source of instability for the mitochondrial genome by perturbing the normal progression of the mitochondrial replication machinery, including DNA unwinding by Twinkle helicase. PMID:25193669

Loss of Hda activity stimulates replication initiation from I-box, but not R4 mutant origins in Escherichia coli.

Science.gov (United States)

Riber, Leise; Fujimitsu, Kazuyuki; Katayama, Tsutomu; Løbner-Olesen, Anders

2009-01-01

Initiation of chromosome replication in Escherichia coli is limited by the initiator protein DnaA associated with ATP. Within the replication origin, binding sites for DnaA associated with ATP or ADP (R boxes) and the DnaA(ATP) specific sites (I-boxes, tau-boxes and 6-mer sites) are found. We analysed chromosome replication of cells carrying mutations in conserved regions of oriC. Cells carrying mutations in DnaA-boxes I2, I3, R2, R3 and R5 as well as FIS and IHF binding sites resembled wild-type cells with respect to origin concentration. Initiation of replication in these mutants occurred in synchrony or with slight asynchrony only. Furthermore, lack of Hda stimulated initiation in all these mutants. The DnaA(ATP) containing complex that leads to initiation can therefore be formed in the absence of several of the origin DnaA binding sites including both DnaA(ATP) specific I-boxes. However, competition between I-box mutant and wild-type origins, revealed a positive role of I-boxes on initiation. On the other hand, mutations affecting DnaA-box R4 were found to be compromised for initiation and could not be augmented by an increase in cellular DnaA(ATP)/DnaA(ADP) ratio. Compared with the sites tested here, R4 therefore seems to contribute to initiation most critically.

Replication protocol analysis: a method for the study of real-world design thinking

DEFF Research Database (Denmark)

Galle, Per; Kovacs, L. B.

1996-01-01

’ is refined into a method called ‘replication protocol analysis’ (RPA), and discussed from a methodological perspective of design research. It is argued that for the study of real-world design thinking this method offers distinct advantages over traditional ‘design protocol analysis’, which seeks to capture......Given the brief of an architectural competition on site planning, and the design awarded the first prize, the first author (trained as an architect but not a participant in the competition) produced a line of reasoning that might have led from brief to design. In the paper, such ‘design replication...... the designer’s authentic line of reasoning. To illustrate how RPA can be used, the site planning case is briefly presented, and part of the replicated line of reasoning analysed. One result of the analysis is a glimpse of a ‘logic of design’; another is an insight which sheds new light on Darke’s classical...

Timeless links replication termination to mitotic kinase activation.

Directory of Open Access Journals (Sweden)

Jayaraju Dheekollu

2011-05-01

Full Text Available The mechanisms that coordinate the termination of DNA replication with progression through mitosis are not completely understood. The human Timeless protein (Tim associates with S phase replication checkpoint proteins Claspin and Tipin, and plays an important role in maintaining replication fork stability at physical barriers, like centromeres, telomeres and ribosomal DNA repeats, as well as at termination sites. We show here that human Tim can be isolated in a complex with mitotic entry kinases CDK1, Auroras A and B, and Polo-like kinase (Plk1. Plk1 bound Tim directly and colocalized with Tim at a subset of mitotic structures in M phase. Tim depletion caused multiple mitotic defects, including the loss of sister-chromatid cohesion, loss of mitotic spindle architecture, and a failure to exit mitosis. Tim depletion caused a delay in mitotic kinase activity in vivo and in vitro, as well as a reduction in global histone H3 S10 phosphorylation during G2/M phase. Tim was also required for the recruitment of Plk1 to centromeric DNA and formation of catenated DNA structures at human centromere alpha satellite repeats. Taken together, these findings suggest that Tim coordinates mitotic kinase activation with termination of DNA replication.

An Adenovirus DNA Replication Factor, but Not Incoming Genome Complexes, Targets PML Nuclear Bodies.

Science.gov (United States)

Komatsu, Tetsuro; Nagata, Kyosuke; Wodrich, Harald

2016-02-01

Promyelocytic leukemia protein nuclear bodies (PML-NBs) are subnuclear domains implicated in cellular antiviral responses. Despite the antiviral activity, several nuclear replicating DNA viruses use the domains as deposition sites for the incoming viral genomes and/or as sites for viral DNA replication, suggesting that PML-NBs are functionally relevant during early viral infection to establish productive replication. Although PML-NBs and their components have also been implicated in the adenoviral life cycle, it remains unclear whether incoming adenoviral genome complexes target PML-NBs. Here we show using immunofluorescence and live-cell imaging analyses that incoming adenovirus genome complexes neither localize at nor recruit components of PML-NBs during early phases of infection. We further show that the viral DNA binding protein (DBP), an early expressed viral gene and essential DNA replication factor, independently targets PML-NBs. We show that DBP oligomerization is required to selectively recruit the PML-NB components Sp100 and USP7. Depletion experiments suggest that the absence of one PML-NB component might not affect the recruitment of other components toward DBP oligomers. Thus, our findings suggest a model in which an adenoviral DNA replication factor, but not incoming viral genome complexes, targets and modulates PML-NBs to support a conducive state for viral DNA replication and argue against a generalized concept that PML-NBs target incoming viral genomes. The immediate fate upon nuclear delivery of genomes of incoming DNA viruses is largely unclear. Early reports suggested that incoming genomes of herpesviruses are targeted and repressed by PML-NBs immediately upon nuclear import. Genome localization and/or viral DNA replication has also been observed at PML-NBs for other DNA viruses. Thus, it was suggested that PML-NBs may immediately sense and target nuclear viral genomes and hence serve as sites for deposition of incoming viral genomes and

Repair replication in replicating and nonreplicating DNA after irradiation with uv light

Energy Technology Data Exchange (ETDEWEB)

Slor, H.; Cleaver, J.E.

1978-06-01

Ultraviolet light induces more pyrimidine dimers and more repair replication in DNA that replicates within 2 to 3 h of irradiation than in DNA that does not replicate during this period. This difference may be due to special conformational changes in DNA and chromatin that might be associated with semiconservative DNA replication.

