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Sample records for accurate transcript normalization

  1. Identification and Evaluation of Reference Genes for Accurate Transcription Normalization in Safflower under Different Experimental Conditions

    PubMed Central

    Li, Dandan; Hu, Bo; Wang, Qing; Liu, Hongchang; Pan, Feng; Wu, Wei

    2015-01-01

    Safflower (Carthamus tinctorius L.) has received a significant amount of attention as a medicinal plant and oilseed crop. Gene expression studies provide a theoretical molecular biology foundation for improving new traits and developing new cultivars. Real-time quantitative PCR (RT-qPCR) has become a crucial approach for gene expression analysis. In addition, appropriate reference genes (RGs) are essential for accurate and rapid relative quantification analysis of gene expression. In this study, fifteen candidate RGs involved in multiple metabolic pathways of plants were finally selected and validated under different experimental treatments, at different seed development stages and in different cultivars and tissues for real-time PCR experiments. These genes were ABCS, 60SRPL10, RANBP1, UBCL, MFC, UBCE2, EIF5A, COA, EF1-β, EF1, GAPDH, ATPS, MBF1, GTPB and GST. The suitability evaluation was executed by the geNorm and NormFinder programs. Overall, EF1, UBCE2, EIF5A, ATPS and 60SRPL10 were the most stable genes, and MBF1, as well as MFC, were the most unstable genes by geNorm and NormFinder software in all experimental samples. To verify the validation of RGs selected by the two programs, the expression analysis of 7 CtFAD2 genes in safflower seeds at different developmental stages under cold stress was executed using different RGs in RT-qPCR experiments for normalization. The results showed similar expression patterns when the most stable RGs selected by geNorm or NormFinder software were used. However, the differences were detected using the most unstable reference genes. The most stable combination of genes selected in this study will help to achieve more accurate and reliable results in a wide variety of samples in safflower. PMID:26457898

  2. LSM Proteins Provide Accurate Splicing and Decay of Selected Transcripts to Ensure Normal Arabidopsis Development[W

    PubMed Central

    Perea-Resa, Carlos; Hernández-Verdeja, Tamara; López-Cobollo, Rosa; Castellano, María del Mar; Salinas, Julio

    2012-01-01

    In yeast and animals, SM-like (LSM) proteins typically exist as heptameric complexes and are involved in different aspects of RNA metabolism. Eight LSM proteins, LSM1 to 8, are highly conserved and form two distinct heteroheptameric complexes, LSM1-7 and LSM2-8,that function in mRNA decay and splicing, respectively. A search of the Arabidopsis thaliana genome identifies 11 genes encoding proteins related to the eight conserved LSMs, the genes encoding the putative LSM1, LSM3, and LSM6 proteins being duplicated. Here, we report the molecular and functional characterization of the Arabidopsis LSM gene family. Our results show that the 11 LSM genes are active and encode proteins that are also organized in two different heptameric complexes. The LSM1-7 complex is cytoplasmic and is involved in P-body formation and mRNA decay by promoting decapping. The LSM2-8 complex is nuclear and is required for precursor mRNA splicing through U6 small nuclear RNA stabilization. More importantly, our results also reveal that these complexes are essential for the correct turnover and splicing of selected development-related mRNAs and for the normal development of Arabidopsis. We propose that LSMs play a critical role in Arabidopsis development by ensuring the appropriate development-related gene expression through the regulation of mRNA splicing and decay. PMID:23221597

  3. Transcriptional network control of normal and leukaemic haematopoiesis

    PubMed Central

    Sive, Jonathan I.; Göttgens, Berthold

    2014-01-01

    Transcription factors (TFs) play a key role in determining the gene expression profiles of stem/progenitor cells, and defining their potential to differentiate into mature cell lineages. TF interactions within gene-regulatory networks are vital to these processes, and dysregulation of these networks by TF overexpression, deletion or abnormal gene fusions have been shown to cause malignancy. While investigation of these processes remains a challenge, advances in genome-wide technologies and growing interactions between laboratory and computational science are starting to produce increasingly accurate network models. The haematopoietic system provides an attractive experimental system to elucidate gene regulatory mechanisms, and allows experimental investigation of both normal and dysregulated networks. In this review we examine the principles of TF-controlled gene regulatory networks and the key experimental techniques used to investigate them. We look in detail at examples of how these approaches can be used to dissect out the regulatory mechanisms controlling normal haematopoiesis, as well as the dysregulated networks associated with haematological malignancies. PMID:25014893

  4. Transcriptional network control of normal and leukaemic haematopoiesis.

    PubMed

    Sive, Jonathan I; Göttgens, Berthold

    2014-12-10

    Transcription factors (TFs) play a key role in determining the gene expression profiles of stem/progenitor cells, and defining their potential to differentiate into mature cell lineages. TF interactions within gene-regulatory networks are vital to these processes, and dysregulation of these networks by TF overexpression, deletion or abnormal gene fusions have been shown to cause malignancy. While investigation of these processes remains a challenge, advances in genome-wide technologies and growing interactions between laboratory and computational science are starting to produce increasingly accurate network models. The haematopoietic system provides an attractive experimental system to elucidate gene regulatory mechanisms, and allows experimental investigation of both normal and dysregulated networks. In this review we examine the principles of TF-controlled gene regulatory networks and the key experimental techniques used to investigate them. We look in detail at examples of how these approaches can be used to dissect out the regulatory mechanisms controlling normal haematopoiesis, as well as the dysregulated networks associated with haematological malignancies.

  5. Does transcription factor induced pluripotency accurately mimic embryo derived pluripotency?

    PubMed

    Lowry, William E

    2012-10-01

    When Takahashi and Yamanaka first demonstrated that just four transcription factors could reprogram a fibroblast to a pluripotent state, the first wave of data to emerge focused on how similar these induced pluripotent stem cells (iPSCs) were to embryo-derived pluripotent stem cells (ESCs) [1]. The next wave of data focused on determining the degree of difference between iPSCs and ESCs [2]. Now the focus is on tweaking the process to generate iPSCs that are more similar to ESCs [3,4]. Because transcription factor based reprogramming allows for nearly any type of cell to be created from any donor cell, there is obviously enormous interest in this technique as a tool for both basic developmental biology and for clinical applications. In this review, I will attempt to summarize the data that serve to distinguish these types of pluripotent stem cells and speculate on the ramifications of any differences.

  6. Reference genes for normalizing transcription in diploid and tetraploid Arabidopsis.

    PubMed

    Wang, Haibin; Wang, Jingjing; Jiang, Jiafu; Chen, Sumei; Guan, Zhiyong; Liao, Yuan; Chen, Fadi

    2014-10-27

    Published transcription data from a set of 19 diploid Arabidopsis thaliana and 5 tetraploid (3 allo- and 2 auto- tetraploid) Arabidopsis accessions were re-analysed to identify reliable reference genes for normalization purposes. Five conventional and 16 novel reference genes previously derived from microarray data covering a wide range of abundance in absolute expression levels in diploid A. thaliana Col-0 were employed. Transcript abundance was well conserved for all 21 potential reference genes in the diploid A. thaliana accessions, with geNorm and NormFinder analysis indicating that AT5G46630, AT1G13320, AT4G26410, AT5G60390 and AT5G08290 were the most stable. However, conservation was less good among the tetraploid accessions, with the transcription of seven of the 21 genes being undetectable in all allotetraploids. The most stable gene was AT5G46630, while AT1G13440 was the unstable one. Hence, the choice of reference gene(s) for A. thaliana is quite wide, but with respect to the analysis of transcriptomic data derived from the tetraploids, it is probably necessary to select more than one reference gene.

  7. Analysis of transcriptional responses of normalizing genes on Crassostrea brasiliana under different experimental conditions.

    PubMed

    Müller, Gabrielle do Amaral E Silva; de Lima, Daína; Zacchi, Flávia Lucena; Piazza, Rômi Sharon; Lüchmann, Karim Hahn; Mattos, Jacó Joaquim; Schlenk, Daniel; Bainy, Afonso Celso Dias

    2017-02-04

    Bivalves show remarkable plasticity to environmental changes and have been proposed as sentinel organisms in biomonitoring. Studies related to transcriptional analysis using quantitative real-time PCR (qRT-PCR) in these organisms have notably increased, imposing a need to identify and validate adequate reference genes for an accurate and reliable analysis. In the present study, nine reference genes were selected from transcriptome data of Crassostrea brasiliana in order to identify their suitability as qRT-PCR normalizer genes. The transcriptional patterns were analyzed in gills of oysters under three different conditions: different temperatures (18°C, 24°C or 32°C) and phenanthrene (PHE) (100 µg.L(-1) ) combined exposure; different salinities (10, 25 or 35 ‰) and PHE combined exposure and 10% of diesel fuel water-accommodated fraction (diesel-WAF) exposure. Reference gene stability was calculated using five algorithms (geNorm, NormFinder, BestKeeper, ΔCt, RefFinder). Transcripts of ankyrin-like (ANK), GAPDH-like and α tubulin-like (TUBA) genes showed minor changes in different temperature/PHE treatment. Transcripts of ANK, β actin-like and β tubulin-like genes showed better stability at salinity/PHE treatment, and ANK, TUBA and 28S ribosomal protein-like genes showed the most stable transcription pattern in oysters exposed to diesel-WAF exposure. This study constitutes the first systematic analysis on reference gene selection for qRT-PCR normalization in C. brasiliana. These genes could be employed in studies using qRT-PCR analysis under similar experimental conditions. This article is protected by copyright. All rights reserved.

  8. Intragenic DNA methylation in transcriptional regulation, normal differentiation and cancer.

    PubMed

    Kulis, Marta; Queirós, Ana C; Beekman, Renée; Martín-Subero, José I

    2013-11-01

    Ever since the discovery of DNA methylation at cytosine residues, the role of this so called fifth base has been extensively studied and debated. Until recently, the majority of DNA methylation studies focused on the analysis of CpG islands associated to promoter regions. However, with the upcoming possibilities to study DNA methylation in a genome-wide context, this epigenetic mark can now be studied in an unbiased manner. As a result, recent studies have shown that not only promoters but also intragenic and intergenic regions are widely modulated during physiological processes and disease. In particular, it is becoming increasingly clear that DNA methylation in the gene body is not just a passive witness of gene transcription but it seems to be actively involved in multiple gene regulation processes. In this review we discuss the potential role of intragenic DNA methylation in alternative promoter usage, regulation of short and long non-coding RNAs, alternative RNA processing, as well as enhancer activity. Furthermore, we summarize how the intragenic DNA methylome is modified both during normal cell differentiation and neoplastic transformation.

  9. Are Normally Sighted, Visually Impaired, and Blind Pedestrians Accurate and Reliable at Making Street Crossing Decisions?

    PubMed Central

    Hassan, Shirin E.

