Sample records for lethality screen reveals

  1. Synthetic lethal screening reveals FGFR as one of the combinatorial targets to overcome resistance to Met-targeted therapy.

    PubMed

    Kim, B; Wang, S; Lee, J M; Jeong, Y; Ahn, T; Son, D-S; Park, H W; Yoo, H-s; Song, Y-J; Lee, E; Oh, Y M; Lee, S B; Choi, J; Murray, J C; Zhou, Y; Song, P H; Kim, K-A; Weiner, L M

    2015-02-26

    Met is a receptor tyrosine kinase that promotes cancer progression. In addition, Met has been implicated in resistance of tumors to various targeted therapies such as epidermal growth factor receptor inhibitors in lung cancers, and has been prioritized as a key molecular target for cancer therapy. However, the underlying mechanism of resistance to Met-targeting drugs is poorly understood. Here, we describe screening of 1310 genes to search for key regulators related to drug resistance to an anti-Met therapeutic antibody (SAIT301) by using a small interfering RNA-based synthetic lethal screening method. We found that knockdown of 69 genes in Met-amplified MKN45 cells sensitized the antitumor activity of SAIT301. Pathway analysis of these 69 genes implicated fibroblast growth factor receptor (FGFR) as a key regulator for antiproliferative effects of Met-targeting drugs. Inhibition of FGFR3 increased target cell apoptosis through the suppression of Bcl-xL expression, followed by reduced cancer cell growth in the presence of Met-targeting drugs. Treatment of cells with the FGFR inhibitors substantially restored the efficacy of SAIT301 in SAIT301-resistant cells and enhanced the efficacy in SAIT301-sensitive cells. In addition to FGFR3, integrin ?3 is another potential target for combination treatment with SAIT301. Suppression of integrin ?3 decreased AKT phosphorylation in SAIT301-resistant cells and restored SAIT301 responsiveness in HCC1954 cells, which are resistant to SAIT301. Gene expression analysis using CCLE database shows that cancer cells with high levels of FGFR and integrin ?3 are resistant to crizotinib treatment, suggesting that FGFR and integrin ?3 could be used as predictive markers for Met-targeted therapy and provide a potential therapeutic option to overcome acquired and innate resistance for the Met-targeting drugs. PMID:24662823

  2. Parallel in vivo and in vitro melanoma RNAi dropout screens reveal synthetic lethality between hypoxia and DNA damage response inhibition.

    PubMed

    Possik, Patricia A; Müller, Judith; Gerlach, Carmen; Kenski, Juliana C N; Huang, Xinyao; Shahrabi, Aida; Krijgsman, Oscar; Song, Ji-Ying; Smit, Marjon A; Gerritsen, Bram; Lieftink, Cor; Kemper, Kristel; Michaut, Magali; Beijersbergen, Roderick L; Wessels, Lodewyk; Schumacher, Ton N; Peeper, Daniel S

    2014-11-20

    To identify factors preferentially necessary for driving tumor expansion, we performed parallel in vitro and in vivo negative-selection short hairpin RNA (shRNA) screens. Melanoma cells harboring shRNAs targeting several DNA damage response (DDR) kinases had a greater selective disadvantage in vivo than in vitro, indicating an essential contribution of these factors during tumor expansion. In growing tumors, DDR kinases were activated following hypoxia. Correspondingly, depletion or pharmacologic inhibition of DDR kinases was toxic to melanoma cells, including those that were resistant to BRAF inhibitor, and this could be enhanced by angiogenesis blockade. These results reveal that hypoxia sensitizes melanomas to targeted inhibition of the DDR and illustrate the utility of in vivo shRNA dropout screens for the identification of pharmacologically tractable targets. PMID:25456132

  3. Actin Dosage Lethality Screening in Yeast Mediated by Selective Ploidy Ablation Reveals Links to Urmylation/Wobble Codon Recognition and Chromosome Stability

    PubMed Central

    Haarer, Brian; Mi-Mi, Lei; Cho, Jessica; Cortese, Matthew; Viggiano, Susan; Burke, Daniel; Amberg, David

    2013-01-01

    The actin cytoskeleton exists in a dynamic equilibrium with monomeric and filamentous states of its subunit protein actin. The spatial and temporal regulation of actin dynamics is critical to the many functions of actin. Actin levels are remarkably constant, suggesting that cells have evolved to function within a narrow range of actin concentrations. Here we report the results of screens in which we have increased actin levels in strains deleted for the ~4800 nonessential yeast genes using a technical advance called selective ploidy ablation. We detected 83 synthetic dosage interactions with actin, 78 resulted in reduced growth, whereas in 5 cases overexpression of actin suppressed the growth defects caused by the deleted genes. The genes were highly enriched in several classes, including transfer RNA wobble uridine modification, chromosome stability and segregation, cell growth, and cell division. We show that actin overexpression sequesters a limited pool of eEF1A, a bifunctional protein involved in aminoacyl-transfer RNA recruitment to the ribosome and actin filament cross-linking. Surprisingly, the largest class of genes is involved in chromosome stability and segregation. We show that actin mutants have chromosome segregation defects, suggesting a possible role in chromosome structure and function. Monomeric actin is a core component of the INO80 and SWR chromatin remodeling complexes and the NuA4 histone modification complex, and our results suggest these complexes may be sensitive to actin stoichiometry. We propose that the resulting effects on chromatin structure can lead to synergistic effects on chromosome stability in strains lacking genes important for chromosome maintenance. PMID:23450344

  4. Synthetic lethal screening in the mammalian central nervous system identifies Gpx6 as a modulator of Huntington's disease.

    PubMed

    Shema, Reut; Kulicke, Ruth; Cowley, Glenn S; Stein, Rachael; Root, David E; Heiman, Myriam

    2015-01-01

    Huntington's disease, the most common inherited neurodegenerative disease, is characterized by a dramatic loss of deep-layer cortical and striatal neurons, as well as morbidity in midlife. Human genetic studies led to the identification of the causative gene, huntingtin. Recent genomic advances have also led to the identification of hundreds of potential interacting partners for huntingtin protein and many hypotheses as to the molecular mechanisms whereby mutant huntingtin leads to cellular dysfunction and death. However, the multitude of possible interacting partners and cellular pathways affected by mutant huntingtin has complicated efforts to understand the etiology of this disease, and to date no curative therapeutic exists. To address the general problem of identifying the disease-phenotype contributing genes from a large number of correlative studies, here we develop a synthetic lethal screening methodology for the mammalian central nervous system, called SLIC, for synthetic lethal in the central nervous system. Applying SLIC to the study of Huntington's disease, we identify the age-regulated glutathione peroxidase 6 (Gpx6) gene as a modulator of mutant huntingtin toxicity and show that overexpression of Gpx6 can dramatically alleviate both behavioral and molecular phenotypes associated with a mouse model of Huntington's disease. SLIC can, in principle, be used in the study of any neurodegenerative disease for which a mouse model exists, promising to reveal modulators of neurodegenerative disease in an unbiased fashion, akin to screens in simpler model organisms. PMID:25535386

  5. Synthetic lethal screens as a means to understand and treat MYC-driven cancers

    PubMed Central

    Cermelli, Silvia; Jang, In Sock; Bernard, Brady; Grandori, Carla

    2014-01-01

    While therapeutics against MYC could potentially be employed against a wide range of human cancers, MYC targeted therapies have proven difficult to develop. The convergence of breakthroughs in human genomics and in gene silencing using RNA interference (RNAi), have recently allowed functional interrogation of the genome and systematic identification of synthetic lethal interactions with hyper-active MYC. Here, we focus on the pathways that have emerged through RNAi screens and present evidence that a subset of genes exhibiting synthetic lethality with MYC are significantly interconnected and linked to chromatin, and transcriptional processes, as well as to DNA repair and cell-cycle checkpoints. Other synthetic lethal interactions with MYC point to novel pathways and potentially broaden the repertoire of targeted therapies. The elucidation of MYC synthetic lethal interactions is still in its infancy and how these interactions may be influenced by tissue specific programs and by concurrent genetic change will require further investigation. Nevertheless, we predict that these studies may lead the way to novel therapeutic approaches and new insights into the role of MYC in cancer. PMID:24591535

  6. Evaluation of Caenorhabditis elegans as an acute lethality and a neurotoxicity screening model

    SciTech Connect

    Williams, P.L.

    1988-01-01

    This investigation evaluated C. elegans as a lethality and neurotoxicity screening model. The lethality experiments were performed in both agar and an aquatic medium. The salts of 8 metals (Hg, Be, Al, Cu, Zn, Pb, Cd, and Sr) were used in the agar studies and the salts of 14 metals (Ag, Hg, Cu, Be, Al, Pb, Cr, As, Tl, Zn, Cd, Ni, Sr, and Sb) were used in the aquatic tests. In each of these tests an LC50 value was determined. The data from the agar plates were compared to the published mammalian oral LD50 values for salts of the same metals. Within this set of chemicals C. elegans was found to be a predictor of mammalian acute lethality, generating LC50 values parallel to the rat and mouse LD50 values. The aquatic data were compared to data from EPA Ambient Water Quality Criteria documents. C. elegans was found to be less sensitive than Daphnia but generally more sensitive than the other invertebrate organisms that are presently used. The neurotoxicity testing also was performed in both agar and an aquatic media. The testing in agar was conducted with the salts of 4 metals (Cu, Be, Pb, and Hg) and 2 organophosphate pesticides (malathion and vapona). The studies in an aquatic medium tested the salts of 4 metals (Cu, Be, Pb, and Hg).

  7. Identification of exosite-targeting inhibitors of anthrax lethal factor by high throughput screening

    PubMed Central

    Bannwarth, Ludovic; Goldberg, Allison B.; Chen, Catherine; Turk, Benjamin E.

    2012-01-01

    SUMMARY Protease inhibitor discovery has focused almost exclusively on compounds that bind to the active site. Inhibitors targeting protease exosites, regions outside of the active site that influence catalysis, offer potential advantages of increased specificity but are difficult to systematically discover. Here we describe an assay suitable for detecting exosite-targeting inhibitors of the metalloproteinase anthrax lethal factor (LF) based on cleavage of a full length mitogen-activated protein kinase kinase (MKK) substrate. We used this assay to screen a small molecule library, and then subjected hits to a secondary screen to exclude compounds that efficiently blocked cleavage of a peptide substrate. We identified a compound that preferentially inhibited cleavage of MKKs compared with peptide substrates and could suppress LF-induced macrophage cytolysis. This approach should be generally applicable to the discovery of exosite-targeting inhibitors of many additional proteases. PMID:22840775

  8. RNA Interference Screen Identifies VEGFR1 as Potentially Synthetic Lethal to Aberrant Wnt/?-catenin Activation in Colon Cancer

    PubMed Central

    Naik, Snehal; Dothager, Robin S.; Marasa, Jayne; Lewis, Cory L.; Piwnica-Worms, David

    2009-01-01

    Purpose: The Wnt/?-catenin (?-cat) signaling cascade is a key regulator of development, and dysregulation of Wnt/?-cat contributes to selected cancers, such as colorectal, breast and hepatocellular carcinoma, through abnormal activation of Wnt target genes. To identify novel modulators of the Wnt/?-cat pathway which may emerge as therapeutic targets, we performed an unbiased high-throughput RNA interference screen. Experimental Design: A synthetic oligonucleotide siRNA library targeting 691 known and predicted human kinases was screened in Wnt3a-stimulated human cells in a live cell luciferase assay for modulation of Wnt/?-cat-dependent transcription. Follow-up studies of a selected high-confidence “hit” were conducted. Results: A robust quartile-based statistical analysis and secondary screen yielded several kinases worthy of further investigation, including Cdc2L1, Lmtk3, Pank2, ErbB3, and of note, vascular endothelial growth factor receptor-1 (VEGFR1/Flt1), a receptor tyrosine kinase with putative weak kinase activity conventionally believed to be a negative regulator of angiogenesis. A series of loss-of-function, genetic null, and VEGFR tyrosine kinase inhibitor assays further revealed that VEGFR1 is a positive regulator of Wnt signaling that functions in a glycogen synthase kinase-3? (GSK3?)-independent manner as a potential synthetic lethal target in Wnt/?-cat-addicted colon carcinoma cells. Conclusion: This unanticipated non-endothelial link between VEGFR1 tyrosine kinase activity and Wnt/?-cat signaling may refine our understanding of aberrant Wnt signaling in colon carcinoma and points to new combinatorial therapeutics targeted to the tumor cell compartment, rather than angiogenesis, in the context of colon cancer. PMID:20008853

  9. Lethality in PARP-1/Ku80 double mutant mice reveals physiologicalsynergy during early embryogenesis

    SciTech Connect

    Henrie, Melinda S.; Kurimasa, Akihiro; Burma, Sandeep; Menissier-de Murcia, Josiane; de Murcia, Gilbert; Li, Gloria C.; Chen,David J.

    2002-09-24

    Ku is an abundant heterodimeric nuclear protein, consisting of 70-kDa and 86-kDa tightly associated subunits that comprise the DNA binding component of DNA-dependent protein kinase. Poly(ADP)ribose polymerase-1 (PARP-1) is a 113-kDa protein that catalyzes the synthesis of poly(ADP-ribose) on target proteins. Both Ku and PARP-1 recognize and bind to DNA ends. Ku functions in the non-homologous end joining (NHEJ) repair pathway whereas PARP-1 functions in the single strand break repair and base excision repair (BER) pathways. Recent studies have revealed that PARP-1 and Ku80 interact in vitro. To determine whether the association of PARP-1 and Ku80 has any physiological significance or synergistic function in vivo, mice lacking both PARP-1 and Ku80 were generated. The resulting offspring died during embryonic development displaying abnormalities around the gastrulation stage. In addition, PARP-1-/-Ku80-/- cultured blastocysts had an increased level of apoptosis. These data suggest that the functions of both Ku80 and PARP-1 are essential for normal embryogenesis and that a loss of genomic integrity leading to cell death through apoptosis is likely the cause of the embryonic lethality observed in these mice.

  10. A Synthetic Lethal Screen Identifies DNA Repair Pathways that Sensitize Cancer Cells to Combined ATR Inhibition and Cisplatin Treatments

    PubMed Central

    Mohni, Kareem N.; Thompson, Petria S.; Luzwick, Jessica W.; Glick, Gloria G.; Pendleton, Christopher S.; Lehmann, Brian D.; Pietenpol, Jennifer A.; Cortez, David

    2015-01-01

    The DNA damage response kinase ATR may be a useful cancer therapeutic target. ATR inhibition synergizes with loss of ERCC1, ATM, XRCC1 and DNA damaging chemotherapy agents. Clinical trials have begun using ATR inhibitors in combination with cisplatin. Here we report the first synthetic lethality screen with a combination treatment of an ATR inhibitor (ATRi) and cisplatin. Combination treatment with ATRi/cisplatin is synthetically lethal with loss of the TLS polymerase ? and 53BP1. Other DNA repair pathways including homologous recombination and mismatch repair do not exhibit synthetic lethal interactions with ATRi/cisplatin, even though loss of some of these repair pathways sensitizes cells to cisplatin as a single-agent. We also report that ATRi strongly synergizes with PARP inhibition, even in homologous recombination-proficient backgrounds. Lastly, ATR inhibitors were able to resensitize cisplatin-resistant cell lines to cisplatin. These data provide a comprehensive analysis of DNA repair pathways that exhibit synthetic lethality with ATR inhibitors when combined with cisplatin chemotherapy, and will help guide patient selection strategies as ATR inhibitors progress into the cancer clinic. PMID:25965342

  11. A Synthetic Lethal Screen Identifies DNA Repair Pathways that Sensitize Cancer Cells to Combined ATR Inhibition and Cisplatin Treatments.

    PubMed

    Mohni, Kareem N; Thompson, Petria S; Luzwick, Jessica W; Glick, Gloria G; Pendleton, Christopher S; Lehmann, Brian D; Pietenpol, Jennifer A; Cortez, David

    2015-01-01

    The DNA damage response kinase ATR may be a useful cancer therapeutic target. ATR inhibition synergizes with loss of ERCC1, ATM, XRCC1 and DNA damaging chemotherapy agents. Clinical trials have begun using ATR inhibitors in combination with cisplatin. Here we report the first synthetic lethality screen with a combination treatment of an ATR inhibitor (ATRi) and cisplatin. Combination treatment with ATRi/cisplatin is synthetically lethal with loss of the TLS polymerase ? and 53BP1. Other DNA repair pathways including homologous recombination and mismatch repair do not exhibit synthetic lethal interactions with ATRi/cisplatin, even though loss of some of these repair pathways sensitizes cells to cisplatin as a single-agent. We also report that ATRi strongly synergizes with PARP inhibition, even in homologous recombination-proficient backgrounds. Lastly, ATR inhibitors were able to resensitize cisplatin-resistant cell lines to cisplatin. These data provide a comprehensive analysis of DNA repair pathways that exhibit synthetic lethality with ATR inhibitors when combined with cisplatin chemotherapy, and will help guide patient selection strategies as ATR inhibitors progress into the cancer clinic. PMID:25965342

  12. Statistical analysis, optimization, and prioritization of virtual screening parameters for zinc enzymes including the anthrax toxin lethal factor.

    PubMed

    Maize, Kimberly M; Zhang, Xia; Amin, Elizabeth Ambrose

    2014-01-01

    The anthrax toxin lethal factor (LF) and matrix metalloproteinase-3 (MMP-3, stromelysin-1) are popular zinc metalloenzyme drug targets, with LF primarily responsible for anthrax-related toxicity and host death, while MMP-3 is involved in cancer- and rheumatic disease-related tissue remodeling. A number of in silico screening techniques, most notably docking and scoring, have proven useful for identifying new potential drug scaffolds targeting LF and MMP-3, as well as for optimizing lead compounds and investigating mechanisms of action. However, virtual screening outcomes can vary significantly depending on the specific docking parameters chosen, and systematic statistical significance analyses are needed to prioritize key parameters for screening small molecules against these zinc systems. In the current work, we present a series of chi-square statistical analyses of virtual screening outcomes for cocrystallized LF and MMP-3 inhibitors docked into their respective targets, evaluated by predicted enzyme-inhibitor dissociation constant and root-mean-square deviation (RMSD) between predicted and experimental bound configurations, and we present a series of preferred parameters for use with these systems in the industry-standard Surflex-Dock screening program, for use by researchers utilizing in silico techniques to discover and optimize new scaffolds. PMID:25373478

  13. Rescue of the embryonic lethal hematopoietic defect reveals a critical role for GATA-2 in urogenital development.

    PubMed Central

    Zhou, Y; Lim, K C; Onodera, K; Takahashi, S; Ohta, J; Minegishi, N; Tsai, F Y; Orkin, S H; Yamamoto, M; Engel, J D

    1998-01-01

    Mutations resulting in embryonic or early postnatal lethality could mask the activities of any gene in unrelated and temporally distinct developmental pathways. Targeted inactivation of the transcription factor GATA-2 gene leads to mid-gestational death as a consequence of hematopoietic failure. We show here that a 250 kbp GATA-2 yeast artificial chromosome (YAC) is expressed strongly in both the primitive and definitive hematopoietic compartments, while two smaller YACs are not. This largest YAC also rescues hematopoiesis in vitro and in vivo, thereby localizing the hematopoietic regulatory cis element(s) to between 100 and 150 kbp 5' to the GATA-2 structural gene. Introducing the YAC transgene into the GATA-2(-/-) genetic background allows the embryos to complete gestation; however, newborn rescued pups quickly succumb to lethal hydroureternephrosis, and display a complex array of genitourinary abnormalities. These findings reveal that GATA-2 plays equally vital roles in urogenital and hematopoietic development. PMID:9822612

  14. A Synthetic Lethal Screen Identifies a Role for Lin-44/Wnt in C. elegans Embryogenesis

    E-print Network

    Hartin, Samantha N.; Hudson, Martin L.; Yingling, Curtis; Ackley, Brian D.

    2015-05-04

    in ptp-3B exhibit a low level of embryonic (Emb) and larval lethality (Lva) as well as Variable-abnormal body morphology defects (Vab) (Fig 2 and Table 1). How- ever, most ptp-3 LOF mutants are superficially normal in appearance. Similarly, sdn-1 mutants...002 Table 1. Lethality by genotype analyses. Genotype a Brood Size Avg (St Dev) b Phenotype c Avg (St Dev) N d Emb Lva Vab WT wild type 247.6 (35.6) 0.5 (0.6) 1.0 (0.7) 0.0 (0.0) 98.5 (0.9) 1238 ptp-3(mu245) 82.0 (51.6) 2.1 (1.7) 8.4 (4.7) 2.2 (2.8) 87...

  15. Genome-wide RNAi screen for synthetic lethal interactions with the C. elegans kinesin-5 homolog BMK-1

    PubMed Central

    Maia, André F.; Tanenbaum, Marvin E.; Galli, Matilde; Lelieveld, Daphne; Egan, David A.; Gassmann, Reto; Sunkel, Claudio E.; van den Heuvel, Sander; Medema, René H.

    2015-01-01

    Kinesins are a superfamily of microtubule-based molecular motors that perform various transport needs and have essential roles in cell division. Among these, the kinesin-5 family has been shown to play a major role in the formation and maintenance of the bipolar mitotic spindle. Moreover, recent work suggests that kinesin-5 motors may have additional roles. In contrast to most model organisms, the sole kinesin-5 gene in Caenorhabditis elegans, bmk-1, is not required for successful mitosis and animals lacking bmk-1 are viable and fertile. To gain insight into factors that may act redundantly with BMK-1 in spindle assembly and to identify possible additional cellular pathways involving BMK-1, we performed a synthetic lethal screen using the bmk-1 deletion allele ok391. We successfully knocked down 82% of the C. elegans genome using RNAi and assayed viability in bmk-1(ok391) and wild type strains using an automated high-throughput approach based on fluorescence microscopy. The dataset includes a final list of 37 synthetic lethal interactions whose further study is likely to provide insight into kinesin-5 function. PMID:25984351

  16. Lethal Kinesin Mutations Reveal Amino Acids Important for ATPase Activation and Structural Coupling*

    PubMed Central

    Brendza, Katherine M.; Rose, Debra J.; Gilbert, Susan P.; Saxton, William M.

    2011-01-01

    To study the relationship between conventional kinesin’s structure and function, we identified 13 lethal mutations in the Drosophila kinesin heavy chain motor domain and tested a subset for effects on mechanochemistry. S246F is a moderate mutation that occurs in loop 11 between the ATP- and microtubule-binding sites. While ATP and microtubule binding appear normal, there is a 3-fold decrease in the rate of ATP turnover. This is consistent with the hypothesis that loop 11 provides a structural link that is important for the activation of ATP turnover by microtubule binding. T291M is a severe mutation that occurs in ?-helix 5 near the center of the microtubule-binding surface. It impairs the microtubule-kinesin interaction and directly effects the ATP-binding pocket, allowing an increase in ATP turnover in the absence of microtubules. The T291M mutation may mimic the structure of a microtubule-bound, partially activated state. E164K is a moderate mutation that occurs at the ?-sheet 5a/loop 8b junction, remote from the ATP pocket. Surprisingly, it causes both tighter ATP-binding and a 2-fold decrease in ATP turnover. We propose that E164 forms an ionic bridge with ?-helix 5 and speculate that it helps coordinate the alternating site catalysis of dimerized kinesin heavy chain motor domains. PMID:10531353

  17. A genetic screen for high copy number suppressors of the synthetic lethality between elg1? and srs2? in yeast.

    PubMed

    Gazy, Inbal; Liefshitz, Batia; Bronstein, Alex; Parnas, Oren; Atias, Nir; Sharan, Roded; Kupiec, Martin

    2013-05-01

    Elg1 and Srs2 are two proteins involved in maintaining genome stability in yeast. After DNA damage, the homotrimeric clamp PCNA, which provides stability and processivity to DNA polymerases and serves as a docking platform for DNA repair enzymes, undergoes modification by the ubiquitin-like molecule SUMO. PCNA SUMOylation helps recruit Srs2 and Elg1 to the replication fork. In the absence of Elg1, both SUMOylated PCNA and Srs2 accumulate at the chromatin fraction, indicating that Elg1 is required for removing SUMOylated PCNA and Srs2 from DNA. Despite this interaction, which suggests that the two proteins work together, double mutants elg1? srs2? have severely impaired growth as haploids and exhibit synergistic sensitivity to DNA damage and a synergistic increase in gene conversion. In addition, diploid elg1? srs2? double mutants are dead, which implies that an essential function in the cell requires at least one of the two gene products for survival. To gain information about this essential function, we have carried out a high copy number suppressor screen to search for genes that, when overexpressed, suppress the synthetic lethality between elg1? and srs2?. We report the identification of 36 such genes, which are enriched for functions related to DNA- and chromatin-binding, chromatin packaging and modification, and mRNA export from the nucleus. PMID:23704284

  18. In vivo selection of lethal mutations reveals two functional domains in arginyl-tRNA synthetase.

    PubMed Central

    Geslain, R; Martin, F; Delagoutte, B; Cavarelli, J; Gangloff, J; Eriani, G

    2000-01-01

    Using random mutagenesis and a genetic screening in yeast, we isolated 26 mutations that inactivate Saccharomyces cerevisiae arginyl-tRNA synthetase (ArgRS). The mutations were identified and the kinetic parameters of the corresponding proteins were tested after purification of the expression products in Escherichia coli. The effects were interpreted in the light of the crystal structure of ArgRS. Eighteen functional residues were found around the arginine-binding pocket and eight others in the carboxy-terminal domain of the enzyme. Mutations of these residues all act by strongly impairing the rates of tRNA charging and arginine activation. Thus, ArgRS and tRNA(Arg) can be considered as a kind of ribonucleoprotein, where the tRNA, before being charged, is acting as a cofactor that activates the enzyme. Furthermore, by using different tRNA(Arg) isoacceptors and heterologous tRNA(Asp), we highlighted the crucial role of several residues of the carboxy-terminal domain in tRNA recognition and discrimination. PMID:10744027

  19. Combining chemical genomics screens in yeast to reveal spectrum of effects of chemical inhibition of sphingolipid biosynthesis

    PubMed Central

    2009-01-01

    Background Single genome-wide screens for the effect of altered gene dosage on drug sensitivity in the model organism Saccharomyces cerevisiae provide only a partial picture of the mechanism of action of a drug. Results Using the example of the tumor cell invasion inhibitor dihydromotuporamine C, we show that a more complete picture of drug action can be obtained by combining different chemical genomics approaches – analysis of the sensitivity of ?0 cells lacking mitochondrial DNA, drug-induced haploinsufficiency, suppression of drug sensitivity by gene overexpression and chemical-genetic synthetic lethality screening using strains deleted of nonessential genes. Killing of yeast by this chemical requires a functional mitochondrial electron-transport chain and cytochrome c heme lyase function. However, we find that it does not require genes associated with programmed cell death in yeast. The chemical also inhibits endocytosis and intracellular vesicle trafficking and interferes with vacuolar acidification in yeast and in human cancer cells. These effects can all be ascribed to inhibition of sphingolipid biosynthesis by dihydromotuporamine C. Conclusion Despite their similar conceptual basis, namely altering drug sensitivity by modifying gene dosage, each of the screening approaches provided a distinct set of information that, when integrated, revealed a more complete picture of the mechanism of action of a drug on cells. PMID:19144191

  20. 2D NMR-spectroscopic screening reveals polyketides in ladybugs.

    PubMed

    Deyrup, Stephen T; Eckman, Laura E; McCarthy, Patrick H; Smedley, Scott R; Meinwald, Jerrold; Schroeder, Frank C

    2011-06-14

    Small molecules of biological origin continue to yield the most promising leads for drug design, but systematic approaches for exploring nature's cache of structural diversity are lacking. Here, we demonstrate the use of 2D NMR spectroscopy to screen a library of biorationally selected insect metabolite samples for partial structures indicating the presence of new chemical entities. This NMR-spectroscopic survey enabled detection of novel compounds in complex metabolite mixtures without prior fractionation or isolation. Our screen led to discovery and subsequent isolation of two families of tricyclic pyrones in Delphastus catalinae, a tiny ladybird beetle that is employed commercially as a biological pest control agent. The D. catalinae pyrones are based on 23-carbon polyketide chains forming 1,11-dioxo-2,6,10-trioxaanthracene and 4,8-dioxo-1,9,13-trioxaanthracene derivatives, representing ring systems not previously found in nature. This study highlights the utility of 2D NMR-spectroscopic screening for exploring nature's structure space and suggests that insect metabolomes remain vastly underexplored. PMID:21646540

  1. RESEARCH Open Access Genome-wide RNAi screen reveals the E3

    E-print Network

    Zhao, Huimin

    RESEARCH Open Access Genome-wide RNAi screen reveals the E3 SUMO-protein ligase gene SIZ1 biomass. In this study, we used the RNAi-Assisted Genome Evolution (RAGE) method to select for furfural. Results: By using a genome-wide RNAi (RNA-interference) screen in S. cerevisiae for genes involved

  2. Genome-wide RNAi screen reveals a role for the ESCRT complex in rotavirus cell entry

    E-print Network

    Perrimon, Norbert

    Genome-wide RNAi screen reveals a role for the ESCRT complex in rotavirus cell entry Daniela Silva) Rotavirus (RV) is the major cause of childhood gastroenteritis worldwide. This study presents a functional for comprehen- sively studying the interaction between viruses and their host cells. Rotaviruses (RVs), members

  3. Histopathology reveals correlative and unique phenotypes in a high-throughput mouse phenotyping screen.

    PubMed

    Adissu, Hibret A; Estabel, Jeanne; Sunter, David; Tuck, Elizabeth; Hooks, Yvette; Carragher, Damian M; Clarke, Kay; Karp, Natasha A; Newbigging, Susan; Jones, Nora; Morikawa, Lily; White, Jacqueline K; McKerlie, Colin

    2014-05-01

    The Mouse Genetics Project (MGP) at the Wellcome Trust Sanger Institute aims to generate and phenotype over 800 genetically modified mouse lines over the next 5 years to gain a better understanding of mammalian gene function and provide an invaluable resource to the scientific community for follow-up studies. Phenotyping includes the generation of a standardized biobank of paraffin-embedded tissues for each mouse line, but histopathology is not routinely performed. In collaboration with the Pathology Core of the Centre for Modeling Human Disease (CMHD) we report the utility of histopathology in a high-throughput primary phenotyping screen. Histopathology was assessed in an unbiased selection of 50 mouse lines with (n=30) or without (n=20) clinical phenotypes detected by the standard MGP primary phenotyping screen. Our findings revealed that histopathology added correlating morphological data in 19 of 30 lines (63.3%) in which the primary screen detected a phenotype. In addition, seven of the 50 lines (14%) presented significant histopathology findings that were not associated with or predicted by the standard primary screen. Three of these seven lines had no clinical phenotype detected by the standard primary screen. Incidental and strain-associated background lesions were present in all mutant lines with good concordance to wild-type controls. These findings demonstrate the complementary and unique contribution of histopathology to high-throughput primary phenotyping of mutant mice. PMID:24652767

  4. NCI: SBIR & STTR - Find Funding - Contracts - 290 siRNA Resource for Synthetic Lethal Screening of DNA Repair and Damage Signaling Networks

    Cancer.gov

    Human cancers appear to be highly and differentially vulnerable to targeted attack on individual gene expression within their DNA repair networks, especially those pathways that are closely associated with DNA replication. However such lethal combinations (i.e., synthetic lethals) are generally not obvious or easily predicted a priori.

  5. A sensitised RNAi screen reveals a ch-TOG genetic interaction network required for spindle assembly.

    PubMed

    Barr, Alexis R; Bakal, Chris

    2015-01-01

    How multiple spindle assembly pathways are integrated to drive bipolar spindle assembly is poorly understood. We performed an image-based double RNAi screen to identify genes encoding Microtubule-Associated Proteins (MAPs) that interact with the highly conserved ch-TOG gene to regulate bipolar spindle assembly in human cells. We identified a ch-TOG centred network of genetic interactions which promotes ensures centrosome-mediated microtubule polymerisation, leading to the incorporation of microtubules polymerised by all pathways into a bipolar structure. Our genetic screen also reveals that ch-TOG maintains a dynamic microtubule population, in part, through modulating HSET activity. ch-TOG ensures that spindle assembly is robust to perturbation but sufficiently dynamic such that spindles can explore a diverse shape space in search of structures that can align chromosomes. PMID:26037491

  6. A sensitised RNAi screen reveals a ch-TOG genetic interaction network required for spindle assembly

    PubMed Central

    Barr, Alexis R.; Bakal, Chris

    2015-01-01

    How multiple spindle assembly pathways are integrated to drive bipolar spindle assembly is poorly understood. We performed an image-based double RNAi screen to identify genes encoding Microtubule-Associated Proteins (MAPs) that interact with the highly conserved ch-TOG gene to regulate bipolar spindle assembly in human cells. We identified a ch-TOG centred network of genetic interactions which promotes ensures centrosome-mediated microtubule polymerisation, leading to the incorporation of microtubules polymerised by all pathways into a bipolar structure. Our genetic screen also reveals that ch-TOG maintains a dynamic microtubule population, in part, through modulating HSET activity. ch-TOG ensures that spindle assembly is robust to perturbation but sufficiently dynamic such that spindles can explore a diverse shape space in search of structures that can align chromosomes. PMID:26037491

  7. Genome-Wide Protein Interaction Screens Reveal Functional Networks Involving Sm-Like Proteins

    PubMed Central

    Fromont-Racine, Micheline; Mayes, Andrew E.; Brunet-Simon, Adeline; Rain, Jean-Christophe; Colley, Alan; Dix, Ian; Decourty, Laurence; Joly, Nicolas; Ricard, Florence; Beggs, Jean D.

    2000-01-01

    A set of seven structurally related Sm proteins forms the core of the snRNP particles containing the spliceosomal U1, U2, U4 and U5 snRNAs. A search of the genomic sequence of Saccharomyces cerevisiae has identified a number of open reading frames that potentially encode structurally similar proteins termed Lsm (Like Sm) proteins. With the aim of analysing all possible interactions between the Lsm proteins and any protein encoded in the yeast genome, we performed exhaustive and iterative genomic two-hybrid screens, starting with the Lsm proteins as baits. Indeed, extensive interactions amongst eight Lsm proteins were found that suggest the existence of a Lsm complex or complexes. These Lsm interactions apparently involve the conserved Sm domain that also mediates interactions between the Sm proteins. The screens also reveal functionally significant interactions with splicing factors, in particular with Prp4 and Prp24, compatible with genetic studies and with the reported association of Lsm proteins with spliceosomal U6 and U4/U6 particles. In addition, interactions with proteins involved in mRNA turnover, such as Mrt1, Dcp1, Dcp2 and Xrn1, point to roles for Lsm complexes in distinct RNA metabolic processes, that are confirmed in independent functional studies. These results provide compelling evidence that two-hybrid screens yield functionally meaningful information about protein–protein interactions and can suggest functions for uncharacterized proteins, especially when they are performed on a genome-wide scale. PMID:10900456

  8. Genetic Modifier Screens Reveal New Components that Interact with the Drosophila Dystroglycan-Dystrophin Complex

    PubMed Central

    Yatsenko, Andriy S.; Shcherbata, Halyna R.; Fischer, Karin A.; Maksymiv, Dariya V.; Chernyk, Yaroslava I.; Ruohola-Baker, Hannele

    2008-01-01

    The Dystroglycan-Dystrophin (Dg-Dys) complex has a capacity to transmit information from the extracellular matrix to the cytoskeleton inside the cell. It is proposed that this interaction is under tight regulation; however the signaling/regulatory components of Dg-Dys complex remain elusive. Understanding the regulation of the complex is critical since defects in this complex cause muscular dystrophy in humans. To reveal new regulators of the Dg-Dys complex, we used a model organism Drosophila melanogaster and performed genetic interaction screens to identify modifiers of Dg and Dys mutants in Drosophila wing veins. These mutant screens revealed that the Dg-Dys complex interacts with genes involved in muscle function and components of Notch, TGF-? and EGFR signaling pathways. In addition, components of pathways that are required for cellular and/or axonal migration through cytoskeletal regulation, such as Semaphorin-Plexin, Frazzled-Netrin and Slit-Robo pathways show interactions with Dys and/or Dg. These data suggest that the Dg-Dys complex and the other pathways regulating extracellular information transfer to the cytoskeletal dynamics are more intercalated than previously thought. PMID:18545683

  9. High-throughput sequencing screen reveals novel, transforming RAS mutations in myeloid leukemia patients

    PubMed Central

    Tyner, Jeffrey W.; Erickson, Heidi; Deininger, Michael W. N.; Willis, Stephanie G.; Eide, Christopher A.; Levine, Ross L.; Heinrich, Michael C.; Gattermann, Norbert; Gilliland, D. Gary; Druker, Brian J.

    2009-01-01

    Transforming mutations in NRAS and KRAS are thought to play a causative role in the development of numerous cancers, including myeloid malignancies. Although mutations at amino acids 12, 13, or 61 account for the majority of oncogenic Ras variants, we hypothesized that less frequent mutations at alternate residues may account for disease in some patients with cancer of unexplained genetic etiology. To search for additional, novel RAS mutations, we sequenced all coding exons in NRAS, KRAS, and HRAS in 329 acute myeloid leukemia (AML) patients, 32 chronic myelomonocytic leukemia (CMML) patients, and 96 healthy individuals. We detected 4 “noncanonical” point mutations in 7 patients: N-RasG60E, K-RasV14I, K-RasT74P, and K-RasA146T. All 4 Ras mutants exhibited oncogenic properties in comparison with wild-type Ras in biochemical and functional assays. The presence of transforming RAS mutations outside of positions 12, 13, and 61 reveals that alternate mechanisms of transformation by RAS may be overlooked in screens designed to detect only the most common RAS mutations. Our results suggest that RAS mutations may play a greater role in leukemogenesis than currently believed and indicate that high-throughput screening for mutant RAS alleles in cancer should include analysis of the entire RAS coding region. PMID:19075190

  10. Genome-wide RNAi screen for nuclear actin reveals a network of cofilin regulators.

    PubMed

    Dopie, Joseph; Rajakylä, Eeva K; Joensuu, Merja S; Huet, Guillaume; Ferrantelli, Evelina; Xie, Tiao; Jäälinoja, Harri; Jokitalo, Eija; Vartiainen, Maria K

    2015-07-01

    Nuclear actin plays an important role in many processes that regulate gene expression. Cytoplasmic actin dynamics are tightly controlled by numerous actin-binding proteins, but regulation of nuclear actin has remained unclear. Here, we performed a genome-wide RNA interference (RNAi) screen in Drosophila cells to identify proteins that influence either nuclear polymerization or import of actin. We validate 19 factors as specific hits, and show that Chinmo (known as Bach2 in mammals), SNF4A? (Prkag1 in mammals) and Rab18 play a role in nuclear localization of actin in both fly and mammalian cells. We identify several new regulators of cofilin activity, and characterize modulators of both cofilin kinases and phosphatase. For example, Chinmo/Bach2, which regulates nuclear actin levels also in vivo, maintains active cofilin by repressing the expression of the kinase Cdi (Tesk in mammals). Finally, we show that Nup98 and lamin are candidates for regulating nuclear actin polymerization. Our screen therefore reveals new aspects of actin regulation and links nuclear actin to many cellular processes. PMID:26021350

  11. Physical and genetic-interaction density reveals functional organization and informs significance cutoffs in genome-wide screens

    PubMed Central

    Dittmar, John C.; Pierce, Steven; Rothstein, Rodney; Reid, Robert J. D.

    2013-01-01

    Genome-wide experiments often measure quantitative differences between treated and untreated cells to identify affected strains. For these studies, statistical models are typically used to determine significance cutoffs. We developed a method termed “CLIK” (Cutoff Linked to Interaction Knowledge) that overlays biological knowledge from the interactome on screen results to derive a cutoff. The method takes advantage of the fact that groups of functionally related interacting genes often respond similarly to experimental conditions and, thus, cluster in a ranked list of screen results. We applied CLIK analysis to five screens of the yeast gene disruption library and found that it defined a significance cutoff that differed from traditional statistics. Importantly, verification experiments revealed that the CLIK cutoff correlated with the position in the rank order where the rate of true positives drops off significantly. In addition, the gene sets defined by CLIK analysis often provide further biological perspectives. For example, applying CLIK analysis retrospectively to a screen for cisplatin sensitivity allowed us to identify the importance of the Hrq1 helicase in DNA crosslink repair. Furthermore, we demonstrate the utility of CLIK to determine optimal treatment conditions by analyzing genome-wide screens at multiple rapamycin concentrations. We show that CLIK is an extremely useful tool for evaluating screen quality, determining screen cutoffs, and comparing results between screens. Furthermore, because CLIK uses previously annotated interaction data to determine biologically informed cutoffs, it provides additional insights into screen results, which supplement traditional statistical approaches. PMID:23589890

  12. Disease fingerprinting with cDNA microarrays reveals distinct gene expression profiles in lethal type-1 and type-2 cytokine-mediated inflammatory reactions

    Microsoft Academic Search

    Karl F. Hoffmann; Thomas C. McCarty; David H. Segal; Monica Chiaramonte; Matthias Hesse; Eric M. Davis; Allen W. Cheever; Paul S. Meltzer; Herbert C. Morse; Thomas A. Wynn

    2001-01-01

    Development of polarized immune responses controls resistance and susceptibility to many microorganisms. However, studies of several inf ectious, allergic, and autoimmune diseases have shown that chronic type-1 and type-2 cytokine responses can also cause significant morbidity and mortality if left unchecked. We used mouse cDNA microarrays to molecularly phenotype the gene expression patterns that characterize two disparate but equally lethal

  13. Functional Screening of Hydrolytic Activities Reveals an Extremely Thermostable Cellulase from a Deep-Sea Archaeon

    PubMed Central

    Leis, Benedikt; Heinze, Simon; Angelov, Angel; Pham, Vu Thuy Trang; Thürmer, Andrea; Jebbar, Mohamed; Golyshin, Peter N.; Streit, Wolfgang R.; Daniel, Rolf; Liebl, Wolfgang

    2015-01-01

    Extreme habitats serve as a source of enzymes that are active under extreme conditions and are candidates for industrial applications. In this work, six large-insert mixed genomic libraries were screened for hydrolase activities in a broad temperature range (8–70°C). Among a variety of hydrolytic activities, one fosmid clone, derived from a library of pooled isolates of hyperthermophilic archaea from deep sea vents, displayed hydrolytic activity on carboxymethyl cellulose substrate plates at 70°C but not at lower temperatures. Sequence analysis of the fosmid insert revealed a gene encoding a novel glycoside hydrolase family 12 (GHF12) endo-1,4-?-glucanase, termed Cel12E. The enzyme shares 45% sequence identity with a protein from the archaeon Thermococcus sp. AM4 and displays a unique multidomain architecture. Biochemical characterization of Cel12E revealed a remarkably thermostable protein, which appears to be of archaeal origin. The enzyme displayed maximum activity at 92°C and was active on a variety of linear 1,4-?-glucans like carboxymethyl cellulose, ?-glucan, lichenan, and phosphoric acid swollen cellulose. The protein is able to bind to various insoluble ?-glucans. Product pattern analysis indicated that Cel12E is an endo-cleaving ?-glucanase. Cel12E expands the toolbox of hyperthermostable archaeal cellulases with biotechnological potential.

  14. Functional Screening of Hydrolytic Activities Reveals an Extremely Thermostable Cellulase from a Deep-Sea Archaeon.

    PubMed

    Leis, Benedikt; Heinze, Simon; Angelov, Angel; Pham, Vu Thuy Trang; Thürmer, Andrea; Jebbar, Mohamed; Golyshin, Peter N; Streit, Wolfgang R; Daniel, Rolf; Liebl, Wolfgang

    2015-01-01

    Extreme habitats serve as a source of enzymes that are active under extreme conditions and are candidates for industrial applications. In this work, six large-insert mixed genomic libraries were screened for hydrolase activities in a broad temperature range (8-70°C). Among a variety of hydrolytic activities, one fosmid clone, derived from a library of pooled isolates of hyperthermophilic archaea from deep sea vents, displayed hydrolytic activity on carboxymethyl cellulose substrate plates at 70°C but not at lower temperatures. Sequence analysis of the fosmid insert revealed a gene encoding a novel glycoside hydrolase family 12 (GHF12) endo-1,4-?-glucanase, termed Cel12E. The enzyme shares 45% sequence identity with a protein from the archaeon Thermococcus sp. AM4 and displays a unique multidomain architecture. Biochemical characterization of Cel12E revealed a remarkably thermostable protein, which appears to be of archaeal origin. The enzyme displayed maximum activity at 92°C and was active on a variety of linear 1,4-?-glucans like carboxymethyl cellulose, ?-glucan, lichenan, and phosphoric acid swollen cellulose. The protein is able to bind to various insoluble ?-glucans. Product pattern analysis indicated that Cel12E is an endo-cleaving ?-glucanase. Cel12E expands the toolbox of hyperthermostable archaeal cellulases with biotechnological potential. PMID:26191525

  15. Host Pathways Important for Coxiella burnetii Infection Revealed by Genome-Wide RNA Interference Screening

    PubMed Central

    McDonough, Justin A.; Newton, Hayley J.; Klum, Scott; Swiss, Rachel; Agaisse, Hervé; Roy, Craig R.

    2013-01-01

    ABSTRACT Coxiella burnetii is an intracellular pathogen that replicates within a lysosome-like vacuole. A Dot/Icm type IVB secretion system is used by C. burnetii to translocate effector proteins into the host cytosol that likely modulate host factor function. To identify host determinants required for C. burnetii intracellular growth, a genome-wide screen was performed using gene silencing by small interfering RNA (siRNA). Replication of C. burnetii was measured by immunofluorescence microscopy in siRNA-transfected HeLa cells. Newly identified host factors included components of the retromer complex, which mediates cargo cycling between the endocytic pathway and the Golgi apparatus. Reducing the levels of the retromer cargo-adapter VPS26-VPS29-VPS35 complex or retromer-associated sorting nexins abrogated C. burnetii replication. Several genes, when silenced, resulted in enlarged vacuoles or an increased number of vacuoles within C. burnetii-infected cells. Silencing of the STX17 gene encoding syntaxin-17 resulted in a striking defect in homotypic fusion of vacuoles containing C. burnetii, suggesting a role for syntaxin-17 in regulating this process. Lastly, silencing host genes needed for C. burnetii replication correlated with defects in the translocation of Dot/Icm effectors, whereas, silencing of genes that affected vacuole morphology, but did not impact replication, did not affect Dot/Icm translocation. These data demonstrate that C. burnetii vacuole maturation is important for creating a niche that permits Dot/Icm function. Thus, genome-wide screening has revealed host determinants involved in sequential events that occur during C. burnetii infection as defined by bacterial uptake, vacuole transport and acidification, activation of the Dot/Icm system, homotypic fusion of vacuoles, and intracellular replication. PMID:23362322

  16. A Genetic Screen for Sleep and Circadian Mutants Reveals Mechanisms Underlying Regulation of Sleep in Drosophila

    PubMed Central

    Wu, Mark N.; Koh, Kyunghee; Yue, Zhifeng; Joiner, William J.; Sehgal, Amita

    2008-01-01

    Study Objectives: In order to characterize the genetic mechanisms underlying sleep, we have carried out a large-scale screen in Drosophila to identify short-sleeping mutants. The objectives of this study were as follows: (1) characterize the phenotypes of the shortest-sleeping mutants; (2) examine whether changes in arousal threshold or sleep homeostasis underlie short-sleeping phenotypes; (3) clone a gene affected in one of the shortest sleepers; and (4) investigate whether circadian mutants can be identified using light:dark (L:D) locomotor data obtained for studying sleep behavior. Design: Locomotor activity was measured using the Drosophila Activity Monitoring System in a 12:12 L:D cycle. Setting: Drosophila research laboratory. Participants: Adult flies from the 2nd chromosome Zuker collection, which contain mutations in most of the nonessential genes on the Drosophila 2nd chromosome. Measurements and Results: Our analysis of sleep characteristics suggests that daily activity (but not waking activity) correlates with daily sleep time and that defects in sleep maintenance are more common than defects in sleep initiation. Our shortest sleepers have intact or increased sleep rebound following sleep deprivation but show reduced thresholds for arousal. Molecular analysis of one of the short-sleeping lines indicates that it is a novel allele of a dopamine transporter (DAT). Finally, we describe a novel approach for identifying circadian mutants using L:D data. Conclusions: Our data suggest that most short-sleeping mutant phenotypes in Drosophila are characterized by an inability to stay asleep, most likely because of a reduced arousal threshold. Citation: Wu MN; Koh K; Yue Z; Joiner WJ; Sehgal A. A genetic screen for sleep and circadian mutants reveals mechanisms underlying regulation of sleep in Drosophila. SLEEP 2008;31(4):465-472. PMID:18457233

  17. Elaborate cellulosome architecture of Acetivibrio cellulolyticus revealed by selective screening of cohesin-dockerin interactions.

    PubMed

    Hamberg, Yuval; Ruimy-Israeli, Vered; Dassa, Bareket; Barak, Yoav; Lamed, Raphael; Cameron, Kate; Fontes, Carlos M G A; Bayer, Edward A; Fried, Daniel B

    2014-01-01

    Cellulosic waste represents a significant and underutilized carbon source for the biofuel industry. Owing to the recalcitrance of crystalline cellulose to enzymatic degradation, it is necessary to design economical methods of liberating the fermentable sugars required for bioethanol production. One route towards unlocking the potential of cellulosic waste lies in a highly complex class of molecular machines, the cellulosomes. Secreted mainly by anaerobic bacteria, cellulosomes are structurally diverse, cell surface-bound protein assemblies that can contain dozens of catalytic components. The key feature of the cellulosome is its modularity, facilitated by the ultra-high affinity cohesin-dockerin interaction. Due to the enormous number of cohesin and dockerin modules found in a typical cellulolytic organism, a major bottleneck in understanding the biology of cellulosomics is the purification of each cohesin- and dockerin-containing component, prior to analyses of their interaction. As opposed to previous approaches, the present study utilized proteins contained in unpurified whole-cell extracts. This strategy was made possible due to an experimental design that allowed for the relevant proteins to be "purified" via targeted affinity interactions as a function of the binding assay. The approach thus represents a new strategy, appropriate for future medium- to high-throughput screening of whole genomes, to determine the interactions between cohesins and dockerins. We have selected the cellulosome of Acetivibrio cellulolyticus for this work due to its exceptionally complex cellulosome systems and intriguing diversity of its cellulosomal modular components. Containing 41 cohesins and 143 dockerins, A. cellulolyticus has one of the largest number of potential cohesin-dockerin interactions of any organism, and contains unusual and novel cellulosomal features. We have surveyed a representative library of cohesin and dockerin modules spanning the cellulosome's total cohesin and dockerin sequence diversity, emphasizing the testing of unusual and previously-unknown protein modules. The screen revealed several novel cell-bound cellulosome architectures, thus expanding on those previously known, as well as soluble cellulose systems that are not bound to the bacterial cell surface. This study sets the stage for screening the entire complement of cellulosomal components from A. cellulolyticus and other organisms with large cellulosome systems. The knowledge gained by such efforts brings us closer to understanding the exceptional catalytic abilities of cellulosomes and will allow the use of novel cellulosomal components in artificial assemblies and in enzyme cocktails for sustainable energy-related research programs. PMID:25374780

  18. A Genetic Screen and Transcript Profiling Reveal a Shared Regulatory Program for Drosophila Linker Histone H1 and Chromatin Remodeler CHD1

    PubMed Central

    Kavi, Harsh; Lu, Xingwu; Xu, Na; Bartholdy, Boris A.; Vershilova, Elena; Skoultchi, Arthur I.; Fyodorov, Dmitry V.

    2015-01-01

    Chromatin structure and activity can be modified through ATP-dependent repositioning of nucleosomes and posttranslational modifications of core histone tails within nucleosome core particles and by deposition of linker histones into the oligonucleosome fiber. The linker histone H1 is essential in metazoans. It has a profound effect on organization of chromatin into higher-order structures and on recruitment of histone-modifying enzymes to chromatin. Here, we describe a genetic screen for modifiers of the lethal phenotype caused by depletion of H1 in Drosophila melanogaster. We identify 41 mis-expression alleles that enhance and 20 that suppress the effect of His1 depletion in vivo. Most of them are important for chromosome organization, transcriptional regulation, and cell signaling. Specifically, the reduced viability of H1-depleted animals is strongly suppressed by ubiquitous mis-expression of the ATP-dependent chromatin remodeling enzyme CHD1. Comparison of transcript profiles in H1-depleted and Chd1 null mutant larvae revealed that H1 and CHD1 have common transcriptional regulatory programs in vivo. H1 and CHD1 share roles in repression of numerous developmentally regulated and extracellular stimulus-responsive transcripts, including immunity-related and stress response-related genes. Thus, linker histone H1 participates in various regulatory programs in chromatin to alter gene expression. PMID:25628309

  19. A genetic screen and transcript profiling reveal a shared regulatory program for Drosophila linker histone H1 and chromatin remodeler CHD1.

    PubMed

    Kavi, Harsh; Lu, Xingwu; Xu, Na; Bartholdy, Boris A; Vershilova, Elena; Skoultchi, Arthur I; Fyodorov, Dmitry V

    2015-04-01

    Chromatin structure and activity can be modified through ATP-dependent repositioning of nucleosomes and posttranslational modifications of core histone tails within nucleosome core particles and by deposition of linker histones into the oligonucleosome fiber. The linker histone H1 is essential in metazoans. It has a profound effect on organization of chromatin into higher-order structures and on recruitment of histone-modifying enzymes to chromatin. Here, we describe a genetic screen for modifiers of the lethal phenotype caused by depletion of H1 in Drosophila melanogaster. We identify 41 mis-expression alleles that enhance and 20 that suppress the effect of His1 depletion in vivo. Most of them are important for chromosome organization, transcriptional regulation, and cell signaling. Specifically, the reduced viability of H1-depleted animals is strongly suppressed by ubiquitous mis-expression of the ATP-dependent chromatin remodeling enzyme CHD1. Comparison of transcript profiles in H1-depleted and Chd1 null mutant larvae revealed that H1 and CHD1 have common transcriptional regulatory programs in vivo. H1 and CHD1 share roles in repression of numerous developmentally regulated and extracellular stimulus-responsive transcripts, including immunity-related and stress response-related genes. Thus, linker histone H1 participates in various regulatory programs in chromatin to alter gene expression. PMID:25628309

  20. Characterization of Synthetic-Lethal Mutants Reveals a Role for the Saccharomyces Cerevisiae Guanine-Nucleotide Exchange Factor Cdc24p in Vacuole Function and Na(+) Tolerance

    PubMed Central

    White, W. H.; Johnson, D. I.

    1997-01-01

    Cdc24p is the guanine-nucleotide exchange factor for the Cdc42p GTPase, which controls cell polarity in Saccharomyces cerevisiae. To identify new genes that may affect cell polarity, we characterized six UV-induced csl (CDC24 synthetic-lethal) mutants that exhibited synthetic-lethality with cdc24-4(ts) at 23°. Five mutants were not complemented by plasmid-borne CDC42, RSR1, BUD5, BEM1, BEM2, BEM3 or CLA4 genes, which are known to play a role in cell polarity. The csl3 mutant displayed phenotypes similar to those observed with calcium-sensitive, Pet(-) vma mutants defective in vacuole function. CSL5 was allelic to VMA5, the vacuolar H(+)-ATPase subunit C, and one third of csl5 cdc24-4(ts) cells were elongated or had misshapen buds. A cdc24-4(ts) ?vma5::LEU2 double mutant did not exhibit synthetic lethality, suggesting that the csl5/vma5 cdc24-4(ts) synthetic-lethality was not simply due to altered vacuole function. The cdc24-4(ts) mutant, like ?vma5::LEU2 and csl3 mutants, was sensitive to high levels of Ca(2+) as well as Na(+) in the growth media, which did not appear to be a result of a fragile cell wall because the phenotypes were not remedied by 1 M sorbitol. Our results indicated that Cdc24p was required in one V-ATPase mutant and another mutant affecting vacuole morphology, and also implicated Cdc24p in Na(+) tolerance. PMID:9286667

  1. A single-cell imaging screen reveals multiple effects of secreted small molecules on bacteria

    PubMed Central

    Salje, Jeanne

    2014-01-01

    Bacteria cells exist in close proximity to other cells of both the same and different species. Bacteria secrete a large number of different chemical species, and the local concentrations of these compounds at the surfaces of nearby cells may reach very high levels. It is fascinating to imagine how individual cells might sense and respond to the complex mix of signals at their surface. However, it is difficult to measure exactly what the local environmental composition looks like, or what the effects of individual compounds on nearby cells are. Here, an electron microscopy imaging screen was designed that would detect morphological changes induced by secreted small molecules. This differs from conventional approaches by detecting structural changes in individual cells rather than gene expression or growth rate changes at the population level. For example, one of the changes detected here was an increase in outer membrane vesicle production, which does not necessarily correspond to a change in gene expression. This initial study focussed on Pseudomonas aeruginosa, Escherichia coli, and Burkholderia dolosa, and revealed an intriguing range of effects of secreted small molecules on cells both within and between species. PMID:24910069

  2. In Vivo RNAi Screen Reveals Neddylation Genes as Novel Regulators of Hedgehog Signaling

    PubMed Central

    Su, Ying; Liu, Min; Ospina, Jason K.; Yang, Shengyuan; Zhu, Alan Jian

    2011-01-01

    Hedgehog (Hh) signaling is highly conserved in all metazoan animals and plays critical roles in many developmental processes. Dysregulation of the Hh signaling cascade has been implicated in many diseases, including cancer. Although key components of the Hh pathway have been identified, significant gaps remain in our understanding of the regulation of individual Hh signaling molecules. Here, we report the identification of novel regulators of the Hh pathway, obtained from an in vivo RNA interference (RNAi) screen in Drosophila. By selectively targeting critical genes functioning in post-translational modification systems utilizing ubiquitin (Ub) and Ub-like proteins, we identify two novel genes (dUba3 and dUbc12) that negatively regulate Hh signaling activity. We provide in vivo and in vitro evidence illustrating that dUba3 and dUbc12 are essential components of the neddylation pathway; they function in an enzyme cascade to conjugate the ubiquitin-like NEDD8 modifier to Cullin proteins. Neddylation activates the Cullin-containing ubiquitin ligase complex, which in turn promotes the degradation of Cubitus interruptus (Ci), the downstream transcription factor of the Hh pathway. Our study reveals a conserved molecular mechanism of the neddylation pathway in Drosophila and sheds light on the complex post-translational regulations in Hh signaling. PMID:21931660

  3. Cell array-based intracellular localization screening reveals novel functional features of human chromosome 21 proteins

    PubMed Central

    Hu, Yu-Hui; Warnatz, Hans-Jörg; Vanhecke, Dominique; Wagner, Florian; Fiebitz, Andrea; Thamm, Sabine; Kahlem, Pascal; Lehrach, Hans; Yaspo, Marie-Laure; Janitz, Michal

    2006-01-01

    Background Trisomy of human chromosome 21 (Chr21) results in Down's syndrome, a complex developmental and neurodegenerative disease. Molecular analysis of Down's syndrome, however, poses a particular challenge, because the aneuploid region of Chr21 contains many genes of unknown function. Subcellular localization of human Chr21 proteins may contribute to further understanding of the functions and regulatory mechanisms of the genes that code for these proteins. Following this idea, we used a transfected-cell array technique to perform a rapid and cost-effective analysis of the intracellular distribution of Chr 21 proteins. Results We chose 89 genes that were distributed over the majority of 21q, ranging from RBM11 (14.5 Mb) to MCM3AP (46.6 Mb), with part of them expressed aberrantly in the Down's syndrome mouse model. Open reading frames of these genes were cloned into a mammalian expression vector with an amino-terminal His6 tag. All of the constructs were arrayed on glass slides and reverse transfected into HEK293T cells for protein expression. Co-localization detection using a set of organelle markers was carried out for each Chr21 protein. Here, we report the subcellular localization properties of 52 proteins. For 34 of these proteins, their localization is described for the first time. Furthermore, the alteration in cell morphology and growth as a result of protein over-expression for claudin-8 and claudin-14 genes has been characterized. Conclusion The cell array-based protein expression and detection approach is a cost-effective platform for large-scale functional analyses, including protein subcellular localization and cell phenotype screening. The results from this study reveal novel functional features of human Chr21 proteins, which should contribute to further understanding of the molecular pathology of Down's syndrome. PMID:16780588

  4. A genome-wide visual screen reveals a role for sphingolipids and ergosterol in cell surface delivery in yeast

    PubMed Central

    Proszynski, Tomasz J.; Klemm, Robin W.; Gravert, Maike; Hsu, Peggy P.; Gloor, Yvonne; Wagner, Jan; Kozak, Karol; Grabner, Hannes; Walzer, Karen; Bagnat, Michel; Simons, Kai; Walch-Solimena, Christiane

    2005-01-01

    Recently synthesized proteins are sorted at the trans-Golgi network into specialized routes for exocytosis. Surprisingly little is known about the underlying molecular machinery. Here, we present a visual screen to search for proteins involved in cargo sorting and vesicle formation. We expressed a GFP-tagged plasma membrane protein in the yeast deletion library and identified mutants with altered marker localization. This screen revealed a requirement of several enzymes regulating the synthesis of sphingolipids and ergosterol in the correct and efficient delivery of the marker protein to the cell surface. Additionally, we identified mutants regulating the actin cytoskeleton (Rvs161p and Vrp1p), known membrane traffic regulators (Kes1p and Chs5p), and several unknown genes. This visual screening method can now be used for different cargo proteins to search in a genome-wide fashion for machinery involved in post-Golgi sorting. PMID:16330752

  5. MicroSCALE Screening Reveals Genetic Modifiers of Therapeutic Response in Melanoma

    E-print Network

    Sabatini, David M.

    Cell microarrays are a promising tool for performing large-scale functional genomic screening in mammalian cells at reasonable cost, but owing to technical limitations they have been restricted for use with a narrow range ...

  6. Antileishmanial High-Throughput Drug Screening Reveals Drug Candidates with New Scaffolds

    E-print Network

    Paris-Sud XI, Université de

    ), Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea, 2 Screening Technology & Pharmacology Group, Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea, 3 Active Compound Space Group, Institut Pasteur Korea, Seongnam-si, Gyeonggi-do, South Korea, 4 Image Mining Group, Institut Pasteur Korea

  7. High-throughput random mutagenesis screen reveals TRPM8 residues specifically required for activation by menthol

    Microsoft Academic Search

    Michael Bandell; Adrienne E Dubin; Matt J Petrus; Anthony Orth; Jayanti Mathur; Sun Wook Hwang; Ardem Patapoutian

    2006-01-01

    Menthol is a cooling compound derived from mint leaves and is extensively used as a flavoring chemical. Menthol activates transient receptor potential melastatin 8 (TRPM8), an ion channel also activated by cold, voltage and phosphatidylinositol-4,5-bisphosphate (PIP2). Here we investigated the mechanism by which menthol activates mouse TRPM8. Using a new high-throughput approach, we screened a random mutant library consisting of

  8. An integrated functional genomics screening program reveals a role for BMP9 in glucose homeostasis

    Microsoft Academic Search

    Cecil Chen; Krzysztof J. Grzegorzewski; Steve Barash; Qinghai Zhao; Helmut Schneider; Qi Wang; Mallika Singh; Laurie Pukac; Adam C. Bell; Roxanne Duan; Tim Coleman; Alokesh Duttaroy; Susan Cheng; Jon Hirsch; Linyi Zhang; Yanick Lazard; Carrie Fischer; Melisa Carey Barber; Zhi-Dong Ma; Ya-Qin Zhang; Peter Reavey; Lilin Zhong; Baiqin Teng; Indra Sanyal; Steve M. Ruben; Olivier Blondel; Charles E. Birse

    2003-01-01

    A coordinated functional genomics program was implemented to identify secreted polypeptides with therapeutic applications in the treatment of diabetes. Secreted factors were predicted from a diverse expressed-sequence tags (EST) database, representing >1,000 cDNA libraries, using a combination of bioinformatic algorithms. Subsequently, ?8,000 human proteins were screened in high-throughput cell-based assays designed to monitor key physiological transitions known to be centrally

  9. Systematic combination screening reveals synergism between rapamycin and sunitinib against human lung cancer.

    PubMed

    Li, Xian; Tong, Lin-Jiang; Ding, Jian; Meng, Ling-Hua

    2014-01-01

    Mammalian target of rapamycin (mTOR) acts as a hub integrating signals from nutrient availability and growth factors and plays central roles in regulating protein synthesis and cell growth, which has been validated as a promising target for cancer therapy. Rapamycin and its analogues have emerged as the first generation of mTOR inhibitors, but their efficacy is modest in clinical settings. Combinatorial use of rapamycin with other drugs is a promising strategy to improve its anticancer activity. Here we developed an unbiased systematic binary screening platform aiming to discover new remedy for rapamycin-based cancer therapy. We found that sunitinib emerged as one of the clinically available anticancer drugs screened that displayed significant synergy with rapamycin in NSCLC cells. Combination of rapamycin with sunitinib resulted in enhanced cell cycle arrest in G1 phase, which was accompanied with enhanced suppression of mTOR signaling and disruption of the negative feedback loop that activate AKT upon mTORC1 inhibition. Furthermore, sunitinib and rapamycin displayed synergistic activity against tube formation by human microvessel endothelial cells as well as outgrowth of endothelial tubes and microvessels both in vitro and in vivo, which is associated with down-regulation of VEGF secretion and HIF1? expression. Our study demonstrated that new combinatorial regimen could be identified via systematic drug combination screening and established a mechanistic rationale for a combination approach using rapalogs and sunitinib in the treatment of human NSCLC. PMID:24018642

  10. A Targeted RNA Interference Screen Reveals Novel Epigenetic Factors That Regulate Herpesviral Gene Expression

    PubMed Central

    Oh, Hyung Suk; Bryant, Kevin F.; Nieland, Thomas J. F.; Mazumder, Aprotim; Bagul, Mukta; Bathe, Mark; Root, David E.; Knipe, David M.

    2014-01-01

    ABSTRACT Herpes simplex virus (HSV) utilizes and subverts host chromatin mechanisms to express its lytic gene products in mammalian cells. The host cell attempts to silence the incoming viral genome by epigenetic mechanisms, but the viral VP16 and ICP0 proteins promote active chromatin on the viral genome by recruiting other host epigenetic factors. However, the dependence on VP16 and ICP0 differs in different cell lines, implying cell type-dependent functional contributions of epigenetic factors for HSV gene expression. In this study, we performed a targeted RNA interference (RNAi) screen for cellular chromatin factors that are involved in regulation of herpes simplex virus (HSV) gene expression in U2OS osteosarcoma cells, a cell line that complements ICP0 mutant and VP16 mutant virus replication. In this screen, we found the same general classes of chromatin factors that regulate HSV gene expression in U2OS cells as in other cell types, including histone demethylases (HDMs), histone deacetylases (HDACs), histone acetyltransferases (HATs), and chromatin-remodeling factors, but the specific factors within these classes are different from those identified previously for other cell types. For example, KDM3A and KDM1A (LSD1) both demethylate mono- and dimethylated H3K9, but KDM3A emerged in our screen of U2OS cells. Further, small interfering RNA (siRNA) and inhibitor studies support the idea that KDM1A is more critical in HeLa cells, as observed previously, while KDM3A is more critical in U2OS cells. These results argue that different cellular chromatin factors are critical in different cell lines to carry out the positive and negative epigenetic effects exerted on the HSV genome. PMID:24496796

  11. Combined zebrafish-yeast chemical-genetic screens reveal gene-copper-nutrition interactions that modulate melanocyte pigmentation.

    PubMed

    Ishizaki, Hironori; Spitzer, Michaela; Wildenhain, Jan; Anastasaki, Corina; Zeng, Zhiqiang; Dolma, Sonam; Shaw, Michael; Madsen, Erik; Gitlin, Jonathan; Marais, Richard; Tyers, Mike; Patton, E Elizabeth

    2010-01-01

    Hypopigmentation is a feature of copper deficiency in humans, as caused by mutation of the copper (Cu(2+)) transporter ATP7A in Menkes disease, or an inability to absorb copper after gastric surgery. However, many causes of copper deficiency are unknown, and genetic polymorphisms might underlie sensitivity to suboptimal environmental copper conditions. Here, we combined phenotypic screens in zebrafish for compounds that affect copper metabolism with yeast chemical-genetic profiles to identify pathways that are sensitive to copper depletion. Yeast chemical-genetic interactions revealed that defects in intracellular trafficking pathways cause sensitivity to low-copper conditions; partial knockdown of the analogous Ap3s1 and Ap1s1 trafficking components in zebrafish sensitized developing melanocytes to hypopigmentation in low-copper environmental conditions. Because trafficking pathways are essential for copper loading into cuproproteins, our results suggest that hypomorphic alleles of trafficking components might underlie sensitivity to reduced-copper nutrient conditions. In addition, we used zebrafish-yeast screening to identify a novel target pathway in copper metabolism for the small-molecule MEK kinase inhibitor U0126. The zebrafish-yeast screening method combines the power of zebrafish as a disease model with facile genome-scale identification of chemical-genetic interactions in yeast to enable the discovery and dissection of complex multigenic interactions in disease-gene networks. PMID:20713646

  12. Novel skin phenotypes revealed by a genome-wide mouse reverse genetic screen

    PubMed Central

    Liakath-Ali, Kifayathullah; Vancollie, Valerie E.; Heath, Emma; Smedley, Damian P.; Estabel, Jeanne; Sunter, David; DiTommaso, Tia; White, Jacqueline K.; Ramirez-Solis, Ramiro; Smyth, Ian; Steel, Karen P.; Watt, Fiona M.

    2014-01-01

    Permanent stop-and-shop large-scale mouse mutant resources provide an excellent platform to decipher tissue phenogenomics. Here we analyse skin from 538 knockout mouse mutants generated by the Sanger Institute Mouse Genetics Project. We optimize immunolabelling of tail epidermal wholemounts to allow systematic annotation of hair follicle, sebaceous gland and interfollicular epidermal abnormalities using ontology terms from the Mammalian Phenotype Ontology. Of the 50 mutants with an epidermal phenotype, 9 map to human genetic conditions with skin abnormalities. Some mutant genes are expressed in the skin, whereas others are not, indicating systemic effects. One phenotype is affected by diet and several are incompletely penetrant. In-depth analysis of three mutants, Krt76, Myo5a (a model of human Griscelli syndrome) and Mysm1, provides validation of the screen. Our study is the first large-scale genome-wide tissue phenotype screen from the International Knockout Mouse Consortium and provides an open access resource for the scientific community. PMID:24721909

  13. A Sleeping Beauty screen reveals NF-kB activation in CLL mouse model

    PubMed Central

    Zanesi, Nicola; Balatti, Veronica; Riordan, Jesse; Burch, Aaron; Rizzotto, Lara; Palamarchuk, Alexey; Cascione, Luciano; Lagana, Alessandro; Dupuy, Adam J.; Croce, Carlo M.