Active flow control of the laminar separation bubble on a plunging airfoil near stall

Science.gov (United States)

Pande, Arth; Agate, Mark; Little, Jesse; Fasel, Hermann

2017-11-01

The effects of small amplitude (A/c = 0.048) high frequency (πfc/U∞ = 0.70) plunging motion on the X-56A airfoil are examined experimentally at Re = 200,000 for 12° angle of attack (CL,MAX = 12.25°) . The purpose of this research is to study the aerodynamic influence of structural motion when the wing is vibrating close to its eigenfrequency near static stall. Specific focus is placed on the laminar separation bubble (LSB) near the leading edge and its control via plasma actuation. In the baseline case, the leading edge bubble bursts during the oscillation cycle causing moment stall. A collaborative computational effort has shown that small amplitude forcing at a frequency that is most amplified by the primary instability of the LSB (FLSB+= 1, Fc+= 52) generates coherent spanwise vortices that entrain freestream momentum, thus reducing separation all while maintaining a laminar flow state. Results (PIV and surface pressure) indicate that a similar control mechanism is effective in the experiments. This is significant given the existence of freestream turbulence in the wind tunnel which has been shown to limit the efficacy of this active flow control technique in a model problem using Direct Numerical Simulation. The implications of these results are discussed.

Human Parvovirus B19 Utilizes Cellular DNA Replication Machinery for Viral DNA Replication.

Science.gov (United States)

Zou, Wei; Wang, Zekun; Xiong, Min; Chen, Aaron Yun; Xu, Peng; Ganaie, Safder S; Badawi, Yomna; Kleiboeker, Steve; Nishimune, Hiroshi; Ye, Shui Qing; Qiu, Jianming

2018-03-01

Human parvovirus B19 (B19V) infection of human erythroid progenitor cells (EPCs) induces a DNA damage response and cell cycle arrest at late S phase, which facilitates viral DNA replication. However, it is not clear exactly which cellular factors are employed by this single-stranded DNA virus. Here, we used microarrays to systematically analyze the dynamic transcriptome of EPCs infected with B19V. We found that DNA metabolism, DNA replication, DNA repair, DNA damage response, cell cycle, and cell cycle arrest pathways were significantly regulated after B19V infection. Confocal microscopy analyses revealed that most cellular DNA replication proteins were recruited to the centers of viral DNA replication, but not the DNA repair DNA polymerases. Our results suggest that DNA replication polymerase δ and polymerase α are responsible for B19V DNA replication by knocking down its expression in EPCs. We further showed that although RPA32 is essential for B19V DNA replication and the phosphorylated forms of RPA32 colocalized with the replicating viral genomes, RPA32 phosphorylation was not necessary for B19V DNA replication. Thus, this report provides evidence that B19V uses the cellular DNA replication machinery for viral DNA replication. IMPORTANCE Human parvovirus B19 (B19V) infection can cause transient aplastic crisis, persistent viremia, and pure red cell aplasia. In fetuses, B19V infection can result in nonimmune hydrops fetalis and fetal death. These clinical manifestations of B19V infection are a direct outcome of the death of human erythroid progenitors that host B19V replication. B19V infection induces a DNA damage response that is important for cell cycle arrest at late S phase. Here, we analyzed dynamic changes in cellular gene expression and found that DNA metabolic processes are tightly regulated during B19V infection. Although genes involved in cellular DNA replication were downregulated overall, the cellular DNA replication machinery was tightly

Simulation model of a transient fault controller for an active-stall wind turbine

Energy Technology Data Exchange (ETDEWEB)

Jauch, C.; Soerensen, P.; Bak Jensen, B.

2005-01-01

This paper describes the simulation model of a controller that enables an active-stall wind turbine to ride through transient faults. The simulated wind turbine is connected to a simple model of a power system. Certain fault scenarios are specified and the turbine shall be able to sustain operation in case of such faults. The design of the controller is described and its performance assessed by simulations. The control strategies are explained and the behaviour of the turbine discussed. (author)

Active unsteady aerodynamic suppression of rotating stall in an incompressible flow centrifugal compressor with vaned diffuser

Science.gov (United States)

Lawless, Patrick B.; Fleeter, Sanford

1991-01-01

A mathematical model is developed to analyze the suppression of rotating stall in an incompressible flow centrifugal compressor with a vaned diffuser, thereby addressing the important need for centrifugal compressor rotating stall and surge control. In this model, the precursor to to instability is a weak rotating potential velocity perturbation in the inlet flow field that eventually develops into a finite disturbance. To suppress the growth of this potential disturbance, a rotating control vortical velocity disturbance is introduced into the impeller inlet flow. The effectiveness of this control is analyzed by matching the perturbation pressure in the compressor inlet and exit flow fields with a model for the unsteady behavior of the compressor. To demonstrate instability control, this model is then used to predict the control effectiveness for centrifugal compressor geometries based on a low speed research centrifugal compressor. These results indicate that reductions of 10 to 15 percent in the mean inlet flow coefficient at instability are possible with control waveforms of half the magnitude of the total disturbance at the inlet.

A Beddoes-Leishman type dynamic stall model in state-space and indicial formulations[Wind turbines

Energy Technology Data Exchange (ETDEWEB)

Hansen, M.H.; Gaunaa, M.; Aagaard Madsen, H.

2004-06-01

This report contains a description of a Beddoes-Leishman type dynamic stall model in both a state-space and an indicial function formulation. The m odel predicts the unsteady aerodynamic foreces and moment on an airfoil section undergoing arbitrary motion in heavy, lead-lag, and pitch. The model includes the effects of shed vorticity from the trailing edge (Theodorsen Theory), and the effects of an instationary trailing edge separation point. The governing equations of the model are nonlinear, and they are linearized about a steady state for application in stability analyzes. A validation is carried out by comparing the response of the model with inviscid solutions and observing the general behavior of the model using known airfoil data as input. The proposed dyanmic model gives results identical to inviscid solutions within the attached-flow region; and it exhibits the expected dynamic features, such as overshoot of the lift, in the stall region. The linearized model is shown to give identical results to the full model for small amplitude oscillations. furthermore, it is shown that the response of finite thickness airfoils can be reproduced to a high accuracy by the use of specific inviscid response functions. (au)

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

Mms1 binds to G-rich regions in Saccharomyces cerevisiae and influences replication and genome stability

NARCIS (Netherlands)

Wanzek, Katharina; Schwindt, Eike; Capra, John A.; Paeschke, Katrin

2017-01-01

The regulation of replication is essential to preserve genome integrity. Mms1 is part of the E3 ubiquitin ligase complex that is linked to replication fork progression. By identifying Mms1 binding sites genome-wide in Saccharomyces cerevisiae we connected Mms1 function to genome integrity and

X-irradiation affects all DNA replication intermediates when inhibiting replication initiation

International Nuclear Information System (INIS)

Loenn, U.; Karolinska Hospital, Stockholm

1982-01-01

When a human melanoma line was irradiated with 10 Gy, there was, after 30 to 60 min, a gradual reduction in the DNA replication rate. Ten to twelve hours after the irradiation, the DNA replication had returned to near normal rate. The results showed tht low dose-rate X-irradiation inhibits preferentially the formation of small DNA replication intermediates. There is no difference between the inhibition of these replication intermediates formed only in the irradiated cells and those formed also in untreated cells. (U.K.)