    2012-01-01

    Purpose. The purpose of this study is to measure the accuracy and reliability of normally sighted, visually impaired, and blind pedestrians at making street crossing decisions using visual and/or auditory information. Methods. Using a 5-point rating scale, safety ratings for vehicular gaps of different durations were measured along a two-lane street of one-way traffic without a traffic signal. Safety ratings were collected from 12 normally sighted, 10 visually impaired, and 10 blind subjects for eight different gap times under three sensory conditions: (1) visual plus auditory information, (2) visual information only, and (3) auditory information only. Accuracy and reliability in street crossing decision-making were calculated for each subject under each sensory condition. Results. We found that normally sighted and visually impaired pedestrians were accurate and reliable in their street crossing decision-making ability when using either vision plus hearing or vision only (P > 0.05). Under the hearing only condition, all subjects were reliable (P > 0.05) but inaccurate with their street crossing decisions (P < 0.05). Compared to either the normally sighted (P = 0.018) or visually impaired subjects (P = 0.019), blind subjects were the least accurate with their street crossing decisions under the hearing only condition. Conclusions. Our data suggested that visually impaired pedestrians can make accurate and reliable street crossing decisions like those of normally sighted pedestrians. When using auditory information only, all subjects significantly overestimated the vehicular gap time. Our finding that blind pedestrians performed significantly worse than either the normally sighted or visually impaired subjects under the hearing only condition suggested that they may benefit from training to improve their detection ability and/or interpretation of vehicular gap times. PMID:22427593

  10. Reference Genes in the Pathosystem Phakopsora pachyrhizi/ Soybean Suitable for Normalization in Transcript Profiling

    PubMed Central

    Hirschburger, Daniela; Müller, Manuel; Voegele, Ralf T.; Link, Tobias

    2015-01-01

    Phakopsora pachyrhizi is a devastating pathogen on soybean, endangering soybean production worldwide. Use of Host Induced Gene Silencing (HIGS) and the study of effector proteins could provide novel strategies for pathogen control. For both approaches quantification of transcript abundance by RT-qPCR is essential. Suitable stable reference genes for normalization are indispensable to obtain accurate RT-qPCR results. According to the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines and using algorithms geNorm and NormFinder we tested candidate reference genes from P. pachyrhizi and Glycine max for their suitability in normalization of transcript levels throughout the infection process. For P. pachyrhizi we recommend a combination of CytB and PDK or GAPDH for in planta experiments. Gene expression during in vitro stages and over the whole infection process was found to be highly unstable. Here, RPS14 and UbcE2 are ranked best by geNorm and NormFinder. Alternatively CytB that has the smallest Cq range (Cq: quantification cycle) could be used. We recommend specification of gene expression relative to the germ tube stage rather than to the resting urediospore stage. For studies omitting the resting spore and the appressorium stages a combination of Elf3 and RPS9, or PKD and GAPDH should be used. For normalization of soybean genes during rust infection Ukn2 and cons7 are recommended. PMID:26404265

  11. Accurate Gene Expression-Based Biodosimetry Using a Minimal Set of Human Gene Transcripts

    SciTech Connect

    Tucker, James D.; Joiner, Michael C.; Thomas, Robert A.; Grever, William E.; Bakhmutsky, Marina V.; Chinkhota, Chantelle N.; Smolinski, Joseph M.; Divine, George W.; Auner, Gregory W.

    2014-03-15

    Purpose: Rapid and reliable methods for conducting biological dosimetry are a necessity in the event of a large-scale nuclear event. Conventional biodosimetry methods lack the speed, portability, ease of use, and low cost required for triaging numerous victims. Here we address this need by showing that polymerase chain reaction (PCR) on a small number of gene transcripts can provide accurate and rapid dosimetry. The low cost and relative ease of PCR compared with existing dosimetry methods suggest that this approach may be useful in mass-casualty triage situations. Methods and Materials: Human peripheral blood from 60 adult donors was acutely exposed to cobalt-60 gamma rays at doses of 0 (control) to 10 Gy. mRNA expression levels of 121 selected genes were obtained 0.5, 1, and 2 days after exposure by reverse-transcriptase real-time PCR. Optimal dosimetry at each time point was obtained by stepwise regression of dose received against individual gene transcript expression levels. Results: Only 3 to 4 different gene transcripts, ASTN2, CDKN1A, GDF15, and ATM, are needed to explain ≥0.87 of the variance (R{sup 2}). Receiver-operator characteristics, a measure of sensitivity and specificity, of 0.98 for these statistical models were achieved at each time point. Conclusions: The actual and predicted radiation doses agree very closely up to 6 Gy. Dosimetry at 8 and 10 Gy shows some effect of saturation, thereby slightly diminishing the ability to quantify higher exposures. Analyses of these gene transcripts may be advantageous for use in a field-portable device designed to assess exposures in mass casualty situations or in clinical radiation emergencies.

  12. Discovery and Classification of Fusion Transcripts in Prostate Cancer and Normal Prostate Tissue.

    PubMed

    Luo, Jian-Hua; Liu, Silvia; Zuo, Ze-Hua; Chen, Rui; Tseng, George C; Yu, Yan P

    2015-07-01

    Fusion transcript formation is one of the fundamental mechanisms that drives the development of prostate cancer. Because of the advance of high-throughput parallel sequencing, many fusion transcripts have been discovered. However, the discovery rate of fusion transcripts specific for prostate cancer is lagging behind the discoveries made on chromosome abnormalities of prostate cancer. Recent analyses suggest that many fusion transcripts are present in both benign and cancerous tissues. Some of these fusion transcripts likely represent important components of normal gene expression in cells. It is necessary to identify the criteria and features of fusion transcripts that are specific for cancer. In this review, we discuss optimization of RNA sequencing depth for fusion transcript discovery and the characteristics of fusion transcripts in normal prostate tissues and prostate cancer. We also propose a new classification of cancer-specific fusion transcripts on the basis of their tail gene fusion protein product and the roles that these fusions may play in cancer development.

  13. Combining transcription factor binding affinities with open-chromatin data for accurate gene expression prediction.

    PubMed

    Schmidt, Florian; Gasparoni, Nina; Gasparoni, Gilles; Gianmoena, Kathrin; Cadenas, Cristina; Polansky, Julia K; Ebert, Peter; Nordström, Karl; Barann, Matthias; Sinha, Anupam; Fröhler, Sebastian; Xiong, Jieyi; Dehghani Amirabad, Azim; Behjati Ardakani, Fatemeh; Hutter, Barbara; Zipprich, Gideon; Felder, Bärbel; Eils, Jürgen; Brors, Benedikt; Chen, Wei; Hengstler, Jan G; Hamann, Alf; Lengauer, Thomas; Rosenstiel, Philip; Walter, Jörn; Schulz, Marcel H

    2017-01-09

    The binding and contribution of transcription factors (TF) to cell specific gene expression is often deduced from open-chromatin measurements to avoid costly TF ChIP-seq assays. Thus, it is important to develop computational methods for accurate TF binding prediction in open-chromatin regions (OCRs). Here, we report a novel segmentation-based method, TEPIC, to predict TF binding by combining sets of OCRs with position weight matrices. TEPIC can be applied to various open-chromatin data, e.g. DNaseI-seq and NOMe-seq. Additionally, Histone-Marks (HMs) can be used to identify candidate TF binding sites. TEPIC computes TF affinities and uses open-chromatin/HM signal intensity as quantitative measures of TF binding strength. Using machine learning, we find low affinity binding sites to improve our ability to explain gene expression variability compared to the standard presence/absence classification of binding sites. Further, we show that both footprints and peaks capture essential TF binding events and lead to a good prediction performance. In our application, gene-based scores computed by TEPIC with one open-chromatin assay nearly reach the quality of several TF ChIP-seq data sets. Finally, these scores correctly predict known transcriptional regulators as illustrated by the application to novel DNaseI-seq and NOMe-seq data for primary human hepatocytes and CD4+ T-cells, respectively.

  14. Combining transcription factor binding affinities with open-chromatin data for accurate gene expression prediction

    PubMed Central

    Schmidt, Florian; Gasparoni, Nina; Gasparoni, Gilles; Gianmoena, Kathrin; Cadenas, Cristina; Polansky, Julia K.; Ebert, Peter; Nordström, Karl; Barann, Matthias; Sinha, Anupam; Fröhler, Sebastian; Xiong, Jieyi; Dehghani Amirabad, Azim; Behjati Ardakani, Fatemeh; Hutter, Barbara; Zipprich, Gideon; Felder, Bärbel; Eils, Jürgen; Brors, Benedikt; Chen, Wei; Hengstler, Jan G.; Hamann, Alf; Lengauer, Thomas; Rosenstiel, Philip; Walter, Jörn; Schulz, Marcel H.

    2017-01-01

    The binding and contribution of transcription factors (TF) to cell specific gene expression is often deduced from open-chromatin measurements to avoid costly TF ChIP-seq assays. Thus, it is important to develop computational methods for accurate TF binding prediction in open-chromatin regions (OCRs). Here, we report a novel segmentation-based method, TEPIC, to predict TF binding by combining sets of OCRs with position weight matrices. TEPIC can be applied to various open-chromatin data, e.g. DNaseI-seq and NOMe-seq. Additionally, Histone-Marks (HMs) can be used to identify candidate TF binding sites. TEPIC computes TF affinities and uses open-chromatin/HM signal intensity as quantitative measures of TF binding strength. Using machine learning, we find low affinity binding sites to improve our ability to explain gene expression variability compared to the standard presence/absence classification of binding sites. Further, we show that both footprints and peaks capture essential TF binding events and lead to a good prediction performance. In our application, gene-based scores computed by TEPIC with one open-chromatin assay nearly reach the quality of several TF ChIP-seq data sets. Finally, these scores correctly predict known transcriptional regulators as illustrated by the application to novel DNaseI-seq and NOMe-seq data for primary human hepatocytes and CD4+ T-cells, respectively. PMID:27899623

  15. AtRTD - a comprehensive reference transcript dataset resource for accurate quantification of transcript-specific expression in Arabidopsis thaliana.

    PubMed

    Zhang, Runxuan; Calixto, Cristiane P G; Tzioutziou, Nikoleta A; James, Allan B; Simpson, Craig G; Guo, Wenbin; Marquez, Yamile; Kalyna, Maria; Patro, Rob; Eyras, Eduardo; Barta, Andrea; Nimmo, Hugh G; Brown, John W S

    2015-10-01

    RNA-sequencing (RNA-seq) allows global gene expression analysis at the individual transcript level. Accurate quantification of transcript variants generated by alternative splicing (AS) remains a challenge. We have developed a comprehensive, nonredundant Arabidopsis reference transcript dataset (AtRTD) containing over 74 000 transcripts for use with algorithms to quantify AS transcript isoforms in RNA-seq. The AtRTD was formed by merging transcripts from TAIR10 and novel transcripts identified in an AS discovery project. We have estimated transcript abundance in RNA-seq data using the transcriptome-based alignment-free programmes Sailfish and Salmon and have validated quantification of splicing ratios from RNA-seq by high resolution reverse transcription polymerase chain reaction (HR RT-PCR). Good correlations between splicing ratios from RNA-seq and HR RT-PCR were obtained demonstrating the accuracy of abundances calculated for individual transcripts in RNA-seq. The AtRTD is a resource that will have immediate utility in analysing Arabidopsis RNA-seq data to quantify differential transcript abundance and expression.

  16. A General Pairwise Interaction Model Provides an Accurate Description of In Vivo Transcription Factor Binding Sites

    PubMed Central

    Santolini, Marc; Mora, Thierry; Hakim, Vincent

    2014-01-01

    The identification of transcription factor binding sites (TFBSs) on genomic DNA is of crucial importance for understanding and predicting regulatory elements in gene networks. TFBS motifs are commonly described by Position Weight Matrices (PWMs), in which each DNA base pair contributes independently to the transcription factor (TF) binding. However, this description ignores correlations between nucleotides at different positions, and is generally inaccurate: analysing fly and mouse in vivo ChIPseq data, we show that in most cases the PWM model fails to reproduce the observed statistics of TFBSs. To overcome this issue, we introduce the pairwise interaction model (PIM), a generalization of the PWM model. The model is based on the principle of maximum entropy and explicitly describes pairwise correlations between nucleotides at different positions, while being otherwise as unconstrained as possible. It is mathematically equivalent to considering a TF-DNA binding energy that depends additively on each nucleotide identity at all positions in the TFBS, like the PWM model, but also additively on pairs of nucleotides. We find that the PIM significantly improves over the PWM model, and even provides an optimal description of TFBS statistics within statistical noise. The PIM generalizes previous approaches to interdependent positions: it accounts for co-variation of two or more base pairs, and predicts secondary motifs, while outperforming multiple-motif models consisting of mixtures of PWMs. We analyse the structure of pairwise interactions between nucleotides, and find that they are sparse and dominantly located between consecutive base pairs in the flanking region of TFBS. Nonetheless, interactions between pairs of non-consecutive nucleotides are found to play a significant role in the obtained accurate description of TFBS statistics. The PIM is computationally tractable, and provides a general framework that should be useful for describing and predicting TFBSs beyond

  17. Identification and validation of reference genes for transcript normalization in strawberry (Fragaria × ananassa) defense responses.