    2013-01-01

    TCL1 oncogene is overexpressed in aggressive form of human chronic lymphocytic leukemia (CLL) and its dysregulation in mouse B cells causes a CD5-positive leukemia similar to the aggressive form of human CLLs. To identify oncogenes that cooperate with Tcl1, we performed genetic screen in E??TCL1 mice using Sleeping Beauty transposon-mediated mutagenesis. Analysis of transposon common insertion sites identified 7 genes activated by transposon insertions. Overexpression of these genes in mouse CLL was confirmed by real time reverse transcription-polymerase chain reaction. Interestingly, the main known function of 4 of 7 genes (Nfkb1, Tab2, Map3K14, and Nfkbid) is participation in or activation of the nuclear factor-kB (NF-kB) pathway. In addition, activation of the NF-kB is 1 of main functions of Akt2, also identified in the screen. These findings demonstrate cooperation of Tcl1 and the NF-kB pathway in the pathogenesis of aggressive CLL. Identification cooperating cancer genes will result in the development of combinatorial therapies to treat CLL. PMID:23591791

  14. A Cell-Based Screen Reveals that the Albendazole Metabolite, Albendazole Sulfone, Targets Wolbachia

    PubMed Central

    Bray, Walter M.; White, Pamela M.; Ruybal, Jordan; Lokey, R. Scott; Debec, Alain; Sullivan, William

    2012-01-01

    Wolbachia endosymbionts carried by filarial nematodes give rise to the neglected diseases African river blindness and lymphatic filariasis afflicting millions worldwide. Here we identify new Wolbachia-disrupting compounds by conducting high-throughput cell-based chemical screens using a Wolbachia-infected, fluorescently labeled Drosophila cell line. This screen yielded several Wolbachia-disrupting compounds including three that resembled Albendazole, a widely used anthelmintic drug that targets nematode microtubules. Follow-up studies demonstrate that a common Albendazole metabolite, Albendazole sulfone, reduces intracellular Wolbachia titer both in Drosophila melanogaster and Brugia malayi, the nematode responsible for lymphatic filariasis. Significantly, Albendazole sulfone does not disrupt Drosophila microtubule organization, suggesting that this compound reduces titer through direct targeting of Wolbachia. Accordingly, both DNA staining and FtsZ immunofluorescence demonstrates that Albendazole sulfone treatment induces Wolbachia elongation, a phenotype indicative of binary fission defects. This suggests that the efficacy of Albendazole in treating filarial nematode-based diseases is attributable to dual targeting of nematode microtubules and their Wolbachia endosymbionts. PMID:23028321

  15. Mass spectrometry screening reveals widespread diversity in trichome specialized metabolites of tomato chromosomal substitution lines

    PubMed Central

    Schilmiller, Anthony; Shi, Feng; Kim, Jeongwoon; Charbonneau, Amanda L; Holmes, Daniel; Daniel Jones, A; Last, Robert L

    2010-01-01

    Glandular secreting trichomes of cultivated tomato (Solanum lycopersicum) and close relatives produce a variety of structurally diverse volatile and non-volatile specialized (‘secondary’) metabolites, including terpenes, flavonoids and acyl sugars. A genetic screen is described here to profile leaf trichome and surface metabolite extracts of nearly isogenic chromosomal substitution lines covering the tomato genome. These lines contain specific regions of the Solanum pennellii LA0716 genome in an otherwise ‘wild-type’ M82 tomato genetic background. Regions that have an impact on the total amount of extractable mono- and sesquiterpenes (IL2-2) or only sesquiterpenes (IL10-3) or specifically influence accumulation of the monoterpene ?-thujene (IL1-3 and IL1-4) were identified using GC-MS. A rapid LC-TOF-MS method was developed and used to identify changes in non-volatile metabolites through non-targeted analysis. Metabolite profiles generated using this approach led to the discovery of introgression lines producing different acyl chain substitutions on acyl sugar metabolites (IL1-3/1-4 and IL8-1/8-1-1), as well as two regions that influence the quantity of acyl sugars (IL5-3 and IL11-3). Chromosomal region 1-1/1-1-3 was found to influence the types of glycoalkaloids that are detected in leaf surface extracts. These results show that direct chemical screening is a powerful way to characterize genetic diversity in trichome specialized metabolism. PMID:20113441

  16. Genetic Screens for Enhancers of brahma Reveal Functional Interactions Between the BRM Chromatin-Remodeling Complex and the Delta-Notch Signal Transduction Pathway in Drosophila

    PubMed Central

    Armstrong, Jennifer A.; Sperling, Adam S.; Deuring, Renate; Manning, Laurina; Moseley, Sarah L.; Papoulas, Ophelia; Piatek, Caroline I.; Doe, Chris Q.; Tamkun, John W.

    2005-01-01

    The Drosophila trithorax group gene brahma (brm) encodes the ATPase subunit of a 2-MDa chromatin-remodeling complex. brm was identified in a screen for transcriptional activators of homeotic genes and subsequently shown to play a global role in transcription by RNA polymerase II. To gain insight into the targeting, function, and regulation of the BRM complex, we screened for mutations that genetically interact with a dominant-negative allele of brm (brmK804R). We first screened for dominant mutations that are lethal in combination with a brmK804R transgene under control of the brm promoter. In a distinct but related screen, we identified dominant mutations that modify eye defects resulting from expression of brmK804R in the eye-antennal imaginal disc. Mutations in three classes of genes were identified in our screens: genes encoding subunits of the BRM complex (brm, moira, and osa), other proteins directly involved in transcription (zerknullt and RpII140), and signaling molecules (Delta and vein). Expression of brmK804R in the adult sense organ precursor lineage causes phenotypes similar to those resulting from impaired Delta-Notch signaling. Our results suggest that signaling pathways may regulate the transcription of target genes by regulating the activity of the BRM complex. PMID:15944353

  17. Genetic screens reveal RAS/MAPK/MSK1 modulate ataxin 1 protein levels and toxicity in SCA1

    PubMed Central

    Tan, Qiumin; Mollema, Nissa; Diaz-Garcia, Javier R.; Gallego-Flores, Tatiana; Lu, Hsiang-Chih; Lagalwar, Sarita; Duvick, Lisa; Kang, Hyojin; Lee, Yoontae; Jafar-Nejad, Paymaan; Sayegh, Layal S.; Richman, Ronald; Liu, Xiuyun; Gao, Yan; Shaw, Chad A.; Arthur, J. Simon C.; Orr, Harry T.; Westbrook, Thomas F.; Botas, Juan; Zoghbi, Huda Y.

    2014-01-01

    Many neurodegenerative disorders such as Alzheimer’s, Parkinson’s and polyglutamine diseases share a common pathogenic mechanism: the abnormal accumulation of disease-causing proteins, due to either the mutant protein’s resistance to degradation or overexpression of the wild-type protein. We developed a strategy to identify therapeutic entry points for such neurodegenerative disorders by screening for genetic networks that influence the levels of disease-driving proteins. We applied this approach, which integrates parallel cell-based and Drosophila genetic screens, to spinocerebellar ataxia type 1 (SCA1), a disease caused by expansion of a polyglutamine tract in ataxin 1 (ATXN1). Our approach revealed that downregulation of several components of the RAS–MAPK–MSK1 pathway decreases ATXN1 levels and suppresses neurodegeneration in Drosophila and mice. Importantly, pharmacologic inhibitors of components of this pathway also decrease ATXN1 levels, suggesting that these components represent new therapeutic targets in mitigating SCA1. Collectively, these data reveal new therapeutic entry points for SCA1 and provide a proof-of-principle for tackling other classes of intractable neurodegenerative diseases. PMID:23719381

  18. A functional genomic screen reveals novel host genes that mediate interferon-alpha's effects against hepatitis C virus

    PubMed Central

    Zhao, Hong; Lin, Wenyu; Kumthip, Kattareeya; Cheng, Du; Fusco, Dahlene N; Hofmann, Oliver; Jilg, Nikolaus; Tai, Andrew W.; Goto, Kaku; Zhang, Leiliang; Hide, Winston; Jang, Jae Young; Peng, Lee F; Chung, Raymond T

    2011-01-01

    Background & Aims The precise mechanisms by which IFN exerts its antiviral effect against HCV have not yet been elucidated. We sought to identify host genes that mediate the antiviral effect of IFN-? by conducting a whole-genome siRNA library screen. Methods High throughput screening was performed using an HCV genotype 1b replicon, pRep-Feo. Those pools with replicate robust Z scores ? 2.0 entered secondary validation in full-length OR6 replicon cells. Huh7.5.1 cells infected with JFH1 were then used to validate the rescue efficacy of selected genes for HCV replication under IFN-? treatment. Results We identified and confirmed 93 human genes involved in the IFN-? anti-HCV effect using a whole-genome siRNA library. Gene ontology analysis revealed that mRNA processing (23 genes, P=2.756e-22), translation initiation (9 genes, P=2.42e-6), and IFN signaling (5 genes, P=1.00e-3) were the most enriched functional groups. Nine genes were components of U4/U6.U5 tri-snRNP. We confirmed that silencing squamous cell carcinoma antigen recognized by T cells (SART1), a specific factor of tri-snRNP, abrogates IFN-?'s suppressive effects against HCV in both replicon cells and JFH1 infectious cells. We further found that SART1 was not an IFN-? inducible, and its anti-HCV effector in the JFH1 infectious model was through regulation of interferon stimulated genes (ISGs) with or without IFN-?. Conclusions We identified 93 genes that mediate the anti-HCV effect of IFN-? through genome-wide siRNA screening; 23 and 9 genes were involved in mRNA processing and translation initiation, respectively. These findings reveal an unexpected role for mRNA processing in generation of the antiviral state, and suggest a new avenue for therapeutic development in HCV. PMID:21888876

  19. Data Mining of NCI’s Anticancer Screening Database Reveals Mitochondrial Complex I Inhibitors Cytotoxic to Leukemia Cell Lines

    PubMed Central

    Glover, Constance J.; Rabow, Alfred A.; Isgor, Yasemin G.; Shoemaker, Robert H.; Covell, David G.

    2007-01-01

    Mitochondria are principal mediators of apoptosis and thus can be considered molecular targets for new chemotherapeutic agents in the treatment of cancer. Inhibitors of mitochondrial complex I of the electron transport chain have been shown to induce apoptosis and exhibit antitumor activity. In an effort to find novel complex I inhibitors which exhibited anti-cancer activity in the NCI’s tumor cell line screen, we examined organized tumor cytotoxicity screening data available as SOM (self-organized maps) (http://spheroid.ncifcrf.gov) at the Developmental Therapeutics Program (DTP) of the National Cancer Institute (NCI). Our analysis focused on an SOM cluster comprised of compounds which included a number of known mitochondrial complex I (NADH:CoQ oxidoreductase) inhibitors. From these clusters ten compounds whose mechanism of action was unknown were tested for inhibition of complex I activity in bovine heart submitochondrial particles (SMP) resulting in the discovery that five of the ten compounds demonstrated significant inhibition with IC50's in the nM range for three of the five. Examination of screening profiles of the five inhibitors toward the NCI’s tumor cell lines revealed that they were cytotoxic to the leukemia subpanel (particularly K562 cells). Oxygen consumption experiments with permeabilized K562 cells revealed that the five most active compounds inhibited complex I activity in these cells in the same rank order and similar potency as determined with bovine heart SMP. Our findings thus fortify the appeal of mitochondrial Complex I as a possible anti-cancer molecular target and provide a data mining strategy for selecting candidate inhibitors for further testing. PMID:17109823

  20. A directed mutagenesis screen in Drosophila melanogaster reveals new mutants that influence hedgehog signaling.

    PubMed Central

    Haines, N; van den Heuvel, M

    2000-01-01

    The Hedgehog signaling pathway has been recognized as essential for patterning processes in development of metazoan animal species. The signaling pathway is, however, not entirely understood. To start to address this problem, we set out to isolate new mutations that influence Hedgehog signaling. We performed a mutagenesis screen for mutations that dominantly suppress Hedgehog overexpression phenotypes in the Drosophila melanogaster wing. We isolated four mutations that influence Hedgehog signaling. These were analyzed in the amenable wing system using genetic and molecular techniques. One of these four mutations affects the stability of the Hedgehog expression domain boundary, also known as the organizer in the developing wing. Another mutation affects a possible Hedgehog autoregulation mechanism, which stabilizes the same boundary. PMID:11102373

  1. Development of Synthetic Lethality Anticancer Therapeutics

    PubMed Central

    2015-01-01

    The concept of synthetic lethality (the creation of a lethal phenotype from the combined effects of mutations in two or more genes) has recently been exploited in various efforts to develop new genotype-selective anticancer therapeutics. These efforts include screening for novel anticancer agents, identifying novel therapeutic targets, characterizing mechanisms of resistance to targeted therapy, and improving efficacies through the rational design of combination therapy. This review discusses recent developments in synthetic lethality anticancer therapeutics, including poly ADP-ribose polymerase inhibitors for BRCA1- and BRCA2-mutant cancers, checkpoint inhibitors for p53 mutant cancers, and small molecule agents targeting RAS gene mutant cancers. Because cancers are caused by mutations in multiple genes and abnormalities in multiple signaling pathways, synthetic lethality for a specific tumor suppressor gene or oncogene is likely cell context-dependent. Delineation of the mechanisms underlying synthetic lethality and identification of treatment response biomarkers will be critical for the success of synthetic lethality anticancer therapy. PMID:24893124

  2. Genetic signature of histiocytic sarcoma revealed by a sleeping beauty transposon genetic screen in mice.

    PubMed

    Been, Raha A; Linden, Michael A; Hager, Courtney J; DeCoursin, Krista J; Abrahante, Juan E; Landman, Sean R; Steinbach, Michael; Sarver, Aaron L; Largaespada, David A; Starr, Timothy K

    2014-01-01

    Histiocytic sarcoma is a rare, aggressive neoplasm that responds poorly to therapy. Histiocytic sarcoma is thought to arise from macrophage precursor cells via genetic changes that are largely undefined. To improve our understanding of the etiology of histiocytic sarcoma we conducted a forward genetic screen in mice using the Sleeping Beauty transposon as a mutagen to identify genetic drivers of histiocytic sarcoma. Sleeping Beauty mutagenesis was targeted to myeloid lineage cells using the Lysozyme2 promoter. Mice with activated Sleeping Beauty mutagenesis had significantly shortened lifespan and the majority of these mice developed tumors resembling human histiocytic sarcoma. Analysis of transposon insertions identified 27 common insertion sites containing 28 candidate cancer genes. Several of these genes are known drivers of hematological neoplasms, like Raf1, Fli1, and Mitf, while others are well-known cancer genes, including Nf1, Myc, Jak2, and Pten. Importantly, several new potential drivers of histiocytic sarcoma were identified and could serve as targets for therapy for histiocytic sarcoma patients. PMID:24827933

  3. Haploid Genetic Screen Reveals a Profound and Direct Dependence on Cholesterol for Hantavirus Membrane Fusion

    PubMed Central

    Kleinfelter, Lara M.; Jangra, Rohit K.; Jae, Lucas T.; Herbert, Andrew S.; Mittler, Eva; Stiles, Katie M.; Wirchnianski, Ariel S.; Kielian, Margaret; Brummelkamp, Thijn R.

    2015-01-01

    ABSTRACT Hantaviruses cause hemorrhagic fever with renal syndrome (HFRS) in the Old World and a highly fatal hantavirus cardiopulmonary syndrome (HCPS) in the New World. No vaccines or antiviral therapies are currently available to prevent or treat hantavirus disease, and gaps in our understanding of how hantaviruses enter cells challenge the search for therapeutics. We performed a haploid genetic screen in human cells to identify host factors required for entry by Andes virus, a highly virulent New World hantavirus. We found that multiple genes involved in cholesterol sensing, regulation, and biosynthesis, including key components of the sterol response element-binding protein (SREBP) pathway, are critical for Andes virus entry. Genetic or pharmacological disruption of the membrane-bound transcription factor peptidase/site-1 protease (MBTPS1/S1P), an SREBP control element, dramatically reduced infection by virulent hantaviruses of both the Old World and New World clades but not by rhabdoviruses or alphaviruses, indicating that this pathway is broadly, but selectively, required by hantaviruses. These results could be fully explained as arising from the modest depletion of cellular membrane cholesterol that accompanied S1P disruption. Mechanistic studies of cells and with protein-free liposomes suggested that high levels of cholesterol are specifically needed for hantavirus membrane fusion. Taken together, our results indicate that the profound dependence on target membrane cholesterol is a fundamental, and unusual, biophysical property of hantavirus glycoprotein-membrane interactions during entry. PMID:26126854

  4. Screen for mitochondrial DNA copy number maintenance genes reveals essential role for ATP synthase.

    PubMed

    Fukuoh, Atsushi; Cannino, Giuseppe; Gerards, Mike; Buckley, Suzanne; Kazancioglu, Selena; Scialo, Filippo; Lihavainen, Eero; Ribeiro, Andre; Dufour, Eric; Jacobs, Howard T

    2014-01-01

    The machinery of mitochondrial DNA (mtDNA) maintenance is only partially characterized and is of wide interest due to its involvement in disease. To identify novel components of this machinery, plus other cellular pathways required for mtDNA viability, we implemented a genome-wide RNAi screen in Drosophila S2 cells, assaying for loss of fluorescence of mtDNA nucleoids stained with the DNA-intercalating agent PicoGreen. In addition to previously characterized components of the mtDNA replication and transcription machineries, positives included many proteins of the cytosolic proteasome and ribosome (but not the mitoribosome), three proteins involved in vesicle transport, some other factors involved in mitochondrial biogenesis or nuclear gene expression, > 30 mainly uncharacterized proteins and most subunits of ATP synthase (but no other OXPHOS complex). ATP synthase knockdown precipitated a burst of mitochondrial ROS production, followed by copy number depletion involving increased mitochondrial turnover, not dependent on the canonical autophagy machinery. Our findings will inform future studies of the apparatus and regulation of mtDNA maintenance, and the role of mitochondrial bioenergetics and signaling in modulating mtDNA copy number. PMID:24952591

  5. Genetic Signature of Histiocytic Sarcoma Revealed by a Sleeping Beauty Transposon Genetic Screen in Mice

    PubMed Central

    Been, Raha A.; Linden, Michael A.; Hager, Courtney J.; DeCoursin, Krista J.; Abrahante, Juan E.; Landman, Sean R.; Steinbach, Michael; Sarver, Aaron L.; Largaespada, David A.; Starr, Timothy K.

    2014-01-01

    Histiocytic sarcoma is a rare, aggressive neoplasm that responds poorly to therapy. Histiocytic sarcoma is thought to arise from macrophage precursor cells via genetic changes that are largely undefined. To improve our understanding of the etiology of histiocytic sarcoma we conducted a forward genetic screen in mice using the Sleeping Beauty transposon as a mutagen to identify genetic drivers of histiocytic sarcoma. Sleeping Beauty mutagenesis was targeted to myeloid lineage cells using the Lysozyme2 promoter. Mice with activated Sleeping Beauty mutagenesis had significantly shortened lifespan and the majority of these mice developed tumors resembling human histiocytic sarcoma. Analysis of transposon insertions identified 27 common insertion sites containing 28 candidate cancer genes. Several of these genes are known drivers of hematological neoplasms, like Raf1, Fli1, and Mitf, while others are well-known cancer genes, including Nf1, Myc, Jak2, and Pten. Importantly, several new potential drivers of histiocytic sarcoma were identified and could serve as targets for therapy for histiocytic sarcoma patients. PMID:24827933

  6. A genome-wide screen in Saccharomyces cerevisiae Reveals Pathways affected By Arsenic Toxicity

    PubMed Central

    Zhou, Xue; Arita, Adriana; Ellen, Thomas P.; Liu, Xin; Bai, Jingxiang; Rooney, John P.; Kurtz, Adrienne D.; Klein, Catherine B.; Dai, Wei; Begley, Thomas J.; Costa, Max

    2009-01-01

    We have used Saccharomyces cerevisiae to identify toxicologically important proteins and pathways involved in arsenic-induced toxicity and carcinogenicity in humans. We performed a systemic screen of the complete set of 4,733 haploid S. cerevisiae single gene deletion mutants to identify those that have decreased or increased growth, relative to wild-type, after exposure to sodium arsenite (NaAsO2). IC50 values for all mutants were determined to further validate our results. Ultimately we identified 248 mutants sensitive to arsenite and 5 mutants resistant to arsenite exposure. We analyzed the proteins corresponding to arsenite-sensitive mutants and determined that they belonged to functional categories that include protein binding, phosphate metabolism, vacuolar/lysosomal transport, protein targeting, sorting, and translocation, cell growth/morphogenesis, cell polarity and filament formation. Furthermore, these data were mapped onto a protein interactome to identify arsenite toxicity-modulating networks. These networks are associated with the cytoskeleton, ubiquitination, histone acetylation and the MAPK signaling pathway. Our studies have potential implications for understanding toxicity and carcinogenesis in arsenic-induced human conditions, such as cancer and aging. PMID:19631266

  7. Chemoecological Screening Reveals High Bioactivity in Diverse Culturable Portuguese Marine Cyanobacteria

    PubMed Central

    Leão, Pedro N.; Ramos, Vitor; Gonçalves, Patrício B.; Viana, Flávia; Lage, Olga M.; Gerwick, William H.; Vasconcelos, Vitor M.

    2013-01-01

    Marine cyanobacteria, notably those from tropical regions, are a rich source of bioactive secondary metabolites. Tropical marine cyanobacteria often grow to high densities in the environment, allowing direct isolation of many secondary metabolites from field-collected material. However, in temperate environments culturing is usually required to produce enough biomass for investigations of their chemical constituents. In this work, we cultured a selection of novel and diverse cyanobacteria isolated from the Portuguese coast, and tested their organic extracts in a series of ecologically-relevant bioassays. The majority of the extracts showed activity in at least one of the bioassays, all of which were run in very small scale. Phylogenetically related isolates exhibited different activity profiles, highlighting the value of microdiversity for bioprospection studies. Furthermore, LC-MS analyses of selected active extracts suggested the presence of previously unidentified secondary metabolites. Overall, the screening strategy employed here, in which previously untapped cyanobacterial diversity was combined with multiple bioassays, proved to be a successful strategy and allowed the selection of several strains for further investigations based on their bioactivity profiles. PMID:23609580

  8. Genome-wide screen reveals replication pathway for quasi-palindrome fragility dependent on homologous recombination.

    PubMed

    Zhang, Yu; Saini, Natalie; Sheng, Ziwei; Lobachev, Kirill S

    2013-01-01

    Inverted repeats capable of forming hairpin and cruciform structures present a threat to chromosomal integrity. They induce double strand breaks, which lead to gross chromosomal rearrangements, the hallmarks of cancers and hereditary diseases. Secondary structure formation at this motif has been proposed to be the driving force for the instability, albeit the mechanisms leading to the fragility are not well-understood. We carried out a genome-wide screen to uncover the genetic players that govern fragility of homologous and homeologous Alu quasi-palindromes in the yeast Saccharomyces cerevisiae. We found that depletion or lack of components of the DNA replication machinery, proteins involved in Fe-S cluster biogenesis, the replication-pausing checkpoint pathway, the telomere maintenance complex or the Sgs1-Top3-Rmi1 dissolvasome augment fragility at Alu-IRs. Rad51, a component of the homologous recombination pathway, was found to be required for replication arrest and breakage at the repeats specifically in replication-deficient strains. These data demonstrate that Rad51 is required for the formation of breakage-prone secondary structures in situations when replication is compromised while another mechanism operates in DSB formation in replication-proficient strains. PMID:24339793

  9. Screen for mitochondrial DNA copy number maintenance genes reveals essential role for ATP synthase

    PubMed Central

    Fukuoh, Atsushi; Cannino, Giuseppe; Gerards, Mike; Buckley, Suzanne; Kazancioglu, Selena; Scialo, Filippo; Lihavainen, Eero; Ribeiro, Andre; Dufour, Eric; Jacobs, Howard T

    2014-01-01

    The machinery of mitochondrial DNA (mtDNA) maintenance is only partially characterized and is of wide interest due to its involvement in disease. To identify novel components of this machinery, plus other cellular pathways required for mtDNA viability, we implemented a genome-wide RNAi screen in Drosophila S2 cells, assaying for loss of fluorescence of mtDNA nucleoids stained with the DNA-intercalating agent PicoGreen. In addition to previously characterized components of the mtDNA replication and transcription machineries, positives included many proteins of the cytosolic proteasome and ribosome (but not the mitoribosome), three proteins involved in vesicle transport, some other factors involved in mitochondrial biogenesis or nuclear gene expression, > 30 mainly uncharacterized proteins and most subunits of ATP synthase (but no other OXPHOS complex). ATP synthase knockdown precipitated a burst of mitochondrial ROS production, followed by copy number depletion involving increased mitochondrial turnover, not dependent on the canonical autophagy machinery. Our findings will inform future studies of the apparatus and regulation of mtDNA maintenance, and the role of mitochondrial bioenergetics and signaling in modulating mtDNA copy number. PMID:24952591

  10. A modifier screen in the eye reveals control genes for Krüppel activity in the Drosophila embryo

    PubMed Central

    Carrera, Pilar; Abrell, Sarah; Kerber, Birgit; Walldorf, Uwe; Preiss, Anette; Hoch, Michael; Jäckle, Herbert

    1998-01-01

    Irregular facets (If) is a dominant mutation of Drosophila that results in small eyes with fused ommatidia. Previous results showed that the gene Krüppel (Kr), which is best known for its early segmentation function, is expressed ectopically in If mutant eye discs. However, it was not known whether ectopic Kr activity is either the cause or the result of the If mutation. Here, we show that If is a gain-of-function allele of Kr. We then used the If mutation in a genetic screen to identify dominant enhancers and suppressors of Kr activity on the third chromosome. Of 30 identified Kr-interacting loci, two were cloned, and we examined whether they also represent components of a natural Kr-dependent developmental pathway of the embryo. We show that the two genes, eyelid (eld) and extramacrochaetae (emc), which encode a Bright family-type DNA binding protein and a helix-loop-helix factor, respectively, are necessary to achieve the singling-out of a unique Kr-expressing cell during the development of the Malpighian tubules, the excretory organs of the fly. The results indicate that the Kr gain-of-function mutation If provides a tool to identify genes that are active during eye development and that a number of them function also in the control of Kr-dependent developmental processes. PMID:9724781

  11. Screening of Escherichia coli Species Biodiversity Reveals New Biofilm-Associated Antiadhesion Polysaccharides

    PubMed Central

    Rendueles, Olaya; Travier, Laetitia; Latour-Lambert, Patricia; Fontaine, Thierry; Magnus, Julie; Denamur, Erick; Ghigo, Jean-Marc

    2011-01-01

    ABSTRACT Bacterial biofilms often form multispecies communities in which complex but ill-understood competition and cooperation interactions occur. In light of the profound physiological modifications associated with this lifestyle, we hypothesized that the biofilm environment might represent an untapped source of natural bioactive molecules interfering with bacterial adhesion or biofilm formation. We produced cell-free solutions extracted from in vitro mature biofilms formed by 122 natural Escherichia coli isolates, and we screened these biofilm extracts for antiadhesion molecules active on a panel of Gram-positive and Gram-negative bacteria. Using this approach, we showed that 20% of the tested biofilm extracts contained molecules that antagonize bacterial growth or adhesion. We characterized a compound, produced by a commensal animal E. coli strain, for which activity is detected only in biofilm extract. Biochemical and genetic analyses showed that this compound corresponds to a new type of released high-molecular-weight polysaccharide whose biofilm-associated production is regulated by the RfaH protein. We demonstrated that the antiadhesion activity of this polysaccharide was restricted to Gram-positive bacteria and that its production reduced susceptibility to invasion and provided rapid exclusion of Staphylococcus aureus from mixed E. coli and S. aureus biofilms. Our results therefore demonstrate that biofilms contain molecules that contribute to the dynamics of mixed bacterial communities and that are not or only poorly detected in unconcentrated planktonic supernatants. Systematic identification of these compounds could lead to strategies that limit pathogen surface colonization and reduce the burden associated with the development of bacterial biofilms on medical devices. PMID:21558434

  12. A Comprehensive Immunoreceptor Phosphotyrosine-based Signaling Network Revealed by Reciprocal Protein-Peptide Array Screening.

    PubMed

    Liu, Huadong; Li, Lei; Voss, Courtney; Wang, Feng; Liu, Juewen; Li, Shawn Shun-Cheng

    2015-07-01

    Cells of the immune system communicate with their environment through immunoreceptors. These receptors often harbor intracellular tyrosine residues, which, when phosphorylated upon receptor activation, serve as docking sites to recruit downstream signaling proteins containing the Src Homology 2 (SH2) domain. A systematic investigation of interactions between the SH2 domain and the immunoreceptor tyrosine-based regulatory motifs (ITRM), including inhibitory (ITIM), activating (ITAM), or switching (ITSM) motifs, is critical for understanding cellular signal transduction and immune function. Using the B cell inhibitory receptor CD22 as an example, we developed an approach that combines reciprocal or bidirectional phosphopeptide and SH2 domain array screens with in-solution binding assays to identify a comprehensive SH2-CD22 interaction network. Extending this approach to 194 human ITRM sequences and 78 SH2 domains led to the identification of a high-confidence immunoreceptor interactome containing 1137 binary interactions. Besides recapitulating many previously reported interactions, our study uncovered numerous novel interactions. The resulting ITRM-SH2 interactome not only helped to fill many gaps in the immune signaling network, it also allowed us to associate different SH2 domains to distinct immune functions. Detailed analysis of the NK cell ITRM-mediated interactions led to the identification of a network nucleated by the Vav3 and Fyn SH2 domains. We showed further that these SH2 domains have distinct functions in cytotoxicity. The bidirectional protein-peptide array approach described herein may be applied to the numerous other peptide-binding modules to identify potential protein-protein interactions in a systematic and reliable manner. PMID:25907764

  13. A transcriptome-wide RNAi screen in the Drosophila ovary reveals novel factors of the germline piRNA pathway

    PubMed Central

    Czech, Benjamin; Preall, Jonathan B.; McGinn, Jon; Hannon, Gregory J.

    2013-01-01

    Summary The Drosophila piRNA pathway provides an RNA-based immune system that defends the germline genome against selfish genetic elements. Two inter-related branches of the piRNA system exist: somatic cells that support oogenesis only employ Piwi, whereas germ cells utilize a more elaborated pathway centered on the three gonad-specific Argonaute proteins Piwi, Aubergine, and Argonaute3. While several key factors of each branch have been identified, our current knowledge is insufficient to explain the complex workings of the piRNA machinery. Here, we report a reverse genetic screen spanning the ovarian transcriptome in an attempt to uncover the full repertoire of genes required for piRNA-mediated transposon silencing in the female germline. Our screen reveals new key factors of piRNA-mediated transposon silencing, including the novel piRNA biogenesis factors, CG2183 (GASZ) and Deadlock. Last, our data uncovers a previously unanticipated set of factors preferentially required for repression of different transposons types. PMID:23665227

  14. Quantitative Genome-Wide Genetic Interaction Screens Reveal Global Epistatic Relationships of Protein Complexes in Escherichia coli

    PubMed Central

    Kumar, Ashwani; Stewart, Geordie; Samanfar, Bahram; Aoki, Hiroyuki; Wagih, Omar; Vlasblom, James; Phanse, Sadhna; Lad, Krunal; Yeou Hsiung Yu, Angela; Graham, Christopher; Jin, Ke; Brown, Eric; Golshani, Ashkan; Kim, Philip; Moreno-Hagelsieb, Gabriel; Greenblatt, Jack; Houry, Walid A.; Parkinson, John; Emili, Andrew

    2014-01-01

    Large-scale proteomic analyses in Escherichia coli have documented the composition and physical relationships of multiprotein complexes, but not their functional organization into biological pathways and processes. Conversely, genetic interaction (GI) screens can provide insights into the biological role(s) of individual gene and higher order associations. Combining the information from both approaches should elucidate how complexes and pathways intersect functionally at a systems level. However, such integrative analysis has been hindered due to the lack of relevant GI data. Here we present a systematic, unbiased, and quantitative synthetic genetic array screen in E. coli describing the genetic dependencies and functional cross-talk among over 600,000 digenic mutant combinations. Combining this epistasis information with putative functional modules derived from previous proteomic data and genomic context-based methods revealed unexpected associations, including new components required for the biogenesis of iron-sulphur and ribosome integrity, and the interplay between molecular chaperones and proteases. We find that functionally-linked genes co-conserved among ?-proteobacteria are far more likely to have correlated GI profiles than genes with divergent patterns of evolution. Overall, examining bacterial GIs in the context of protein complexes provides avenues for a deeper mechanistic understanding of core microbial systems. PMID:24586182

  15. Screening of odor-receptor pairs in Caenorhabditis elegans reveals different receptors for high and low odor concentrations.

    PubMed

    Taniguchi, Gun; Uozumi, Takayuki; Kiriyama, Keisuke; Kamizaki, Tomoko; Hirotsu, Takaaki

    2014-04-29

    Olfactory systems sense and respond to various odorants. Olfactory receptors, which in most organisms are G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptors, directly bind volatile or soluble odorants. Compared to the genomes of mammals, the genome of the nematode Caenorhabditis elegans contains more putative olfactory receptor genes, suggesting that in nematodes there may be combinatorial complexity to the receptor-odor relationship. We used RNA interference (RNAi) screening to identify nematode olfactory receptors necessary for the response to specific odorants. This screening identified 194 candidate olfactory receptor genes linked to 11 odorants. Additionally, we identified SRI-14 as being involved in sensing high concentrations of diacetyl. Rescue and neuron-specific RNAi experiments demonstrated that SRI-14 functioned in ASH neurons, specific chemosensory neurons, resulting in avoidance responses. Calcium imaging revealed that ASH neurons responded to high diacetyl concentrations only, whereas another class of chemosensory neurons, AWA neurons, reacted to both low and high concentrations. Loss of SRI-14 function hampered ASH responses to high diacetyl concentrations, whereas loss of ODR-10 function reduced AWA responses to low odorant concentrations. Chemosensory neurons ectopically expressing SRI-14 responded to a high concentration of diacetyl. Thus, nematodes have concentration-dependent odor-sensing mechanisms that are segregated at the olfactory receptor and sensory neuron levels. PMID:24782565

  16. Functional Screening of a Metagenomic Library Reveals Operons Responsible for Enhanced Intestinal Colonization by Gut Commensal Microbes

    PubMed Central

    Yoon, Mi Young; Lee, Kang-Mu; Yoon, Yujin; Go, Junhyeok; Park, Yongjin; Cho, Yong-Joon; Tannock, Gerald W.

    2013-01-01

    Evidence suggests that gut microbes colonize the mammalian intestine through propagation as an adhesive microbial community. A bacterial artificial chromosome (BAC) library of murine bowel microbiota DNA in the surrogate host Escherichia coli DH10B was screened for enhanced adherence capability. Two out of 5,472 DH10B clones, 10G6 and 25G1, exhibited enhanced capabilities to adhere to inanimate surfaces in functional screens. DNA segments inserted into the 10G6 and 25G1 clones were 52 and 41 kb and included 47 and 41 protein-coding open reading frames (ORFs), respectively. DNA sequence alignments, tetranucleotide frequency, and codon usage analysis strongly suggest that these two DNA fragments are derived from species belonging to the genus Bacteroides. Consistent with this finding, a large portion of the predicted gene products were highly homologous to those of Bacteroides spp. Transposon mutagenesis and subsequent experiments that involved heterologous expression identified two operons associated with enhanced adherence. E. coli strains transformed with the 10a or 25b operon adhered to the surface of intestinal epithelium and colonized the mouse intestine more vigorously than did the control strain. This study has revealed the genetic determinants of unknown commensals (probably resembling Bacteroides species) that enhance the ability of the bacteria to colonize the murine bowel. PMID:23584783

  17. Identification of ?-hematin inhibitors in a high-throughput screening effort reveals scaffolds with in vitro antimalarial activity

    PubMed Central

    Sandlin, Rebecca D.; Fong, Kim Y.; Wicht, Kathryn J.; Carrell, Holly M.; Egan, Timothy J.; Wright, David W.

    2014-01-01

    The emergence of drug resistant strains of Plasmodium spp. creates a critical need for the development of novel antimalarials. Formation of hemozoin, a crystalline heme detoxification process vital to parasite survival serves as an important drug target. The quinoline antimalarials including chloroquine and amodiaquine owe their antimalarial activity to inhibition of hemozoin formation. Though in vivo formation of hemozoin occurs within the presence of neutral lipids, the lipophilic detergent NP-40 was previously shown to serve as a surrogate in the ?-hematin (synthetic hemozoin) formation process. Consequently, an NP-40 mediated ?-hematin formation assay was developed for use in high-throughput screening. Here, the assay was utilized to screen 144,330 compounds for the identification of inhibitors of crystallization, resulting in 530 hits. To establish the effectiveness of these target-based ?-hematin inhibitors against Plasmodiumfalciparum, each hit was further tested in cultures of parasitized red blood cells. This effort revealed that 171 of the ?-hematin inhibitors are also active against the parasite. Dose–response data identified 73 of these ?-hematin inhibitors have IC50 values ?5 ?M, including 25 compounds with nanomolar activity against P. falciparum. A scaffold-based analysis of this data identified 14 primary scaffolds that represent 46% of the 530 total hits. Representative compounds from each of the classes were further assessed for hemozoin inhibitory activity in P. falciparum infected human erythrocytes. Each of the hit compounds tested were found to be positive inhibitors, while a negative control did not perturb this biological pathway in culture. PMID:25516843

  18. A quantitative imaging-based screen reveals the exocyst as a network hub connecting endocytosis and exocytosis.

    PubMed

    Jose, Mini; Tollis, Sylvain; Nair, Deepak; Mitteau, Romain; Velours, Christophe; Massoni-Laporte, Aurelie; Royou, Anne; Sibarita, Jean-Baptiste; McCusker, Derek

    2015-07-01

    The coupling of endocytosis and exocytosis underlies fundamental biological processes ranging from fertilization to neuronal activity and cellular polarity. However, the mechanisms governing the spatial organization of endocytosis and exocytosis require clarification. Using a quantitative imaging-based screen in budding yeast, we identified 89 mutants displaying defects in the localization of either one or both pathways. High-resolution single-vesicle tracking revealed that the endocytic and exocytic mutants she4? and bud6? alter post-Golgi vesicle dynamics in opposite ways. The endocytic and exocytic pathways display strong interdependence during polarity establishment while being more independent during polarity maintenance. Systems analysis identified the exocyst complex as a key network hub, rich in genetic interactions with endocytic and exocytic components. Exocyst mutants displayed altered endocytic and post-Golgi vesicle dynamics and interspersed endocytic and exocytic domains compared with control cells. These data are consistent with an important role for the exocyst in coordinating endocytosis and exocytosis. PMID:25947137

  19. Screening for hydrolytic enzymes reveals Ayr1p as a novel triacylglycerol lipase in Saccharomyces cerevisiae.

    PubMed

    Ploier, Birgit; Scharwey, Melanie; Koch, Barbara; Schmidt, Claudia; Schatte, Jessica; Rechberger, Gerald; Kollroser, Manfred; Hermetter, Albin; Daum, Günther

    2013-12-13

    Saccharomyces cerevisiae, as well as other eukaryotes, preserves fatty acids and sterols in a biologically inert form, as triacylglycerols and steryl esters. The major triacylglycerol lipases of the yeast S. cerevisiae identified so far are Tgl3p, Tgl4p, and Tgl5p (Athenstaedt, K., and Daum, G. (2003) YMR313c/TGL3 encodes a novel triacylglycerol lipase located in lipid particles of Saccharomyces cerevisiae. J. Biol. Chem. 278, 23317-23323; Athenstaedt, K., and Daum, G. (2005) Tgl4p and Tgl5p, two triacylglycerol lipases of the yeast Saccharomyces cerevisiae, are localized to lipid particles. J. Biol. Chem. 280, 37301-37309). We observed that upon cultivation on oleic acid, triacylglycerol mobilization did not come to a halt in a yeast strain deficient in all currently known triacylglycerol lipases, indicating the presence of additional not yet characterized lipases/esterases. Functional proteome analysis using lipase and esterase inhibitors revealed a subset of candidate genes for yet unknown hydrolytic enzymes on peroxisomes and lipid droplets. Based on the conserved GXSXG lipase motif, putative functions, and subcellular localizations, a selected number of candidates were characterized by enzyme assays in vitro, gene expression analysis, non-polar lipid analysis, and in vivo triacylglycerol mobilization assays. These investigations led to the identification of Ayr1p as a novel triacylglycerol lipase of yeast lipid droplets and confirmed the hydrolytic potential of the peroxisomal Lpx1p in vivo. Based on these results, we discuss a possible link between lipid storage, lipid mobilization, and peroxisomal utilization of fatty acids as a carbon source. PMID:24187129

  20. A Functional Screen Reveals an Extensive Layer of Transcriptional and Splicing Control Underlying RAS/MAPK Signaling in Drosophila

    PubMed Central

    Ashton-Beaucage, Dariel; Udell, Christian M.; Gendron, Patrick; Sahmi, Malha; Lefrançois, Martin; Baril, Caroline; Guenier, Anne-Sophie; Duchaine, Jean; Lamarre, Daniel; Lemieux, Sébastien; Therrien, Marc

    2014-01-01

    The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway—including a new protein complex modulating RAF activation—we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing components can also specifically impact alternative splicing. PMID:24643257

  1. A chemically-defined screening platform reveals behavioral similarities between primary human mesenchymal stem cells and endothelial cells

    PubMed Central

    Koepsel, Justin T.; Loveland, Samuel G.; Schwartz, Michael P.; Zorn, Stefan; Belair, David G.; Le, Ngoc Nhi; Murphy, William L.

    2012-01-01

    Chemically defined substrates, which rigorously control protein-surface and cell-surface interactions, can be used to probe the effects of specific biomolecules on cell behavior. Here we combined a chemically-defined, array-based format with automated, time-lapse microscopy to efficiently screen cell-substrate interactions. Self-assembled monolayers (SAMs) of alkanethiolates bearing oligo(ethylene glycol) units and reactive terminal groups were used to present cell adhesion peptides while minimizing non-specific protein interactions. Specifically, we describe rapid fabrication of arrays of 1 mm spots, which present varied densities of the integrin-binding ligand Gly-Arg-Gly-Asp-Ser-Pro (GRGDSP). Results indicate that cell attachment, cell spreading, and proliferation exhibit strong dependencies on GRGDSP density for both human mesenchymal stem cells (hMSCs) and human umbilical vein endothelial cells (HUVECs). Furthermore, relative spreading and proliferation over a broad range of GRGDSP densities are similar for both primary cell types, and detailed comparison between cell behaviors identified a 1:1 correlation between spreading and proliferation for both HUVECs and hMSCs. Finally, time-lapse microscopy of SAM arrays revealed distinct adhesion-dependent migratory behaviors for HUVECs and hMSCs. These results demonstrate the benefits of using an array-based screening platform for investigating cell function. While the proof-of-concept focuses on simple cellular properties, the quantitative similarities observed for hMSCs and HUVECs provides a direct example of how phenomena that would not easily be predicted can be shown to correlate between different cell types. PMID:23147838

  2. Functional drug screening reveals anticonvulsants as enhancers of mTOR-independent autophagic killing of Mycobacterium tuberculosis through inositol depletion

    PubMed Central

    Schiebler, Mark; Brown, Karen; Hegyi, Krisztina; Newton, Sandra M; Renna, Maurizio; Hepburn, Lucy; Klapholz, Catherine; Coulter, Sarah; Obregón-Henao, Andres; Henao Tamayo, Marcela; Basaraba, Randall; Kampmann, Beate; Henry, Katherine M; Burgon, Joseph; Renshaw, Stephen A; Fleming, Angeleen; Kay, Robert R; Anderson, Karen E; Hawkins, Phillip T; Ordway, Diane J; Rubinsztein, David C; Floto, Rodrigo Andres

    2015-01-01

    Mycobacterium tuberculosis (MTB) remains a major challenge to global health made worse by the spread of multidrug resistance. We therefore examined whether stimulating intracellular killing of mycobacteria through pharmacological enhancement of macroautophagy might provide a novel therapeutic strategy. Despite the resistance of MTB to killing by basal autophagy, cell-based screening of FDA-approved drugs revealed two anticonvulsants, carbamazepine and valproic acid, that were able to stimulate autophagic killing of intracellular M. tuberculosis within primary human macrophages at concentrations achievable in humans. Using a zebrafish model, we show that carbamazepine can stimulate autophagy in vivo and enhance clearance of M. marinum, while in mice infected with a highly virulent multidrug-resistant MTB strain, carbamazepine treatment reduced bacterial burden, improved lung pathology and stimulated adaptive immunity. We show that carbamazepine induces antimicrobial autophagy through a novel, evolutionarily conserved, mTOR-independent pathway controlled by cellular depletion of myo-inositol. While strain-specific differences in susceptibility to in vivo carbamazepine treatment may exist, autophagy enhancement by repurposed drugs provides an easily implementable potential therapy for the treatment of multidrug-resistant mycobacterial infection. PMID:25535254

  3. A complex regulatory network coordinating cell cycles during C. elegans development is revealed by a genome-wide RNAi screen.

    PubMed

    Roy, Sarah H; Tobin, David V; Memar, Nadin; Beltz, Eleanor; Holmen, Jenna; Clayton, Joseph E; Chiu, Daniel J; Young, Laura D; Green, Travis H; Lubin, Isabella; Liu, Yuying; Conradt, Barbara; Saito, R Mako

    2014-05-01

    The development and homeostasis of multicellular animals requires precise coordination of cell division and differentiation. We performed a genome-wide RNA interference screen in Caenorhabditis elegans to reveal the components of a regulatory network that promotes developmentally programmed cell-cycle quiescence. The 107 identified genes are predicted to constitute regulatory networks that are conserved among higher animals because almost half of the genes are represented by clear human orthologs. Using a series of mutant backgrounds to assess their genetic activities, the RNA interference clones displaying similar properties were clustered to establish potential regulatory relationships within the network. This approach uncovered four distinct genetic pathways controlling cell-cycle entry during intestinal organogenesis. The enhanced phenotypes observed for animals carrying compound mutations attest to the collaboration between distinct mechanisms to ensure strict developmental regulation of cell cycles. Moreover, we characterized ubc-25, a gene encoding an E2 ubiquitin-conjugating enzyme whose human ortholog, UBE2Q2, is deregulated in several cancers. Our genetic analyses suggested that ubc-25 acts in a linear pathway with cul-1/Cul1, in parallel to pathways employing cki-1/p27 and lin-35/pRb to promote cell-cycle quiescence. Further investigation of the potential regulatory mechanism demonstrated that ubc-25 activity negatively regulates CYE-1/cyclin E protein abundance in vivo. Together, our results show that the ubc-25-mediated pathway acts within a complex network that integrates the actions of multiple molecular mechanisms to control cell cycles during development. PMID:24584095

  4. Chemical genetic screen in fission yeast reveals roles for vacuolar acidification, mitochondrial fission, and cellular GMP levels in lifespan extension.

    PubMed

    Stephan, Jessica; Franke, Jacqueline; Ehrenhofer-Murray, Ann E

    2013-08-01

    The discovery that genetic mutations in several cellular pathways can increase lifespan has lent support to the notion that pharmacological inhibition of aging pathways can be used to extend lifespan and to slow the onset of age-related diseases. However, so far, only few compounds with such activities have been described. Here, we have conducted a chemical genetic screen for compounds that cause the extension of chronological lifespan of Schizosaccharomyces pombe. We have characterized eight natural products with such activities, which has allowed us to uncover so far unknown anti-aging pathways in S. pombe. The ionophores monensin and nigericin extended lifespan by affecting vacuolar acidification, and this effect depended on the presence of the vacuolar ATPase (V-ATPase) subunits Vma1 and Vma3. Furthermore, prostaglandin J? displayed anti-aging properties due to the inhibition of mitochondrial fission, and its effect on longevity required the mitochondrial fission protein Dnm1 as well as the G-protein-coupled glucose receptor Git3. Also, two compounds that inhibit guanosine monophosphate (GMP) synthesis, mycophenolic acid (MPA) and acivicin, caused lifespan extension, indicating that an imbalance in guanine nucleotide levels impinges upon longevity. We furthermore have identified diindolylmethane (DIM), tschimganine, and the compound mixture mangosteen as inhibiting aging. Taken together, these results reveal unanticipated anti-aging activities for several phytochemicals and open up opportunities for the development of novel anti-aging therapies. PMID:23521895

  5. A genome-wide screen for methyl methanesulfonate-sensitive mutants reveals genes required for S phase progression in the presence of DNA damage

    Microsoft Academic Search

    Michael Chang; Mohammed Bellaoui; Charles Boone; Grant W. Brown

    2002-01-01

    We performed a systematic screen of the set of 5,000 viable Saccharomyces cerevisiae haploid gene deletion mutants and have identified 103 genes whose deletion causes sensitivity to the DNA-damaging agent methyl methanesulfonate (MMS). In total, 40 previously uncharacterized alkylation damage response genes were identified. Comparison with the set of genes known to be transcriptionally induced in response to MMS revealed

  6. An iTRAQ proteomics screen reveals the effects of the MDM2 binding ligand Nutlin-3 on cellular proteostasis.

    PubMed

    Nicholson, Judith; Neelagandan, Kalainanghi; Huart, Anne-Sophie; Ball, Kathryn; Molloy, Mark P; Hupp, Ted

    2012-11-01

    Mouse double minute 2 (MDM2) participates in protein synthesis, folding, and ubiquitin-mediated degradation and is therefore a proteostasic hub protein. The MDM2 interactome contains over 100 proteins, yet stratification of dominant MDM2-interacting proteins has not been achieved. 8-plex iTRAQ (nanoLC-MS/MS) of MCF7 cells treated with the MDM2-binding ligand Nutlin-3 identified the most abundant cellular protein changes over early time points; 1,323 unique proteins were identified including 35 with altered steady-state levels within 2 h of Nutlin-3 treatment, identifying a core group of MDM2 related proteins. Six of these proteins were previously identified MDM2 interactors, and the effects of Nutlin-3 on the MDM2-nucleophosmin interaction (NPM) was further validated. This revealed that Nutlin-3 mediates the in vivo conversion of NPM from an oligomer to a monomer as an MDM2-dependent phenomenon, with Nutlin-3 stimulating MDM2 binding to a peptide motif derived from the oligomerization interface of NPM. These data form the first proteomic screen of Nutlin-3 in cells whereby we (i) identify the most abundant MDM2-interacting proteins whose steady-state levels change early after Nutlin-3 treatment; (ii) identify the first protein apart from p53, nucleophosmin (NPM), whose interaction with MDM2 can be stimulated allosterically by Nutlin-3; and (iii) raise the possibility that Nutlin-3 might act as a general agonist of other MDM2 protein-protein interactions. PMID:23039052

  7. A chromatin immunoprecipitation screen in mouse keratinocytes reveals Runx1 as a direct transcriptional target of DeltaNp63.

    PubMed

    Ortt, Kori; Raveh, Eli; Gat, Uri; Sinha, Satrajit

    2008-07-01

    Development of the skin epidermis and appendages such as hair follicles involves coordinated processes of keratinocyte proliferation and differentiation. The transcription factor p63 plays a critical role in these steps as evident by a complete lack of these structures in p63 null mice. The p63 gene encodes for two proteins TAp63 and DeltaNp63, the latter being the more prevalent and dominant isoform expressed in keratinocytes. Although numerous p63 target genes have been identified, these studies have been limited to transformed human keratinocyte cell lines. Here, we have employed a genomic screening approach of chromatin immunoprecipitation (ChIP) coupled with an enrichment strategy to identify DeltaNp63 response elements in primary mouse keratinocytes. Analysis of p63-ChIP-derived DNA segments has revealed more than 100 potential target genes including novel as well as mouse counterparts of established human p63 targets. Among these is Runx1, a transcription factor important for hair follicle development. We demonstrate that DeltaNp63 binds to a p63-response element located within a well-conserved enhancer of the Runx1 gene. Furthermore, siRNA mediated reduction of DeltaNp63 in mouse keratinocytes reduces Runx1 expression. Consistent with this, endogenous Runx1 levels are lower in the skin of p63(+/-) animals as compared to wild type animals. Lastly, we demonstrate that DeltaNp63 and Runx1 are co-expressed in specific compartments of the hair follicle in a dynamic fashion. Taken together our data demonstrate that p63 directly regulates Runx1 gene expression through a novel enhancer element and suggests a role for these two transcription factors in dictating skin keratinocyte and appendage development. PMID:18275068

  8. A genome-wide RNA interference screen reveals an essential CREB3L2-ATF5-MCL1 survival pathway in malignant glioma with therapeutic implications

    Microsoft Academic Search

    Zhi Sheng; Li Li; Lihua J Zhu; Thomas W Smith; Andrea Demers; Alonzo H Ross; Richard P Moser; Michael R Green

    2010-01-01

    Activating transcription factor-5 (ATF5) is highly expressed in malignant glioma and has a key role in promoting cell survival. Here we perform a genome-wide RNAi screen to identify transcriptional regulators of ATF5. Our results reveal an essential survival pathway in malignant glioma, whereby activation of a RAS–mitogen-activated protein kinase or phosphoinositide-3-kinase signaling cascade leads to induction of the transcription factor

  9. Proteomic screening of variola virus reveals a unique NF-?B inhibitor that is highly conserved among pathogenic orthopoxviruses

    PubMed Central

    Mohamed, Mohamed R.; Rahman, Masmudur M.; Lanchbury, Jerry S.; Shattuck, Donna; Neff, Chris; Dufford, Max; van Buuren, Nick; Fagan, Katharine; Barry, Michele; Smith, Scott; Damon, Inger; McFadden, Grant

    2009-01-01

    Identification of the binary interactions between viral and host proteins has become a valuable tool for investigating viral tropism and pathogenesis. Here, we present the first systematic protein interaction screening of the unique variola virus proteome by using yeast 2-hybrid screening against a variety of human cDNA libraries. Several protein–protein interactions were identified, including an interaction between variola G1R, an ankryin/F-box containing protein, and human nuclear factor kappa-B1 (NF-?B1)/p105. This represents the first direct interaction between a pathogen-encoded protein and NF-?B1/p105. Orthologs of G1R are present in a variety of pathogenic orthopoxviruses, but not in vaccinia virus, and expression of any one of these viral proteins blocks NF-?B signaling in human cells. Thus, proteomic screening of variola virus has the potential to uncover modulators of the human innate antiviral responses. PMID:19451633

  10. Antimicrobial, antitumor and brine shrimp lethality assay of Ranunculus arvensis L. extracts.

    PubMed

    Bhatti, Muhammad Zeeshan; Ali, Amjad; Saeed, Asma; Saeed, Ahmad; Malik, Salman Akbar

    2015-05-01

    To investigate the antitumor activity, brine shrimp lethality assay, antibacterial and antifungal activity of Methanol Extract (ME), Water Extract (WE), Acetone Extract (AE), Chloroform Extract (CE), Methanol-Water Extract (MWE), Methanol-Acetone Extract (MAE), Methanol-Chloroform Extract (MCE) of Ranunculus arvensis (L.). Antitumor activity was evaluated with Agrobacterium tumefaciens (At10) induced potato disc assay. Cytotoxicity was evaluated with brine shrimp lethality assay. Antibacterial activity was evaluated with six bacterial strains including Escherichia coli, Enterobacter aerogenes, Bordetella bronchiseptica, Klebsiella pneumoniae, Micrococcus luteus and Streptococcus anginosus and antifungal screening was done against five fungal strains including Aspergillus niger, A. flavus, A. fumigates, Fusarium solani and Mucor species by using disc diffusion method. Best antitumor activity was obtained with ME and WE, having highest IC50 values 20.27 ± 1.62 and 93.01 ± 1.33?g/disc. Brine shrimp lethality assay showed LC50 values of AE, MAE and ME were obtained as 384.66 ± 9.42?g/ml, 724.11 ± 8.01?g/ml and 978.7 ±8.01 ?g/ml respectively. WE of R. arvensis revealed weak antimicrobial result against the tested microorganisms. On the other hand, the antifungal activity of the plant extracts was found to be insignificant. These findings demonstrate that extracts of R. arvensis possesses significant antitumor activity. Further extensive study is necessary to assess the therapeutic potential of the plant. PMID:26004705

  11. Tasers – Less than Lethal!

    PubMed Central

    Sharma, Abiram; Theivacumar, Nada S; Souka, Hesham M

    2009-01-01

    We report a case of potentially lethal injury associated with the use of Taser. A 42-year-old man was stopped by police for potential detention. He held a large carving knife over his epigasrium threatening to stab himself.With a view to achieving immobilisation, a Taser gun was used. On activation of the Taser, the subject suffered a 7-cm wide and 10-cm deep stab injury to the upper abdomen. In this case, activation of the Taser resulted in the contraction of skeletal muscles, flexors more intensely than extensors, resulting in the stab injury. PMID:19416583

  12. Comparative Haploid Genetic Screens Reveal Divergent Pathways in the Biogenesis and Trafficking of Glycophosphatidylinositol-Anchored Proteins.

    PubMed

    Davis, Eric M; Kim, Jihye; Menasche, Bridget L; Sheppard, Jacob; Liu, Xuedong; Tan, Aik-Choon; Shen, Jingshi

    2015-06-23

    Glycophosphatidylinositol-anchored proteins (GPI-APs) play essential roles in physiology, but their biogenesis and trafficking have not been systematically characterized. Here, we took advantage of the recently available haploid genetics approach to dissect GPI-AP pathways in human cells using prion protein (PrP) and CD59 as model molecules. Our screens recovered a large number of common and unexpectedly specialized factors in the GPI-AP pathways. PIGN, PGAP2, and PIGF, which encode GPI anchor-modifying enzymes, were selectively isolated in the CD59 screen, suggesting that GPI anchor composition significantly influences the biogenesis of GPI-APs in a substrate-dependent manner. SEC62 and SEC63, which encode components of the ER-targeting machinery, were selectively recovered in the PrP screen, indicating that they do not constitute a universal route for the biogenesis of mammalian GPI-APs. Together, these comparative haploid genetic screens demonstrate that, despite their similarity in overall architecture and subcellular localization, GPI-APs follow markedly distinct biosynthetic and trafficking pathways. PMID:26074080

  13. Synthetic Lethal Interaction between Oncogenic KRAS Dependency and STK33 Suppression in Human Cancer Cells

    E-print Network

    Scholl, Claudia

    An alternative to therapeutic targeting of oncogenes is to perform “synthetic lethalityscreens for genes that are essential only in the context of specific cancer-causing mutations. We used high-throughput RNA interference ...

  14. High-throughput screening reveals alsterpaullone, 2-cyanoethyl as a potent p27Kip1 transcriptional inhibitor.

    PubMed

    Walters, Brandon J; Lin, Wenwei; Diao, Shiyong; Brimble, Mark; Iconaru, Luigi I; Dearman, Jennifer; Goktug, Asli; Chen, Taosheng; Zuo, Jian

    2014-01-01

    p27Kip1 is a cell cycle inhibitor that prevents cyclin dependent kinase (CDK)/cyclin complexes from phosphorylating their targets. p27Kip1 is a known tumor suppressor, as the germline loss of p27Kip1 results in sporadic pituitary formation in aged rodents, and its presence in human cancers is indicative of a poor prognosis. In addition to its role in cancer, loss of p27Kip1 results in regenerative phenotypes in some tissues and maintenance of stem cell pluripotency, suggesting that p27Kip1 inhibitors could be beneficial for tissue regeneration. Because p27Kip1 is an intrinsically disordered protein, identifying direct inhibitors of the p27Kip1 protein is difficult. Therefore, we pursued a high-throughput screening strategy to identify novel p27Kip1 transcriptional inhibitors. We utilized a luciferase reporter plasmid driven by the p27Kip1 promoter to transiently transfect HeLa cells and used cyclohexamide as a positive control for non-specific inhibition. We screened a "bioactive" library consisting of 8,904 (4,359 unique) compounds, of which 830 are Food and Drug Administration (FDA) approved. From this screen, we successfully identified 111 primary hits with inhibitory effect against the promoter of p27Kip1. These hits were further refined using a battery of secondary screens. Here we report four novel p27Kip1 transcriptional inhibitors, and further demonstrate that our most potent hit compound (IC50?=?200 nM) Alsterpaullone 2-cyanoethyl, inhibits p27Kip1 transcription by preventing FoxO3a from binding to the p27Kip1 promoter. This screen represents one of the first attempts to identify inhibitors of p27Kip1 and may prove useful for future tissue regeneration studies. PMID:24646893

  15. High-Throughput Screening Reveals Alsterpaullone, 2-Cyanoethyl as a Potent p27Kip1 Transcriptional Inhibitor

    PubMed Central

    Walters, Brandon J.; Lin, Wenwei; Diao, Shiyong; Brimble, Mark; Iconaru, Luigi I.; Dearman, Jennifer; Goktug, Asli; Chen, Taosheng; Zuo, Jian

    2014-01-01

    p27Kip1 is a cell cycle inhibitor that prevents cyclin dependent kinase (CDK)/cyclin complexes from phosphorylating their targets. p27Kip1 is a known tumor suppressor, as the germline loss of p27Kip1 results in sporadic pituitary formation in aged rodents, and its presence in human cancers is indicative of a poor prognosis. In addition to its role in cancer, loss of p27Kip1 results in regenerative phenotypes in some tissues and maintenance of stem cell pluripotency, suggesting that p27Kip1 inhibitors could be beneficial for tissue regeneration. Because p27Kip1 is an intrinsically disordered protein, identifying direct inhibitors of the p27Kip1 protein is difficult. Therefore, we pursued a high-throughput screening strategy to identify novel p27Kip1 transcriptional inhibitors. We utilized a luciferase reporter plasmid driven by the p27Kip1 promoter to transiently transfect HeLa cells and used cyclohexamide as a positive control for non-specific inhibition. We screened a “bioactive” library consisting of 8,904 (4,359 unique) compounds, of which 830 are Food and Drug Administration (FDA) approved. From this screen, we successfully identified 111 primary hits with inhibitory effect against the promoter of p27Kip1. These hits were further refined using a battery of secondary screens. Here we report four novel p27Kip1 transcriptional inhibitors, and further demonstrate that our most potent hit compound (IC50?=?200 nM) Alsterpaullone 2-cyanoethyl, inhibits p27Kip1 transcription by preventing FoxO3a from binding to the p27Kip1 promoter. This screen represents one of the first attempts to identify inhibitors of p27Kip1 and may prove useful for future tissue regeneration studies. PMID:24646893

  16. In vitro screening of compounds against laboratory and field isolates of human hookworm reveals quantitative differences in anthelmintic susceptibility.

    PubMed

    Treger, Rebecca S; Otchere, Joseph; Keil, Martin F; Quagraine, Josephine E; Rai, Ganesha; Mott, Bryan T; Humphries, Debbie L; Wilson, Michael; Cappello, Michael; Vermeire, Jon J

    2014-01-01

    A panel of 80 compounds was screened for anthelmintic activity against a laboratory strain of Ancylostoma ceylanicum and field isolates of hookworm obtained from school children in the Kintampo North District of the Brong Ahafo Region of Ghana. Although the laboratory strain of A. ceylanicum was more susceptible to the compounds tested than the field isolates of hookworm, a twofold increase in compound concentration resulted in comparable egg hatch percent inhibition for select compounds. These data provide evidence that the efficacy of anthelmintic compounds may be species-dependent and that field and laboratory strains of hookworm differ in their sensitivities to the anthelmintics tested. These data also suggest that both compound concentration and hookworm species must be considered when screening to identify novel anthelmintic compounds. PMID:24297811

  17. In vitro Screening of Compounds against Laboratory and Field Isolates of Human Hookworm Reveals Quantitative Differences in Anthelmintic Susceptibility

    PubMed Central

    Treger, Rebecca S.; Otchere, Joseph; Keil, Martin F.; Quagraine, Josephine E.; Rai, Ganesha; Mott, Bryan T.; Humphries, Debbie L.; Wilson, Michael; Cappello, Michael; Vermeire, Jon J.

    2014-01-01

    A panel of 80 compounds was screened for anthelmintic activity against a laboratory strain of Ancylostoma ceylanicum and field isolates of hookworm obtained from school children in the Kintampo North District of the Brong Ahafo Region of Ghana. Although the laboratory strain of A. ceylanicum was more susceptible to the compounds tested than the field isolates of hookworm, a twofold increase in compound concentration resulted in comparable egg hatch percent inhibition for select compounds. These data provide evidence that the efficacy of anthelmintic compounds may be species-dependent and that field and laboratory strains of hookworm differ in their sensitivities to the anthelmintics tested. These data also suggest that both compound concentration and hookworm species must be considered when screening to identify novel anthelmintic compounds. PMID:24297811

  18. A systematic RNAi screen reveals involvement of endocytic pathway in neuronal dysfunction in  -synuclein transgenic C. elegans

    Microsoft Academic Search

    Tomoki Kuwahara; Akihiko Koyama; Shingo Koyama; Sawako Yoshina; Chang-Hong Ren; Takeo Kato; Shohei Mitani; Takeshi Iwatsubo

    2008-01-01

    Mutations or multiplications in a-synuclein gene cause familial forms of Parkinson disease or dementia with Lewy bodies (LB), and the deposition of wild-type a-synuclein as LB occurs as a hallmark lesion of these dis- orders, collectively referred to as synucleinopathies, implicating a-synuclein in the pathogenesis of synuclei- nopathy. To identify modifier genes of a-synuclein-induced neurotoxicity, we conducted an RNAi screen

  19. Comparative RNAi Screens in C. elegans and C. briggsae Reveal the Impact of Developmental System Drift on Gene Function

    PubMed Central

    Verster, Adrian J.; Ramani, Arun K.; McKay, Sheldon J.; Fraser, Andrew G.

    2014-01-01

    Although two related species may have extremely similar phenotypes, the genetic networks underpinning this conserved biology may have diverged substantially since they last shared a common ancestor. This is termed Developmental System Drift (DSD) and reflects the plasticity of genetic networks. One consequence of DSD is that some orthologous genes will have evolved different in vivo functions in two such phenotypically similar, related species and will therefore have different loss of function phenotypes. Here we report an RNAi screen in C. elegans and C. briggsae to identify such cases. We screened 1333 genes in both species and identified 91 orthologues that have different RNAi phenotypes. Intriguingly, we find that recently evolved genes of unknown function have the fastest evolving in vivo functions and, in several cases, we identify the molecular events driving these changes. We thus find that DSD has a major impact on the evolution of gene function and we anticipate that the C. briggsae RNAi library reported here will drive future studies on comparative functional genomics screens in these nematodes. PMID:24516395

  20. Comparative RNAi screens in C. elegans and C. briggsae reveal the impact of developmental system drift on gene function.

    PubMed

    Verster, Adrian J; Ramani, Arun K; McKay, Sheldon J; Fraser, Andrew G

    2014-02-01

    Although two related species may have extremely similar phenotypes, the genetic networks underpinning this conserved biology may have diverged substantially since they last shared a common ancestor. This is termed Developmental System Drift (DSD) and reflects the plasticity of genetic networks. One consequence of DSD is that some orthologous genes will have evolved different in vivo functions in two such phenotypically similar, related species and will therefore have different loss of function phenotypes. Here we report an RNAi screen in C. elegans and C. briggsae to identify such cases. We screened 1333 genes in both species and identified 91 orthologues that have different RNAi phenotypes. Intriguingly, we find that recently evolved genes of unknown function have the fastest evolving in vivo functions and, in several cases, we identify the molecular events driving these changes. We thus find that DSD has a major impact on the evolution of gene function and we anticipate that the C. briggsae RNAi library reported here will drive future studies on comparative functional genomics screens in these nematodes. PMID:24516395

  1. Genome-Wide Small Interfering RNA Screens Reveal VAMP3 as a Novel Host Factor Required for Uukuniemi Virus Late Penetration

    PubMed Central

    Meier, Roger; Franceschini, Andrea; Horvath, Peter; Tetard, Marilou; Mancini, Roberta; von Mering, Christian; Helenius, Ari

    2014-01-01

    ABSTRACT The Bunyaviridae constitute a large family of enveloped animal viruses, many of which are important emerging pathogens. How bunyaviruses enter and infect mammalian cells remains largely uncharacterized. We used two genome-wide silencing screens with distinct small interfering RNA (siRNA) libraries to investigate host proteins required during infection of human cells by the bunyavirus Uukuniemi virus (UUKV), a late-penetrating virus. Sequence analysis of the libraries revealed that many siRNAs in the screens inhibited infection by silencing not only the intended targets but additional genes in a microRNA (miRNA)-like manner. That the 7-nucleotide seed regions in the siRNAs can cause a perturbation in infection was confirmed by using synthetic miRNAs (miRs). One of the miRs tested, miR-142-3p, was shown to interfere with the intracellular trafficking of incoming viruses by regulating the v-SNARE VAMP3, a strong hit shared by both siRNA screens. Inactivation of VAMP3 by the tetanus toxin led to a block in infection. Using fluorescence-based techniques in fixed and live cells, we found that the viruses enter VAMP3+ endosomal vesicles 5 min after internalization and that colocalization was maximal 15 min thereafter. At this time, LAMP1 was associated with the VAMP3+ virus-containing endosomes. In cells depleted of VAMP3, viruses were mainly trapped in LAMP1-negative compartments. Together, our results indicated that UUKV relies on VAMP3 for penetration, providing an indication of added complexity in the trafficking of viruses through the endocytic network. IMPORTANCE Bunyaviruses represent a growing threat to humans and livestock globally. Unfortunately, relatively little is known about these emerging pathogens. We report here the first human genome-wide siRNA screens for a bunyavirus. The screens resulted in the identification of 562 host cell factors with a potential role in cell entry and virus replication. To demonstrate the robustness of our approach, we confirmed and analyzed the role of the v-SNARE VAMP3 in Uukuniemi virus entry and infection. The information gained lays the basis for future research into the cell biology of bunyavirus infection and new antiviral strategies. In addition, by shedding light on serious caveats in large-scale siRNA screening, our experimental and bioinformatics procedures will be valuable in the comprehensive analysis of past and future high-content screening data. PMID:24850728

  2. Lethality test system

    SciTech Connect

    Parsons, W.M.; Sims, J.R.; Parker, J.V.

    1986-01-01

    The Lethality Test System (LTS), presently under construction at Los Alamos, is an electromagnetic launcher facility designed to perform impact experiments at velocities up to 15 km/s. The launcher is a 25 mm round bore, plasma armature railgun extending 22 m in length. Preinjection is accomplished with a two-stage gas gun capable of 7 km/s. The railgun power supply utilizes traction motors, vacuum interrupters, and pulse transformers. An assembly of 28 traction motors, equipped with flywheels, stores approximately 80 MJ at 92% of full speed and energizes the primary windings of three pulse transformers at a current of 50 kA. At peak current an array of vacuum interrupters disconnects the transformer primary windings and forces the current to flow in the secondary windings. The secondary windings are connected to the railgun, and by staging the vacuum interrupter openings, a 1 MA to 1.3 MA ramped current waveform will be delivered to the railgun.

  3. Potential lethal and non-lethal effects of predators on dispersal of spider mites.

    PubMed

    Otsuki, Hatsune; Yano, Shuichi

    2014-11-01

    Predators can affect prey dispersal lethally by direct consumption or non-lethally by making prey hesitate to disperse. These lethal and non-lethal effects are detectable only in systems where prey can disperse between multiple patches. However, most studies have drawn their conclusions concerning the ability of predatory mites to suppress spider mites based on observations of their interactions on a single patch or on heavily infested host plants where spider mites could hardly disperse toward intact patches. In these systems, specialist predatory mites that penetrate protective webs produced by spider mites quickly suppress the spider mites, whereas generalist predators that cannot penetrate the webs were ineffective. By using a connected patch system, we revealed that a generalist ant, Pristomyrmex punctatus Mayr (Hymenoptera: Formicidae), effectively prevented dispersal of spider mites, Tetranychus kanzawai Kishida (Acari: Tetranychidae), by directly consuming dispersing individuals. We also revealed that a generalist predatory mite, Euseius sojaensis Ehara (Acari: Phytoseiidae), prevented between-patch dispersal of T. kanzawai by making them hesitate to disperse. In contrast, a specialist phytoseiid predatory mite, Neoseiulus womersleyi Schicha, allowed spider mites to escape an initial patch, increasing the number of colonized patches within the system. Our results suggest that ants and generalist predatory mites can effectively suppress Tetranychus species under some conditions, and should receive more attention as agents for conservation biological control in agroecosystems. PMID:24867061

  4. High-throughput screening of FDA-approved drugs using oxygen biosensor plates reveals secondary mitofunctional effects

    PubMed Central

    Sahdeo, Sunil; Tomilov, Alexey; Komachi, Kelly; Iwahashi, Christine; Datta, Sandipan; Hughes, Owen; Hagerman, Paul; Cortopassi, Gino

    2014-01-01

    Repurposing of FDA-approved drugs with effects on mitochondrial function might shorten the critical path to mitochondrial disease drug development. We improved a biosensor-based assay of mitochondrial O2 consumption, and identified mitofunctional defects in cell models of LHON and FXTAS. Using this platform, we screened a 1600-compound library of clinically used drugs. The assay identified drugs known to affect mitochondrial function, such as metformin and decoquinate. We also identified several drugs not previously known to affect mitochondrial respiration including acarbose, metaraminol, gallamine triethiodide, and acamprosate. These previously unknown ‘mitoactives’ represent novel links to targets for mitochondrial regulation and potentially therapy, for mitochondrial disease. PMID:25034306

  5. Approaches to Identifying Synthetic Lethal Interactions in Cancer

    PubMed Central

    Thompson, Jordan M.; Nguyen, Quy H.; Singh, Manpreet; Razorenova, Olga V.

    2015-01-01

    Targeting synthetic lethal interactions is a promising new therapeutic approach to exploit specific changes that occur within cancer cells. Multiple approaches to investigate these interactions have been developed and successfully implemented, including chemical, siRNA, shRNA, and CRISPR library screens. Genome-wide computational approaches, such as DAISY, also have been successful in predicting synthetic lethal interactions from both cancer cell lines and patient samples. Each approach has its advantages and disadvantages that need to be considered depending on the cancer type and its molecular alterations. This review discusses these approaches and examines case studies that highlight their use.

  6. Screen of FDA-approved drug library reveals compounds that protect hair cells from aminoglycosides and cisplatin

    PubMed Central

    Vlasits, Anna L.; Simon, Julian A.; Raible, David W.; Rubel, Edwin W; Owens, Kelly N.