Role of DNA Repair Factor Xeroderma Pigmentosum Protein Group C in Response to Replication Stress As Revealed by DNA Fragile Site Affinity Chromatography and Quantitative Proteomics.

Science.gov (United States)

Beresova, Lucie; Vesela, Eva; Chamrad, Ivo; Voller, Jiri; Yamada, Masayuki; Furst, Tomas; Lenobel, Rene; Chroma, Katarina; Gursky, Jan; Krizova, Katerina; Mistrik, Martin; Bartek, Jiri

2016-12-02

Replication stress (RS) fuels genomic instability and cancer development and may contribute to aging, raising the need to identify factors involved in cellular responses to such stress. Here, we present a strategy for identification of factors affecting the maintenance of common fragile sites (CFSs), which are genomic loci that are particularly sensitive to RS and suffer from increased breakage and rearrangements in tumors. A DNA probe designed to match the high flexibility island sequence typical for the commonly expressed CFS (FRA16D) was used as specific DNA affinity bait. Proteins significantly enriched at the FRA16D fragment under normal and replication stress conditions were identified using stable isotope labeling of amino acids in cell culture-based quantitative mass spectrometry. The identified proteins interacting with the FRA16D fragment included some known CFS stabilizers, thereby validating this screening approach. Among the hits from our screen so far not implicated in CFS maintenance, we chose Xeroderma pigmentosum protein group C (XPC) for further characterization. XPC is a key factor in the DNA repair pathway known as global genomic nucleotide excision repair (GG-NER), a mechanism whose several components were enriched at the FRA16D fragment in our screen. Functional experiments revealed defective checkpoint signaling and escape of DNA replication intermediates into mitosis and the next generation of XPC-depleted cells exposed to RS. Overall, our results provide insights into an unexpected biological role of XPC in response to replication stress and document the power of proteomics-based screening strategies to elucidate mechanisms of pathophysiological significance.

The cholesterol, fatty acid and triglyceride synthesis pathways regulated by site 1 protease (S1P) are required for efficient replication of severe fever with thrombocytopenia syndrome virus.

Science.gov (United States)

Urata, Shuzo; Uno, Yukiko; Kurosaki, Yohei; Yasuda, Jiro

2018-06-12

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging infectious disease caused by the SFTS virus (SFTSV), which has a high mortality rate. Currently, no licensed vaccines or therapeutic agents have been approved for use against SFTSV infection. Here, we report that the cholesterol, fatty acid, and triglyceride synthesis pathways regulated by S1P is involved in SFTSV replication, using CHO-K1 cell line (SRD-12B) that is deficient in site 1 protease (S1P) enzymatic activity, PF-429242, a small compound targeting S1P enzymatic activity, and Fenofibrate and Lovastatin, which inhibit triglyceride and cholesterol synthesis, respectively. These results enhance our understanding of the SFTSV replication mechanism and may contribute to the development of novel therapies for SFTSV infection. Copyright © 2018. Published by Elsevier Inc.

Chromatin Structure and Replication Origins: Determinants Of Chromosome Replication And Nuclear Organization

Science.gov (United States)

Smith, Owen K.; Aladjem, Mirit I.

2014-01-01

The DNA replication program is, in part, determined by the epigenetic landscape that governs local chromosome architecture and directs chromosome duplication. Replication must coordinate with other biochemical processes occurring concomitantly on chromatin, such as transcription and remodeling, to insure accurate duplication of both genetic and epigenetic features and to preserve genomic stability. The importance of genome architecture and chromatin looping in coordinating cellular processes on chromatin is illustrated by two recent sets of discoveries. First, chromatin-associated proteins that are not part of the core replication machinery were shown to affect the timing of DNA replication. These chromatin-associated proteins could be working in concert, or perhaps in competition, with the transcriptional machinery and with chromatin modifiers to determine the spatial and temporal organization of replication initiation events. Second, epigenetic interactions are mediated by DNA sequences that determine chromosomal replication. In this review we summarize recent findings and current models linking spatial and temporal regulation of the replication program with epigenetic signaling. We discuss these issues in the context of the genome’s three-dimensional structure with an emphasis on events occurring during the initiation of DNA replication. PMID:24905010

Prelife catalysts and replicators

OpenAIRE

Ohtsuki, Hisashi; Nowak, Martin A.

2009-01-01

Life is based on replication and evolution. But replication cannot be taken for granted. We must ask what there was prior to replication and evolution. How does evolution begin? We have proposed prelife as a generative system that produces information and diversity in the absence of replication. We model prelife as a binary soup of active monomers that form random polymers. ‘Prevolutionary’ dynamics can have mutation and selection prior to replication. Some sequences might have catalytic acti...

Recent advances in the genome-wide study of DNA replication origins in yeast

Directory of Open Access Journals (Sweden)

Chong ePeng

2015-02-01

Full Text Available DNA replication, one of the central events in the cell cycle, is the basis of biological inheritance. In order to be duplicated, a DNA double helix must be opened at defined sites, which are called DNA replication origins (ORIs. Unlike in bacteria, where replication initiates from a single replication origin, multiple origins are utilized in the eukaryotic genome. Among them, the ORIs in budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe have been best characterized. In recent years, advances in DNA microarray and next-generation sequencing technologies have increased the number of yeast species involved in ORIs research dramatically. The ORIs in some nonconventional yeast species such as Kluyveromyces lactis and Pichia pastoris have also been genome-widely identified. Relevant databases of replication origins in yeast were constructed, then the comparative genomic analysis can be carried out. Here, we review several experimental approaches that have been used to map replication origins in yeast and some of the available web resources related to yeast ORIs. We also discuss the sequence characteristics and chromosome structures of ORIs in the four yeast species, which can be utilized to improve the replication origins prediction.

Recent advances in the genome-wide study of DNA replication origins in yeast

Science.gov (United States)

Peng, Chong; Luo, Hao; Zhang, Xi; Gao, Feng

2015-01-01

DNA replication, one of the central events in the cell cycle, is the basis of biological inheritance. In order to be duplicated, a DNA double helix must be opened at defined sites, which are called DNA replication origins (ORIs). Unlike in bacteria, where replication initiates from a single replication origin, multiple origins are utilized in the eukaryotic genomes. Among them, the ORIs in budding yeast Saccharomyces cerevisiae and the fission yeast Schizosaccharomyces pombe have been best characterized. In recent years, advances in DNA microarray and next-generation sequencing technologies have increased the number of yeast species involved in ORIs research dramatically. The ORIs in some non-conventional yeast species such as Kluyveromyces lactis and Pichia pastoris have also been genome-widely identified. Relevant databases of replication origins in yeast were constructed, then the comparative genomic analysis can be carried out. Here, we review several experimental approaches that have been used to map replication origins in yeast and some of the available web resources related to yeast ORIs. We also discuss the sequence characteristics and chromosome structures of ORIs in the four yeast species, which can be utilized to improve yeast replication origins prediction. PMID:25745419

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.