    PubMed

    Amil-Ruiz, Francisco; Garrido-Gala, José; Blanco-Portales, Rosario; Folta, Kevin M; Muñoz-Blanco, Juan; Caballero, José L

    2013-01-01

    Strawberry (Fragaria spp) is an emerging model for the development of basic genomics and recombinant DNA studies among rosaceous crops. Functional genomic and molecular studies involve relative quantification of gene expression under experimental conditions of interest. Accuracy and reliability are dependent upon the choice of an optimal reference control transcript. There is no information available on validated endogenous reference genes for use in studies testing strawberry-pathogen interactions. Thirteen potential pre-selected strawberry reference genes were tested against different tissues, strawberry cultivars, biotic stresses, ripening and senescent conditions, and SA/JA treatments. Evaluation of reference candidate's suitability was analyzed by five different methodologies, and information was merged to identify best reference transcripts. A combination of all five methods was used for selective classification of reference genes. The resulting superior reference genes, FaRIB413, FaACTIN, FaEF1α and FaGAPDH2 are strongly recommended as control genes for relative quantification of gene expression in strawberry. This report constitutes the first systematic study to identify and validate optimal reference genes for accurate normalization of gene expression in strawberry plant defense response studies.

  18. Identification and Validation of Reference Genes for Transcript Normalization in Strawberry (Fragaria × ananassa) Defense Responses

    PubMed Central

    Amil-Ruiz, Francisco; Garrido-Gala, José; Blanco-Portales, Rosario; Folta, Kevin M.; Muñoz-Blanco, Juan; Caballero, José L.

    2013-01-01

    Strawberry (Fragaria spp) is an emerging model for the development of basic genomics and recombinant DNA studies among rosaceous crops. Functional genomic and molecular studies involve relative quantification of gene expression under experimental conditions of interest. Accuracy and reliability are dependent upon the choice of an optimal reference control transcript. There is no information available on validated endogenous reference genes for use in studies testing strawberry-pathogen interactions. Thirteen potential pre-selected strawberry reference genes were tested against different tissues, strawberry cultivars, biotic stresses, ripening and senescent conditions, and SA/JA treatments. Evaluation of reference candidate’s suitability was analyzed by five different methodologies, and information was merged to identify best reference transcripts. A combination of all five methods was used for selective classification of reference genes. The resulting superior reference genes, FaRIB413, FaACTIN, FaEF1α and FaGAPDH2 are strongly recommended as control genes for relative quantification of gene expression in strawberry. This report constitutes the first systematic study to identify and validate optimal reference genes for accurate normalization of gene expression in strawberry plant defense response studies. PMID:23940602

  19. COSMOS: accurate detection of somatic structural variations through asymmetric comparison between tumor and normal samples.

    PubMed

    Yamagata, Koichi; Yamanishi, Ayako; Kokubu, Chikara; Takeda, Junji; Sese, Jun

    2016-05-05

    An important challenge in cancer genomics is precise detection of structural variations (SVs) by high-throughput short-read sequencing, which is hampered by the high false discovery rates of existing analysis tools. Here, we propose an accurate SV detection method named COSMOS, which compares the statistics of the mapped read pairs in tumor samples with isogenic normal control samples in a distinct asymmetric manner. COSMOS also prioritizes the candidate SVs using strand-specific read-depth information. Performance tests on modeled tumor genomes revealed that COSMOS outperformed existing methods in terms of F-measure. We also applied COSMOS to an experimental mouse cell-based model, in which SVs were induced by genome engineering and gamma-ray irradiation, followed by polymerase chain reaction-based confirmation. The precision of COSMOS was 84.5%, while the next best existing method was 70.4%. Moreover, the sensitivity of COSMOS was the highest, indicating that COSMOS has great potential for cancer genome analysis.

  20. COSMOS: accurate detection of somatic structural variations through asymmetric comparison between tumor and normal samples

    PubMed Central

    Yamagata, Koichi; Yamanishi, Ayako; Kokubu, Chikara; Takeda, Junji; Sese, Jun

    2016-01-01

    An important challenge in cancer genomics is precise detection of structural variations (SVs) by high-throughput short-read sequencing, which is hampered by the high false discovery rates of existing analysis tools. Here, we propose an accurate SV detection method named COSMOS, which compares the statistics of the mapped read pairs in tumor samples with isogenic normal control samples in a distinct asymmetric manner. COSMOS also prioritizes the candidate SVs using strand-specific read-depth information. Performance tests on modeled tumor genomes revealed that COSMOS outperformed existing methods in terms of F-measure. We also applied COSMOS to an experimental mouse cell-based model, in which SVs were induced by genome engineering and gamma-ray irradiation, followed by polymerase chain reaction-based confirmation. The precision of COSMOS was 84.5%, while the next best existing method was 70.4%. Moreover, the sensitivity of COSMOS was the highest, indicating that COSMOS has great potential for cancer genome analysis. PMID:26833260

  1. Read-through transcripts in normal human lung parenchyma are down-regulated in lung adenocarcinoma

    PubMed Central

    Cotroneo, Chiara E.; Galvan, Antonella; Noci, Sara; Piazza, Rocco; Pirola, Alessandra; Spinelli, Roberta; Incarbone, Matteo; Palleschi, Alessandro; Rosso, Lorenzo; Santambrogio, Luigi; Dragani, Tommaso A.; Colombo, Francesca

    2016-01-01

    Read-through transcripts result from the continuous transcription of adjacent, similarly oriented genes, with the splicing out of the intergenic region. They have been found in several neoplastic and normal tissues, but their pathophysiological significance is unclear. We used high-throughput sequencing of cDNA fragments (RNA-Seq) to identify read-through transcripts in the non-involved lung tissue of 64 surgically treated lung adenocarcinoma patients. A total of 52 distinct read-through species was identified, with 24 patients having at least one read-through event, up to a maximum of 17 such transcripts in one patient. Sanger sequencing validated 28 of these transcripts and identified an additional 15, for a total of 43 distinct read-through events involving 35 gene pairs. Expression levels of 10 validated read-through transcripts were measured by quantitative PCR in pairs of matched non-involved lung tissue and lung adenocarcinoma tissue from 45 patients. Higher expression levels were observed in normal lung tissue than in the tumor counterpart, with median relative quantification ratios between normal and tumor varying from 1.90 to 7.78; the difference was statistically significant (P < 0.001, Wilcoxon's signed-rank test for paired samples) for eight transcripts: ELAVL1–TIMM44, FAM162B–ZUFSP, IFNAR2–IL10RB, INMT–FAM188B, KIAA1841–C2orf74, NFATC3–PLA2G15, SIRPB1–SIRPD, and SHANK3–ACR. This report documents the presence of read-through transcripts in apparently normal lung tissue, with inter-individual differences in patterns and abundance. It also shows their down-regulation in tumors, suggesting that these chimeric transcripts may function as tumor suppressors in lung tissue. PMID:27058892

  2. Signaling Proteins and Transcription Factors in Normal and Malignant Early B Cell Development

    PubMed Central

    Pérez-Vera, Patricia; Reyes-León, Adriana; Fuentes-Pananá, Ezequiel M.

    2011-01-01

    B cell development starts in bone marrow with the commitment of hematopoietic progenitors to the B cell lineage. In murine models, the IL-7 and preBCR receptors, and the signaling pathways and transcription factors that they regulate, control commitment and maintenance along the B cell pathway. E2A, EBF1, PAX5, and Ikaros are among the most important transcription factors controlling early development and thereby conditioning mice homeostatic B cell lymphopoiesis. Importantly, their gain or loss of function often results in malignant development in humans, supporting conserved roles for these transcription factors. B cell acute lymphoblastic leukemia is the most common cause of pediatric cancer, and it is characterized by unpaired early B cell development resulting from genetic lesions in these critical signaling pathways and transcription factors. Fine mapping of these genetic abnormalities is allowing more specific treatments, more accurately predicting risk profiles for this disease, and improving survival rates. PMID:22046564

  3. Signaling proteins and transcription factors in normal and malignant early B cell development.

    PubMed

    Pérez-Vera, Patricia; Reyes-León, Adriana; Fuentes-Pananá, Ezequiel M

    2011-01-01

    B cell development starts in bone marrow with the commitment of hematopoietic progenitors to the B cell lineage. In murine models, the IL-7 and preBCR receptors, and the signaling pathways and transcription factors that they regulate, control commitment and maintenance along the B cell pathway. E2A, EBF1, PAX5, and Ikaros are among the most important transcription factors controlling early development and thereby conditioning mice homeostatic B cell lymphopoiesis. Importantly, their gain or loss of function often results in malignant development in humans, supporting conserved roles for these transcription factors. B cell acute lymphoblastic leukemia is the most common cause of pediatric cancer, and it is characterized by unpaired early B cell development resulting from genetic lesions in these critical signaling pathways and transcription factors. Fine mapping of these genetic abnormalities is allowing more specific treatments, more accurately predicting risk profiles for this disease, and improving survival rates.

  4. Occupancy by key transcription factors is a more accurate predictor of enhancer activity than histone modifications or chromatin accessibility

    SciTech Connect

    Dogan, Nergiz; Wu, Weisheng; Morrissey, Christapher S.; Chen, Kuan-Bei; Stonestrom, Aaron; Long, Maria; Keller, Cheryl A.; Cheng, Yong; Jain, Deepti; Visel, Axel; Pennacchio, Len A.; Weiss, Mitchell J.; Blobel, Gerd A.; Hardison, Ross C.

    2015-04-23

    Regulated gene expression controls organismal development, and variation in regulatory patterns has been implicated in complex traits. Thus accurate prediction of enhancers is important for further understanding of these processes. Genome-wide measurement of epigenetic features, such as histone modifications and occupancy by transcription factors, is improving enhancer predictions, but the contribution of these features to prediction accuracy is not known. Given the importance of the hematopoietic transcription factor TAL1 for erythroid gene activation, we predicted candidate enhancers based on genomic occupancy by TAL1 and measured their activity. Contributions of multiple features to enhancer prediction were evaluated based on the results of these and other studies. Results: TAL1-bound DNA segments were active enhancers at a high rate both in transient transfections of cultured cells (39 of 79, or 56%) and transgenic mice (43 of 66, or 65%). The level of binding signal for TAL1 or GATA1 did not help distinguish TAL1-bound DNA segments as active versus inactive enhancers, nor did the density of regulation-related histone modifications. A meta-analysis of results from this and other studies (273 tested predicted enhancers) showed that the presence of TAL1, GATA1, EP300, SMAD1, H3K4 methylation, H3K27ac, and CAGE tags at DNase hypersensitive sites gave the most accurate predictors of enhancer activity, with a success rate over 80% and a median threefold increase in activity. Chromatin accessibility assays and the histone modifications H3K4me1 and H3K27ac were sensitive for finding enhancers, but they have high false positive rates unless transcription factor occupancy is also included. Conclusions: Occupancy by key transcription factors such as TAL1, GATA1, SMAD1, and EP300, along with evidence of transcription, improves the accuracy of enhancer predictions based on epigenetic features.