    2012-01-01

    Loss of mechanosensory hair cells in the inner ear accounts for many hearing loss and balance disorders. Several beneficial pharmaceutical drugs cause hair cell death as a side effect. These include aminoglycoside antibiotics, such as neomycin, kanamycin and gentamicin, and several cancer chemotherapy drugs, such as cisplatin. Discovering new compounds that protect mammalian hair cells from toxic insults is experimentally difficult because of the inaccessibility of the inner ear. We used the zebrafish lateral line sensory system as an in vivo screening platform to survey a library of FDA-approved pharmaceuticals for compounds that protect hair cells from neomycin, gentamicin, kanamycin and cisplatin. Ten compounds were identified that provide protection from at least two of the four toxins. The resulting compounds fall into several drug classes, including serotonin and dopamine-modulating drugs, adrenergic receptor ligands, and estrogen receptor modulators. The protective compounds show different effects against the different toxins, supporting the idea that each toxin causes hair cell death by distinct, but partially overlapping, mechanisms. Furthermore, some compounds from the same drug classes had different protective properties, suggesting that they might not prevent hair cell death by their known target mechanisms. Some protective compounds blocked gentamicin uptake into hair cells, suggesting that they may block mechanotransduction or other routes of entry. The protective compounds identified in our screen will provide a starting point for studies in mammals as well as further research discovering the cellular signaling pathways that trigger hair cell death. PMID:22967486

  7. High-throughput 3D screening reveals differences in drug sensitivities between culture models of JIMT1 breast cancer cells.

    PubMed

    Hongisto, Vesa; Jernström, Sandra; Fey, Vidal; Mpindi, John-Patrick; Kleivi Sahlberg, Kristine; Kallioniemi, Olli; Perälä, Merja

    2013-01-01

    The traditional method for studying cancer in vitro is to grow immortalized cancer cells in two-dimensional monolayers on plastic. However, many cellular features are impaired in these artificial conditions, and large changes in gene expression compared to tumors have been reported. Three-dimensional cell culture models have become increasingly popular and are suggested to be better models than two-dimensional monolayers due to improved cell-to-cell contact and structures that resemble in vivo architecture. The aim of this study was to develop a simple high-throughput three-dimensional drug screening method and to compare drug responses in JIMT1 breast cancer cells when grown in two dimensions, in poly(2-hydroxyethyl methacrylate) induced anchorage-independent three-dimensional models, and in Matrigel three-dimensional cell culture models. We screened 102 compounds with multiple concentrations and biological replicates for their effects on cell proliferation. The cells were either treated immediately upon plating, or they were allowed to grow in three-dimensional cultures for 4 days before the drug treatment. Large variations in drug responses were observed between the models indicating that comparisons of culture model-influenced drug sensitivities cannot be made based on the effects of a single drug. However, we show with the 63 most prominent drugs that, in general, JIMT1 cells grown on Matrigel were significantly more sensitive to drugs than cells grown in two-dimensional cultures, while the responses of cells grown in poly(2-hydroxyethyl methacrylate) resembled those of the two-dimensional cultures. Furthermore, comparing the gene expression profiles of the cell culture models to xenograft tumors indicated that cells cultured in Matrigel and as xenografts most closely resembled each other. In this study, we also suggest that three-dimensional cultures can provide a platform for systematic experimentation of larger compound collections in a high-throughput mode and be used as alternatives to traditional two-dimensional screens for better comparability to the in vivo state. PMID:24194875

  8. High-Throughput 3D Screening Reveals Differences in Drug Sensitivities between Culture Models of JIMT1 Breast Cancer Cells

    PubMed Central

    Fey, Vidal; Mpindi, John-Patrick; Kleivi Sahlberg, Kristine; Kallioniemi, Olli; Perälä, Merja

    2013-01-01

    The traditional method for studying cancer in vitro is to grow immortalized cancer cells in two-dimensional monolayers on plastic. However, many cellular features are impaired in these artificial conditions, and large changes in gene expression compared to tumors have been reported. Three-dimensional cell culture models have become increasingly popular and are suggested to be better models than two-dimensional monolayers due to improved cell-to-cell contact and structures that resemble in vivo architecture. The aim of this study was to develop a simple high-throughput three-dimensional drug screening method and to compare drug responses in JIMT1 breast cancer cells when grown in two dimensions, in poly(2-hydroxyethyl methacrylate) induced anchorage-independent three-dimensional models, and in Matrigel three-dimensional cell culture models. We screened 102 compounds with multiple concentrations and biological replicates for their effects on cell proliferation. The cells were either treated immediately upon plating, or they were allowed to grow in three-dimensional cultures for 4 days before the drug treatment. Large variations in drug responses were observed between the models indicating that comparisons of culture model-influenced drug sensitivities cannot be made based on the effects of a single drug. However, we show with the 63 most prominent drugs that, in general, JIMT1 cells grown on Matrigel were significantly more sensitive to drugs than cells grown in two-dimensional cultures, while the responses of cells grown in poly(2-hydroxyethyl methacrylate) resembled those of the two-dimensional cultures. Furthermore, comparing the gene expression profiles of the cell culture models to xenograft tumors indicated that cells cultured in Matrigel and as xenografts most closely resembled each other. In this study, we also suggest that three-dimensional cultures can provide a platform for systematic experimentation of larger compound collections in a high-throughput mode and be used as alternatives to traditional two-dimensional screens for better comparability to the in vivo state. PMID:24194875

  9. A Chemical Screening Approach Reveals that Indole Flourescence is Quenched by Pre-Fibrillar But Not Fibrillar Amyloid-?

    PubMed Central

    Reinke, Ashley A.; Seh, Han Yiau; Gestwicki, Jason E.

    2009-01-01

    Aggregated amyloid-? (A?) peptide is implicated in the pathology of Alzheimer’s disease. In vitro and in vivo, these aggregates are found in a variety of morphologies, including globular oligomers and linear fibrils, which possess distinct biological activities. However, known chemical probes, including the dyes thioflavin T and Congo Red, appear to lack selectivity for specific amyloid structures. To identify molecules that might differentiate between these architectures, we employed a fluorescence-based interaction assay to screen a collection of 68 known A? ligands against pre-formed oligomers and fibrils. In these studies, we found that the fluorescence of five indole-based compounds was selectively quenched (~15%) in the presence of oligomers, but remained unchanged after addition of fibrils. These results suggest that indoles might be complementary to existing chemical probes for studying amyloid formation in vitro. PMID:19640715

  10. Screening Active Compounds from Garcinia Species Native to China Reveals Novel Compounds Targeting the STAT/JAK Signaling Pathway

    PubMed Central

    Xu, Linfeng; Lao, Yuanzhi; Zhao, Yanhui; Qin, Jian; Fu, Wenwei; Zhang, Yingjia; Xu, Hongxi

    2015-01-01

    Natural compounds from medicinal plants are important resources for drug development. In a panel of human tumor cells, we screened a library of the natural products from Garcinia species which have anticancer potential to identify new potential therapeutic leads and discovered that caged xanthones were highly effective at suppressing multiple cancer cell lines. Their anticancer activities mainly depended on apoptosis pathways. For compounds in sensitive cancer line, their mechanisms of mode of action were evaluated. 33-Hydroxyepigambogic acid and 35-hydroxyepigambogic acid exhibited about 1??M IC50 values against JAK2/JAK3 kinases and less than 1??M IC50 values against NCI-H1650 cell which autocrined IL-6. Thus these two compounds provided a new antitumor molecular scaffold. Our report describes 33-hydroxyepigambogic acid and 35-hydroxyepigambogic acid that inhibited NCI-H1650 cell growth by suppressing constitutive STAT3 activation via direct inhibition of JAK kinase activity. PMID:26090459

  11. Genomewide Screen for Negative Regulators of Sirtuin Activity in Saccharomyces cerevisiae Reveals 40 Loci and Links to Metabolism

    PubMed Central

    Raisner, Ryan M.; Madhani, Hiten D.

    2008-01-01

    Sirtuins are conserved proteins implicated in myriad key processes including gene control, aging, cell survival, metabolism, and DNA repair. In Saccharomyces cerevisiae, the sirtuin Silent information regulator 2 (Sir2) promotes silent chromatin formation, suppresses recombination between repeats, and inhibits senescence. We performed a genomewide screen for factors that negatively regulate Sir activity at a reporter gene placed immediately outside a silenced region. After linkage analysis, assessment of Sir dependency, and knockout tag verification, 40 loci were identified, including 20 that have not been previously described to regulate Sir. In addition to chromatin-associated factors known to prevent ectopic silencing (Bdf1, SAS-I complex, Rpd3L complex, Ku), we identified the Rtt109 DNA repair-associated histone H3 lysine 56 acetyltransferase as an anti-silencing factor. Our findings indicate that Rtt109 functions independently of its proposed effectors, the Rtt101 cullin, Mms1, and Mms22, and demonstrate unexpected interplay between H3K56 and H4K16 acetylation. The screen also identified subunits of mediator (Soh1, Srb2, and Srb5) and mRNA metabolism factors (Kem1, Ssd1), thus raising the possibility that weak silencing affects some aspect of mRNA structure. Finally, several factors connected to metabolism were identified. These include the PAS-domain metabolic sensor kinase Psk2, the mitochondrial homocysteine detoxification enzyme Lap3, and the Fe-S cluster protein maturase Isa2. We speculate that PAS kinase may integrate metabolic signals to control sirtuin activity. PMID:18689887

  12. Genomewide screen for negative regulators of sirtuin activity in Saccharomyces cerevisiae reveals 40 loci and links to metabolism.

    PubMed

    Raisner, Ryan M; Madhani, Hiten D

    2008-08-01

    Sirtuins are conserved proteins implicated in myriad key processes including gene control, aging, cell survival, metabolism, and DNA repair. In Saccharomyces cerevisiae, the sirtuin Silent information regulator 2 (Sir2) promotes silent chromatin formation, suppresses recombination between repeats, and inhibits senescence. We performed a genomewide screen for factors that negatively regulate Sir activity at a reporter gene placed immediately outside a silenced region. After linkage analysis, assessment of Sir dependency, and knockout tag verification, 40 loci were identified, including 20 that have not been previously described to regulate Sir. In addition to chromatin-associated factors known to prevent ectopic silencing (Bdf1, SAS-I complex, Rpd3L complex, Ku), we identified the Rtt109 DNA repair-associated histone H3 lysine 56 acetyltransferase as an anti-silencing factor. Our findings indicate that Rtt109 functions independently of its proposed effectors, the Rtt101 cullin, Mms1, and Mms22, and demonstrate unexpected interplay between H3K56 and H4K16 acetylation. The screen also identified subunits of mediator (Soh1, Srb2, and Srb5) and mRNA metabolism factors (Kem1, Ssd1), thus raising the possibility that weak silencing affects some aspect of mRNA structure. Finally, several factors connected to metabolism were identified. These include the PAS-domain metabolic sensor kinase Psk2, the mitochondrial homocysteine detoxification enzyme Lap3, and the Fe-S cluster protein maturase Isa2. We speculate that PAS kinase may integrate metabolic signals to control sirtuin activity. PMID:18689887

  13. A direct screen for c-di-GMP modulators reveals a Salmonella Typhimurium periplasmic ?-arginine-sensing pathway.

    PubMed

    Mills, Erez; Petersen, Erik; Kulasekara, Bridget R; Miller, Samuel I

    2015-01-01

    Cyclic-di-GMP (c-di-GMP) is a bacterial second messenger that transduces internal and external signals and regulates bacterial motility and biofilm formation. Some organisms encode more than 100 c-di-GMP-modulating enzymes, but only for a few has a signal been defined that modulates their activity. We developed and applied a high-throughput, real-time flow cytometry method that uses a fluorescence resonance energy transfer (FRET)-based biosensor of free c-di-GMP to screen for signals that modulate its concentration within Salmonella Typhimurium. We identified multiple compounds, including glucose, N-acetyl-d-glucosamine, salicylic acid, and ?-arginine, that modulated the FRET signal and therefore the free c-di-GMP concentration. By screening a library of mutants, we identified proteins required for the c-di-GMP response to each compound. Furthermore, low micromolar concentrations of ?-arginine induced a rapid translation-independent increase in c-di-GMP concentrations and c-di-GMP-dependent cellulose synthesis, responses that required the regulatory periplasmic domain of the diguanylate cyclase STM1987. ?-Arginine signaling also required the periplasmic putative ?-arginine-binding protein ArtI, implying that ?-arginine sensing occurred in the periplasm. Among the 20 commonly used amino acids, S. Typhimurium specifically responded to ?-arginine with an increase in c-di-GMP, suggesting that ?-arginine may serve as a signal during S. Typhimurium infection. Our results demonstrate that a second-messenger biosensor can be used to identify environmental signals and define pathways that alter microbial behavior. PMID:26060330

  14. Generalized lethality criteria for beam weapon systems

    NASA Astrophysics Data System (ADS)

    Tsai, J.

    The lethality criteria which define the interaciton between the directed energy systems and the targets have been derived. For high energy laser systems, the lethality criteria are defined in terms of a lethal fluence. For neutral particle beam systems, the lathality criteria can be specified either as lethal charge fluence or as lethal dose. Derivations are all based on an assumed circular profile in the intensity distribution along the radial direction. Three different methods to calculate the lethality criteria are introduced. Lethality criteria and dwell time requirements have been deduced. A relationship between the different lethality concepts has been formulated.

  15. Screening of lactic acid bacteria from Indonesia reveals glucansucrase and fructansucrase genes in two different Weissella confusa strains from soya.

    PubMed

    Malik, Amarila; Radji, Maksum; Kralj, Slavko; Dijkhuizen, Lubbert

    2009-11-01

    Homopolysaccharide (glucan and fructan) synthesis from sucrose by sucrase enzymes in lactic acid bacteria (LAB) has been well studied in the genera Leuconostoc, Streptococcus and Lactobacillus. This study aimed to identify and characterize genes encoding glucansucrase/glucosyltransferase (GTF) and fructansucrases/fructosyltransferase (FTF) enzymes from genomic DNA of 'rare' Indonesian exopolysaccharide-producing LAB. From a total of 63 exopolysaccharide-producing LAB isolates obtained from foods, beverages and environmental samples, 18 isolates showing the most slimy and mucoid colony morphologies on sucrose were chosen for further study. By comparing bacterial growth on De Man, Rogosa and Sharpe (MRS)-sucrose with that on MRS-raffinose, and using the results of a previous PCR screening study with degenerate primer pairs targeting the conserved catalytic domain of GTFs, various strains were identified as producers of fructan (13), of glucan only (five) or as potential producers of both glucan and fructan (nine). Here, we report the characteristics of three gtf genes and one ftf gene obtained from Weissella confusa strains MBF8-1 and MBF8-2. Strain MBF8-1 harbored two putative gtf genes with high sequence similarity to GTFB of Lactobacillus reuteri 121 and GTF180 of L. reuteri 180, respectively. Strain MBF8-2 possessed single gtf and ftf genes with high sequence similarity to GTFKg3 of Lactobacillus fermentum Kg3 and DSRWC of Weissella cibaria, and FTF levansucrase of L. reuteri 121, respectively. PMID:19758326

  16. Large-scale screening of transcription factor-promoter interactions in spruce reveals a transcriptional network involved in vascular development.

    PubMed

    Duval, Isabelle; Lachance, Denis; Giguère, Isabelle; Bomal, Claude; Morency, Marie-Josée; Pelletier, Gervais; Boyle, Brian; MacKay, John J; Séguin, Armand

    2014-06-01

    This research aimed to investigate the role of diverse transcription factors (TFs) and to delineate gene regulatory networks directly in conifers at a relatively high-throughput level. The approach integrated sequence analyses, transcript profiling, and development of a conifer-specific activation assay. Transcript accumulation profiles of 102 TFs and potential target genes were clustered to identify groups of coordinately expressed genes. Several different patterns of transcript accumulation were observed by profiling in nine different organs and tissues: 27 genes were preferential to secondary xylem both in stems and roots, and other genes were preferential to phelloderm and periderm or were more ubiquitous. A robust system has been established as a screening approach to define which TFs have the ability to regulate a given promoter in planta. Trans-activation or repression effects were observed in 30% of TF-candidate gene promoter combinations. As a proof of concept, phylogenetic analysis and expression and trans-activation data were used to demonstrate that two spruce NAC-domain proteins most likely play key roles in secondary vascular growth as observed in other plant species. This study tested many TFs from diverse families in a conifer tree species, which broadens the knowledge of promoter-TF interactions in wood development and enables comparisons of gene regulatory networks found in angiosperms and gymnosperms. PMID:24713992

  17. Multi-Mycotoxin Screening Reveals the Occurrence of 139 Different Secondary Metabolites in Feed and Feed Ingredients

    PubMed Central

    Streit, Elisabeth; Schwab, Christina; Sulyok, Michael; Naehrer, Karin; Krska, Rudolf; Schatzmayr, Gerd

    2013-01-01

    The development of liquid chromatography-mass spectrometry (LC-MS)/mass spectrometry (MS) methods for the simultaneous detection and quantification of a broad spectrum of mycotoxins has facilitated the screening of a larger number of samples for contamination with a wide array of less well-known “emerging” mycotoxins and other metabolites. In this study, 83 samples of feed and feed raw materials were analysed. All of them were found to contain seven to 69 metabolites. The total number of detected metabolites amounts to 139. Fusarium mycotoxins were most common, but a number of Alternaria toxins also occurred very often. Furthermore, two so-called masked mycotoxins (i.e., mycotoxin conjugates), namely deoxynivalenol-3-glucoside (75% positives) and zearalenone-4-sulfate (49% positives), were frequently detected. Although the observed median concentrations of the individual analytes were generally in the low ?g/kg range, evaluating the toxicological potential of a given sample is difficult. Toxicity data on less well-known mycotoxins and other detected metabolites are notoriously scarce, as an overview on the available information on the most commonly detected metabolites shows. Besides, the possible synergistic effects of co-occurring substances have to be considered. PMID:23529186

  18. Mutational screening of the USH2A gene in Spanish USH patients reveals 23 novel pathogenic mutations

    PubMed Central

    2011-01-01

    Background Usher Syndrome type II (USH2) is an autosomal recessive disorder, characterized by moderate to severe hearing impairment and retinitis pigmentosa (RP). Among the three genes implicated, mutations in the USH2A gene account for 74-90% of the USH2 cases. Methods To identify the genetic cause of the disease and determine the frequency of USH2A mutations in a cohort of 88 unrelated USH Spanish patients, we carried out a mutation screening of the 72 coding exons of this gene by direct sequencing. Moreover, we performed functional minigene studies for those changes that were predicted to affect splicing. Results As a result, a total of 144 DNA sequence variants were identified. Based upon previous studies, allele frequencies, segregation analysis, bioinformatics' predictions and in vitro experiments, 37 variants (23 of them novel) were classified as pathogenic mutations. Conclusions This report provide a wide spectrum of USH2A mutations and clinical features, including atypical Usher syndrome phenotypes resembling Usher syndrome type I. Considering only the patients clearly diagnosed with Usher syndrome type II, and results obtained in this and previous studies, we can state that mutations in USH2A are responsible for 76.1% of USH2 disease in patients of Spanish origin. PMID:22004887

  19. RNAi screen in apoptotic cancer cell-stimulated human macrophages reveals co-regulation of IL-6/IL-10 expression.

    PubMed

    Ley, Stephanie; Weigert, Andreas; Hériché, Jean-Karim; Mille-Baker, Blandine; Janssen, Richard A J; Brüne, Bernhard

    2013-01-01

    Tumor-associated macrophages (TAM) are a major supportive component within neoplasms and are characterized by a plethora of functions that facilitate tumor outgrowth. Mechanisms of macrophage attraction and differentiation to a tumor-promoting phenotype, defined among others by distinct cytokine patterns such as pronounced interleukin (IL-10) production, are ill-defined. We aimed to identify signaling pathways that contribute to the generation of TAM-like macrophages using an adenoviral RNAi-based approach. Primary human monocyte-derived macrophages were stimulated with apoptotic tumor cell supernatants (ACM) to induce a TAM-like phenotype, characterized by secretion of IL-10, IL-6, IL-8 but repression of IL-12. For the high-throughput screen, macrophages were transduced with 8495 constructs of the adenoviral shRNA SilenceSelect(®) library of Galapagos BV, which aims at identifying druggable targets. We identified 96 genes involved in IL-10 production in response to ACM and observed a pronounced cluster of targets regulating both IL-10 and IL-6. Validation of five targets within the IL-10/IL-6 cluster was performed using siRNA or pharmacological inhibitors in human primary macrophages. Among those, interleukin 4 receptor-? and cannabinoid receptor 2 were confirmed as regulators of IL-10 and IL-6 secretion by ACM-stimulated macrophages. Our approach characterizes cellular functions of transfection-resistant, highly plastic and versatile cells and identifies novel targets involved in the generation of a TAM-like phenotype in human macrophages. PMID:22445721

  20. A genetic screen based on in vivo RNA imaging reveals centrosome-independent mechanisms for localizing gurken transcripts in Drosophila.

    PubMed

    Hayashi, Rippei; Wainwright, S Mark; Liddell, Sophie J; Pinchin, Sheena M; Horswell, Stuart; Ish-Horowicz, David

    2014-04-01

    We have screened chromosome arm 3L for ethyl methanesulfonate-induced mutations that disrupt localization of fluorescently labeled gurken (grk) messenger (m)RNA, whose transport along microtubules establishes both major body axes of the developing Drosophila oocyte. Rapid identification of causative mutations by single-nucleotide polymorphism recombinational mapping and whole-genomic sequencing allowed us to define nine complementation groups affecting grk mRNA localization and other aspects of oogenesis, including alleles of elg1, scaf6, quemao, nudE, Tsc2/gigas, rasp, and Chd5/Wrb, and several null alleles of the armitage Piwi-pathway gene. Analysis of a newly induced kinesin light chain allele shows that kinesin motor activity is required for both efficient grk mRNA localization and oocyte centrosome integrity. We also show that initiation of the dorsoanterior localization of grk mRNA precedes centrosome localization, suggesting that microtubule self-organization contributes to breaking axial symmetry to generate a unique dorsoventral axis. PMID:24531791

  1. Systematic screening reveals a role for BRCA1 in the response to transcription-associated DNA damage

    PubMed Central

    Hill, Sarah J.; Rolland, Thomas; Adelmant, Guillaume; Xia, Xianfang; Owen, Matthew S.; Dricot, Amélie; Zack, Travis I.; Sahni, Nidhi; Jacob, Yves; Hao, Tong; McKinney, Kristine M.; Clark, Allison P.; Reyon, Deepak; Tsai, Shengdar Q.; Joung, J. Keith; Beroukhim, Rameen; Marto, Jarrod A.; Vidal, Marc; Gaudet, Suzanne; Hill, David E.

    2014-01-01

    BRCA1 is a breast and ovarian tumor suppressor. Given its numerous incompletely understood functions and the possibility that more exist, we performed complementary systematic screens in search of new BRCA1 protein-interacting partners. New BRCA1 functions and/or a better understanding of existing ones were sought. Among the new interacting proteins identified, genetic interactions were detected between BRCA1 and four of the interactors: TONSL, SETX, TCEANC, and TCEA2. Genetic interactions were also detected between BRCA1 and certain interactors of TONSL, including both members of the FACT complex. From these results, a new BRCA1 function in the response to transcription-associated DNA damage was detected. Specifically, new roles for BRCA1 in the restart of transcription after UV damage and in preventing or repairing damage caused by stabilized R loops were identified. These roles are likely carried out together with some of the newly identified interactors. This new function may be important in BRCA1 tumor suppression, since the expression of several interactors, including some of the above-noted transcription proteins, is repeatedly aberrant in both breast and ovarian cancers. PMID:25184681

  2. Systematic screening reveals a role for BRCA1 in the response to transcription-associated DNA damage.

    PubMed

    Hill, Sarah J; Rolland, Thomas; Adelmant, Guillaume; Xia, Xianfang; Owen, Matthew S; Dricot, Amélie; Zack, Travis I; Sahni, Nidhi; Jacob, Yves; Hao, Tong; McKinney, Kristine M; Clark, Allison P; Reyon, Deepak; Tsai, Shengdar Q; Joung, J Keith; Beroukhim, Rameen; Marto, Jarrod A; Vidal, Marc; Gaudet, Suzanne; Hill, David E; Livingston, David M

    2014-09-01

    BRCA1 is a breast and ovarian tumor suppressor. Given its numerous incompletely understood functions and the possibility that more exist, we performed complementary systematic screens in search of new BRCA1 protein-interacting partners. New BRCA1 functions and/or a better understanding of existing ones were sought. Among the new interacting proteins identified, genetic interactions were detected between BRCA1 and four of the interactors: TONSL, SETX, TCEANC, and TCEA2. Genetic interactions were also detected between BRCA1 and certain interactors of TONSL, including both members of the FACT complex. From these results, a new BRCA1 function in the response to transcription-associated DNA damage was detected. Specifically, new roles for BRCA1 in the restart of transcription after UV damage and in preventing or repairing damage caused by stabilized R loops were identified. These roles are likely carried out together with some of the newly identified interactors. This new function may be important in BRCA1 tumor suppression, since the expression of several interactors, including some of the above-noted transcription proteins, is repeatedly aberrant in both breast and ovarian cancers. PMID:25184681

  3. Systematic screening of polyphosphate (poly P) levels in yeast mutant cells reveals strong interdependence with primary metabolism

    PubMed Central

    Freimoser, Florian M; Hürlimann, Hans Caspar; Jakob, Claude A; Werner, Thomas P; Amrhein, Nikolaus

    2006-01-01

    Background Inorganic polyphosphate (poly P) occurs universally in all organisms from bacteria to man. It functions, for example, as a phosphate and energy store, and is involved in the activation and regulation of proteins. Despite its ubiquitous occurrence and important functions, it is unclear how poly P is synthesized or how poly P metabolism is regulated in higher eukaryotes. This work describes a systematic analysis of poly P levels in yeast knockout strains mutated in almost every non-essential gene. Results After three consecutive screens, 255 genes (almost 4% of the yeast genome) were found to be involved in the maintenance of normal poly P content. Many of these genes encoded proteins functioning in the cytoplasm, the vacuole or in transport and transcription. Besides reduced poly P content, many strains also exhibited reduced total phosphate content, showed altered ATP and glycogen levels and were disturbed in the secretion of acid phosphatase. Conclusion Cellular energy and phosphate homeostasis is suggested to result from the equilibrium between poly P, ATP and free phosphate within the cell. Poly P serves as a buffer for both ATP and free phosphate levels and is, therefore, the least essential and consequently most variable component in this network. However, strains with reduced poly P levels are not only affected in their ATP and phosphate content, but also in other components that depend on ATP or free phosphate content, such as glycogen or secreted phosphatase activity. PMID:17107617

  4. DNA repair and synthetic lethality

    PubMed Central

    Guo, Gong-she; Zhang, Feng-mei; Gao, Rui-jie; Delsite, Robert; Feng, Zhi-hui; Powell, Simon N

    2011-01-01

    Tumors often have DNA repair defects, suggesting additional inhibition of other DNA repair pathways in tumors may lead to synthetic lethality. Accumulating data demonstrate that DNA repair-defective tumors, in particular homologous recombination (HR), are highly sensitive to DNA-damaging agents. Thus, HR-defective tumors exhibit potential vulnerability to the synthetic lethality approach, which may lead to new therapeutic strategies. It is well known that poly (adenosine diphosphate (ADP)-ribose) polymerase (PARP) inhibitors show the synthetically lethal effect in tumors defective in BRCA1 or BRCA2 genes encoded proteins that are required for efficient HR. In this review, we summarize the strategies of targeting DNA repair pathways and other DNA metabolic functions to cause synthetic lethality in HR-defective tumor cells. PMID:22010575

  5. Syn-Lethality: An Integrative Knowledge Base of Synthetic Lethality towards Discovery of Selective Anticancer Therapies

    PubMed Central

    Li, Xue-juan; Mishra, Shital K.; Wu, Min; Zhang, Fan

    2014-01-01

    Synthetic lethality (SL) is a novel strategy for anticancer therapies, whereby mutations of two genes will kill a cell but mutation of a single gene will not. Therefore, a cancer-specific mutation combined with a drug-induced mutation, if they have SL interactions, will selectively kill cancer cells. While numerous SL interactions have been identified in yeast, only a few have been known in human. There is a pressing need to systematically discover and understand SL interactions specific to human cancer. In this paper, we present Syn-Lethality, the first integrative knowledge base of SL that is dedicated to human cancer. It integrates experimentally discovered and verified human SL gene pairs into a network, associated with annotations of gene function, pathway, and molecular mechanisms. It also includes yeast SL genes from high-throughput screenings which are mapped to orthologous human genes. Such an integrative knowledge base, organized as a relational database with user interface for searching and network visualization, will greatly expedite the discovery of novel anticancer drug targets based on synthetic lethality interactions. The database can be downloaded as a stand-alone Java application. PMID:24864230

  6. Syn-lethality: an integrative knowledge base of synthetic lethality towards discovery of selective anticancer therapies.

    PubMed

    Li, Xue-juan; Mishra, Shital K; Wu, Min; Zhang, Fan; Zheng, Jie

    2014-01-01

    Synthetic lethality (SL) is a novel strategy for anticancer therapies, whereby mutations of two genes will kill a cell but mutation of a single gene will not. Therefore, a cancer-specific mutation combined with a drug-induced mutation, if they have SL interactions, will selectively kill cancer cells. While numerous SL interactions have been identified in yeast, only a few have been known in human. There is a pressing need to systematically discover and understand SL interactions specific to human cancer. In this paper, we present Syn-Lethality, the first integrative knowledge base of SL that is dedicated to human cancer. It integrates experimentally discovered and verified human SL gene pairs into a network, associated with annotations of gene function, pathway, and molecular mechanisms. It also includes yeast SL genes from high-throughput screenings which are mapped to orthologous human genes. Such an integrative knowledge base, organized as a relational database with user interface for searching and network visualization, will greatly expedite the discovery of novel anticancer drug targets based on synthetic lethality interactions. The database can be downloaded as a stand-alone Java application. PMID:24864230

  7. A Screen of Coxiella burnetii Mutants Reveals Important Roles for Dot/Icm Effectors and Host Autophagy in Vacuole Biogenesis

    PubMed Central

    Newton, Hayley J.; Kohler, Lara J.; McDonough, Justin A.; Temoche-Diaz, Morayma; Crabill, Emerson; Hartland, Elizabeth L.; Roy, Craig R.

    2014-01-01

    Coxiella burnetii is an intracellular pathogen that replicates in a lysosome-derived vacuole. The molecular mechanisms used by this bacterium to create a pathogen-occupied vacuole remain largely unknown. Here, we conducted a visual screen on an arrayed library of C. burnetii NMII transposon insertion mutants to identify genes required for biogenesis of a mature Coxiella-containing vacuole (CCV). Mutants defective in Dot/Icm secretion system function or the PmrAB regulatory system were incapable of intracellular replication. Several mutants with intracellular growth defects were found to have insertions in genes encoding effector proteins translocated into host cells by the Dot/Icm system. These included mutants deficient in the effector proteins Cig57, CoxCC8 and Cbu1754. Mutants that had transposon insertions in genes important in central metabolism or encoding tRNA modification enzymes were identified based on the appearance filamentous bacteria intracellularly. Lastly, mutants that displayed a multi-vacuolar phenotype were identified. All of these mutants had a transposon insertion in the gene encoding the effector protein Cig2. Whereas vacuoles containing wild type C. burnetii displayed robust accumulation of the autophagosome protein LC3, the vacuoles formed by the cig2 mutant did not contain detectible amounts of LC3. Furthermore, interfering with host autophagy during infection by wild type C. burnetii resulted in a multi-vacuolar phenotype similar to that displayed by the cig2 mutant. Thus, a functional Cig2 protein is important for interactions between the CCV and host autophagosomes and this drives a process that enhances the fusogenic properties of this pathogen-occupied organelle. PMID:25080348

  8. An RNAi screen for Aire cofactors reveals a role for Hnrnpl in polymerase release and Aire-activated ectopic transcription

    PubMed Central

    Giraud, Matthieu; Jmari, Nada; Du, Lina; Carallis, Floriane; Nieland, Thomas J. F.; Perez-Campo, Flor M.; Bensaude, Olivier; Root, David E.; Hacohen, Nir; Mathis, Diane; Benoist, Christophe

    2014-01-01

    Aire induces the expression of a large set of autoantigen genes in the thymus, driving immunological tolerance in maturing T cells. To determine the full spectrum of molecular mechanisms underlying the Aire transactivation function, we screened an AIRE-dependent gene-expression system with a genome-scale lentiviral shRNA library, targeting factors associated with chromatin architecture/function, transcription, and mRNA processing. Fifty-one functional allies were identified, with a preponderance of factors that impact transcriptional elongation compared with initiation, in particular members of the positive transcription elongation factor b (P-TEFb) involved in the release of “paused” RNA polymerases (CCNT2 and HEXIM1); mRNA processing and polyadenylation factors were also highlighted (HNRNPL/F, SFRS1, SFRS3, and CLP1). Aire’s functional allies were validated on transfected and endogenous target genes, including the generation of lentigenic knockdown (KD) mice. We uncovered the effect of the splicing factor Hnrnpl on Aire-induced transcription. Transcripts sensitive to the P-TEFb inhibitor flavopiridol were reduced by Hnrnpl knockdown in thymic epithelial cells, independently of their dependence on Aire, therefore indicating a general effect of Hnrnpl on RNA elongation. This conclusion was substantiated by demonstration of HNRNPL interactions with P-TEFb components (CDK9, CCNT2, HEXIM1, and the small 7SK RNA). Aire-containing complexes include 7SK RNA, the latter interaction disrupted by HNRNPL knockdown, suggesting that HNRNPL may partake in delivering inactive P-TEFb to Aire. Thus, these results indicate that mRNA processing factors cooperate with Aire to release stalled polymerases and to activate ectopic expression of autoantigen genes in the thymus. PMID:24434558

  9. Functional centrality: detecting lethality of proteins in protein interaction networks.

    PubMed

    Tew, Kar Leong; Li, Xiao-Li; Tan, Soon-Heng

    2007-01-01

    Identifying lethal proteins is important for understanding the intricate mechanism governing life. Researchers have shown that the lethality of a protein can be computed based on its topological position in the protein-protein interaction (PPI) network. Performance of current approaches has been less than satisfactory as the lethality of a protein is a functional characteristic that cannot be determined solely by network topology. Furthermore, a significant number of lethal proteins have low connectivity in the interaction networks but are overlooked by most current methods. Our work reveals that a protein's lethality correlates more strongly with its "functional centrality" than pure topological centrality. We define functional centrality as the topological centrality within a subnetwork of proteins with similar functions. Evaluation experiments on four Saccharomyces cerevisiae PPI datasets showed that NFC performed significantly better than all the other existing computational techniques. Our method was able to detect low connectivity lethal proteins that were previously undetected by conventional methods. The results and an online version of NFC is available at http://lethalproteins.i2r.a-star.edu.sg. PMID:18546514

  10. In silico screening reveals structurally diverse, nanomolar inhibitors of NQO2 that are functionally active in cells and can modulate NF?B signalling

    PubMed Central

    Nolan, Karen A.; Dunstan, Mark S.; Caraher, Mary C.; Scott, Katherine A.; Leys, David; Stratford, Ian J.

    2011-01-01

    The NCI chemical database has been screened using in silico docking to identify novel nanomolar inhibitors of NRH:quinone oxidoreductase 2 (NQO2). The inhibitors identified from the screen exhibit a diverse range of scaffolds and the structure of one of the inhibitors, NSC13000 co-crystalized with NQO2, has been solved. This has been used to aid the generation of a structure/activity relationship between the computationally derived binding affinity and experimentally measured enzyme inhibitory potency. Many of the compounds are functionally active as inhibitors of NQO2 in cells at non toxic concentrations. To demonstrate this, advantage was taken of the NQO2-mediated toxicity of the chemotherapeutic drug CB1954. The toxicity of this drug is substantially reduced when the function of NQO2 is inhibited and many of the compounds achieve this in cells at nanomolar concentrations. The NQO2 inhibitors also attenuated TNF?-mediated, NF?B-driven transcriptional activity. The link between NQO2 and the regulation of NF?B was confirmed by using siRNA to NQO2 and by the observation that NRH, the cofactor for NQO2 enzyme activity, could regulate NF?B activity in an NQO2 dependent manner. NF?B is a potential therapeutic target and this study reveals an underlying mechanism that may exploitable for developing new anti-cancer drugs. PMID:22090421

  11. NMR-spectroscopic screening of spider venom reveals sulfated nucleosides as major components for the brown recluse and related species

    PubMed Central

    Schroeder, Frank C.; Taggi, Andrew E.; Gronquist, Matthew; Malik, Rabia U.; Grant, Jacqualine B.; Eisner, Thomas; Meinwald, Jerrold

    2008-01-01

    Extensive chemical analyses of spider venoms from many species have revealed complex mixtures of biologically active compounds, of which several have provided important leads for drug development. We have recently shown that NMR spectroscopy can be used advantageously for a direct structural characterization of the small-molecule content of such complex mixtures. Here, we report the application of this strategy to a larger-scale analysis of a collection of spider venoms representing >70 species, which, in combination with mass spectrometric analyses, allowed the identification of a wide range of known, and several previously undescribed, small molecules. These include polyamines, common neurotransmitters, and amino acid derivatives as well as two additional members of a recently discovered family of natural products, the sulfated nucleosides. In the case of the well studied brown recluse spider, Loxosceles reclusa, sulfated guanosine derivatives were found to comprise the major small-molecule components of the venom. PMID:18794518

  12. Screening of Metagenomic and Genomic Libraries Reveals Three Classes of Bacterial Enzymes That Overcome the Toxicity of Acrylate

    PubMed Central

    Curson, Andrew R. J.; Burns, Oliver J.; Voget, Sonja; Daniel, Rolf; Todd, Jonathan D.; McInnis, Kathryn; Wexler, Margaret; Johnston, Andrew W. B.

    2014-01-01

    Acrylate is produced in significant quantities through the microbial cleavage of the highly abundant marine osmoprotectant dimethylsulfoniopropionate, an important process in the marine sulfur cycle. Acrylate can inhibit bacterial growth, likely through its conversion to the highly toxic molecule acrylyl-CoA. Previous work identified an acrylyl-CoA reductase, encoded by the gene acuI, as being important for conferring on bacteria the ability to grow in the presence of acrylate. However, some bacteria lack acuI, and, conversely, many bacteria that may not encounter acrylate in their regular environments do contain this gene. We therefore sought to identify new genes that might confer tolerance to acrylate. To do this, we used functional screening of metagenomic and genomic libraries to identify novel genes that corrected an E. coli mutant that was defective in acuI, and was therefore hyper-sensitive to acrylate. The metagenomic libraries yielded two types of genes that overcame this toxicity. The majority encoded enzymes resembling AcuI, but with significant sequence divergence among each other and previously ratified AcuI enzymes. One other metagenomic gene, arkA, had very close relatives in Bacillus and related bacteria, and is predicted to encode an enoyl-acyl carrier protein reductase, in the same family as FabK, which catalyses the final step in fatty-acid biosynthesis in some pathogenic Firmicute bacteria. A genomic library of Novosphingobium, a metabolically versatile alphaproteobacterium that lacks both acuI and arkA, yielded vutD and vutE, two genes that, together, conferred acrylate resistance. These encode sequential steps in the oxidative catabolism of valine in a pathway in which, significantly, methacrylyl-CoA is a toxic intermediate. These findings expand the range of bacteria for which the acuI gene encodes a functional acrylyl-CoA reductase, and also identify novel enzymes that can similarly function in conferring acrylate resistance, likely, again, through the removal of the toxic product acrylyl-CoA. PMID:24848004

  13. Antibacterial and brine shrimp lethality effect of marine actinobacterium Streptomyces sp. CAS72 against human pathogenic bacteria

    PubMed Central

    Sivasankar, Palaniappan; Manivasagan, Panchanathan; Vijayanand, Packiyaraj; Sivakumar, Kannan; Sugesh, Shanmugam; Poongodi, Subramaniam; Maharani, Viswanathan; Vijayalakshmi, Shanmugam; Balasubramanian, Thangavel

    2013-01-01

    Objective To investigate the in vitro antibacterial activity against human pathogenic bacteria and brine shrimp lethality bioassay of the marine actinobacterium. Methods Forty six marine actinobacterial strains were isolated from sediment samples of Uppanar estuary, Cuddalore, India. Preliminary screening was done by cross-streak method and the potential strain was identified by morphological, chemotaxonomical and molecular methods. Fermentation was done and the metabolite was obtained by liquid-liquid extraction using ethyl acetate and purified by silica gel (100-200 mesh) column chromatography. The purified metabolite was tested for antibacterial activity, minimal inhibitory concentration and brine shrimp lethality bioassay. Results Among the forty six strains, CAS72 was found effective against human pathogenic bacteria. The strain CAS72 was identified as Streptomyces sp. The purified metabolite exhibited a significant in vitro antibacterial activity. The MIC value was also determined against human pathogenic bacteria and a strong cytotoxic activity in brine shrimp lethality assay was observed and the LC50 value was 23.5 µg/mL. Conclusions The present investigation reveals that the marine actinobacteria are well obtainable in Uppanar estuary environment and it can provide a definite source for novel bioactive metabolites.

  14. Virtual and In Vitro Screens Reveal a Potential Pharmacophore that Avoids the Fibrillization of A?1–42

    PubMed Central

    Hernández-Rodríguez, Maricarmen; Correa-Basurto, José; Nicolás-Vázquez, María Inés; Miranda-Ruvalcaba, René; Benítez-Cardoza, Claudia Guadalupe; Reséndiz-Albor, Aldo Arturo; Méndez-Méndez, Juan Vicente; Rosales-Hernández, Martha C.

    2015-01-01

    Among the multiple factors that induce Alzheimer’s disease, aggregation of the amyloid ? peptide (A?) is considered the most important due to the ability of the 42-amino acid A? peptides (A?1–42) to form oligomers and fibrils, which constitute A? pathological aggregates. For this reason, the development of inhibitors of A?1–42 pathological aggregation represents a field of research interest. Several A?1–42 fibrillization inhibitors possess tertiary amine and aromatic moieties. In the present study, we selected 26 compounds containing tertiary amine and aromatic moieties with or without substituents and performed theoretical studies that allowed us to select four compounds according to their free energy values for A?1–42 in ?-helix (A?-?), random coil (A?-RC) and ?-sheet (A?-?) conformations. Docking studies revealed that compound 5 had a higher affinity for A?-? and A?-RC than the other compounds. In vitro, this compound was able to abolish Thioflavin T fluorescence and favored an RC conformation of A?1–42 in circular dichroism studies, resulting in the formation of amorphous aggregates as shown by atomic force microscopy. The results obtained from quantum studies allowed us to identify a possible pharmacophore that can be used to design A?1–42 aggregation inhibitors. In conclusion, compounds with higher affinity for A?-? and A?-RC prevented the formation of oligomeric species. PMID:26172152

  15. Complete genome-wide screening and subtractive genomic approach revealed new virulence factors, potential drug targets against bio-war pathogen Brucella melitensis 16M

    PubMed Central

    Pradeepkiran, Jangampalli Adi; Sainath, Sri Bhashyam; Kumar, Konidala Kranthi; Bhaskar, Matcha

    2015-01-01

    Brucella melitensis 16M is a Gram-negative coccobacillus that infects both animals and humans. It causes a disease known as brucellosis, which is characterized by acute febrile illness in humans and causes abortions in livestock. To prevent and control brucellosis, identification of putative drug targets is crucial. The present study aimed to identify drug targets in B. melitensis 16M by using a subtractive genomic approach. We used available database repositories (Database of Essential Genes, Kyoto Encyclopedia of Genes and Genomes Automatic Annotation Server, and Kyoto Encyclopedia of Genes and Genomes) to identify putative genes that are nonhomologous to humans and essential for pathogen B. melitensis 16M. The results revealed that among 3 Mb genome size of pathogen, 53 putative characterized and 13 uncharacterized hypothetical genes were identified; further, from Basic Local Alignment Search Tool protein analysis, one hypothetical protein showed a close resemblance (50%) to Silicibacter pomeroyi DUF1285 family protein (2RE3). A further homology model of the target was constructed using MODELLER 9.12 and optimized through variable target function method by molecular dynamics optimization with simulating annealing. The stereochemical quality of the restrained model was evaluated by PROCHECK, VERIFY-3D, ERRAT, and WHATIF servers. Furthermore, structure-based virtual screening was carried out against the predicted active site of the respective protein using the glycerol structural analogs from the PubChem database. We identified five best inhibitors with strong affinities, stable interactions, and also with reliable drug-like properties. Hence, these leads might be used as the most effective inhibitors of modeled protein. The outcome of the present work of virtual screening of putative gene targets might facilitate design of potential drugs for better treatment against brucellosis. PMID:25834405

  16. A drug resistance screen using a selective MET inhibitor reveals a spectrum of mutations that partially overlap with activating mutations found in cancer patients.

    PubMed

    Tiedt, Ralph; Degenkolbe, Elisa; Furet, Pascal; Appleton, Brent A; Wagner, Sabrina; Schoepfer, Joseph; Buck, Emily; Ruddy, David A; Monahan, John E; Jones, Michael D; Blank, Jutta; Haasen, Dorothea; Drueckes, Peter; Wartmann, Markus; McCarthy, Clive; Sellers, William R; Hofmann, Francesco

    2011-08-01

    The emergence of drug resistance is a primary concern in any cancer treatment, including with targeted kinase inhibitors as exemplified by the appearance of Bcr-Abl point mutations in chronic myeloid leukemia (CML) patients treated with imatinib. In vitro approaches to identify resistance mutations in Bcr-Abl have yielded mutation spectra that faithfully recapitulated clinical observations. To predict resistance mutations in the receptor tyrosine kinase MET that could emerge during inhibitor treatment in patients, we conducted a resistance screen in BaF3 TPR-MET cells using the novel selective MET inhibitor NVP-BVU972. The observed spectrum of mutations in resistant cells was dominated by substitutions of tyrosine 1230 but also included other missense mutations and partially overlapped with activating MET mutations that were previously described in cancer patients. Cocrystallization of the MET kinase domain in complex with NVP-BVU972 revealed a key role for Y1230 in binding of NVP-BVU972, as previously reported for multiple other selective MET inhibitors. A second resistance screen in the same format with the MET inhibitor AMG 458 yielded a distinct spectrum of mutations rich in F1200 alterations, which is consistent with a different predicted binding mode. Our findings suggest that amino acid substitutions in the MET kinase domain of cancer patients need to be carefully monitored before and during treatment with MET inhibitors, as resistance may preexist or emerge. Compounds binding in the same manner as NVP-BVU972 might be particularly susceptible to the development of resistance through mutations in Y1230, a condition that may be addressed by MET inhibitors with alternative binding modes. PMID:21697284

  17. Complete genome-wide screening and subtractive genomic approach revealed new virulence factors, potential drug targets against bio-war pathogen Brucella melitensis 16M.

    PubMed

    Pradeepkiran, Jangampalli Adi; Sainath, Sri Bhashyam; Kumar, Konidala Kranthi; Bhaskar, Matcha

    2015-01-01

    Brucella melitensis 16M is a Gram-negative coccobacillus that infects both animals and humans. It causes a disease known as brucellosis, which is characterized by acute febrile illness in humans and causes abortions in livestock. To prevent and control brucellosis, identification of putative drug targets is crucial. The present study aimed to identify drug targets in B. melitensis 16M by using a subtractive genomic approach. We used available database repositories (Database of Essential Genes, Kyoto Encyclopedia of Genes and Genomes Automatic Annotation Server, and Kyoto Encyclopedia of Genes and Genomes) to identify putative genes that are nonhomologous to humans and essential for pathogen B. melitensis 16M. The results revealed that among 3 Mb genome size of pathogen, 53 putative characterized and 13 uncharacterized hypothetical genes were identified; further, from Basic Local Alignment Search Tool protein analysis, one hypothetical protein showed a close resemblance (50%) to Silicibacter pomeroyi DUF1285 family protein (2RE3). A further homology model of the target was constructed using MODELLER 9.12 and optimized through variable target function method by molecular dynamics optimization with simulating annealing. The stereochemical quality of the restrained model was evaluated by PROCHECK, VERIFY-3D, ERRAT, and WHATIF servers. Furthermore, structure-based virtual screening was carried out against the predicted active site of the respective protein using the glycerol structural analogs from the PubChem database. We identified five best inhibitors with strong affinities, stable interactions, and also with reliable drug-like properties. Hence, these leads might be used as the most effective inhibitors of modeled protein. The outcome of the present work of virtual screening of putative gene targets might facilitate design of potential drugs for better treatment against brucellosis. PMID:25834405

  18. The effects of anthrax lethal toxin on host barrier function.

    PubMed

    Xie, Tao; Auth, Roger D; Frucht, David M

    2011-06-01

    The pathological actions of anthrax toxin require the activities of its edema factor (EF) and lethal factor (LF) enzyme components, which gain intracellular access via its receptor-binding component, protective antigen (PA). LF is a metalloproteinase with specificity for selected mitogen-activated protein kinase kinases (MKKs), but its activity is not directly lethal to many types of primary and transformed cells in vitro. Nevertheless, in vivo treatment of several animal species with the combination of LF and PA (termed lethal toxin or LT) leads to morbidity and mortality, suggesting that LT-dependent toxicity is mediated by cellular interactions between host cells. Decades of research have revealed that a central hallmark of this toxicity is the disruption of key cellular barriers required to maintain homeostasis. This review will focus on the current understanding of the effects of LT on barrier function, highlighting recent progress in establishing the molecular mechanisms underlying these effects. PMID:22069727

  19. Arenavirus extinction through lethal mutagenesis

    Microsoft Academic Search

    Juan Carlos de la Torre

    2005-01-01

    Viral hemorrhagic fevers represent serious human public health problems causing devastating and often lethal disease. Several hemorrhagic fevers are caused by arenaviruses including Lassa fever virus (LFV) and the South American viral hemorrhagic fevers (SAHF). In recent years, increased air travel between Africa and other areas has led to the importation of LFV into the US, Europe, Japan, and Canada.

  20. A trans-Amazonian screening of mtDNA reveals deep intraspecific divergence in forest birds and suggests a vast underestimation of species diversity.

    PubMed

    Milá, Borja; Tavares, Erika S; Muñoz Saldaña, Alberto; Karubian, Jordan; Smith, Thomas B; Baker, Allan J

    2012-01-01

    The Amazonian avifauna remains severely understudied relative to that of the temperate zone, and its species richness is thought to be underestimated by current taxonomy. Recent molecular systematic studies using mtDNA sequence reveal that traditionally accepted species-level taxa often conceal genetically divergent subspecific lineages found to represent new species upon close taxonomic scrutiny, suggesting that intraspecific mtDNA variation could be useful in species discovery. Surveys of mtDNA variation in Holarctic species have revealed patterns of variation that are largely congruent with species boundaries. However, little information exists on intraspecific divergence in most Amazonian species. Here we screen intraspecific mtDNA genetic variation in 41 Amazonian forest understory species belonging to 36 genera and 17 families in 6 orders, using 758 individual samples from Ecuador and French Guiana. For 13 of these species, we also analyzed trans-Andean populations from the Ecuadorian Chocó. A consistent pattern of deep intraspecific divergence among trans-Amazonian haplogroups was found for 33 of the 41 taxa, and genetic differentiation and genetic diversity among them was highly variable, suggesting a complex range of evolutionary histories. Mean sequence divergence within families was the same as that found in North American birds (13%), yet mean intraspecific divergence in Neotropical species was an order of magnitude larger (2.13% vs. 0.23%), with mean distance between intraspecific lineages reaching 3.56%. We found no clear relationship between genetic distances and differentiation in plumage color. Our results identify numerous genetically and phenotypically divergent lineages which may result in new species-level designations upon closer taxonomic scrutiny and thorough sampling, although lineages in the tropical region could be older than those in the temperate zone without necessarily representing separate species. In-depth phylogeographic surveys are urgently needed to avoid underestimating tropical diversity, and the use of mtDNA markers can be instrumental in identifying and prioritizing taxa for species discovery. PMID:22815761

  1. Cultivation-Independent Screening Revealed Hot Spots of IncP-1, IncP-7 and IncP-9 Plasmid Occurrence in Different Environmental Habitats

    PubMed Central

    Dealtry, Simone; Ding, Guo-Chun; Weichelt, Viola; Dunon, Vincent; Schlüter, Andreas; Martini, María Carla; Papa, María Florencia Del; Lagares, Antonio; Amos, Gregory Charles Auton; Wellington, Elizabeth Margaret Helen; Gaze, William Hugo; Sipkema, Detmer; Sjöling, Sara; Springael, Dirk; Heuer, Holger; van Elsas, Jan Dirk; Thomas, Christopher; Smalla, Kornelia

    2014-01-01

    IncP-1, IncP-7 and IncP-9 plasmids often carry genes encoding enzymes involved in the degradation of man-made and natural contaminants, thus contributing to bacterial survival in polluted environments. However, the lack of suitable molecular tools often limits the detection of these plasmids in the environment. In this study, PCR followed by Southern blot hybridization detected the presence of plasmid-specific sequences in total community (TC-) DNA or fosmid DNA from samples originating from different environments and geographic regions. A novel primer system targeting IncP-9 plasmids was developed and applied along with established primers for IncP-1 and IncP-7. Screening TC-DNA from biopurification systems (BPS) which are used on farms for the purification of pesticide-contaminated water revealed high abundances of IncP-1 plasmids belonging to different subgroups as well as IncP-7 and IncP-9. The novel IncP-9 primer-system targeting the rep gene of nine IncP-9 subgroups allowed the detection of a high diversity of IncP-9 plasmid specific sequences in environments with different sources of pollution. Thus polluted sites are “hot spots” of plasmids potentially carrying catabolic genes. PMID:24587126

  2. Cultivation-independent screening revealed hot spots of IncP-1, IncP-7 and IncP-9 plasmid occurrence in different environmental habitats.

    PubMed

    Dealtry, Simone; Ding, Guo-Chun; Weichelt, Viola; Dunon, Vincent; Schlüter, Andreas; Martini, María Carla; Del Papa, María Florencia; Lagares, Antonio; Amos, Gregory Charles Auton; Wellington, Elizabeth Margaret Helen; Gaze, William Hugo; Sipkema, Detmer; Sjöling, Sara; Springael, Dirk; Heuer, Holger; van Elsas, Jan Dirk; Thomas, Christopher; Smalla, Kornelia

    2014-01-01

    IncP-1, IncP-7 and IncP-9 plasmids often carry genes encoding enzymes involved in the degradation of man-made and natural contaminants, thus contributing to bacterial survival in polluted environments. However, the lack of suitable molecular tools often limits the detection of these plasmids in the environment. In this study, PCR followed by Southern blot hybridization detected the presence of plasmid-specific sequences in total community (TC-) DNA or fosmid DNA from samples originating from different environments and geographic regions. A novel primer system targeting IncP-9 plasmids was developed and applied along with established primers for IncP-1 and IncP-7. Screening TC-DNA from biopurification systems (BPS) which are used on farms for the purification of pesticide-contaminated water revealed high abundances of IncP-1 plasmids belonging to different subgroups as well as IncP-7 and IncP-9. The novel IncP-9 primer-system targeting the rep gene of nine IncP-9 subgroups allowed the detection of a high diversity of IncP-9 plasmid specific sequences in environments with different sources of pollution. Thus polluted sites are "hot spots" of plasmids potentially carrying catabolic genes. PMID:24587126

  3. Identification of cetrimonium bromide and irinotecan as compounds with synthetic lethality against NDRG1 deficient prostate cancer cells

    PubMed Central

    Wissing, Michel D.; Mendonca, Janet; Kim, Eunice; Kim, Eugene; Shim, Joong S.; Kaelber, Nadine S.; Kant, Huub; Hammers, Hans; Commes, Therese; Van Diest, Paul J.; Liu, Jun O.; Kachhap, Sushant K.

    2013-01-01

    The N-myc downstream regulated gene 1 (NDRG1) has been identified as a metastasis-suppressor gene in prostate cancer (PCa). Compounds targeting PCa cells deficient in NDRG1 could potentially decrease invasion/metastasis of PCa. A cell based screening strategy was employed to identify small molecules that selectively target NDRG1 deficient PCa cells. DU-145 PCa cells rendered deficient in NDRG1 expression by a lentiviral shRNA-mediated knockdown strategy were used in the primary screen. Compounds filtered from the primary screen were further validated through proliferation and clonogenic survival assays in parental and NDRG1 knockdown PCa cells. Screening of 3360 compounds revealed irinotecan and cetrimonium bromide (CTAB) as compounds that exhibited synthetic lethality against NDRG1 deficient PCa cells. A three-dimensional (3-D) invasion assay was utilized to test the ability of CTAB to inhibit invasion of DU-145 cells. CTAB was found to remarkably decrease invasion of DU-145 cells in collagen matrix. Our results suggest that CTAB and irinotecan could be further explored for their potential clinical benefit in patients with NDRG1 deficient PCa. PMID:23377825

  4. Harnessing synthetic lethal interactions in anticancer drug discovery

    PubMed Central

    Chan, Denise A.; Giaccia, Amato J.

    2013-01-01

    Unique features of tumours that can be exploited by targeted therapies are a key focus of current cancer research. One such approach is known as synthetic lethality screening, which involves searching for genetic interactions of two mutations whereby the presence of either mutation alone has no effect on cell viability but the combination of the two mutations results in cell death. The presence of one of these mutations in cancer cells but not in normal cells can therefore create opportunities to selectively kill cancer cells by mimicking the effect of the second genetic mutation with targeted therapy. Here, we summarize strategies that can be used to identify synthetic lethal interactions for anticancer drug discovery, describe examples of such interactions that are currently being investigated in preclinical and clinical studies of targeted anticancer therapies, and discuss the challenges of realizing the full potential of such therapies. PMID:21532565

  5. Introduction to Lethal School Violence

    Microsoft Academic Search

    Jeffrey A. Daniels; Mary C. Bradley

    \\u000a In this chapter, we offer an introduction to the topic of the book, lethal school violence (LSV). We begin with an introduction\\u000a to and definition of LSV, and then highlight five different situations that often result in fatalities (i.e., suicide, rampage\\u000a shootings, gang-related deaths, domestic murder\\/suicide that occurs on campus, and barricaded captive events). We then turn\\u000a our attention to

  6. Early events of lethal action by tobramycin in Pseudomonas aeruginosa

    SciTech Connect

    Raulston, J.E.

    1988-01-01

    The immediate activities of the aminoglycoside antibiotic, tobramycin, were investigated in Pseudomonas aeruginosa PAO1. The influence of carbon growth substate and the antibiotic exposure environment in the magnitude of activity were examined. Lethality by 8 {mu}g/ml tobramycin occurred rapidly (1 to 3 minutes). The release of specific cellular components into the supernatant was associated with lethality. This material was initially detected as an increase in UV-absorbance. Magnesium in the reaction mixture provided protection against lethality and leakage, but did not reverse lethal damage after a 3 minute tobramycin treatment. Also, uptake of {sup 3}H-tobramycin was reduced in the presence of magnesium. Cells grown with glucose as a carbon source were more susceptible than organic acid grown cells as was the rapidity and amount of cell damage. Analyses of the leakage material revealed a 2-fold increase of protein in the supernatant after a 1-3 minute treatment which paralleled lethality. A prominent 29 kDa protein was observed by SDS-PAGE in the released material, which has been identified as the periplasmic enzyme, {beta}-lactamase. The immediate activities of tobramycin did not involve (i) release of overall cell protein, (ii) massive loss of total pool amino acids, (iii) cell lysis, (iv) inhibition of proline uptake, (v) release of lipopolysaccharide, or (vi) leakage of ATP. Electron microscopy showed no apparent damage after a 3 minute exposure. 40% inhibition of protein synthesis had occurred by 3 minutes of exposure, while release of UV-absorbing material and lethality were detectable after only 1 minute. Resistant cystic fibrosis isolates of P. aeruginosa did not leak under the same experimental conditions, but one of two susceptible strains examined did show increased UV-absorbance following treatment.

  7. The Genetic Analysis of Snf: A Drosophila Sex Determination Gene Required for Activation of Sex-Lethal in Both the Germline and the Soma

    PubMed Central

    Salz, H. K.

    1992-01-01

    Our analysis demonstrates that snf is a positive regulator of Sex-lethal in both the germline and the soma. In the germline, unregulated expression of Sex-lethal can bypass the requirement for snf(+) gene function, implying that snf is required for Sex-lethal activity in the germline. This conclusion is supported by the finding that the Sex-lethal transcription pattern is abnormal in a snf mutant background. In the soma, activation of Sex-lethal appears to be sensitive to snf gene dosage only when the probability of Sex-lethal activation has been otherwise reduced. We also show that the activity of one of the constitutive Sex-lethal alleles (Sxl(M1)) is sensitive to snf gene dosage, demonstrating that, in spite of its constitutive behavior in some assays, Sxl(M1) is still subject to some regulation. In spite of snf's role in the somatic activation of Sex-lethal, no lethal alleles of snf were isolated in a screen of ~25,000 chromosomes. The observation that the existing snf mutations present a lethal phenotype only in certain genetic backgrounds suggests that snf is required, but is not essential, for the activation of Sex-lethal in the soma. In contrast, snf does appear to be essential for activation of Sex-lethal in the germline, as evidenced by its female-sterile phenotype. PMID:1551576

  8. Ethical language and decision-making for prenatally diagnosed lethal malformations.

    PubMed

    Wilkinson, Dominic; de Crespigny, Lachlan; Xafis, Vicki

    2014-10-01

    In clinical practice, and in the medical literature, severe congenital malformations such as trisomy 18, anencephaly, and renal agenesis are frequently referred to as 'lethal' or as 'incompatible with life'. However, there is no agreement about a definition of lethal malformations, nor which conditions should be included in this category. Review of outcomes for malformations commonly designated 'lethal' reveals that prolonged survival is possible, even if rare. This article analyses the concept of lethal malformations and compares it to the problematic concept of 'futility'. We recommend avoiding the term 'lethal' and suggest that counseling should focus on salient prognostic features instead. For conditions with a high chance of early death or profound impairment in survivors despite treatment, perinatal and neonatal palliative care would be ethical. However, active obstetric and neonatal management, if desired, may also sometimes be appropriate. PMID:25200733

  9. Ethical language and decision-making for prenatally diagnosed lethal malformations

    PubMed Central

    Wilkinson, Dominic; de Crespigny, Lachlan; Xafis, Vicki

    2014-01-01

    Summary In clinical practice, and in the medical literature, severe congenital malformations such as trisomy 18, anencephaly, and renal agenesis are frequently referred to as ‘lethal’ or as ‘incompatible with life’. However, there is no agreement about a definition of lethal malformations, nor which conditions should be included in this category. Review of outcomes for malformations commonly designated ‘lethalreveals that prolonged survival is possible, even if rare. This article analyses the concept of lethal malformations and compares it to the problematic concept of ‘futility’. We recommend avoiding the term ‘lethal’ and suggest that counseling should focus on salient prognostic features instead. For conditions with a high chance of early death or profound impairment in survivors despite treatment, perinatal and neonatal palliative care would be ethical. However, active obstetric and neonatal management, if desired, may also sometimes be appropriate. PMID:25200733

  10. Acute and sub-lethal response to mercury in Arctic and boreal calanoid copepods.

    PubMed

    Overjordet, Ida Beathe; Altin, Dag; Berg, Torunn; Jenssen, Bjørn Munro; Gabrielsen, Geir Wing; Hansen, Bjørn Henrik

    2014-10-01

    Acute lethal toxicity, expressed as LC50 values, is a widely used parameter in risk assessment of chemicals, and has been proposed as a tool to assess differences in species sensitivities to chemicals between climatic regions. Arctic Calanus glacialis and boreal Calanus finmarchicus were exposed to mercury (Hg(2+)) under natural environmental conditions including sea temperatures of 2° and 10°C, respectively. Acute lethal toxicity (96 h LC50) and sub-lethal molecular response (GST expression; in this article gene expression is used as a synonym of gene transcription, although it is acknowledged that gene expression is also regulated, e.g., at translation and protein stability level) were studied. The acute lethal toxicity was monitored for 96 h using seven different Hg concentrations. The sub-lethal experiment was set up on the basis of nominal LC50 values for each species using concentrations equivalent to 50, 5 and 0.5% of their 96 h LC50 value. No significant differences were found in acute lethal toxicity between the two species. The sub-lethal molecular response revealed large differences both in response time and the fold induction of GST, where the Arctic species responded both faster and with higher mRNA levels of GST after 48 h exposure. Under the natural exposure conditions applied in the present study, the Arctic species C. glacialis may potentially be more susceptible to mercury exposure on the sub-lethal level. PMID:25036619

  11. Tasers--less than lethal!

    PubMed

    Sharma, Abiram; Theivacumar, Nada S; Souka, Hesham M

    2009-05-01

    We report a case of potentially lethal injury associated with the use of Taser. A 42-year-old man was stopped by police for potential detention. He held a large carving knife over his epigasrium threatening to stab himself. With a view to achieving immobilisation, a Taser gun was used. On activation of the Taser, the subject suffered a 7-cm wide and 10-cm deep stab injury to the upper abdomen. In this case, activation of the Taser resulted in the contraction of skeletal muscles, flexors more intensely than extensors, resulting in the stab injury. PMID:19416583

  12. Lethal mitochondrial cardiomyopathy in a hypomorphic Med30 mouse mutant is ameliorated by ketogenic diet

    PubMed Central

    Krebs, Philippe; Fan, Weiwei; Chen, Yen-Hui; Tobita, Kimimasa; Downes, Michael R.; Wood, Malcolm R.; Sun, Lei; Xia, Yu; Ding, Ning; Spaeth, Jason M.; Moresco, Eva Marie Y.; Boyer, Thomas G.; Lo, Cecilia Wen Ya; Yen, Jeffrey; Evans, Ronald M.; Beutler, Bruce

    2011-01-01

    Deficiencies of subunits of the transcriptional regulatory complex Mediator generally result in embryonic lethality, precluding study of its physiological function. Here we describe a missense mutation in Med30 causing progressive cardiomyopathy in homozygous mice that, although viable during lactation, show precipitous lethality 2–3 wk after weaning. Expression profiling reveals pleiotropic changes in transcription of cardiac genes required for oxidative phosphorylation and mitochondrial integrity. Weaning mice to a ketogenic diet extends viability to 8.5 wk. Thus, we establish a mechanistic connection between Mediator and induction of a metabolic program for oxidative phosphorylation and fatty acid oxidation, in which lethal cardiomyopathy is mitigated by dietary intervention. PMID:22106289

  13. Electroshock weapons can be lethal!

    NASA Astrophysics Data System (ADS)

    Lundquist, Marjorie

    2008-03-01

    Electroshock weapons (EWs)-stun guns, tasers, riot shields-are electroconductive devices designed to safely incapacitate healthy men neuromuscularly, so they are called nonlethal or less-lethal. EW firms seeking large nonmilitary markets targeted law enforcement and corrections personnel, who began using EWs in prisons/jails and on public patrol in 1980 in the USA. This shifted the EW-shocked population from healthy soldiers to a heterogeneous mix of both sexes, ages 6-92, in a wide variety of health conditions! An EW operates by disrupting normal physiological processes, producing transient effects in healthy people. But if a person's health is sufficiently compromised, the margin of safety can be lost, resulting in death or permanent health problems. 325 people have died after EW shock since 1980. Did the EW cause these deaths? Evidence indicates that EWs do play a causal role in most such deaths. EWs can be lethal for people in diabetic shock^1 (hypoglycemia), which may be why Robert Dziekanski-a Polish immigrant to Canada-died so quickly after he was tasered at Vancouver Airport: not having eaten for over 10 hours, he likely was severely hypoglycemic. The EW death rate in North America is 30 times higher than need be, because EW users have not been properly trained to use EWs on a heterogeneous population safely! ^1J. Clinical Engineering 30(3):111(2005).

  14. Lethality and Entropy of Protein Interaction Networks

    Microsoft Academic Search

    Thomas Manke; Lloyd Demetrius; Martin Vingron

    Abstract We characterize protein interaction networks in terms of network entropy. This approach sug- gests a ranking principle, which strongly correlates with elements of functional importance, such as lethal proteins. Our combined analysis of protein interaction networks and functional proflles in single cellular yeast and mulit-cellular worm shows that proteins with large contribution to net- work entropy are preferentially lethal.

  15. Crisis Intervention with Highly Lethal Suicidal People.

    ERIC Educational Resources Information Center

    Leenaars, Antoon A.

    1994-01-01

    Outlines model for crisis intervention with highly lethal suicidal people. Explores idea that crisis is a perception, including issues of lethality and perturbation, object relations, responsibility, weapon availability, and active versus passive response. Highlights specific problems with transference and countertransference. Suggests that there…

  16. Alcohol Consumption and Nearly Lethal Suicide Attempts.

    ERIC Educational Resources Information Center

    Powell, Kenneth E.; Kresnow, Marcie-jo; Mercy, James A.; Potter, Lloyd B.; Swann, Alan C.; Frankowski, Ralph F.; Lee, Roberta K.; Bayer, Timothy L.

    2002-01-01

    Presents a case-control study of the association between nearly lethal suicide attempts and facets of alcohol consumption; namely, drinking frequency, drinking quantity, binge drinking, alcoholism, drinking within 3 hours of suicide attempt, and age began drinking. In bivariate analyses, all measures were associated with nearly lethal suicide…

  17. Highly variable recessive lethal or nearly lethal mutation rates during germ-line development of male Drosophila melanogaster

    PubMed Central

    Gao, Jian-Jun; Pan, Xue-Rong; Hu, Jing; Ma, Li; Wu, Jian-Min; Shao, Ye-Lin; Barton, Sara A.; Woodruff, Ronny C.; Zhang, Ya-Ping; Fu, Yun-Xin

    2011-01-01

    Each cell of higher organism adults is derived from a fertilized egg through a series of divisions, during which mutations can occur. Both the rate and timing of mutations can have profound impacts on both the individual and the population, because mutations that occur at early cell divisions will affect more tissues and are more likely to be transferred to the next generation. Using large-scale multigeneration screening experiments for recessive lethal or nearly lethal mutations of Drosophila melanogaster and recently developed statistical analysis, we show for male D. melanogaster that (i) mutation rates (for recessive lethal or nearly lethal) are highly variable during germ cell development; (ii) first cell cleavage has the highest mutation rate, which drops substantially in the second cleavage or the next few cleavages; (iii) the intermediate stages, after a few cleavages to right before spermatogenesis, have at least an order of magnitude smaller mutation rate; and (iv) spermatogenesis also harbors a fairly high mutation rate. Because germ-line lineage shares some (early) cell divisions with somatic cell lineage, the first conclusion is readily extended to a somatic cell lineage. It is conceivable that the first conclusion is true for most (if not all) higher organisms, whereas the other three conclusions are widely applicable, although the extent may differ from species to species. Therefore, conclusions or analyses that are based on equal mutation rates during development should be taken with caution. Furthermore, the statistical approach developed can be adopted for studying other organisms, including the human germ-line or somatic mutational patterns. PMID:21890796

  18. Yeast Three-Hybrid Screening of Rous Sarcoma Virus Mutants with Randomly Mutagenized Minimal Packaging Signals Reveals Regions Important for Gag Interactions

    Microsoft Academic Search

    EUN-GYUNG LEE; MAXINE L. LINIAL

    2000-01-01

    We previously showed that the yeast three-hybrid system provides a genetic assay of both RNA and protein components for avian retroviral RNA encapsidation. In the current study, we used this assay to precisely define cis-acting determinants involved in avian leukosis sarcoma virus packaging RNA binding to Gag protein. In vivo screening of Rous sarcoma virus mutants was performed with randomly

  19. Comparative analysis of RNAi screening technologies at genome-scale reveals an inherent processing inefficiency of the plasmid-based shRNA hairpin.