Phosphorylation of Large T Antigen Regulates Merkel Cell Polyomavirus Replication

International Nuclear Information System (INIS)

Diaz, Jason; Wang, Xin; Tsang, Sabrina H.; Jiao, Jing; You, Jianxin

2014-01-01

Merkel Cell Polyomavirus (MCPyV) was recently discovered as a novel human polyomavirus that is associated with ~80% of Merkel Cell Carcinomas. The Large Tumor antigen (LT) is an early viral protein which has a variety of functions, including manipulation of the cell cycle and initiating viral DNA replication. Phosphorylation plays a critical regulatory role for polyomavirus LT proteins, but no investigation of MCPyV LT phosphorylation has been performed to date. In this report mass spectrometry analysis reveals three unique phosphorylation sites: T271, T297 and T299. In vivo replication assays confirm that phosphorylation of T271 does not play a role in viral replication, while modification at T297 and T299 have dramatic and opposing effects on LT’s ability to initiate replication from the viral origin. We test these mutants for their ability to bind, unwind, and act as a functional helicase at the viral origin. These studies provide a framework for understanding how phosphorylation of LT may dynamically regulate viral replication. Although the natural host cell of MCPyV has not yet been established, this work provides a foundation for understanding how LT activity is regulated and provides tools for better exploring this regulation in both natural host cells and Merkel cells

Phosphorylation of Large T Antigen Regulates Merkel Cell Polyomavirus Replication

Energy Technology Data Exchange (ETDEWEB)

Diaz, Jason; Wang, Xin; Tsang, Sabrina H. [Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104 (United States); Jiao, Jing [Department of Pathology and Laboratory Medicine, Children’s Hospital of Philadelphia, Philadelphia, PA 19104 (United States); You, Jianxin, E-mail: jianyou@mail.med.upenn.edu [Department of Microbiology, University of Pennsylvania Perelman School of Medicine, Philadelphia, PA 19104 (United States)

2014-07-08

Merkel Cell Polyomavirus (MCPyV) was recently discovered as a novel human polyomavirus that is associated with ~80% of Merkel Cell Carcinomas. The Large Tumor antigen (LT) is an early viral protein which has a variety of functions, including manipulation of the cell cycle and initiating viral DNA replication. Phosphorylation plays a critical regulatory role for polyomavirus LT proteins, but no investigation of MCPyV LT phosphorylation has been performed to date. In this report mass spectrometry analysis reveals three unique phosphorylation sites: T271, T297 and T299. In vivo replication assays confirm that phosphorylation of T271 does not play a role in viral replication, while modification at T297 and T299 have dramatic and opposing effects on LT’s ability to initiate replication from the viral origin. We test these mutants for their ability to bind, unwind, and act as a functional helicase at the viral origin. These studies provide a framework for understanding how phosphorylation of LT may dynamically regulate viral replication. Although the natural host cell of MCPyV has not yet been established, this work provides a foundation for understanding how LT activity is regulated and provides tools for better exploring this regulation in both natural host cells and Merkel cells.

SARS-coronavirus replication is supported by a reticulovesicular network of modified endoplasmic reticulum.

Directory of Open Access Journals (Sweden)

Kèvin Knoops

2008-09-01

Full Text Available Positive-strand RNA viruses, a large group including human pathogens such as SARS-coronavirus (SARS-CoV, replicate in the cytoplasm of infected host cells. Their replication complexes are commonly associated with modified host cell membranes. Membrane structures supporting viral RNA synthesis range from distinct spherular membrane invaginations to more elaborate webs of packed membranes and vesicles. Generally, their ultrastructure, morphogenesis, and exact role in viral replication remain to be defined. Poorly characterized double-membrane vesicles (DMVs were previously implicated in SARS-CoV RNA synthesis. We have now applied electron tomography of cryofixed infected cells for the three-dimensional imaging of coronavirus-induced membrane alterations at high resolution. Our analysis defines a unique reticulovesicular network of modified endoplasmic reticulum that integrates convoluted membranes, numerous interconnected DMVs (diameter 200-300 nm, and "vesicle packets" apparently arising from DMV merger. The convoluted membranes were most abundantly immunolabeled for viral replicase subunits. However, double-stranded RNA, presumably revealing the site of viral RNA synthesis, mainly localized to the DMV interior. Since we could not discern a connection between DMV interior and cytosol, our analysis raises several questions about the mechanism of DMV formation and the actual site of SARS-CoV RNA synthesis. Our data document the extensive virus-induced reorganization of host cell membranes into a network that is used to organize viral replication and possibly hide replicating RNA from antiviral defense mechanisms. Together with biochemical studies of the viral enzyme complex, our ultrastructural description of this "replication network" will aid to further dissect the early stages of the coronavirus life cycle and its virus-host interactions.

Wide area data replication in an ITER-relevant data environment

International Nuclear Information System (INIS)

Centioli, C.; Iannone, F.; Panella, M.; Vitale, V.; Bracco, G.; Guadagni, R.; Migliori, S.; Steffe, M.; Eccher, S.; Maslennikov, A.; Mililotti, M.; Molowny, M.; Palumbo, G.; Carboni, M.

2005-01-01

The next generation of tokamak experiments will require a new way of approaching data sharing issues among fusion organizations. In the fusion community, many researchers at different worldwide sites will analyse data produced by International Thermonuclear Experimental Reactor (ITER), wherever it will be built. In this context, an efficient availability of the data in the sites where the computational resources are located becomes a major architectural issue for the deployment of ITER computational infrastructure. The approach described in this paper goes beyond the usual site-centric model mainly devoted to granting access exclusively to experimental data stored at the device sites. To this aim, we propose a new data replication architecture relying on a wide area network, based on a Master/Slave model and on synchronization techniques producing mirrored data sites. In this architecture, data replication will affect large databases (TB) as well as large UNIX-like file systems, using open source-based software components, namely MySQL, as database management system, and RSYNC and BBFTP for data transfer. A test-bed has been set up to evaluate the performance of the software components underlying the proposed architecture. The test-bed hardware layout deploys a cluster of four Dual-Xeon Supermicro each with a raid array of 1 TB. High performance network line (1 Gbit over 400 km) provides the infrastructure to test the components on a wide area network. The results obtained will be thoroughly discussed

Chromosomal context and replication properties of ARS plasmids in ...