  5. Occupancy by key transcription factors is a more accurate predictor of enhancer activity than histone modifications or chromatin accessibility

    DOE PAGES

    Dogan, Nergiz; Wu, Weisheng; Morrissey, Christapher S.; ...

    2015-04-23

    Regulated gene expression controls organismal development, and variation in regulatory patterns has been implicated in complex traits. Thus accurate prediction of enhancers is important for further understanding of these processes. Genome-wide measurement of epigenetic features, such as histone modifications and occupancy by transcription factors, is improving enhancer predictions, but the contribution of these features to prediction accuracy is not known. Given the importance of the hematopoietic transcription factor TAL1 for erythroid gene activation, we predicted candidate enhancers based on genomic occupancy by TAL1 and measured their activity. Contributions of multiple features to enhancer prediction were evaluated based on the resultsmore » of these and other studies. Results: TAL1-bound DNA segments were active enhancers at a high rate both in transient transfections of cultured cells (39 of 79, or 56%) and transgenic mice (43 of 66, or 65%). The level of binding signal for TAL1 or GATA1 did not help distinguish TAL1-bound DNA segments as active versus inactive enhancers, nor did the density of regulation-related histone modifications. A meta-analysis of results from this and other studies (273 tested predicted enhancers) showed that the presence of TAL1, GATA1, EP300, SMAD1, H3K4 methylation, H3K27ac, and CAGE tags at DNase hypersensitive sites gave the most accurate predictors of enhancer activity, with a success rate over 80% and a median threefold increase in activity. Chromatin accessibility assays and the histone modifications H3K4me1 and H3K27ac were sensitive for finding enhancers, but they have high false positive rates unless transcription factor occupancy is also included. Conclusions: Occupancy by key transcription factors such as TAL1, GATA1, SMAD1, and EP300, along with evidence of transcription, improves the accuracy of enhancer predictions based on epigenetic features.« less

  6. Infrequently transcribed long genes depend on the Set2/Rpd3S pathway for accurate transcription

    PubMed Central

    Li, Bing; Gogol, Madelaine; Carey, Mike; Pattenden, Samantha G.; Seidel, Chris; Workman, Jerry L.

    2007-01-01

    The presence of Set2-mediated methylation of H3K36 (K36me) correlates with transcription frequency throughout the yeast genome. K36me targets the Rpd3S complex to deacetylate transcribed regions and suppress cryptic transcription initiation at certain genes. Here, using a genome-wide approach, we report that the Set2–Rpd3S pathway is generally required for controlling acetylation at coding regions. When using acetylation as a functional readout for this pathway, we discovered that longer genes and, surprisingly, genes transcribed at lower frequency exhibit a stronger dependency. Moreover, a systematic screen using high-resolution tiling microarrays allowed us to identify a group of genes that rely on Set2–Rpd3S to suppress spurious transcripts. Interestingly, most of these genes are within the group that depend on the same pathway to maintain a hypoacetylated state at coding regions. These data highlight the importance of using the functional readout of histone codes to define the roles of specific pathways. PMID:17545470

  7. Accurate elevation and normal moveout corrections of seismic reflection data on rugged topography

    USGS Publications Warehouse

    Liu, J.; Xia, J.; Chen, C.; Zhang, G.

    2005-01-01

    The application of the seismic reflection method is often limited in areas of complex terrain. The problem is the incorrect correction of time shifts caused by topography. To apply normal moveout (NMO) correction to reflection data correctly, static corrections are necessary to be applied in advance for the compensation of the time distortions of topography and the time delays from near-surface weathered layers. For environment and engineering investigation, weathered layers are our targets, so that the static correction mainly serves the adjustment of time shifts due to an undulating surface. In practice, seismic reflected raypaths are assumed to be almost vertical through the near-surface layers because they have much lower velocities than layers below. This assumption is acceptable in most cases since it results in little residual error for small elevation changes and small offsets in reflection events. Although static algorithms based on choosing a floating datum related to common midpoint gathers or residual surface-consistent functions are available and effective, errors caused by the assumption of vertical raypaths often generate pseudo-indications of structures. This paper presents the comparison of applying corrections based on the vertical raypaths and bias (non-vertical) raypaths. It also provides an approach of combining elevation and NMO corrections. The advantages of the approach are demonstrated by synthetic and real-world examples of multi-coverage seismic reflection surveys on rough topography. ?? The Royal Society of New Zealand 2005.

  8. Accurate Accumulation of Dose for Improved Understanding of Radiation Effects in Normal Tissue

    SciTech Connect

    Jaffray, David A.; Lindsay, Patricia E.; Brock, Kristy K.; Deasy, Joseph O.; Tome, W.A.

    2010-03-01

    The actual distribution of radiation dose accumulated in normal tissues over the complete course of radiation therapy is, in general, poorly quantified. Differences in the patient anatomy between planning and treatment can occur gradually (e.g., tumor regression, resolution of edema) or relatively rapidly (e.g., bladder filling, breathing motion) and these undermine the accuracy of the planned dose distribution. Current efforts to maximize the therapeutic ratio require models that relate the true accumulated dose to clinical outcome. The needed accuracy can only be achieved through the development of robust methods that track the accumulation of dose within the various tissues in the body. Specific needs include the development of segmentation methods, tissue-mapping algorithms, uncertainty estimation, optimal schedules for image-based monitoring, and the development of informatics tools to support subsequent analysis. These developments will not only improve radiation outcomes modeling but will address the technical demands of the adaptive radiotherapy paradigm. The next 5 years need to see academia and industry bring these tools into the hands of the clinician and the clinical scientist.

  9. Accurate Proteome-wide Label-free Quantification by Delayed Normalization and Maximal Peptide Ratio Extraction, Termed MaxLFQ *

    PubMed Central

    Cox, Jürgen; Hein, Marco Y.; Luber, Christian A.; Paron, Igor; Nagaraj, Nagarjuna; Mann, Matthias

    2014-01-01

    Protein quantification without isotopic labels has been a long-standing interest in the proteomics field. However, accurate and robust proteome-wide quantification with label-free approaches remains a challenge. We developed a new intensity determination and normalization procedure called MaxLFQ that is fully compatible with any peptide or protein separation prior to LC-MS analysis. Protein abundance profiles are assembled using the maximum possible information from MS signals, given that the presence of quantifiable peptides varies from sample to sample. For a benchmark dataset with two proteomes mixed at known ratios, we accurately detected the mixing ratio over the entire protein expression range, with greater precision for abundant proteins. The significance of individual label-free quantifications was obtained via a t test approach. For a second benchmark dataset, we accurately quantify fold changes over several orders of magnitude, a task that is challenging with label-based methods. MaxLFQ is a generic label-free quantification technology that is readily applicable to many biological questions; it is compatible with standard statistical analysis workflows, and it has been validated in many and diverse biological projects. Our algorithms can handle very large experiments of 500+ samples in a manageable computing time. It is implemented in the freely available MaxQuant computational proteomics platform and works completely seamlessly at the click of a button. PMID:24942700

  10. Normal dendrite growth in Drosophila motor neurons requires the AP-1 transcription factor.

    PubMed

    Hartwig, Cortnie L; Worrell, Jason; Levine, Richard B; Ramaswami, Mani; Sanyal, Subhabrata

    2008-09-01

    During learning and memory formation, information flow through networks is regulated significantly through structural alterations in neurons. Dendrites, sites of signal integration, are key targets of activity-mediated modifications. Although local mechanisms of dendritic growth ensure synapse-specific changes, global mechanisms linking neural activity to nuclear gene expression may have profound influences on neural function. Fos, being an immediate-early gene, is ideally suited to be an initial transducer of neural activity, but a precise role for the AP-1 transcription factor in dendrite growth remains to be elucidated. Here we measure changes in the dendritic fields of identified Drosophila motor neurons in vivo and in primary culture to investigate the role of the immediate-early transcription factor AP-1 in regulating endogenous and activity-induced dendrite growth. Our data indicate that (a) increased neural excitability or depolarization stimulates dendrite growth, (b) AP-1 (a Fos, Jun hetero-dimer) is required for normal motor neuron dendritic growth during development and in response to activity induction, and (c) neuronal Fos protein levels are rapidly but transiently induced in motor neurons following neural activity. Taken together, these results show that AP-1 mediated transcription is important for dendrite growth, and that neural activity influences global dendritic growth through a gene-expression dependent mechanism gated by AP-1.

  11. Human BLCAP transcript: new editing events in normal and cancerous tissues.

    PubMed

    Galeano, Federica; Leroy, Anne; Rossetti, Claudia; Gromova, Irina; Gautier, Philippe; Keegan, Liam P; Massimi, Luca; Di Rocco, Concezio; O'Connell, Mary A; Gallo, Angela

    2010-07-01

    Bladder cancer-associated protein (BLCAP) is a highly conserved protein among species, and it is considered a novel candidate tumor suppressor gene originally identified from human bladder carcinoma. However, little is known about the regulation or the function of this protein. Here, we show that the human BLCAP transcript undergoes multiple A-to-I editing events. Some of the new editing events alter the highly conserved amino terminus of the protein creating alternative protein isoforms by changing the genetically coded amino acids. We found that both ADAR1 and ADAR2-editing enzymes cooperate to edit this transcript and that different tissues displayed distinctive ratios of edited and unedited BLCAP transcripts. Moreover, we observed a general decrease in BLCAP-editing level in astrocytomas, bladder cancer and colorectal cancer when compared with the related normal tissues. The newly identified editing events, found to be downregulated in cancers, could be useful for future studies as a diagnostic tool to distinguish malignancies or epigenetic changes in different tumors.

  12. MGSA/GRO transcription is differentially regulated in normal retinal pigment epithelial and melanoma cells.

    PubMed Central

    Shattuck, R L; Wood, L D; Jaffe, G J; Richmond, A

    1994-01-01

    We have characterized constitutive and cytokine-regulated MGSA/GRO alpha, -beta, and -gamma gene expression in normal retinal pigment epithelial (RPE) cells and a malignant melanoma cell line (Hs294T) to discern the mechanism for MGSA/GRO constitutive expression in melanoma. In RPE cells, constitutive MGSA/GRO alpha, -beta, and -gamma mRNAs are not detected by Northern (RNA) blot analysis although nuclear runoff experiments show that all three genes are transcribed. In Hs294T cells, constitutive MGSA/GRO alpha expression is detectable by Northern blot analysis, and the level of basal MGSA/GRO alpha transcription is 8- to 30-fold higher than in RPE cells. In contrast, in Hs294T cells, basal MGSA/GRO beta and -gamma transcription is only twofold higher than in RPE cells and no beta or gamma mRNA is detected by Northern blot. These data suggest that the constitutive MGSA/GRO alpha mRNA in Hs294T cells is due to increased basal MGSA/GRO alpha gene transcription. The cytokines interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF alpha) significantly increase the mRNA levels for all three MGSA/GRO isoforms in Hs294T and RPE cells, and both transcriptional and posttranscriptional mechanisms are operational. Nuclear runoff assays indicate that in RPE cells, a 1-h IL-1 treatment induces a 10- to 20-fold increase in transcription of MGSA/GRO alpha, -beta and -gamma but only a 2-fold increase in Hs294T cells. Similarly, chloramphenicol acetyltransferase (CAT) reporter gene analysis using the MGSA/GRO alpha, -beta, and -gamma promoter regions demonstrates that IL-1 treatment induces an 8- to 14-fold increase in CAT activity in RPE cells but only a 2-fold increase in Hs294T cells. The effect of deletion or mutation of the MGSA/GRO alpha NF-kappa B element, combined with data from gel mobility shift analyses, indicates that the NF-kappa B p50/p65 heterodimer in RPE cells plays an important role in IL-1- and TNF alpha-enhanced gene transcription. In Hs294T cells, gel shift