    PubMed

    Bhinder, Bhavneet; Shum, David; Djaballah, Hakim

    2014-02-01

    RNAi screening in combination with the genome-sequencing projects would constitute the Holy Grail of modern genetics; enabling discovery and validation towards a better understanding of fundamental biology leading to novel targets to combat disease. Hit discordance at inter-screen level together with the lack of reproducibility is emerging as the technology's main pitfalls. To examine some of the underlining factors leading to such discrepancies, we reasoned that perhaps there is an inherent difference in knockdown efficiency of the various RNAi technologies. For this purpose, we utilized the two most popular ones, chemically synthesized siRNA duplex and plasmid-based shRNA hairpin, in order to perform a head to head comparison. Using a previously developed gain-of-function assay probing modulators of the miRNA biogenesis pathway, we first executed on a siRNA screen against the Silencer Select V4.0 library (AMB) nominating 1,273, followed by an shRNA screen against the TRC1 library (TRC1) nominating 497 gene candidates. We observed a poor overlap of only 29 hits given that there are 15,068 overlapping genes between the two libraries; with DROSHA as the only common hit out of the seven known core miRNA biogenesis genes. Distinct genes interacting with the same biogenesis regulators were observed in both screens, with a dismal cross-network overlap of only 3 genes (DROSHA, TGFBR1, and DIS3). Taken together, our study demonstrates differential knockdown activities between the two technologies, possibly due to the inefficient intracellular processing and potential cell-type specificity determinants in generating intended targeting sequences for the plasmid-based shRNA hairpins; and suggests this observed inefficiency as potential culprit in addressing the lack of reproducibility. PMID:24433414

  20. Comparative analysis of RNAi screening technologies at genome-scale reveals an inherent processing inefficiency of the plasmid-based shRNA hairpin

    PubMed Central

    Bhinder, Bhavneet; Shum, David; Djaballah, Hakim

    2014-01-01

    RNAi screening in combination with the genome-sequencing projects would constitute the Holy Grail of modern genetics; enabling discovery and validation towards a better understanding of fundamental biology leading to novel targets to combat disease. Hit discordance at inter-screen level together with the lack of reproducibility is emerging as the technology's main pitfalls. To examine some of the underlining factors leading to such discrepancies, we reasoned that perhaps there is an inherent difference in knockdown efficiency of the various RNAi technologies. For this purpose, we utilized the two most popular ones, chemically synthesized siRNA duplex and plasmid-based shRNA hairpin, in order to perform a head to head comparison. Using a previously developed gain-of-function assay probing modulators of the miRNA biogenesis pathway, we first executed on a siRNA screen against the Silencer Select V4.0 library (AMB) nominating 1,273, followed by an shRNA screen against the TRC1 library (TRC1) nominating 497 gene candidates. We observed a poor overlap of only 29 hits given that there are 15,068 overlapping genes between the two libraries; with DROSHA as the only common hit out of the seven known core miRNA biogenesis genes. Distinct genes interacting with the same biogenesis regulators were observed in both screens, with a dismal cross-network overlap of only 3 genes (DROSHA, TGFBR1, and DIS3). Taken together, our study demonstrates differential knockdown activities between the two technologies, possibly due to the inefficient intracellular processing and potential cell-type specificity determinants in generating intended targeting sequences for the plasmid-based shRNA hairpins; and suggests this observed inefficiency as potential culprit in addressing the lack of reproducibility. PMID:24433414

  1. Lethal mutagenesis failure may augment viral adaptation.

    PubMed

    Paff, Matthew L; Stolte, Steven P; Bull, James J

    2014-01-01

    Lethal mutagenesis, the attempt to extinguish a population by elevating its mutation rate, has been endorsed in the virology literature as a promising approach for treating viral infections. In support of the concept, in vitro studies have forced viral extinction with high doses of mutagenic drugs. However, the one known mutagenic drug used on patients commonly fails to cure infections, and in vitro studies typically find a wide range of mutagenic conditions permissive for viral growth. A key question becomes how subsequent evolution is affected if the viral population is mutated but avoids extinction--Is viral adaptation augmented rather than suppressed? Here we consider the evolution of highly mutated populations surviving mutagenesis, using the DNA phage T7. In assays using inhibitory hosts, whenever resistance mutants were observed, the mutagenized populations exhibited higher frequencies, but some inhibitors blocked plaque formation by even the mutagenized stock. Second, outgrowth of previously mutagenized populations led to rapid and potentially complete fitness recovery but polymorphism was slow to decay, and mutations exhibited inconsistent patterns of change. Third, the combination of population bottlenecks with mutagenesis did cause fitness declines, revealing a vulnerability that was not apparent from mutagenesis of large populations. The results show that a population surviving high mutagenesis may exhibit enhanced adaptation in some environments and experience little negative fitness consequences in many others. PMID:24092771

  2. Lethal Mutagenesis Failure May Augment Viral Adaptation

    PubMed Central

    Paff, Matthew L.; Stolte, Steven P.; Bull, James J.

    2014-01-01

    Lethal mutagenesis, the attempt to extinguish a population by elevating its mutation rate, has been endorsed in the virology literature as a promising approach for treating viral infections. In support of the concept, in vitro studies have forced viral extinction with high doses of mutagenic drugs. However, the one known mutagenic drug used on patients commonly fails to cure infections, and in vitro studies typically find a wide range of mutagenic conditions permissive for viral growth. A key question becomes how subsequent evolution is affected if the viral population is mutated but avoids extinction—Is viral adaptation augmented rather than suppressed? Here we consider the evolution of highly mutated populations surviving mutagenesis, using the DNA phage T7. In assays using inhibitory hosts, whenever resistance mutants were observed, the mutagenized populations exhibited higher frequencies, but some inhibitors blocked plaque formation by even the mutagenized stock. Second, outgrowth of previously mutagenized populations led to rapid and potentially complete fitness recovery but polymorphism was slow to decay, and mutations exhibited inconsistent patterns of change. Third, the combination of population bottlenecks with mutagenesis did cause fitness declines, revealing a vulnerability that was not apparent from mutagenesis of large populations. The results show that a population surviving high mutagenesis may exhibit enhanced adaptation in some environments and experience little negative fitness consequences in many others. PMID:24092771

  3. Lethal photosensitization of Helicobacter species

    NASA Astrophysics Data System (ADS)

    Millson, Charles E.; Wilson, Michael; MacRobert, Alexander J.; Thurrell, Wendy; Mlkvy, Peter; Davies, Claire; Bown, Stephen G.

    1995-01-01

    Helicobacter pylori (H. pylori) is associated with a large number of gastroduodenal disorders. Clearance of the bacteria has been shown to benefit patients with duodenal ulcers, gastric ulcers, and certain rare types of gastric tumors. Broad-spectrum antibiotics are the mainstay of current treatment strategies but side-effects, poor compliance, and drug resistance limit their usefulness. We sensitized H. pylori with toluidine blue, haematoporphyrin derivative, aluminum disulphonated phthalocyanine, methylene blue or protoporphyrin IX prior to exposure to low-power laser light from either a gallium aluminum arsenide laser or a helium neon gas laser. All 5 sensitizers caused reductions of greater than 1000-fold in the number of viable bacteria. Light alone had no effect and only HpD caused a significant decrease in bacterial numbers without laser light. Next, we sensitized H. mustelae on explanted ferret gastric mucosa (ex vivo) with the same sensitizers and exposed them to light from a copper vapor pumped dye laser tuned appropriately. MB caused significant reductions in bacterial counts. Successful lethal photosensitization of Helicobacter pylori both in vitro and ex vivo raises the possibility of a local method for eradicating the bacteria, especially as the bacteria are only found in those parts of the upper gastrointestinal tract that are accessible to the endoscope.

  4. Structural Basis for a Lethal Mutation in U6 RNA†,‡

    PubMed Central

    Sashital, Dipali G.; Allmann, Anne M.; Van Doren, Steven R.; Butcher, Samuel E.

    2011-01-01

    U6 RNA is essential for nuclear pre-mRNA splicing and has been implicated directly in catalysis of intron removal. The U80G mutation at the essential magnesium binding site of the U6 3? intramolecular stem–loop region (ISL) is lethal in yeast. To further understand the structure and function of the U6 ISL, we have investigated the structural basis for the lethal U80G mutation by NMR and optical spectroscopy. The NMR structure reveals that the U80G mutation causes a structural rearrangement within the ISL resulting in the formation of a new Watson–Crick base pair (C67·G80), and disrupts a protonated C67·A79 wobble pair that forms in the wild-type structure. Despite the structural change, the accessibility of the metal binding site is unperturbed, and cadmium titration produces similar phosphorus chemical shift changes for both the U80G mutant and wild-type RNAs. The thermodynamic stability of the U80G mutant is significantly increased (??Gfold = ?3.6 ± 1.9 kcal/mol), consistent with formation of the Watson–Crick pair. Our structural and thermodynamic data, in combination with previous genetic data, suggest that the lethal basis for the U80G mutation is stem–loop hyperstabilization. This hyperstabilization may prevent the U6 ISL melting and rearrangement necessary for association with U4. PMID:12578359

  5. Crystallographic studies of the Anthrax lethal toxin. Annual report

    SciTech Connect

    Frederick, C.A.

    1996-07-01

    The lethal form of Anthrax results from the inhalation of anthrax spores. Death is primarily due to the effects of the lethal toxin (Protective Antigen (PA) + Lethal Factor) from the causative agent, Bacillus anthracis. All the Anthrax vaccines currently in use or under development contain or produce PA, the major antigenic component of anthrax toxin, and there is a clear need for an improved vaccine for human use. In the previous report we described the first atomic resolution structure of PA, revealing that the molecule is composed largely of beta-sheets organized into four domains. This information can be used in the design. of recombinant PA vaccines. In this report we describe additional features of the full-length PA molecule derived from further crystallographic refinement and careful examination of the structure. We compare two crystal forms of PA grown at different pH values and discuss the functional implications. A complete definition of the function of each domain must await the crystal structure of the PA63 heptamer. We have grown crystals of the heptamer under both detergent and detergent-free conditions, and made substantial progress towards the crystal structure. The mechanism of anthrax intoxication in the light of our results is reviewed.

  6. Reaming experiments for the lethality test system

    SciTech Connect

    Hooten, D.; Stanley, P.

    1988-01-01

    Various reaming techniques were tried for use on the barrel of the Lethality Test System railgun. This report covers the successes and failures of the reamers and the techniques that were tried. 5 figs.

  7. [Synthetic lethal genes to mutant p53].

    PubMed

    Tongyang, Liu; Haiqiang, Guo; Meiyan, Zhu; Yingze, Huang; Shuting, Jia; Ying, Luo; Jihong, Zhang

    2015-04-01

    Targeted therapy has become a powerful approach for cancer treatment. Better understanding of oncogenes as well as synthetic lethal interactions with oncogenes will lead to new strategies for tumor-specific treatment. It is well known that mutant p53 plays an important role in tumorigenesis and tumor development. Thus, understanding the synthetic lethal relationship between p53 mutations and interacting genes in tumor is critical for the personalized treatments of p53 mutant tumors. Synthetic lethal genes to mutant p53 can be divided into cell cycle regulators and non-cell cycle regulators. This paper review show these two types of target genes contribute to synthetic lethal interactions with p53 mutations and potential applications of these interactions in anticancer therapy. PMID:25881697

  8. A high-throughput screen for ligand binding reveals the specificities of three amino acid chemoreceptors from Pseudomonas syringae pv. actinidiae.

    PubMed

    McKellar, James L O; Minnell, Jordan J; Gerth, Monica L

    2015-05-01

    Chemoreceptors play a central role in chemotaxis, allowing bacteria to detect chemical gradients and bias their swimming behavior in order to navigate toward favorable environments. The genome of the kiwifruit pathogen, Pseudomonas syringae pv. actinidiae (Psa) strain NZ-V13 encodes 43 predicted chemoreceptors, none of which has been characterized. We developed a high-throughput fluorescence-based thermal shift assay for identifying the signal molecules that are recognized by a given chemoreceptor ligand binding domain (LBD). Using this assay, we characterized the ligand binding profiles of three Psa homologs of the P.?aeruginosa?PAO1 amino acid chemoreceptors PctA, PctB and PctC. Each recombinant LBD was screened against 95 potential ligands. The three Psa homologs, named pscA, pscB and pscC (Psa chemoreceptors A, B and C) bound 3, 10 and 3 amino acids respectively. In each case, their binding profiles were distinct from their P.?aeruginosa?PAO1 homologs. Notably, Psa?PscA-LBD only bound the acidic amino acids l-aspartate, d-aspartate and l-glutamate, whereas P.?aeruginosa?PctA-LBD binds all of the l-proteinogenic amino acids except for l-aspartate and l-glutamate. A combination of homology modeling, site-directed mutagenesis and functional screening identified a single amino acid residue in the Psa?PscA-LBD (Ala146) that is critically important for determining its narrow specificity. PMID:25656450

  9. Genome-wide microRNA screening reveals that the evolutionary conserved miR-9a regulates body growth by targeting sNPFR1/NPYR.

    PubMed

    Suh, Yoon Seok; Bhat, Shreelatha; Hong, Seung-Hyun; Shin, Minjung; Bahk, Suhyoung; Cho, Kyung Sang; Kim, Seung-Whan; Lee, Kyu-Sun; Kim, Young-Joon; Jones, Walton D; Yu, Kweon

    2015-01-01

    MicroRNAs (miRNAs) regulate many physiological processes including body growth. Insulin/IGF signalling is the primary regulator of animal body growth, but the extent to which miRNAs act in insulin-producing cells (IPCs) is unclear. Here we generate a UAS-miRNA library of Drosophila stocks and perform a genetic screen to identify miRNAs whose overexpression in the IPCs inhibits body growth in Drosophila. Through this screen, we identify miR-9a as an evolutionarily conserved regulator of insulin signalling and body growth. IPC-specific miR-9a overexpression reduces insulin signalling and body size. Of the predicted targets of miR-9a, we find that loss of miR-9a enhances the level of sNPFR1. We show via an in vitro binding assay that miR-9a binds to sNPFR1 mRNA in insect cells and to the mammalian orthologue NPY2R in rat insulinoma cells. These findings indicate that the conserved miR-9a regulates body growth by controlling sNPFR1/NPYR-mediated modulation of insulin signalling. PMID:26138755

  10. Temperature-Sensitive Cell-Lethal Mutants of Drosophila: Isolation and Characterization

    PubMed Central

    Arking, Robert

    1975-01-01

    One hundred and twenty-one temperature-sensitive (ts) sex-linked lethals were screened by means of X-ray-induced somatic crossing over to determine if any were ts cell-lethal mutants. Cell-lethal mutations were identified by their ability to block the development of homozygous clones when raised under restrictive conditions (29°). Twenty-two ts cell-lethal mutants were isolated and categorized into three classes, depending upon the patterns of damage observed in larval and imaginal tissues. The phenotypes produced by these mutations ranged from those which affected only a limited set of structures (i.e., genital discs only) to those which affected diverse tissues at all stages of the life cycle. Each mutation has its own characteristic time-dependent pattern, frequency, and type of damage. All the mutations affect imaginal tissue, but only one-third of the mutations affect both larval and imaginal tissue. The fastest-acting lethals need 15 hours at the restrictive temperature to kill the cells and the slowest-acting lethals require at least 48 hours. By choosing the appropriate mutant and by manipulating the times of exposure to the restrictive temperature, it has proven possible to produce duplications and deficiencies in specific structures of the adult. A mechanism by which lethality might yield such structures is suggested. In addition, 15 of the mutants are ts female sterile mutants. Only one of these 15 mutants can recover its fertility when shifted back down to the permissive temperature (22°). PMID:821813

  11. High content image-based screening of a protease inhibitor library reveals compounds broadly active against Rift Valley fever virus and other highly pathogenic RNA viruses.

    PubMed

    Mudhasani, Rajini; Kota, Krishna P; Retterer, Cary; Tran, Julie P; Whitehouse, Chris A; Bavari, Sina

    2014-08-01

    High content image-based screening was developed as an approach to test a protease inhibitor small molecule library for antiviral activity against Rift Valley fever virus (RVFV) and to determine their mechanism of action. RVFV is the causative agent of severe disease of humans and animals throughout Africa and the Arabian Peninsula. Of the 849 compounds screened, 34 compounds exhibited ? 50% inhibition against RVFV. All of the hit compounds could be classified into 4 distinct groups based on their unique chemical backbone. Some of the compounds also showed broad antiviral activity against several highly pathogenic RNA viruses including Ebola, Marburg, Venezuela equine encephalitis, and Lassa viruses. Four hit compounds (C795-0925, D011-2120, F694-1532 and G202-0362), which were most active against RVFV and showed broad-spectrum antiviral activity, were selected for further evaluation for their cytotoxicity, dose response profile, and mode of action using classical virological methods and high-content imaging analysis. Time-of-addition assays in RVFV infections suggested that D011-2120 and G202-0362 targeted virus egress, while C795-0925 and F694-1532 inhibited virus replication. We showed that D011-2120 exhibited its antiviral effects by blocking microtubule polymerization, thereby disrupting the Golgi complex and inhibiting viral trafficking to the plasma membrane during virus egress. While G202-0362 also affected virus egress, it appears to do so by a different mechanism, namely by blocking virus budding from the trans Golgi. F694-1532 inhibited viral replication, but also appeared to inhibit overall cellular gene expression. However, G202-0362 and C795-0925 did not alter any of the morphological features that we examined and thus may prove to be good candidates for antiviral drug development. Overall this work demonstrates that high-content image analysis can be used to screen chemical libraries for new antivirals and to determine their mechanism of action and any possible deleterious effects on host cellular biology. PMID:25144302

  12. Lethal potency and fractionation of Duvernoy's secretion from the brown tree snake, Boiga irregularis.

    PubMed

    Weinstein, S A; Chiszar, D; Bell, R C; Smith, L A

    1991-01-01

    The liquid secretion contained only 15% protein and had relatively low proteolytic activity. The reconstituted crude secretion had a murine i.p. LD50 of 10.33 mg/kg and was not hemorrhagic in doses up to 200 micrograms. Fast Protein Liquid Chromatographic (FPLC) cation exchange analysis of reconstituted crude secretion resulted in resolution of 16 peaks. Lethal activity was identified in three peaks. The major lethal fraction was 12.5% of the secretion protein and had a murine i.p. LD50 of 7.3 mg/kg. A pooled fraction containing two lethal peaks which comprised 9.4% of secretion protein had moderate proteolytic activity and produced myoglobinuria in mice. The fraction had an approximate murine i.p. LD50 of 3.7 mg/kg. Microscopic examination of muscle tissue from mice succumbing to this fraction revealed multifocal myofiber degeneration and necrosis. SDS-PAGE indicated that the major lethal fraction contained three proteins with mol. wts of 12,500, 18,000 and 52,000 and the myotoxic fraction contained two proteins with mol. wts of 14,500 and 17,000. While B. irregularis Duvernoy's secretion has a low lethal index, it does contain a myotoxic fraction with moderate lethal potency. These observations and recent data describing clinical envenomation of several infant patients suggest that large specimens may pose a hazard to infants and small children. PMID:1862518

  13. Functional screening in Drosophila reveals the conserved role of REEP1 in promoting stress resistance and preventing the formation of Tau aggregates.

    PubMed

    Appocher, Chiara; Klima, Raffaella; Feiguin, Fabian

    2014-12-20

    Pathological modifications in the microtubule-associated protein Tau is a common characteristic observed in different neurological diseases, suggesting that analogous metabolic pathways might be similarly affected during neurodegeneration. To identify these molecules and mechanisms, we utilized Drosophila models of human Tau-mediated neurodegeneration to perform an RNA interference functional screening against genes considered to be implicated in the pathogenesis of different neurodegenerative disorders. We found that the downregulation of the Drosophila REEP1 homolog protein enhanced Tau toxicity with increased formation of insoluble aggregates. On the contrary, the overexpression of either the Drosophila or the human REEP1 protein was able to revert these phenotypes and promote neuronal resistance to ER stress. These studies identify a new function for the REEP1 protein in vivo and a novel cellular mechanism to prevent Tau toxicity. PMID:25096240

  14. Functional Screening of the Cronobacter sakazakii BAA-894 Genome reveals a role for ProP (ESA_02131) in carnitine uptake.

    PubMed

    Feeney, Audrey; Sleator, Roy D

    2015-05-01

    Cronobacter sakazakii is a neonatal pathogen responsible for up to 80% of fatalities in infected infants. Low birth weight infants and neonates infected with C. sakazakii suffer necrotizing enterocolitis, bacteraemia and meningitis. The mode of transmission most often associated with infection is powdered infant formula (PIF) which, with an aw of ?0.2, is too low to allow most microorganisms to persist. Survival of C. sakazakii in environments subject to extreme hyperosmotic stress has previously been attributed to the uptake of compatible solutes including proline and betaine. Herein, we report the construction and screening of a C. sakazakii genome bank and the identification of ProP (ESA_02131) as a carnitine uptake system. PMID:25915804

  15. Identification and classification of genes required for tolerance to high-sucrose stress revealed by genome-wide screening of Saccharomyces cerevisiae.

    PubMed

    Ando, Akira; Tanaka, Fumiko; Murata, Yoshinori; Takagi, Hiroshi; Shima, Jun

    2006-03-01

    Yeasts used in bread making are exposed to high concentrations of sucrose during sweet dough fermentation. Despite its importance, tolerance to high-sucrose stress is poorly understood at the gene level. To clarify the genes required for tolerance to high-sucrose stress, genome-wide screening was undertaken using the complete deletion strain collection of diploid Saccharomyces cerevisiae. The screening identified 273 deletions that yielded high sucrose sensitivity, approximately 20 of which were previously uncharacterized. These 273 deleted genes were classified based on their cellular function and localization of their gene products. Cross-sensitivity of the high-sucrose-sensitive mutants to high concentrations of NaCl and sorbitol was studied. Among the 273 sucrose-sensitive deletion mutants, 269 showed cross-sensitivities to sorbitol or NaCl, and four (i.e. ade5,7, ade6, ade8, and pde2) were specifically sensitive to high sucrose. The general stress response pathways via high-osmolarity glycerol and stress response element pathways and the function of the invertase in the ade mutants were similar to those in the wild-type strain. In the presence of high-sucrose stress, intracellular contents of ATP in ade mutants were at least twofold lower than that of the wild-type cells, suggesting that depletion of ATP is a factor in sensitivity to high-sucrose stress. The genes identified in this study might be important for tolerance to high-sucrose stress, and therefore should be target genes in future research into molecular modification for breeding of yeast tolerant to high-sucrose stress. PMID:16487347

  16. Functional RNAi screen targeting cytokine and growth factor receptors reveals oncorequisite role for interleukin-2 gamma receptor in JAK3-mutation-positive leukemia.

    PubMed

    Agarwal, A; MacKenzie, R J; Eide, C A; Davare, M A; Watanabe-Smith, K; Tognon, C E; Mongoue-Tchokote, S; Park, B; Braziel, R M; Tyner, J W; Druker, B J

    2015-06-01

    To understand the role of cytokine and growth factor receptor-mediated signaling in leukemia pathogenesis, we designed a functional RNA interference (RNAi) screen targeting 188 cytokine and growth factor receptors that we found highly expressed in primary leukemia specimens. Using this screen, we identified interleukin-2 gamma receptor (IL2R?) as a critical growth determinant for a JAK3(A572V) mutation-positive acute myeloid leukemia cell line. We observed that knockdown of IL2R? abrogates phosphorylation of JAK3 and downstream signaling molecules, JAK1, STAT5, MAPK and pS6 ribosomal protein. Overexpression of IL2R? in murine cells increased the transforming potential of activating JAK3 mutations, whereas absence of IL2R? completely abrogated the clonogenic potential of JAK3(A572V), as well as the transforming potential of additional JAK3-activating mutations such as JAK3(M511I). In addition, mutation at the IL2R? interaction site in the FERM domain of JAK3 (Y100C) completely abrogated JAK3-mediated leukemic transformation. Mechanistically, we found IL2R? contributes to constitutive JAK3 mutant signaling by increasing JAK3 expression and phosphorylation. Conversely, we found that mutant, but not wild-type JAK3, increased the expression of IL2R?, indicating IL2R? and JAK3 contribute to constitutive JAK/STAT signaling through their reciprocal regulation. Overall, we demonstrate a novel role for IL2R? in potentiating oncogenesis in the setting of JAK3-mutation-positive leukemia. In addition, our study highlights an RNAi-based functional assay that can be used to facilitate the identification of non-kinase cytokine and growth factor receptor targets for inhibiting leukemic cell growth. PMID:25109334

  17. A High-Throughput Fatty Acid Profiling Screen Reveals Novel Variations in Fatty Acid Biosynthesis in Chlamydomonas reinhardtii and Related Algae

    PubMed Central

    Pflaster, Erin L.; Schwabe, Michael J.; Becker, Joyanne; Wilkinson, Melissa S.; Parmer, Ashley; Clemente, Thomas E.; Cahoon, Edgar B.

    2014-01-01

    Analysis of fatty acid methyl esters (FAMEs) by gas chromatography (GC) is a common technique for the quantitative and qualitative analysis of acyl lipids. Methods for FAME preparation are typically time-consuming and labor-intensive and require multiple transfers of reagents and products between reaction tubes and autosampler vials. In order to increase throughput and lower the time and materials costs required for FAME preparation prior to GC analysis, we have developed a method in which 10-to-20-mg samples of microbial biomass are transferred to standard GC autosampler vials, transesterified using an emulsion of methanolic trimethylsulfonium hydroxide and hexane, and analyzed directly by GC without further sample handling. This method gives results that are essentially identical to those obtained by the more labor- and material-intensive FAME preparation methods, such as transmethylation with methanolic HCl. We applied this method to the screening of laboratory and environmental isolates of the green alga Chlamydomonas for variations in fatty acid composition. This screening method facilitated two novel discoveries. First, we identified a common laboratory strain of C. reinhardtii, CC-620, completely lacking all ?-3 fatty acids normally found in this organism and showed that this strain contains an inactivating mutation in the CrFAD7 gene, encoding the sole ?-3 desaturase activity in this organism. Second, we showed that some species of Chlamydomonas make ?6-unsaturated polyunsaturated fatty acids (PUFA) rather than the ?5 species normally made by the previously characterized laboratory strains of Chlamydomonas, suggesting that there is species-specific variation in the regiospecificity and substrate selectivity of front-end desaturases in this algal genus. PMID:25239975

  18. Drug Repurposing Screen Reveals FDA-Approved Inhibitors of Human HMG-CoA Reductase and Isoprenoid Synthesis That Block Cryptosporidium parvum Growth

    PubMed Central

    Bessoff, Kovi; Sateriale, Adam; Lee, K. Kyungae

    2013-01-01

    Cryptosporidiosis, a diarrheal disease usually caused by Cryptosporidium parvum or Cryptosporidium hominis in humans, can result in fulminant diarrhea and death in AIDS patients and chronic infection and stunting in children. Nitazoxanide, the current standard of care, has limited efficacy in children and is no more effective than placebo in patients with advanced AIDS. Unfortunately, the lack of financial incentives and the technical difficulties associated with working with Cryptosporidium parasites have crippled efforts to develop effective treatments. In order to address these obstacles, we developed and validated (Z? score = 0.21 to 0.47) a cell-based high-throughput assay and screened a library of drug repurposing candidates (the NIH Clinical Collections), with the hopes of identifying safe, FDA-approved drugs to treat cryptosporidiosis. Our screen yielded 21 compounds with confirmed activity against C. parvum growth at concentrations of <10 ?M, many of which had well-defined mechanisms of action, making them useful tools to study basic biology in addition to being potential therapeutics. Additional work, including structure-activity relationship studies, identified the human 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitor itavastatin as a potent inhibitor of C. parvum growth (50% inhibitory concentration [IC50] = 0.62 ?M). Bioinformatic analysis of the Cryptosporidium genomes indicated that the parasites lack all known enzymes required for the synthesis of isoprenoid precursors. Additionally, itavastatin-induced growth inhibition of C. parvum was partially reversed by the addition of exogenous isopentenyl pyrophosphate, suggesting that itavastatin reduces Cryptosporidium growth via on-target inhibition of host HMG-CoA reductase and that the parasite is dependent on the host cell for synthesis of isoprenoid precursors. PMID:23380723

  19. Drug repurposing screen reveals FDA-approved inhibitors of human HMG-CoA reductase and isoprenoid synthesis that block Cryptosporidium parvum growth.

    PubMed

    Bessoff, Kovi; Sateriale, Adam; Lee, K Kyungae; Huston, Christopher D

    2013-04-01

    Cryptosporidiosis, a diarrheal disease usually caused by Cryptosporidium parvum or Cryptosporidium hominis in humans, can result in fulminant diarrhea and death in AIDS patients and chronic infection and stunting in children. Nitazoxanide, the current standard of care, has limited efficacy in children and is no more effective than placebo in patients with advanced AIDS. Unfortunately, the lack of financial incentives and the technical difficulties associated with working with Cryptosporidium parasites have crippled efforts to develop effective treatments. In order to address these obstacles, we developed and validated (Z' score = 0.21 to 0.47) a cell-based high-throughput assay and screened a library of drug repurposing candidates (the NIH Clinical Collections), with the hopes of identifying safe, FDA-approved drugs to treat cryptosporidiosis. Our screen yielded 21 compounds with confirmed activity against C. parvum growth at concentrations of <10 ?M, many of which had well-defined mechanisms of action, making them useful tools to study basic biology in addition to being potential therapeutics. Additional work, including structure-activity relationship studies, identified the human 3-hydroxy-3-methyl-glutaryl-coenzyme A (HMG-CoA) reductase inhibitor itavastatin as a potent inhibitor of C. parvum growth (50% inhibitory concentration [IC(50)] = 0.62 ?M). Bioinformatic analysis of the Cryptosporidium genomes indicated that the parasites lack all known enzymes required for the synthesis of isoprenoid precursors. Additionally, itavastatin-induced growth inhibition of C. parvum was partially reversed by the addition of exogenous isopentenyl pyrophosphate, suggesting that itavastatin reduces Cryptosporidium growth via on-target inhibition of host HMG-CoA reductase and that the parasite is dependent on the host cell for synthesis of isoprenoid precursors. PMID:23380723

  20. Screening for Cutaneous Melanoma by Skin Self-Examination

    Microsoft Academic Search

    Marianne Berwick; Colin B. Begg; Judith A. Fine; George C. Roush; Raymond L. Barnhill

    1996-01-01

    Background: Although some evidence indicates that early detection protects against the development of lethal melanoma, no randomized clinical trials have been con- ducted to measure the efficacy of early detection (or screen- ing) in preventing death from this disease. Since melanoma incidence in the United States is relatively rare, a ran- domized clinical trial to test the efficacy of screening

  1. Preparation and characterization of cobalt-substituted anthrax lethal factor

    SciTech Connect

    Saebel, Crystal E.; Carbone, Ryan; Dabous, John R.; Lo, Suet Y. [Department of Chemistry and Biochemistry, Laurentian University, 935 Ramsey Lake Rd., Sudbury, Ontario, Canada P3E 2C6 (Canada)] [Department of Chemistry and Biochemistry, Laurentian University, 935 Ramsey Lake Rd., Sudbury, Ontario, Canada P3E 2C6 (Canada); Siemann, Stefan, E-mail: ssiemann@laurentian.ca [Department of Chemistry and Biochemistry, Laurentian University, 935 Ramsey Lake Rd., Sudbury, Ontario, Canada P3E 2C6 (Canada)] [Department of Chemistry and Biochemistry, Laurentian University, 935 Ramsey Lake Rd., Sudbury, Ontario, Canada P3E 2C6 (Canada)

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer Cobalt-substituted anthrax lethal factor (CoLF) is highly active. Black-Right-Pointing-Pointer CoLF can be prepared by bio-assimilation and direct exchange. Black-Right-Pointing-Pointer Lethal factor binds cobalt tightly. Black-Right-Pointing-Pointer The electronic spectrum of CoLF reveals penta-coordination. Black-Right-Pointing-Pointer Interaction of CoLF with thioglycolic acid follows a 2-step mechanism. -- Abstract: Anthrax lethal factor (LF) is a zinc-dependent endopeptidase involved in the cleavage of mitogen-activated protein kinase kinases near their N-termini. The current report concerns the preparation of cobalt-substituted LF (CoLF) and its characterization by electronic spectroscopy. Two strategies to produce CoLF were explored, including (i) a bio-assimilation approach involving the cultivation of LF-expressing Bacillus megaterium cells in the presence of CoCl{sub 2}, and (ii) direct exchange by treatment of zinc-LF with CoCl{sub 2}. Independent of the method employed, the protein was found to contain one Co{sup 2+} per LF molecule, and was shown to be twice as active as its native zinc counterpart. The electronic spectrum of CoLF suggests the Co{sup 2+} ion to be five-coordinate, an observation similar to that reported for other Co{sup 2+}-substituted gluzincins, but distinct from that documented for the crystal structure of native LF. Furthermore, spectroscopic studies following the exposure of CoLF to thioglycolic acid (TGA) revealed a sequential mechanism of metal removal from LF, which likely involves the formation of an enzyme: Co{sup 2+}:TGA ternary complex prior to demetallation of the active site. CoLF reported herein constitutes the first spectroscopic probe of LF's active site, which may be utilized in future studies to gain further insight into the enzyme's mechanism and inhibitor interactions.

  2. A genome-wide RNAi screen reveals a Trio-regulated Rho GTPase circuitry transducing mitogenic signals initiated by G protein-coupled receptors.

    PubMed

    Vaqué, Jose P; Dorsam, Robert T; Feng, Xiaodong; Iglesias-Bartolome, Ramiro; Forsthoefel, David J; Chen, Qianming; Debant, Anne; Seeger, Mark A; Ksander, Bruce R; Teramoto, Hidemi; Gutkind, J Silvio

    2013-01-10

    Activating mutations in GNAQ and GNA11, encoding members of the G?(q) family of G protein ? subunits, are the driver oncogenes in uveal melanoma, and mutations in Gq-linked G protein-coupled receptors have been identified recently in numerous human malignancies. How G?(q) and its coupled receptors transduce mitogenic signals is still unclear because of the complexity of signaling events perturbed upon Gq activation. Using a synthetic-biology approach and a genome-wide RNAi screen, we found that a highly conserved guanine nucleotide exchange factor, Trio, is essential for activating Rho- and Rac-regulated signaling pathways acting on JNK and p38, and thereby transducing proliferative signals from G?(q) to the nucleus independently of phospholipase C-?. Indeed, whereas many biological responses elicited by Gq depend on the transient activation of second-messenger systems, Gq utilizes a hard-wired protein-protein-interaction-based signaling circuitry to achieve the sustained stimulation of proliferative pathways, thereby controlling normal and aberrant cell growth. PMID:23177739

  3. InVivo Loss of Function Screening Reveals Carbonic Anhydrase IX as a Key Modulator of Tumor Initiating Potential in Primary Pancreatic Tumors.

    PubMed

    Pore, Nabendu; Jalla, Sanjoo; Liu, Zheng; Higgs, Brandon; Sorio, Claudio; Scarpa, Aldo; Hollingsworth, Robert; Tice, David A; Michelotti, Emil

    2015-06-01

    Reprogramming of energy metabolism is one of the emerging hallmarks of cancer. Up-regulation of energy metabolism pathways fuels cell growth and division, a key characteristic of neoplastic disease, and can lead to dependency on specific metabolic pathways. Thus, targeting energy metabolism pathways might offer the opportunity for novel therapeutics. Here, we describe the application of a novel in vivo screening approach for the identification of genes involved in cancer metabolism using a patient-derived pancreatic xenograft model. Lentiviruses expressing short hairpin RNAs (shRNAs) targeting 12 different cell surface protein transporters were separately transduced into the primary pancreatic tumor cells. Transduced cells were pooled and implanted into mice. Tumors were harvested at different times, and the frequency of each shRNA was determined as a measure of which ones prevented tumor growth. Several targets including carbonic anhydrase IX (CAIX), monocarboxylate transporter 4, and anionic amino acid transporter light chain, xc- system (xCT) were identified in these studies and shown to be required for tumor initiation and growth. Interestingly, CAIX was overexpressed in the tumor initiating cell population. CAIX expression alone correlated with a highly tumorigenic subpopulation of cells. Furthermore, CAIX expression was essential for tumor initiation because shRNA knockdown eliminated the ability of cells to grow in vivo. To the best of our knowledge, this is the first parallel in vivo assessment of multiple novel oncology target genes using a patient-derived pancreatic tumor model. PMID:26152355

  4. A high-throughput chemical screen reveals that harmine-mediated inhibition of DYRK1A increases human pancreatic beta cell replication.

    PubMed

    Wang, Peng; Alvarez-Perez, Juan-Carlos; Felsenfeld, Dan P; Liu, Hongtao; Sivendran, Sharmila; Bender, Aaron; Kumar, Anil; Sanchez, Roberto; Scott, Donald K; Garcia-Ocaña, Adolfo; Stewart, Andrew F

    2015-04-01

    Types 1 and 2 diabetes affect some 380 million people worldwide. Both ultimately result from a deficiency of functional pancreatic insulin-producing beta cells. Beta cells proliferate in humans during a brief temporal window beginning around the time of birth, with a peak percentage (?2%) engaged in the cell cycle in the first year of life. In embryonic life and after early childhood, beta cell replication is barely detectable. Whereas beta cell expansion seems an obvious therapeutic approach to beta cell deficiency, adult human beta cells have proven recalcitrant to such efforts. Hence, there remains an urgent need for antidiabetic therapeutic agents that can induce regeneration and expansion of adult human beta cells in vivo or ex vivo. Here, using a high-throughput small-molecule screen (HTS), we find that analogs of the small molecule harmine function as a new class of human beta cell mitogenic compounds. We also define dual-specificity tyrosine-regulated kinase-1a (DYRK1A) as the likely target of harmine and the nuclear factors of activated T cells (NFAT) family of transcription factors as likely mediators of human beta cell proliferation and differentiation. Using three different mouse and human islet in vivo-based models, we show that harmine is able to induce beta cell proliferation, increase islet mass and improve glycemic control. These observations suggest that harmine analogs may have unique therapeutic promise for human diabetes therapy. Enhancing the potency and beta cell specificity of these compounds are important future challenges. PMID:25751815

  5. Two-color cell array screen reveals interdependent roles for histone chaperones and a chromatin boundary regulator in histone gene repression.

    PubMed

    Fillingham, Jeffrey; Kainth, Pinay; Lambert, Jean-Philippe; van Bakel, Harm; Tsui, Kyle; Peña-Castillo, Lourdes; Nislow, Corey; Figeys, Daniel; Hughes, Timothy R; Greenblatt, Jack; Andrews, Brenda J

    2009-08-14

    We describe a fluorescent reporter system that exploits the functional genomic tools available in budding yeast to systematically assess consequences of genetic perturbations on gene expression. We used our Reporter-Synthetic Genetic Array (R-SGA) method to screen for regulators of core histone gene expression. We discovered that the histone chaperone Rtt106 functions in a pathway with two other chaperones, Asf1 and the HIR complex, to create a repressive chromatin structure at core histone promoters. We found that activation of histone (HTA1) gene expression involves both relief of Rtt106-mediated repression by the activity of the histone acetyltransferase Rtt109 and restriction of Rtt106 to the promoter region by the bromodomain-containing protein Yta7. We propose that the maintenance of Asf1/HIR/Rtt106-mediated repressive chromatin domains is the primary mechanism of cell-cycle regulation of histone promoters. Our data suggest that this pathway may represent a chromatin regulatory mechanism that is broadly used across the genome. PMID:19683497

  6. A screen for modifiers of cyclin E function in Drosophila melanogaster identifies Cdk2 mutations, revealing the insignificance of putative phosphorylation sites in Cdk2.

    PubMed

    Lane, M E; Elend, M; Heidmann, D; Herr, A; Marzodko, S; Herzig, A; Lehner, C F

    2000-05-01

    In higher eukaryotes, cyclin E is thought to control the progression from G1 into S phase of the cell cycle by associating as a regulatory subunit with cdk2. To identify genes interacting with cyclin E, we have screened in Drosophila melanogaster for mutations that act as dominant modifiers of an eye phenotype caused by a Sevenless-CycE transgene that directs ectopic Cyclin E expression in postmitotic cells of eye imaginal disc and causes a rough eye phenotype in adult flies. The majority of the EMS-induced mutations that we have identified fall into four complementation groups corresponding to the genes split ends, dacapo, dE2F1, and Cdk2(Cdc2c). The Cdk2 mutations in combination with mutant Cdk2 transgenes have allowed us to address the regulatory significance of potential phosphorylation sites in Cdk2 (Thr 18 and Tyr 19). The corresponding sites in the closely related Cdk1 (Thr 14 and Tyr 15) are of crucial importance for regulation of the G2/M transition by myt1 and wee1 kinases and cdc25 phosphatases. In contrast, our results demonstrate that the equivalent sites in Cdk2 play no essential role. PMID:10790398

  7. Loss of Desmoplakin Tail Causes Lethal Acantholytic Epidermolysis Bullosa*

    PubMed Central

    Jonkman, Marcel F.; Pasmooij, Anna M. G.; Pasmans, Suzanne G. M. A.; van den Berg, Maarten P.; ter Horst, Henk J.; Timmer, Albertus; Pas, Hendri H.

    2005-01-01

    The cytoplasmic plaque protein desmoplakin (DP), which is located in desmosomes, plays a major role in epithelial and muscle cell adhesion by linking the transmembrane cadherins to the cytoplasmic intermediate filament network. Mutations of DP may cause striate palmoplantar keratoderma, arrhythmogenic right ventricular dysplasia, skin fragility/woolly hair syndrome, Naxos-like disease, and Carvajal syndrome. DP must be indispensable, because DP-/- mice are early abortive. Here, we report a patient with severe fragility of skin and mucous membranes caused by genetic truncation of the DP tail. The new phenotype is lethal in the neonatal period because of immense transcutaneous fluid loss. The phenotype also comprised universal alopecia, neonatal teeth, and nail loss. Histology showed suprabasal clefting and acantholysis throughout the spinous layer, mimicking pemphigus. Electron microscopy revealed disconnection of keratin intermediate filaments from desmosomes. Immunofluorescence staining of DP showed a distinct punctate intercellular pattern in the patient’s skin. Protein analysis revealed expression of truncated DP polypeptides. Mutational analysis of the patient demonstrated compound heterozygosity for two DP mutations, 6079C?T (R1934X) and 6370delTT, respectively. Aberrant mRNA transcripts that predict premature termination of translation with loss of the three intermediate filament-binding subdomains in the DP tail were detected by RT-PCR. The new dramatic phenotype, which we named “lethal acantholytic epidermolysis bullosa,” underscores the paramount role of DP in epidermal integrity. PMID:16175511

  8. The Spatial Concentration of Southern Whites and Argument-Based Lethal Violence

    ERIC Educational Resources Information Center

    Lee, Matthew R.; Shihadeh, Edward S.

    2009-01-01

    This analysis examines how the spatial concentration of Southern whites is associated with white argument-based lethal violence. Using a well-known measure of spatial segregation (V, the adjusted P* index) among Southern-born whites in U.S. counties in 2000, the results reveal that the spatial concentration of Southern-born whites is only…

  9. A genome-wide miRNA screen revealed miR-603 as a MGMT-regulating miRNA in glioblastomas.

    PubMed

    Kushwaha, Deepa; Ramakrishnan, Valya; Ng, Kimberly; Steed, Tyler; Nguyen, Thien; Futalan, Diahnn; Akers, Johnny C; Sarkaria, Jann; Jiang, Tao; Chowdhury, Dipanjan; Carter, Bob S; Chen, Clark C

    2014-06-30

    MGMT expression is a critical determinant for therapeutic resistance to DNA alkylating agents. We previously demonstrated that MGMT expression is post-transcriptionally regulated by miR-181d and other miRNAs. Here, we performed a genome-wide screen to identify MGMT regulating miRNAs. Candidate miRNAs were further tested for inverse correlation with MGMT expression in clinical specimens. We identified 15 candidate miRNAs and characterized the top candidate, miR-603. Transfection of miR-603 suppressed MGMT mRNA/protein expression in vitro and in vivo; this effect was reversed by transfection with antimiR-603. miR-603 affinity-precipitated with MGMT mRNA and suppressed luciferase activity in an MGMT-3'UTR-luciferase assay, suggesting direct interaction between miR-603 and MGMT 3'UTR. miR-603 transfection enhanced the temozolomide (TMZ) sensitivity of MGMT-expressing glioblastoma cell lines. Importantly, miR-603 mediated MGMT suppression and TMZ resistance were reversed by expression of an MGMT cDNA. In a collection of 74 clinical glioblastoma specimens, both miR-603 and miR-181d levels inversely correlated with MGMT expression. Moreover, a combined index of the two miRNAs better reflected MGMT expression than each individually. These results suggest that MGMT is co-regulated by independent miRNAs. Characterization of these miRNAs should contribute toward strategies for enhancing the efficacy of DNA alkylating agents. PMID:24994119

  10. TRAIL-Based High Throughput Screening Reveals a Link between TRAIL-Mediated Apoptosis and Glutathione Reductase, a Key Component of Oxidative Stress Response

    PubMed Central

    Rozanov, Dmitri; Cheltsov, Anton; Sergienko, Eduard; Vasile, Stefan; Golubkov, Vladislav; Aleshin, Alexander E.; Levin, Trevor; Traer, Elie; Hann, Byron; Freimuth, Julia; Alexeev, Nikita; Alekseyev, Max A.; Budko, Sergey P; Bächinger, Hans Peter; Spellman, Paul

    2015-01-01

    A high throughput screen for compounds that induce TRAIL-mediated apoptosis identified ML100 as an active chemical probe, which potentiated TRAIL activity in prostate carcinoma PPC-1 and melanoma MDA-MB-435 cells. Follow-up in silico modeling and profiling in cell-based assays allowed us to identify NSC130362, pharmacophore analog of ML100 that induced 65-95% cytotoxicity in cancer cells and did not affect the viability of human primary hepatocytes. In agreement with the activation of the apoptotic pathway, both ML100 and NSC130362 synergistically with TRAIL induced caspase-3/7 activity in MDA-MB-435 cells. Subsequent affinity chromatography and inhibition studies convincingly demonstrated that glutathione reductase (GSR), a key component of the oxidative stress response, is a target of NSC130362. In accordance with the role of GSR in the TRAIL pathway, GSR gene silencing potentiated TRAIL activity in MDA-MB-435 cells but not in human hepatocytes. Inhibition of GSR activity resulted in the induction of oxidative stress, as was evidenced by an increase in intracellular reactive oxygen species (ROS) and peroxidation of mitochondrial membrane after NSC130362 treatment in MDA-MB-435 cells but not in human hepatocytes. The antioxidant reduced glutathione (GSH) fully protected MDA-MB-435 cells from cell lysis induced by NSC130362 and TRAIL, thereby further confirming the interplay between GSR and TRAIL. As a consequence of activation of oxidative stress, combined treatment of different oxidative stress inducers and NSC130362 promoted cell death in a variety of cancer cells but not in hepatocytes in cell-based assays and in in vivo, in a mouse tumor xenograft model. PMID:26075913

  11. Differential screening of mutated SOD1 transgenic mice reveals early up-regulation of a fast axonal transport component in spinal cord motor neurons.

    PubMed

    Dupuis, L; de Tapia, M; René, F; Lutz-Bucher, B; Gordon, J W; Mercken, L; Pradier, L; Loeffler, J P

    2000-08-01

    In the present study we analyze the molecular mechanisms underlying motor neuron degeneration in familial amyotrophic lateral sclerosis (FALS). For this, we used a transgenic mouse model expressing the Cu/Zn superoxide dismutase (SOD1) gene with a Gly(86) to Arg (G86R) mutation equivalent to that found in a subset of human FALS. Using an optimized suppression subtractive hybridization method, a cDNA specifically up-regulated during the asymptomatic phase in the lumbar spinal cord of G86R mice was identified by sequence analysis as the KIF3-associated protein (KAP3), a regulator of fast axonal transport. RT-PCR analysis revealed that KAP3 induction was an early event arising long before axonal degeneration. Immunohistochemical studies further revealed that KAP3 protein predominantly accumulates in large motor neurons of the ventral spinal cord. We further demonstrated that KAP3 up-regulation occurs independent of any change in the other components of the kinesin II complex. However, since the ubiquitous KIF1A motor is up-regulated, our results show an early and complex rearrangement of the fast axonal transport machinery in the course of FALS pathology. PMID:10964600

  12. Vascular Endothelial Growth Factor Receptor-1 Is Synthetic Lethal to Aberrant {beta}-Catenin Activation in Colon Cancer.

    PubMed

    Naik, Snehal; Dothager, Robin S; Marasa, Jayne; Lewis, Cory L; Piwnica-Worms, David

    2009-12-15

    PURPOSE: The Wnt/beta-catenin (beta-cat) signaling cascade is a key regulator of development, and dysregulation of Wnt/beta-cat contributes to selected cancers, such as colorectal, breast, and hepatocellular carcinoma, through abnormal activation of Wnt target genes. To identify novel modulators of the Wnt/beta-cat pathway that may emerge as therapeutic targets, we did an unbiased high-throughput RNA interference screen. EXPERIMENTAL DESIGN: A synthetic oligonucleotide small interfering RNA library targeting 691 known and predicted human kinases was screened in Wnt3a-stimulated human cells in a live cell luciferase assay for modulation of Wnt/beta-cat-dependent transcription. Follow-up studies of a selected high-confidence "hit" were conducted. RESULTS: A robust quartile-based statistical analysis and secondary screen yielded several kinases worthy of further investigation, including Cdc2L1, Lmtk3, Pank2, ErbB3, and, of note, vascular endothelial growth factor receptor (VEGFR)1/Flt1, a receptor tyrosine kinase (TK) with putative weak kinase activity conventionally believed to be a negative regulator of angiogenesis. A series of loss-of-function, genetic null, and VEGFR TK inhibitor assays further revealed that VEGFR1 is a positive regulator of Wnt signaling that functions in a glycogen synthase kinase-3beta (GSK3beta)-independent manner as a potential synthetic lethal target in Wnt/beta-cat-addicted colon carcinoma cells. CONCLUSIONS: This unanticipated non-endothelial link between VEGFR1 TK activity and Wnt/beta-cat signaling may refine our understanding of aberrant Wnt signaling in colon carcinoma and points to new combinatorial therapeutics targeted to the tumor cell compartment, rather than angiogenesis, in the context of colon cancer. (Clin Cancer Res 2009;15(24):7529-37). PMID:20008853

  13. CSNK1E/CTNNB1 are synthetic lethal to TP53 in colorectal cancer and are markers for prognosis.

    PubMed

    Tiong, Khong-Loon; Chang, Kuo-Ching; Yeh, Kun-Tu; Liu, Ting-Yuan; Wu, Jia-Hong; Hsieh, Ping-Heng; Lin, Shu-Hui; Lai, Wei-Yun; Hsu, Yu-Chin; Chen, Jeou-Yuan; Chang, Jan-Gowth; Shieh, Grace S

    2014-05-01

    Two genes are called synthetic lethal (SL) if their simultaneous mutations lead to cell death, but each individual mutation does not. Targeting SL partners of mutated cancer genes can kill cancer cells specifically, but leave normal cells intact. We present an integrated approach to uncovering SL pairs in colorectal cancer (CRC). Screening verified SL pairs using microarray gene expression data of cancerous and normal tissues, we first identified potential functionally relevant (simultaneously differentially expressed) gene pairs. From the top-ranked pairs, ~20 genes were chosen for immunohistochemistry (IHC) staining in 171 CRC patients. To find novel SL pairs, all 169 combined pairs from the individual IHC were synergistically correlated to five clinicopathological features, e.g. overall survival. Of the 11 predicted SL pairs, MSH2-POLB and CSNK1E-MYC were consistent with literature, and we validated the top two pairs, CSNK1E-TP53 and CTNNB1-TP53 using RNAi knockdown and small molecule inhibitors of CSNK1E in isogenic HCT-116 and RKO cells. Furthermore, synthetic lethality of CSNK1E and TP53 was verified in mouse model. Importantly, multivariate analysis revealed that CSNK1E-P53, CTNNB1-P53, MSH2-RB1, and BRCA1-WNT5A were independent prognosis markers from stage, with CSNK1E-P53 applicable to early-stage and the remaining three throughout all stages. Our findings suggest that CSNK1E is a promising target for TP53-mutant CRC patients which constitute ~40% to 50% of patients, while to date safety regarding inhibition of TP53 is controversial. Thus the integrated approach is useful in finding novel SL pairs for cancer therapeutics, and it is readily accessible and applicable to other cancers. PMID:24947187

  14. Therapeutically targeting RNA viruses via lethal mutagenesis

    PubMed Central

    Graci, Jason D; Cameron, Craig E

    2008-01-01

    RNA viruses exhibit increased mutation frequencies relative to other organisms. Recent work has attempted to exploit this unique feature by increasing the viral mutation frequency beyond an extinction threshold, an antiviral strategy known as lethal mutagenesis. A number of novel nucleoside analogs have been designed around this premise. Herein, we review the quasispecies nature of RNA viruses and survey the antiviral, biological and biochemical characteristics of mutagenic nucleoside analogs, including clinically-used ribavirin. Biological implications of modulating viral replication fidelity are discussed in the context of translating lethal mutagenesis into a clinically-useful antiviral strategy. PMID:19727424

  15. Disease screening of three breeding populations of adult exhibition budgerigars (Melopsittacus undulatus) in New Zealand reveals a high prevalence of a novel polyomavirus and avian malaria infection.

    PubMed

    Baron, Hamish R; Howe, Laryssa; Varsani, Arvind; Doneley, Robert J T

    2014-03-01

    Disease surveillance is vital to the management of New Zealand's endemic and threatened avian species. Three infectious agents that are potential threats to New Zealand's endemic birds include avian polyomavirus (APV), beak and feather disease virus (BFDV), and avian malaria. All three agents have been reported in New Zealand; however, possible reservoir populations have not been identified. In this communication, we report the first study of APV, BFDV, and avian malaria in introduced adult exhibition budgerigars (Melopsittacus undulatus) in New Zealand. Blood samples were collected from 90 living adult budgerigars from three breeding locations in the North Island of New Zealand. An overall APV prevalence of 22% was determined using a broad-spectrum nested PCR that amplified the major capsid protein VP1 gene of polyomavirus. Phylogenetic analysis of the VP1 gene revealed a unique isolate of APV, which had a sequence divergence of 32% to previously reported budgerigar fledgling disease strains and 33% to the recently reported New Zealand finch isolate. All of the budgerigars sampled were found to be PCR negative for BFDV, and an overall prevalence of 30% was detected by PCR for avian malaria. Sequencing revealed the presence of ubiquitous malarial strains and also the potentially destructive Plasmodium relictum strain. The results of this study suggest that both APV and avian malaria are present in New Zealand adult budgerigars, and our study highlights the need for further studies to determine whether these pathogens in captive bird populations may be a threat or spill over into New Zealand's endemic and threatened avifauna and whether prevention and control methods need to be implemented. PMID:24758122

  16. Structure-Based Systematic Isolation of Conditional-Lethal Mutations in the Single Yeast Calmodulin Gene

    PubMed Central

    Ohya, Y.; Botstein, D.

    1994-01-01

    Conditional-lethal mutations of the single calmodulin gene in Saccharomyces cerevisiae have been very difficult to isolate by random and systematic methods, despite the fact that deletions cause recessive lethality. We report here the isolation of numerous conditional-lethal mutants that were recovered by systematically altering phenylalanine residues. The phenylalanine residues of calmodulin were implicated in function both by structural studies of calmodulin bound to target peptides and by their extraordinary conservation in evolution. Seven single and 26 multiple Phe -> Ala mutations were constructed. Mutant phenotypes were examined in a haploid cmd1 disrupted strain under three conditions: single copy, low copy, and overexpressed. Whereas all but one of the single mutations caused no obvious phenotype, most of the multiple mutations caused obvious growth phenotypes. Five were lethal, 6 were lethal only in synthetic medium, 13 were temperature-sensitive lethal and 2 had no discernible phenotypic consequences. Overexpression of some of the mutant genes restored the phenotype to nearly wild type. Several temperature-sensitive calmodulin mutations were suppressed by elevated concentration of CaCl(2) in the medium. Mutant calmodulin protein was detected at normal levels in extracts of most of the lethal mutant cells, suggesting that the deleterious phenotypes were due to loss of the calmodulin function and not protein instability. Analysis of diploid strains heterozygous for all combinations of cmd1-ts alleles revealed four intragenic complementation groups. The contributions of individual phe->ala changes to mutant phenotypes support the idea of internal functional redundancy in the symmetrical calmodulin protein molecule. These results suggest that the several phenylalanine residues in calmodulin are required to different extents in different combinations in order to carry out each of the several essential tasks. PMID:7896089

  17. A cancer screen in zebrafish identifies many ribosomal proteins as haploinsufficient tumor suppressors

    E-print Network

    Lai, Kevin

    2006-01-01

    A collection of over 500 lines of zebrafish (Danio rerio), each heterozygous for a recessive embryonic lethal mutation caused by a retroviral insertion, was screened for lines that displayed early mortality and externally ...

  18. Candidate Gene Screen in the Red Flour Beetle Tribolium Reveals Six3 as Ancient Regulator of Anterior Median Head and Central Complex Development

    PubMed Central

    Hein, Hendrikje Jeannette; Bucher, Gregor

    2011-01-01

    Several highly conserved genes play a role in anterior neural plate patterning of vertebrates and in head and brain patterning of insects. However, head involution in Drosophila has impeded a systematic identification of genes required for insect head formation. Therefore, we use the red flour beetle Tribolium castaneum in order to comprehensively test the function of orthologs of vertebrate neural plate patterning genes for a function in insect head development. RNAi analysis reveals that most of these genes are indeed required for insect head capsule patterning, and we also identified several genes that had not been implicated in this process before. Furthermore, we show that Tc-six3/optix acts upstream of Tc-wingless, Tc-orthodenticle1, and Tc-eyeless to control anterior median development. Finally, we demonstrate that Tc-six3/optix is the first gene known to be required for the embryonic formation of the central complex, a midline-spanning brain part connected to the neuroendocrine pars intercerebralis. These functions are very likely conserved among bilaterians since vertebrate six3 is required for neuroendocrine and median brain development with certain mutations leading to holoprosencephaly. PMID:22216011

  19. Crystal structure of the anthrax lethal factor

    Microsoft Academic Search

    Andrew D. Pannifer; Thiang Yian Wong; Robert Schwarzenbacher; Martin Renatus; Carlo Petosa; Jadwiga Bienkowska; D. Borden Lacy; R. John Collier; Stephen H. Leppla; Philip Hanna; Robert C. Liddington

    2001-01-01

    Lethal factor (LF) is a protein (relative molecular mass 90,000) that is critical in the pathogenesis of anthrax. It is a highly specific protease that cleaves members of the mitogen-activated protein kinase kinase (MAPKK) family near to their amino termini, leading to the inhibition of one or more signalling pathways. Here we describe the crystal structure of LF and its

  20. Medical Conditions and Nearly Lethal Suicide Attempts.

    ERIC Educational Resources Information Center

    Ikeda, Robin M.; Kresnow, Marcie-jo; Mercy, James A.; Powell, Kenneth E.; Simon, Thomas R.; Potter, Lloyd B.; Durant, Tonji M.; Swahn, Monica H.

    2002-01-01

    This population-based, case-control study examined physical illness as a risk factor for suicidal behavior. Case patients were more likely than controls to report having any serious medical conditions. Results suggest that young men with medical conditions are at increased risk for nearly lethal suicide attempts. (Contains 33 references and 3…

  1. Non-lethal technologies—an overview

    Microsoft Academic Search

    Nick LEWER; Neil DAVISON

    hilst the focus for this issue of Disarmament Forum is on chemical and biological weapons, sight should not be lost of the spectrum of non-lethal technologies that are being deployed or under development. These technologies will have an increasing impact on war fighting, peace support operations, civil policing and prison control. It is our purpose here to briefly review the

  2. ATR pathway inhibition is synthetically lethal in cancer cells with ERCC1 deficiency

    PubMed Central

    Mohni, Kareem N.; Kavanaugh, Gina M.; Cortez, David

    2014-01-01

    The DNA damage response kinase ATR and its effector kinase CHEK1 are required for cancer cells to survive oncogene-induced replication stress. ATR inhibitors exhibit synthetic lethal interactions with deficiencies in the DNA damage response enzymes ATM and XRCC1 and with overexpression of the cell cycle kinase Cyclin E. Here we report a systematic screen to identify synthetic lethal interactions with ATR-pathway targeted drugs, rationalized by their predicted therapeutic utility in the oncology clinic. We found that reduced function in the ATR pathway itself provided the strongest synthetic lethal interaction. In addition, we found that loss of the structure specific-endonuclease ERCC1-XPF (ERCC4) is synthetic lethal with ATR pathway inhibitors. ERCC1-deficient cells exhibited elevated levels of DNA damage, which was increased further by ATR inhibition. When treated with ATR or CHEK1 inhibitors, ERCC1-deficient cells arrested in S phase and failed to complete cell cycle transit even after drug removal. Notably, triple-negative breast cancer cells and non-small cell lung cancer cells depleted of ERCC1 exhibited increased sensitivity to ATR-pathway targeted drugs. Overall, we concluded that ATR pathway-targeted drugs may offer particular utility in cancers with reduced ATR pathway function or reduced levels of ERCC4 activity. PMID:24662920

  3. A genetic screen for temperature-sensitive cell-division mutants of Caenorhabditis elegans.

    PubMed Central

    O'Connell, K F; Leys, C M; White, J G

    1998-01-01

    A novel screen to isolate conditional cell-division mutants in Caenorhabditis elegans has been developed. The screen is based on the phenotypes associated with existing cell-division mutations: some disrupt postembryonic divisions and affect formation of the gonad and ventral nerve cord-resulting in sterile, uncoordinated animals-while others affect embryonic divisions and result in lethality. We obtained 19 conditional mutants that displayed these phenotypes when shifted to the restrictive temperature at the appropriate developmental stage. Eighteen of these mutations have been mapped; 17 proved to be single alleles of newly identified genes, while 1 proved to be an allele of a previously identified gene. Genetic tests on the embryonic lethal phenotypes indicated that for 13 genes, embryogenesis required maternal expression, while for 6, zygotic expression could suffice. In all cases, maternal expression of wild-type activity was found to be largely sufficient for embryogenesis. Cytological analysis revealed that 10 mutants possessed embryonic cell-division defects, including failure to properly segregate DNA, failure to assemble a mitotic spindle, late cytokinesis defects, prolonged cell cycles, and improperly oriented mitotic spindles. We conclude that this approach can be used to identify mutations that affect various aspects of the cell-division cycle. PMID:9649522

  4. Zygotic Lethal Mutations with Maternal Effect Phenotypes in Drosophila Melanogaster. II. Loci on the Second and Third Chromosomes Identified by P-Element-Induced Mutations

    PubMed Central

    Perrimon, N.; Lanjuin, A.; Arnold, C.; Noll, E.

    1996-01-01

    Screens for zygotic lethal mutations that are associated with specific maternal effect lethal phenotypes have only been conducted for the X chromosome. To identify loci on the autosomes, which represent four-fifths of the Drosophila genome, we have used the autosomal ``FLP-DFS'' technique to screen a collection of 496 P element-induced mutations established by the Berkeley Drosophila Genome Project. We have identified 64 new loci whose gene products are required for proper egg formation or normal embryonic development. PMID:8978055

  5. Fiducial Generalized Confidence Interval for Median Lethal Dose (LD50)

    E-print Network

    Hannig, Jan

    Fiducial Generalized Confidence Interval for Median Lethal Dose (LD50) Lidong E , Jan Hannig and Hari Iyer§ July 20, 2009 Abstract Median lethal dose (LD50) is a common measure of acute toxicity lethal dose (LD50), Fiducial Generalized Confidence In- terval (FGCI), Gibbs sampling. This work

  6. Gene function prediction from congruent synthetic lethal interactions in yeast

    Microsoft Academic Search

    Ping Ye; Brian D Peyser; Xuewen Pan; Jef D Boeke; Forrest A Spencer; Joel S Bader

    2005-01-01

    We predicted gene function using synthetic lethal genetic interactions between null alleles in Saccharomyces cerevisiae. Phenotypic and protein interaction data indicate that synthetic lethal gene pairs function in parallel or compensating pathways. Congruent gene pairs, defined as sharing synthetic lethal partners, are in single pathway branches. We predicted benomyl sensitivity and nuclear migration defects using congruence; these phenotypes were uncorrelated

  7. Crystallographic studies of the Anthrax lethal toxin. Annual report

    Microsoft Academic Search

    1996-01-01

    The lethal form of Anthrax results from the inhalation of anthrax spores. Death is primarily due to the effects of the lethal toxin (Protective Antigen (PA) + Lethal Factor) from the causative agent, Bacillus anthracis. All the Anthrax vaccines currently in use or under development contain or produce PA, the major antigenic component of anthrax toxin, and there is a

  8. Health Screening

    MedlinePLUS

    Screenings are tests that look for diseases before you have symptoms. Screening tests can find diseases early, when they're easier ... Overweight and obesity Prostate cancer in men Which tests you need depends on your age, your sex, ...

  9. Whole-Transcriptome Shotgun Sequencing (RNA-seq) Screen Reveals Upregulation of Cellobiose and Motility Operons of Lactobacillus ruminis L5 during Growth on Tetrasaccharides Derived from Barley ?-Glucan

    PubMed Central

    Lawley, Blair; Sims, Ian M.

    2013-01-01

    Lactobacillus ruminis is an inhabitant of human bowels and bovine rumens. None of 10 isolates (three from bovine rumen, seven from human feces) of L. ruminis that were tested could utilize barley ?-glucan for growth. Seven of the strains of L. ruminis were, however, able to utilize tetrasaccharides (3-O-?-cellotriosyl-d-glucose [LDP4] or 4-O-?-laminaribiosyl-d-cellobiose [CDP4]) present in ?-glucan hydrolysates for growth. The tetrasaccharides were generated by the use of lichenase or cellulase, respectively. To learn more about the utilization of tetrasaccharides by L. ruminis, whole-transcriptome shotgun sequencing (RNA-seq) was tested as a transcriptional screen to detect altered gene expression when an autochthonous human strain (L5) was grown in medium containing CDP4. RNA-seq results were confirmed and extended by reverse transcription-quantitative PCR assays of selected genes in two upregulated operons when cells were grown as batch cultures in medium containing either CDP4 or LDP4. The cellobiose utilization operon had increased transcription, particularly in early growth phase, whereas the chemotaxis/motility operon was upregulated in late growth phase. Phenotypic changes were seen in relation to upregulation of chemotaxis/flagellar operons: flagella were rarely seen by electron microscopy on glucose-grown cells but cells cultured in tetrasaccharide medium were commonly flagellated. Chemotactic movement toward tetrasaccharides was demonstrated in capillary cultures. L. ruminis utilized 3-O-?-cellotriosyl-d-glucose released by ?-glucan hydrolysis due to bowel commensal Coprococcus sp., indicating that cross feeding of tetrasaccharide between bacteria could occur. Therefore, the RNA-seq screen and subsequent experiments had utility in revealing foraging attributes of gut commensal Lactobacillus ruminis. PMID:23851085

  10. In Silico Screening for Palmitoyl Substrates Reveals a Role for DHHC1/3/10 (zDHHC1/3/11)-mediated Neurochondrin Palmitoylation in Its Targeting to Rab5-positive Endosomes*

    PubMed Central

    Oku, Shinichiro; Takahashi, Naoki; Fukata, Yuko; Fukata, Masaki

    2013-01-01

    Protein palmitoylation, a common post-translational lipid modification, plays an important role in protein trafficking and functions. Recently developed palmitoyl-proteomic methods identified many novel substrates. However, the whole picture of palmitoyl substrates has not been clarified. Here, we performed global in silico screening using the CSS-Palm 2.0 program, free software for prediction of palmitoylation sites, and selected 17 candidates as novel palmitoyl substrates. Of the 17 candidates, 10 proteins, including 6 synaptic proteins (Syd-1, transmembrane AMPA receptor regulatory protein (TARP) ?-2, TARP ?-8, cornichon-2, Ca2+/calmodulin-dependent protein kinase II?, and neurochondrin (Ncdn)/norbin), one focal adhesion protein (zyxin), two ion channels (TRPM8 and TRPC1), and one G-protein-coupled receptor (orexin 2 receptor), were palmitoylated. Using the DHHC palmitoylating enzyme library, we found that all tested substrates were palmitoylated by the Golgi-localized DHHC3/7 subfamily. Ncdn, a regulator for neurite outgrowth and synaptic plasticity, was robustly palmitoylated by the DHHC1/10 (zDHHC1/11; z1/11) subfamily, whose substrate has not yet been reported. As predicted by CSS-Palm 2.0, Cys-3 and Cys-4 are the palmitoylation sites for Ncdn. Ncdn was specifically localized in somato-dendritic regions, not in the axon of rat cultured neurons. Stimulated emission depletion microscopy revealed that Ncdn was localized to Rab5-positive early endosomes in a palmitoylation-dependent manner, where DHHC1/10 (z1/11) were also distributed. Knockdown of DHHC1, -3, or -10 (z11) resulted in the loss of Ncdn from Rab5-positive endosomes. Thus, through in silico screening, we demonstrate that Ncdn and the DHHC1/10 (z1/11) and DHHC3/7 subfamilies are novel palmitoyl substrate-enzyme pairs and that Ncdn palmitoylation plays an essential role in its specific endosomal targeting. PMID:23687301

  11. Rapidly lethal dermatomyositis associated with cutaneous lymphangitis carcinomatosa

    PubMed Central

    Resende, Cristina; Araújo, Catarina; Duarte, Maria Luz; Brito, Celeste

    2013-01-01

    A 70-year-old woman with a recent diagnosis of dermatomyositis (DM) presented to the dermatology department for study of a probably paraneoplastic syndrome. On examination, we observed discrete, indurated, reddish, painful plaques and nodules on her abdomen and both thighs. A cutaneous biopsy from an abdominal nodule, performed as part of the paraneoplastic workup, was suggestive of cutaneous lymphangitis carcinomatosa, secondary to unknown malignancy. An extensive investigation to locate the site of the primary tumour revealed no specific findings. A course of palliative chemotherapy with cisplatin and 5-fluorouracil was then given, but the patient’s condition deteriorated and 6?months after her initial observation the patient died. We describe this case because, to our knowledge, the association between DM and cutaneous lymphangitis carcinomatosa has not been described yet in the literature and to highlight that, DM can be a rapidly lethal disease. PMID:23761617

  12. Efficient synthetic inhibitors of anthrax lethal factor

    PubMed Central

    Forino, Martino; Johnson, Sherida; Wong, Thiang Y.; Rozanov, Dmitri V.; Savinov, Alexei Y.; Li, Wei; Fattorusso, Roberto; Becattini, Barbara; Orry, Andrew J.; Jung, Dawoon; Abagyan, Ruben A.; Smith, Jeffrey W.; Alibek, Ken; Liddington, Robert C.; Strongin, Alex Y.; Pellecchia, Maurizio

    2005-01-01

    Inhalation anthrax is a deadly disease for which there is currently no effective treatment. Bacillus anthracis lethal factor (LF) metalloproteinase is an integral component of the tripartite anthrax lethal toxin that is essential for the onset and progression of anthrax. We report here on a fragment-based approach that allowed us to develop inhibitors of LF. The small-molecule inhibitors we have designed, synthesized, and tested are highly potent and selective against LF in both in vitro tests and cell-based assays. These inhibitors do not affect the prototype human metalloproteinases that are structurally similar to LF. Initial in vivo evaluation of postexposure efficacy of our inhibitors combined with antibiotic ciprofloxican against B. anthracis resulted in significant protection. Our data strongly indicate that the scaffold of inhibitors we have identified is the foundation for the development of novel, safe, and effective emergency therapy of postexposure inhalation anthrax. PMID:15983377

  13. Bullying and Lethal Acts of School Violence

    Microsoft Academic Search

    Jeffrey A. Daniels; Mary C. Bradley

    \\u000a In Chap. 3, we address a topic that has received considerable research attention over the past decade and has been implicated as a causal\\u000a factor in LSV. It has been reported that many lethal school shootings were in part in retaliation for being bullied. Within\\u000a this chapter, we address prevalence of bullying in the USA and then discuss different forms

  14. Substrate specificity of the anthrax lethal factor

    Microsoft Academic Search

    M. Yu. Zakharova; S. A. Dubiley; D. M. Chudakov; A. G. Gabibov; I. G. Shemyakin; A. V. Kolesnikov

    2008-01-01

    The lethal factor (LF), a high-specific metalloprotease, is a subunit of the B. anthracis toxin. In susceptible cells, LF has a toxic effect due to inactivation of MKK-family kinases (mitogen-activated serine?threonine kinase kinases) and presumably of some other proteins. To develop an approach to neutralization of the anthrax toxin effect, the entire pattern of LF cell targets should be determined.