Indian Academy of Sciences (India)

2015-11-28

Nov 28, 2015 ... plasmid but only a subset of them functions as replication origins in their ... except that they are rich in A + T content (As on one strand and Ts .... different unique, terminal, PCR-generated restriction sites used for cloning each fragment are ..... Hall TA 1999 BioEdit: a user-friendly biological sequence align-.

Epigenetically-inherited centromere and neocentromere DNA replicates earliest in S-phase.

Directory of Open Access Journals (Sweden)

Amnon Koren

2010-08-01

Full Text Available Eukaryotic centromeres are maintained at specific chromosomal sites over many generations. In the budding yeast Saccharomyces cerevisiae, centromeres are genetic elements defined by a DNA sequence that is both necessary and sufficient for function; whereas, in most other eukaryotes, centromeres are maintained by poorly characterized epigenetic mechanisms in which DNA has a less definitive role. Here we use the pathogenic yeast Candida albicans as a model organism to study the DNA replication properties of centromeric DNA. By determining the genome-wide replication timing program of the C. albicans genome, we discovered that each centromere is associated with a replication origin that is the first to fire on its respective chromosome. Importantly, epigenetic formation of new ectopic centromeres (neocentromeres was accompanied by shifts in replication timing, such that a neocentromere became the first to replicate and became associated with origin recognition complex (ORC components. Furthermore, changing the level of the centromere-specific histone H3 isoform led to a concomitant change in levels of ORC association with centromere regions, further supporting the idea that centromere proteins determine origin activity. Finally, analysis of centromere-associated DNA revealed a replication-dependent sequence pattern characteristic of constitutively active replication origins. This strand-biased pattern is conserved, together with centromere position, among related strains and species, in a manner independent of primary DNA sequence. Thus, inheritance of centromere position is correlated with a constitutively active origin of replication that fires at a distinct early time. We suggest a model in which the distinct timing of DNA replication serves as an epigenetic mechanism for the inheritance of centromere position.

Replication stress interferes with histone recycling and predeposition marking of new histones

DEFF Research Database (Denmark)

Jasencakova, Zuzana; Scharf, Annette N D; Ask, Katrine

2010-01-01

To restore chromatin on new DNA during replication, recycling of histones evicted ahead of the fork is combined with new histone deposition. The Asf1 histone chaperone, which buffers excess histones under stress, is a key player in this process. Yet how histones handled by human Asf1 are modified...... remains unclear. Here we identify marks on histones H3-H4 bound to Asf1 and changes induced upon replication stress. In S phase, distinct cytosolic and nuclear Asf1b complexes show ubiquitous H4K5K12diAc and heterogeneous H3 marks, including K9me1, K14ac, K18ac, and K56ac. Upon acute replication arrest......, the predeposition mark H3K9me1 and modifications typical of chromatin accumulate in Asf1 complexes. In parallel, ssDNA is generated at replication sites, consistent with evicted histones being trapped with Asf1. During recovery, histones stored with Asf1 are rapidly used as replication resumes. This shows...

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…

Global conformational dynamics of a Y-family DNA polymerase during catalysis.

Directory of Open Access Journals (Sweden)

Cuiling Xu

2009-10-01

Full Text Available Replicative DNA polymerases are stalled by damaged DNA while the newly discovered Y-family DNA polymerases are recruited to rescue these stalled replication forks, thereby enhancing cell survival. The Y-family DNA polymerases, characterized by low fidelity and processivity, are able to bypass different classes of DNA lesions. A variety of kinetic and structural studies have established a minimal reaction pathway common to all DNA polymerases, although the conformational intermediates are not well defined. Furthermore, the identification of the rate-limiting step of nucleotide incorporation catalyzed by any DNA polymerase has been a matter of long debate. By monitoring time-dependent fluorescence resonance energy transfer (FRET signal changes at multiple sites in each domain and DNA during catalysis, we present here a real-time picture of the global conformational transitions of a model Y-family enzyme: DNA polymerase IV (Dpo4 from Sulfolobus solfataricus. Our results provide evidence for a hypothetical DNA translocation event followed by a rapid protein conformational change prior to catalysis and a subsequent slow, post-chemistry protein conformational change. Surprisingly, the DNA translocation step was induced by the binding of a correct nucleotide. Moreover, we have determined the directions, rates, and activation energy barriers of the protein conformational transitions, which indicated that the four domains of Dpo4 moved in a synchronized manner. These results showed conclusively that a pre-chemistry conformational change associated with domain movements was too fast to be the rate-limiting step. Rather, the rearrangement of active site residues limited the rate of correct nucleotide incorporation. Collectively, the conformational dynamics of Dpo4 offer insights into how the inter-domain movements are related to enzymatic function and their concerted interactions with other proteins at the replication fork.

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.

An evaluation of the Bayesian approach to fitting the N-mixture model for use with pseudo-replicated count data

Science.gov (United States)

Toribo, S.G.; Gray, B.R.; Liang, S.

2011-01-01

The N-mixture model proposed by Royle in 2004 may be used to approximate the abundance and detection probability of animal species in a given region. In 2006, Royle and Dorazio discussed the advantages of using a Bayesian approach in modelling animal abundance and occurrence using a hierarchical N-mixture model. N-mixture models assume replication on sampling sites, an assumption that may be violated when the site is not closed to changes in abundance during the survey period or when nominal replicates are defined spatially. In this paper, we studied the robustness of a Bayesian approach to fitting the N-mixture model for pseudo-replicated count data. Our simulation results showed that the Bayesian estimates for abundance and detection probability are slightly biased when the actual detection probability is small and are sensitive to the presence of extra variability within local sites.

A stochastic model for the simulation of wind turbine blades in static stall

DEFF Research Database (Denmark)

Bertagnolio, Franck; Rasmussen, Flemming; Sørensen, Niels N.

2010-01-01

The aim of this work is to improve aeroelastic simulation codes by accounting for the unsteady aerodynamic forces that a blade experiences in static stall. A model based on a spectral representation of the aerodynamic lift force is defined. The drag and pitching moment are derived using...... a conditional simulation technique for stochastic processes. The input data for the model can be collected either from measurements or from numerical results from a Computational Fluid Dynamics code for airfoil sections at constant angles of attack. An analysis of such data is provided, which helps to determine...

Suppression of dynamic stall with a leading-edge slat on a VR-7 airfoil

Science.gov (United States)

Mcalister, K. W.; Tung, C.