  13. Differential Transcriptional Response in Macrophages Infected with Cell Wall Deficient versus Normal Mycobacterium Tuberculosis

    PubMed Central

    Fu, Yu-Rong; Gao, Kun-Shan; Ji, Rui; Yi, Zheng-Jun

    2015-01-01

    Host-pathogen interactions determine the outcome following infection by mycobacterium tuberculosis (Mtb). Under adverse circumstances, normal Mtb can form cell-wall deficient (CWD) variants within macrophages, which have been considered an adaptive strategy for facilitating bacterial survival inside macrophages. However, the molecular mechanism by which infection of macrophages with different phenotypic Mtb elicits distinct responses of macrophages is not fully understood. To explore the molecular events triggered upon Mtb infection of macrophages, differential transcriptional responses of RAW264.7 cells infected with two forms of Mtb, CWD-Mtb and normal Mtb, were studied by microarray analysis. Some of the differentially regulated genes were confirmed by RT-qPCR in both RAW264.7 cells and primary macrophages. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway was used to analyze functions of differentially expressed genes. Distinct gene expression patterns were observed between CWD-Mtb and normal Mtb group. Mapt was up-regulated, while NOS2 and IL-11 were down-regulated in CWD-Mtb infected RAW264.7 cells and primary macrophages compared with normal Mtb infected ones. Many deregulated genes were found to be related to macrophages activation, immune response, phagosome maturation, autophagy and lipid metabolism. KEGG analysis showed that the differentially expressed genes were mainly involved in MAPK signaling pathway, nitrogen metabolism, cytokine-cytokine receptor interaction and focal adhesion. Taken together, the present study showed that differential macrophage responses were induced by intracellular CWD-Mtb an normal Mtb infection, which suggested that interactions between macrophages and different phenotypic Mtb are very complex. The results provide evidence for further understanding of pathogenesis of CWD-Mtb and may help in improving strategies to eliminate intracellular CWD-Mtb. PMID:25552926

  14. Normal stroma suppresses cancer cell proliferation via mechanosensitive regulation of JMJD1a-mediated transcription

    PubMed Central

    Kaukonen, Riina; Mai, Anja; Georgiadou, Maria; Saari, Markku; De Franceschi, Nicola; Betz, Timo; Sihto, Harri; Ventelä, Sami; Elo, Laura; Jokitalo, Eija; Westermarck, Jukka; Kellokumpu-Lehtinen, Pirkko-Liisa; Joensuu, Heikki; Grenman, Reidar; Ivaska, Johanna

    2016-01-01

    Tissue homeostasis is dependent on the controlled localization of specific cell types and the correct composition of the extracellular stroma. While the role of the cancer stroma in tumour progression has been well characterized, the specific contribution of the matrix itself is unknown. Furthermore, the mechanisms enabling normal—not cancer—stroma to provide tumour-suppressive signals and act as an antitumorigenic barrier are poorly understood. Here we show that extracellular matrix (ECM) generated by normal fibroblasts (NFs) is softer than the CAF matrix, and its physical and structural features regulate cancer cell proliferation. We find that normal ECM triggers downregulation and nuclear exit of the histone demethylase JMJD1a resulting in the epigenetic growth restriction of carcinoma cells. Interestingly, JMJD1a positively regulates transcription of many target genes, including YAP/TAZ (WWTR1), and therefore gene expression in a stiffness-dependent manner. Thus, normal stromal restricts cancer cell proliferation through JMJD1a-dependent modulation of gene expression. PMID:27488962

  15. Capping of vesicular stomatitis virus pre-mRNA is required for accurate selection of transcription stop-start sites and virus propagation.

    PubMed

    Ogino, Tomoaki

    2014-10-29

    The multifunctional RNA-dependent RNA polymerase L protein of vesicular stomatitis virus catalyzes unconventional pre-mRNA capping via the covalent enzyme-pRNA intermediate formation, which requires the histidine-arginine (HR) motif in the polyribonucleotidyltransferase domain. Here, the effects of cap-defective mutations in the HR motif on transcription were analyzed using an in vitro reconstituted transcription system. The wild-type L protein synthesized the leader RNA from the 3'-end of the genome followed by 5'-capped and 3'-polyadenylated mRNAs from internal genes by a stop-start transcription mechanism. Cap-defective mutants efficiently produced the leader RNA, but displayed aberrant stop-start transcription using cryptic termination and initiation signals within the first gene, resulting in sequential generation of ∼40-nucleotide transcripts with 5'-ATP from a correct mRNA-start site followed by a 28-nucleotide transcript and long 3'-polyadenylated transcript initiated with non-canonical GTP from atypical start sites. Frequent transcription termination and re-initiation within the first gene significantly attenuated the production of downstream mRNAs. Consistent with the inability of these mutants in in vitro mRNA synthesis and capping, these mutations were lethal to virus replication in cultured cells. These findings indicate that viral mRNA capping is required for accurate stop-start transcription as well as mRNA stability and translation and, therefore, for virus replication in host cells.

  16. Validation of reference genes for accurate normalization of gene expression for real time-quantitative PCR in strawberry fruits using different cultivars and osmotic stresses.

    PubMed

    Galli, Vanessa; Borowski, Joyce Moura; Perin, Ellen Cristina; Messias, Rafael da Silva; Labonde, Julia; Pereira, Ivan dos Santos; Silva, Sérgio Delmar Dos Anjos; Rombaldi, Cesar Valmor

    2015-01-10

    The increasing demand of strawberry (Fragaria×ananassa Duch) fruits is associated mainly with their sensorial characteristics and the content of antioxidant compounds. Nevertheless, the strawberry production has been hampered due to its sensitivity to abiotic stresses. Therefore, to understand the molecular mechanisms highlighting stress response is of great importance to enable genetic engineering approaches aiming to improve strawberry tolerance. However, the study of expression of genes in strawberry requires the use of suitable reference genes. In the present study, seven traditional and novel candidate reference genes were evaluated for transcript normalization in fruits of ten strawberry cultivars and two abiotic stresses, using RefFinder, which integrates the four major currently available software programs: geNorm, NormFinder, BestKeeper and the comparative delta-Ct method. The results indicate that the expression stability is dependent on the experimental conditions. The candidate reference gene DBP (DNA binding protein) was considered the most suitable to normalize expression data in samples of strawberry cultivars and under drought stress condition, and the candidate reference gene HISTH4 (histone H4) was the most stable under osmotic stresses and salt stress. The traditional genes GAPDH (glyceraldehyde-3-phosphate dehydrogenase) and 18S (18S ribosomal RNA) were considered the most unstable genes in all conditions. The expression of phenylalanine ammonia lyase (PAL) and 9-cis epoxycarotenoid dioxygenase (NCED1) genes were used to further confirm the validated candidate reference genes, showing that the use of an inappropriate reference gene may induce erroneous results. This study is the first survey on the stability of reference genes in strawberry cultivars and osmotic stresses and provides guidelines to obtain more accurate RT-qPCR results for future breeding efforts.

  17. Fractionation and reconstitution of factors required for accurate transcription of mammalian ribosomal RNA genes: identification of a species-dependent initiation factor.

    PubMed Central

    Mishima, Y; Financsek, I; Kominami, R; Muramatsu, M

    1982-01-01

    Mouse and human cell extracts (S100) can support an accurate and efficient transcription initiation on homologous ribosomal RNA gene (rDNA) templates. The cell extracts were fractionated with the aid of a phosphocellulose column into four fractions (termed A, B, C and D), including one containing a major part of the RNA polymerase I activity. Various reconstitution experiments indicate that fraction D is an absolute requirement for the correct and efficient transcription initiation by RNA polymerase I on both mouse and human genes. Fraction B effectively suppresses random initiation on these templates. Fraction A appears to further enhance the transcription which takes place with fractions C and D. Although fractions A, B and C are interchangeable between mouse and human extracts, fraction D is not; i.e. initiation of transcription required the presence of a homologous fraction D for both templates. The factor(s) in fraction D, however, is not literally species-specific, since mouse D fraction is capable of supporting accurate transcription initiation on a rat rDNA template in the presence of all the other fractions from human cell extract under the conditions where human D fraction is unable to support it. We conclude from these experiments that a species-dependent factor in fraction D plays an important role in the initiation of rDNA transcription in each animal species. Images PMID:7177852

  18. Identification and evaluation of new reference genes in Gossypium hirsutum for accurate normalization of real-time quantitative RT-PCR data

    PubMed Central

    2010-01-01

    Background Normalizing through reference genes, or housekeeping genes, can make more accurate and reliable results from reverse transcription real-time quantitative polymerase chain reaction (qPCR). Recent studies have shown that no single housekeeping gene is universal for all experiments. Thus, suitable reference genes should be the first step of any qPCR analysis. Only a few studies on the identification of housekeeping gene have been carried on plants. Therefore qPCR studies on important crops such as cotton has been hampered by the lack of suitable reference genes. Results By the use of two distinct algorithms, implemented by geNorm and NormFinder, we have assessed the gene expression of nine candidate reference genes in cotton: GhACT4, GhEF1α5, GhFBX6, GhPP2A1, GhMZA, GhPTB, GhGAPC2, GhβTUB3 and GhUBQ14. The candidate reference genes were evaluated in 23 experimental samples consisting of six distinct plant organs, eight stages of flower development, four stages of fruit development and in flower verticils. The expression of GhPP2A1 and GhUBQ14 genes were the most stable across all samples and also when distinct plants organs are examined. GhACT4 and GhUBQ14 present more stable expression during flower development, GhACT4 and GhFBX6 in the floral verticils and GhMZA and GhPTB during fruit development. Our analysis provided the most suitable combination of reference genes for each experimental set tested as internal control for reliable qPCR data normalization. In addition, to illustrate the use of cotton reference genes we checked the expression of two cotton MADS-box genes in distinct plant and floral organs and also during flower development. Conclusion We have tested the expression stabilities of nine candidate genes in a set of 23 tissue samples from cotton plants divided into five different experimental sets. As a result of this evaluation, we recommend the use of GhUBQ14 and GhPP2A1 housekeeping genes as superior references for normalization of gene

  19. Selection of reference genes for quantitative reverse-transcription polymerase chain reaction normalization in Brassica napus under various stress conditions.

    PubMed

    Wang, Zheng; Chen, Yu; Fang, Hedi; Shi, Haifeng; Chen, Keping; Zhang, Zhiyan; Tan, Xiaoli

    2014-10-01

    Data normalization is essential for reliable output of quantitative real-time reverse-transcription polymerase chain reaction (qRT-PCR) assays, as the unsuitable choice of reference gene(s), whose expression might be influenced by exogenous treatments in plant tissues, could cause misinterpretation of results. To date, no systematic studies on reference genes have been performed in stressed Brassica napus. In this study, we investigated the expression variations of nine candidate reference genes in 40 samples of B. napus leaves subjected to various exogenous treatments. Parallel analyses by geNorm and NormFinder revealed that optimal reference genes differed across the different sets of samples. The best-ranked reference genes were PP2A and TIP41 for salt stress, TIP41 and ACT7 for heavy metal (Cr(6+)) stress, PP2A and UBC21 for drought stress, F-box and SAND for cold stress, F-box and ZNF for salicylic acid stress, TIP41, ACT7, and PP2A for methyl jasmonate stress, TIP41 and ACT7 for abscisic acid stress, and TIP41, UBC21, and PP2A for Sclerotinia sclerotiorum stress. Two newly employed reference genes, TIP41 and PP2A, showed better performances, suggesting their suitability in multiple conditions. To further validate the suitability of the reference genes, the expression patterns of BnWRKY40 and BnMKS1 were studied in parallel. This study is the first systematic analysis of reference gene selection for qRT-PCR normalization in B. napus, an agriculturally important crop, under different stress conditions. The results will contribute toward more accurate and widespread use of qRT-PCR in gene analysis of the genus Brassica.