  15. Lethal interpersonal violence in the middle pleistocene.

    PubMed

    Sala, Nohemi; Arsuaga, Juan Luis; Pantoja-Pérez, Ana; Pablos, Adrián; Martínez, Ignacio; Quam, Rolf M; Gómez-Olivencia, Asier; Bermúdez de Castro, José María; Carbonell, Eudald

    2015-01-01

    Evidence of interpersonal violence has been documented previously in Pleistocene members of the genus Homo, but only very rarely has this been posited as the possible manner of death. Here we report the earliest evidence of lethal interpersonal violence in the hominin fossil record. Cranium 17 recovered from the Sima de los Huesos Middle Pleistocene site shows two clear perimortem depression fractures on the frontal bone, interpreted as being produced by two episodes of localized blunt force trauma. The type of injuries, their location, the strong similarity of the fractures in shape and size, and the different orientations and implied trajectories of the two fractures suggest they were produced with the same object in face-to-face interpersonal conflict. Given that either of the two traumatic events was likely lethal, the presence of multiple blows implies an intention to kill. This finding shows that the lethal interpersonal violence is an ancient human behavior and has important implications for the accumulation of bodies at the site, supporting an anthropic origin. PMID:26018668

  16. Lethal Interpersonal Violence in the Middle Pleistocene

    PubMed Central

    Sala, Nohemi; Arsuaga, Juan Luis; Pantoja-Pérez, Ana; Pablos, Adrián; Martínez, Ignacio; Quam, Rolf M.; Gómez-Olivencia, Asier; Bermúdez de Castro, José María; Carbonell, Eudald

    2015-01-01

    Evidence of interpersonal violence has been documented previously in Pleistocene members of the genus Homo, but only very rarely has this been posited as the possible manner of death. Here we report the earliest evidence of lethal interpersonal violence in the hominin fossil record. Cranium 17 recovered from the Sima de los Huesos Middle Pleistocene site shows two clear perimortem depression fractures on the frontal bone, interpreted as being produced by two episodes of localized blunt force trauma. The type of injuries, their location, the strong similarity of the fractures in shape and size, and the different orientations and implied trajectories of the two fractures suggest they were produced with the same object in face-to-face interpersonal conflict. Given that either of the two traumatic events was likely lethal, the presence of multiple blows implies an intention to kill. This finding shows that the lethal interpersonal violence is an ancient human behavior and has important implications for the accumulation of bodies at the site, supporting an anthropic origin. PMID:26018668

  17. Lethal and sub-lethal effects of UVB on juvenile Biomphalaria glabrata (Mollusca: Pulmonata).

    PubMed

    Ruelas, Debbie S; Karentz, Deneb; Sullivan, John T

    2006-11-01

    Although Schistosoma mansoni occurs mainly in the tropics, where intense levels of solar radiation are present, the impact of ultraviolet (UV) light on schistosome transmission is not known. The purpose of this study was to investigate potential effects of UVB (290-320nm) on juvenile Biomphalaria glabrata, the snail intermediate host of S. mansoni. Albino and wild-type snails were exposed to doses of UVB from UV-fluorescent lamps, and the following were measured: survival, photoreactivation (light-mediated DNA repair), effects on feeding behavior, and morphological tissue abnormalities. Irradiation with UVB is lethal to B. glabrata in a dose-dependent manner. Exposure to white light subsequent to UVB irradiation enhances survival, probably by photoreactivation. The shell offers some, but not complete, protection. Experiments in which UVB transmittance through the shell was blocked with black nail polish suggest that injury to both exposed (headfoot) and shell-enclosed (mantle and visceral mass) tissues contributes to mortality in lethally irradiated snails. Wild-type (pigmented) snails are less susceptible to lethal effects of UVB than albino snails, and they may be more capable of photoreactivation. UVB exposure inhibits snail feeding behavior, and causes tentacle forks and growths on the headfoot. Thus, UVB may influence the life cycle of S. mansoni by both lethal and sub-lethal damage to the snail intermediate host. However, the ability of snails to photoreactivate may mitigate these effects. PMID:16996081

  18. Lethal and Sub-lethal Effects of UVB on Juvenile Biomphalaria glabrata (Mollusca: Pulmonata)

    PubMed Central

    Ruelas, Debbie S.; Karentz, Deneb; Sullivan, John T.

    2007-01-01

    Although Schistosoma mansoni occurs mainly in the tropics, where intense levels of solar radiation are present, the impact of ultraviolet (UV) light on schistosome transmission is not known. The purpose of this study was to investigate potential effects of UVB (290–320 nm) on juvenile Biomphalaria glabrata, the snail intermediate host of S. mansoni. Albino and wild type snails were exposed to doses of UVB from UV-fluorescent lamps, and the following were measured: survival, photoreactivation (light-mediated DNA repair), effects on feeding behavior, and morphological tissue abnormalities. Irradiation with UVB is lethal to B. glabrata in a dose-dependent manner. Exposure to white light subsequent to UVB irradiation enhances survival, probably by photoreactivation. The shell offers some, but not complete, protection. Experiments in which UVB transmittance through the shell was blocked with black nail polish suggest that injury to both exposed (headfoot) and shell-enclosed (mantle and visceral mass) tissues contributes to mortality in lethally-irradiated snails. Wild-type (pigmented) snails are less susceptible to lethal effects of UVB than albino snails, and they may be more capable of photoreactivation. UVB exposure inhibits snail feeding behavior, and causes tentacle forks and growths on the headfoot. Thus, UVB may influence the life cycle of S. mansoni by both lethal and sub-lethal damage to the snail intermediate host. However, the ability of snails to photoreactivate may mitigate these effects. PMID:16996081

  19. Establishment and Characterization of a Lethal Mouse Model for the Angola Strain of Marburg Virus

    PubMed Central

    Wong, Gary; Audet, Jonathan; Cutts, Todd; Niu, Yulian; Booth, Stephanie

    2014-01-01

    ABSTRACT Infections with Marburg virus (MARV) and Ebola virus (EBOV) cause severe hemorrhagic fever in humans and nonhuman primates (NHPs) with fatality rates up to 90%. A number of experimental vaccine and treatment platforms have previously been shown to be protective against EBOV infection. However, the rate of development for prophylactics and therapeutics against MARV has been lower in comparison, possibly because a small-animal model is not widely available. Here we report the development of a mouse model for studying the pathogenesis of MARV Angola (MARV/Ang), the most virulent strain of MARV. Infection with the wild-type virus does not cause disease in mice, but the adapted virus (MARV/Ang-MA) recovered from liver homogenates after 24 serial passages in severe combined immunodeficient (SCID) mice caused severe disease when administered intranasally (i.n.) or intraperitoneally (i.p.). The median lethal dose (LD50) was determined to be 0.015 50% TCID50 (tissue culture infective dose) of MARV/Ang-MA in SCID mice, and i.p. infection at a dose of 1,000× LD50 resulted in death between 6 and 8 days postinfection in SCID mice. Similar results were obtained with immunocompetent BALB/c and C57BL/6 mice challenged i.p. with 2,000× LD50 of MARV/Ang-MA. Virological and pathological analyses of MARV/Ang-MA-infected BALB/c mice revealed that the associated pathology was reminiscent of observations made in NHPs with MARV/Ang. MARV/Ang-MA-infected mice showed most of the clinical hallmarks observed with Marburg hemorrhagic fever, including lymphopenia, thrombocytopenia, marked liver damage, and uncontrolled viremia. Virus titers reached 108 TCID50/ml in the blood and between 106 and 1010 TCID50/g tissue in the intestines, kidney, lungs, brain, spleen, and liver. This model provides an important tool to screen candidate vaccines and therapeutics against MARV infections. IMPORTANCE The Angola strain of Marburg virus (MARV/Ang) was responsible for the largest outbreak ever documented for Marburg viruses. With a 90% fatality rate, it is similar to Ebola virus, which makes it one of the most lethal viruses known to humans. There are currently no approved interventions for Marburg virus, in part because a small-animal model that is vulnerable to MARV/Ang infection is not available to screen and test potential vaccines and therapeutics in a quick and economical manner. To address this need, we have adapted MARV/Ang so that it causes illness in mice resulting in death. The signs of disease in these mice are reminiscent of wild-type MARV/Ang infections in humans and nonhuman primates. We believe that this will be of help in accelerating the development of life-saving measures against Marburg virus infections. PMID:25142608

  20. A rational approach to prenatal screening and intervention.

    PubMed

    Yagel, S; Anteby, E

    1998-05-01

    Improved testing procedures now allow prenatal screening for a wide range of congenital defects, including cystic fibrosis and muscular dystrophy. An emerging technique also allows diagnosis of congenital anomalies during the pre-implantation stage of in vitro fertilization. Thus, clinicians need an established criteria to use as a guide when counseling parents about what prenatal testing is possible, feasible, and desirable. For example, there are limits to prenatal testing for conditions like mutations in the breast cancer gene because affected individuals do not necessary develop the condition and a cure may be found by the time the condition develops. It is even questionable if parents should be given this information until the child reaches an appropriate age. One approach to development of guidelines is to classify congenital abnormalities according to severity, age of onset, and type (structural-functional versus mental). This system reveals anomalies that are clearly lethal, lead to moderate or severe disability with little or no prospect of improvement or cure, are characterized by early onset, and/or involve obvious mental retardation. This approach is particularly relevant in cases of trisomy 21, and progressively invasive screening techniques are available to detect this most common pattern of malformation in humans. PMID:9647530

  1. Lethal Forethought: Delayed Reward Discounting Differentiates High- and Low-Lethality Suicide Attempts in Old Age

    PubMed Central

    Dombrovski, Alexandre Y.; Szanto, Katalin; Siegle, Greg J.; Wallace, Meredith L.; Forman, Steven D.; Sahakian, Barbara; Reynolds, Charles F.; Clark, Luke

    2011-01-01

    Background The decision to commit suicide may be impulsive, but lethal suicidal acts often involve planning and forethought. People who attempt suicide make disadvantageous decisions in other contexts, but nothing is known about the way they decide about the future. Can the willingness to postpone future gratification differentiate between individuals prone to serious, premeditated and less serious, unplanned suicidal acts? Methods Four groups of depressed participants aged 60+ made choices between smaller immediate and larger delayed monetary rewards: 15 who made high-lethality suicide attempts, 14 who made low-lethality suicide attempts, 12 who seriously contemplated suicide, and 42 people with depression but no history of suicidal thoughts. The reference group was 31 psychiatrically healthy elders. Results Individuals who had made low-lethality attempts displayed an exaggerated preference for immediate rewards compared to non-suicidal depressed and healthy controls. Those who had carried out high-lethality suicide attempts were more willing to delay future rewards, compared to low-lethality attempters. Better planned suicide attempts were also associated with willingness to wait for larger rewards. These effects were unchanged after accounting for education, global cognitive function, substance use disorders, psychotropic medications, and possible brain injury from attempts. Discount rates were correlated with having debt but were not significantly associated with income, hopelessness, depressive severity, premorbid IQ, age at first attempt, or choice of violent means. Conclusions While clinicians often focus on impulsivity in patients at risk for suicide, these data suggest that identifying biological characteristics and treatments for non-impulsive suicidal older people may be even more important. PMID:21329911

  2. Touch Screens

    Microsoft Academic Search

    Michael McRoberts

    \\u000a You are now going to take a look at a cool gadget that you can use easily with an Arduino—a touch screen. Since the advent\\u000a of smart phones and handheld game consoles, touch screens are now inexpensive and readily available. A touch screen allows\\u000a you to make an easy touch interface for a device or it can be overlaid onto

  3. Screening for prostate cancer: estimating the magnitude of overdetection

    PubMed Central

    McGregor, M; Hanley, J A; Boivin, J F; McLean, R G

    1998-01-01

    BACKGROUND: No randomized controlled trial of prostate cancer screening has been reported and none is likely to be completed in the near future. In the absence of direct evidence, the decision to screen must therefore be based on estimates of benefits and risks. The main risk of screening is overdetection--the detection of cancer that, if left untreated, would not cause death. In this study the authors estimate the level of overdetection that might result from annual screening of men aged 50-70. METHODS: The annual rate of lethal screen-detectable cancer (detectable cancer that would prove fatal before age 85 if left untreated) was calculated from the observed prostate cancer mortality rate in Quebec; the annual rate of all cases of screen-detectable prostate cancer was calculated from 2 recent screening studies. RESULTS: The annual rate of lethal screen-detectable prostate cancer was estimated to be 1.3 per 1000 men. The annual rate of all cases of screen-detectable prostate cancer was estimated to be 8.0 per 1000 men. The estimated case-fatality rate among men up to 85 years of age was 16% (1.3/8.0) (sensitivity analysis 13% to 22%). INTERPRETATION: Of every 100 men with screen-detected prostate cancer, only 16 on average (13 to 22) could have their lives extended by surgery, since the prostate cancer would not cause death before age 85 in the remaining 84 (78 to 87). PMID:9861205

  4. Bacillus anthracis Lethal Toxin Reduces Human Alveolar Epithelial Barrier Function

    PubMed Central

    Langer, Marybeth; Duggan, Elizabeth Stewart; Booth, John Leland; Patel, Vineet Indrajit; Zander, Ryan A.; Silasi-Mansat, Robert; Ramani, Vijay; Veres, Tibor Zoltan; Prenzler, Frauke; Sewald, Katherina; Williams, Daniel M.; Coggeshall, Kenneth Mark; Awasthi, Shanjana; Lupu, Florea; Burian, Dennis; Ballard, Jimmy Dale; Braun, Armin

    2012-01-01

    The lung is the site of entry for Bacillus anthracis in inhalation anthrax, the deadliest form of the disease. Bacillus anthracis produces virulence toxins required for disease. Alveolar macrophages were considered the primary target of the Bacillus anthracis virulence factor lethal toxin because lethal toxin inhibits mouse macrophages through cleavage of MEK signaling pathway components, but we have reported that human alveolar macrophages are not a target of lethal toxin. Our current results suggest that, unlike human alveolar macrophages, the cells lining the respiratory units of the lung, alveolar epithelial cells, are a target of lethal toxin in humans. Alveolar epithelial cells expressed lethal toxin receptor protein, bound the protective antigen component of lethal toxin, and were subject to lethal-toxin-induced cleavage of multiple MEKs. These findings suggest that human alveolar epithelial cells are a target of Bacillus anthracis lethal toxin. Further, no reduction in alveolar epithelial cell viability was observed, but lethal toxin caused actin rearrangement and impaired desmosome formation, consistent with impaired barrier function as well as reduced surfactant production. Therefore, by compromising epithelial barrier function, lethal toxin may play a role in the pathogenesis of inhalation anthrax by facilitating the dissemination of Bacillus anthracis from the lung in early disease and promoting edema in late stages of the illness. PMID:23027535

  5. Discovery and development of anthrax lethal factor metalloproteinase inhibitors.

    PubMed

    Turk, Benjamin E

    2008-02-01

    Anthrax is caused by infection with Bacillus anthracis, a spore forming, rod-shaped, encapsulated gram positive bacteria. The disease manifests itself in distinct ways depending on the route of entry of infective bacterial spores: cutaneous, inhalational, and gastrointestinal. Though rare in humans, inhalational anthrax has become a major concern due to the capacity for spores to be weaponized. The limited success of antibiotic therapy has motivated investigation of complementary therapeutic strategies that target the bacteria's secreted toxin. The zinc-dependent metalloproteinase lethal factor (LF) is a critical component of anthrax toxin and an important potential target for small molecule drugs. In the past few years, a number of approaches have been taken to identify LF inhibitors, from generating conventional metal chelating substrate analogs to random screening of diverse compound libraries. These efforts have produced several different classes of specific nanomolar range inhibitors. Some compounds have fared well in animal models for anthrax toxemia and infection, and these inhibitors and their derivatives may form the basis for future therapies to treat the disease in humans. PMID:18289054

  6. Lethal mobilization of DDT by cowbirds

    USGS Publications Warehouse

    Van Velzen, A.C.; Stiles, W.B.; Stickel, L.F.

    1972-01-01

    This study is an experimental demonstration of lethal mobilization of DDT by brown-headed cowbirds (Molothrus ater) and the effects of food deprivation on the distribution and loss of DDT, DDD, and DDE. The principal experimental group consisted of 20 birds fed a dietary dosage of 100 ppm of DDT for 13 days. After 2 days of full rations of untreated food, they were subjected to food restriction. Food was reduced to 43 percent of normal. Seven of the 20 birds died within 4 days. No birds died in the three control groups, treated as follows: ( 1 ) 20 birds fed 100 ppm DDT for 13 days and full rations of untreated food thereafter, (2) 20 birds fed only untreated food but subjected to food restriction, and (3) 20 birds fed full rations of untreated food throughout. In a pilot study, birds were fed 100, 200, or 300 ppm of DDT and subjected to two periods of food restriction, the first of these immediately after dosage ceased and the second 4 months later. DDT-dosed birds from all dosage levels died in each period of food restriction. Before the weight loss that accompanied food restriction, the brains of DDT-dosed birds had concentrations of DDT and DDD that were far below the lethal range. Concentrations increased rapidly to lethal levels. In these birds, DDT in carcasses decreased while DDD increased. DDT-dosed birds that died during food restriction lost 16 percent of their total body burden of DDT + DDD + DDE, 21 percent of their weight, and 81 percent of their fat. The DDT-dosed birds that were subjected to food restriction but survived lost a significantly greater proportion of their body burden of residues than similarly dosed birds not subjected to weight loss. Brain levels of DDT and DDD in birds that died during food restriction soon after dosage did not differ significantly from brain levels of birds that died in a period of food restriction 4 months after dosage. Concentrations of DDE were significantly higher in the latter group, although they were lower than concentrations considered to be lethal. In contrast, carcass levels of DDT and DDD were significantly lower, and DDE was only slightly higher, in the birds that died in the second period of food restriction. It is concluded that stored DDT residues present a hazard to birds, which utilize stored fat during periods of stress due to reproduction, cold weather, disease, injury, limited food supply, or migration.

  7. Genetic analysis and complementation by germ-line transformation of lethal mutations in the unc-22 IV region of Caenorhabditis elegans

    Microsoft Academic Search

    Denise V. Clark; David L. Baillie

    1992-01-01

    The subject of this study is the organization of essential genes in the 2 map-unit unc-22 IV region of the Caenorhabditis elegans genome. With the goal of achieving mutational saturation of essential genes in this region, 6491 chromosomes mutagenized with ethyl methanesulfonate (EMS) were screened for the presence of lethal mutations in the unc-22 region. The genetic analysis of 21

  8. Carotid Artery Screening

    MedlinePLUS

    Carotid Artery Screening What is carotid artery screening? Who should consider heart screening – and why? How are the procedures ... more information about heart screening? What is carotid artery screening? Screening examinations are tests performed to find ...

  9. Suicide Intent and Accurate Expectations of Lethality: Predictors of Medical Lethality of Suicide Attempts

    ERIC Educational Resources Information Center

    Brown, Gregory K.; Henriques, Gregg R.; Sosdjan, Daniella; Beck, Aaron T.

    2004-01-01

    The degree of intent to commit suicide and the severity of self-injury were examined in individuals (N = 180) who had recently attempted suicide. Although a minimal association was found between the degree of suicide intent and the degree of lethality of the attempt, the accuracy of expectations about the likelihood of dying was found to moderate…

  10. Screen Time

    NSDL National Science Digital Library

    OMSI

    2007-01-01

    This game asks you a series of questions about how much time you spend in front of a screen, not being active. It begins by pointing out that since we spend a lot of time in front of computer screens at work or school, additional time at home can really affect how healthy we are. It asks how much time you spend watching TV, playing computer games, and using the computer each day. It then adds up the total amount of screen time you spend every day, and calculates how many hours you spend a year in front of a screen. It also tells you if that's a healthy amount, and suggests ways to stay active while in front of screens.

  11. Rapid, optimized interactomic screening.

    PubMed

    Hakhverdyan, Zhanna; Domanski, Michal; Hough, Loren E; Oroskar, Asha A; Oroskar, Anil R; Keegan, Sarah; Dilworth, David J; Molloy, Kelly R; Sherman, Vadim; Aitchison, John D; Fenyö, David; Chait, Brian T; Jensen, Torben Heick; Rout, Michael P; LaCava, John

    2015-06-01

    We must reliably map the interactomes of cellular macromolecular complexes in order to fully explore and understand biological systems. However, there are no methods to accurately predict how to capture a given macromolecular complex with its physiological binding partners. Here, we present a screening method that comprehensively explores the parameters affecting the stability of interactions in affinity-captured complexes, enabling the discovery of physiological binding partners in unparalleled detail. We have implemented this screen on several macromolecular complexes from a variety of organisms, revealing novel profiles for even well-studied proteins. Our approach is robust, economical and automatable, providing inroads to the rigorous, systematic dissection of cellular interactomes. PMID:25938370

  12. Apparent lethal concentrations of pyrolysis products of some polymeric materials

    NASA Technical Reports Server (NTRS)

    Hilado, C. J.; Marcussen, W. H.; Furst, A.; Kourtides, D. A.; Parker, J. A.

    1976-01-01

    Thirty-nine samples of polymeric materials were evaluated to determine the apparent lethal concentrations of their pyrolysis products. The materials were compared on the basis of the apparent lethal concentration for 50 percent of the test animals. Relative toxicity rankings based o apparent lethal concentration values can differ significantly depending on whether they are based on weight of sample charged or weight of sample pyrolyzed. The ranking of polyphenylene sulfide is particularly sensitive to this difference.

  13. Antenatal diagnosis of lethal skeletal dysplasias.

    PubMed

    Tretter, A E; Saunders, R C; Meyers, C M; Dungan, J S; Grumbach, K; Sun, C C; Campbell, A B; Wulfsberg, E A

    1998-02-17

    Lethal skeletal dysplasias (LSD) are a heterogeneous group of rare but important genetic disorders characterized by abnormal growth and development of bone and cartilage. We describe the diagnosis and outcome of 29 cases of lethal skeletal dysplasias evaluated between January 1989 and December 1996 at the University of Maryland Medical Center and the Ultrasound Institute of Baltimore. Two cases presented at delivery with no prenatal care while the remaining 27 cases were identified by antenatal sonography. Final diagnoses included thanatophoric dysplasia (14), osteogenesis imperfecta, type II (6), achondrogenesis (2), short rib syndromes (3), campomelic syndrome (2), atelosteogenesis (1), and no evidence of a skeletal dysplasia (1). Twenty out of 27 pregnancies were terminated with an average at detection of 21.6 weeks. The other 7 pregnancies that went on to deliver had an average age at detection of 29.2 weeks. Fetal abnormalities in the terminated pregnancies were identified at a significantly earlier gestational age (P = 0.0016) than the pregnancies that continued. While the identification of LSD by sonography was excellent (26/27), only 13/27 (48%) were given an accurate specific antenatal diagnosis. In 8/14 (57%) cases with an inaccurate or nonspecific diagnosis there was a significant or crucial change in the genetic counseling. Thus, while antenatal sonography is an excellent method for discovering LSD, clinical examination, radiographs, and autopsy are mandatory for making a specific diagnosis. PMID:9489797

  14. Hyperactivated Wnt Signaling Induces Synthetic Lethal Interaction with Rb Inactivation by Elevating TORC1 Activities

    PubMed Central

    Hsu, Fu-Ning; Zhang, Robin; Searle, Jennifer S.; Pei, Xun; Li, Xuan; Ryoo, Hyung Don; Ji, Jun-Yuan; Du, Wei

    2014-01-01

    Inactivation of the Rb tumor suppressor can lead to increased cell proliferation or cell death depending on specific cellular context. Therefore, identification of the interacting pathways that modulate the effect of Rb loss will provide novel insights into the roles of Rb in cancer development and promote new therapeutic strategies. Here, we identify a novel synthetic lethal interaction between Rb inactivation and deregulated Wg/Wnt signaling through unbiased genetic screens. We show that a weak allele of axin, which deregulates Wg signaling and increases cell proliferation without obvious effects on cell fate specification, significantly alters metabolic gene expression, causes hypersensitivity to metabolic stress induced by fasting, and induces synergistic apoptosis with mutation of fly Rb ortholog, rbf. Furthermore, hyperactivation of Wg signaling by other components of the Wg pathway also induces synergistic apoptosis with rbf. We show that hyperactivated Wg signaling significantly increases TORC1 activity and induces excessive energy stress with rbf mutation. Inhibition of TORC1 activity significantly suppressed synergistic cell death induced by hyperactivated Wg signaling and rbf inactivation, which is correlated with decreased energy stress and decreased induction of apoptotic regulator expression. Finally the synthetic lethality between Rb and deregulated Wnt signaling is conserved in mammalian cells and that inactivation of Rb and APC induces synergistic cell death through a similar mechanism. These results suggest that elevated TORC1 activity and metabolic stress underpin the evolutionarily conserved synthetic lethal interaction between hyperactivated Wnt signaling and inactivated Rb tumor suppressor. PMID:24809668

  15. Mutations synthetically lethal with cep1 target S. cerevisiae kinetochore components.

    PubMed Central

    Baker, R E; Harris, K; Zhang, K

    1998-01-01

    CP1 (encoded by CEP1) is a Saccharomyces cerevisiae chromatin protein that binds a DNA element conserved in centromeres and in the 5'-flanking DNA of methionine biosynthetic (MET) genes. Strains lacking CP1 are defective in chromosome segregation and MET gene transcription, leading to the hypothesis that CP1 plays a general role in assembling higher order chromatin structures at genomic sites where it is bound. A screen for mutations synthetically lethal with a cep1 null allele yielded five recessive csl (cep1 synthetic lethal) mutations, each defining a unique complementation group. Four of the five mutations synergistically increased the loss rate of marker chromosomes carrying a centromere lacking the CP1 binding site, suggesting that the cep1 synthetic lethality was due to chromosome segregation defects. Three of these four CSL genes were subsequently found to be known or imputed kinetochore genes: CEP3, NDC10, and CSE4. The fourth, CSL4, corresponded to ORF YNL232w on chromosome XIV, and was found to be essential. A human cDNA was identified that encoded a protein homologous to Csl4 and that complemented the csl4-1 mutation. The results are consistent with the view that the major cellular role of CP1 is to safeguard the biochemical integrity of the kinetochore. PMID:9584087

  16. Genome-wide siRNA screen reveals a new cellular partner of NK cell receptor KIR2DL4: heparan sulfate directly modulates KIR2DL4-mediated responses

    PubMed Central

    Brusilovsky, Michael; Cordoba, Moti; Rosental, Benyamin; Hershkovitz, Oren; Andrake, Mark D.; Pecherskaya, Anna; Einarson, Margret B.; Zhou, Yan; Braiman, Alex

    2013-01-01

    KIR2DL4 (CD158d) is a distinct member of the killer cell Ig-like receptor (KIR) family in human NK cells that can induce cytokine production and cytolytic activity in resting NK cells. Soluble HLA-G, normally expressed only by fetal-derived trophoblast cells, was reported to be a ligand for KIR2DL4; however, KIR2DL4 expression is not restricted to the placenta and can be found in CD56high subset of peripheral blood NK cells. We demonstrated that KIR2DL4 can interact with alternative ligand(s), expressed by cells of epithelial or fibroblast origin. A genome-wide high-throughput siRNA screen revealed that KIR2DL4 recognition of cells surface ligand(s) is directly regulated by heparan sulfate (HS) glucosamine 3-O-sulfotransferase 3B1 (HS3ST3B1). KIR2DL4 was found to directly interact with HS/heparin, and the D0-domain of KIR2DL4 was essential for this interaction. Accordingly, exogenous HS/heparin can regulate cytokine production by KIR2DL4-expressing NK cells and HEK293T cells (HEK293T-2DL4) and induces differential localization of KIR2DL4 to rab5+ and rab7+ endosomes, thus leading to down-regulation of cytokine production and degradation of the receptor. Furthermore, we showed that intimate interaction of syndecan-4 (SDC4) HS Proteo-Glycan (HSPG) and KIR2DL4 directly affects receptor endocytosis and membrane trafficking. PMID:24127555

  17. Gill lesions in the major carp, Labeo rohita exposed to lethal and sublethal concentrations of tannery effluent.

    PubMed

    Dhanapakiam, P; Sampoorani, V; Kavitha, M; Ramasamy, V K; Chandrakala, A; Aruna, K C

    2004-07-01

    The major carp, Labeo rohita were exposed to (0.873%) lethal and sublethal (0.073%) concentrations of tannery effluent for 24h and 40 days respectively under static bioassay condition. The surface architecture of gill revealed severe damages such as : fusion and clumping in the middle and distal parts of the primary lamellae, swelling and deterioration of the cells. The interlamellar space was filled either with hyperplastic epithelial or mucous cells. Secondary lamellae lost their identity and appeared finger like in structure in the lethal concentration and necrosis was observed in the primary and secondary epithelium. Swelling of primary and secondary epithelial cells was evident in sublethal concentration. PMID:15847345

  18. Vision Screening

    MedlinePLUS

    ... office. Some community screenings use this method. Corneal light reflex testing This simple test can be performed ... focuses on a penlight, the position of the light reflection from the front surface (cornea) of the ...

  19. Ants defend aphids against lethal disease.

    PubMed

    Nielsen, Charlotte; Agrawal, Anurag A; Hajek, Ann E

    2010-04-23

    Social insects defend their own colonies and some species also protect their mutualist partners. In mutualisms with aphids, ants typically feed on honeydew produced by aphids and, in turn guard and shelter aphid colonies from insect natural enemies. Here we report that Formica podzolica ants tending milkweed aphids, Aphis asclepiadis, protect aphid colonies from lethal fungal infections caused by an obligate aphid pathogen, Pandora neoaphidis. In field experiments, bodies of fungal-killed aphids were quickly removed from ant-tended aphid colonies. Ant workers were also able to detect infective conidia on the cuticle of living aphids and responded by either removing or grooming these aphids. Our results extend the long-standing view of ants as mutualists and protectors of aphids by demonstrating focused sanitizing and quarantining behaviour that may lead to reduced disease transmission in aphid colonies. PMID:19923138

  20. Comparison of Lethal Zone Characteristics of Several Natural Fragmenting Warheads

    Microsoft Academic Search

    Berko ZECEVIC; Alan CATOVIC; Jasmin TERZIC

    Research of HE warheads lethal zone is very complex topic because of large number of controlled and independent, sometimes correlated, influencing factors. Capability for prediction of lethal zone is based on complexity of databases regarding natural fragmentation parameters, which should contain data about warhead body material characteristics, types of explosive charge, number, mass, initial velocity and spatial distribution of fragments,

  1. Anthrax lethal toxin: a weapon of multisystem destruction

    Microsoft Academic Search

    A. Agrawal; B. Pulendran

    2004-01-01

    Lethal toxin (LT) is a major virulence factor secreted by anthrax bacteria. It is composed of two proteins, PA (protective antigen) and LF (lethal factor). PA transports the LF inside the cell, where LF, a zinc-dependent metalloprotease cleaves the mitogen activated protein kinase kinase (MAPKK) enzymes of the mitogen activated protein kinase (MAPK) signaling pathway, thereby impairing their function. This

  2. A Single Phosphorodiamidate Morpholino Oligomer Targeting VP24 Protects Rhesus Monkeys against Lethal Ebola Virus Infection

    PubMed Central

    Warren, Travis K.; Whitehouse, Chris A.; Wells, Jay; Welch, Lisa; Heald, Alison E.; Charleston, Jay S.; Sazani, Pete; Reid, St. Patrick; Iversen, Patrick L.

    2015-01-01

    ABSTRACT Ebola viruses (EBOV) cause severe disease in humans and nonhuman primates with high mortality rates and continue to emerge in new geographic locations, including several countries in West Africa, the site of a large ongoing outbreak. Phosphorodiamidate morpholino oligomers (PMOs) are synthetic antisense molecules that are able to target mRNAs in a sequence-specific fashion and suppress translation through steric hindrance. We previously showed that the use of PMOs targeting a combination of VP35 and VP24 protected rhesus monkeys from lethal EBOV infection. Surprisingly, the present study revealed that a PMOplus compound targeting VP24 alone was sufficient to confer protection from lethal EBOV infection but that a PMOplus targeting VP35 alone resulted in no protection. This study further substantiates recent data demonstrating that VP24 may be a key virulence factor encoded by EBOV and suggests that VP24 is a promising target for the development of effective anti-EBOV countermeasures. PMID:25670780

  3. Sub-lethal effects of Roundup™ on tadpole anti-predator responses.

    PubMed

    Moore, Harrison; Chivers, Douglas P; Ferrari, Maud C O

    2015-01-01

    Roundup™ is a commonly used pesticide applied to agriculture and forest habitats. These areas are generally ideal for amphibians due to the presence of small, ephemeral water bodies. While Roundup™ has been shown to have lethal effects on many species of amphibians, effects on behaviour and sensory perception have yet to be considered. Here, we exposed wood frog tadpoles to a sub-lethal concentration of Roundup™ and showed that the ability of tadpoles to respond to injured conspecific cues, an important source of information regarding local predation risk, was impaired. Subsequent experiments revealed that impaired responses likely result from a chemical reaction between the Roundup™ and the cues and that tadpoles chronically exposed to Roundup™ had reduced basal movement rates compared with unexposed tadpoles. Our data demonstrate that environmentally-relevant concentrations of Roundup™ can drastically alter movement and anti-predator responses of tadpoles, with potential negative consequences for the population. PMID:25450945

  4. Synthetic lethality in ATM-deficient RAD50-mutant tumors underlie outlier response to cancer therapy

    PubMed Central

    Al-Ahmadie, Hikmat; Iyer, Gopa; Hohl, Marcel; Asthana, Saurabh; Inagaki, Akiko; Schultz, Nikolaus; Hanrahan, Aphrothiti J.; Scott, Sasinya N.; Brannon, A. Rose; McDermott, Gregory C.; Pirun, Mono; Ostrovnaya, Irina; Kim, Philip; Socci, Nicholas D.; Viale, Agnes; Schwartz, Gary K.; Reuter, Victor; Bochner, Bernard H.; Rosenberg, Jonathan E.; Bajorin, Dean F.; Berger, Michael F.; Petrini, John H.J.; Solit, David B.; Taylor, Barry S.

    2014-01-01

    Metastatic solid tumors are almost invariably fatal. Patients with disseminated small-cell cancers have a particularly unfavorable prognosis with most succumbing to their disease within two years. Here, we report on the genetic and functional analysis of an outlier curative response of a patient with metastatic small cell cancer to combined checkpoint kinase 1 (Chk1) inhibition and DNA damaging chemotherapy. Whole-genome sequencing revealed a clonal hemizygous mutation in the Mre11 complex gene RAD50 that attenuated ATM signaling which in the context of Chk1 inhibition contributed, via synthetic lethality, to extreme sensitivity to irinotecan. As Mre11 mutations occur in a diversity of human tumors, the results suggest a tumor-specific combination therapy strategy whereby checkpoint inhibition in combination with DNA damaging chemotherapy is synthetically lethal in tumor but not normal cells with somatic mutations that impair Mre11 complex function. PMID:24934408

  5. Phytochemical screening and anticonvulsant studies of ethyl acetate fraction of Globimetula braunii on laboratory animals

    PubMed Central

    Aliyu, Musa Mumammad; Musa, Abdullahi Isma'il; Kamal, Muhammad Ja'afar; Mohammed, Magaji Garba

    2014-01-01

    Objective To investigate the phytochemical properties and the anticonvulsant potential of the ethyl acetate soluble fraction of ethanol leaf extract of Globimetula braunii, a plant used in ethnomedicine for the treatment of epilepsy. Methods The phytochemical screening was carried out using standard protocol while the anticonvulsant activity was studied using maximal electroshock test in chicks, pentylenetetrazole and 4-aminopyridine-induced seizures in mice. Results The preliminary phytochemical screening carried out on the crude ethanol extract revealed the presence of saponins, carbohydrates, flavonoids, tannins, anthraquinones and steroids. Similarly, tannins, flavonoids and steroids/terpenes were found to be present in the ethyl acetate fraction. In the pharmacological screening, 150 mg/kg of the fraction protected 83.33% of animals against pentylenetetrazole-induced seizure in mice whereas sodium valproate a standard anti-epileptic drug offered 100% protection. In the 4-aminopyridine-induced seizure model, the fraction produced a significant (P<0.05) increase in the mean onset of seizure in unprotected animals. The fraction did not exhibit a significant activity against maximal electroshock convulsion. The median lethal dose of the fraction was found to be 1?261.91 mg/kg. Conclusions These results suggest that the ethyl acetate fraction of Globimetula braunii leaves extract possesses psychoactive compound that may be useful in the management of petit mal epilepsy and lend credence to the ethnomedical use of the plant in the management of epilepsy. PMID:25182552

  6. Chronic Exposure of Corals to Fine Sediments: Lethal and Sub-Lethal Impacts

    PubMed Central

    Flores, Florita; Hoogenboom, Mia O.; Smith, Luke D.; Cooper, Timothy F.; Abrego, David; Negri, Andrew P.

    2012-01-01

    Understanding the sedimentation and turbidity thresholds for corals is critical in assessing the potential impacts of dredging projects in tropical marine systems. In this study, we exposed two species of coral sampled from offshore locations to six levels of total suspended solids (TSS) for 16 weeks in the laboratory, including a 4 week recovery period. Dose-response relationships were developed to quantify the lethal and sub-lethal thresholds of sedimentation and turbidity for the corals. The sediment treatments affected the horizontal foliaceous species (Montipora aequituberculata) more than the upright branching species (Acropora millepora). The lowest sediment treatments that caused full colony mortality were 30 mg l?1 TSS (25 mg cm?2 day?1) for M. aequituberculata and 100 mg l?1 TSS (83 mg cm?2 day?1) for A. millepora after 12 weeks. Coral mortality generally took longer than 4 weeks and was closely related to sediment accumulation on the surface of the corals. While measurements of damage to photosystem II in the symbionts and reductions in lipid content and growth indicated sub-lethal responses in surviving corals, the most reliable predictor of coral mortality in this experiment was long-term sediment accumulation on coral tissue. PMID:22662225

  7. Prediction of lethal and synthetically lethal knock-outs in regulatory networks.

    PubMed

    Boldhaus, Gunnar; Greil, Florian; Klemm, Konstantin

    2013-03-01

    The complex interactions involved in regulation of a cell's function are captured by its interaction graph. More often than not, detailed knowledge about enhancing or suppressive regulatory influences and cooperative effects is lacking and merely the presence or absence of directed interactions is known. Here, we investigate to which extent such reduced information allows to forecast the effect of a knock-out or a combination of knock-outs. Specifically, we ask in how far the lethality of eliminating nodes may be predicted by their network centrality, such as degree and betweenness, without knowing the function of the system. The function is taken as the ability to reproduce a fixed point under a discrete Boolean dynamics. We investigate two types of stochastically generated networks: fully random networks and structures grown with a mechanism of node duplication and subsequent divergence of interactions. On all networks we find that the out-degree is a good predictor of the lethality of a single node knock-out. For knock-outs of node pairs, the fraction of successors shared between the two knocked-out nodes (out-overlap) is a good predictor of synthetic lethality. Out-degree and out-overlap are locally defined and computationally simple centrality measures that provide a predictive power close to the optimal predictor. PMID:22918565

  8. Lethal and Pre-Lethal Effects of a Fungal Biopesticide Contribute to Substantial and Rapid Control of Malaria

    E-print Network

    Read, Andrew

    Lethal and Pre-Lethal Effects of a Fungal Biopesticide Contribute to Substantial and Rapid Control ingredients to control the adult mosquitoes that vector malaria. Biopesticides based on the spores that `slow acting' fungal biopesticides are, therefore, incapable of delivering malaria control in real

  9. 76 FR 6054 - Use of Less-Than-Lethal Force: Delegation

    Federal Register 2010, 2011, 2012, 2013, 2014

    2011-02-03

    ...less-than-lethal weapons, including those containing chemical agents, may not...less-than-lethal weapons, including chemical agents. (a) The...less-than-lethal weapons, including those containing chemical agents, only...

  10. 28 CFR 552.25 - Use of less-than-lethal weapons, including chemical agents.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... false Use of less-than-lethal weapons, including chemical agents. 552...552.25 Use of less-than-lethal weapons, including chemical agents. (a...authorize the use of less-than-lethal weapons, including those containing...

  11. Received: 2 April 2009, Revised: 7 July 2009, Accepted: 15 July 2009, Published online in Wiley InterScience: 2009 Library screening by fragment-based docking

    E-print Network

    Caflisch, Amedeo

    reviews which contain a large number of applications.15­20 Successful in vitro screening campaigns have protein kinases,23­27 DNA gyrase,28 caspase,29,30 anthrax lethal factor,31 and phosphodi- esterase.32

  12. Human cooperation by lethal group competition.

    PubMed

    Egas, Martijn; Kats, Ralph; van der Sar, Xander; Reuben, Ernesto; Sabelis, Maurice W

    2013-01-01

    Why humans are prone to cooperate puzzles biologists, psychologists and economists alike. Between-group conflict has been hypothesized to drive within-group cooperation. However, such conflicts did not have lasting effects in laboratory experiments, because they were about luxury goods, not needed for survival ("looting"). Here, we find within-group cooperation to last when between-group conflict is implemented as "all-out war" (eliminating the weakest groups). Human subjects invested in helping group members to avoid having the lowest collective pay-off, whereas they failed to cooperate in control treatments with random group elimination or with no subdivision in groups. When the game was repeated, experience was found to promote helping. Thus, not within-group interactions alone, not random group elimination, but pay-off-dependent group elimination was found to drive within-group cooperation in our experiment. We suggest that some forms of human cooperation are maintained by multi-level selection: reciprocity within groups and lethal competition among groups acting together. PMID:23459158

  13. Lethal body burdens of polar narcotics: Chlorophenols

    SciTech Connect

    Wezel, A.P. van; Punte, S.S. [Utrecht Univ. (Netherlands). Environmental Chemistry Group; Opperhuizen, A. [Ministry of Transport, Public Works and Water Management, The Hague (Netherlands). National Institute for Coastal and Marine Management

    1995-09-01

    The goal of the present study was to measure in fathead minnow (Pimephales promelas) the lethal body burden (LBB) of three chlorophenols that are known as polar narcotic chemicals. The LBBs of the chlorophenols were compared to LBBs of nonpolar narcotic chemicals to consider if the two classes of narcotic chemicals differ on a body burden level. The LBB of the most acidic chlorophenol was measured at two different levels of pH exposure to determine the influence of the degree of ionization on the magnitude of the LBB. Both n-octanol/water partition coefficients and n-hexane/water partition coefficients of the chlorophenols were determined at different pH levels to consider the influence of ionization on the partition coefficient and to determine the importance of a polar group in the organic phase on the partitioning behavior. Partitioning to n-octanol and n-hexane was used as input in a model to simulate the equilibrium partitioning between hydrophobic and nonhydrophobic and target and nontarget compartments in the fish.

  14. Tumor clone dynamics in lethal prostate cancer

    PubMed Central

    Carreira, Suzanne; Romanel, Alessandro; Goodall, Jane; Grist, Emily; Ferraldeschi, Roberta; Miranda, Susana; Prandi, Davide; Lorente, David; Frenel, Jean-Sebastien; Pezaro, Carmel; Omlin, Aurelius; Rodrigues, Daniel Nava; Flohr, Penelope; Tunariu, Nina; de Bono, Johann S.; Demichelis, Francesca; Attard, Gerhardt

    2015-01-01

    It is unclear whether a single clone metastasizes and remains dominant over the course of lethal prostate cancer. We describe the clonal architectural heterogeneity at different stages of disease progression by sequencing serial plasma and tumor samples from 16 ERG-positive patients. By characterizing the clonality of commonly occurring deletions at 21q22, 8p21, and 10q23, we identified multiple independent clones in metastatic disease that are differentially represented in tissue and circulation. To exemplify the clinical utility of our studies, we then showed a temporal association between clinical progression and emergence of androgen receptor (AR) mutations activated by glucocorticoids in about 20% of patients progressing on abiraterone and prednisolone or dexamethasone. Resistant clones showed a complex dynamic with temporal and spatial heterogeneity, suggesting distinct mechanisms of resistance at different sites that emerged and regressed depending on treatment selection pressure. This introduces a management paradigm requiring sequential monitoring of advanced prostate cancer patients with plasma and tumor biopsies to ensure early discontinuation of agents when they become potential disease drivers. PMID:25232177

  15. Hearing Screening

    ERIC Educational Resources Information Center

    Johnson-Curiskis, Nanette

    2012-01-01

    Hearing levels are threatened by modern life--headsets for music, rock concerts, traffic noises, etc. It is crucial we know our hearing levels so that we can draw attention to potential problems. This exercise requires that students receive a hearing screening for their benefit as well as for making the connection of hearing to listening.

  16. SCREENING TECHNOLOGIES

    EPA Science Inventory

    Field screening methods are used to determine whether a characteristic of interest is present or absent, above and below a predetermined threshold at a given site, or in a concentration within a predetermined range of interest. or the U.S. EPA and the regulated community much emp...

  17. Lethal and sub lethal effects of the biocide chlorhexidine on aquatic organisms.

    PubMed

    Jesus, Fátima T; Oliveira, Rhaul; Silva, Andreia; Catarino, Ana L; Soares, Amadeu M V M; Nogueira, António J A; Domingues, Inês

    2013-11-01

    Chlorhexidine is among the most used biocides in Europe, however its toxicity to aquatic organisms is scarcely known. The main objective of this study was to assess the lethal and sub lethal effects of chlorhexidine digluconate (ChD) on four aquatic model organisms: the bacteria Vibrio fischeri, the algae Pseudokirchneriella subcapitata, the crustacean Daphnia magna and the embryos of the fish Danio rerio. ChD was very toxic to algae and crustaceans, with a 72 h-EC50 of 62.2 ?g/l and a 48 h-EC50 of 45.0 ?g/l, respectively. Toxicity to fish embryos and the bacteria was lower, with a 96 h-EC50 of 804.0 ?g/l and a 15 min-EC50 of 1,694.0 ?g/l, respectively. Concerning sub lethal effects on D. magna (feeding inhibition) a 6 h-EC50 of 503.7 ?g/l was obtained. In fish, ChD caused developmental abnormalities, namely alterations in the amniotic fluid (48 h-EC20 of 753.6 ?g/l) and early hatching. Moreover, enzymatic biomarkers on fish embryos showed an induction of cholinesterase activity in all ChD tested concentrations (80-900 ?g/l). The catalase activity was also induced at the highest concentration tested (900 ?g/l) whereas no changes were observed for glutathione-S-transferase and lactate dehydrogenase activities. The toxicity of ChD to the algae and crustacean raises concerns about its potential effects in aquatic food webs, since these organisms are in the base of trophic chains, and highlights the need for further studies on ChD toxicity to aquatic organisms. PMID:24026526

  18. [The characteristics of the expression of temperature-dependent dominant lethal mutations in Drosophila melanogaster].

    PubMed

    Miasniankina, E N; Generalova, M V

    1993-01-01

    The results of a series of investigations on obtaining and analysing dominant temperature-sensitive lethals (DTS-lethals) are summarized. Using EMS, both cold- and heat sensitive mutations of this type were induced in large autosomes. The cold-sensitive mutations of chromosome 3 were revealed for the first time. The effect of genetic phone on DTS lethal penetrance and expressivity was studied. The character of expression of two mutations possessing a pleiotropic action was examined. The heat-sensitive 1(2)M90DTS mutation disturbed the structure of abdominal tergites in all imago and, besides, caused an abrupt decrease in the female reproductive period. The pattern of alterations observed in oogenesis along with the result of phenogenetic analysis suggest that this gene takes part in the genetic control over the proliferation of stem oogonial cells. The cold-sensitive 1(2)M66DCS mutation caused a number of disturbances in imago thoracic structures: inability to flight, raised up wings, extremity fracture, etc. Mutant flies showed serious disorders of indirect flight muscles both at morphological and at ultrastructural levels. It was established that this mutation was a cold-sensitive allele of Mhc gene which controls the synthesis of myosine heavy chains in drosophila. PMID:8471970

  19. Metabolic Response of Escherichia coli upon Treatment with Hypochlorite at Sub-Lethal Concentrations

    PubMed Central

    Winter, Jeannette; Eisenreich, Wolfgang

    2015-01-01

    Hypochlorite is a reactive oxygen species that is worldwide as an antibacterial disinfectant. Hypochlorite exposure is known to cause oxidative damage to DNA and proteins. As a response to these effects, the metabolite profiles of organisms treated with sub-lethal doses of hypochlorite are assumed to be severely modified; however, the nature of these changes is hardly understood. Therefore, using nuclear magnetic resonance spectroscopy and gas chromatography-coupled mass spectrometry, we analyzed the time-dependent impact of hypochlorite exposure with a sub-lethal concentration (50 µM) on the metabolite profile of the Escherichia coli strain MG1655. Principle component analysis clearly distinguished between the metabolite profiles of bacteria treated for 0, 5,10, 20, 40, or 60 min. Major changes in the relative amounts of fatty acids, acetic acid, and formic acid occurred within the first 5 min. Comparative gas chromatography-coupled mass spectrometry analyses revealed that the amounts of free methionine and alanine were significantly decreased in the treated cells, demonstrating their susceptibility to hypochlorite exposure. The concentrations of succinate, urea, orotic acid, 2-aminobutyric acid, and 2-hydroxybutyric acid were also severely affected, indicating general changes in the metabolic network by hypochlorite. However, most metabolite levels relaxed to the reference values of untreated cells after 40–60 min, reflecting the capability of E. coli to rapidly adapt to environmental stress factors such as the presence of sub-lethal oxidant levels. PMID:25932918

  20. Exploiting synthetic lethal interactions for targeted cancer therapy

    E-print Network

    Jiang, Hai

    Emerging data suggests that synthetic lethal interactions between mutated oncogenes/tumor suppressor genes and molecules involved in DNA damage signaling and repair can be therapeutically exploited to preferentially kill ...

  1. Anthrax lethal factor inhibitors as potential countermeasure of the infection.

    PubMed

    Kumar, B V S Suneel; Malik, Siddharth; Grandhi, Pradeep; Dayam, Raveendra; Sarma, J A R P

    2014-01-01

    Anthrax Lethal Factor (LF) is a zinc-dependent metalloprotease, one of the virulence factor of anthrax infection. Three forms of the anthrax infection have been identified: cutaneous (through skin), gastrointestinal (through alimentary tract), and pulmonary (by inhalation of spores). Anthrax toxin is composed of protective antigen (PA), lethal factor (LF), and edema factor (EF). Protective antigen mediates the entry of Lethal Factor/Edema Factor into the cytosol of host cells. Lethal factor (LF) inactivates mitogen-activated protein kinase kinase inducing cell death, and EF is an adenylyl cyclase impairing host defenses. In the past few years, extensive studies are undertaken to design inhibitors targeting LF. The current review focuses on the small molecule inhibitors targeting LF activity and its structure activity relationships (SAR). PMID:25262802

  2. Lethal and sublethal effects of cypermethrin to Hypsiboas pulchellus tadpoles

    Microsoft Academic Search

    M. Gabriela Agostini; Guillermo S. Natale; Alicia E. Ronco

    2010-01-01

    The study of the effects of the insecticide cypermethrin (CY) technical grade and its Sherpa® commercial formulation on Hypsiboas pulchellus tadpoles assessing lethality, behavior, growth, and abnormalities under standardized laboratory conditions is reported. Observed\\u000a behaviors were identified and categorized by means of a ranking system according to the loss of mobility. Results of acute\\u000a lethal effects indicate higher potency for

  3. Internal Lethal Concentrations of Halobenzenes with Fish ( Gambusia affinis)

    Microsoft Academic Search

    Yupadee Chaisuksant; Qiming Yu; Des Connell

    1997-01-01

    The internal lethal concentration is a potential measure of toxicity which could be usefully applied in environmental toxicology and risk assessment. Using halobenzenes, which are common environmental contaminants, and represented test compounds, experiments were conducted in aquaria with the mosquitofish (Gambusia affinis). The average internal lethal concentration (ILC50) for four representative halohydrocarbons, 1,4-diBB, 1,2,3-triCB, 1,2,4-triBB, and pentaCB, were consistent with

  4. Production and proteolytic assay of lethal factor from Bacillus anthracis

    Microsoft Academic Search

    Joungmok Kim; Young-Myung Kim; Bon-Sung Koo; Young-Kyu Chae; Moon-Young Yoon

    2003-01-01

    Bacillus anthracis is the causative agent of anthrax. The major virulence factors are a poly-d-glutamic acid capsule and three-protein component exotoxin, protective antigen (PA, 83kDa), lethal factor (LF, 90kDa), and edema factor (EF, 89kDa), respectively. These three proteins individually have no known toxic activities, but in combination with PA form two toxins (lethal toxin or edema toxin), causing different pathogenic

  5. Purified Influenza Virus Nucleoprotein Protects Mice from Lethal Infection

    Microsoft Academic Search

    DAVID C. WRAITH; ANGELA E. VESSEY; BRIGITTE A. ASKONAS

    1987-01-01

    SUMMARY Local administration of nucleoprotein purified from X31 (H3N2) influenza A virus primed for A virus cross-reactive cytotoxic T cells and resulted in substantial protection (75 ~) of mice from a lethal challenge with the heterologous mouse-adapted A\\/PR\\/8\\/34 (H1N1) virus. By following the course of a lethal virus challenge we found that nucleoprotein priming did not prevent virus infection but

  6. Low molecular weight inhibitors of the protease anthrax lethal factor.

    PubMed

    Dalkas, Georgios A; Papakyriakou, Athanasios; Vlamis-Gardikas, Alexios; Spyroulias, Georgios A

    2008-03-01

    Anthrax Lethal Factor (LF) is a zinc-dependent metalloprotease that together with the protective antigen constitute the anthrax lethal toxin, the most prominent virulence factor of the disease anthrax. This review summarizes the current knowledge on anthrax toxicity and defense in relation to LF. Particular emphasis is placed on the structural aspects of LF, the properties of its substrates and the achievements in the design of low molecular weight inhibitors of the catalytic activity of the metalloenzyme. PMID:18336349

  7. Late-acting dominant lethal genetic systems and mosquito control

    PubMed Central

    Phuc, Hoang Kim; Andreasen, Morten H; Burton, Rosemary S; Vass, Céline; Epton, Matthew J; Pape, Gavin; Fu, Guoliang; Condon, Kirsty C; Scaife, Sarah; Donnelly, Christl A; Coleman, Paul G; White-Cooper, Helen; Alphey, Luke

    2007-01-01

    Background Reduction or elimination of vector populations will tend to reduce or eliminate transmission of vector-borne diseases. One potential method for environmentally-friendly, species-specific population control is the Sterile Insect Technique (SIT). SIT has not been widely used against insect disease vectors such as mosquitoes, in part because of various practical difficulties in rearing, sterilization and distribution. Additionally, vector populations with strong density-dependent effects will tend to be resistant to SIT-based control as the population-reducing effect of induced sterility will tend to be offset by reduced density-dependent mortality. Results We investigated by mathematical modeling the effect of manipulating the stage of development at which death occurs (lethal phase) in an SIT program against a density-dependence-limited insect population. We found late-acting lethality to be considerably more effective than early-acting lethality. No such strains of a vector insect have been described, so as a proof-of-principle we constructed a strain of the principal vector of the dengue and yellow fever viruses, Aedes (Stegomyia) aegypti, with the necessary properties of dominant, repressible, highly penetrant, late-acting lethality. Conclusion Conventional SIT induces early-acting (embryonic) lethality, but genetic methods potentially allow the lethal phase to be tailored to the program. For insects with strong density-dependence, we show that lethality after the density-dependent phase would be a considerable improvement over conventional methods. For density-dependent parameters estimated from field data for Aedes aegypti, the critical release ratio for population elimination is modeled to be 27% to 540% greater for early-acting rather than late-acting lethality. Our success in developing a mosquito strain with the key features that the modeling indicated were desirable demonstrates the feasibility of this approach for improved SIT for disease control. PMID:17374148

  8. Mechanism by Which Caffeine Potentiates Lethality of Nitrogen Mustard

    Microsoft Academic Search

    Ching C. Lau; Arthur B. Pardee

    1982-01-01

    Caffeine is synergistic with many DNA-damaging agents in increasing lethality to mammalian cells. The mechanism is not well understood. Our results show that caffeine potentiates the lethality of the nitrogen mustard 2-chloro-N-(2-chloroethyl)-N-methylethanamine (HN2) by inducing damaged cells to undergo mitosis before properly repairing lesions in their DNA. Treatment with low doses of HN2 (0.5 mu M for 1 hr) caused

  9. Newborn Screening

    NSDL National Science Digital Library

    American Association for the Advancement of Science (; )

    2005-03-07

    There's been a lot of talk recently about national testing standards for students. But now, some doctors are calling for national testing standards for newborns. In some states, every newborn baby gets a blood test that can detect more than thirty serious diseases. But in other states, it's a very different story. According to geneticist Piero Rinaldo of the Mayo Clinic, just having a state senator with an affected child could make the difference. Now, he and dozens of other experts have developed uniform standards for newborn screening. And they're calling on the federal government to adopt them. This Science update details many of the obstacles by which individual states and the U.S. government face when challenged with newborn screening practices. Additional links for further inquiry are provided.

  10. Vision Screening

    NASA Technical Reports Server (NTRS)

    1993-01-01

    The Visi Screen OSS-C, marketed by Vision Research Corporation, incorporates image processing technology originally developed by Marshall Space Flight Center. Its advantage in eye screening is speed. Because it requires no response from a subject, it can be used to detect eye problems in very young children. An electronic flash from a 35 millimeter camera sends light into a child's eyes, which is reflected back to the camera lens. The photorefractor then analyzes the retinal reflexes generated and produces an image of the child's eyes, which enables a trained observer to identify any defects. The device is used by pediatricians, day care centers and civic organizations that concentrate on children with special needs.

  11. Revertant Mutation Releases Confined Lethal Mutation, Opening Pandora's Box: A Novel Genetic Pathogenesis

    PubMed Central

    Ogawa, Yasushi; Takeichi, Takuya; Kono, Michihiro; Hamajima, Nobuyuki; Yamamoto, Toshimichi; Sugiura, Kazumitsu; Akiyama, Masashi

    2014-01-01

    When two mutations, one dominant pathogenic and the other “confining” nonsense, coexist in the same allele, theoretically, reversion of the latter may elicit a disease, like the opening of Pandora's box. However, cases of this hypothetical pathogenic mechanism have never been reported. We describe a lethal form of keratitis-ichthyosis-deafness (KID) syndrome caused by the reversion of the GJB2 nonsense mutation p.Tyr136X that would otherwise have confined the effect of another dominant lethal mutation, p.Gly45Glu, in the same allele. The patient's mother had the identical misssense mutation which was confined by the nonsense mutation. The biological relationship between the parents and the child was confirmed by genotyping of 15 short tandem repeat loci. Haplotype analysis using 40 SNPs spanning the >39 kbp region surrounding the GJB2 gene and an extended SNP microarray analysis spanning 83,483 SNPs throughout chromosome 13 in the family showed that an allelic recombination event involving the maternal allele carrying the mutations generated the pathogenic allele unique to the patient, although the possibility of coincidental accumulation of spontaneous point mutations cannot be completely excluded. Previous reports and our mutation screening support that p.Gly45Glu is in complete linkage disequilibrium with p.Tyr136X in the Japanese population. Estimated from statisitics in the literature, there may be approximately 11,000 p.Gly45Glu carriers in the Japanese population who have this second-site confining mutation, which acts as natural genetic protection from the lethal disease. The reversion-triggered onset of the disesase shown in this study is a previously unreported genetic pathogenesis based on Mendelian inheritance. PMID:24785414

  12. Lung cancer screening: rationale and background

    PubMed Central

    2011-01-01

    Abstract The poor outcome in symptomatic lung cancer patients and the much better prognosis when lung cancer is diagnosed and treated at early asymptomatic stages call for screening. As lung cancer predominantly affects smokers and individuals exposed to other carcinogens, screening programs need not include the whole population but only these risk groups. Every screening program will tend to better identify the more indolent tumours that grow slowly enough to be detected by screening before symptoms develop, whereas aggressive fast-growing tumours may present as interval cancers despite screening (length-time bias). Some malignant tumours detected with screening may never cause the person’s death due to competing causes for death, particularly in heavy smokers, such as cardiovascular disease or other cancers (overdiagnosis bias). If a cancer is still lethal despite detection through screening, the affected individual may live longer with the diagnosis of cancer but not longer altogether (lead-time bias). It is likely that this will have a negative effect on that individual’s quality of life. Participation in screening programs may have beneficial as well as adverse effects on smoking habits; in the worst case it may encourage people to continue smoking. Trials assessing chest radiography or sputum microscopy have not demonstrated a reduction in lung cancer mortality through screening, probably because the tests were not sensitive enough. computed tomography promises better sensitivity. Other modern tests such as fibre optic bronchoscopy, analysis of molecular markers or genetic testing in serum, sputum or exhaled air are not yet ready for clinical practice. PMID:22185788

  13. Quadruple screen test

    MedlinePLUS

    ... screen; Multiple marker screening; AFP plus; Triple screen test; AFP maternal; MSAFP; 4-marker screen ... This test is most often done between the 15th and 22nd weeks of the pregnancy. It is most accurate ...

  14. Lung Cancer Screening

    MedlinePLUS

    ... Prevention Lung Cancer Screening Lung Cancer Research Lung Cancer Screening (PDQ®) What is screening? Screening is looking ... are called diagnostic tests . General Information About Lung Cancer Key Points Lung cancer is a disease in ...

  15. Skin Cancer Screening

    MedlinePLUS

    ... Cancer Skin Cancer Screening Skin Cancer Research Skin Cancer Screening (PDQ®) What is screening? Screening is looking ... are called diagnostic tests . General Information About Skin Cancer Key Points Skin cancer is a disease in ...

  16. Protection against Anthrax Lethal Toxin Challenge by Genetic Immunization with a Plasmid Encoding the Lethal Factor Protein

    Microsoft Academic Search

    BRIAN M. PRICE; ADRIANE L. LINER; STEPHEN H. LEPPLA; ALFRED MATECZUN; DARRELL R. GALLOWAY

    2001-01-01

    The ability of genetic vaccination to protect against a lethal challenge of anthrax toxin was evaluated. BALB\\/c mice were immunized via gene gun inoculation with eucaryotic expression vector plasmids encoding either a fragment of the protective antigen (PA) or a fragment of lethal factor (LF). Plasmid pCLF4 contains the N-terminal region (amino acids (aa) 10 to 254) of Bacillus anthracis

  17. Differential screening of mitochondrial cDNA libraries from male-fertile and cytoplasmic male-sterile sugar-beet reveals genome rearrangements at atp6 and atpA loci

    Microsoft Academic Search

    Yongbiao Xue; Sylvie Collin; D. Roy Davies; Colwyn M. Thomas

    1994-01-01

    As part of a strategy to define differences in genome organization and expression between cytoplasmic male-sterile (CMS) and male-fertile (MF) sugar-beet mitochondria, cDNA libraries from both mitochondrial genotypes were constructed. Preliminary screening with ribosomal RNA gene probes identified candidate cDNA clones corresponding to structural genes. In addition, reciprocal hybridization experiments were performed using labelled first-strand cDNA to identify uniquely transcribed

  18. Screening for skin cancer

    Microsoft Academic Search

    Mark Helfand; Susan M Mahon; Karen B Eden; Paul S Frame; C. Tracy Orleans

    2001-01-01

    Context: Malignant melanoma is often lethal, and its incidence in the United States has increased rapidly over the past 2 decades. Nonmelanoma skin cancer is seldom lethal, but, if advanced, can cause severe disfigurement and morbidity. Early detection and treatment of melanoma might reduce mortality, while early detection and treatment of nonmelanoma skin cancer might prevent major disfigurement and to

  19. Bacterial community composition of three candidate insect vectors of palm phytoplasma (Texas Phoenix Palm Decline and Lethal Yellowing).

    PubMed

    Powell, Christopher M; Hail, Daymon; Potocnjak, Julia; Hanson, J Delton; Halbert, Susan H; Bextine, Blake R

    2015-02-01

    Texas Phoenix Palm Decline (TPPD) and Lethal Yellowing are two phytoplasma-linked diseases in palms. The phytoplasma causing TPPD is thought to be transmitted by three putative planthopper vectors, Ormenaria rufifascia, Omolicna joi, and Haplaxius crudus. These insects have been morphologically and molecularly described, and have screened positive for Candidatus Phytoplasma palmae. Individuals from each species were subjected to 16S bacterial community sequencing using the Roche 454 platform, providing new information regarding the previously unexplored bacterial communities present in putative vectors. PMID:25298076

  20. Determinants of the lethality of climate-related disasters in the Caribbean Community (CARICOM): a cross-country analysis.

    PubMed

    Andrewin, Aisha N; Rodriguez-Llanes, Jose M; Guha-Sapir, Debarati

    2015-01-01

    Floods and storms are climate-related hazards posing high mortality risk to Caribbean Community (CARICOM) nations. However risk factors for their lethality remain untested. We conducted an ecological study investigating risk factors for flood and storm lethality in CARICOM nations for the period 1980-2012. Lethality - deaths versus no deaths per disaster event- was the outcome. We examined biophysical and social vulnerability proxies and a decadal effect as predictors. We developed our regression model via multivariate analysis using a generalized logistic regression model with quasi-binomial distribution; removal of multi-collinear variables and backward elimination. Robustness was checked through subset analysis. We found significant positive associations between lethality, percentage of total land dedicated to agriculture (odds ratio [OR] 1.032; 95% CI: 1.013-1.053) and percentage urban population (OR 1.029, 95% CI 1.003-1.057). Deaths were more likely in the 2000-2012 period versus 1980-1989 (OR 3.708, 95% CI 1.615-8.737). Robustness checks revealed similar coefficients and directions of association. Population health in CARICOM nations is being increasingly impacted by climate-related disasters connected to increasing urbanization and land use patterns. Our findings support the evidence base for setting sustainable development goals (SDG). PMID:26153115

  1. Determinants of the lethality of climate-related disasters in the Caribbean Community (CARICOM): a cross-country analysis

    PubMed Central

    Andrewin, Aisha N.; Rodriguez-Llanes, Jose M.; Guha-Sapir, Debarati

    2015-01-01

    Floods and storms are climate-related hazards posing high mortality risk to Caribbean Community (CARICOM) nations. However risk factors for their lethality remain untested. We conducted an ecological study investigating risk factors for flood and storm lethality in CARICOM nations for the period 1980–2012. Lethality - deaths versus no deaths per disaster event- was the outcome. We examined biophysical and social vulnerability proxies and a decadal effect as predictors. We developed our regression model via multivariate analysis using a generalized logistic regression model with quasi-binomial distribution; removal of multi-collinear variables and backward elimination. Robustness was checked through subset analysis. We found significant positive associations between lethality, percentage of total land dedicated to agriculture (odds ratio [OR] 1.032; 95% CI: 1.013–1.053) and percentage urban population (OR 1.029, 95% CI 1.003–1.057). Deaths were more likely in the 2000–2012 period versus 1980–1989 (OR 3.708, 95% CI 1.615–8.737). Robustness checks revealed similar coefficients and directions of association. Population health in CARICOM nations is being increasingly impacted by climate-related disasters connected to increasing urbanization and land use patterns. Our findings support the evidence base for setting sustainable development goals (SDG). PMID:26153115

  2. Functional Genomics Analysis of the Saccharomyces cerevisiae Iron Responsive Transcription Factor Aft1 Reveals Iron-Independent Functions

    PubMed Central

    Berthelet, Sharon; Usher, Jane; Shulist, Kristian; Hamza, Akil; Maltez, Nancy; Johnston, Anne; Fong, Ying; Harris, Linda J.; Baetz, Kristin

    2010-01-01

    The Saccharomyces cerevisiae transcription factor Aft1 is activated in iron-deficient cells to induce the expression of iron regulon genes, which coordinate the increase of iron uptake and remodel cellular metabolism to survive low-iron conditions. In addition, Aft1 has been implicated in numerous cellular processes including cell-cycle progression and chromosome stability; however, it is unclear if all cellular effects of Aft1 are mediated through iron homeostasis. To further investigate the cellular processes affected by Aft1, we identified >70 deletion mutants that are sensitive to perturbations in AFT1 levels using genome-wide synthetic lethal and synthetic dosage lethal screens. Our genetic network reveals that Aft1 affects a diverse range of cellular processes, including the RIM101 pH pathway, cell-wall stability, DNA damage, protein transport, chromosome stability, and mitochondrial function. Surprisingly, only a subset of mutants identified are sensitive to extracellular iron fluctuations or display genetic interactions with mutants of iron regulon genes AFT2 or FET3. We demonstrate that Aft1 works in parallel with the RIM101 pH pathway and the role of Aft1 in DNA damage repair is mediated by iron. In contrast, through both directed studies and microarray transcriptional profiling, we show that the role of Aft1 in chromosome maintenance and benomyl resistance is independent of its iron regulatory role, potentially through a nontranscriptional mechanism. PMID:20439772

  3. Vancomycin minimum inhibitory concentrations and lethality in Staphylococcus aureus bacteremia

    PubMed Central

    Sulla, Felipe; Bussius, Daniel T.; Acquesta, Felipe; Navarini, Alessandra; Sasagawa, Suzethe M.; Mimica, Marcelo J.

    2015-01-01

    Background After the dissemination of penicillin and oxacillin resistance in Staphylococcus aureus, vancomycin-intermediate and vancomycin resistant isolates have been reported. Even between isolates with minimum inhibitory concentrations (MICs) within the susceptible range, some authors have demonstrated that higher MICs correlate with higher lethality. Methods To test this hypothesis in our setting, we compared vancomycin MICs evaluated by two methods and clinical outcomes in hospitalized patients with S. aureus bacteremia. Results We compared lethality in patients infected with isolates that had MICs under or over 2 mg/L. Among patients infected with isolates that had microdilution MICs <2 mg/L, the lethality was 25%; among patients infected with strains that had microdilution MICs ?2 mg/L, 33% died. Among patients infected with isolates that had Etest MICs <2 mg/L, 23% died; in comparison, patients infected with strains that had Etest MICs ?2 mg/L had a lethality of 44%. Conclusion Our results showed a slight tendency of higher lethality when higher MICs were present. However, this difference did not reach statistical significance, possibly due to the relatively small number of patients included in the study. Future prospective studies are needed to further evaluate this correlation and to help clinicians guide antimicrobial therapy. PMID:26097833

  4. On quantifying nonthermal effects on the lethality of pressure-assisted heat preservation processes.

    PubMed

    Peleg, Micha; Corradini, Maria G; Normand, Mark D

    2012-01-01

    Direct experimental identification and quantification of the pressure contribution to a pressure-assisted sterilization process efficacy is difficult. However, dynamic kinetic models of thermal inactivation can be used to assess the lethality of a purely thermal process having the same temperature profile. Thus, a pressure-assisted process' temperature record can be used to generate a corresponding purely thermal survival curve with parameters determined in conventional heating experiments. Comparison of the actual final survival ratio with that calculated for the purely thermal process would reveal whether the hydrostatic pressure had synergistic or antagonistic effect on bacterial spores survival. The effect would be manifested in the number of log cycles subtracted or added to the survival ratio, and in the length of time at the holding temperature needed to produce the final survival ratio of the combined process. A set of combined treatments would reveal how the temperature and pressure profiles affect the pressure's influence on the process' lethality to either vegetative cells or spores. The need to withdraw samples during the thermal and combined processes would be avoided if the thermal survival parameters could be calculated by the "three endpoints method," which does not require the entire survival curve determination. Currently however, this method is limited to thermal inactivation patterns characterized by up to 3 survival parameters, the Weibull-Log logistic (WeLL) model, for example. PMID:22260125

  5. A Survey of New Temperature-Sensitive, Embryonic-Lethal Mutations in C. elegans: 24 Alleles of Thirteen Genes

    PubMed Central

    O'Rourke, Sean M.; Garner, Aleena R.; Hamill, Danielle R.; Osterberg, Valerie R.; Lyczak, Rebecca; Madison, Erin E.; Nguyen, Michael H.; Sandberg, Nathan A.; Sedghi, Noushin; Willis, John H.; Yochem, John; Johnson, Eric A.; Bowerman, Bruce

    2011-01-01

    To study essential maternal gene requirements in the early C. elegans embryo, we have screened for temperature-sensitive, embryonic lethal mutations in an effort to bypass essential zygotic requirements for such genes during larval and adult germline development. With conditional alleles, multiple essential requirements can be examined by shifting at different times from the permissive temperature of 15°C to the restrictive temperature of 26°C. Here we describe 24 conditional mutations that affect 13 different loci and report the identity of the gene mutations responsible for the conditional lethality in 22 of the mutants. All but four are mis-sense mutations, with two mutations affecting splice sites, another creating an in-frame deletion, and one creating a premature stop codon. Almost all of the mis-sense mutations affect residues conserved in orthologs, and thus may be useful for engineering conditional mutations in other organisms. We find that 62% of the mutants display additional phenotypes when shifted to the restrictive temperature as L1 larvae, in addition to causing embryonic lethality after L4 upshifts. Remarkably, we also found that 13 out of the 24 mutations appear to be fast-acting, making them particularly useful for careful dissection of multiple essential requirements. Our findings highlight the value of C. elegans for identifying useful temperature-sensitive mutations in essential genes, and provide new insights into the requirements for some of the affected loci. PMID:21390299

  6. Genome-wide screening of human T-cell epitopes in influenza A virus reveals a broad-spectrum of CD4+ T cell responses to internal proteins, hemagglutinins and neuraminidases

    PubMed Central

    Babon, Jenny Aurielle B.; Cruz, John; Orphin, Laura; Pazoles, Pamela; Co, Mary Dawn T.; Ennis, Francis A.; Terajima, Masanori

    2009-01-01

    We performed a genome-wide screening for T cell epitopes using synthetic peptides that encompass all of the influenza A viral proteins, including subtype variants for hemagglutinin (HA) (H1, H3 and H5) and neuraminidase (NA) (human and avian N1 and N2) proteins, based on the sequence information of recently circulating strains. We found a total of 83 peptides, 54 of them novel, to which specific T cells were detectable in IFN-? ELISPOT assays using peripheral blood mononuclear cells from four healthy adult donors. The surface glycoproteins, HA and NA, major components of vaccines, had many T cell epitopes. HA and matrix protein 1 had more T cell epitopes than other viral proteins, most of which were recognized by CD4+ T cells. We established several cytotoxic CD4+ T cell lines from these donors. We also analyzed H1 and H3 HA-specific T cell responses using the peripheral blood mononuclear cells of 30 hospital workers. 53% of donors gave a positive response to H3 HA peptides, while 17% gave a positive response to H1 HA peptides. Our genome-wide screening is useful in identifying T cell epitopes and complementary to the approach based on the predicted binding peptides to well-studied HLA-A, B and DR alleles. PMID:19524006

  7. Lethal effects of short-wavelength visible light on insects

    PubMed Central

    Hori, Masatoshi; Shibuya, Kazuki; Sato, Mitsunari; Saito, Yoshino

    2014-01-01

    We investigated the lethal effects of visible light on insects by using light-emitting diodes (LEDs). The toxic effects of ultraviolet (UV) light, particularly shortwave (i.e., UVB and UVC) light, on organisms are well known. However, the effects of irradiation with visible light remain unclear, although shorter wavelengths are known to be more lethal. Irradiation with visible light is not thought to cause mortality in complex animals including insects. Here, however, we found that irradiation with short-wavelength visible (blue) light killed eggs, larvae, pupae, and adults of Drosophila melanogaster. Blue light was also lethal to mosquitoes and flour beetles, but the effective wavelength at which mortality occurred differed among the insect species. Our findings suggest that highly toxic wavelengths of visible light are species-specific in insects, and that shorter wavelengths are not always more toxic. For some animals, such as insects, blue light is more harmful than UV light. PMID:25488603

  8. Advantages of less-tech, less-than-lethal technologies

    NASA Astrophysics Data System (ADS)

    Marts, Donna J.; Overlin, Trudy K.