1993-01-01

The VR-7 airfoil was experimentally studied with and without a leading-edge slat at fixed angles of attack from 0 deg to 30 deg at Re = 200,000 and for unsteady pitching motions described by alpha equals alpha(sub m) + 10 deg(sin(wt)). The models were two dimensional, and the test was performed in a water tunnel at Ames Research Center. The unsteady conditions ranged over Re equals 100,000 to 250,000, k equals 0.001 to 0.2, and alpha(sub m) = 10 deg to 20 deg. Unsteady lift, drag, and pitching-moment measurements were obtained along with fluorescent-dye flow visualizations. The addition of the slat was found to delay the static-drag and static-moment stall by about 5 degrees and to eliminate completely the development of a dynamic-stall vortex during unsteady motions that reached angles as high as 25 degrees. In all of the unsteady cases studied, the slat caused a significant reduction in the force and moment hysteresis amplitudes. The reduced frequency was found to have the greatest effect on the results, whereas the Reynolds number had little effect on the behavior of either the basic or the slatted airfoil. The slat caused a slight drag penalty at low angles of attack, but generally increased the lift/drag ratio when averaged over the full cycle of oscillation.

Experimental Methods Applied in a Study of Stall Flutter in an Axial Flow Fan

Directory of Open Access Journals (Sweden)

John D. Gill

2004-01-01

Full Text Available Flutter testing is an integral part of aircraft gas turbine engine development. In typical flutter testing blade mounted sensors in the form of strain gages and casing mounted sensors in the form of light probes (NSMS are used. Casing mounted sensors have the advantage of being non-intrusive and can detect the vibratory response of each rotating blade. Other types of casing mounted sensors can also be used to detect flutter of rotating blades. In this investigation casing mounted high frequency response pressure transducers are used to characterize the part-speed stall flutter response of a single stage unshrouded axial-flow fan. These dynamic pressure transducers are evenly spaced around the circumference at a constant axial location upstream of the fan blade leading edge plane. The pre-recorded experimental data at 70% corrected speed is analyzed for the case where the fan is back-pressured into the stall flutter zone. The experimental data is analyzed using two probe and multi-probe techniques. The analysis techniques for each method are presented. Results from these two analysis methods indicate that flutter occurred at a frequency of 411 Hz with a dominant nodal diameter of 2. The multi-probe analysis technique is a valuable method that can be used to investigate the initiation of flutter in turbomachines.

Compliance with NAGCAT work practices recommendations for youth cleaning service alleys in stall barns.

Science.gov (United States)

Canan, B D; Asti, L; Heaney, C; Ashida, S; Renick, K; Xiang, H; Stallones, L; Jepsen, S D; Crawford, J M; Wilkins, J R

2011-04-01

Unintentional injury is the leading cause of death in the U.S. among persons 1 to 44 years of age. Over one million children and adolescents in the U.S. live, work, and/or play on farms, where injury risk is relatively high compared to other settings. In an attempt to reduce the number of childhood agricultural injuries occurring on farms, the North American Guidelines for Children's Agricultural Tasks (NAGCAT) was developed to assist parents or other caregivers in assigning developmentally appropriate chores to youth exposed to agricultural hazards. The results presented here are from a longitudinal study in which we obtained (self-reported) daily chore, injury, and safety behavior data from children and adolescents. We focused on one NAGCAT chore, cleaning a service alley in a stall barn, in order to estimate the extent of compliance with specific work practice recommendations contained in the NAGCAT. Our results indicated that among the four NAGCAT-recommended safety practices for cleaning service alleys in stall barns (wearing nonskid shoes, leather gloves, a respirator, and eye protection), wearing non-skid shoes was the only safety practice reported with any degree of regularity. Overall, boys were more likely to wear non-skid shoes compared to girls. In addition, older youth were generally more likely to report higher work practice compliance compared to younger youth.

Formation and repair of DNA-protein cross-links (DPCs) in newly replicated DNA

International Nuclear Information System (INIS)

Chiu, S.; Friedman, L.R.; Oleinick, N.L.

1987-01-01

DPCs preferentially involve proteins of the nuclear matrix, the site of replication and transcription. To elucidate the relationship with replication, the formation and repair of DPCs has been studied in newly replicated DNA. Log-phase V79 cells were pulsed with /sup 3/H-TdR (10-20 μCi/ml) for 30-90 sec at 22 0 followed by up to a 60 min chase at 37 0 . Irradiation (0-100 Gy) immediately after the pulse increases the labeled DNA in DPCs with a dose-dependence that is unaffected by the initial level of labeled DPC or by chase time. When cells are irradiated before the pulse, DNA synthesis is inhibited; however, release of pulse-labeled DPCs appears normal. The data suggest that during replication, DNA is cross-linked to (matrix) protein, contributing to background DPCs

DNA Replication in Engineered Escherichia coli Genomes with Extra Replication Origins.

Science.gov (United States)

Milbredt, Sarah; Farmani, Neda; Sobetzko, Patrick; Waldminghaus, Torsten

2016-10-21

The standard outline of bacterial genomes is a single circular chromosome with a single replication origin. From the bioengineering perspective, it appears attractive to extend this basic setup. Bacteria with split chromosomes or multiple replication origins have been successfully constructed in the last few years. The characteristics of these engineered strains will largely depend on the respective DNA replication patterns. However, the DNA replication has not been investigated systematically in engineered bacteria with multiple origins or split replicons. Here we fill this gap by studying a set of strains consisting of (i) E. coli strains with an extra copy of the native replication origin (oriC), (ii) E. coli strains with an extra copy of the replication origin from the secondary chromosome of Vibrio cholerae (oriII), and (iii) a strain in which the E. coli chromosome is split into two linear replicons. A combination of flow cytometry, microarray-based comparative genomic hybridization (CGH), and modeling revealed silencing of extra oriC copies and differential timing of ectopic oriII copies compared to the native oriC. The results were used to derive construction rules for future multiorigin and multireplicon projects.