  20. Transcription factor regulation can be accurately predicted from the presence of target gene signatures in microarray gene expression data

    PubMed Central

    Essaghir, Ahmed; Toffalini, Federica; Knoops, Laurent; Kallin, Anders; van Helden, Jacques; Demoulin, Jean-Baptiste

    2010-01-01

    Deciphering transcription factor networks from microarray data remains difficult. This study presents a simple method to infer the regulation of transcription factors from microarray data based on well-characterized target genes. We generated a catalog containing transcription factors associated with 2720 target genes and 6401 experimentally validated regulations. When it was available, a distinction between transcriptional activation and inhibition was included for each regulation. Next, we built a tool (www.tfacts.org) that compares submitted gene lists with target genes in the catalog to detect regulated transcription factors. TFactS was validated with published lists of regulated genes in various models and compared to tools based on in silico promoter analysis. We next analyzed the NCI60 cancer microarray data set and showed the regulation of SOX10, MITF and JUN in melanomas. We then performed microarray experiments comparing gene expression response of human fibroblasts stimulated by different growth factors. TFactS predicted the specific activation of Signal transducer and activator of transcription factors by PDGF-BB, which was confirmed experimentally. Our results show that the expression levels of transcription factor target genes constitute a robust signature for transcription factor regulation, and can be efficiently used for microarray data mining. PMID:20215436

  1. Reference genes selection for transcript normalization in kenaf (Hibiscus cannabinus L.) under salinity and drought stress.

    PubMed

    Niu, Xiaoping; Qi, Jianmin; Chen, Meixia; Zhang, Gaoyang; Tao, Aifen; Fang, Pingping; Xu, Jiantang; Onyedinma, Sandra A; Su, Jianguang

    2015-01-01

    Kenaf (Hibiscus cannabinus) is an economic and ecological fiber crop but suffers severe losses in fiber yield and quality under the stressful conditions of excess salinity and drought. To explore the mechanisms by which kenaf responds to excess salinity and drought, gene expression was performed at the transcriptomic level using RNA-seq. Thus, it is crucial to have a suitable set of reference genes to normalize target gene expression in kenaf under different conditions using real-time quantitative reverse transcription-PCR (qRT-PCR). In this study, we selected 10 candidate reference genes from the kenaf transcriptome and assessed their expression stabilities by qRT-PCR in 14 NaCl- and PEG-treated samples using geNorm, NormFinder, and BestKeeper. The results indicated that TUBα and 18S rRNA were the optimum reference genes under conditions of excess salinity and drought in kenaf. Moreover, TUBα and 18S rRNA were used singly or in combination as reference genes to validate the expression levels of WRKY28 and WRKY32 in NaCl- and PEG-treated samples by qRT-PCR. The results further proved the reliability of the two selected reference genes. This work will benefit future studies on gene expression and lead to a better understanding of responses to excess salinity and drought in kenaf.

  2. Reference genes selection for transcript normalization in kenaf (Hibiscus cannabinus L.) under salinity and drought stress

    PubMed Central

    Niu, Xiaoping; Chen, Meixia; Zhang, Gaoyang; Tao, Aifen; Fang, Pingping; Xu, Jiantang; Onyedinma, Sandra A.

    2015-01-01

    Kenaf (Hibiscus cannabinus) is an economic and ecological fiber crop but suffers severe losses in fiber yield and quality under the stressful conditions of excess salinity and drought. To explore the mechanisms by which kenaf responds to excess salinity and drought, gene expression was performed at the transcriptomic level using RNA-seq. Thus, it is crucial to have a suitable set of reference genes to normalize target gene expression in kenaf under different conditions using real-time quantitative reverse transcription-PCR (qRT-PCR). In this study, we selected 10 candidate reference genes from the kenaf transcriptome and assessed their expression stabilities by qRT-PCR in 14 NaCl- and PEG-treated samples using geNorm, NormFinder, and BestKeeper. The results indicated that TUBα and 18S rRNA were the optimum reference genes under conditions of excess salinity and drought in kenaf. Moreover, TUBα and 18S rRNA were used singly or in combination as reference genes to validate the expression levels of WRKY28 and WRKY32 in NaCl- and PEG-treated samples by qRT-PCR. The results further proved the reliability of the two selected reference genes. This work will benefit future studies on gene expression and lead to a better understanding of responses to excess salinity and drought in kenaf. PMID:26644967

  3. A simple analytical and experimental procedure for selection of reference genes for reverse-transcription quantitative PCR normalization data.

    PubMed

    Manjarin, R; Trottier, N L; Weber, P S; Liesman, J S; Taylor, N P; Steibel, J P

    2011-10-01

    Variation in cellular activity in a tissue induces changes in RNA concentration, which affects the validity of gene mRNA abundance analyzed by reverse transcription quantitative PCR (RT-qPCR). A common way of accounting for such variation consists of the use of reference genes for normalization. Programs such as geNorm may be used to select suitable reference genes, although a large set of genes that are not co-regulated must be analyzed to obtain accurate results. The objective of this study was to propose an alternative experimental and analytical protocol to assess the invariance of reference genes in porcine mammary tissue using mammary RNA and DNA concentrations as correction factors. Mammary glands were biopsied from 4 sows on d 110 of gestation (prepartum), on d 5 (early) and 17 (peak) of lactation, and on d 5 after weaning (postweaning). Relative expression of 7 potential reference genes, API5, MRPL39, VAPB, ACTB, GAPDH, RPS23, and MTG1, and one candidate gene, SLC7A1, was quantified by RT-qPCR using a relative standard curve approach. Variation in gene expression levels, measured as cycles to threshold at each stage of mammary physiological activity, was tested using a linear mixed model fitting RNA and DNA concentrations as covariates. Results were compared with those obtained with geNorm analysis, and genes selected by each method were used to normalize SLC7A1. Quantified relative mRNA abundance of GAPDH and MRPL39 remained unchanged across stages of mammary physiological activity after accounting for changes in tissue RNA and DNA concentration. In contrast, geNorm analysis selected MTG1, MRPL39, and VAPB as the best reference genes. However, when target gene SLC7A1 was normalized with genes selected either based on our proposed protocol or by geNorm, fold changes in mRNA abundance did not differ. In conclusion, the proposed analytical protocol assesses expression invariance of potential reference genes by accounting for variation in tissue RNA and DNA

  4. DNA methylation profiling of transcription factor genes in normal lymphocyte development and lymphomas.

    PubMed

    Ivascu, Claudia; Wasserkort, Reinhold; Lesche, Ralf; Dong, Jun; Stein, Harald; Thiel, Andreas; Eckhardt, Florian

    2007-01-01

    Transcription factors play a crucial role during hematopoiesis by orchestrating lineage commitment and determining cellular fate. Although tight regulation of transcription factor expression appears to be essential, little is known about the epigenetic mechanisms involved in transcription factor gene regulation. We have analyzed DNA methylation profiles of 13 key transcription factor genes in primary cells of the hematopoietic cascade, lymphoma cell lines and lymph node biopsies of diffuse large B-cell- and T-cell-non-Hodgkin lymphoma patients. Several of the transcription factor genes (SPI1, GATA3, TCF-7, Etv5, c-maf and TBX21) are differentially methylated in specific cell lineages and stages of the hematopoietic cascade. For some genes, such as SPI1, Etv5 and Eomes, we found an inverse correlation between the methylation of the 5' untranslated region and expression of the associated gene suggesting that these genes are regulated by DNA methylation. Differential methylation is not limited to cells of the healthy hematopoietic cascade, as we observed aberrant methylation of c-maf, TCF7, Eomes and SPI1 in diffuse large B-cell lymphomas. Our results suggest that epigenetic remodelling of transcription factor genes is a frequent mechanism during hematopoietic development. Aberrant methylation of transcription factor genes is frequently observed in diffuse large B-cell lymphomas and might have a functional role during tumorigenesis.

  5. Identification and validation of reference genes for accurate normalization of real-time quantitative PCR data in kiwifruit.

    PubMed

    Ferradás, Yolanda; Rey, Laura; Martínez, Óscar; Rey, Manuel; González, Ma Victoria

    2016-05-01

    Identification and validation of reference genes are required for the normalization of qPCR data. We studied the expression stability produced by eight primer pairs amplifying four common genes used as references for normalization. Samples representing different tissues, organs and developmental stages in kiwifruit (Actinidia chinensis var. deliciosa (A. Chev.) A. Chev.) were used. A total of 117 kiwifruit samples were divided into five sample sets (mature leaves, axillary buds, stigmatic arms, fruit flesh and seeds). All samples were also analysed as a single set. The expression stability of the candidate primer pairs was tested using three algorithms (geNorm, NormFinder and BestKeeper). The minimum number of reference genes necessary for normalization was also determined. A unique primer pair was selected for amplifying the 18S rRNA gene. The primer pair selected for amplifying the ACTIN gene was different depending on the sample set. 18S 2 and ACT 2 were the candidate primer pairs selected for normalization in the three sample sets (mature leaves, fruit flesh and stigmatic arms). 18S 2 and ACT 3 were the primer pairs selected for normalization in axillary buds. No primer pair could be selected for use as the reference for the seed sample set. The analysis of all samples in a single set did not produce the selection of any stably expressing primer pair. Considering data previously reported in the literature, we validated the selected primer pairs amplifying the FLOWERING LOCUS T gene for use in the normalization of gene expression in kiwifruit.

  6. Deactivation of the GATA Transcription Factor ELT-2 Is a Major Driver of Normal Aging in C. elegans

    PubMed Central

    Mann, Frederick G.; Van Nostrand, Eric L.; Friedland, Ari E.; Liu, Xiao; Kim, Stuart K.

    2016-01-01

    To understand the molecular processes underlying aging, we screened modENCODE ChIP-seq data to identify transcription factors that bind to age-regulated genes in C. elegans. The most significant hit was the GATA transcription factor encoded by elt-2, which is responsible for inducing expression of intestinal genes during embryogenesis. Expression of ELT-2 decreases during aging, beginning in middle age. We identified genes regulated by ELT-2 in the intestine during embryogenesis, and then showed that these developmental genes markedly decrease in expression as worms grow old. Overexpression of elt-2 extends lifespan and slows the rate of gene expression changes that occur during normal aging. Thus, our results identify the developmental regulator ELT-2 as a major driver of normal aging in C. elegans. PMID:27070429

  7. Deactivation of the GATA Transcription Factor ELT-2 Is a Major Driver of Normal Aging in C. elegans.

    PubMed

    Mann, Frederick G; Van Nostrand, Eric L; Friedland, Ari E; Liu, Xiao; Kim, Stuart K

    2016-04-01

    To understand the molecular processes underlying aging, we screened modENCODE ChIP-seq data to identify transcription factors that bind to age-regulated genes in C. elegans. The most significant hit was the GATA transcription factor encoded by elt-2, which is responsible for inducing expression of intestinal genes during embryogenesis. Expression of ELT-2 decreases during aging, beginning in middle age. We identified genes regulated by ELT-2 in the intestine during embryogenesis, and then showed that these developmental genes markedly decrease in expression as worms grow old. Overexpression of elt-2 extends lifespan and slows the rate of gene expression changes that occur during normal aging. Thus, our results identify the developmental regulator ELT-2 as a major driver of normal aging in C. elegans.