    1995-05-01

    This paper illustrates the advantages of developing less-tech technologies by reporting on two less-tech, less-than-lethal prototype law enforcement tools developed at the Idaho National Engineering Laboratory. The devices were developed for the National Institute of Justice, less- than-lethal weapons program: 1) an air bag restraint device for use in restraining suspects who become violent during transport in patrol vehicles, and 2) a retractable spiked barrier strip for stopping fleeing vehicles during high-speed pursuit. The success of both projects relied on developing design requirements in conjunction with the actual users of the devices.

  9. Medicare Preventive and Screening Services

    MedlinePLUS

    ... Hepatitis C screening test HIV screening Mammograms (screening) Nutrition therapy services Obesity screenings & counseling One-time “Welcome to Medicare” preventive visit Prostate cancer screenings Sexually transmitted infections screening & counseling Shots: Flu ...

  10. Risks of Breast Cancer Screening

    MedlinePLUS

    ... of Breast & Gynecologic Cancers Breast Cancer Screening Research Breast Cancer Screening (PDQ®) What is screening? Screening is looking ... cancer screening: Cancer Screening Overview General Information About Breast Cancer Key Points Breast cancer is a disease in ...

  11. Antidotes to anthrax lethal factor intoxication. Part 3: Evaluation of core structures and further modifications to the C2-side chain

    PubMed Central

    Jiao, Guan-Sheng; Kim, Seongjin; Moayeri, Mahtab; Crown, Devorah; Thai, April; Cregar-Hernandez, Lynne; McKasson, Linda; Sankaran, Banumathi; Lehrer, Axel; Wong, Teri; Johns, Lisa; Margosiak, Stephen A.; Leppla, Stephen H.; Johnson, Alan T.

    2012-01-01

    Four core structures capable of providing sub-nanomolar inhibitors of anthrax lethal factor (LF) were evaluated by comparing the potential for toxicity, physicochemical properties, in vitro ADME profiles, and relative efficacy in a rat lethal toxin (LT) model of LF intoxication. Poor efficacy in the rat LT model exhibited by the phenoxyacetic acid series (3) correlated with low rat microsome and plasma stability. Specific molecular interactions contributing to the high affinity of inhibitors with a secondary amine in the C2-side chain were revealed by x-ray crystallography. PMID:22342144

  12. Strategies for Molecularly Enhanced Chemotherapy to Achieve Synthetic Lethality in Endometrial Tumors with Mutant p53

    PubMed Central

    Meng, Xiangbing; Dizon, Don S.; Yang, Shujie; Wang, Xinjun; Zhu, Danlin; Thiel, Kristina W.; Leslie, Kimberly K.

    2013-01-01

    Serous uterine endometrial carcinomas are aggressive type II cancers with poor outcomes for which new treatment strategies are urgently needed, in particular, strategies that augment sensitivity to established chemotherapy regimens. The tumor suppressor gene TP53 is dysregulated in more than 90% of serous tumors, altering master regulators of the G2/M cell cycle checkpoint in unique and predictable ways and desensitizing cells to chemotherapy. We hypothesized that synthetic lethality can be achieved in endometrial cancer cells with mutant p53 by combining paclitaxel with agents to overcome G2/M arrest and induce mitotic catastrophe. The combination of BIBF1120, an investigational VEGFR, PDGFR, and FGFR multityrosine kinase inhibitor with established anti-angiogenic activity, with paclitaxel abrogated the G2/M checkpoint in p53-null endometrial cancer cells via modulation of G2/M checkpoint regulators followed by induction of mitotic cell death. In endometrial cancer cells harboring an oncogenic gain-of-function p53 mutation, synthetic lethality was created by combining paclitaxel with BIBF1120 and a histone deacetylase inhibitor, which serves to destabilize mutant p53. These cells were also sensitive to an inhibitor of the G2/M kinase Wee1 in combination with paclitaxel. These findings reveal that, in addition to antiangiogenic activity, the angiokinase inhibitor BIBF1120 can be used to restore sensitivity to paclitaxel and induce mitotic cell death in endometrial cancer cells with non-functional p53. These preclinical data serve as a critical platform for the creative design of future clinical trials utilizing molecularly enhanced chemotherapy to achieve synthetic lethality based on the mutational landscape. PMID:24381593

  13. Dissociation of lethal toxicity and enzymic activity of notexin from Notechis scutatus scutatus (Australian-tiger-snake) venom by modification of tyrosine residues.

    PubMed Central

    Yang, C C; Chang, L S

    1991-01-01

    Notexin from Notechis scutatus scutatus snake venom was subjected to tyrosine modification with p-nitrobenzenesulphonyl fluoride (NBSF), and four modified derivatives were separated by h.p.l.c. The results of amino acid analysis and sequence determination revealed that only Tyr-7, Tyr-70 and Tyr-77 were modified in notexin. Modification of Tyr-7 resulted in decreases in lethal toxicity and enzymic activity by 70.2% and 22.7% respectively. Conversely, modification of Tyr-77 caused a 1.8-fold increase in enzymic activity, in contrast with the loss of 52.5% of lethality. A drastic decrease in lethal toxicity was observed when both Tyr-7 and Tyr-70 were modified, whereas the enzymic activity decreased by only 35.8%. Likewise, the derivative in which Tyr-7 and Tyr-77 were modified retained 44.4% of enzymic activity, but showed a marked decrease in lethal toxicity. It is obvious that modification of tyrosine residues causes a decrease in lethal toxicity of notexin, which does not directly correlate with the change in enzymic activity. On the other hand, the antigenicity of NBS derivatives remained unchanged. The modified derivatives retained their affinity for Ca2+, indicating that the modified tyrosine residues did not participate in Ca2+ binding. These results indicate that modification of tyrosine residues can differentially influence the enzymic activity and lethal toxicity of notexin, and suggest that notexin might possess two functional sites, one being responsible for the catalytic activity and the other associated with its lethal effect. PMID:1764038

  14. Screen time and children

    MedlinePLUS

    "Screen time" is a term used for activities done in front of a screen, such as watching TV, working on a computer, or playing video games. Screen time is sedentary activity, meaning you are being physically ...

  15. Health Screenings and Immunizations

    MedlinePLUS

    ... and Immunizations: What Are Health Screenings? In This Topic What Are Health Screenings? Recommended Screenings For Women ... for More Information National Institute on Aging Related Topics Talking with Your Doctor The information in this ...

  16. Mental Health Screening Center

    MedlinePLUS

    Mental Health Screening Center These online screening tools are not a substitute for consultation with a health professional. ... you have any concerns, see your doctor or mental health professional. Depression This screening form was developed from ...

  17. Anthrax Toxin Receptor 2Dependent Lethal Toxin Killing In Vivo

    E-print Network

    Sejnowski, Terrence J.

    Anthrax Toxin Receptor 2­Dependent Lethal Toxin Killing In Vivo Heather M. Scobie1,2 , Darran J Jolla, California, United States of America Anthrax toxin receptors 1 and 2 (ANTXR1 and ANTXR2) have by residue D683 of the protective antigen (PA) subunit of anthrax toxin. The receptor-bound metal ion and PA

  18. Bureaucracy, Safety and Software: a potentially lethal cocktail

    E-print Network

    Hatton, Les

    Bureaucracy, Safety and Software: a potentially lethal cocktail Les Hatton CISM, University of software controlled safety critical systems. It observes that the rapid growth of bureaucracy in society of such bureaucracy are generally mitigated because the rules naturally devolve from the exercise of the scientific

  19. The Prevalence, Lethality and Intent of Suicide Attempts among Adolescents.

    ERIC Educational Resources Information Center

    Andrews, Judy A.; Lewinsohn, Peter M.

    Although suicide is the second leading cause of death among adolescents in the United States, little is known about the prevalence or characteristics of suicide attempts among adolescents. Data from 1,710 adolescents attending 9 high schools in 5 communities were examined to determine the prevalence of suicide attempts and the lethality and intent…

  20. Regulation of apoptosis by lethal cytokines in human mesothelial cells

    Microsoft Academic Search

    Marina Penélope Catalan; Dolores Subirá; Ana Reyero; Rafael Selgas; Arturo Ortiz-Gonzalez; Jesús Egido; Alberto Ortiz

    2003-01-01

    Regulation of apoptosis by lethal cytokines in human mesothelial cells.BackgroundDysregulation of peritoneal cell death may contribute to the complications of peritoneal dialysis (PD). Chronic peritoneal dialysis and acute peritonitis are both associated with loss of mesothelial cells. In addition, acute peritonitis is characterized by sudden changes in the number of peritoneal leukocytes. However, the factors regulating peritoneal cell survival are

  1. ACUTE LETHALITY OF COPPER, CADMIUM, AND ZINC TO NORTHERN SQUAWFISH

    EPA Science Inventory

    Flow-through acute toxicity tests on juvenile northern squawfish (Ptychocheilus oregonensis) were conducted with copper, cadmium, and zinc. The 96-hour median lethal concentrations were 18 micrograms/liter for copper, 1,104 micrograms/liter for cadmium, and 3,693 micrograms/liter...

  2. Hyperinsulinaemia: A prospective risk factor for lethal clinical prostate cancer

    Microsoft Academic Search

    Jan Hammarsten; Benkt Högstedt

    2005-01-01

    Previous studies have suggested that hyperinsulinaemia and other components of metabolic syndrome are risk factors for clinical prostate cancer. This prospective study tested the hypothesis that hyperinsulinaemia and other components of metabolic syndrome are risk factors for lethal clinical prostate cancer. The clinical, haemodynamic, anthropometric, metabolic and insulin profile at baseline in men who had died from clinical prostate cancer

  3. Prison Inmate Characteristics and Suicide Attempt Lethality: An Exploratory Study

    Microsoft Academic Search

    Philip R. Magaletta; Marc W. Patry; Ben Wheat; Jeffery Bates

    2008-01-01

    Working with suicidal inmates is among the most demanding elements of clinical practice in corrections, yet few studies regarding the characteristics of prison inmate suicide attempters or their attempts exist. This represents a significant gap as the method of attempt, the prison context, and the resulting lethality of these incidents may be different from attempts made outside of prison. This

  4. HSP70 Protects against TNF-Induced Lethal Inflammatory Shock

    Microsoft Academic Search

    Wim Van Molle; Ben Wielockx; Tina Mahieu; Masuhiro Takada; Takahide Taniguchi; Kenji Sekikawa; Claude Libert

    2002-01-01

    The heat shock (HS) response is a universal response activated after exposure to various stimuli. The major HS protein (HSP) is the 72 kDa HSP70 with strong homology in different eukaryotic species. We demonstrate that HS treatment of mice leads to a strong induction of HSP70 in several organs and confers significant protection against lethality induced by tumor necrosis factor

  5. Help-Seeking Behavior Prior to Nearly Lethal Suicide Attempts.

    ERIC Educational Resources Information Center

    Barnes, Lauren Seymour; Ikeda, Robin M.; Kresnow, Marcie-jo

    2002-01-01

    The association between help-seeking and nearly lethal suicide attempts was evaluated using data from a population-based, case-control study. Measures of help-seeking included type of consultant contacted, and whether suicide was discussed. Findings suggest efforts to better understand the role of help-seeking in suicide prevention deserves…

  6. Lethal Autonomous Systems and the Plight of the Noncombatant

    E-print Network

    Lethal Autonomous Systems and the Plight of the Non­combatant by Ronald Arkin (Georgia Institute gathering, surveil­ lance, reconnaissance, target acquisi­ tion, designation and engagement ca­ pabilities­ manisation of the enemy through the use of derogatory names and epithets; poorly trained or inexperienced

  7. Science or slaughter: need for lethal sampling of sharks.

    PubMed

    Heupel, M R; Simpfendorfer, C A

    2010-10-01

    General consensus among scientists, commercial interests, and the public regarding the status of shark populations is leading to an increasing need for the scientific community to provide information to help guide effective management and conservation actions. Experience from other marine vertebrate taxa suggests that public, political, and media pressures will play an increasingly important part in setting research, management, and conservation priorities. We examined the potential implications of nonscientific influences on shark research. In particular, we considered whether lethal research sampling of sharks is justified. Although lethal sampling comes at a cost to a population, especially for threatened species, the conservation benefits from well-designed studies provide essential data that cannot be collected currently in any other way. Methods that enable nonlethal collection of life-history data on sharks are being developed (e.g., use of blood samples to detect maturity), but in the near future they will not provide widespread or significant benefits. Development of these techniques needs to continue, as does the way in which scientists coordinate their use of material collected during lethal sampling. For almost half of the known shark species there are insufficient data to determine their population status; thus, there is an ongoing need for further collection of scientific data to ensure all shark populations have a future. Shark populations will benefit most when decisions about the use of lethal sampling are made on the basis of scientific evidence that is free from individual, political, public, and media pressures. PMID:20337690

  8. Small Molecule Inhibitors of Anthrax Lethal Factor Toxin

    PubMed Central

    Williams, John D.; Khan, Atiyya R.; Cardinale, Steven C.; Butler, Michelle M.; Bowlin, Terry L.; Peet, Norton P.

    2014-01-01

    This manuscript describes the preparation of new small molecule inhibitors of Bacillus anthracis lethal factor. Our starting point was the symmetrical, bis-quinolinyl compound 1 (NSC 12155). Optimization of one half of this molecule led to new LF inhibitors that were desymmetrized to afford more drug-like compounds. PMID:24290062

  9. An overview of the future of non-lethal weapons.

    PubMed

    Alexander, J B

    2001-01-01

    During the past decade, vast changes have occurred in the geopolitical landscape and the nature of the types of conflicts in which technologically developed countries have been involved. While the threat of conventional war remains, forces have been more frequently deployed in situations that require great restraint. Adversaries are often likely to be elusive and commingled with noncombatants. There has been some shift in public opinion away from tolerance of collateral casualties. Therefore there is a need to be able to apply force while limiting casualties. Non-lethal weapons provide part of the solution. Among the changes that will influence the future have been studies by the US and NATO concerning the use of non-lethal weapons, coincidental with increased funding for their development and testing. New concepts and policies have recently been formalized. Surprisingly, the most strident objections to the implementation of non-lethal weapons have come from organizations that are ostensibly designed to protect non-combatants. These arguments are specious and, while technically and academically challenging, actually serve to foster an environment that will result in the deaths of many more innocent civilians. They misconstrue technology with human intent. The reasons for use of force will not abate. Alternatives to bombs, missiles, tanks and artillery must therefore be found. Non-lethal weapons are not a panacea but do offer the best hope of minimizing casualties while allowing nations or alliances the means to use force in protection of national or regional interests. PMID:11578037

  10. Analysis of Ebola Glycoprotein Sequences from Strains of Varying Lethality

    E-print Network

    Analysis of Ebola Glycoprotein Sequences from Strains of Varying Lethality Biochem 218 Spring 2002 Tammy Doukas tdoukas@stanford.edu I. Background and Significance Ebola hemorrhagic fever is a disease in humans, chimpanzees, and monkeys, caused by infection with Ebola virus, and associated with high

  11. Physicians' Attitudes About Involvement in Lethal Injection for Capital Punishment

    Microsoft Academic Search

    Neil Farber; Elizabeth B. Davis; Joan Weiner; Janine Jordan; E. Gil Boyer; Peter A. Ubel

    2000-01-01

    Background: Physicians could play various roles in car- rying out capital punishment via lethal injection. Medi- cal societies like the American Medical Association (AMA) and American College of Physicians have established which roles are acceptable and which are disallowed. No one has explored physicians' attitudes toward their po- tential roles in this process. Methods: We surveyed physicians about how accept-

  12. America Revealed

    NSDL National Science Digital Library

    2013-04-22

    The tagline on the America Revealed website says it all: "America Revealed explores the hidden patterns and rhythms that make America work." A remarkable series from PBS, the show talks about everything from how fresh seafood is sourced to how farmers combat crop pests. The Stories section includes a collage of images that, when scrolled over, provide accounts from a variety of people and industries. First-time visitors might want to watch the "Introduction to Manufacturing" series, which explores items that are made in the United States. Visitors can also use the Map section to look for stories of note from around the country, from Long Island to Southern California. The Teachers area includes ten lesson plans and links to additional resources. Finally, visitors can click on the Episodes area to watch complete episodes of the program.

  13. A Multivariate Model of Stakeholder Preference for Lethal Cat Management

    PubMed Central

    Wald, Dara M.; Jacobson, Susan K.

    2014-01-01

    Identifying stakeholder beliefs and attitudes is critical for resolving management conflicts. Debate over outdoor cat management is often described as a conflict between two groups, environmental advocates and animal welfare advocates, but little is known about the variables predicting differences among these critical stakeholder groups. We administered a mail survey to randomly selected stakeholders representing both of these groups (n?=?1,596) in Florida, where contention over the management of outdoor cats has been widespread. We used a structural equation model to evaluate stakeholder intention to support non-lethal management. The cognitive hierarchy model predicted that values influenced beliefs, which predicted general and specific attitudes, which in turn, influenced behavioral intentions. We posited that specific attitudes would mediate the effect of general attitudes, beliefs, and values on management support. Model fit statistics suggested that the final model fit the data well (CFI?=?0.94, RMSEA?=?0.062). The final model explained 74% of the variance in management support, and positive attitudes toward lethal management (humaneness) had the largest direct effect on management support. Specific attitudes toward lethal management and general attitudes toward outdoor cats mediated the relationship between positive (p<0.05) and negative cat-related impact beliefs (p<0.05) and support for management. These results supported the specificity hypothesis and the use of the cognitive hierarchy to assess stakeholder intention to support non-lethal cat management. Our findings suggest that stakeholders can simultaneously perceive both positive and negative beliefs about outdoor cats, which influence attitudes toward and support for non-lethal management. PMID:24736744

  14. Lethal poisoning with ethiofencarb and ethanol.

    PubMed

    Al-Samarraie, Muhammad S J; Karinen, Ritva; Rognum, Torleiv; Hasvold, Inger; Opdal Stokke, Mimi; Christophersen, Asbjørg S

    2009-09-01

    Ethiofencarb is one of the carbamate compounds, which are, in general, less toxic than organophosphorus insecticides. This is due to their reversible acetylcholinesterase inhibition and relative inability to cross the blood-brain barrier. Generally, ethiofencarb is regarded to be of low toxicity (LD(50) > 200 mg/kg); however, severe poisoning and death are not uncommon. To our knowledge, no measurements of ethiofencarb and its metabolites in human postmortem whole blood have been published. We present here a case report of fatal ethiofencarb intoxication with quantitative analysis of ethiofencarb and its metabolites in ante- and postmortem blood. In addition, postmortem urine was collected and analyzed. A 56-year-old man, who worked as a gardener, was found in poor condition, sitting in his car seat. He had been vomiting. The man was admitted to the local hospital about 1 h later. At admission, he was conscious, but unable to speak clearly. His condition deteriorated, and he developed severe pulmonary edema. Resuscitation with atropine and adrenaline were attempted, but he died approximately 3 h after admission. The analysis of postmortem peripheral blood revealed 0.12 g/100 mL ethanol, 26.4 mg/L ethiofencarb, 37.9 mg/L ethiofencarbsulfoxide, and 0.9 mg/L ethiofencarbsulfone. Ethanol (0.26 g/100 mL), ethiofencarb, ethiofencarbsulfoxide, and ethiofencarbsulfone were also detected in urine. PMID:19796510

  15. Genome-Wide siRNA Screen Identifies Complementary Signaling Pathways Involved in Listeria Infection and Reveals Different Actin Nucleation Mechanisms during Listeria Cell Invasion and Actin Comet Tail Formation

    PubMed Central

    Kühbacher, Andreas; Emmenlauer, Mario; Rämo, Pauli; Kafai, Natasha; Dehio, Christoph

    2015-01-01

    ABSTRACT Listeria monocytogenes enters nonphagocytic cells by a receptor-mediated mechanism that is dependent on a clathrin-based molecular machinery and actin rearrangements. Bacterial intra- and intercellular movements are also actin dependent and rely on the actin nucleating Arp2/3 complex, which is activated by host-derived nucleation-promoting factors downstream of the cell receptor Met during entry and by the bacterial nucleation-promoting factor ActA during comet tail formation. By genome-wide small interfering RNA (siRNA) screening for host factors involved in bacterial infection, we identified diverse cellular signaling networks and protein complexes that support or limit these processes. In addition, we could precise previously described molecular pathways involved in Listeria invasion. In particular our results show that the requirements for actin nucleators during Listeria entry and actin comet tail formation are different. Knockdown of several actin nucleators, including SPIRE2, reduced bacterial invasion while not affecting the generation of comet tails. Most interestingly, we observed that in contrast to our expectations, not all of the seven subunits of the Arp2/3 complex are required for Listeria entry into cells or actin tail formation and that the subunit requirements for each of these processes differ, highlighting a previously unsuspected versatility in Arp2/3 complex composition and function. PMID:25991686

  16. Strategy for enhanced transgenic strain development for embryonic conditionnal lethality in Anastrepha suspensa

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Here the first reproductive sterility system for the tephritid pest, Anastrepha suspensa, is presented, based on lethality primarily in embryos heterozygous for a lethal conditional transgene combination. The tetracycline-suppressible system uses the cellularization-specific A. suspensa serendipity...

  17. IL-6 trans-signaling promotes pancreatitis-associated lung injury and lethality.

    PubMed

    Zhang, Hong; Neuhöfer, Patrick; Song, Liang; Rabe, Björn; Lesina, Marina; Kurkowski, Magdalena U; Treiber, Matthias; Wartmann, Thomas; Regnér, Sara; Thorlacius, Henrik; Saur, Dieter; Weirich, Gregor; Yoshimura, Akihiko; Halangk, Walter; Mizgerd, Joseph P; Schmid, Roland M; Rose-John, Stefan; Algül, Hana

    2013-03-01

    Acute lung injury (ALI) is an inflammatory disease with a high mortality rate. Although typically seen in individuals with sepsis, ALI is also a major complication in severe acute pancreatitis (SAP). The pathophysiology of SAP-associated ALI is poorly understood, but elevated serum levels of IL-6 is a reliable marker for disease severity. Here, we used a mouse model of acute pancreatitis-associated (AP-associated) ALI to determine the role of IL-6 in ALI lethality. Il6-deficient mice had a lower death rate compared with wild-type mice with AP, while mice injected with IL-6 were more likely to develop lethal ALI. We found that inflammation-associated NF-?B induced myeloid cell secretion of IL-6, and the effects of secreted IL-6 were mediated by complexation with soluble IL-6 receptor, a process known as trans-signaling. IL-6 trans-signaling stimulated phosphorylation of STAT3 and production of the neutrophil attractant CXCL1 in pancreatic acinar cells. Examination of human samples revealed expression of IL-6 in combination with soluble IL-6 receptor was a reliable predictor of ALI in SAP. These results demonstrate that IL-6 trans-signaling is an essential mediator of ALI in SAP across species and suggest that therapeutic inhibition of IL-6 may prevent SAP-associated ALI. PMID:23426178

  18. Synthetic lethality in the tobacco plastid ribosome and its rescue at elevated growth temperatures.

    PubMed

    Ehrnthaler, Miriam; Scharff, Lars B; Fleischmann, Tobias T; Hasse, Claudia; Ruf, Stephanie; Bock, Ralph

    2014-02-01

    Consistent with their origin from cyanobacteria, plastids (chloroplasts) perform protein biosynthesis on bacterial-type 70S ribosomes. The plastid genomes of seed plants contain a conserved set of ribosomal protein genes. Three of these have proven to be nonessential for translation and, thus, for cellular viability: rps15, rpl33, and rpl36. To help define the minimum ribosome, here, we examined whether more than one of these nonessential plastid ribosomal proteins can be removed from the 70S ribosome. To that end, we constructed all possible double knockouts for the S15, L33, and L36 ribosomal proteins by stable transformation of the tobacco (Nicotiana tabacum) plastid genome. We find that, although S15 and L33 function in different ribosomal particles (30S and 50S, respectively), their combined deletion from the plastid genome results in synthetic lethality under autotrophic conditions. Interestingly, the lethality can be overcome by growth under elevated temperatures due to an improved efficiency of plastid ribosome biogenesis. Our results reveal functional interactions between protein and RNA components of the 70S ribosome and uncover the interdependence of the biogenesis of the two ribosomal subunits. In addition, our findings suggest that defining a minimal set of plastid genes may prove more complex than generally believed. PMID:24563204

  19. Malignant gangliocytic paraganglioma of the duodenum with distant metastases and a lethal course

    PubMed Central

    Li, Bin; Li, Yang; Tian, Xiao-Ying; Luo, Bo-Ning; Li, Zhi

    2014-01-01

    Gangliocytic paraganglioma (GP) is rare and has been regarded as benign in general with a good prognosis. We present a patient with duodenal GP showing a malignant and lethal clinical course. A 47-year-old male patient was found to have a duodenal tumor and enlarged regional lymph nodes. The patient initially underwent a pancreaticoduodenectomy to resect the tumor and involved lymph nodes completely. Histological and immunohistochemical analyses showed findings typical of GP. However, the distant metastatic lesions in the liver and pelvic cavity were rapidly observed after surgery. The patient underwent chemotherapy and radiotherapy, as well as a second surgery to partly remove the metastatic mass in the pelvic cavity. The histological examination revealed no significant difference in histological features between the primary duodenal tumor and the metastatic pelvic mass. However, the patient finally died of the tumor due to the recurrence of the residual pelvic lesion and increased liver mass. To our knowledge, this is the first report of lethal GP with multifocal metastases. Our case confirms that GP should be regarded as a malignant potential tumor with behavior code of “1”, rather than a benign tumor of “0”. PMID:25386095

  20. Purification and biophysical characterization of the core protease domain of anthrax lethal factor

    SciTech Connect

    Gkazonis, Petros V.; Dalkas, Georgios A.; Chasapis, Christos T. [Department of Pharmacy, University of Patras, GR-26504 Patras (Greece)] [Department of Pharmacy, University of Patras, GR-26504 Patras (Greece); Vlamis-Gardikas, Alexios [Department of Biochemistry, Foundation for Biomedical Research (BRFAA), Academy of Athens, GR-11527 Athens (Greece)] [Department of Biochemistry, Foundation for Biomedical Research (BRFAA), Academy of Athens, GR-11527 Athens (Greece); Bentrop, Detlef [Institute of Physiology II, University of Freiburg, D-79108 Freiburg (Germany)] [Institute of Physiology II, University of Freiburg, D-79108 Freiburg (Germany); Spyroulias, Georgios A., E-mail: G.A.Spyroulias@upatras.gr [Department of Pharmacy, University of Patras, GR-26504 Patras (Greece)

    2010-06-04

    Anthrax lethal toxin (LeTx) stands for the major virulence factor of the anthrax disease. It comprises a 90 kDa highly specific metalloprotease, the anthrax lethal factor (LF). LF possesses a catalytic Zn{sup 2+} binding site and is highly specific against MAPK kinases, thus representing the most potent native biomolecule to alter and inactivate MKK [MAPK (mitogen-activated protein kinase) kinases] signalling pathways. Given the importance of the interaction between LF and substrate for the development of anti-anthrax agents as well as the potential treatment of nascent tumours, the analysis of the structure and dynamic properties of the LF catalytic site are essential to elucidate its enzymatic properties. Here we report the recombinant expression and purification of a C-terminal part of LF (LF{sub 672-776}) that harbours the enzyme's core protease domain. The biophysical characterization and backbone assignments ({sup 1}H, {sup 13}C, {sup 15}N) of the polypeptide revealed a stable, well folded structure even in the absence of Zn{sup 2+}, suitable for high resolution structural analysis by NMR.

  1. Erythropoiesis Suppression Is Associated with Anthrax Lethal Toxin-Mediated Pathogenic Progression

    PubMed Central

    Chang, Hsin-Hou; Wang, Tsung-Pao; Chen, Po-Kong; Lin, Yo-Yin; Liao, Chih-Hsien; Lin, Ting-Kai; Chiang, Ya-Wen; Lin, Wen-Bin; Chiang, Chih-Yu; Kau, Jyh-Hwa; Huang, Hsin-Hsien; Hsu, Hui-Ling; Liao, Chi-Yuan; Sun, Der-Shan

    2013-01-01

    Anthrax is a disease caused by the bacterium Bacillus anthracis, which results in high mortality in animals and humans. Although some of the mechanisms are already known such as asphyxia, extensive knowledge of molecular pathogenesis of this disease is deficient and remains to be further investigated. Lethal toxin (LT) is a major virulence factor of B. anthracis and a specific inhibitor/protease of mitogen-activated protein kinase kinases (MAPKKs). Anthrax LT causes lethality and induces certain anthrax-like symptoms, such as anemia and hypoxia, in experimental mice. Mitogen-activated protein kinases (MAPKs) are the downstream pathways of MAPKKs, and are important for erythropoiesis. This prompted us to hypothesize that anemia and hypoxia may in part be exacerbated by erythropoietic dysfunction. As revealed by colony-forming cell assays in this study, LT challenges significantly reduced mouse erythroid progenitor cells. In addition, in a proteolytic activity-dependent manner, LT suppressed cell survival and differentiation of cord blood CD34+-derived erythroblasts in vitro. Suppression of cell numbers and the percentage of erythroblasts in the bone marrow were detected in LT-challenged C57BL/6J mice. In contrast, erythropoiesis was provoked through treatments of erythropoietin, significantly ameliorating the anemia and reducing the mortality of LT-treated mice. These data suggested that suppressed erythropoiesis is part of the pathophysiology of LT-mediated intoxication. Because specific treatments to overcome LT-mediated pathogenesis are still lacking, these efforts may help the development of effective treatments against anthrax. PMID:23977125

  2. Putting the Screen in Screening

    PubMed Central

    Harris, Sion Kim; Knight, John R.

    2014-01-01

    Alcohol is strongly linked to the leading causes of adolescent and adult mortality and health problems, making medical settings such as primary care and emergency departments important venues for addressing alcohol use. Extensive research evidence supports the effectiveness of alcohol screening and brief interventions (SBIs) in medical settings, but this valuable strategy remains underused, with medical staff citing lack of time and training as major implementation barriers. Technology-based tools may offer a way to improve efficiency and quality of SBI delivery in such settings. This review describes the latest research examining the feasibility and efficacy of computer- or other technology-based alcohol SBI tools in medical settings, as they relate to the following three patient populations: adults (18 years or older); pregnant women; and adolescents (17 years or younger). The small but growing evidence base generally shows strong feasibility and acceptability of technology-based SBI in medical settings. However, evidence for effectiveness in changing alcohol use is limited in this young field.

  3. A Random Screen Using a Novel Reporter Assay System Reveals a Set of Sequences That Are Preferred as the TATA or TATA-Like Elements in the CYC1 Promoter of Saccharomyces cerevisiae

    PubMed Central

    Watanabe, Kiyoshi; Yabe, Makoto; Kasahara, Koji; Kokubo, Tetsuro

    2015-01-01

    In Saccharomyces cerevisiae, the core promoters of class II genes contain either TATA or TATA-like elements to direct accurate transcriptional initiation. Genome-wide analyses show that the consensus sequence of the TATA element is TATAWAWR (8 bp), whereas TATA-like elements carry one or two mismatches to this consensus. The fact that several functionally distinct TATA sequences have been identified indicates that these elements may function, at least to some extent, in a gene-specific manner. The purpose of the present study was to identify functional TATA sequences enriched in one particular core promoter and compare them with the TATA or TATA-like elements that serve as the pre-initiation complex (PIC) assembly sites on the yeast genome. For this purpose, we conducted a randomized screen of the TATA element in the CYC1 promoter by using a novel reporter assay system and identified several hundreds of unique sequences that were tentatively classified into nine groups. The results indicated that the 7 bp TATA element (i.e., TATAWAD) and several sets of TATA-like sequences are preferred specifically by this promoter. Furthermore, we find that the most frequently isolated TATA-like sequence, i.e., TATTTAAA, is actually utilized as a functional core promoter element for the endogenous genes, e.g., ADE5,7 and ADE6. Collectively, these results indicate that the sequence requirements for the functional TATA or TATA-like elements in one particular core promoter are not as stringent. However, the variation of these sequences differs significantly from that of the PIC assembly sites on the genome, presumably depending on promoter structures and reflecting the gene-specific function of these sequences. PMID:26046838

  4. Identification of a Novel RNA Virus Lethal to Tilapia

    PubMed Central

    Eyngor, Marina; Zamostiano, Rachel; Kembou Tsofack, Japhette Esther; Berkowitz, Asaf; Bercovier, Hillel; Tinman, Simon; Lev, Menachem; Hurvitz, Avshalom; Galeotti, Marco; Eldar, Avi

    2014-01-01

    Tilapines are important for the sustainability of ecological systems and serve as the second most important group of farmed fish worldwide. Significant mortality of wild and cultured tilapia has been observed recently in Israel. The etiological agent of this disease, a novel RNA virus, is described here, and procedures allowing its isolation and detection are revealed. The virus, denominated tilapia lake virus (TiLV), was propagated in primary tilapia brain cells or in an E-11 cell line, and it induced a cytopathic effect at 5 to 10 days postinfection. Electron microscopy revealed enveloped icosahedral particles of 55 to 75 nm. Low-passage TiLV, injected intraperitoneally in tilapia, induced a disease resembling the natural disease, which typically presents with lethargy, ocular alterations, and skin erosions, with >80% mortality. Histological changes included congestion of the internal organs (kidneys and brain) with foci of gliosis and perivascular cuffing of lymphocytes in the brain cortex; ocular inflammation included endophthalmitis and cataractous changes of the lens. The cohabitation of healthy and diseased fish demonstrated that the disease is contagious and that mortalities (80 to 100%) occur within a few days. Fish surviving the initial mortality were immune to further TiLV infections, suggesting the mounting of a protective immune response. Screening cDNA libraries identified a TiLV-specific sequence, allowing the design of a PCR-based diagnostic test. This test enables the specific identification of TiLV in tilapines and should help control the spread of this virus worldwide. PMID:25232154

  5. Earth Revealed

    NSDL National Science Digital Library

    1992-01-01

    What goes on during an earthquake? Who came up with the theory of plate tectonics? What can the fossil record tell us about the evolution of life on Earth? These are all fine questions, and students and educators with a thirst for geological knowledge will find the answers to these (and many more) questions in the "Earth Revealed" television series. Offered as part of the Annenberg Media website, the 26-part series includes such episodes as "Geologic Time", "Mountain Building", and "The Birth of a Theory". As with many of the Annenberg Media offerings, visitors can view entire episodes here, and they can also take a look at a list of additional resources.

  6. 40 CFR 798.5275 - Sex-linked recessive lethal test in drosophila melanogaster.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ...2014-07-01 2014-07-01 false Sex-linked recessive lethal test in drosophila...GUIDELINES Genetic Toxicity § 798.5275 Sex-linked recessive lethal test in drosophila melanogaster. (a) Purpose. The sex-linked recessive lethal (SLRL)...

  7. 40 CFR 798.5275 - Sex-linked recessive lethal test in drosophila melanogaster.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ...2013-07-01 2013-07-01 false Sex-linked recessive lethal test in drosophila...GUIDELINES Genetic Toxicity § 798.5275 Sex-linked recessive lethal test in drosophila melanogaster. (a) Purpose. The sex-linked recessive lethal (SLRL)...

  8. 40 CFR 798.5275 - Sex-linked recessive lethal test in drosophila melanogaster.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ...2012-07-01 2012-07-01 false Sex-linked recessive lethal test in drosophila...GUIDELINES Genetic Toxicity § 798.5275 Sex-linked recessive lethal test in drosophila melanogaster. (a) Purpose. The sex-linked recessive lethal (SLRL)...

  9. To Laugh in the Face of Death: The Games That Lethal People Play.

    ERIC Educational Resources Information Center

    Thorson, James A.; Powell, F. C.

    1990-01-01

    A total of 399 individuals completed a lethal behaviors scale and a measure of death anxiety, which were found to have no significant correlation. Predictors of lethalness included doing dangerous things for the fun of it and having ever driven a motorcycle. The most lethal individuals were young, male, and less educated. (Author/ABL)

  10. A Methodology to Assess Lethality and Collateral Damage for Nonfragmenting Precision-Guided Weapons

    Microsoft Academic Search

    Amanda Humphrey; David Faulkner

    2008-01-01

    A methodology was developed to assess lethality and collateral damage for the Focused Lethality Munition (FLM) program. FLM is a new nonfragmenting, precision-guided weapon with damage effects mechanisms that differ from the principal fragmentation damage effects for traditional weapons. To date, guidelines to determine lethality, based on mannequin test data, have not been articulated for nonfragmenting warheads such as FLM.

  11. 28 CFR 552.25 - Use of chemical agents or non-lethal weapons.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... false Use of chemical agents or non-lethal weapons. 552.25 Section...552.25 Use of chemical agents or non-lethal weapons. The Warden may authorize the use of chemical agents or non-lethal weapons only when the...

  12. Plasma Screen Floating Mount

    DOEpatents

    Eakle, Robert F. (New Ellenton, SC); Pak, Donald J. (Martine, GA)

    2004-10-26

    A mounting system for a flat display screen, particularly a plasma display screen, suspends the screen separately in each of the x-, y- and z-directions. A series of frames located by linear bearings and isolated by springs and dampers allows separate controlled movement in each axis. The system enables the use of relatively larger display screens in vehicles in which plasma screen are subject to damage from vibration.

  13. Investigation of Phenolic Profiles, Cytotoxic Potential and Phytochemical Screening of Different Extracts of Drynaria quercifolia J. Smith (Leaves)

    PubMed Central

    Runa, Jannatul Ferdous; Hossain, Marjan; Hasanuzzaman, Md.; Ali, Md. Ramjan

    2013-01-01

    Purpose: The present study is aimed to evaluate phenolic profiles, cytotoxic activity and phytochemical screening of different extracts of Drynaria quercifolia leaves. Methods: The dried and powder leaves were extracted with methanol at room temperature and the concentrated methanolic extract was fractionated by the modified Kupchan partitioning method to provide pet-ether, carbon tetrachloride, chloroform and aqueous soluble fractions. Phenolic profiles were determined by using Folin-Ciocalteau reagent, which results were expressed in gallic acid equivalent (mg of GAE/g of sample). Phytochemical properties of different extractives of plant materials were tested by the method of Trease and Evans. Brine shrimp lethality bioassay was used to investigate the cytotoxic potential of D. quercifolia. Results: The phytochemical screening revealed the potent source of different phytochemical constituents on different extractives including alkaloid, glycosides, tannin, saponins, proteins and amino acids, flavonoids, triterpenes, phenols, phytosterols and carbohydrate. In the determination of phenolic profiles, different extractives showed a significant content of phenolic compounds ranging from 103.43 -132.23 mg of GAE/g of extractive. Compared to vincristine sulfate different extractives of plant materials demonstrated moderate cytotoxic potential (having LC50 of 12.45 ?g/ml, 13.02 ?g/ml 15.83 ?g/ml, 14.95 ?g/ml and 7.612 ?g/ml, respectively). Conclusion: It is concluded from this study that D. quercifolia is an excellent source of phenolic content and phytoconstitutes as well as possesses moderate cytotoxic activity. PMID:24312880

  14. Georgians Revealed

    NSDL National Science Digital Library

    What was life like during the Georgian era in Britain? During the period between 1714 and 1830, cities and towns were transformed, conspicuous consumption became the pastime of the emerging middle classes, and gardening and shopping for leisure became commonplace. This digital companion to the British Library's "Georgians Revealed" exhibit brings together some of the key books and newspapers from the period, along with details about guided tours through the physical exhibitions, a Georgian London walking tour, and more. For those unable to view the exhibit in person, this companion site provides brief but detailed narratives on interesting facets of the exhibit, including dancing with the Georgians and celebrity culture. The site is rounded out by an excellent timeline of key events from the time of George I (1714-1727) to George IV (1820-1830) accompanied by vivid illustrations and portraiture.

  15. An improved brine shrimp larvae lethality microwell test method.

    PubMed

    Zhang, Yi; Mu, Jun; Han, Jinyuan; Gu, Xiaojie

    2012-01-01

    This article described an improved brine shrimp larvae lethality microwell test method. A simply designed connecting vessel with alternative photoperiod was used to culture and collect high yield of active Artemia parthenogenetica nauplii for brine shrimp larvae lethality microwell test. Using this method, pure A. parthenogenetica nauplii suspension was easily cultured and harvested with high density about 100-150 larvae per milliliter and the natural mortality was reduced to near zero by elimination of unnecessary artificial disturbance. And its sensitivity was validated by determination of LC(50)-24 h of different reference toxicants including five antitumor agents, two pesticides, three organic pollutants, and four heavy metals salts, most of which exhibited LC(50)-24 h between 0.07 and 58.43 mg/L except for bleomycin and mitomycin C with LC(50)-24 h over 300 mg/L. PMID:21859360

  16. Invasive pneumococcal disease: association between serotype, clinical presentation and lethality.

    PubMed

    Rodríguez, M Angeles Gutiérrez; González, Amai Varela; Gavín, María Ascensión Ordobás; Martínez, Fernando Martín; Marín, Natividad García; Blázquez, Belén Ramos; Moreno, Juan Carlos Sanz

    2011-08-01

    To ascertain the factors linked to invasive pneumococcal disease (IPD) caused by the different serotypes in the period 2007-2009, following the conjugate vaccine's inclusion in the childhood vaccination schedule, a total of 2013 IPD cases were reviewed. The mean annual incidence in this period was 10.74 cases per 100,000 inhabitans and the lethality was 8.8%. Overall serotype distribution displayed certain peculiarities, such as the high frequency of serotype 5. Serotype 3, male gender, sepsis and presence of risk factors were significantly associated with lethality. Vaccinated children under 5 years of age had a higher risk of disease due to serotype 19A. Serotype 8 was associated with the presence of underlying risk factors. PMID:21683112

  17. A framework for the assessment of non-lethal weapons.

    PubMed

    Rappert, Brian

    2004-01-01

    In many government, police and military circles, attention is being given to so-called 'non-lethal' weapons as means of reducing many of the negative effects directly or indirectly associated with the use of force. Despite the purported ability of the adoption of such weaponry to lessen grounds for contention and concern, past experience suggests the need for scepticism regarding the purported benefits. Rather than relying on poorly substantiated claims, comprehensive procedures are needed to ensure the appropriateness of force options. This article outlines some of the institutional structures required for 'carefully evaluating' and 'carefully controlling' non-lethal weapons, with a discussion of the perennial tensions associated with ensuring the relative 'acceptability' of the use of force. PMID:15015546

  18. Loss of the Mono-ADP-ribosyltransferase, Tiparp, Increases Sensitivity to Dioxin-induced Steatohepatitis and Lethality.

    PubMed

    Ahmed, Shaimaa; Bott, Debbie; Gomez, Alvin; Tamblyn, Laura; Rasheed, Adil; Cho, Tiffany; MacPherson, Laura; Sugamori, Kim S; Yang, Yang; Grant, Denis M; Cummins, Carolyn L; Matthews, Jason

    2015-07-01

    The aryl hydrocarbon receptor (AHR) mediates the toxic effects of the environmental contaminant dioxin (2,3,7,8-tetrachlorodibenzo-p-dioxin; TCDD). Dioxin causes a range of toxic responses, including hepatic damage, steatohepatitis, and a lethal wasting syndrome; however, the mechanisms are still unknown. Here, we show that the loss of TCDD-inducible poly(ADP-ribose) polymerase (Tiparp), an ADP-ribosyltransferase and AHR repressor, increases sensitivity to dioxin-induced toxicity, steatohepatitis, and lethality. Tiparp(-/-) mice given a single injection of 100 ?g/kg dioxin did not survive beyond day 5; all Tiparp(+/+) mice survived the 30-day treatment. Dioxin-treated Tiparp(-/-) mice exhibited increased liver steatosis and hepatotoxicity. Tiparp ADP-ribosylated AHR but not its dimerization partner, the AHR nuclear translocator, and the repressive effects of TIPARP on AHR were reversed by the macrodomain containing mono-ADP-ribosylase MACROD1 but not MACROD2. These results reveal previously unidentified roles for Tiparp, MacroD1, and ADP-ribosylation in AHR-mediated steatohepatitis and lethality in response to dioxin. PMID:25975270

  19. Lethal and nonlethal murine malarial infections differentially affect apoptosis, proliferation, and CD8 expression on thymic T cells.

    PubMed

    Khanam, S; Sharma, S; Pathak, S

    2015-07-01

    Although thymic atrophy and apoptosis of the double-positive (DP) T cells have been reported in murine malaria, comparative studies investigating the effect of lethal and nonlethal Plasmodium infections on the thymus are lacking. We assessed the effects of P. yoelii lethal (17XL) and nonlethal (17XNL) infections on thymic T cells. Both strains affected the thymus. 17XL infection induced DP T-cell apoptosis and a selective decrease in surface CD8 expression on developing thymocytes. By contrast, more severe but reversible effects were observed during 17XNL infection. DP T cells underwent apoptosis, and proliferation of both DN and DP cells was affected around peak parasitemia. A transient increase in surface CD8 expression on thymic T cells was also observed. Adult thymic organ culture revealed that soluble serum factors, but not IFN-? or TNF-?, contributed to the observed effects. Thus, lethal and nonlethal malarial infections led to multiple disparate effects on thymus. These parasite-induced thymic changes are expected to impact the naïve T-cell repertoire and the subsequent control of the immune response against the parasite. Further investigations are required to elucidate the mechanism responsible for these disparate effects, especially the reversible involution of the thymus in case of nonlethal infection. PMID:25886201

  20. Optimized Production and Purification of Bacillus anthracis Lethal Factor

    Microsoft Academic Search

    Stephen H. Leppla

    2000-01-01

    Bacillus anthracis lethal factor (LF) is a 90-kDa zinc metalloprotease that plays an important role in the virulence of the organism. LF has previously been purified from Escherichia coli and Bacillus anthracis. The yields and purities of these preparations were inadequate for crystal structure determination. In this study, the genes encoding wild-type LF and a mutated, inactive LF (LF-E687C) were

  1. EXCALIBIR - A space experiment in orbital debris lethality

    Microsoft Academic Search

    Robert D. Culp; Michael R. Dickey

    1991-01-01

    The study proposes a space experiment using extended Space Shuttle external tanks to test the impact of orbital debris. The External Tank Calibrated Impact Response test, EXCALIBIR, is a low-cost low-risk, high-payoff approach to investigating the threat to resident space objects posed by untrackable orbital debris, to provide lethality data to the kinetic energy weapons community, and to aid in

  2. Intact alternation performance in high lethality suicide attempters.

    PubMed

    Keilp, John G; Wyatt, Gwinne; Gorlyn, Marianne; Oquendo, Maria A; Burke, Ainsley K; John Mann, J

    2014-09-30

    Suicide attempters often perform poorly on tasks linked to ventral prefrontal cortical (VPFC) function. Object Alternation (OA) - a VPFC probe - has not been used in these studies. In this study, currently depressed medication-free past suicide attempters whose most severe attempt was of high (n=31) vs. low (n=64) lethality, 114 medication-free depressed non-attempters, and 86 non-patients completed a computerized OA task. Participants also completed comparison tasks assessing the discriminant validity of OA (Wisconsin Card Sort), its concurrent validity relative to tasks associated with past attempt status (computerized Stroop task, Buschke Selective Reminding Test), and its construct validity as a VPFC measure (Go-No Go and Iowa Gambling Task). Against expectations, high lethality suicide attempters - the majority of whom used non-violent methods in their attempts with some planning - outperformed other depressed groups on OA, with no group differences observed on Wisconsin Card Sort. Despite intact performance on OA, past attempters exhibited deficits on the Stroop and Buschke. OA performance was associated with performance on Go-No Go and Iowa Gambling, confirming that OA measures a similar construct. VPFC dysfunction may not be a characteristic of all suicide attempters, especially those who make more carefully planned, non-violent - though potentially lethal - attempts. PMID:24878299

  3. Cold shock lethality and injury in Clostridium perfringens.

    PubMed

    Traci, P A; Duncan, C L

    1974-11-01

    Several observations have been made in regard to cold shock lethality of Clostridium perfringens: (i) loss of viability was not consequence of exposure of the cells to air; (ii) stationary-phase cells were much more resistant to cold shock at 4 C than exponential-phase cells; (iii) at 4 C 96% of an initial population of exponential-phase cells was killed upon cold shock and 95% of the remaining population was killed within 90 min of continued exposure at 4 C; (iv) the minimal temperature differential for detectable cold shock lethality was between 17 and 23 C, and the maximum beyond which lethality was not appreciably increased was between 28 and 33 C. Up to 75% of viable cold-shocked cells were injured, as demonstrated by cold shocking late exponential-phase cells at 10 C and using differential plating procedure for recovery. Repair of injury was temperature dependent, and occurred in a complex medium and 0.1% peptone but not water. Nalidixic acid, chloramphenicol, and rifampin did not inhibit repair of injury. PMID:4374121

  4. Anthrax lethal toxin: a weapon of multisystem destruction.

    PubMed

    Agrawal, A; Pulendran, B

    2004-11-01

    Lethal toxin (LT) is a major virulence factor secreted by anthrax bacteria. It is composed of two proteins, PA (protective antigen) and LF (lethal factor). PA transports the LF inside the cell, where LF, a zinc-dependent metalloprotease cleaves the mitogen activated protein kinase kinase (MAPKK) enzymes of the mitogen activated protein kinase (MAPK) signaling pathway, thereby impairing their function. This disruption of the MAPK pathway, which serves essential functions such as proliferation, survival and inflammation in all cell types, results in multisystem dysfunction in the host. The inactivation of the MAPK pathway in both macrophages and dendritic cells leads to inhibition of proinflammatory cytokine secretion, downregulation of costimulatory molecules such as CD80 and CD86, and ineffective T cell priming. The net result is an impaired innate and adaptive immune response. Endothelial cells of the vascular system undergo apoptosis upon LT exposure, also likely due to inactivation of the MAPK pathway. The activity of various hormone receptors such as glucocorticoids, progesterone and estrogen is also blocked, due to inhibition of p38 MAPK phosphorylation, thus affecting the body's response to stress. The present review summarizes the various disarming effects of Bacillus anthracis through the use of a single weapon, the lethal toxin. PMID:15558214

  5. Quantitative high throughput screening identifies inhibitors of anthrax-induced cell death

    PubMed Central

    Zhu, Ping Jun; Hobson, Peyton; Southall, Noel; Qiu, Cunping; Thomas, Craig J.; Lu, Jiamo; Inglese, James; Zheng, Wei; Leppla, Stephen H.; Bugge, Thomas H.; Austin, Christopher P.; Liu, Shihui

    2009-01-01

    Here, we report the results of a quantitative high-throughput screen (qHTS) measuring the endocytosis and translocation of a ?-lactamase-fused-lethal factor and the identification of small molecules capable of obstructing the process of anthrax toxin internalization. Several small molecules protect RAW264.7 macrophages and CHO cells from anthrax lethal toxin and protected cells from an LF-Pseudomonas exotoxin fusion protein and diphtheria toxin. Further efforts demonstrated that these compounds impaired the PA heptamer pre-pore to pore conversion in cells expressing the CMG2 receptor, but not the related TEM8 receptor, indicating that these compounds likely interfere with toxin internalization. PMID:19540764

  6. [Lung cancer screening].

    PubMed

    Sánchez González, M

    2014-01-01

    Lung cancer is a very important disease, curable in early stages. There have been trials trying to show the utility of chest x-ray or computed tomography in Lung Cancer Screening for decades. In 2011, National Lung Screening Trial results were published, showing a 20% reduction in lung cancer mortality in patients with low dose computed tomography screened for three years. These results are very promising and several scientific societies have included lung cancer screening in their guidelines. Nevertheless we have to be aware of lung cancer screening risks, such as: overdiagnosis, radiation and false positive results. Moreover, there are many issues to be solved, including choosing the appropriate group to be screened, the duration of the screening program, intervals between screening and its cost-effectiveness. Ongoing trials will probably answer some of these questions. This article reviews the current evidence on lung cancer screening. PMID:23830728

  7. Cost-effectiveness of breast cancer screening policies using simulation.

    PubMed

    Gocgun, Y; Banjevic, D; Taghipour, S; Montgomery, N; Harvey, B J; Jardine, A K S; Miller, A B

    2015-08-01

    In this paper, we study breast cancer screening policies using computer simulation. We developed a multi-state Markov model for breast cancer progression, considering both the screening and treatment stages of breast cancer. The parameters of our model were estimated through data from the Canadian National Breast Cancer Screening Study as well as data in the relevant literature. Using computer simulation, we evaluated various screening policies to study the impact of mammography screening for age-based subpopulations in Canada. We also performed sensitivity analysis to examine the impact of certain parameters on number of deaths and total costs. The analysis comparing screening policies reveals that a policy in which women belonging to the 40-49 age group are not screened, whereas those belonging to the 50-59 and 60-69 age groups are screened once every 5 years, outperforms others with respect to cost per life saved. Our analysis also indicates that increasing the screening frequencies for the 50-59 and 60-69 age groups decrease mortality, and that the average number of deaths generally decreases with an increase in screening frequency. We found that screening annually for all age groups is associated with the highest costs per life saved. Our analysis thus reveals that cost per life saved increases with an increase in screening frequency. PMID:25866350

  8. Lack of dominant lethality in mice following 1-bromopropane treatment.

    PubMed

    Yu, Wook-Joon; Kim, Jong-Choon; Chung, Moon-Koo

    2008-03-29

    1-Bromopropane (1-BP) is widely used in spray adhesives, precision cleaner, and degreaser. This study was conducted to investigate the potential of 1-BP to induce dominant lethality in mice. 1-BP was orally administered to males at doses of 300 and 600 mg/kg for 10 days before mating. Cyclophosphamide was used as a positive control (PC), which was administered intraperitoneally to males at 40 mg/kg for 5 days. The vehicle control (VC) group received corn oil only. Thereafter, males were mated with untreated females during six sequential mating periods of a week each. Males were sacrificed at the end of mating and so were the pregnant females on days 15-17 of gestation. Clinical signs, gross findings, mating index, gestation index, the numbers of corpora lutea, implantations, live fetuses, resorptions and dead fetuses, pre- and post-implantation losses, and dominant lethal mutation rate were examined. There were no treatment-related changes in clinical signs, gross findings, mating index, gestation index, number of corpora lutea and implantations, pre-implantation loss, live fetuses, resorptions, dead fetuses, post-implantation loss at any 1-BP doses tested. In the PC group, there were no treatment-related changes in mating index, gestation index, number of corpora lutea, and dead fetuses. However, a decrease in the number of implantations and an increase in pre-implantation loss were observed during the first 2 weeks as compared to those of the VC group. No treatment-related changes were observed in the third to sixth weeks. Increases in resorptions, fetal deaths and post-implantation loss, and a decrease in the number of live fetuses were observed in the first 3 weeks of the PC group compared to those of the VC group. However, no treatment-related changes were observed during the forth to sixth weeks. An increase in dominant lethal mutation rate was observed in 1-3 weeks of mating of the PC group, but there was no significant difference in 1-6 weeks of mating of the 1-BP treatment groups. In conclusion, 1-BP did not induce dominant lethality in mice. These results are in agreement with the report of Saito-Suzuki et al., demonstrating that no dominant lethality of 1-BP was observed in Sprague-Dawley rats. PMID:18291709

  9. Two Cases of Lethal Complications Following Ultrasound-Guided Percutaneous Fine-Needle Biopsy of the Liver

    SciTech Connect

    Drinkovic, Ivan; Brkljacic, Boris [Department of Radiology, Ultrasonic Center, Medical School of the University of Zagreb, University Hospital 'Merkur', Zajceva 19, 10000 Zagreb (Croatia)

    1996-09-15

    Two cases with lethal complications are reported among 1750 ultrasound (US)-guided percutaneous fine-needle liver biopsies performed in our department. The first patient had angiosarcoma of the liver which was not suspected after computed tomography (CT) and US studies had been performed. The other patient had hepatocellular carcinoma in advanced hepatic cirrhosis. Death was due to bleeding in both cases. Pre-procedure laboratory tests did not reveal the existence of major bleeding disorders in either case. Normal liver tissue was interposed in the needle track between the liver capsule and the lesions which were targeted.

  10. Synthetic lethality with fibrillarin identifies NOP77p, a nucleolar protein required for pre-rRNA processing and modification.

    PubMed Central

    Bergès, T; Petfalski, E; Tollervey, D; Hurt, E C

    1994-01-01

    The nucleolar protein fibrillarin (encoded by the NOP1 gene in yeast), is required for many post-transcriptional steps in yeast ribosome synthesis. A screen for mutations showing synthetic lethality with a temperature sensitive nop1-5 allele led to the identification of the NOP77 gene. NOP77 is essential for viability and encodes a nucleolar protein with a predicted molecular weight of 77 kDa. Depletion of NOP77p impairs both the processing and methylation of the pre-rRNA. The processing defect is greatest for the pathway leading to 25S rRNA synthesis, and is distinctly different from that observed for mutations in other nucleolar components. NOP77p contains three canonical RNA recognition motifs (RRMs), suggesting that it is an RNA binding protein. The NOP77 allele which complements the synthetic lethal nop1 strains has an alanine at position 308, predicted to lie in helix alpha 1 of RRM3, whereas the non-complementing nop77-1 allele contains a proline at the corresponding position. We propose that NOP77p mediates specific interactions between NOP1p and the pre-rRNA. Images PMID:8039506

  11. Synthetic lethal targeting of PTEN-deficient cancer cells using selective disruption of polynucleotide kinase/phosphatase

    PubMed Central

    Mereniuk, Todd R.; El Gendy, Mohamed A.M.; Mendes-Pereira, Ana M.; Lord, Christopher J.; Ghosh, Sunita; Foley, Edan; Ashworth, Alan; Weinfeld, Michael

    2013-01-01

    A recent screen of 6961 siRNAs to discover possible synthetic lethal partners of the DNA repair protein polynucleotide kinase/phosphatase (PNKP) led to the identification of the potent tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (PTEN). Here we have confirmed the PNKP/PTEN synthetic lethal partnership in a variety of different cell lines including the PC3 prostate cancer cell line, which is naturally deficient in PTEN. We provide evidence that co-depletion of PTEN and PNKP induces apoptosis. In HCT116 colon cancer cells the loss of PTEN is accompanied by an increased background level of DNA double strand breaks, which accumulate in the presence of an inhibitor of PNKP DNA 3?-phosphatase activity. Complementation of PC3 cells with several well-characterized mutated PTEN cDNAs indicated that the critical function of PTEN required to prevent toxicity induced by an inhibitor of PNKP is most likely associated with its cytoplasmic lipid phosphatase activity. Finally, we show that modest inhibition of PNKP in a PTEN knockout background enhances cellular radiosensitivity, suggesting that such a “synthetic sickness” approach involving the combination of PNKP inhibition with radiotherapy may be applicable to PTEN-deficient tumors. PMID:23883586

  12. Synthetic lethal targeting of PTEN-deficient cancer cells using selective disruption of polynucleotide kinase/phosphatase.

    PubMed

    Mereniuk, Todd R; El Gendy, Mohamed A M; Mendes-Pereira, Ana M; Lord, Christopher J; Ghosh, Sunita; Foley, Edan; Ashworth, Alan; Weinfeld, Michael

    2013-10-01

    A recent screen of 6,961 siRNAs to discover possible synthetic lethal partners of the DNA repair protein polynucleotide kinase/phosphatase (PNKP) led to the identification of the potent tumor suppressor phosphatase and tensin homolog deleted on chromosome 10 (PTEN). Here, we have confirmed the PNKP/PTEN synthetic lethal partnership in a variety of different cell lines including the PC3 prostate cancer cell line, which is naturally deficient in PTEN. We provide evidence that codepletion of PTEN and PNKP induces apoptosis. In HCT116 colon cancer cells, the loss of PTEN is accompanied by an increased background level of DNA double-strand breaks, which accumulate in the presence of an inhibitor of PNKP DNA 3'-phosphatase activity. Complementation of PC3 cells with several well-characterized mutated PTEN cDNAs indicated that the critical function of PTEN required to prevent toxicity induced by an inhibitor of PNKP is most likely associated with its cytoplasmic lipid phosphatase activity. Finally, we show that modest inhibition of PNKP in a PTEN knockout background enhances cellular radiosensitivity, suggesting that such a "synthetic sickness" approach involving the combination of PNKP inhibition with radiotherapy may be applicable to PTEN-deficient tumors. PMID:23883586

  13. Stretching Screens and Imaginations.

    ERIC Educational Resources Information Center

    Douthwaite, Shelaugh

    1983-01-01

    Secondary students utilize a simplified technique to make silk screen prints, which can be printed onto T-shirts. The only materials needed from art suppliers are a few squeegees and a few yards of polyester screen mesh. (RM)

  14. Breast Cancer Screening

    MedlinePLUS

    ... is available from the NCI Web site . Three tests are used by health care providers to screen for breast cancer: Mammogram Mammography is the most common screening test for breast cancer . A mammogram is an x- ...

  15. Screening for Birth Defects

    MedlinePLUS

    ... extra chromosome. A common trisomy is trisomy 21 (Down syndrome) . Other trisomies include trisomy 13 (Patau syndrome) and ... Test Test Type What Does It Screen for? Down Syndrome Detection Rate Combined firsttrimester screening Blood test for ...

  16. Cervical Cancer Screening

    MedlinePLUS

    ... Cancer found early may be easier to treat. Cervical cancer screening is usually part of a woman's ... may do more tests, such as a biopsy. Cervical cancer screening has risks. The results can sometimes ...

  17. International Cancer Screening Network

    Cancer.gov

    Skip to Main Content Search International Cancer Screening Network Sponsored by the National Cancer Institute Working Together to Evaluate Cancer Screening and Improve Outcomes Internationally About the ICSN Overview Participating Countries Contact

  18. Lethal and sublethal effects of marine sediment extracts on fish cells and chromosomes

    NASA Astrophysics Data System (ADS)

    Landolt, Marsha L.; Kocan, Richard M.

    1984-03-01

    The cost of conducting conventional chronic bioassays with every potentially toxic compound found in marine ecosystems is prohibitive; therefore short-term toxicity tests which can be used for rapid screening were developed. The tests employ cultured fish cells to measure lethal, sublethal or genotoxic effects of pure compounds and complex mixtures. The sensitivity of these tests has been proven under laboratory conditions; the following study used two of these tests, the anaphase aberration test and a cytotoxicity assay, under field conditions. Sediment was collected from 97 stations within Puget Sound, Washington. Serial washings of the sediment in methanol and dichloromethane yielded an organic extract which was dried, dissolved in DMSO and incubated as a series of dilutions with rainbow trout gonad (RTG-2) cells. The toxic effects of the extract were measured by examining the rate of cell proliferation and the percentage of damaged anaphase figures. Anaphase figures were considered to be abnormal if they exhibited non-disjunctions, chromosome fragments, or chromosome bridges. A second cell line (bluegill fry, BF-2) was also tested for cell proliferation and was included because, unlike the RTG-2 cell line, it contains little or no mixed function oxygenase activity. Of 97 stations tested, 35 showed no genotoxic activity, 42 showed high genotoxic activity (P?.01) and the remainder were intermediate. Among the toxic sites were several deep water stations adjacent to municipal sewage outfalls and four urban waterways contaminated by industrial and municipal effluents. Extracts from areas that showed genotoxic effects also inhibited cell proliferation and were cytotoxic to RTG-2 cells. Few effects were noted in the MFO deficient BF-2 cells. Short term in vitro tests provide aquatic toxicologists with a versatile and cost effective tool for screening complex environments. Through these tests one can identify compounds or geographic regions that exhibit high cytotoxic or genotoxic potential.

  19. Colorectal cancer screening.

    PubMed

    Burt, Randall W; Cannon, Jamie A; David, Donald S; Early, Dayna S; Ford, James M; Giardiello, Francis M; Halverson, Amy L; Hamilton, Stanley R; Hampel, Heather; Ismail, Mohammad K; Jasperson, Kory; Klapman, Jason B; Lazenby, Audrey J; Lynch, Patrick M; Mayer, Robert J; Ness, Reid M; Provenzale, Dawn; Rao, M Sambasiva; Shike, Moshe; Steinbach, Gideon; Terdiman, Jonathan P; Weinberg, David; Dwyer, Mary; Freedman-Cass, Deborah

    2013-12-01

    Mortality from colorectal cancer can be reduced by early diagnosis and by cancer prevention through polypectomy. These NCCN Guidelines for Colorectal Cancer Screening describe various colorectal screening modalities and recommended screening schedules for patients at average or increased risk of developing colorectal cancer. In addition, the guidelines provide recommendations for the management of patients with high-risk colorectal cancer syndromes, including Lynch syndrome. Screening approaches for Lynch syndrome are also described. PMID:24335688

  20. Triple Screening in Pregnancy

    MedlinePLUS

    MENU Return to Web version Triple Screening in Pregnancy What is a triple screen? A triple screen is a blood test that measures three things called alpha-fetoprotein, human chorionic gonadotropin and unconjugated estriol. The results of the blood test ...

  1. VLHC Beam Screen Cooling

    Microsoft Academic Search

    C. Darve; P. Bauer; P. Limon; T. Peterson

    In the framework of the Very Large Hadron Collider (VLHC) R&D studies, vacuum and beam screen issues are being investigated at Fermilab. This report focuses on the cooling system for the VLHC beam screen and its integration into the VLHC cryo-system. Beam screen cooling is a key issue in the VLHC, due to the large synchrotron radiation (SR) power generated

  2. The Mutational Landscape of Lethal Castrate Resistant Prostate Cancer

    PubMed Central

    Grasso, Catherine S.; Wu, Yi-Mi; Robinson, Dan R.; Cao, Xuhong; Dhanasekaran, Saravana M.; Khan, Amjad P.; Quist, Michael J.; Jing, Xiaojun; Lonigro, Robert J.; Brenner, J. Chad; Asangani, Irfan A.; Ateeq, Bushra; Chun, Sang Y.; Siddiqui, Javed; Sam, Lee; Anstett, Matt; Mehra, Rohit; Prensner, John R.; Palanisamy, Nallasivam; Ryslik, Gregory A.; Vandin, Fabio; Raphael, Benjamin J.; Kunju, Lakshmi P.; Rhodes, Daniel R.; Pienta, Kenneth J.; Chinnaiyan, Arul M.; Tomlins, Scott A.

    2012-01-01

    Characterization of the prostate cancer transcriptome and genome has identified chromosomal rearrangements and copy number gains/losses, including ETS gene fusions, PTEN loss and androgen receptor (AR) amplification, that drive prostate cancer development and progression to lethal, metastatic castrate resistant prostate cancer (CRPC)1. As less is known about the role of mutations2–4, here we sequenced the exomes of 50 lethal, heavily-pretreated metastatic CRPCs obtained at rapid autopsy (including three different foci from the same patient) and 11 treatment naïve, high-grade localized prostate cancers. We identified low overall mutation rates even in heavily treated CRPC (2.00/Mb) and confirmed the monoclonal origin of lethal CRPC. Integrating exome copy number analysis identified disruptions of CHD1, which define a subtype of ETS fusionnegative prostate cancer. Similarly, we demonstrate that ETS2, which is deleted in ~1/3 of CRPCs (commonly through TMPRSS2:ERG fusions), is also deregulated through mutation. Further, we identified recurrent mutations in multiple chromatin/histone modifying genes, including MLL2 (mutated in 8.6% of prostate cancers), and demonstrate interaction of the MLL complex with AR, which is required for AR-mediated signaling. We also identified novel recurrent mutations in the AR collaborating factor FOXA1, which is mutated in 5 of 147 (3.4%) prostate cancers (both untreated localized prostate cancer and CRPC), and showed that mutated FOXA1 represses androgen signaling and increases tumour growth. Proteins that physically interact with AR, such as the ERG gene fusion product, FOXA1, MLL2, UTX, and ASXL1 were found to be mutated in CRPC. In summary, we describe the mutational landscape of a heavily treated metastatic cancer, identify novel mechanisms of AR signaling deregulated in prostate cancer, and prioritize candidates for future study. PMID:22722839

  3. Left ventricular function during lethal and sublethal endotoxemia in swine

    SciTech Connect

    Goldfarb, R.D.; Nightingale, L.M.; Kish, P.; Weber, P.B.; Loegering, D.J.

    1986-08-01

    Previous studies suggested that after a median lethal dose (LD50) of endotoxin, cardiac contractility was depressed in nonsurviving dogs. The canine cardiovascular system is unlike humans in that dogs have a hepatic vein sphincter that is susceptible to adrenergic stimulation capable of raising hepatic and splanchnic venous pressures. The authors retested the hypothesis that lethality after endotoxin administration is associated with cardiac contractile depression in pigs, because of the hepatic circulation in this species is similar to that of humans. They compared cardiac mechanical function of pigs administered a high dose (250 g/kg) or a low dose (100 g/kg) endotoxin by use of the slope of the end-systolic pressure-diameter relationship (ESPDR) as well as other measurements of cardiac performance. In all the pigs administered a high dose, ESPDR demonstrated a marked, time-dependent depression whereas we observed no significant ESPDR changes after low endotoxin doses. The other cardiodynamic variables were uninterpretable, due to the significant changes in heart rate, end-diastolic diameter (preload), and aortic diastolic pressure (afterload). Plasma myocardia depressant factor activity accumulated in all endotoxin-administered animals, tending to be greater in the high-dose group. In this group, both subendocardial blood flow and global function were depressed, whereas pigs administered the low dose endotoxin demonstrated slight, but nonsignificant, increases in flow and function. These observations indicate that myocardial contractile depression is associated with a lethal outcome to high doses of endotoxin. Myocardial perfusion was measured using radiolabeled microspheres infused into the left atria.