Role for a region of helically unstable DNA within the Epstein-Barr virus latent cycle origin of DNA replication oriP in origin function

International Nuclear Information System (INIS)

Polonskaya, Zhanna; Benham, Craig J.; Hearing, Janet

2004-01-01

The minimal replicator of the Epstein-Barr virus (EBV) latent cycle origin of DNA replication oriP is composed of two binding sites for the Epstein-Barr virus nuclear antigen-1 (EBNA-1) and flanking inverted repeats that bind the telomere repeat binding factor TRF2. Although not required for minimal replicator activity, additional binding sites for EBNA-1 and TRF2 and one or more auxiliary elements located to the right of the EBNA-1/TRF2 sites are required for the efficient replication of oriP plasmids. Another region of oriP that is predicted to be destabilized by DNA supercoiling is shown here to be an important functional component of oriP. The ability of DNA fragments of unrelated sequence and possessing supercoiled-induced DNA duplex destabilized (SIDD) structures, but not fragments characterized by helically stable DNA, to substitute for this component of oriP demonstrates a role for the SIDD region in the initiation of oriP-plasmid DNA replication

Hydroxyurea-Induced Replication Stress

Directory of Open Access Journals (Sweden)

Kenza Lahkim Bennani-Belhaj

2010-01-01

Full Text Available Bloom's syndrome (BS displays one of the strongest known correlations between chromosomal instability and a high risk of cancer at an early age. BS cells combine a reduced average fork velocity with constitutive endogenous replication stress. However, the response of BS cells to replication stress induced by hydroxyurea (HU, which strongly slows the progression of replication forks, remains unclear due to publication of conflicting results. Using two different cellular models of BS, we showed that BLM deficiency is not associated with sensitivity to HU, in terms of clonogenic survival, DSB generation, and SCE induction. We suggest that surviving BLM-deficient cells are selected on the basis of their ability to deal with an endogenous replication stress induced by replication fork slowing, resulting in insensitivity to HU-induced replication stress.

MMS exposure promotes increased MtDNA mutagenesis in the presence of replication-defective disease-associated DNA polymerase γ variants.

Science.gov (United States)

Stumpf, Jeffrey D; Copeland, William C

2014-10-01

Mitochondrial DNA (mtDNA) encodes proteins essential for ATP production. Mutant variants of the mtDNA polymerase cause mutagenesis that contributes to aging, genetic diseases, and sensitivity to environmental agents. We interrogated mtDNA replication in Saccharomyces cerevisiae strains with disease-associated mutations affecting conserved regions of the mtDNA polymerase, Mip1, in the presence of the wild type Mip1. Mutant frequency arising from mtDNA base substitutions that confer erythromycin resistance and deletions between 21-nucleotide direct repeats was determined. Previously, increased mutagenesis was observed in strains encoding mutant variants that were insufficient to maintain mtDNA and that were not expected to reduce polymerase fidelity or exonuclease proofreading. Increased mutagenesis could be explained by mutant variants stalling the replication fork, thereby predisposing the template DNA to irreparable damage that is bypassed with poor fidelity. This hypothesis suggests that the exogenous base-alkylating agent, methyl methanesulfonate (MMS), would further increase mtDNA mutagenesis. Mitochondrial mutagenesis associated with MMS exposure was increased up to 30-fold in mip1 mutants containing disease-associated alterations that affect polymerase activity. Disrupting exonuclease activity of mutant variants was not associated with increased spontaneous mutagenesis compared with exonuclease-proficient alleles, suggesting that most or all of the mtDNA was replicated by wild type Mip1. A novel subset of C to G transversions was responsible for about half of the mutants arising after MMS exposure implicating error-prone bypass of methylated cytosines as the predominant mutational mechanism. Exposure to MMS does not disrupt exonuclease activity that suppresses deletions between 21-nucleotide direct repeats, suggesting the MMS-induce mutagenesis is not explained by inactivated exonuclease activity. Further, trace amounts of CdCl2 inhibit mtDNA replication but

Mutational analysis of varicella-zoster virus (VZV) immediate early protein (IE62) subdomains and their importance in viral replication

Energy Technology Data Exchange (ETDEWEB)

Khalil, Mohamed I., E-mail: mkhalil2@stanford.edu [Departments of Pediatrics and Microbiology & Immunology, Stan ford University School of Medicine, Stanford, CA (United States); Department of Molecular Biology, National Research Centre, El-Buhouth St., Cairo (Egypt); Che, Xibing; Sung, Phillip; Sommer, Marvin H. [Departments of Pediatrics and Microbiology & Immunology, Stan ford University School of Medicine, Stanford, CA (United States); Hay, John [Department of Microbiology and Immunology, School of Medicine and Biomedical Science, University at Buffalo, Buffalo, NY (United States); Arvin, Ann M. [Departments of Pediatrics and Microbiology & Immunology, Stan ford University School of Medicine, Stanford, CA (United States)

2016-05-15

VZV IE62 is an essential, immediate-early, tegument protein and consists of five domains. We generated recombinant viruses carrying mutations in the first three IE62 domains and tested their influence on VZV replication kinetics. The mutations in domain I did not affect replication kinetics while domain II mutations, disrupting the DNA binding and dimerization domain (DBD), were lethal for VZV replication. Mutations in domain III of the nuclear localization signal (NLS) and the two phosphorylation sites S686A/S722A resulted in slower growth in early and late infection respectively and were associated with IE62 accumulation in the cytoplasm and nucleus respectively. This study mapped the functional domains of IE62 in context of viral infection, indicating that DNA binding and dimerization domain is essential for VZV replication. In addition, the correct localization of IE62, whether nuclear or cytoplasmic, at different points in the viral life cycle, is important for normal progression of VZV replication. - Highlights: • Mutation of IE62 domain I did not affect VZV replication in melanoma cells. • IE62 domain II and III are important for VZV replication in melanoma cells. • Mutations of IE62 domain II (DBD) were lethal for virus replication. • Mutations of IE62 NLS and phosphorylation sites inhibited VZV replication. • NLS and S686A/S722A mutations altered localization of IE62 during early and late infection.

Mutational analysis of varicella-zoster virus (VZV) immediate early protein (IE62) subdomains and their importance in viral replication

International Nuclear Information System (INIS)

Khalil, Mohamed I.; Che, Xibing; Sung, Phillip; Sommer, Marvin H.; Hay, John; Arvin, Ann M.

2016-01-01

VZV IE62 is an essential, immediate-early, tegument protein and consists of five domains. We generated recombinant viruses carrying mutations in the first three IE62 domains and tested their influence on VZV replication kinetics. The mutations in domain I did not affect replication kinetics while domain II mutations, disrupting the DNA binding and dimerization domain (DBD), were lethal for VZV replication. Mutations in domain III of the nuclear localization signal (NLS) and the two phosphorylation sites S686A/S722A resulted in slower growth in early and late infection respectively and were associated with IE62 accumulation in the cytoplasm and nucleus respectively. This study mapped the functional domains of IE62 in context of viral infection, indicating that DNA binding and dimerization domain is essential for VZV replication. In addition, the correct localization of IE62, whether nuclear or cytoplasmic, at different points in the viral life cycle, is important for normal progression of VZV replication. - Highlights: • Mutation of IE62 domain I did not affect VZV replication in melanoma cells. • IE62 domain II and III are important for VZV replication in melanoma cells. • Mutations of IE62 domain II (DBD) were lethal for virus replication. • Mutations of IE62 NLS and phosphorylation sites inhibited VZV replication. • NLS and S686A/S722A mutations altered localization of IE62 during early and late infection.