  8. A support vector machine model provides an accurate transcript-level-based diagnostic for major depressive disorder

    PubMed Central

    Yu, J S; Xue, A Y; Redei, E E; Bagheri, N

    2016-01-01

    Major depressive disorder (MDD) is a critical cause of morbidity and disability with an economic cost of hundreds of billions of dollars each year, necessitating more effective treatment strategies and novel approaches to translational research. A notable barrier in addressing this public health threat involves reliable identification of the disorder, as many affected individuals remain undiagnosed or misdiagnosed. An objective blood-based diagnostic test using transcript levels of a panel of markers would provide an invaluable tool for MDD as the infrastructure—including equipment, trained personnel, billing, and governmental approval—for similar tests is well established in clinics worldwide. Here we present a supervised classification model utilizing support vector machines (SVMs) for the analysis of transcriptomic data readily obtained from a peripheral blood specimen. The model was trained on data from subjects with MDD (n=32) and age- and gender-matched controls (n=32). This SVM model provides a cross-validated sensitivity and specificity of 90.6% for the diagnosis of MDD using a panel of 10 transcripts. We applied a logistic equation on the SVM model and quantified a likelihood of depression score. This score gives the probability of a MDD diagnosis and allows the tuning of specificity and sensitivity for individual patients to bring personalized medicine closer in psychiatry. PMID:27779627

  9. Retroviral insertions 90 kilobases proximal to the Evi-1 myeloid transforming gene activate transcription from the normal promoter.

    PubMed Central

    Bartholomew, C; Ihle, J N

    1991-01-01

    The inappropriate production of the Evi-1 zinc finger protein occurs in retrovirus-induced murine myeloid leukemias and human acute myelogenous leukemias. In murine leukemias, expression of the Evi-1 gene is associated with retroviral insertions either in the Evi-1 locus, which is immediately 5' of the coding region of the gene, or in the genetically linked Cb-1/fim-3 locus. In these studies, we demonstrate by chromosomal walking and pulse field electrophoresis that the Cb-1/fim-3 locus is located 90 kb 5' of the Evi-1 locus. Primary structure analysis of Evi-1 cDNA clones from a Cb-1/fim-3 rearranged cell line (DA-3) demonstrates that transcription initiates 5' of the Evi-1 locus and that the first noncoding exon of the gene is 681 bp larger than previously defined. S1 nuclease protection studies reveal multiple transcription initiation sites within this region. Comparable transcriptional initiation sites were identified in RNA from kidney and ovary, in which the gene is normally expressed, suggesting that retroviral insertions in the Cb-1/fim-3 locus activate transcription from the normal promoter. In one myeloid cell line (DA-3), a single long terminal repeat (LTR) is present in the Cb-1/fim-3 locus. No stable transcripts were detectable from this LTR. In cells with retroviral insertions in the Cb-1/fim-3 locus, one allele of the Evi-1 locus becomes hypermethylated in the 5' region of the gene. Together, these results are most consistent with an LTR-mediated, long-range cis activation of Evi-1 gene expression. Images PMID:1848663

  10. The update on transcriptional regulation of autophagy in normal and pathologic cells: A novel therapeutic target.

    PubMed

    Zhang, Zili; Guo, Mei; Zhao, Shifeng; Xu, Wenxuan; Shao, Jiangjuan; Zhang, Feng; Wu, Li; Lu, Yin; Zheng, Shizhong

    2015-08-01

    Autophagy is a genetically programmed, evolutionarily conserved catabolic process that occurs in response to stress. While short-lived proteins are degraded via the ubiquitin/proteasome pathway in higher eukaryotes, autophagy refers to intracellular pathway for development, differentiation, survival, and homeostasis through the lysosomal-dependent machinery, which regulates the synthesis, degradation, and recycling of long-lived proteins and even whole cytoplasmic organelles. The newly discovery shows that autophagy plays an important role in recycling nutrients upon starvation and maintaining cellular energy homeostasis. Recent basic and clinical investigations further demonstrate that autophagic abnormalities have been considered underlying reasons for lots of human diseases, including liver disease, cardiovascular and cerebrovascular diseases, neurodegenerative diseases, neoplastic diseases, and cancers. Pharmacological modulation of autophagy might point to possible therapeutic strategies for combating a broad range of diseases. Regrettably, although the mechanisms underlying the regulation of autophagy have long been attracted extensive concern of numerous scholars and its acute regulation by nutrient-sensing signaling pathways is well described, the overview on longer-term transcriptional regulation of autophagy is still filled with confusion. This review summarizes current knowledge on transcriptional regulation of autophagy and expands our horizons on the autophagy repertoire. On the other hand, it also offers a glimpse into different strategies that have been used in experimental models to counteract excessive pathological autophagy via transcriptional regulation mechanism in the prevention and treatment of disease.

  11. A human endogenous long terminal repeat provides a polyadenylation signal to a novel, alternatively spliced transcript in normal placenta.

    PubMed

    Goodchild, N L; Wilkinson, D A; Mager, D L

    1992-11-16

    We have been investigating the impact that the long terminal repeats (LTRs) of the RTVL-H family of human endogenous retroviral-like elements may have on the expression of adjacent cellular genes. Using a differential hybridization strategy, we have screened a cDNA library from a normal full-term human placenta and have identified two clones containing non-RTVL-H-related cellular sequences that have been polyadenylated within an RTVL-H LTR. One of these clones, cPj-LTR, contains an open reading frame (ORF) of 223 amino acids. Southern analysis indicated that the corresponding gene, termed PLT, is most probably a single multi-exon locus and that related sequences are present in the mouse genome, suggesting that this gene has been evolutionarily conserved. Database searches detected no significant homology to previously published sequences, indicating that PLT is a novel gene. Northern analysis identified several PLT-related transcripts in placental RNA samples, one of which is associated with the LTR. The presence of this PLT-LTR fusion transcript in normal placenta was also confirmed by PCR. Additional hybridization studies with RNAs from various cell lines suggested that the PLT locus is differentially expressed in different cell types. To investigate the structure of the non-LTR-associated PLT-related transcripts, additional clones were isolated from the placental cDNA library. Analysis of these clones suggests that the PLT mRNA undergoes alternative splicing at its 3' end, with polyadenylation within an RTVL-H LTR occurring in one of the resulting transcripts.

  12. Accurate discrimination of Alzheimer's disease from other dementia and/or normal subjects using SPECT specific volume analysis

    NASA Astrophysics Data System (ADS)

    Iyatomi, Hitoshi; Hashimoto, Jun; Yoshii, Fumuhito; Kazama, Toshiki; Kawada, Shuichi; Imai, Yutaka

    2014-03-01

    Discrimination between Alzheimer's disease and other dementia is clinically significant, however it is often difficult. In this study, we developed classification models among Alzheimer's disease (AD), other dementia (OD) and/or normal subjects (NC) using patient factors and indices obtained by brain perfusion SPECT. SPECT is commonly used to assess cerebral blood flow (CBF) and allows the evaluation of the severity of hypoperfusion by introducing statistical parametric mapping (SPM). We investigated a total of 150 cases (50 cases each for AD, OD, and NC) from Tokai University Hospital, Japan. In each case, we obtained a total of 127 candidate parameters from: (A) 2 patient factors (age and sex), (B) 12 CBF parameters and 113 SPM parameters including (C) 3 from specific volume analysis (SVA), and (D) 110 from voxel-based analysis stereotactic extraction estimation (vbSEE). We built linear classifiers with a statistical stepwise feature selection and evaluated the performance with the leave-one-out cross validation strategy. Our classifiers achieved very high classification performances with reasonable number of selected parameters. In the most significant discrimination in clinical, namely those of AD from OD, our classifier achieved both sensitivity (SE) and specificity (SP) of 96%. In a similar way, our classifiers achieved a SE of 90% and a SP of 98% in AD from NC, as well as a SE of 88% and a SP of 86% in AD from OD and NC cases. Introducing SPM indices such as SVA and vbSEE, classification performances improved around 7-15%. We confirmed that these SPM factors are quite important for diagnosing Alzheimer's disease.

  13. Transcription profiles of Streptococcus pneumoniae grown under different conditions of normal gravitation

    NASA Astrophysics Data System (ADS)

    Allen, C. A.; Galindo, C. L.; Pandya, U.; Watson, D. A.; Chopra, A. K.; Niesel, D. W.

    2007-02-01

    High-aspect rotating vessels (HARVs) are used to study the effects low-shear modeled microgravity (LSMMG) on bacterial gene expression. LSMMG is generated by orienting HARVs with the axis of rotation perpendicular to the gravity vector while gravitational controls are oriented with the axis of rotation parallel to the gravity vector. Microarray analysis was performed on Streptococcus pneumoniae TIGR4 grown in HARVs under three conditions (LSMMG, 1×g, and static) to determine if global transcriptional activity is altered between different gravitational controls and LSMMG. Results revealed 101 differentially expressed genes under static conditions compared to 1×g, 46 genes between 1×g and LSMMG, and nine genes between static and LSMMG. Hierarchical cluster analysis revealed 15 genes exhibiting similar expression patterns under static conditions compared to 1×g. These results indicate that rotation, in addition to low-shear forces, might contribute to bacterial adaptation to the LSMMG.

  14. Non-nuclear Pool of Splicing Factor SFPQ Regulates Axonal Transcripts Required for Normal Motor Development.

    PubMed

    Thomas-Jinu, Swapna; Gordon, Patricia M; Fielding, Triona; Taylor, Richard; Smith, Bradley N; Snowden, Victoria; Blanc, Eric; Vance, Caroline; Topp, Simon; Wong, Chun-Hao; Bielen, Holger; Williams, Katherine L; McCann, Emily P; Nicholson, Garth A; Pan-Vazquez, Alejandro; Fox, Archa H; Bond, Charles S; Talbot, William S; Blair, Ian P; Shaw, Christopher E; Houart, Corinne

    2017-04-04

    Recent progress revealed the complexity of RNA processing and its association to human disorders. Here, we unveil a new facet of this complexity. Complete loss of function of the ubiquitous splicing factor SFPQ affects zebrafish motoneuron differentiation cell autonomously. In addition to its nuclear localization, the protein unexpectedly localizes to motor axons. The cytosolic version of SFPQ abolishes motor axonal defects, rescuing key transcripts, and restores motility in the paralyzed sfpq null mutants, indicating a non-nuclear processing role in motor axons. Novel variants affecting the conserved coiled-coil domain, so far exclusively found in fALS exomes, specifically affect the ability of SFPQ to localize in axons. They broadly rescue morphology and motility in the zebrafish mutant, but alter motor axon morphology, demonstrating functional requirement for axonal SFPQ. Altogether, we uncover the axonal function of the splicing factor SFPQ in motor development and highlight the importance of the coiled-coil domain in this process.

  15. A putative APSES transcription factor is necessary for normal growth and development of Aspergillus nidulans.

    PubMed

    Lee, Ji-Yeon; Kim, Lee-Han; Kim, Ha-Eun; Park, Jae-Sin; Han, Kap-Hoon; Han, Dong-Min

    2013-12-01

    The nsdD gene encoding a GATA type transcription factor positively controls sexual development in Aspergillus nidulans. According to microarray data, 20 genes that were upregulated by deleting nsdD during various life cycle stages were randomly selected and deleted for functional analysis. None of the mutants showed apparent changes in growth or development compared with those of the wild-type except the AN3154 gene that encodes a putative APSES transcription factor and is an ortholog of Saccharomyces cerevisiae swi4. Deleting AN3154 resulted in retarded growth and development, and the gene was named rgdA (retared growth and development). The rgdA deletion mutant developed a reduced number of conidia even under favorable conditions for asexual development. The retarded growth and development was partially suppressed by the veA1 mutation. The conidial heads of the mutant aborted, showing reduced and irregular shaped phialides. Fruiting body development was delayed compared with that in the wild-type. The mutant did not respond to various nutritional or environmental factors that affected the development patterns. The rgdA gene was expressed at low levels throughout the life cycle and was not significantly affected by several regulators of sexual and asexual development such as nsdD, veA, stuA, or brlA. However, the rgdA gene affected brlA and abaA expression, which function as key regulators of asexual sporulation, suggesting that rgdA functions upstream of those genes.