  4. What Are Reasons for the Large Gender Differences in the Lethality of Suicidal Acts? An Epidemiological Analysis in Four European Countries

    PubMed Central

    Heinrichs, Katherina; Székely, András; Tóth, Mónika Ditta; Coyne, James; Quintão, Sónia; Arensman, Ella; Coffey, Claire; Maxwell, Margaret; Värnik, Airi; van Audenhove, Chantal; McDaid, David; Sarchiapone, Marco; Schmidtke, Armin; Genz, Axel; Gusmão, Ricardo; Hegerl, Ulrich

    2015-01-01

    Background In Europe, men have lower rates of attempted suicide compared to women and at the same time a higher rate of completed suicides, indicating major gender differences in lethality of suicidal behaviour. The aim of this study was to analyse the extent to which these gender differences in lethality can be explained by factors such as choice of more lethal methods or lethality differences within the same suicide method or age. In addition, we explored gender differences in the intentionality of suicide attempts. Methods and Findings Methods. Design: Epidemiological study using a combination of self-report and official data. Setting: Mental health care services in four European countries: Germany, Hungary, Ireland, and Portugal. Data basis: Completed suicides derived from official statistics for each country (767 acts, 74.4% male) and assessed suicide attempts excluding habitual intentional self-harm (8,175 acts, 43.2% male). Main Outcome Measures and Data Analysis. We collected data on suicidal acts in eight regions of four European countries participating in the EU-funded “OSPI-Europe”-project (www.ospi-europe.com). We calculated method-specific lethality using the number of completed suicides per method * 100 / (number of completed suicides per method + number of attempted suicides per method). We tested gender differences in the distribution of suicidal acts for significance by using the ?2-test for two-by-two tables. We assessed the effect sizes with phi coefficients (?). We identified predictors of lethality with a binary logistic regression analysis. Poisson regression analysis examined the contribution of choice of methods and method-specific lethality to gender differences in the lethality of suicidal acts. Findings Main Results Suicidal acts (fatal and non-fatal) were 3.4 times more lethal in men than in women (lethality 13.91% (regarding 4106 suicidal acts) versus 4.05% (regarding 4836 suicidal acts)), the difference being significant for the methods hanging, jumping, moving objects, sharp objects and poisoning by substances other than drugs. Median age at time of suicidal behaviour (35–44 years) did not differ between males and females. The overall gender difference in lethality of suicidal behaviour was explained by males choosing more lethal suicide methods (odds ratio (OR) = 2.03; 95% CI = 1.65 to 2.50; p < 0.000001) and additionally, but to a lesser degree, by a higher lethality of suicidal acts for males even within the same method (OR = 1.64; 95% CI = 1.32 to 2.02; p = 0.000005). Results of a regression analysis revealed neither age nor country differences were significant predictors for gender differences in the lethality of suicidal acts. The proportion of serious suicide attempts among all non-fatal suicidal acts with known intentionality (NFSAi) was significantly higher in men (57.1%; 1,207 of 2,115 NFSAi) than in women (48.6%; 1,508 of 3,100 NFSAi) (?2 = 35.74; p < 0.000001). Main limitations of the study Due to restrictive data security regulations to ensure anonymity in Ireland, specific ages could not be provided because of the relatively low absolute numbers of suicide in the Irish intervention and control region. Therefore, analyses of the interaction between gender and age could only be conducted for three of the four countries. Attempted suicides were assessed for patients presenting to emergency departments or treated in hospitals. An unknown rate of attempted suicides remained undetected. This may have caused an overestimation of the lethality of certain methods. Moreover, the detection of attempted suicides and the registration of completed suicides might have differed across the four countries. Some suicides might be hidden and misclassified as undetermined deaths. Conclusions Men more often used highly lethal methods in suicidal behaviour, but there was also a higher method-specific lethality which together explained the large gender differences in the lethality of suicidal acts. Gender differences in the lethality of suicidal acts were

  5. Studies on the Chick-lethal Toxin of Escherichia coli

    PubMed Central

    Truscott, R. B.

    1973-01-01

    A toxin which is lethal for two week old chicks has been recovered from strains of Escherichia coli O78:K80 of bovine and avian origin and from avian isolates of serogroups O2, O45 and O109. The toxin is heat-labile, antigenic, high in protein, inactivated by pronase, trypsin, amylase, and pancreatic lipase. The toxin may be precipitated by ammonium sulfate or TCA treatment from the supernatant obtained by repeated centrifugation of sonicated cells. Considerable purification has been obtained by column chromatography using Sepharose 6B. PMID:4270809

  6. Ocular manifestations of the potentially lethal rheumatologic and vasculitic disorders.

    PubMed

    Foster, C Stephen

    2013-06-01

    Vision threatening ocular inflammation may occur in patients with any of the acquired connective tissue disorders and vasculitic diseases. Additionally, the ocular inflammation may be the presenting manifestation of the disease, which leads the patient to seek medical care. Other manifestations of the potentially lethal disease may be subtle or absent, presenting the thoughtful ophthalmologist with the opportunity to make life saving discoveries. Necrotizing scleritis, peripheral ulcerative keratitis, and retinal vasculitis are the ocular findings which should prompt the ophthalmologist to initiate very aggressive measures aimed at discovering any evidence of extra-ocular abnormalities, laboratory or otherwise. Appropriate therapy will be sight saving and may be life saving. PMID:23688612

  7. Use of a High Throughput Screening Approach Coupled With In Vivo Zebrafish Embryo Screening to Develop Hazard Ranking for Engineered Nanomaterials

    PubMed Central

    George, Saji; Xia, Tian; Rallo, Robert; Zhao, Yan; Ji, Zhaoxia; Lin, Sijie; Wang, Xiang; Zhang, Haiyuan; France, Bryan; Schoenfeld, David; Damoiseaux, Robert; Liu, Rong; Lin, Shuo; Bradley, Kenneth A; Cohen, Yoram; Nel, André E

    2014-01-01

    Because of concerns about the safety of a growing number of engineered nanomaterials (ENM), it is necessary to develop high throughput screening and in silico data transformation tools that can speed up in vitro hazard ranking. Here, we report the use of a multi-parametric, automated screening assay that incorporates sub-lethal and lethal cellular injury responses to perform high throughput analysis of a batch of commercial metal/metal oxide nanoparticles (NP) with the inclusion of a quantum dot (QD1). The responses chosen for tracking cellular injury through automated epifluorescence microscopy included ROS production, intracellular calcium flux, mitochondrial depolarization, and plasma membrane permeability. The z-score transformed high volume data set was used to construct heat maps for in vitro hazard ranking as well as showing the similarity patterns of NPs and response parameters through the use of self-organizing maps (SOM). Among the materials analyzed, QD1 and nano-ZnO showed the most prominent lethality, while Pt, Ag, SiO2, Al2O3, and Au triggered sub-lethal effects but without cytotoxicity. In order to compare the in vitro with the in vivo response outcomes in zebrafish embryos, NPs were used to assess their impact on mortality rate, hatching rate, cardiac rate, and morphological defects. While QDs, ZnO, and Ag induced morphological abnormalities or interfered in embryo hatching, Pt and Ag exerted inhibitory effects on cardiac rate. Ag toxicity in zebrafish differed from the in vitro results, which is congruent with this material's designation as extremely dangerous in the environment. Interestingly, while toxicity in the initially selected QD formulation was due to a solvent (toluene), supplementary testing of additional QDs selections yielded in vitro hazard profiling that reflect the release of chalcogenides. In conclusion, the use of a high throughput screening, in silico data handling and zebrafish testing may constitute a paradigm for rapid and integrated ENM toxicological screening. PMID:21323332

  8. Amifostine alleviates radiation-induced lethal small bowel damage via promotion of 14-3-3?-mediated nuclear p53 accumulation

    PubMed Central

    Chen, Yu-Min; Chen, Yi-Fan; Wang, Chung-Chi; Lin, I-Hui; Huang, Yu-Jie; Yang, Kuender D.

    2014-01-01

    Amifostine (AM) is a radioprotector that scavenges free radicals and is used in patients undergoing radiotherapy. p53 has long been implicated in cell cycle arrest for cellular repair after radiation exposure. We therefore investigated the protective p53-dependent mechanism of AM on small bowel damage after lethal whole-abdominal irradiation (WAI). AM increased both the survival rate of rats and crypt survival following lethal 18 Gy WAI. The p53 inhibitor PFT-? compromised AM-mediated effects when administered prior to AM administration. AM significantly increased clonogenic survival in IEC-6 cells expressing wild type p53 but not in p53 knockdown cells. AM significantly increased p53 nuclear accumulation and p53 tetramer expression before irradiation through the inhibition of p53 degradation. AM inhibited p53 interactions with MDM2 but enhanced p53 interactions with 14-3-3?. Knockdown of 14-3-3? also compromised the effect of AM on clonogenic survival and p53 nuclear accumulation in IEC-6 cells. For the first time, our data reveal that AM alleviates lethal small bowel damage through the induction of 14-3-3? and subsequent accumulation of p53. Enhancement of the p53/14-3-3? interaction results in p53 tetramerization in the nucleus that rescues lethal small bowel damage. PMID:25230151

  9. Using biomarkers to stage disease progression in a lethal mousepox model treated with CMX001

    PubMed Central

    Parker, Scott; Schriewer, Jill; Oberle, Christina; Robertson, Alice; Lanier, Randall; Painter, George; Buller, R Mark

    2013-01-01

    Background The emergence of human monkeypox and the potential use of recombinant variola and monkeypox viruses as biological terrorist agents have necessitated the development of therapeutic and prophylactic therapies. The primary, or index, cases of smallpox and/or human monkeypox will likely be identified by a characteristic rash. Effective biomarkers will be required to monitor disease progression, guide the choice of therapeutic intervention strategies and evaluate their efficacies. To address this we have evaluated several biomarkers of disease in a lethal mousepox model. Methods The efficacy of a single dose of a hexadecyloxypropyl ester of cidofovir (CMX001) at 20, 25 and 30 mg/ kg doses administered on days 4, 5, 6 and 7 post-infection was evaluated in A/Ncr mice intranasally infected with low doses of ectromelia virus (<20 plaque-forming units). Mice were monitored for weight loss, blood interferon-? levels, alanine aminotransferase (ALT), aspartate aminotransferase, viral DNA copies and neutrophilia levels to stage disease progression. Results We have used these biomarkers to establish the optimal dosing regimen for treatment and reveal that a single dose of 25 mg/kg of CMX001 can be efficacious at treating lethal mousepox when administered on days 4 or 5 post-infection. This dose significantly reduces ALT, interferon-? and DNA copies found in the blood of infected animals. Conclusions A single dose regimen of CMX001 is efficacious at treating mousepox. Disease progression and antiviral efficacy can be monitored using several biomarkers that could readily be used in the case of a human monkeypox or smallpox outbreak. PMID:19043920

  10. Bacillus anthracis’ lethal toxin induces broad transcriptional responses in human peripheral monocytes

    PubMed Central

    2012-01-01

    Background Anthrax lethal toxin (LT), produced by the Gram-positive bacterium Bacillus anthracis, is a highly effective zinc dependent metalloprotease that cleaves the N-terminus of mitogen-activated protein kinase kinases (MAPKK or MEKs) and is known to play a role in impairing the host immune system during an inhalation anthrax infection. Here, we present the transcriptional responses of LT treated human monocytes in order to further elucidate the mechanisms of LT inhibition on the host immune system. Results Western Blot analysis demonstrated cleavage of endogenous MEK1 and MEK3 when human monocytes were treated with 500?ng/mL LT for four hours, proving their susceptibility to anthrax lethal toxin. Furthermore, staining with annexin V and propidium iodide revealed that LT treatment did not induce human peripheral monocyte apoptosis or necrosis. Using Affymetrix Human Genome U133 Plus 2.0 Arrays, we identified over 820 probe sets differentially regulated after LT treatment at the p <0.001 significance level, interrupting the normal transduction of over 60 known pathways. As expected, the MAPKK signaling pathway was most drastically affected by LT, but numerous genes outside the well-recognized pathways were also influenced by LT including the IL-18 signaling pathway, Toll-like receptor pathway and the IFN alpha signaling pathway. Multiple genes involved in actin regulation, signal transduction, transcriptional regulation and cytokine signaling were identified after treatment with anthrax LT. Conclusion We conclude LT directly targets human peripheral monocytes and causes multiple aberrant gene responses that would be expected to be associated with defects in human monocyte’s normal signaling transduction pathways and function. This study provides further insights into the mechanisms associated with the host immune system collapse during an anthrax infection, and suggests that anthrax LT may have additional downstream targets outside the well-known MAPK pathway. PMID:22747600

  11. Procalcitonin as a predictive biomarker for total body irradiation induced bacterial load and lethality in mice

    PubMed Central

    Biju, Prabath G.; Garg, Sarita; Wang, Wenze; Choudhry, Mashkoor A.; Kovacs, Elizabeth J.; Fink, Louis M.; Hauer-Jensen, Martin

    2012-01-01

    Sepsis is the leading cause of mortality in intensive care units. Early detection and intervention are critical to prevent death. The acute radiation syndrome is characterized by damage of the gastrointestinal and hematopoietic systems. Translocation of intestinal microflora combined with immune system compromise may lead to septicemia and death. This work examined the utility of procalcitonin, a clinical sepsis biomarker, in a mouse model of radiation toxicity. C57/BL6 mice were exposed to total body irradiation (TBI). Intestinal mucosal permeability was measured in vivo, and liver bacterial load and plasma levels of procalcitonin (PCT), lipopolysaccharide (LPS), and lipopolysaccharide binding protein (LBP) were measured at baseline and 3.5, 7, and 10 days after TBI. The value of early PCT in predicting subsequent lethality was determined by receiver operator characteristics (ROC) analysis. Four days after TBI a dose-dependent increase in permeability of the intestinal mucosa was observed, while bacterial translocation was present from day 7 onward. There was a high positive correlation between bacterial translocation and all sepsis biomarkers, with PCT exhibiting the strongest correlation. Moreover, plasma PCT levels were elevated already from day 3.5 onwards, whereas, LPS was elevated from day 7 and LBP only 10 days after TBI. ROC analysis revealed that PCT levels measured 3.5 days after TBI predicted lethality at 10 days. These data demonstrate the value of PCT as an early biomarker in radiation-induced bacteremia for mouse studies and suggest that clinical results from other septic conditions may apply to post-radiation septicemia in humans. PMID:22576002

  12. Liquid environmental stress screening

    NASA Astrophysics Data System (ADS)

    Walker, J. L.

    1992-05-01

    This report describes a method of performing Environmental Stress Screening in an inert fluid. Because the liquid has a high dielectric strength, the items being screened can be energized and operational while submerged. Transmitting vibrations through the liquid to the item being screened permits screening without the use of fixtures, and the liquid's high heat capacity allows very rapid temperature changes, either by pumping hot or cold liquid from remote reservoirs or by moving the device from a hot tank to a cold tank. A prototype screening system has been built at NRaD, and faults induced in a small (47 pieces) lot of printed wiring boards have been detected.

  13. Serum Amyloid A Protects Murine Macrophages from Lethal Toxin-Mediated Death1

    PubMed Central

    Rose, Kira; Long, Paul; Shankar, Malini; Ballard, Jimmy D.; Webb, Carol F.

    2011-01-01

    Lethal toxin, a key virulence factor produced by Bacillus anthracis, induces cell death, in part by disrupting numerous signaling pathways, in mouse macrophages. However, exposure to sublethal doses of lethal toxin allows some cells to survive. Because these pro-survival signaling events occur within a few hours after exposure to sublethal doses, we hypothesized that acute phase proteins might influence macrophage survival. Our data show that serum amyloid A (SAA) is produced in response to lethal toxin treatment. Moreover, pre-treatment of macrophages with exogenous SAA protected macrophages from lethal toxin-mediated death. Exogenous SAA activated the p38 mitogen activated protein kinase (MAP) kinase pathway, while lethal toxin mutants incapable of p38 activation were incapable of causing cell death. Chemical inhibition of the p38 activation pathway abrogated the protective effects of SAA. These data show that SAA affords protection against lethal toxin in mouse macrophages and link this response to the p38 pathway. PMID:22082566

  14. Virtual Screening Against ?-Cobratoxin

    PubMed Central

    UTSINTONG, MALEERUK; TALLEY, TODD T.; TAYLOR, PALMER W.; OLSON, ARTHUR J.; VAJRAGUPTA, OPA

    2011-01-01

    ?-Cobratoxin (Cbtx), the neurotoxin isolated from the venom of the Thai cobra Naja kaouthia, causes paralysis by preventing acetylcholine (ACh) binding to nicotinic acetylcholine receptors (nAChRs). In the current study, the region of the Cbtx molecule that is directly involved in binding to nAChRs is used as the target for anticobratoxin drug design. The crystal structure (1YI5) of Cbtx in complex with the acetylcholine binding protein (AChBP), a soluble homolog of the extracellular binding domain of nAChRs, was selected to prepare an ?-cobratoxin active binding site for docking. The amino acid residues (Ser182-Tyr192) of the AChBP structure, the binding site of Cbtx, were used as the positive control to validate the prepared Cbtx active binding site (root mean square deviation < 1.2 Å). Virtual screening of the National Cancer Institute diversity set, a library of 1990 compounds with nonredundant pharmacophore profiles, using AutoDock against the Cbtx active site, revealed 39 potential inhibitor candidates. The adapted in vitro radioligand competition assays using [3H]epibatidine and [125I]bungarotoxin against the AChBPs from the marine species, Aplysia californica (Ac), and from the freshwater snails, Lymnaea stagnalis (Ls) and Bolinus truncates (Bt), revealed 4 compounds from the list of inhibitor candidates that had micromolar to nanomolar interferences for the toxin binding to AChBPs. Three hits (NSC42258, NSC121865, and NSC134754) can prolong the survival time of the mice if administered 30 min before injection with Cbtx, but only NSC121865 and NSC134754 can prolong the survival time if injected immediately after injection with Cbtx. These inhibitors serve as novel templates/scaffolds for the development of more potent and specific anticobratoxin. PMID:19734437

  15. Lethal body burdens of polar narcotic chemicals: Chlorophenols

    SciTech Connect

    Wezel, A. van; Punte, S.; Opperhuizen, A. [RITOX, Utrecht (Netherlands). Environmental Chemistry Group

    1994-12-31

    Lethal body burdens (LBBs) were measured of low chlorinated phenols to obtain insight in the intrinsic toxicity of these compounds and in the origins of differences in toxicity between polar and nonpolar narcotic chemicals. Fathead minnows were exposed until death to 2-chlorophenol, 4-chlorophenol or 2,4-chlorophenol at fixed pH. LBBs of chlorophenols were shown to be significantly lower than LBBs of the nonpolar chlorobenzenes. The pH of the exposure doesn`t influence the LBB of chlorophenols, whereas it does influence chemical speciation and thus the uptake and the LC50 of the chlorophenols. Octanol-water partition coefficients were measured at different pH, to gain insight in the distribution of the chlorophenols between aqueous and fatty compartments inside the organism. It is shown that at a physiological pH only a fraction of the total amount of chlorophenols inside the organism is accumulated in the fatty parts of the organism. These fatty parts, including the membranes, are usually considered as the target site for narcotic chemicals. It is concluded that the concentration at the membrane needed for lethality is lower for polar narcotic chemicals than for nonpolar narcotic chemicals, implying either a different mode of action at the same target site or another target site.

  16. An outbreak of lethal adenovirus infection among different otariid species.

    PubMed

    Inoshima, Yasuo; Murakami, Tomoaki; Ishiguro, Naotaka; Hasegawa, Kazuhiro; Kasamatsu, Masahiko

    2013-08-30

    An outbreak of fatal fulminant hepatitis at a Japanese aquarium involved 3 otariids: a California sea lion (Zalophus californianus), a South African fur seal (Arctocephalus pusillus) and a South American sea lion (Otaria flavescens). In a span of about a week in February 2012, 3 otariids showed diarrhea and were acutely low-spirited; subsequently, all three animals died within a period of 3 days. Markedly increased aspartate amino transferase and alanine amino transferase activities were observed. Necrotic hepatitis and eosinophilic intranuclear inclusion bodies in liver hepatocytes and intestinal epithelial cells were observed in the South American sea lion on histological examination. Otarine adenovirus DNA was detected from the livers of all three animals by polymerase chain reaction and determination of the sequences showed that all were identical. These results suggest that a single otarine adenovirus strain may have been the etiological agent of this outbreak of fatal fulminant hepatitis among the different otariid species, and it may be a lethal threat to wild and captive otariids. This is the first evidence of an outbreak of lethal adenovirus infection among different otariid species. PMID:23643878

  17. A virulent parasite can provide protection against a lethal parasitoid.

    PubMed

    Sternberg, Eleanore D; Lefèvre, Thierry; Rawstern, Amanda H; de Roode, Jacobus C

    2011-03-01

    Hosts often become infected with multiple parasite strains or species. Previous work has shown that the outcome of infections with multiple parasite strains or species often differs significantly from that of single infections, making them a potentially important factor in determining the prevalence and spread of disease. Here we show that infection with a virulent parasite increases host survival during later exposure to a lethal parasitoid. Specifically, when monarch butterfly larvae (Danaus plexippus) are inoculated with the virulent protozoan parasite Ophryocystis elektroscirrha and then attacked by the lethal parasitoid fly Lespesia archippivora, survival is higher than when the larvae are exposed to the parasitoid only. This is potentially a result of the protozoan's requirement for host survival to obtain between-host transmission. Our findings suggest that a virulent parasite can play a protective role for its host and indicate that parasites can act as mutualists depending on the presence of other parasites. We emphasize the importance of considering infection in an ecological context, including the presence of competing parasites. PMID:21145987

  18. Suppressive Effects of Anthrax Lethal Toxin on Megakaryopoiesis

    PubMed Central

    Lin, Guan-Ling; Wang, Tsung-Pao; Lai, Yi-Ling; Lin, Ting-Kai; Hsieh, Ming-Chun; Kau, Jyh-Hwa; Huang, Hsin-Hsien; Hsu, Hui-Ling; Liao, Chi-Yuan; Sun, Der-Shan

    2013-01-01

    Anthrax lethal toxin (LT) is a major virulence factor of Bacillus anthracis. LT challenge suppresses platelet counts and platelet function in mice, however, the mechanism responsible for thrombocytopenia remains unclear. LT inhibits cellular mitogen-activated protein kinases (MAPKs), which are vital pathways responsible for cell survival, differentiation, and maturation. One of the MAPKs, the MEK1/2-extracellular signal-regulated kinase pathway, is particularly important in megakaryopoiesis. This study evaluates the hypothesis that LT may suppress the progenitor cells of platelets, thereby inducing thrombocytopenic responses. Using cord blood-derived CD34+ cells and mouse bone marrow mononuclear cells to perform in vitro differentiation, this work shows that LT suppresses megakaryopoiesis by reducing the survival of megakaryocytes. Thrombopoietin treatments can reduce thrombocytopenia, megakaryocytic suppression, and the quick onset of lethality in LT-challenged mice. These results suggest that megakaryocytic suppression is one of the mechanisms by which LT induces thrombocytopenia. These findings may provide new insights for developing feasible approaches against anthrax. PMID:23555687

  19. Effectiveness of lethal, directed wolf-depredation control in Minnesota

    USGS Publications Warehouse

    Harper, E.K.; Paul, W.J.; Mech, L.D.; Weisberg, S.

    2008-01-01

    Wolf (Canis lupus) depredations on livestock in Minnesota, USA, are an economic problem for many livestock producers, and depredating wolves are lethally controlled. We sought to determine the effectiveness of lethal control through the analysis of data from 923 government-verified wolf depredations from 1979 to 1998. We analyzed the data by 1) assessing the correlations between the number of wolves killed in response to depredations with number of depredations the following year at state and local levels, and 2) the time to the next depredation. No analysis indicated that trapping wolves substantially reduced the following year's depredations at state or local levels. However, more specific analyses indicated that in certain situations, killing wolves was more effective than no action (i.e., not trapping). For example, trapping and killing adult males decreased the re-depredation risk. At sheep farms, killing wolves was generally effective. Attempting to trap, regardless of the results, seemed more effective at reducing depredations than not trapping, suggesting that mere human activity near depredation sites might deter future depredations.

  20. Injury Risk Assessment of Non-Lethal Projectile Head Impacts

    PubMed Central

    Oukara, Amar; Nsiampa, Nestor; Robbe, Cyril; Papy, Alexandre

    2014-01-01

    Kinetic energy non-lethal projectiles are used to impart sufficient effect onto a person in order to deter uncivil or hazardous behavior with a low probability of permanent injury. Since their first use, real cases indicate that the injuries inflicted by such projectiles may be irreversible and sometimes lead to death, especially for the head impacts. Given the high velocities and the low masses involved in such impacts, the assessment approaches proposed in automotive crash tests and sports may not be appropriate. Therefore, there is a need of a specific approach to assess the lethality of these projectiles. In this framework, some recent research data referred in this article as “force wall approach” suggest the use of three lesional thresholds (unconsciousness, meningeal damages and bone damages) that depend on the intracranial pressure. Three corresponding critical impact forces are determined for a reference projectile. Based on the principle that equal rigid wall maximal impact forces will produce equal damage on the head, these limits can be determined for any other projectile. In order to validate the consistence of this innovative method, it is necessary to compare the results with other existing assessment methods. This paper proposes a comparison between the “force wall approach” and two different head models. The first one is a numerical model (Strasbourg University Finite Element Head Model-SUFEHM) from Strasbourg University; the second one is a mechanical surrogate (Ballistics Load Sensing Headform-BLSH) from Biokinetics. PMID:25400712

  1. Injury risk assessment of non-lethal projectile head impacts.

    PubMed

    Oukara, Amar; Nsiampa, Nestor; Robbe, Cyril; Papy, Alexandre

    2014-01-01

    Kinetic energy non-lethal projectiles are used to impart sufficient effect onto a person in order to deter uncivil or hazardous behavior with a low probability of permanent injury. Since their first use, real cases indicate that the injuries inflicted by such projectiles may be irreversible and sometimes lead to death, especially for the head impacts. Given the high velocities and the low masses involved in such impacts, the assessment approaches proposed in automotive crash tests and sports may not be appropriate. Therefore, there is a need of a specific approach to assess the lethality of these projectiles. In this framework, some recent research data referred in this article as "force wall approach" suggest the use of three lesional thresholds (unconsciousness, meningeal damages and bone damages) that depend on the intracranial pressure. Three corresponding critical impact forces are determined for a reference projectile. Based on the principle that equal rigid wall maximal impact forces will produce equal damage on the head, these limits can be determined for any other projectile. In order to validate the consistence of this innovative method, it is necessary to compare the results with other existing assessment methods. This paper proposes a comparison between the "force wall approach" and two different head models. The first one is a numerical model (Strasbourg University Finite Element Head Model-SUFEHM) from Strasbourg University; the second one is a mechanical surrogate (Ballistics Load Sensing Headform-BLSH) from Biokinetics. PMID:25400712

  2. Best Practice in Preschool Screening.

    ERIC Educational Resources Information Center

    Rosenkoetter, Sharon E.; Wanska, Susan K.

    This brief guide outlines best practices in preschool screening for the presence of possible disabilities. It covers: a definition of screening, its history, the rationale for screening, requirements for effective screening measures (reliability, validity, fairness, utility), models for preschool screening, evaluation of screening procedures, and…

  3. A homoallelic Gly[sup 317] [yields] Asp mutation in ALPL causes the perinatal (lethal) form of hypophosphatasia in Canadian mennonites

    SciTech Connect

    Greenberg, C.R.; Taylor, C.D.; Haworth, J.C.; Seargeant, L.E.; Phillipps, S.; Triggs-Raine, B.; Chodirker, B.N. (Univ. of Manitoba, Winnepeg (Canada))

    1993-07-01

    The authors have discovered a single homoallelic nucleotide substitution as the putative cause of the perinatal (lethal) form of hypophosphatasia in Canadian Mennonites. Previous linkage and haplotype analysis in this population suggested that a single mutational event was responsible for this autosomal recessive form of hypophosphatasia. The mutation is a guanosine-to-adenosine substitution at nucleotide position 1177 in exon 10 of the tissue nonspecific (liver/bone/kidney) alkaline phosphatase gene. This Gly[sup 317] [yields] Asp mutation segregates exclusively with the heterozygote phenotype previously assigned by biochemical testing (maximum combined lod score of 18.24 at [theta] = 0.00). This putative disease-causing mutation has not been described in controls nor in other non-Mennonite probands with both lethal and nonlethal forms of hypophosphatasia studied to date. This Gly[sup 317] [yields] Asp mutation changes a polar glycine to an acidic aspartate at amino acid position 317 within the highly conserved active site region of the 507-amino-acid polypeptide. Carrier screening for this lethal mutation in a high-risk population is now feasible. 15 refs., 2 figs.

  4. Impulsivity, aggression and brain structure in high and low lethality suicide attempters with borderline personality disorder

    PubMed Central

    Soloff, Paul; White, Richard; Diwadkar, Vaibhav A.

    2014-01-01

    Impulsivity and aggressiveness are trait dispositions associated with the vulnerability to suicidal behavior across diagnoses. They are associated with structural and functional abnormalities in brain networks involved in regulation of mood, impulse and behavior. They are also core characteristics of borderline personality disorder (BPD), a disorder defined, in part, by recurrent suicidal behavior. We assessed the relationships between personality traits, brain structure and lethality of suicide attempts in 51 BPD attempters using multiple regression analyses on structural MRI data. BPD was diagnosed by the Diagnostic Interview for Borderline Patients-revised, impulsivity by the Barratt Impulsiveness Scale (BIS), aggression by the Brown-Goodwin Lifetime History of Aggression (LHA), and high lethality by a score of 4 or more on the Lethality Rating Scale (LRS). Sixteen High Lethality attempters were compared to 35 Low Lethality attempters, with no significant differences noted in gender, co-morbidity, childhood abuse, BIS or LHA scores. Degree of medical lethality (LRS) was negatively related to gray matter volumes across multiple fronto-temporal-limbic regions. Effects of impulsivity and aggression on gray matter volumes discriminated High from Low Lethality attempters and differed markedly within lethality groups. Lethality of suicide attempts in BPD may be related to the mediation of these personality traits by specific neural networks. PMID:24656768

  5. The role of sub-lethal weapons in human rights abuse.

    PubMed

    Wright, S

    2001-01-01

    This article is based on two recent reports contracted by the European Parliament (EP), which assessed sub-lethal weapons as flexible tools of political control. It analyses the role and function of existing weapons systems in human rights abuses using examples from Indonesia, Israel, Kenya, Northern Ireland and Turkey. These weapons are designed to 'appear' rather than 'be' safe and, since they augment rather than replace lethal technologies, their use can distort conflicts and actually bridge the firewall between use of less-lethal and lethal technologies. PMID:11578040

  6. Active versus Passive Screen Time for Young Children

    ERIC Educational Resources Information Center

    Sweetser, Penelope; Johnson, Daniel; Ozdowska, Anne; Wyeth, Peta

    2012-01-01

    In this paper we report some initial findings from our investigations into the Australian Government's Longitudinal Study of Australian Children dataset. It is revealed that the majority of Australian children are exceeding the government's Screen Time recommendations and that most of their screen time is spent as TV viewing, as opposed to video…

  7. The art and design of genetic screens: zebrafish

    Microsoft Academic Search

    E. Elizabeth Patton; Leonard I. Zon

    2001-01-01

    Inventive genetic screens in zebrafish are revealing new genetic pathways that control vertebrate development, disease and behaviour. By exploiting the versatility of zebrafish, biological processes that had been previously obscured can be visualized and many of the responsible genes can be isolated. Coupled with gene knockdown and overexpression technologies, and small-molecule-induced phenotypes, genetic screens in zebrafish provide a powerful system

  8. Screening Spouse Abusers for Child Abuse Potential.

    ERIC Educational Resources Information Center

    Milner, Joel S.; Gold, Ruth G.

    1986-01-01

    Investigated the ability of the Child Abuse Potential Inventory to screen for child abuse in a group of spouse abusers. The completed, valid protocols revealed that 36.5 percent of the spouse abusers had elevated child abuse scores, while only 9.1 percent of the nonabusers had elevated abuse scores. (Author/BL)

  9. Automated Groundwater Screening

    SciTech Connect

    Taylor, Glenn A.; Collard, Leonard, B.

    2005-10-31

    The Automated Intruder Analysis has been extended to include an Automated Ground Water Screening option. This option screens 825 radionuclides while rigorously applying the National Council on Radiation Protection (NCRP) methodology. An extension to that methodology is presented to give a more realistic screening factor for those radionuclides which have significant daughters. The extension has the promise of reducing the number of radionuclides which must be tracked by the customer. By combining the Automated Intruder Analysis with the Automated Groundwater Screening a consistent set of assumptions and databases is used. A method is proposed to eliminate trigger values by performing rigorous calculation of the screening factor thereby reducing the number of radionuclides sent to further analysis. Using the same problem definitions as in previous groundwater screenings, the automated groundwater screening found one additional nuclide, Ge-68, which failed the screening. It also found that 18 of the 57 radionuclides contained in NCRP Table 3.1 failed the screening. This report describes the automated groundwater screening computer application.

  10. Composite scintillator screen

    DOEpatents

    Zeman, Herbert D. (1687 Peach St., Memphis, TN 38112)

    1994-01-01

    A scintillator screen for an X-ray system includes a substrate of low-Z material and bodies of a high-Z material embedded within the substrate. By preselecting the size of the bodies embedded within the substrate, the spacial separation of the bodies and the thickness of the screen, the sensitivity of the screen to X-rays within a predetermined energy range can be predicted.

  11. ScreenFlow

    NSDL National Science Digital Library

    2008-01-01

    Capturing images and screen shots can be tricky, and it's nice to hear about new applications that can help out with such tasks. ScreenFlow allows users to create screen recordings and it also includes an array of editing options. Visitors should note that this is a trial version, and that the full-featured version costs $99. This version is compatible with computers running Mac OS X 10.5 Leopard.

  12. Breast screening revisited.

    PubMed

    Agrawal, Alka; Tripathi, Prem; Sahu, Abhinav; Daftary, Jalpa

    2014-01-01

    Breast screening is the medical screening of asymptomatic, apparently healthy women for breast lump in an attempt to achieve an earlier diagnosis. The assumption is that the early detection will improve outcomes. In western countries, breast screening programs have led to a significant reduction in mortality and improved prognosis of patients with breast cancer. However in India, although the number of breast cancer are on the rise there is no such organized program. This article emphasizes on the importance of breast screening and protocol to be followed in our country where it can have significant impact on the prognosis. PMID:25657940

  13. Batrachochytrium dendrobatidis infection and lethal chytridiomycosis in caecilian amphibians (Gymnophiona).

    PubMed

    Gower, David J; Doherty-Bone, Thomas; Loader, Simon P; Wilkinson, Mark; Kouete, Marcel T; Tapley, Benjamin; Orton, Frances; Daniel, Olivia Z; Wynne, Felicity; Flach, Edmund; Müller, Hendrik; Menegon, Michele; Stephen, Ian; Browne, Robert K; Fisher, Mathew C; Cunningham, Andrew A; Garner, Trenton W J

    2013-06-01

    Batrachochytrium dendrobatidis (Bd) is commonly termed the 'amphibian chytrid fungus' but thus far has been documented to be a pathogen of only batrachian amphibians (anurans and caudatans). It is not proven to infect the limbless, generally poorly known, and mostly soil-dwelling caecilians (Gymnophiona). We conducted the largest qPCR survey of Bd in caecilians to date, for more than 200 field-swabbed specimens from five countries in Africa and South America, representing nearly 20 species, 12 genera, and 8 families. Positive results were recovered for 58 specimens from Tanzania and Cameroon (4 families, 6 genera, 6+ species). Quantities of Bd were not exceptionally high, with genomic equivalent (GE) values of 0.052-17.339. In addition, we report the first evidence of lethal chytridiomycosis in caecilians. Mortality in captive (wild-caught, commercial pet trade) Geotrypetes seraphini was associated with GE scores similar to those we detected for field-swabbed, wild animals. PMID:23677560

  14. Clinical Effects and Lethal and Forensic Aspects of Propofol*

    PubMed Central

    Levy, Richard J.

    2010-01-01

    Propofol is a potent intravenous anesthetic agent that rapidly induces sedation and unconsciousness. The potential for propofol dependency, recreational use and abuse has only recently been recognized and several cases of accidental overdose and suicide have emerged. In addition, the first documented case of murder using propofol was reported a few months ago and a high profile case of suspected homicide with propofol is currently under investigation. A number of analytical methods have been employed to detect and quantify propofol concentrations in biological specimens. The reported propofol related deaths and post-mortem blood and tissue levels are reviewed. Importantly, limitations of propofol detection are discussed and future considerations are presented. Because propofol has the potential for diversion with lethal consequences, the forensic scientist must have a basic understanding of its clinical indications and uses, pharmacologic properties, and detection methods. In addition, medical institutions should develop systems to prevent and detect diversion of this potential drug of abuse. PMID:20950316

  15. DDE in birds: Lethal residues and loss rates

    USGS Publications Warehouse

    Stickel, W.H.; Stickel, L.F.; Dyrland, R.A.; Hughes, D.L.

    1984-01-01

    Lethal brain residues of DDE were determined experimentally in four species of wild birds (male common grackels (Quiscalus quiscula ), immature female red-winged blackbirds (Agelaius phoeniceus ), adult male brown-headed cowbirds (Molathrus ater ), and immature female starlings (Sturnus vulgaris ) given dietary dosage of 1,500 ppm DDE until one-half had died, then sacrificing the survivors, chemically analyzing the tissues, and comparing results in dead birds and survivors. In all species, residues of 300 to 400 ppm of DDE in the brain were considered to show increasing likelihood of death from DDE, confirming results of an earlier study with a single species. Body residues (ppm wet weight) were not diagnostic, overlapping grossly in dead birds and survivors, but averaging higher in survivors.

  16. Aroclor 1254 residues in birds: Lethal levels and loss rates

    USGS Publications Warehouse

    Stickel, W.H.; Stickel, L.F.; Dyrland, R.A.; Hughes, D.L.

    1984-01-01

    Lethal residues of polychlorinated biphenyls (PCBs) were determined experimentally in four species of wild birds (male common grackles (Quiscalus quiscula ), immature female red-winged blackbirds (Agelaius phoeniceus ), adult male brown-headed cowbirds (Molothrus ater ) and immature female starlings (Sturnus vulgaris)) given dietary dosage of 1,500 ppm of Aroclor 1254) until one-half had died, sacrificing the survivors, chemically analyzing the tissues, and comparing results in dead birds and survivors. For all species, residues of 310 ppm or higher in the brain showed increasing likelihood of death from PCB poisoning. Residues in dead birds did not differ among species except for starlings (Sturnus vulgaris ), which averaged slightly lower than the others. However, the species differed in the length of time to 50% mortality and in the levels of PCBs in brains at sacrifice.

  17. Autotaxin Overexpression Causes Embryonic Lethality and Vascular Defects

    PubMed Central

    Yukiura, Hiroshi; Kano, Kuniyuki; Kise, Ryoji; Inoue, Asuka; Aoki, Junken

    2015-01-01

    Autotaxin (ATX) is a secretory protein, which converts lysophospholipids to lysophosphatidic acid (LPA), and is essential for embryonic vascular formation. ATX is abundantly detected in various biological fluids and its level is elevated in some pathophysiological conditions. However, the roles of elevated ATX levels remain to be elucidated. In this study, we generated conditional transgenic (Tg) mice overexpressing ATX and examined the effects of excess LPA signalling. We found that ATX overexpression in the embryonic period caused severe vascular defects and was lethal around E9.5. ATX was conditionally overexpressed in the neonatal period using the Cre/loxP system, which resulted in a marked increase in the plasma LPA level. This resulted in retinal vascular defects including abnormal vascular plexus and increased vascular regression. Our findings indicate that the ATX level must be carefully regulated to ensure coordinated vascular formation PMID:25992708

  18. Arsenic-induced sub-lethal stress reprograms human bronchial epithelial cells to CD61¯ cancer stem cells.

    PubMed

    Chang, Qingshan; Chen, Bailing; Thakur, Chitra; Lu, Yongju; Chen, Fei

    2014-03-15

    In the present report, we demonstrate that sub-lethal stress induced by consecutive exposure to 0.25 µM arsenic (As3+) for six months can trigger reprogramming of the human bronchial epithelial cell (BEAS-2B) to form cancer stem cells (CSCs) without forced introduction of the stemness transcription factors. These CSCs formed from As3+-induced sub-lethal stress featured with an increased expression of the endogenous stemness genes, including Oct4, Sox2, Klf4, Myc, and others that are associated with the pluripotency and self-renewal of the CSCs. Flow cytometry analysis indicated that 90% of the CSC cells are CD61¯, whereas 100% of the parental cells are CD61+. These CD61¯ CSCs are highly tumorigenic and metastatic to the lung in xenotransplantation tests in NOD/SCID Il2r?-/- mice. Additional tests also revealed that the CD61¯ CSCs showed a significant decrease in the expression of the genes important for DNA repair and oxidative phosphorylation. To determine the clinical relevance of the above findings, we stratified human lung cancers based on the level of CD61 protein and found that CD61low cancer correlates with poorer survival of the patients. Such a correlation was also observed in human breast cancer and ovarian cancer. Taken together, our findings suggest that in addition to the traditional approaches of enforced introduction of the exogenous stemness circuit transcription factors, sub-lethal stress induced by consecutive low dose As3+ is also able to convert non-stem cells to the CSCs. PMID:24675390

  19. Single Amino Acid Exchanges in Separate Domains of the Drosophila Serendipity ? Zinc Finger Protein Cause Embryonic and Sex Biased Lethality

    PubMed Central

    Crozatier, M.; Kongsuwan, K.; Ferrer, P.; Merriam, J. R.; Lengyel, J. A.; Vincent, A.

    1992-01-01

    The Drosophila serendipity (sry) delta (?) zinc finger protein is a sequence-specific DNA binding protein, maternally inherited by the embryo and present in nuclei of transcriptionally active cells throughout fly development. We report here the isolation and characterization of four ethyl methanesulfate-induced zygotic lethal mutations of different strengths in the sry ? gene. For the stronger allele, all of the lethality occurs during late embryogenesis or the first larval instar. In the cases of the three weaker alleles, most of the lethality occurs during pupation; moreover, those adult escapers that emerge are sterile males lacking partially or completely in spermatozoa bundles. Genetic analysis of sry ? thus indicates that it is an essential gene, whose continued expression throughout the life cycle, notably during embryogenesis and pupal stage, is required for viability. Phenotypic analysis of sry ? hemizygote escaper males further suggests that sry ? may be involved in regulation of two different sets of genes: genes required for viability and genes involved in gonadal development. All four sry ? alleles are fully rescued by a wild-type copy of sry ?, but not by an additional copy of the sry ? gene, reinforcing the view that, although structurally related, these two genes exert distinct functions. Molecular characterization of the four sry ? mutations revealed that these mutations correspond to single amino acid replacements in the sry ? protein. Three of these replacements map to the same (third out of seven) zinc finger in the carboxy-terminal DNA binding domain; interestingly, none affects the zinc finger consensus residues. The fourth mutation is located in the NH(2)-proximal part of the protein, in a domain proposed to be involved in specific protein-protein interactions. PMID:1516821

  20. Lethal Mutagenesis of Poliovirus Mediated by a Mutagenic Pyrimidine Analogue?

    PubMed Central

    Graci, Jason D.; Harki, Daniel A.; Korneeva, Victoria S.; Edathil, Jocelyn P.; Too, Kathleen; Franco, David; Smidansky, Eric D.; Paul, Aniko V.; Peterson, Blake R.; Brown, Daniel M.; Loakes, David; Cameron, Craig E.

    2007-01-01

    Lethal mutagenesis is the mechanism of action of ribavirin against poliovirus (PV) and numerous other RNA viruses. However, there is still considerable debate regarding the mechanism of action of ribavirin against a variety of RNA viruses. Here we show by using T7 RNA polymerase-mediated production of PV genomic RNA, PV polymerase-catalyzed primer extension, and cell-free PV synthesis that a pyrimidine ribonucleoside triphosphate analogue (rPTP) with ambiguous base-pairing capacity is an efficient mutagen of the PV genome. The in vitro incorporation properties of rPTP are superior to ribavirin triphosphate. We observed a log-linear relationship between virus titer reduction and the number of rPMP molecules incorporated. A PV genome encoding a high-fidelity polymerase was more sensitive to rPMP incorporation, consistent with diminished mutational robustness of high-fidelity PV. The nucleoside (rP) did not exhibit antiviral activity in cell culture, owing to the inability of rP to be converted to rPMP by cellular nucleotide kinases. rP was also a poor substrate for herpes simplex virus thymidine kinase. The block to nucleoside phosphorylation could be bypassed by treatment with the P nucleobase, which exhibited both antiviral activity and mutagenesis, presumably a reflection of rP nucleotide formation by a nucleotide salvage pathway. These studies provide additional support for lethal mutagenesis as an antiviral strategy, suggest that rPMP prodrugs may be highly efficacious antiviral agents, and provide a new tool to determine the sensitivity of RNA virus genomes to mutagenesis as well as interrogation of the impact of mutational load on the population dynamics of these viruses. PMID:17686844

  1. Tumor expression of adiponectin receptor 2 and lethal prostate cancer.

    PubMed

    Rider, Jennifer R; Fiorentino, Michelangelo; Kelly, Rachel; Gerke, Travis; Jordahl, Kristina; Sinnott, Jennifer A; Giovannucci, Edward L; Loda, Massimo; Mucci, Lorelei A; Finn, Stephen

    2015-06-01

    To investigate the role of adiponectin receptor 2 (AdipoR2) in aggressive prostate cancer we used immunohistochemistry to characterize AdipoR2 protein expression in tumor tissue for 866 men with prostate cancer from the Physicians' Health Study and the Health Professionals Follow-up Study. AdipoR2 tumor expression was not associated with measures of obesity, pathological tumor stage or prostate-specific antigen (PSA) at diagnosis. However, AdipoR2 expression was positively associated with proliferation as measured by Ki-67 expression quartiles (P-trend < 0.0001), with expression of fatty acid synthase (P-trend = 0.001), and with two measures of angiogenesis (P-trend < 0.1). An inverse association was observed with apoptosis as assessed by the TUNEL assay (P-trend = 0.006). Using Cox proportional hazards regression and controlling for age at diagnosis, Gleason score, year of diagnosis category, cohort and baseline BMI, we identified a statistically significant trend for the association between quartile of AdipoR2 expression and lethal prostate cancer (P-trend = 0.02). The hazard ratio for lethal prostate cancer for the two highest quartiles, as compared to the two lowest quartiles, of AdipoR2 expression was 1.9 (95% confidence interval [CI]: 1.2-3.0). Results were similar when additionally controlling for categories of PSA at diagnosis and Ki-67 expression quartiles. These results strengthen the evidence for the role of AdipoR2 in prostate cancer progression. PMID:25863129

  2. Lysosome and Phagosome Stability in Lethal Cell Injury

    PubMed Central

    Hawkins, Hal K.; Ericsson, Jan L. E.; Biberfeld, Peter; Trump, Benjamin F.

    1972-01-01

    In two types of cell injury in a tissue culture system, the possibility was tested that lysosome rupture may be a lethal cellular reaction to injury, and thus an important general cause of irreversibility of damage in injured tissue. Prior labeling of secondary lysosomes with the fluorochrome acridine orange, or with ferritin, was used to trace changes in lysosomes after applying an injury. The metabolic inhibitors iodoacetate and cyanide were used together to block the cell's energy supply, or attachment of antiserum and subsequent complement attack were used to damage the surface membrane, producing rapid loss of cell volume control. Living cells were studied by time-lapse phase-contrast cinemicrography and fluorescence microscopy, and samples were fixed at intervals for electron microscopy. The cytolytic action of complement was lethal to sensitized cells within 2 hours, but results showed that lysosomes did not rupture for approximately 4 hours and in fact did not release the fluorescent dye until after reaching the postmortem necrotic phase of injury. Cells treated with metabolic inhibitors also showed irreversible alterations, while lysosomes remained intact and retained the ferritin marker. The fluorochrome marker, acridine orange, escaped from lysosomes early after metabolic injury, but the significance of this observation is not clear. The results are interpreted as evidence against the concept that lysosome rupture threatens the survival of injured cells. The original suicide bag mechanism of cell damage thus is apparently not operative in the systems studied. Lysosomes appear to be relatively stable organelles which, following injury of the types studied, burst only after cell death, acting then as scavengers which help to clear cellular debris. ImagesFigs 5-7Fig 18Fig 19Fig 20Figs 21-23Fig 8Fig 9Fig 10Fig 11Figs 24-27Fig 12Figs 13 and 14Fig 1Fig 2Fig 3Fig 4Fig 15Fig 16Fig 17 PMID:4340333

  3. An exome sequencing strategy to diagnose lethal autosomal recessive disorders.

    PubMed

    Ellard, Sian; Kivuva, Emma; Turnpenny, Peter; Stals, Karen; Johnson, Matthew; Xie, Weijia; Caswell, Richard; Lango Allen, Hana

    2015-03-01

    Rare disorders resulting in prenatal or neonatal death are genetically heterogeneous. For some conditions, affected fetuses can be diagnosed by ultrasound scan, but this is not usually possible until mid-gestation. There is often limited fetal DNA available for investigation. We investigated a strategy for diagnosing autosomal recessive lethal disorders in non-consanguineous pedigrees with multiple affected fetuses. Exome sequencing was performed to identify genes where each parent is heterozygous for a rare non-synonymous-coding or splicing variant. Putative pathogenic variants were tested for cosegregation in affected fetuses and unaffected siblings. In eight couples of European ancestry, we found on average 1.75 genes (range 0-4) where both parents were heterozygous for rare potentially deleterious variants. A proof-of-principle study detected heterozygous DYNC2H1 variants in a couple whose five fetuses had short-rib polydactyly. Prospective analysis of two couples with multiple pregnancy terminations for fetal akinesia syndrome was performed and a diagnosis was obtained in both the families. The first couple were each heterozygous for a previously reported GLE1 variant, p.Arg569His or p.Val617Met; both were inherited by their two affected fetuses. The second couple were each heterozygous for a novel RYR1 variant, c.14130-2A>G or p.Ser3074Phe; both were inherited by their three affected fetuses but not by their unaffected child. Biallelic GLE1 and RYR1 disease-causing variants have been described in other cases with fetal akinesia syndrome. We conclude that exome sequencing of parental samples can be an effective tool for diagnosing lethal recessive disorders in outbred couples. This permits early prenatal diagnosis in future pregnancies. PMID:24961629

  4. Foal with Overo lethal white syndrome born to a registered quarter horse mare

    PubMed Central

    Lightbody, Tamara

    2002-01-01

    A 16-hour-old white foal, born to a registered quarter horse mare, was examined for signs of colic. The foal had Overo lethal white syndrome, which causes ileocolonic agangliosis. This was confirmed by DNA testing. Since there is no treatment for Overo lethal white syndrome, the foal was euthanized. PMID:12240532

  5. Examining the Impact of Psychiatric Diagnosis and Comorbidity on the Medical Lethality of Adolescent "Suicide Attempts"

    ERIC Educational Resources Information Center

    Mc Manama O'Brien, Kimberly H.; Berzin, Stephanie C.

    2012-01-01

    Specific psychiatric diagnoses and comorbidity patterns were examined to determine if they were related to the medical lethality of "suicide attempts" among adolescents presenting to an urban general hospital (N = 375). Bivariate analysis showed that attempters with substance abuse disorders had higher levels of lethality than attempters without…

  6. Experimental studies on effects of sub-lethal dose of pulsed electric field on Hela cells

    Microsoft Academic Search

    Chengxiang Li; Chenguo Yao; Yanshan Qin; Yan Mi; Xiaoyun Liu; Zhengai Xiong

    2010-01-01

    In this paper, we studied the effects of sub-lethal dose of pulsed electric field on cervical cancer cells to achieve an efficient IRE by experiments on Hela cells. MTT and flow cytometry were used to detect the cell viability and cell circle. From the experiment results, it could be concluded that cell proliferation is inhibited by the sub-lethal dose field

  7. PEX11  Deficiency Is Lethal and Impairs Neuronal Migration but Does Not Abrogate Peroxisome Function

    Microsoft Academic Search

    Xiaoling Li; Eveline Baumgart; James C. Morrell; Gerardo Jimenez-Sanchez; David Valle; Stephen J. Gould

    2002-01-01

    Zellweger syndrome is a lethal neurological disorder characterized by severe defects in peroxisomal protein import. The resulting defects in peroxisome metabolism and the accumulation of peroxisomal substrates are thought to cause the other Zellweger syndrome phenotypes, including neuronal migration defects, hypotonia, a developmental delay, and neonatal lethality. These phenotypes are also manifested in mouse models of Zellweger syndrome generated by

  8. Co-lethality studied as an asset against viral drug escape: the HIV protease case

    PubMed Central

    2010-01-01

    Background Co-lethality, or synthetic lethality is the documented genetic situation where two, separately non-lethal mutations, become lethal when combined in one genome. Each mutation is called a "synthetic lethal" (SL) or a co-lethal. Like invariant positions, SL sets (SL linked couples) are choice targets for drug design against fast-escaping RNA viruses: mutational viral escape by loss of affinity to the drug may induce (synthetic) lethality. Results From an amino acid sequence alignment of the HIV protease, we detected the potential SL couples, potential SL sets, and invariant positions. From the 3D structure of the same protein we focused on the ones that were close to each other and accessible on the protein surface, to possibly bind putative drugs. We aligned 24,155 HIV protease amino acid sequences and identified 290 potential SL couples and 25 invariant positions. After applying the distance and accessibility filter, three candidate drug design targets of respectively 7 (under the flap), 4 (in the cantilever) and 5 (in the fulcrum) amino acid positions were found. Conclusions These three replication-critical targets, located outside of the active site, are key to our anti-escape strategy. Indeed, biological evidence shows that 2/3 of those target positions perform essential biological functions. Their mutational variations to escape antiviral medication could be lethal, thus limiting the apparition of drug-resistant strains. Reviewers This article was reviewed by Arcady Mushegian, Shamil Sunyaev and Claus Wilke. PMID:20565756

  9. BIOASSAY PROTOCOL FOR LETHAL AND SUBLETHAL EFFECTS OF FUNGAL PATHOGENS ON CHRYSOPERLA CARNEA

    EPA Science Inventory

    This practice describes procedures for evaluating the lethal and sub-lethal effects of exposure to fungal pathogens on larvae and adults of the predatory insect Chrysoperla carnea (Stephens). his practice was developed and tested with the fungal insect pathogen Beauveria bassiana...

  10. Evaluating the Predictive Validity of Suicidal Intent and Medical Lethality in Youth

    ERIC Educational Resources Information Center

    Sapyta, Jeffrey; Goldston, David B.; Erkanli, Alaattin; Daniel, Stephanie S.; Heilbron, Nicole; Mayfield, Andrew; Treadway, S. Lyn

    2012-01-01

    Objectives: To examine whether suicidal intent and medical lethality of past suicide attempts are predictive of future attempts, the association between intent and lethality, and the consistency of these characteristics across repeated attempts among youth. Method: Suicide attempts in a 15-year prospective study of 180 formerly psychiatrically…

  11. Impairment of dendritic cells and adaptive immunity by anthrax lethal toxin

    Microsoft Academic Search

    Anshu Agrawal; Jai Lingappa; Stephen H. Leppla; Sudhanshu Agrawal; Abdul Jabbar; Conrad Quinn; Bali Pulendran

    2003-01-01

    Anthrax poses a clear and present danger as an agent of biological terrorism. Infection with Bacillus anthracis, the causative agent of anthrax, if untreated can result in rampant bacteraemia, multisystem dysfunction and death. Anthrax lethal toxin (LT) is a critical virulence factor of B. anthracis, which occurs as a complex of protective antigen and lethal factor. Here we demonstrate that

  12. The Danger Assessment: Validation of a Lethality Risk Assessment Instrument for Intimate Partner Femicide

    ERIC Educational Resources Information Center

    Campbell, Jacquelyn C.; Webster, Daniel W.; Glass, Nancy

    2009-01-01

    The Danger Assessment (DA) is an instrument designed to assess the likelihood of lethality or near lethality occurring in a case of intimate partner violence. This article describes the development, psychometric validation, and suggestions for use of the DA. An 11-city study of intimate partner femicide used multivariate analysis to test the…

  13. Effects of Training with Lethal Chemicals on Job Proficiency and Job Confidence.

    ERIC Educational Resources Information Center

    Smith, Paula; And Others

    A study was designed to determine if soldiers trained to use chemical agents are more proficient in performing their jobs in an environment where lethal chemical agents are used and more confident of their ability to survive. A treatment group, composed of 150 soldiers, knew that their training would involve lethal agents in the Chemical…

  14. Genetically modified anthrax lethal toxin safely delivers whole HIV protein antigens into the

    E-print Network

    Lieberman, Judy

    , 2000 (received for review January 24, 2000) Bacillus anthrax lethal toxin can be engineered to deliverGenetically modified anthrax lethal toxin safely delivers whole HIV protein antigens compartment of mammalian cells. The engineered anthrax toxin vaccine appears unlikely to induce an antibody

  15. A fragment of anthrax lethal factor delivers proteins to the cytosol without requiring protective antigen

    E-print Network

    Lieberman, Judy

    A fragment of anthrax lethal factor delivers proteins to the cytosol without requiring protective by Elkan R. Blout, Harvard Medical School, Cambridge, MA, April 2, 2003 (received for review March 10, 2003) Anthrax protective antigen (PA) is a 735-aa polypeptide that facili- tates the exit of anthrax lethal

  16. Cellular and systemic effects of anthrax lethal toxin and edema toxin

    Microsoft Academic Search

    Mahtab Moayeri; Stephen H. Leppla

    2009-01-01

    Anthrax lethal toxin (LT) and edema toxin (ET) are the major virulence factors of anthrax and can replicate the lethality and symptoms associated with the disease. This review provides an overview of our current understanding of anthrax toxin effects in animal models and the cytotoxicity (necrosis and apoptosis) induced by LT in different cells. A brief reexamination of early historic

  17. Anthrax Lethal Toxin-Mediated Killing of Human and Murine Dendritic Cells Impairs

    E-print Network

    Brojatsch, Jürgen

    Anthrax Lethal Toxin-Mediated Killing of Human and Murine Dendritic Cells Impairs the Adaptive anthracis interferes with host defenses by releasing anthrax lethal toxin (LT), which inactivates mitogen that anthrax LT impairs adaptive immunity by specifically targeting DCs. This may represent an immune- evasion

  18. Human synthetic lethal inference as potential anti-cancer target gene detection

    Microsoft Academic Search

    Nuria Conde-Pueyo; Andreea Munteanu; Ricard V Solé; Carlos Rodríguez-Caso

    2009-01-01

    BACKGROUND: Two genes are called synthetic lethal (SL) if mutation of either alone is not lethal, but mutation of both leads to death or a significant decrease in organism's fitness. The detection of SL gene pairs constitutes a promising alternative for anti-cancer therapy. As cancer cells exhibit a large number of mutations, the identification of these mutated genes' SL partners

  19. ORIGINAL ARTICLE Evolution of the Sex-lethal Gene in Insects and Origin

    E-print Network

    Nei, Masatoshi

    ORIGINAL ARTICLE Evolution of the Sex-lethal Gene in Insects and Origin of the Sex-Determination Sex-lethal (Sxl) functions as the switch gene for sex-determination in Drosophila melanogaster protein's sex- determination function have remained largely unknown. In this study, we explore

  20. Newborn screening in Indonesia.

    PubMed

    Rustama, Diet S; Fadil, M Ryadi; Harahap, Elly R; Primadi, Aris

    2003-01-01

    In Indonesia, newborn screening is not yet a policy, and the incidence of preventable causes of mental retardation detected by newborn screening is not known. Congenital hypothyroidism (CH) is not infrequent. Without a screening program, unrecognized CH patients were neglected for years. Since May 1999, the International Atomic Energy Agency (IAEA) has assisted in starting a CH Newborn Screening Project to estimate the local incidence of CH and to evaluate the problems associated with the screening. In June 2000, a pilot study was conducted using primary TSH measurement, supplemented by T4 in infants with elevated TSH. The target was to screen 12,000 newborn infants, using cord blood serum taken at birth, or a heel prick between 2 to 6 days of age. Between June 2000 and February 2001, 3,534 neonates born in 4 hospitals were screened using cord blood serum taken at birth (recall rate 3.3%). From March 2001 onwards, the heel prick method was used and participating hospitals increased from 4 to 7. Using this approach, until August 2001, 3,309 samples were analysed and the recall rate was much lower (0.64%). The number of unsatisfactory samples was relatively high due to an unstable process of blood collection. Parental refusal and low acceptance of screening among policy makers resulted from lack of awareness of the dangers of CH, and the screening program was not considered a health priority. Recall of patients after screening was a major barrier, with problems in tracking patients arising from urbanization and a high rate of relocation. To advance the CH screening program nationwide, infrastructure must be improved along with the recall system, and education as well as information campaigns for parents and medical professionals must be intensified. The Department of Health must be persuaded to give a national mandate. PMID:15906701

  1. Cost-effectiveness of colon cancer screening.

    PubMed

    Lieberman, D

    1991-12-01

    The cost-effectiveness of two colon cancer-screening strategies was compared. The first strategy mirrors the recommendations of the American Cancer Society and includes sigmoidoscopy starting at age 50, and yearly fecal occult blood testing. The second strategy is screening with colonoscopy. The analysis revealed that the 10-yr cost of screening with sigmoidoscopy is nearly $1,700, compared with nearly $2,500 for colonoscopy, using prevailing procedure costs. This difference can be reduced by lowering the cost of normal colonoscopies. The cost of identifying one patient with an adenomatous polyp is $8,766 with sigmoidoscopy, compared to $5,988 with colonoscopy because of the higher detection rate with colonoscopy. The calculated cost of preventing one death from colon cancer is $444,133 with sigmoidoscopy versus $347,214 with colonoscopy. In conclusion, colon cancer prevention with current screening methods is very costly. Screening with sigmoidoscopy and fecal occult blood testing may not be cost-effective, compared to screening with colonoscopy. PMID:1962624

  2. Lethal infection by Bordetella pertussis mutants in the infant mouse model.

    PubMed Central

    Weiss, A A; Goodwin, M S

    1989-01-01

    Different aspects of lethal infection of infant mice with Bordetella pertussis were examined. Mutants deficient in vir-regulated genes were tested for the ability to cause a lethal infection in the infant mouse model. Adenylate cyclase toxin-hemolysin and pertussis toxin were required to cause a lethal infection at low doses. Mixed infection caused by challenging the mice with an equal number of pertussis toxin and adenylate cyclase toxin-hemolysin mutants at a dose at which neither alone was lethal was also unable to cause a lethal infection. Production of the filamentous hemagglutinin and the dermonecrotic toxin was not required to cause a lethal infection. Nine other mutants in vir-regulated genes whose phenotypes have yet to be determined were also tested. Only two of these mutants were impaired in the ability to cause a lethal infection. Expression of fimbriae does not appear to affect the dose required to cause a lethal infection; however, fimbrial expression was correlated with the later stages of a nonlethal, persistent infection. Growth of the bacteria in MgSO4, a condition which reversibly suppresses expression of the genes required for virulence, did not alter the ability of the bacteria to cause a lethal infection. Auxotrophic mutants deficient in leucine biosynthesis were as virulent as the parental strain; however, mutants deficient in methionine biosynthesis were less virulent. A B. parapertussis strain was much less effective in promoting a lethal infection than any of the wild-type B. pertussis strains examined. A persistent infection in the lungs was observed for weeks after challenge for mice given a sublethal dose of B. pertussis, and transmission from infected infants to the mother was never observed. PMID:2572561

  3. Film Screening and Conversation

    E-print Network

    Mathis, Wayne N.

    Film Screening and Conversation 2011 6-9pm Smithsonian Asian Paci c American Program Rasmuson Director John Sayles Film Run Time: 124 minutes Closest Metro: L'Enfant Plaza Related Traveling Exhibition, the Smithsonian Asian Pacific American Program presents a screening of the film Amigo and a conversation

  4. Screening for Skin Cancer

    Microsoft Academic Search

    Mark Helfand; Susan M. Mahon; Karen B. Eden; Paul S. Frame; C. Tracy Orleans

    We searched the MEDLINE database for papers published between 1994 and June 1999, using search terms for screening, physical examination, morbidity, and skin neoplasms. For information on accuracy of screening tests, we used the search terms sensitivity and specificity. We identified the most important studies from before 1994 from the Guide to Clinical Preventive Services, second edition, and from high-quality

  5. DCCPS: BRP: PCRB: Screening

    Cancer.gov

    PCRB is interested in research that promotes optimal use of breast, cervical, and colorectal cancer screenings in populations. Optimal use balances the benefit of screening for people who will get cancer in their lifetime against the harms to those who will not get cancer.

  6. Newborn screening fact sheets.

    PubMed

    Kaye, Celia I; Accurso, Frank; La Franchi, Stephen; Lane, Peter A; Hope, Northrup; Sonya, Pang; G Bradley, Schaefer; Michele A, Lloyd-Puryear

    2006-09-01

    Newborn screening fact sheets were last revised in 1996 by the American Academy of Pediatrics Committee on Genetics. This revision was prompted by advances in the field since 1996, including technologic innovations, as well as greater appreciation of ethical issues such as those surrounding informed consent. The following disorders are discussed in this revision of the newborn screening fact sheets: biotinidase deficiency, congenital adrenal hyperplasia, congenital hearing loss, congenital hypothyroidism, cystic fibrosis, galactosemia, homocystinuria, maple syrup urine disease, medium-chain acyl-coenzyme A dehydrogenase deficiency, phenylketonuria, sickle cell disease and other hemoglobinopathies, and tyrosinemia. A series of topics related to newborn screening is discussed in a companion publication to this electronic publication of the fact sheets (available at: www.pediatrics.org/cgi/content/full/118/3/1304). These topics are newborn screening as a public health system; factors contributing to the need for review of the newborn screening system; informed consent; tandem mass spectrometry; DNA analysis in newborn screening; status of newborn screening in the United States; and the effect of sample timing, preterm birth, diet, transfusion, and total parenteral nutrition on newborn screening results. PMID:16950973

  7. A Nutritional Conditional Lethal Mutant Due to Pyridoxine 5?-Phosphate Oxidase Deficiency in Drosophila melanogaster

    PubMed Central

    Chi, Wanhao; Zhang, Li; Du, Wei; Zhuang, Xiaoxi

    2014-01-01

    The concept of auxotrophic complementation has been proposed as an approach to identify genes in essential metabolic pathways in Drosophila melanogaster. However, it has achieved limited success to date, possibly due to the low probability of finding mutations fit with the chemically defined profile. Instead of using the chemically defined culture media lacking specific nutrients, we used bare minimum culture medium, i.e., 4% sucrose, for adult Drosophila. We identified a nutritional conditional lethal mutant and localized a c.95C > A mutation in the Drosophila pyridoxine 5?-phosphate oxidase gene [dPNPO or sugarlethal (sgll)] using meiotic recombination mapping, deficiency mapping, and whole genome sequencing. PNPO converts dietary vitamin B6 such as pyridoxine to its active form pyridoxal 5?-phosphate (PLP). The missense mutation (sgll95) results in the substitution of alanine to aspartate (p.Ala32Asp). The sgll95 flies survive well on complete medium but all die within 6 d on 4% sucrose only diet, which can be rescued by pyridoxine or PLP supplement, suggesting that the mutation does not cause the complete loss of PNPO activity. The sgll knockdown further confirms its function as the Drosophila PNPO. Because better tools for positional cloning and cheaper whole genome sequencing have made the identification of point mutations much easier than before, alleviating the necessity to pinpoint specific metabolic pathways before gene identification, we propose that nutritional conditional screens based on bare minimum growth media like ours represent promising approaches for discovering important genes and mutations in metabolic pathways, thereby accelerating the establishment of in vivo models that recapitulate human metabolic diseases. PMID:24739647

  8. Spinal Cord Compression Revealing an Intraosseous Schwannoma

    PubMed Central

    Metoui, Leila; Ajili, Faïda; Maiza, Mouna; Ben Ammar, Mehdi; Gharsallah, Imen; M'sakni, Issam; Louzir, Bassem; Othmani, Salah

    2013-01-01

    A 68-year-old female presented with inflammatory lumbalgia and cruralgia. Physical examination revealed a lumbar stiffness without neurological deficit. Secondarily, paraplegia and urinary retention appeared. Magnetic resonance imaging showed a vertebral compaction of L3 vertebra with medullar compression. Emergent surgery revealed an epidural tumor involving largely the L3 vertebral body. Histology found schwannoma with positive protein S100 on the immunohistochemical study. Metastasis screening revealed bilateral nodular lesions of the lungs and a trochanter high scintigraphic signal. It was a malignant schwannoma. The patient underwent radiotherapy in addition to the total tumor resection. PMID:24381595

  9. Screening for melanoma.

    PubMed

    Collins, Mary-Katharine M; Secrest, Aaron M; Ferris, Laura K

    2014-10-01

    Although melanoma is a deadly cancer that is rising in incidence, the USA does not have uniform guidelines for melanoma screening. Screening for melanoma requires no specialized equipment and has little associated morbidity. However, screening has the greatest impact when performed among patients with the highest risk for melanoma incidence and mortality. Screening lower-risk patients may result in prohibitively high costs, unnecessary biopsies of benign lesions, and decreased access to a dermatologic specialist for patients who are actually at a higher risk. We advocate targeting melanoma screening efforts toward those patients at high risk of developing and dying from melanoma, as well as toward those at-risk patients who are least likely to detect their own melanoma. PMID:24999755

  10. Colorectal cancer screening.

    PubMed

    Leggett, B A; Hewett, D G

    2015-01-01

    Colorectal cancer is one of the most common malignancies in Australia, and screening to detect it an earlier stage is cost-effective. Furthermore, detection and removal of precursor polyps can reduce incidence. Currently, there are limited data to determine the screening rate in Australia, but it is certainly lower than the 80% screening rate considered desirable. Whether colonoscopy is used as the screening test or to follow up positive results of an initial non-invasive test, it plays a fundamental role. Despite high sensitivity and specificity, it is expensive and invasive with measurable risk and is not acceptable as an initial test to many participants. It does not provide complete protection, and interval cancers between planned colonoscopies are associated with proximal location, origin in sessile serrated adenomas and operator-dependent factors. An essential component of colorectal screening is the measurement of colonoscopy quality indicators, such as caecal intubation and adenoma detection rates, which are known to be associated with the rate of interval cancer. The non-invasive screening test currently recommended in Australia is biennial testing for faecal occult blood between the ages of 50 and 75 using a faecal immunochemical test, with positives evaluated by colonoscopy. This is provided through the National Bowel Cancer Screening Programme, currently for those at the ages of 50, 55, 60 and 65 years, with full implementation of biennial screening by 2020. To improve screening in Australia, the most fruitful approach may be to acknowledge that there is a choice of screening tests and to focus on the goal of improving overall participation rate and being able to measure this. PMID:25582937

  11. Lethal action of the nitrothiazolyl-salicylamide derivative nitazoxanide via induction of oxidative stress in Leishmania (L.) infantum.