ATEFlap aerodynamic model, a dynamic stall model including the effects of trailing edge flap deflection

Energy Technology Data Exchange (ETDEWEB)

Bergami, L.; Gaunaa, M.

2012-02-15

The report presents the ATEFlap aerodynamic model, which computes the unsteady lift, drag and moment on a 2D airfoil section equipped with Adaptive Trailing Edge Flap. The model captures the unsteady response related to the effects of the vorticity shed into the wake, and the dynamics of flow separation a thin-airfoil potential flow model is merged with a dynamic stall model of the Beddoes-Leishmann type. The inputs required by the model are steady data for lift, drag, and moment coefficients as function of angle of attack and flap deflection. Further steady data used by the Beddoes- Leishmann dynamic stall model are computed in an external preprocessor application, which gives the user the possibility to verify, and eventually correct, the steady data passed to the aerodynamic model. The ATEFlap aerodynamic model is integrated in the aeroelastic simulation tool HAWC2, thus al- lowing to simulate the response of a wind turbine with trailing edge flaps on the rotor. The algorithms used by the preprocessor, and by aerodynamic model are presented, and modifications to previous implementations of the aerodynamic model are briefly discussed. The performance and the validity of the model are verified by comparing the dynamic response computed by the ATEFlap with solutions from CFD simulations. (Author)

Analysis of the temporal program of replication initiation in yeast chromosomes.

Science.gov (United States)

Friedman, K L; Raghuraman, M K; Fangman, W L; Brewer, B J

1995-01-01

The multiple origins of eukaryotic chromosomes vary in the time of their initiation during S phase. In the chromosomes of Saccharomyces cerevisiae the presence of a functional telomere causes nearby origins to delay initiation until the second half of S phase. The key feature of telomeres that causes the replication delay is the telomeric sequence (C(1-3)A/G(1-3)T) itself and not the proximity of the origin to a DNA end. A second group of late replicating origins has been found at an internal position on chromosome XIV. Four origins, spanning approximately 140 kb, initiate replication in the second half of S phase. At least two of these internal origins maintain their late replication time on circular plasmids. Each of these origins can be separated into two functional elements: those sequences that provide origin function and those that impose late activation. Because the assay for determining replication time is costly and laborious, it has not been possible to analyze in detail these 'late' elements. We report here the development of two new assays for determining replication time. The first exploits the expression of the Escherichia coli dam methylase in yeast and the characteristic period of hemimethylation that transiently follows the passage of a replication fork. The second uses quantitative hybridization to detect two-fold differences in the amount of specific restriction fragments as a function of progress through S phase. The novel aspect of this assay is the creation in vivo of a non-replicating DNA sequence by site-specific pop-out recombination. This non-replicating fragment acts as an internal control for copy number within and between samples. Both of these techniques are rapid and much less costly than the more conventional density transfer experiments that require CsCl gradients to detect replicated DNA. With these techniques it should be possible to identify the sequences responsible for late initiation, to search for other late replicating

Effect of paddock vs. stall housing on 24 hour gastric pH within the proximal and ventral equine stomach.

Science.gov (United States)

Husted, L; Sanchez, L C; Olsen, S N; Baptiste, K E; Merritt, A M

2008-06-01

Stall housing has been suggested as a risk factor for ulcer development in the equine stomach; however, the exact pathogenesis for this has not been established. To investigate the effect of 3 environmental situations (grass paddock, stall alone or stall with adjacent companion) on pH in the proximal and the ventral stomach. Six horses with permanently implanted gastric cannulae were used in a randomised, cross-over, block design. Each horse rotated through each of three 24 h environmental situations. Horses remained on their normal diet (grass hay ad libitum and grain b.i.d.) throughout the study. Intragastric pH was measured continuously for 72 h just inside the lower oesophageal sphincter (proximal stomach) and via a pH probe in the gastric cannula (ventral stomach). Neither proximal nor ventral 24 h gastric pH changed significantly between the 3 environmental situations. Mean hourly proximal gastric pH decreased significantly in the interval from 01.00-09.00 h compared to the interval from 13.00-20.00 h, regardless of environmental situation. Median hourly proximal pH only differed in the interval from 06.00-07.00 h compared to the interval 14.00-19.00 h. Neither mean nor median hourly ventral gastric pH varied significantly with the time of day. The change in housing status used in the current study did not affect acid exposure within either region of the equine stomach. The pH in the ventral stomach was uniformly stable throughout the study, while the proximal pH demonstrated a 24 h circadian pattern.

Identification of Thrust, Lift, and Drag for Deep-stall Flight Data of a Fixed-wing Unmanned Aircraft

DEFF Research Database (Denmark)

Cunis, Torbjørn; Leth, Tobias; Totu, Luminita Cristiana

2018-01-01

In this paper, we consider a small unmanned aircraft and data collected during regular and deep-stall flight. We present an identification method for the thrust force generated by the propulsion system based on the in-flight measurements where we make use of the well-known linear and quadratic...... force estimation in the full flight envelope....

Regulatory cross-talk links Vibrio cholerae chromosome II replication and segregation.

Directory of Open Access Journals (Sweden)

Yoshiharu Yamaichi

2011-07-01

Full Text Available There is little knowledge of factors and mechanisms for coordinating bacterial chromosome replication and segregation. Previous studies have revealed that genes (and their products that surround the origin of replication (oriCII of Vibrio cholerae chromosome II (chrII are critical for controlling the replication and segregation of this chromosome. rctB, which flanks one side of oriCII, encodes a protein that initiates chrII replication; rctA, which flanks the other side of oriCII, inhibits rctB activity. The chrII parAB2 operon, which is essential for chrII partitioning, is located immediately downstream of rctA. Here, we explored how rctA exerts negative control over chrII replication. Our observations suggest that RctB has at least two DNA binding domains--one for binding to oriCII and initiating replication and the other for binding to rctA and thereby inhibiting RctB's ability to initiate replication. Notably, the inhibitory effect of rctA could be alleviated by binding of ParB2 to a centromere-like parS site within rctA. Furthermore, by binding to rctA, ParB2 and RctB inversely regulate expression of the parAB2 genes. Together, our findings suggest that fluctuations in binding of the partitioning protein ParB2 and the chrII initiator RctB to rctA underlie a regulatory network controlling both oriCII firing and the production of the essential chrII partitioning proteins. Thus, by binding both RctB and ParB2, rctA serves as a nexus for regulatory cross-talk coordinating chrII replication and segregation.

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.

Caffeine Abolishes the Ultraviolet-Induced REV3 Translesion Replication Pathway in Mouse Cells

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