    PubMed

    Mesquita, Juliana Tonini; Pinto, Erika Gracielle; Taniwaki, Noemi Nosomi; Galisteo, Andres Jimenez; Tempone, Andre Gustavo

    2013-12-01

    Studying the cellular death pathways in Leishmania is an important aspect of discovering new antileishmanials. While using a drug repositioning approach, the lethal action of the nitrothiazolyl-salicylamide derivative nitazoxanide (NTZ) was investigated against Leishmania (L.) infantum. The in vitro antileishmanial activity and cytotoxicity were assessed using both parasite stages and mammalian NCTC cells, respectively. The lethal action of NTZ was investigated by detecting the phosphatidylserine (PS) exposure, reactive oxygen species (ROS) regulation, plasma membrane permeability, mitochondrial membrane potential and ultrastructural modifications by transmission electron microscopy. NTZ's activity against L. infantum was confirmed, producing IC50 values of 42.71?g/mL against promastigotes and 6.78?g/mL against intracellular amastigotes. NTZ rapidly altered the cellular metabolism of promastigotes by depolarising the mitochondrial membrane and up-regulating the reactive oxygen species (ROS). In addition, the flow cytometry data revealed an intense and time-dependent exposure of PS in promastigotes. When using SYTOX(®) Green as a fluorescent probe, NTZ demonstrated no interference in plasma membrane permeability. The ultrastructural alterations in promastigotes were time-dependent and caused chromatin condensation, plasma membrane blebbing and mitochondrial swelling. These data suggest that NTZ induced oxidative stress in L. (L.) infantum and might be a useful compound for investigating new therapeutic targets. PMID:24071379

  12. Hyperactivation of Alk induces neonatal lethality in knock-in AlkF1178L mice

    PubMed Central

    Lopez-Delisle, Lucille; Pierre-Eugène, Cécile; Bloch-Gallego, Evelyne; Birling, Marie-Christine; Duband, Jean-Loup; Durand, Estelle; Bourgeois, Thomas; Matrot, Boris; Sorg, Tania; Huerre, Michel; Meziane, Hamid; Roux, Michel J.; Champy, Marie-France; Gallego, Jorge; Delattre, Olivier; Janoueix-Lerosey, Isabelle

    2014-01-01

    The ALK (Anaplastic Lymphoma Kinase) gene encodes a tyrosine kinase receptor preferentially expressed in the central and peripheral nervous systems. A syndromic presentation associating congenital neuroblastoma with severe encephalopathy and an abnormal shape of the brainstem has been described in patients harbouring de novo germline F1174V and F1245V ALK mutations. Here, we investigated the phenotype of knock-in (KI) mice bearing the AlkF1178L mutation (F1174L in human). Although heterozygous KI mice did not reproduce the severe breathing and feeding difficulties observed in human patients, behavioral tests documented a reduced activity during dark phases and an increased anxiety of mutated mice. Matings of heterozygotes yielded the expected proportions of wild-type, heterozygotes and homozygotes at birth but a high neonatal lethality was noticed for homozygotes. We documented Alk expression in several motor nuclei of the brainstem involved in the control of sucking and swallowing. Evaluation of basic physiological functions 12 hours after birth revealed slightly more apneas but a dramatic reduced milk intake for homozygotes compared to control littermates. Overall, our data demonstrate that Alk activation above a critical threshold is not compatible with survival in mice, in agreement with the extremely severe phenotype of patients carrying aggressive de novo ALK germline mutations. PMID:24811761

  13. Peri-Implantation Lethality in Mice Lacking the Sm Motif-Containing Protein Lsm4

    PubMed Central

    Hirsch, Emilio; Oohashi, Toshitaka; Ahmad, Marianne; Stamm, Stefan; Fässler, Reinhard

    2000-01-01

    Small nuclear ribonucleoproteins (snRNPs) are particles present only in eukaryotic cells. They are involved in a large variety of RNA maturation processes, most notably in pre-mRNA splicing. Several of the proteins typically found in snRNPs contain a sequence signature, the Sm domain, conserved from yeast to mammals. By using a promoter trap strategy to target actively transcribed loci in murine embryonic stem cells, a new murine gene encoding an Sm motif-containing protein was identified. Database searches revealed that it is the mouse orthologue of Lsm4p, a protein found in yeast and human cells and putatively associated with U6 snRNA. Introduction of the geo reporter gene cassette under the control of the murine Lsm4 (mLsm4) endogenous promoter showed that the gene was ubiquitously transcribed in embryonic and adult tissues. The insertion of the geo cassette disrupted the mLsm4 allele, and homozygosity for the mutation led to a recessive embryonic lethal phenotype. mLsm4-null zygotes survived to the blastocyst stages, implanted into the uterus, but died shortly thereafter. The early death of mLsm4p-null mice suggests that the role of mLsm4p in splicing is essential and cannot be compensated by other Lsm proteins. PMID:10629062

  14. A LigA Three-Domain Region Protects Hamsters from Lethal Infection by Leptospira interrogans

    PubMed Central

    Coutinho, Mariana L.; Choy, Henry A.; Kelley, Melissa M.; Matsunaga, James; Babbitt, Jane T.; Lewis, Michael S.; Aleixo, Jose Antonio G.; Haake, David A.

    2011-01-01

    The leptospiral LigA protein consists of 13 bacterial immunoglobulin-like (Big) domains and is the only purified recombinant subunit vaccine that has been demonstrated to protect against lethal challenge by a clinical isolate of Leptospira interrogans in the hamster model of leptospirosis. We determined the minimum number and location of LigA domains required for immunoprotection. Immunization with domains 11 and 12 was found to be required but insufficient for protection. Inclusion of a third domain, either 10 or 13, was required for 100% survival after intraperitoneal challenge with Leptospira interrogans serovar Copenhageni strain Fiocruz L1-130. As in previous studies, survivors had renal colonization; here, we quantitated the leptospiral burden by qPCR to be 1.2×103 to 8×105 copies of leptospiral DNA per microgram of kidney DNA. Although renal histopathology in survivors revealed tubulointerstitial changes indicating an inflammatory response to the infection, blood chemistry analysis indicated that renal function was normal. These studies define the Big domains of LigA that account for its vaccine efficacy and highlight the need for additional strategies to achieve sterilizing immunity to protect the mammalian host from leptospiral infection and its consequences. PMID:22180800

  15. Determination of Median Lethal Dose of Combination of Endosulfan and Cypermethrin in Wistar Rat

    PubMed Central

    Raj, Jaya; Chandra, Mohineesh; Dogra, Tirath D.; Pahuja, Monika; Raina, Anupuma

    2013-01-01

    The present study was designed to determine the lethal dose 50 (LD50) of combination of cypermethrin, a pyrethroid, and endosulfan, an organochlorine compound in Wistar rats. LD50 is the amount (dose) of a chemical, calculated as per the concentration of chemicals that produces death in 50% of a population of test animals to which it is administered by any of a variety of methods. A single oral dose of combination of cypermethrin and endosulfan were dissolved in dimethyl sulfoxide (DMSO) in a ratio of 1:1 and administered orally at the concentration of 165 mg/kg body weight (b.w), 330 mg/kg b.w, 660 mg/kg b.w, and 1320 mg/kg b.w to experimental animals. LD50 was calculated according to the method described by Miller and Tainter (1994) and was observed as 691.83 mg/kg b.w for this combination. Single dose of test article at 165 mg/kg b.w did not reveal any toxic signs or behavioral alterations, hence considered as No observed Adverse Effect level (NOAEL). PMID:23833430

  16. Purification and partial characterization of stonustoxin (lethal factor) from Synanceja horrida venom.

    PubMed

    Poh, C H; Yuen, R; Khoo, H E; Chung, M; Gwee, M; Gopalakrishnakone, P

    1991-01-01

    1. The lethal factor of the stonefish (Synanceja horrida) venom, designated as the stonustoxin, was purified to homogeneity by a two-step procedure on Sephacryl S-200 High Resolution (HR) gel permeation and DEAE Bio-Gel A anion exchange chromatography. 2. Stonustoxin has a native mol. wt of 148,000 and an isoelectric point of 6.9. 3. SDS-polyacrylamide gel electrophoresis revealed two subunits (designated alpha and beta) with mol. wts of 71,000 and 79,000, respectively. 4. The amino acid composition of both subunits and the N-terminal amino acid sequence of the beta subunit were also determined. 5. Purified stonustoxin had an LD50 of 0.017 microgram/g which is 22-fold more potent than that of the crude venom. 6. The toxin exhibited potent haemolytic activity in vitro and edema-inducing activity with a minimum edema dose (MED) of 0.15 micrograms in mouse paw. The edema effect was not antagonized by diphenhydramine. PMID:1790672

  17. A History of Vision Screening.

    ERIC Educational Resources Information Center

    Appelboom, Tina M.

    1985-01-01

    The vision screening program has a long and interesting history involving educators, pediatricians, optometrists, and ophthamologists. This historical review of vision screening in the schools includes a discussion of amblyopia and screening of preschool students. (Author/CB)

  18. Temperature-Sensitive Lethal Mutations on Yeast Chromosome I Appear to Define Only a Small Number of Genes

    PubMed Central

    Kaback, David B.; Oeller, Paul W.; Steensma, H. Yde; Hirschman, Janet; Ruezinsky, Diane; Coleman, Kevin G.; Pringle, John R.

    1984-01-01

    A method was developed for isolating large numbers of mutations on chromosome I of the yeast Saccharomyces cerevisiae. A strain monosomic for chromosome I (i.e., haploid for chromosome I and diploid for all other chromosomes) was mutagenized with either ethyl methanesulfonate or N-methyl-N'-nitro-N -nitrosoguanidine and screened for temperature-sensitive (Ts- ) mutants capable of growth on rich, glucose-containing medium at 25° but not at 37°. Recessive mutations induced on chromosome I are expressed, whereas those on the diploid chromosomes are usually not expressed because of the presence of wild-type alleles on the homologous chromosomes. Dominant ts mutations on all chromosomes should also be expressed, but these appeared rarely. — Of the 41 ts mutations analyzed, 32 mapped on chromosome I. These 32 mutations fell into only three complementation groups, which proved to be the previously described genes CDC15, CDC24 and PYK1 (or CDC19). We recovered 16 or 17 independent mutations in CDC15, 12 independent mutations in CDC24 and three independent mutations in PYK1. A fourth gene on chromosome I, MAK16, is known to be capable of giving rise to a ts-lethal allele, but we recovered no mutations in this gene. The remaining nine mutations isolated using the monosomic strain appeared not to map on chromosome I and were apparently expressed in the original mutants because they had become homozygous or hemizygous by mitotic recombination or chromosome loss. — The available information about the size of chromosome I suggests that it should contain approximately 60–100 genes. However, our isolation in the monosomic strain of multiple, independent alleles of just three genes suggests that only a small proportion of the genes on chromosome I is easily mutable to give a Ts--lethal phenotype. — During these studies, we located CDC24 on chromosome I and determined that it is centromere distal to PYK1 on the left arm of the chromosome. PMID:6383953

  19. Pegfilgrastim Improves Survival of Lethally Irradiated Nonhuman Primates.

    PubMed

    Hankey, Kim G; Farese, Ann M; Blaauw, Erica C; Gibbs, Allison M; Smith, Cassandra P; Katz, Barry P; Tong, Yan; Prado, Karl L; MacVittie, Thomas J

    2015-06-01

    Leukocyte growth factors (LGF), such as filgrastim, pegfilgrastim and sargramostim, have been used to mitigate the hematologic symptoms of acute radiation syndrome (ARS) after radiation accidents. Although these pharmaceuticals are currently approved for treatment of chemotherapy-induced myelosuppression, such approval has not been granted for myelosuppression resulting from acute radiation exposure. Regulatory approval of drugs used to treat radiological or nuclear exposure injuries requires their development and testing in accordance with the Animal Efficacy Rule, set forth by the U.S. Food and Drug Administration. To date, filgrastim is the only LGF that has undergone efficacy assessment conducted under the Animal Efficacy Rule. To confirm the efficacy of another LGF with a shorter dosing regimen compared to filgrastim, we evaluated the use of pegfilgrastim (Neulasta®) in a lethal nonhuman primate (NHP) model of hematopoietic acute radiation syndrome (H-ARS). Rhesus macaques were exposed to 7.50 Gy total-body irradiation (the LD50/60), delivered at 0.80 Gy/min using linear accelerator 6 MV photons. Pegfilgrastim (300 ?g/kg, n = 23) or 5% dextrose in water (n = 23) was administered on day 1 and 8 postirradiation and all animals received medical management. Hematologic and physiologic parameters were evaluated for 60 days postirradiation. The primary, clinically relevant end point was survival to day 60; secondary end points included hematologic-related parameters. Pegfilgrastim significantly (P = 0.0014) increased 60 day survival to 91.3% (21/23) from 47.8% (11/23) in the control. Relative to the controls, pegfilgrastim also significantly: 1. decreased the median duration of neutropenia and thrombocytopenia; 2. improved the median time to recovery of absolute neutrophil count (ANC) ?500/?L, ANC ?1,000/?L and platelet (PLT) count ?20,000/?L; 3. increased the mean ANC at nadir; and 4. decreased the incidence of Gram-negative bacteremia. These data demonstrate that pegfilgrastim is an additional medical countermeasure capable of increasing survival and neutrophil-related parameters when administered in an abbreviated schedule to a NHP model of lethal H-ARS. PMID:26035709

  20. Breast cancer screening.

    PubMed

    Sirovich, B E; Sox, H C

    1999-10-01

    Randomized controlled trials involving nearly 500,000 women on two continents have confirmed the early promise that screening mammography can reduce breast cancer mortality. The observed benefits of mammographic screening, however, are not the same in all women. The mortality reduction in women over age 70 is unknown, and women aged 40 to 49 do not appear to benefit from mammographic screening to the same extent as those over age 50. The reasons for this disparity are incompletely understood, but it depends in part upon differing tumor biology and mammographic test characteristics in younger women. Even if relative survival benefits were equal for women under and over age 50, absolute reduction in risk would remain considerably lower for younger women, a disparity that would not be corrected by improved screening technology or adjustment of interscreening intervals. The authors' review of the evidence leads them to strongly support mammographic screening of women aged 50 to 69 at an interval not longer than 2 years. The authors also feel it is reasonable to screen women over age 70 who have a favorable life expectancy. They conclude, however, that the evidence does not support a blanket recommendation in favor of screening women aged 40 to 49. Instead, they advocate a well-informed conversation between physician and patient regarding the present knowledge and the risks and benefits of screening for each individual woman. Definitive answers await the results of ongoing RCTs designed to study the survival benefit conferred by screening women aged 40 to 49. Disagreement will undoubtedly persist regarding which recommendations should determine private practice and public policy. PMID:10572546

  1. Retrocyclins Kill Bacilli and Germinating Spores of Bacillus anthracis and Inactivate Anthrax Lethal Toxin*

    PubMed Central

    Wang, Wei; Mulakala, Chandrika; Ward, Sabrina C.; Jung, Grace; Luong, Hai; Pham, Duy; Waring, Alan J.; Kaznessis, Yiannis; Lu, Wuyuan; Bradley, Kenneth A.; Lehrer, Robert I.

    2008-01-01

    ?-defensins are cyclic octadecapeptides encoded by the modified ?-defensin genes of certain nonhuman primates. The recent demonstration that human ?-defensins could prevent deleterious effects of anthrax lethal toxin in vitro and in vivo led us to examine the effects of ?-defensins on Bacillus anthracis (Sterne). We tested rhesus ?-defensins 1–3, retrocyclins 1–3, and several analogues of RC-1. Low concentrations of ?-defensins not only killed vegetative cells of B. anthracis (Sterne) and rendered their germinating spores nonviable, they also inactivated the enzymatic activity of anthrax lethal factor and protected murine RAW-264.7 cells from lethal toxin, a mixture of lethal factor and protective antigen. Structure-function studies indicated that the cyclic backbone, intramolecular tri-disulfide ladder, and arginine residues of ?-defensins contributed substantially to these protective effects. Surface plasmon resonance studies showed that retrocyclins bound the lethal factor rapidly and with high affinity. Retrocyclin-mediated inhibition of the enzymatic activity of lethal factor increased substantially if the enzyme and peptide were preincubated before substrate was added. The temporal discrepancy between the rapidity of binding and the slowly progressive extent of lethal factor inhibition suggest that post-binding events, perhaps in situ oligomerization, contribute to the antitoxic properties of retrocyclins. Overall, these findings suggest that ?-defensins provide molecular templates that could be used to create novel agents effective against B. anthracis and its toxins. PMID:16790431

  2. Retrocyclins kill bacilli and germinating spores of Bacillus anthracis and inactivate anthrax lethal toxin.

    PubMed

    Wang, Wei; Mulakala, Chandrika; Ward, Sabrina C; Jung, Grace; Luong, Hai; Pham, Duy; Waring, Alan J; Kaznessis, Yiannis; Lu, Wuyuan; Bradley, Kenneth A; Lehrer, Robert I

    2006-10-27

    Theta-defensins are cyclic octadecapeptides encoded by the modified alpha-defensin genes of certain nonhuman primates. The recent demonstration that human alpha-defensins could prevent deleterious effects of anthrax lethal toxin in vitro and in vivo led us to examine the effects of theta-defensins on Bacillus anthracis (Sterne). We tested rhesus theta-defensins 1-3, retrocyclins 1-3, and several analogues of RC-1. Low concentrations of theta-defensins not only killed vegetative cells of B. anthracis (Sterne) and rendered their germinating spores nonviable, they also inactivated the enzymatic activity of anthrax lethal factor and protected murine RAW-264.7 cells from lethal toxin, a mixture of lethal factor and protective antigen. Structure-function studies indicated that the cyclic backbone, intramolecular tri-disulfide ladder, and arginine residues of theta-defensins contributed substantially to these protective effects. Surface plasmon resonance studies showed that retrocyclins bound the lethal factor rapidly and with high affinity. Retrocyclin-mediated inhibition of the enzymatic activity of lethal factor increased substantially if the enzyme and peptide were preincubated before substrate was added. The temporal discrepancy between the rapidity of binding and the slowly progressive extent of lethal factor inhibition suggest that post-binding events, perhaps in situ oligomerization, contribute to the antitoxic properties of retrocyclins. Overall, these findings suggest that theta-defensins provide molecular templates that could be used to create novel agents effective against B. anthracis and its toxins. PMID:16790431

  3. Screening for psychosocial distress: a national survey of oncology social workers.

    PubMed

    BrintzenhofeSzoc, Karlynn; Davis, Cindy; Kayser, Karen; Lee, Hee Yun; Nedjat-Haiem, Frances; Oktay, Julianne S; Zabora, James; Zebrack, Bradley J

    2015-01-01

    Oncology social workers are the primary providers of psychosocial care for cancer patients, thus they are well positioned to implement and oversee psychosocial distress screening. A national survey of members of the Association of Oncology Social Work was conducted to examine screening knowledge, attitudes, behaviors, and perceived competency. The findings indicated that most participants screened for psychosocial distress using a standardized instrument and identified institutional and individual barriers to implementing screening. Analyses revealed that social workers who perceived patient benefits from screening and were knowledgeable about guidelines were significantly more likely to implement screening procedures and use a standardized instrument. PMID:25420574

  4. Proteinuria and Perinatal Lethality in Mice Lacking NEPH1, a Novel Protein with Homology to NEPHRIN

    PubMed Central

    Donoviel, Dorit B.; Freed, Deon D.; Vogel, Hannes; Potter, David G.; Hawkins, Edith; Barrish, James P.; Mathur, Brian N.; Turner, C. Alexander; Geske, Robert; Montgomery, Charles A.; Starbuck, Michael; Brandt, Mary; Gupta, Anupma; Ramirez-Solis, Ramiro; Zambrowicz, Brian P.; Powell, David R.

    2001-01-01

    A high-throughput, retrovirus-mediated mutagenesis method based on gene trapping in embryonic stem cells was used to identify a novel mouse gene. The human ortholog encodes a transmembrane protein containing five extracellular immunoglobulin-like domains that is structurally related to human NEPHRIN, a protein associated with congenital nephrotic syndrome. Northern analysis revealed wide expression in humans and mice, with highest expression in kidney. Based on similarity to NEPHRIN and abundant expression in kidney, this protein was designated NEPH1 and embryonic stem cells containing the retroviral insertion in the Neph1 locus were used to generate mutant mice. Analysis of kidney RNA from Neph1?/? mice showed that the retroviral insertion disrupted expression of Neph1 transcripts. Neph1?/? pups were represented at the expected normal Mendelian ratios at 1 to 3 days of age but at only 10% of the expected frequency at 10 to 12 days after birth, suggesting an early postnatal lethality. The Neph1?/? animals that survived beyond the first week of life were sickly and small but without edema, and all died between 3 and 8 weeks of age. Proteinuria ranging from 300 to 2,000 mg/dl was present in all Neph1?/? mice. Electron microscopy demonstrated NEPH1 expression in glomerular podocytes and revealed effacement of podocyte foot processes in Neph1?/? mice. These findings suggest that NEPH1, like NEPHRIN, may play an important role in maintaining the structure of the filtration barrier that prevents proteins from freely entering the glomerular urinary space. PMID:11416156

  5. Anthrax Lethal Toxin Induces Acute Diastolic Dysfunction in Rats Through Disruption of the Phospholamban Signaling Network

    PubMed Central

    Golden, Honey B.; Watson, Linley E.; Nizamutdinov, Damir; Feng, Hao; Gerilechaogetu, Fnu; Lal, Hind; Verma, Suresh K.; Mukhopadhyay, Swagoto; Foster, Donald M.; Dillmann, Wolfgang H.; Dostal, D.E.

    2013-01-01

    Background Anthrax lethal toxin (LT), secreted by Bacillus anthracis, causes severe cardiac dysfunction by unknown mechanisms. LT specifically cleaves the docking domains of MAPKK (MEKs); thus, we hypothesized that LT directly impairs cardiac function through dysregulation of MAPK signaling mechanisms. Methods and Results In a time-course study of LT toxicity, echocardiography revealed acute diastolic heart failure accompanied by pulmonary regurgitation and left atrial dilation in adult Sprague-Dawley rats at time points corresponding to dysregulated JNK, phospholamban (PLB) and protein phosphatase 2A (PP2A) myocardial signaling. Using isolated rat ventricular myocytes, we identified the MEK7-JNK1-PP2A-PLB signaling axis to be important for regulation of intracellular calcium (Ca2+i) handling, PP2A activation and targeting of PP2A-B56? to Ca2+i handling proteins, such as PLB. Through a combination of gain-of-function and loss-of-function studies, we demonstrated that over-expression of MEK7 protects against LT-induced PP2A activation and Ca2+i dysregulation through activation of JNK1. Moreover, targeted phosphorylation of PLB-Thr17 by Akt improved sarcoplasmic reticulum Ca2+i release and reuptake during LT toxicity. Co-immunoprecipitation experiments further revealed the pivotal role of MEK7-JNK-Akt complex formation for phosphorylation of PLB-Thr17 during acute LT toxicity. Conclusions Our findings support a cardiogenic mechanism of LT-induced diastolic dysfunction, by which LT disrupts JNK1 signaling and results in Ca2+i dysregulation through diminished phosphorylation of PLB by Akt and increased dephosphorylation of PLB by PP2A. Integration of the MEK7-JNK1 signaling module with Akt represents an important stress-activated signalosome that may confer protection to sustain cardiac contractility and maintain normal levels of Ca2+i through PLB-T17 phosphorylation. PMID:23907041

  6. Interferon alfacon-1 protects hamsters from lethal pichinde virus infection.

    PubMed

    Gowen, Brian B; Barnard, Dale L; Smee, Donald F; Wong, Min-Hui; Pace, Anne M; Jung, Kie-Hoon; Winslow, Scott G; Bailey, Kevin W; Blatt, Lawrence M; Sidwell, Robert W

    2005-06-01

    Hemorrhagic fever of arenaviral origin is a frequently fatal infectious disease of considerable priority to the biodefense mission. Historically, the treatment of arenaviral infections with alpha interferons has not yielded favorable results. Here we present evidence that interferon alfacon-1, a nonnaturally occurring bioengineered alpha interferon approved for the treatment of chronic hepatitis C, is active against Pichinde and Tacaribe arenaviruses in cell culture. In the hamster model of Pichinde virus (PCV) infection, interferon alfacon-1 treatment significantly protected animals from death, prolonged the survival of those that eventually died, reduced virus titers, and limited liver damage characteristic of PCV-induced disease. Moreover, interferon alfacon-1 also demonstrated therapeutic activity, to a lesser degree, when the initiation of treatment was delayed up to 2 days post-virus challenge. Despite the observed advantages of interferon alfacon-1 therapy, efforts to stimulate the immune system with the known interferon inducer poly(I:C12U) (Ampligen) offered only limited protection against lethal PCV challenge. Taken together, these data suggest that the increased potency of the bio-optimized interferon alfacon-1 molecule may be critical to the observed antiviral effects. These data are the first report demonstrating efficacious treatment of acute arenaviral disease with alpha interferon therapy, and further study is warranted. PMID:15917537

  7. Sticky foam as a less-than-lethal technology

    SciTech Connect

    Scott, S.H.

    1996-12-31

    Sandia National Labs (SNL) in 1994 completed a project funded by the National Institute of Justice (NIJ) to determine the applicability of sticky foam for correctional applications. Sticky foam is an extremely tacky, tenacious material used to block, entangle, and impair individuals. The NIJ project developed a gun capable of firing multiple shots of sticky foam, tested the gun and sticky foam effectiveness on SNL volunteers acting out prison and law enforcement scenarios, and had the gun and sticky foam evaluated by correctional representatives. Based on the NIJ project work, SNL supported the Marine Corps Mission, Operation United Shield, with sticky foam guns and supporting equipment to assist in the withdrawal of UN Peacekeepers from Somalia. Prior to the loan of the equipment, the Marines were given training in sticky foam characterization, toxicology, safety issues, cleanup and waste disposal, use limitations, use protocol and precautions, emergency facial clean-up, skin cleanup, gun filling, targeting and firing, and gun cleaning. The Marine Corps successfully used the sticky foam guns as part of that operation. This paper describes these recent developments of sticky foam for non-lethal uses and some of the lessons learned from scenario and application testing.

  8. Hemolysis-induced lethality involves inflammasome activation by heme.

    PubMed

    Dutra, Fabianno F; Alves, Letícia S; Rodrigues, Danielle; Fernandez, Patricia L; de Oliveira, Rosane B; Golenbock, Douglas T; Zamboni, Dario S; Bozza, Marcelo T

    2014-09-30

    The increase of extracellular heme is a hallmark of hemolysis or extensive cell damage. Heme has prooxidant, cytotoxic, and inflammatory effects, playing a central role in the pathogenesis of malaria, sepsis, and sickle cell disease. However, the mechanisms by which heme is sensed by innate immune cells contributing to these diseases are not fully characterized. We found that heme, but not porphyrins without iron, activated LPS-primed macrophages promoting the processing of IL-1? dependent on nucleotide-binding domain and leucine rich repeat containing family, pyrin domain containing 3 (NLRP3). The activation of NLRP3 by heme required spleen tyrosine kinase, NADPH oxidase-2, mitochondrial reactive oxygen species, and K(+) efflux, whereas it was independent of heme internalization, lysosomal damage, ATP release, the purinergic receptor P2X7, and cell death. Importantly, our results indicated the participation of macrophages, NLRP3 inflammasome components, and IL-1R in the lethality caused by sterile hemolysis. Thus, understanding the molecular pathways affected by heme in innate immune cells might prove useful to identify new therapeutic targets for diseases that have heme release. PMID:25225402

  9. Precursors of lethal violence: a death row sample.

    PubMed

    Freedman, D; Hemenway, D

    2000-06-01

    A qualitative methodology based on the standards of criminal defense investigation was used to analyze the social and family histories of 16 men sentenced to death in California. Using a multisource cross-validation methodology, we assessed patterns of impairment, injury and deficit at each of four ecological levels: family, individual, community and social institutions. Investigation documented consistent and pervasive patterns of serious impairment, injury and deficit across the cases and levels. The men share numerous risk factors and few resiliency factors associated with violence. We found family violence in all 16 cases, including severe physical and/or sexual abuse in 14 cases; individual impairments in 16, including 14 with post-traumatic stress disorder, 13 with severe depression and 12 with histories of traumatic brain injury; community isolation and violence in 12; and institutional failure in 15, including 13 cases of severe physical and/or sexual abuse while in foster care or under state youth authority jurisdiction. Appropriate interventions might have made a difference in reducing lethal violence and its precursor conditions. PMID:10798330

  10. Novel repression of the glucocorticoid receptor by anthrax lethal toxin.

    PubMed

    Webster, Jeanette I; Moayeri, Mahtab; Sternberg, Esther M

    2004-06-01

    Death from anthrax has been reported to occur from systemic shock. The lethal toxin (LeTx) is the major effector of anthrax mortality. Although the mechanism of entry of this toxin into cells is well understood, its actions once inside the cell are not as well understood. LeTx is known to cleave and inactivate MAPKKs. We have recently shown that LeTx represses the glucocorticoid receptor (GR) both in vitro and in vivo. This repression is partial and specific, repressing the glucocorticoid, progesterone, and estrogen receptor alpha, but not the mineralocorticoid or estrogen receptor beta. This toxin does not affect GR ligand or DNA binding, and we have suggested that it may function by removing/inactivating one or more of the many cofactors involved in nuclear hormone receptor signaling. Although the precise involvement of this nuclear hormone receptor repression in LeTx toxicity is unknown, examples of blunted HPA axis and glucocorticoid signaling in numerous autoimmune/inflammatory diseases suggest that such repression of critically important receptors could have deleterious effects on health. PMID:15265771

  11. The anthrax lethal factor and its MAPK kinase-specific metalloprotease activity.

    PubMed

    Tonello, Fiorella; Montecucco, Cesare

    2009-12-01

    The anthrax lethal factor is a multi-domain protein toxin released by Bacillus anthracis which enters cells in a process mediated by the protective antigen and specific cell receptors. In the cytosol, the lethal factor cleaves the N-terminal tail of many MAPK kinases, thus deranging a major cell signaling pathway. The structural features at the basis of these activities of LF are reviewed here with particular attention to the proteolytic activity and to the identification of specific inhibitors. A significant similarity between the metalloprotease domain of the lethal factor and of that of the clostridial neurotoxins has been noted and is discussed. PMID:19665472

  12. Potentially-lethal damage and radioprotection in human cells exposed to californium-252

    SciTech Connect

    Schroy, C.B.; Goud, S.N.; Magura, C.; Feola, J.M.; Maruyama, Y.

    1986-01-01

    Cultured human T-1E cells were irradiated with californium-252 neutrons and gamma rays. When 2 mm caffeine was present in the medium for 47 h after irradiation cell survival (assayed by colony formation) was decreased significantly. When 2 m dimethylsulfoxide was present during the irradiations radioprotection was observed using the same assay. The caffeine data indicate that potentially-lethal lesions exist in cells after californium exposure and that these lesions can be made lethal when they would otherwise be repaired. The DMSO data indicate that radioprotection from californium exposure can be achieved and that scanvengable free radicals play an important role in Cf-252 lethality.

  13. A high-throughput screen identifies PARP1/2 inhibitors as a potential therapy for ERCC1-deficient non-small cell lung cancer.

    PubMed

    Postel-Vinay, S; Bajrami, I; Friboulet, L; Elliott, R; Fontebasso, Y; Dorvault, N; Olaussen, K A; André, F; Soria, J-C; Lord, C J; Ashworth, A

    2013-11-21

    Excision repair cross-complementation group 1 (ERCC1) is a DNA repair enzyme that is frequently defective in non-small cell lung cancer (NSCLC). Although low ERCC1 expression correlates with platinum sensitivity, the clinical effectiveness of platinum therapy is limited, highlighting the need for alternative treatment strategies. To discover new mechanism-based therapeutic strategies for ERCC1-defective tumours, we performed high-throughput drug screens in an isogenic NSCLC model of ERCC1 deficiency and dissected the mechanism underlying ERCC1-selective effects by studying molecular biomarkers of tumour cell response. The high-throughput screens identified multiple clinical poly (ADP-ribose) polymerase 1 and 2 (PARP1/2) inhibitors, such as olaparib (AZD-2281), niraparib (MK-4827) and BMN 673, as being selective for ERCC1 deficiency. We observed that ERCC1-deficient cells displayed a significant delay in double-strand break repair associated with a profound and prolonged G?/M arrest following PARP1/2 inhibitor treatment. Importantly, we found that ERCC1 isoform 202, which has recently been shown to mediate platinum sensitivity, also modulated PARP1/2 sensitivity. A PARP1/2 inhibitor-synthetic lethal siRNA screen revealed that ERCC1 deficiency was epistatic with homologous recombination deficiency. However, ERCC1-deficient cells did not display a defect in RAD51 foci formation, suggesting that ERCC1 might be required to process PARP1/2 inhibitor-induced DNA lesions before DNA strand invasion. PARP1 silencing restored PARP1/2 inhibitor resistance in ERCC1-deficient cells but had no effect in ERCC1-proficient cells, supporting the hypothesis that PARP1 might be required for the ERCC1 selectivity of PARP1/2 inhibitors. This study suggests that PARP1/2 inhibitors as a monotherapy could represent a novel therapeutic strategy for NSCLC patients with ERCC1-deficient tumours. PMID:23934192

  14. Colorectal Cancer Screening

    MedlinePLUS

    ... rectum small intestine colon (large intestine) What Is Colorectal Cancer? Colorectal cancer is cancer that occurs in the ... or older, start getting screened now. Who Gets Colorectal Cancer? • Both men and women can get it. • It ...

  15. Health Screenings at School

    MedlinePLUS

    ... ability to learn. In some states these screening tests are mandated by law and may also include dental checks, scoliosis evaluations, blood pressure readings, and height and weight measurements. In school districts in which ...

  16. Screening and Risk Factors

    Cancer.gov

    Close Window State Cancer Profiles Quick Reference Guides ? Quick Reference Guides Index Screening and Risk Factors Send to Printer Text description of this image. Site Home Policies Accessibility Viewing Files FOIA Contact Us U.S. Department of Health

  17. Screening for Prostate Cancer

    MedlinePLUS

    ... the patient’s preferences, prostate cancer risk, health, and life expectancy. Doctors should not screen for prostate cancer using ... than 69 years, or any man with a life expectancy less than 10 to 15 years. What are ...

  18. Stomach (Gastric) Cancer Screening

    MedlinePLUS

    ... available from the NCI Web site . There is no standard or routine screening test for stomach cancer. ... cancers would help you live longer than if no treatment were given, and treatments for cancer may ...

  19. Newborn screening tests

    MedlinePLUS

    ... about 40 disorders. All 50 states screen for congenital hypothyroidism , galactosemia , and phenylketonuria (PKU). In addition to the ... acid metabolism disorders Biotinidase deficiency Congenital adrenal ... fibrosis Fatty acid metabolism disorders Galactosemia Glucose- ...

  20. Endometrial Cancer Screening

    MedlinePLUS

    ... Follow-up tests may detect endometrial cancer. Transvaginal ultrasound No studies have shown that screening by transvaginal ... of the latest published information on cancer prevention, detection, genetics, treatment, supportive care, and complementary and alternative ...

  1. Screening for Cervical Cancer

    MedlinePLUS

    ... 17 PM You are here: Home Recommendations for Primary Care Practice Published Recommendations Recommendation Summary Cervical Cancer: Screening ... are one-page documents that provide guidance to primary care clinicians for using recommendations in practice. This summary ...

  2. A genomewide overexpression screen identifies genes involved in the phosphatidylinositol 3-kinase pathway in the human protozoan parasite Entamoeba histolytica.

    PubMed

    Koushik, Amrita B; Welter, Brenda H; Rock, Michelle L; Temesvari, Lesly A

    2014-03-01

    Entamoeba histolytica is a protozoan parasite that causes amoebic dysentery and liver abscess. E. histolytica relies on motility, phagocytosis, host cell adhesion, and proteolysis of extracellular matrix for virulence. In eukaryotic cells, these processes are mediated in part by phosphatidylinositol 3-kinase (PI3K) signaling. Thus, PI3K may be critical for virulence. We utilized a functional genomics approach to identify genes whose products may operate in the PI3K pathway in E. histolytica. We treated a population of trophozoites that were overexpressing genes from a cDNA library with a near-lethal dose of the PI3K inhibitor wortmannin. This screen was based on the rationale that survivors would be overexpressing gene products that directly or indirectly function in the PI3K pathway. We sequenced the overexpressed genes in survivors and identified a cDNA encoding a Rap GTPase, a protein previously shown to participate in the PI3K pathway. This supports the validity of our approach. Genes encoding a coactosin-like protein, EhCoactosin, and a serine-rich E. histolytica protein (SREHP) were also identified. Cells overexpressing EhCoactosin or SREHP were also less sensitive to a second PI3K inhibitor, LY294002. This corroborates the link between these proteins and PI3K. Finally, a mutant cell line with an increased level of phosphatidylinositol (3,4,5)-triphosphate, the product of PI3K activity, exhibited increased expression of SREHP and EhCoactosin. This further supports the functional connection between these proteins and PI3K in E. histolytica. To our knowledge, this is the first forward-genetics screen adapted to reveal genes participating in a signal transduction pathway in this pathogen. PMID:24442890

  3. A Genomewide Overexpression Screen Identifies Genes Involved in the Phosphatidylinositol 3-Kinase Pathway in the Human Protozoan Parasite Entamoeba histolytica

    PubMed Central

    Koushik, Amrita B.; Welter, Brenda H.; Rock, Michelle L.

    2014-01-01

    Entamoeba histolytica is a protozoan parasite that causes amoebic dysentery and liver abscess. E. histolytica relies on motility, phagocytosis, host cell adhesion, and proteolysis of extracellular matrix for virulence. In eukaryotic cells, these processes are mediated in part by phosphatidylinositol 3-kinase (PI3K) signaling. Thus, PI3K may be critical for virulence. We utilized a functional genomics approach to identify genes whose products may operate in the PI3K pathway in E. histolytica. We treated a population of trophozoites that were overexpressing genes from a cDNA library with a near-lethal dose of the PI3K inhibitor wortmannin. This screen was based on the rationale that survivors would be overexpressing gene products that directly or indirectly function in the PI3K pathway. We sequenced the overexpressed genes in survivors and identified a cDNA encoding a Rap GTPase, a protein previously shown to participate in the PI3K pathway. This supports the validity of our approach. Genes encoding a coactosin-like protein, EhCoactosin, and a serine-rich E. histolytica protein (SREHP) were also identified. Cells overexpressing EhCoactosin or SREHP were also less sensitive to a second PI3K inhibitor, LY294002. This corroborates the link between these proteins and PI3K. Finally, a mutant cell line with an increased level of phosphatidylinositol (3,4,5)-triphosphate, the product of PI3K activity, exhibited increased expression of SREHP and EhCoactosin. This further supports the functional connection between these proteins and PI3K in E. histolytica. To our knowledge, this is the first forward-genetics screen adapted to reveal genes participating in a signal transduction pathway in this pathogen. PMID:24442890

  4. Non-Lethal Heat Shock Increased Hsp70 and Immune Protein Transcripts but Not Vibrio Tolerance in the White-Leg Shrimp

    PubMed Central

    Loc, Nguyen Hong; MacRae, Thomas H.; Musa, Najiah; Bin Abdullah, Muhd Danish Daniel; Abdul Wahid, Mohd. Effendy; Sung, Yeong Yik

    2013-01-01

    Non-lethal heat shock boosts bacterial and viral disease tolerance in shrimp, possibly due to increases in endogenous heat shock protein 70 (Hsp70) and/or immune proteins. To further understand the mechanisms protecting shrimp against infection, Hsp70 and the mRNAs encoding the immune-related proteins prophenoloxidase (proPO), peroxinectin, penaeidin, crustin and hemocyanin were studied in post-larvae of the white-leg shrimp Litopenaeus vannamei, following a non-lethal heat shock. As indicated by RT-qPCR, a 30 min abrupt heat shock increased Hsp70 mRNA in comparison to non-heated animals. Immunoprobing of western blots and quantification by ELISA revealed that Hsp70 production after heat shock was correlated with enhanced Hsp70 mRNA. proPO and hemocyanin mRNA levels were augmented, whereas peroxinectin and crustin mRNA levels were unchanged following non-lethal heat shock. Penaeidin mRNA was decreased by all heat shock treatments. Thirty min abrupt heat shock failed to improve survival of post-larvae in a standardized challenge test with Vibrio harveyi, indicating that under the conditions of this study, L. vannamei tolerance to Vibrio infection was influenced neither by Hsp70 accumulation nor the changes in the immune-related proteins, observations dissimilar to other shrimp species examined. PMID:24039886

  5. Pre-exposure to yeast protects larvae of Galleria mellonella from a subsequent lethal infection by Candida albicans and is mediated by the increased expression of antimicrobial peptides.

    PubMed

    Bergin, David; Murphy, Lisa; Keenan, Joanne; Clynes, Martin; Kavanagh, Kevin

    2006-07-01

    Pre-exposure of the larvae of Galleria mellonella to Candida albicans or Saccharomyces cerevisiae protects against a subsequent infection with 10(6) C. albicans cells. This protection can also be induced by exposing larvae to glucan or laminarin prior to the administration of the potentially lethal inoculum. Analysis of the genes coding for galiomicin, a defensin in G. mellonella, a cysteine-rich antifungal peptide gallerimycin, an iron-binding protein transferrin and an inducible metalloproteinase inhibitor (IMPI) from G. mellonella demonstrated increased expression, which is at its highest after 24 h of the initial inoculum. Examination of the expression of proteins in the insect haemolymph using 2D electrophoresis and MALDI TOF analysis revealed an increased expression of a number of proteins associated with the insect immune response to infection 24 h after the initial exposure. This study demonstrates that the larvae of G. mellonella can withstand a lethal inoculum of C. albicans if pre-exposed to a non-lethal dose of yeast or polysaccharide 24 h previously which is mediated by increased expression of a number of antimicrobial peptides and the appearance of a number of peptides in the challenged larvae. PMID:16782387

  6. Ultraviolet radiation screening compounds

    Microsoft Academic Search

    CHARLES S. COCKELL; JOHN KNOWLAND

    1988-01-01

    Amongst the diversity of methods used by organisms to reduce damage caused by ultraviolet (UV) radiation, the synthesis of UV-screening compounds is almost ubiquitous. UV-screening compounds provide a passive method for the reduction of UV-induced damage and they are widely distributed across the microbial, plant and animal kingdoms. They share some common chemical features. It is likely that on early

  7. 8. DETAIL OF COMPUTER SCREEN AND CONTROL BOARDS: LEFT SCREEN ...

    Library of Congress Historic Buildings Survey, Historic Engineering Record, Historic Landscapes Survey

    8. DETAIL OF COMPUTER SCREEN AND CONTROL BOARDS: LEFT SCREEN TRACKS RESIDUAL CHLORINE; INDICATES AMOUNT OF SUNLIGHT WHICH ENABLES OPERATOR TO ESTIMATE NEEDED CHLORINE; CENTER SCREEN SHOWS TURNOUT STRUCTURES; RIGHT SCREEN SHOWS INDICATORS OF ALUMINUM SULFATE TANK FARM. - F. E. Weymouth Filtration Plant, 700 North Moreno Avenue, La Verne, Los Angeles County, CA

  8. Students' Compulsion To Screen: Research on Kenneth Burke's Terministic Screens.

    ERIC Educational Resources Information Center

    Strasma, Kip

    Kenneth Burke suggests that language operates from ultimate motives centered around "god-terms" through terministic screens. God-terms represent the strongest terministic screens in any culture: they screen attention to selected realities while screening or deflecting away others. A model of composition can be constructed from these theoretical…

  9. The role of cationic amino acid residues in the lethal activity of stonustoxin from stonefish (Synanceja horrida) venom.

    PubMed

    Khoo, H E; Chen, D; Yuen, R

    1998-03-01

    Stonustoxin (SNTX) is a two subunit pore-forming cytolytic protein purified from the venom of the stonefish (Synanceja horrida). SNTX also possesses lethal activity. Since cationic residues contribute significantly to the cytolytic activity of several pore-forming toxins, we examined the role of lysine and arginine residues in the lethal activity of SNTX. SNTX lost its lethal activity when the positively-charged side chains of lysine residues were converted to negatively-charged side chains upon succinylation. When the arginine residues were modified using 2,3-butanedione, SNTX also lost its lethal activity. However, the domains for cytolytic and lethal activity may not necessarily be the same. PMID:9556226

  10. Screening of solar cells

    NASA Technical Reports Server (NTRS)

    Appelbaum, J.; Chait, A.; Thompson, D. A.

    1993-01-01

    Because solar cells in a production batch are not identical, screening is performed to obtain similar cells for aggregation into arrays. A common technique for screening is based on a single operating point of the I-V characteristic of the cell, usually the maximum power point. As a result, inferior cell matching may occur at the actual operating points. Screening solar cells based on the entire I-V characteristic will inherently result in more similar cells in the array. An array consisting of more similar cells is likely to have better overall characteristics and more predictable performance. Solar cell screening methods and cell ranking are discussed. The concept of a mean cell is defined as a cell 'best' representing all the cells in the production batch. The screening and ranking of all cells are performed with respect to the mean cell. The comparative results of different screening methods are illustrated on a batch of 50 silicon cells of the Space Station Freedom.

  11. Non-Lethal Ionizing Radiation Promotes Aging-Like Phenotypic Changes of Human Hematopoietic Stem and Progenitor Cells in Humanized Mice

    PubMed Central

    Wang, Changshan; Oshima, Motohiko; Sashida, Goro; Tomioka, Takahisa; Hasegawa, Nagisa; Mochizuki-Kashio, Makiko; Nakajima-Takagi, Yaeko; Kusunoki, Yoichiro; Kyoizumi, Seishi; Imai, Kazue; Nakachi, Kei; Iwama, Atsushi

    2015-01-01

    Precise understanding of radiation effects is critical to develop new modalities for the prevention and treatment of radiation-induced damage. We previously reported that non-lethal doses of X-ray irradiation induce DNA damage in human hematopoietic stem and progenitor cells (HSPCs) reconstituted in NOD/Shi-scid IL2r?null (NOG) immunodeficient mice and severely compromise their repopulating capacity. In this study, we analyzed in detail the functional changes in human HSPCs in NOG mice following non-lethal radiation. We transplanted cord blood CD34+ HSPCs into NOG mice. At 12 weeks post-transplantation, the recipients were irradiated with 0, 0.5, or 1.0 Gy. At 2 weeks post-irradiation, human CD34+ HSPCs recovered from the primary recipient mice were transplanted into secondary recipients. CD34+ HSPCs from irradiated mice showed severely impaired reconstitution capacity in the secondary recipient mice. Of interest, non-lethal radiation compromised contribution of HSPCs to the peripheral blood cells, particularly to CD19+ B lymphocytes, which resulted in myeloid-biased repopulation. Co-culture of limiting numbers of CD34+ HSPCs with stromal cells revealed that the frequency of B cell-producing CD34+ HSPCs at 2 weeks post-irradiation was reduced more than 10-fold. Furthermore, the key B-cell regulator genes such as IL-7R and EBF1 were downregulated in HSPCs upon 0.5 Gy irradiation. Given that compromised repopulating capacity and myeloid-biased differentiation are representative phenotypes of aged HSCs, our findings indicate that non-lethal ionizing radiation is one of the critical external stresses that promote aging of human HSPCs in the bone marrow niche. PMID:26161905

  12. 40 CFR 798.5275 - Sex-linked recessive lethal test in drosophila melanogaster.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ...approximately one-fifth of the entire haploid genome. (b) Definitions. (1) Lethal mutation is a change in the genome which, when expressed, causes death...Recessive mutation is a change in the genome which is expressed in the...

  13. 40 CFR 798.5275 - Sex-linked recessive lethal test in drosophila melanogaster.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ...approximately one-fifth of the entire haploid genome. (b) Definitions. (1) Lethal mutation is a change in the genome which, when expressed, causes death...Recessive mutation is a change in the genome which is expressed in the...

  14. Lethal aggression in mobile forager bands and implications for the origins of war.

    PubMed

    Fry, Douglas P; Söderberg, Patrik

    2013-07-19

    It has been argued that warfare evolved as a component of early human behavior within foraging band societies. We investigated lethal aggression in a sample of 21 mobile forager band societies (MFBS) derived systematically from the standard cross-cultural sample. We hypothesized, on the basis of mobile forager ethnography, that most lethal events would stem from personal disputes rather than coalitionary aggression against other groups (war). More than half of the lethal aggression events were perpetrated by lone individuals, and almost two-thirds resulted from accidents, interfamilial disputes, within-group executions, or interpersonal motives such as competition over a particular woman. Overall, the findings suggest that most incidents of lethal aggression among MFBS may be classified as homicides, a few others as feuds, and a minority as war. PMID:23869015

  15. ORIGINAL PAPER Evaluating the impact of non-lethal DNA sampling on two

    E-print Network

    Landis, Doug

    ORIGINAL PAPER Evaluating the impact of non-lethal DNA sampling on two butterflies, Vanessa cardui of Vanessa cardui and Satyrodes eurydice. Based on these studies we were successful in obtaining a permit

  16. A model of anthrax toxin lethal factor bound to protective antigen

    E-print Network

    Baker, David

    translocation in an N- to C-terminal direction. computation docking electrostatic Bacillus anthracis), and lethal factor (LF), that are collectively referred to as anthrax toxin (1). After its proteolytic

  17. A systems analysis identifies a feedforward inflammatory circuit leading to lethal influenza infection.

    PubMed

    Brandes, Marlène; Klauschen, Frederick; Kuchen, Stefan; Germain, Ronald N

    2013-07-01

    For acutely lethal influenza infections, the relative pathogenic contributions of direct viral damage to lung epithelium versus dysregulated immunity remain unresolved. Here, we take a top-down systems approach to this question. Multigene transcriptional signatures from infected lungs suggested that elevated activation of inflammatory signaling networks distinguished lethal from sublethal infections. Flow cytometry and gene expression analysis involving isolated cell subpopulations from infected lungs showed that neutrophil influx largely accounted for the predictive transcriptional signature. Automated imaging analysis, together with these gene expression and flow data, identified a chemokine-driven feedforward circuit involving proinflammatory neutrophils potently driven by poorly contained lethal viruses. Consistent with these data, attenuation, but not ablation, of the neutrophil-driven response increased survival without changing viral spread. These findings establish the primacy of damaging innate inflammation in at least some forms of influenza-induced lethality and provide a roadmap for the systematic dissection of infection-associated pathology. PMID:23827683

  18. Dystrophin deficiency causes lethal muscle hypertrophy in cats.

    PubMed

    Gaschen, F P; Hoffman, E P; Gorospe, J R; Uhl, E W; Senior, D F; Cardinet, G H; Pearce, L K

    1992-07-01

    Two 5-month-old male Domestic Shorthair littermates showed general skeletal muscle hypertrophy, multifocal submucosal lingual calcification with lingual enlargement, and excessive salivation. Both cats had a reduced level of activity, walked with a stiff gait, and tended to "bunny hop" when they ran. These clinical features were similar to those of previously reported dystrophin-deficient cats. Using multiple dystrophin antibodies, we found that the cats described in this report also showed marked dystrophin deficiency. The histopathology was remarkable for hypertrophy and splitting of fibers, and progressive accumulation of calcium deposits within the muscle. There was little or no endomysial fibrosis at 2 years of age. The natural history of dystrophin-deficiency in cats has not been described: both previous cats had been euthanized at 2 years of age prior to experiencing any life-threatening problems. At 6 months of age, one of the new cats developed megaesophagus because of severe progressive hypertrophy of the diaphragmatic muscles. The diaphragm completely occluded the esophagus, and the cat was euthanized for humane reasons. The second cat remained in good condition until age 18 months when it developed acute renal failure attributed to severe prolonged dehydration and hyperosmolality. The cat recovered after receiving supportive treatment but was unable to maintain fluid homeostasis. The insufficient water intake was attributed to glossal hypertrophy and dysfunction. At age 2 years, the cat received regular subcutaneous injections of low-sodium fluids to maintain proper hydration. The clinical consequence of dystrophin deficiency in cats is lethal muscle hypertrophy. We have called the feline disease "hypertrophic feline muscular dystrophy" (HFMD). PMID:1506854

  19. Peptidomimetic fluoromethylketone rescues mice from lethal endotoxic shock.

    PubMed Central

    Grobmyer, S. R.; Armstrong, R. C.; Nicholson, S. C.; Gabay, C.; Arend, W. P.; Potter, S. H.; Melchior, M.; Fritz, L. C.; Nathan, C. F.

    1999-01-01

    BACKGROUND: Septic shock is a leading cause of mortality in intensive care units. No new interventions in the last 20 years have made a substantial impact on the outcome of patients with septic shock. Identification of inhibitable pathways that mediate death in shock is an important goal. MATERIALS AND METHODS: Two novel caspase inhibitors, (2-indolyl)-carbonyl-Ala-Asp-fluoromethylketone (IDN 1529) and (1-methyl-3-methyl-2-indolyl)-carbonyl-Val-Asp-fluoromethylketone (IDN 1965), were studied in a murine model of endotoxic shock. RESULTS: IDN 1529 prolonged survival when given before or up to 3 hr after high-dose LPS (p < 0.01) and increased by 2.2-fold the number of animals surviving longterm after a lower dose of LPS (p < 0.01). Despite its similar chemical structure, IDN 1965 lacked these protective effects. Both compounds inhibited caspases 1, 2, 3, 6, 8, and 9, and both afforded comparable reduction in Fas- and LPS-induced caspase 3-like activity and apoptosis. Paradoxically, administration of IDN 1529 but not IDN 1965 led to an increase in the LPS-induced elevation of serum cytokines related directly (IL-1beta, IL-18) or indirectly (IL-1alpha, IL-1Ra) to the action of caspase 1. CONCLUSIONS: A process that appears to be distinct from both apoptosis and the release of inflammatory cytokines is a late-acting requirement for lethality in endotoxic shock. Inhibition of this process can rescue mice even when therapy is initiated after LPS has made the mice severely ill. Images Fig. 4 Fig. 5 PMID:10551900

  20. MYC-mediated synthetic lethality for treatment of hematological malignancies.

    PubMed

    Li, Xin; Zhang, Xin A; Xie, Wei; Li, Xiaoqing; Huang, Shiang

    2015-01-01

    Deregulated c-MYC expression is found in many human malignancies. MYC activation induces multiple lineages of hematological malignancies in single Myc transgenic mice. MYC inactivation causes tumor regression. MYC is therefore an attractive target for cancer treatment. However, little progress has been made in the development and application of targeted MYC inactivation in clinical practice. In double Myc transgenic mouse models, Myc-driven leukemogenesis and lymphomagenesis can be accelerated by transduction of non-MYC oncogenes, leading to dual addiction to MYC and the non-MYC oncogenes. Wang et al. (2004) first established the concept of MYC-mediated synthetic lethality (MYC-SL). MYC overexpression sensitized cells to TRAILand DR5-agonist-induced apoptosis. This suggests that MYC-dependent tumor cells may be killed by targeting partner oncogenes of MYC. Many small molecule inhibitors (SMIs) have been proven to induce MYC-SL by targeting AUK-B, Brd4, CDK1, CHK1, MCL-1, the mTOR/4E-BP1/eIF4E pathway, and PIM1/2. Compared with conventional treatment approaches, SMI-induced MYC-SL displays highly selective anticancer activity and much lower cytotoxicity to normal cells. SMI-induced MYC-SL can reverse eIF4F- and PIM2-induced multiple chemoresistance. The combination of an SMI with chemotherapeutic agents can elevate chemotherapy efficacy by enhancing chemosensitivity. This combination will be a promising novel approach to treating MYC-dependent tumors by inducing MYC-SL. PMID:25564254

  1. Origin of the lethal gas burst from Lake Monoun, Cameroun

    USGS Publications Warehouse

    Sigurdsson, H.; Devine, J.D.; Tchua, F.M.; Presser, F.M.; Pringle, M.K.W.; Evans, William C.

    1987-01-01

    On 15 August, 1984, a lethal gas burst issued from a submerged 96-m-deep crater in Lake Monoun in Cameroun, western Africa, killing 37 people. The event was associated with a landslide from the eastern crater rim, which slumped into deep water. Waters below 50 m are anoxic, dominated by high Fe2+ (???600 mg/l) and HCO3- (??? 1900 mg/l), anoxic and supersaturated with siderite, which is a major component of the crater floor sediments. The unusually high Fe2+ levels are attributed to reduction of laterite-derived ferric iron gradually brought into the lake as loess and in river input. Sulfur compounds are below detection limits in both water and gas. Gases effervescing from depressurized deep waters are dominantly CO2 with minor CH4, having ??13C of -7.18 and -54.8 per mil, respectively. Bacterial decomposition of organic matter may account for the methane, but 14C of lake water indicates that only 10% of the carbon is modern, giving an apparent age of 18,000 years. The dominant source of carbon is therefore attributed to long-term emission of CO2 as volcanic exhalation from vents within the crater, which led to gradual build-up of HCO3- in the lake. The density stratification of the lake may have been upset by an earthquake and underwater landslide on 15 August, which triggered overturn of the lake and caused nucleation of CO2 in the deep water. The resultant ebullition of CO2 from deep lake waters led to a gas burst at the surface and locally generated a water wave up to 5 m high. People travelling through the gas cloud were asphyxiated, presumably from CO2, and suffered skin discoloration from unidentified components. ?? 1987.

  2. Determination of Radiation-induced Mutation Rates of Recessive Lethal Alleles in Saccharomyces

    Microsoft Academic Search

    W. Laskowski; K. Haefner

    1963-01-01

    COMPARING X-ray inactivation curves of haploid and diploid strains of Saccharomyces, Latarjet and Ephrussi1 were the first to suggest that radiation inactivation of yeast cells may be caused by an induction of recessive and dominant lethal mutations. The first evidence of X-ray-induced recessive and dominant lethal mutations was obtained by Mortimer2 by means of tetrad analysis and specific mating experiments

  3. Mice lacking functional STAT1 are highly susceptible to lethal infection with Lassa virus.

    PubMed

    Yun, Nadezhda E; Seregin, Alexey V; Walker, David H; Popov, Vsevolod L; Walker, Aida G; Smith, Jeanon N; Miller, Milagros; de la Torre, Juan C; Smith, Jennifer K; Borisevich, Viktoriya; Fair, Joseph N; Wauquier, Nadia; Grant, Donald S; Bockarie, Bayon; Bente, Dennis; Paessler, Slobodan

    2013-10-01

    Lassa fever (LF) is a potentially lethal human disease that is caused by the arenavirus Lassa virus (LASV). Annually, around 300,000 infections with up to 10,000 deaths occur in regions of Lassa fever endemicity in West Africa. Here we demonstrate that mice lacking a functional STAT1 pathway are highly susceptible to infection with LASV and develop lethal disease with pathology similar to that reported in humans. PMID:23903830

  4. Proteasome Activity Is Required for Anthrax Lethal Toxin To Kill Macrophages

    Microsoft Academic Search

    GUANGQING TANG; STEPHEN H. LEPPLA

    1999-01-01

    Anthrax lethal toxin (LeTx), consisting of protective antigen (PA) and lethal factor (LF), rapidly kills primary mouse macrophages and macrophage-like cell lines such as RAW 264.7. LF is translocated by PA into the cytosol of target cells, where it acts as a metalloprotease to cleave mitogen-activated protein kinase kinase 1 (MEK1) and possibly other proteins. In this study, we show

  5. Lethal and mutational effects of solar and UV radiation on Staphylococcus aureus

    Microsoft Academic Search

    Robin M. Chapple; Barbara Inglis; Peter R. Stewart

    1992-01-01

    Strains of Staphylococcus aureus, an opportunistic pathogen commonly found on human skin, were exposed to sunlight and UV C radiation, and the lethal and mutational effects measured. Sunlight killed cells with an inactivation constant of 3×10-5 per joule per square metre; UV C was much more lethal, giving an inactivation constant of approximately 0.1 per joule per square metre. Some

  6. Cytokine Inducing Activities of Rhizobial and Mesorhizobial Lipopolysaccharides of Different Lethal Toxicity

    Microsoft Academic Search

    T. Urbanik-Sypniewska; A. Choma; J. Kutkowska; T. KamiÑska; M. Kandefer-SzerszeÑ; R. Russa; J. Dolecka

    2000-01-01

    The lethality and cytokines-inducing activity of lipopolysaccharides (LPS) obtained from nodulating bacteria, Rhizobium leguminosarum and Mesorhizobium loti, were compared to those of Salmonella typhimurium LPSThe activity of Rleguminosarum LPS was almost comparable to Salmonella endotoxin in terms of lethality, Limulus lysate gelating activity and in vivo tumor necrosis factor-? (TNF-?), interleukin-1ß (IL-lß), interleukin-6 (IL-6) and interferon-? (IFN-?) induction capacityIn contrast

  7. The suicidal process in male prisoners making near-lethal suicide attempts

    Microsoft Academic Search

    Adrienne Rivlin; Seena Fazel; Lisa Marzano; Keith Hawton

    2011-01-01

    Most investigations of factors contributing to prisoner suicide have focused on suicidal behaviour as a discrete event and used official records or ‘psychological autopsy’ methodology. A potentially more informative approach is to study survivors of near-lethal suicide attempts about their suicidal process. We have investigated the suicidal process in male prisoners through semi-structured interviews with 60 prisoners who made near-lethal

  8. Lethal factors of diseases and protective countermeasures of wild and penned Giant pandas

    Microsoft Academic Search

    Zou Xinghuai; Wang Aimin; Zeng Lujin; He Guangxin; Wu Kongju; Chen Yucun; Weng Nina

    1998-01-01

    Statistical analysis of certain major diseases lethal to the animal was carried out through on-spot investigations and consultations\\u000a to related documentary materials. The results show that the first serious lethal factor is digestive system diseases. Next\\u000a comes malnutrition and organic exhaustion. In order of decremental percentages, other diseases are roundworm disease, epilepsy,\\u000a toxicosis, pneumonia, tumor, pericarditis, etc., In relation to

  9. Observations on Experimental Anthrax: Demonstration of a Specific Lethal Factor produced in vivo by Bacillus anthracis

    Microsoft Academic Search

    H. Smith; J. Keppie

    1954-01-01

    AN enigma in the study of the cause of death in anthrax has been that no lethal endo- or exo-toxin has been found in cultures of the organism1-3. Recently, we have been able to demonstrate a factor in the plasma of guinea pigs dying of anthrax which is not only lethal but also specifically neutralized by anthrax antiserum. This communication

  10. Cellular and systemic effects of anthrax lethal toxin and edema toxin.

    PubMed

    Moayeri, Mahtab; Leppla, Stephen H

    2009-12-01

    Anthrax lethal toxin (LT) and edema toxin (ET) are the major virulence factors of anthrax and can replicate the lethality and symptoms associated with the disease. This review provides an overview of our current understanding of anthrax toxin effects in animal models and the cytotoxicity (necrosis and apoptosis) induced by LT in different cells. A brief reexamination of early historic findings on toxin in vivo effects in the context of our current knowledge is also presented. PMID:19638283

  11. Physics in Screening Environments

    NASA Astrophysics Data System (ADS)

    Certik, Ondrej

    In the current study, we investigated atoms in screening environments like plasmas. It is common practice to extract physical data, such as temperature and electron densities, from plasma experiments. We present results that address inherent computational difficulties that arise when the screening approach is extended to include the interaction between the atomic electrons. We show that there may arise an ambiguity in the interpretation of physical properties, such as temperature and charge density, from experimental data due to the opposing effects of electron-nucleus screening and electron-electron screening. The focus of the work, however, is on the resolution of inherent computational challenges that appear in the computation of two-particle matrix elements. Those enter already at the Hartree-Fock level. Furthermore, as examples of post Hartree-Fock calculations, we show second-order Green's function results and many body perturbation theory results of second order. A self-contained derivation of all necessary equations has been included. The accuracy of the implementation of the method is established by comparing standard unscreened results for various atoms and molecules against literature for Hartree-Fock as well as Green's function and many body perturbation theory. The main results of the thesis are presented in the chapter called Screened Results, where the behavior of several atomic systems depending on electron-electron and electron-nucleus Debye screening was studied. The computer code that we have developed has been made available for anybody to use. Finally, we present and discuss results obtained for screened interactions. We also examine thoroughly the computational details of the calculations and particular implementations of the method.

  12. Autosomal recessive lethal mutations in two mutator stocks of Drosophila ananassae.

    PubMed

    Matsuda, M

    1991-12-01

    The frequency of recessive lethals in the 2nd chromosome was examined in two mutator stocks of Drosophila ananassae, ca and ca; px. They are characterized respectively by possessing an extrachromosomal clastogenic mutator in males, and by the retrotransposon "tom", which induces Om mutability only in females. The frequencies of recessive lethal mutations in the 2nd chromosome among progenies from males and females of the ca; px stock are 0.35 and 0.34 percent, respectively. Similarity of these frequencies indicates that tom does not induce recessive lethals in females. In contrast to the ca; px stock, the frequency of recessive lethals in males of the ca mutator stock was estimated to be 1.54 percent for the 2nd chromosome. No visible mutants except Minutes were recovered. Some recessive lethals derived from ca stock males were associated with chromosomal rearrangements. Being consistent with its high rate of Minute mutation it was demonstrated that the ca clastogenic mutator also induced recessive lethals. PMID:1814375

  13. Small-Molecule Inhibitors of Lethal Factor Protease Activity Protect against Anthrax Infection

    PubMed Central

    Crown, Devorah; Jiao, Guan-Sheng; Kim, Seongjin; Johnson, Alan; Leysath, Clinton; Leppla, Stephen H.

    2013-01-01

    Bacillus anthracis, the causative agent of anthrax, manifests its pathogenesis through the action of two secreted toxins. The bipartite lethal and edema toxins, a combination of lethal factor or edema factor with the protein protective antigen, are important virulence factors for this bacterium. We previously developed small-molecule inhibitors of lethal factor proteolytic activity (LFIs) and demonstrated their in vivo efficacy in a rat lethal toxin challenge model. In this work, we show that these LFIs protect against lethality caused by anthrax infection in mice when combined with subprotective doses of either antibiotics or neutralizing monoclonal antibodies that target edema factor. Significantly, these inhibitors provided protection against lethal infection when administered as a monotherapy. As little as two doses (10 mg/kg) administered at 2 h and 8 h after spore infection was sufficient to provide a significant survival benefit in infected mice. Administration of LFIs early in the infection was found to inhibit dissemination of vegetative bacteria to the organs in the first 32 h following infection. In addition, neutralizing antibodies against edema factor also inhibited bacterial dissemination with similar efficacy. Together, our findings confirm the important roles that both anthrax toxins play in establishing anthrax infection and demonstrate the potential for small-molecule therapeutics targeting these proteins. PMID:23774434

  14. Application of Bioluminescence Imaging to the Prediction of Lethality in Vaccinia Virus-Infected Mice? †

    PubMed Central

    Zaitseva, Marina; Kapnick, Senta M.; Scott, John; King, Lisa R.; Manischewitz, Jody; Sirota, Lev; Kodihalli, Shantha; Golding, Hana

    2009-01-01

    To find an alternative endpoint for the efficacy of antismallpox treatments, bioluminescence was measured in live BALB/c mice following lethal challenge with a recombinant WR vaccinia virus expressing luciferase. Intravenous vaccinia immunoglobulin treatments were used to confer protection on a proportion of animals. Using known lethality outcomes in 200 animals and total fluxes recorded daily in live animals, we performed univariate receiver operating characteristic (ROC) curve analysis to assess whether lethality can be predicted based on bioluminescence. Total fluxes in the spleens on day 3 and in the livers on day 5 generated accurate predictive models; the area under the ROC curve (AUC) was 0.91. Multiple logistic regression analysis utilizing a linear combination of six measurements: total flux in the liver on days 2, 3, and 5; in the spleen on days 1 and 3; and in the nasal cavity on day 4 generated the most accurate predictions (AUC = 0.96). This model predicted lethality in 90% of animals with only 10% of nonsurviving animals incorrectly predicted to survive. Compared with bioluminescence, ROC analysis with 25% and 30% weight loss as thresholds accurately predicted survival on day 5, but lethality predictions were low until day 9. Collectively, our data support the use of bioimaging for lethality prediction following vaccinia virus challenge and for gaining insight into protective mechanisms conferred by vaccines and therapeutics. PMID:19656894

  15. The frequency and allelism of lethal chromosomes in isolated desert populations of Drosophila pseudoobscura.

    PubMed

    Bryant, S H

    1976-12-01

    Second-chromosome lethals were extracted from four populations of Drosophila pseudoobscura in Southern California. Two of the populations were from desert oases and two from the classic habitat on Mt. San Jacinto, previously studied by Dobzhansky. Allelism tests were made on the lethals within and between all locations. The frequency of lethal second-chromosomes in each location was 0.18, and this was not different from the results of other workers for samples throughout the species range. Interpopulational allelism rates were about 0.005, and not different from earlier results of Dobzhansky. Intrapopulational rates in this study were, with one exception, the same as the interpopulational rates, and significantly lower than Dobzhansky found using the third chromosome. This may be due to lethals being linked with heterotic third-chromosome inversions. The allelism rate of the exceptional population (about 0.03 and equal to Dobzhansky's intrapopulational results) may be due to heterotic lethals, or a founder effect. Two lethals were found in three populations each, possibly due to migration among these populations, which are up to 334 km apart. PMID:1010313

  16. The Maternally Expressed WRKY Transcription Factor TTG2 Controls Lethality in Interploidy Crosses of Arabidopsis

    PubMed Central

    Dilkes, Brian P; Spielman, Melissa; Weizbauer, Renate; Watson, Brian; Burkart-Waco, Diana; Scott, Rod J; Comai, Luca

    2008-01-01

    The molecular mechanisms underlying lethality of F1 hybrids between diverged parents are one target of speciation research. Crosses between diploid and tetraploid individuals of the same genotype can result in F1 lethality, and this dosage-sensitive incompatibility plays a role in polyploid speciation. We have identified variation in F1 lethality in interploidy crosses of Arabidopsis thaliana and determined the genetic architecture of the maternally expressed variation via QTL mapping. A single large-effect QTL, DR. STRANGELOVE 1 (DSL1), was identified as well as two QTL with epistatic relationships to DSL1. DSL1 affects the rate of postzygotic lethality via expression in the maternal sporophyte. Fine mapping placed DSL1 in an interval encoding the maternal effect transcription factor TTG2. Maternal parents carrying loss-of-function mutations in TTG2 suppressed the F1 lethality caused by paternal excess interploidy crosses. The frequency of cellularization in the endosperm was similarly affected by both natural variation and ttg2 loss-of-function mutants. The simple genetic basis of the natural variation and effects of single-gene mutations suggests that F1 lethality in polyploids could evolve rapidly. Furthermore, the role of the sporophytically active TTG2 gene in interploidy crosses indicates that the developmental programming of the mother regulates the viability of interploidy hybrid offspring. PMID:19071961

  17. Score test variable screening

    PubMed Central

    Zhao, Sihai Dave; Li, Yi

    2015-01-01

    Variable screening has emerged as a crucial first step in the analysis of high-throughput data, but existing procedures can be computationally cumbersome, difficult to justify theoretically, or inapplicable to certain types of analyses. Motivated by a high-dimensional censored quantile regression problem in multiple myeloma genomics, this paper makes three contributions. First, we establish a score test-based screening framework, which is widely applicable, extremely computationally efficient, and relatively simple to justify. Secondly, we propose a resampling-based procedure for selecting the number of variables to retain after screening according to the principle of reproducibility. Finally, we propose a new iterative score test screening method which is closely related to sparse regression. In simulations we apply our methods to four different regression models and show that they can outperform existing procedures. We also apply score test screening to an analysis of gene expression data from multiple myeloma patients using a censored quantile regression model to identify high-risk genes. PMID:25124197

  18. Newborn hearing screening.

    PubMed

    Stewart, D L; Pearlman, A

    1994-11-01

    Congenital deafness is a relatively common problem with an incidence of 1/300 to 1/1000. Most states have no mass screening program for hearing loss, but the state of Kentucky compiles a High Risk Registry which is a historical survey of parents relating to risk factors for hearing loss. Unfortunately this survey can miss 50% of those who have a hearing deficit. If not detected prior to discharge, there is often a delay in diagnosis of deafness which prevents early intervention. We report 2 years' experience at Kosair Children's Hospital where 1,987 infants admitted to well baby, intermediate, or intensive care nurseries were screened using the ALGO-1 screener (Natus Medical Inc, Foster City, CA) which is a modified auditory brain stem evoked response (ABR). Our screening of this population led to an 11% incidence of referral for complete audiological evaluation. There were no significant complications. Forty-eight infants were found to have nonspecified, sensorineural, or conductive hearing loss. The positive predictive value of the test was 96%. Therefore, we feel that the use of the modified ABR in the newborn is a timely, cost efficient method of screening for hearing loss and should be used for mass screening of all newborns. PMID:7806952

  19. Screening for depression.

    PubMed

    Maurer, Douglas M

    2012-01-15

    In the United States, depression affects up to 9 percent of patients and accounts for more than $43 billion in medical care costs. The U.S. Preventive Services Task Force recommends screening in adolescents and adults in clinical practices that have systems in place to ensure accurate diagnosis, effective treatment, and follow-up. It does not recommend for or against screening for depression in children seven to 11 years of age or screening for suicide risk in the general population. The Patient Health Questionnaire (PHQ)-2 and PHQ-9 are commonly used and validated screening tools. The PHQ-2 has a 97 percent sensitivity and 67 percent specificity in adults, whereas the PHQ-9 has a 61 percent sensitivity and 94 percent specificity in adults. If the PHQ-2 is positive for depression, the PHQ-9 should be administered; in older adults, the 15-item Geriatric Depression Scale is also an appropriate follow-up test. If these screening tests are positive for depression, further evaluation is needed to confirm that the patient's symptoms meet the Diagnostic and Statistical Manual of Mental Disorders' criteria for diagnosis. PMID:22335214

  20. Sharing chemical relationships does not reveal structures.

    PubMed

    Matlock, Matthew; Swamidass, S Joshua

    2014-01-27

    In this study, we propose a new, secure method of sharing useful chemical information from small-molecule libraries, without revealing the structures of the libraries' molecules. Our method shares the relationship between molecules rather than structural descriptors. This is an important advance because, over the past few years, several groups have developed and published new methods of analyzing small-molecule screening data. These methods include advanced hit-picking protocols, promiscuous active filters, economic optimization algorithms, and screening visualizations, which can identify patterns in the data that might otherwise be overlooked. Application of these methods to private data requires finding strategies for sharing useful chemical data without revealing chemical structures. This problem has been examined in the context of ADME prediction models, with results from information theory suggesting it is impossible to share useful chemical information without revealing structures. In contrast, we present a new strategy for encoding the relationships between molecules instead of their structures, based on anonymized scaffold networks and trees, that safely shares enough chemical information to be useful in analyzing chemical data, while also sufficiently blinding structures from discovery. We present the details of this encoding, an analysis of the usefulness of the information it conveys, and the security of the structures it encodes. This approach makes it possible to share data across institutions, and may securely enable collaborative analysis that can yield insight into both specific projects and screening technology as a whole. PMID:24289228