  16. Drosophila Uri, a PP1α binding protein, is essential for viability, maintenance of DNA integrity and normal transcriptional activity

    PubMed Central

    Kirchner, Jasmin; Vissi, Emese; Gross, Sascha; Szoor, Balazs; Rudenko, Andrey; Alphey, Luke; White-Cooper, Helen

    2008-01-01

    Background Protein phosphatase 1 (PP1) is involved in diverse cellular processes, and is targeted to substrates via interaction with many different protein binding partners. PP1 catalytic subunits (PP1c) fall into PP1α and PP1β subfamilies based on sequence analysis, however very few PP1c binding proteins have been demonstrated to discriminate between PP1α and PP1β. Results URI (unconventional prefoldin RPB5 interactor) is a conserved molecular chaperone implicated in a variety of cellular processes, including the transcriptional response to nutrient signalling and maintenance of DNA integrity. We show that Drosophila Uri binds PP1α with much higher affinity than PP1β, and that this ability to discriminate between PP1c forms is conserved to humans. Most Uri is cytoplasmic, however we found some protein associated with active RNAPII on chromatin. We generated a uri loss of function allele, and show that uri is essential for viability in Drosophila. uri mutants have transcriptional defects, reduced cell viability and differentiation in the germline, and accumulate DNA damage in their nuclei. Conclusion Uri is the first PP1α specific binding protein to be described in Drosophila. Uri protein plays a role in transcriptional regulation. Activity of uri is required to maintain DNA integrity and cell survival in normal development. PMID:18412953

  17. Growth regulation, imprinting, and epigenetic transcription-related gene expression differs in lung of deceased transgenic cloned and normal goats.

    PubMed

    Meng, Li; Jia, Ruo-Xin; Sun, Yan-Yan; Wang, Zi-Yu; Wan, Yong-Jie; Zhang, Yan-Li; Zhong, Bu-Shuai; Wang, Feng

    2014-02-01

    Somatic cell nuclear transfer (SCNT) is a promising technique to produce mammalian transgenic clones. Only a small proportion of manipulated embryos, however, can develop into viable offspring. The abnormal growth and development of cloned animals, furthermore, are accompanied by aberrant lung development. Our objective was to investigate molecular background of lung developmental problems in transgenic (random insertion of exogenous DNA) cloned goats. We examined expression of 15 genes involved in growth regulation, imprinting, and epigenetic transcription in lung tissue of deceased transgenic cloned and normal goats of various ages. Compared with normal goats of the same age from conventional reproduction, expression of 13 genes (BMP4, FGF10, GHR, HGFR, PDGFR, RABP, VEGF, H19, CDKNIC, PCAF, MeCP2, HDAC1, and Dnmt3b) decreased in transgenic cloned goats that died at or shortly after birth; Expression of eight genes (FGF10, PDGFR, RABP, VEGF, PCAF, HDAC1, MeCP2, and Dnmt3b) decreased in fetal death of transgenic cloned goats. Expression of two epigenetic transcription genes (PCAF and Dnmt3b) decreased in disease death of transgenic cloned goats (1-4 months old). Disruptions in gene expression might be associated with the high neonatal mortality in transgenic cloned animals. These findings have implications in understanding the low efficiency of transgenic cloning.

  18. Single-cell transcriptional analysis of normal, aberrant, and malignant hematopoiesis in zebrafish

    PubMed Central

    Garcia, Elaine G.; Lobbardi, Riadh; Jain, Esha; Tang, Qin; Moore, John C.; Cortes, Mauricio; Molodtsov, Aleksey; Kasheta, Melissa; Luo, Christina C.; Garcia, Amaris J.; Mylvaganam, Ravi; Yoder, Jeffrey A.; Blackburn, Jessica S.; Sadreyev, Ruslan I.; Ceol, Craig J.; North, Trista E.

    2016-01-01

    Hematopoiesis culminates in the production of functionally heterogeneous blood cell types. In zebrafish, the lack of cell surface antibodies has compelled researchers to use fluorescent transgenic reporter lines to label specific blood cell fractions. However, these approaches are limited by the availability of transgenic lines and fluorescent protein combinations that can be distinguished. Here, we have transcriptionally profiled single hematopoietic cells from zebrafish to define erythroid, myeloid, B, and T cell lineages. We also used our approach to identify hematopoietic stem and progenitor cells and a novel NK-lysin 4+ cell type, representing a putative cytotoxic T/NK cell. Our platform also quantified hematopoietic defects in rag2E450fs mutant fish and showed that these fish have reduced T cells with a subsequent expansion of NK-lysin 4+ cells and myeloid cells. These data suggest compensatory regulation of the innate immune system in rag2E450fs mutant zebrafish. Finally, analysis of Myc-induced T cell acute lymphoblastic leukemia showed that cells are arrested at the CD4+/CD8+ cortical thymocyte stage and that a subset of leukemia cells inappropriately reexpress stem cell genes, including bmi1 and cmyb. In total, our experiments provide new tools and biological insights into single-cell heterogeneity found in zebrafish blood and leukemia. PMID:27139488

  19. Expression of Bcl-2 and Bcl-2-Ig fusion transcripts in normal and neoplastic cells.

    PubMed Central

    Graninger, W B; Seto, M; Boutain, B; Goldman, P; Korsmeyer, S J

    1987-01-01

    We examined the expression of the Bcl-2 gene at chromosome segment 18q21, that is translocated into the Ig heavy chain gene locus in t(14;18) bearing lymphomas. Bcl-2, while B cell associated, is expressed in a variety of hematopoietic lineages including T cells. Bcl-2 mRNA levels are high during pre-B cell development, the time at which the t(14;18) translocation occurs, but are down regulated with maturation. Like certain other oncogenes, Bcl-2 is quiescent in resting B cells but up-regulated with B cell activation. Mature B cell lymphomas with a t(14;18) have log-folds more mRNA than matched counterparts without the translocation. A sensitive S1 protection assay revealed that all transcripts in t(14;18) B cells were Bcl-2-Ig fusion mRNAs and originated from the translocated allele. Thus, there is a marked deregulation of Bcl-2 when it is introduced into the Ig locus in t(14;18) lymphomas. Images PMID:3500184

  20. Selection and validation of reference genes for transcript normalization in gene expression studies in Catharanthus roseus.

    PubMed

    Pollier, Jacob; Vanden Bossche, Robin; Rischer, Heiko; Goossens, Alain

    2014-10-01

    Quantitative Real-Time PCR (qPCR), a sensitive and commonly used technique for gene expression analysis, requires stably expressed reference genes for normalization of gene expression. Up to now, only one reference gene for qPCR analysis, corresponding to 40S Ribosomal protein S9 (RPS9), was available for the medicinal plant Catharanthus roseus, the only source of the commercial anticancer drugs vinblastine and vincristine. Here, we screened for additional reference genes for this plant species by mining C. roseus RNA-Seq data for orthologs of 22 genes known to be stably expressed in Arabidopsis thaliana and qualified as superior reference genes for this model plant species. Based on this, eight candidate C. roseus reference genes were identified and, together with RPS9, evaluated by performing qPCR on a series of different C. roseus explants and tissue cultures. NormFinder, geNorm and BestKeeper analyses of the resulting qPCR data revealed that the orthologs of At2g28390 (SAND family protein, SAND), At2g32170 (N2227-like family protein, N2227) and At4g26410 (Expressed protein, EXP) had the highest expression stability across the different C. roseus samples and are superior as reference genes as compared to the traditionally used RPS9. Analysis of publicly available C. roseus RNA-Seq data confirmed the expression stability of SAND and N2227, underscoring their value as reference genes for C. roseus qPCR analysis.

  1. TEAD transcription factors are required for normal primary myoblast differentiation in vitro and muscle regeneration in vivo

    PubMed Central

    Joshi, Shilpy; Le Gras, Stéphanie; Watanabe, Shuichi; Braun, Thomas

    2017-01-01

    The TEAD family of transcription factors (TEAD1-4) bind the MCAT element in the regulatory elements of both growth promoting and myogenic differentiation genes. Defining TEAD transcription factor function in myogenesis has proved elusive due to overlapping expression of family members and their functional redundancy. We show that silencing of either Tead1, Tead2 or Tead4 did not effect primary myoblast (PM) differentiation, but that their simultaneous knockdown strongly impaired differentiation. In contrast, Tead1 or Tead4 silencing impaired C2C12 differentiation showing their different contributions in PMs and C2C12 cells. Chromatin immunoprecipitation identified enhancers associated with myogenic genes bound by combinations of Tead4, Myod1 or Myog. Tead4 regulated distinct gene sets in C2C12 cells and PMs involving both activation of the myogenic program and repression of growth and signaling pathways. ChIP-seq from mature mouse muscle fibres in vivo identified a set of highly transcribed muscle cell-identity genes and sites bound by Tead1 and Tead4. Although inactivation of Tead4 in mature muscle fibres caused no obvious phenotype under normal conditions, notexin-induced muscle regeneration was delayed in Tead4 mutants suggesting an important role in myogenic differentiation in vivo. By combining knockdown in cell models in vitro with Tead4 inactivation in muscle in vivo, we provide the first comprehensive description of the specific and redundant roles of Tead factors in myogenic differentiation. PMID:28178271

  2. Selection and Validation of Reference Genes for Accurate RT-qPCR Data Normalization in Coffea spp. under a Climate Changes Context of Interacting Elevated [CO2] and Temperature

    PubMed Central

    Martins, Madlles Q.; Fortunato, Ana S.; Rodrigues, Weverton P.; Partelli, Fábio L.; Campostrini, Eliemar; Lidon, Fernando C.; DaMatta, Fábio M.; Ramalho, José C.; Ribeiro-Barros, Ana I.

    2017-01-01

    World coffee production has faced increasing challenges associated with ongoing climatic changes. Several studies, which have been almost exclusively based on temperature increase, have predicted extensive reductions (higher than half by 2,050) of actual coffee cropped areas. However, recent studies showed that elevated [CO2] can strongly mitigate the negative impacts of heat stress at the physiological and biochemical levels in coffee leaves. In addition, it has also been shown that coffee genotypes can successfully cope with temperatures above what has been traditionally accepted. Altogether, this information suggests that the real impact of climate changes on coffee growth and production could be significantly lower than previously estimated. Gene expression studies are an important tool to unravel crop acclimation ability, demanding the use of adequate reference genes. We have examined the transcript stability of 10 candidate reference genes to normalize RT-qPCR expression studies using a set of 24 cDNAs from leaves of three coffee genotypes (CL153, Icatu, and IPR108), grown under 380 or 700 μL CO2 L−1, and submitted to increasing temperatures from 25/20°C (day/night) to 42/34°C. Samples were analyzed according to genotype, [CO2], temperature, multiple stress interaction ([CO2], temperature) and total stress interaction (genotype, [CO2], and temperature). The transcript stability of each gene was assessed through a multiple analytical approach combining the Coeficient of Variation method and three algorithms (geNorm, BestKeeper, NormFinder). The transcript stability varied according to the type of stress for most genes, but the consensus ranking obtained with RefFinder, classified MDH as the gene with the highest mRNA stability to a global use, followed by ACT and S15, whereas α-TUB and CYCL showed the least stable mRNA contents. Using the coffee expression profiles of the gene encoding the large-subunit of ribulose-1,5-bisphosphate carboxylase

  3. The Homeodomain Transcription Factor Cdx1 Does Not Behave as an Oncogene in Normal Mouse Intestine1

    PubMed Central

    Crissey, Mary Ann S; Guo, Rong-Jun; Fogt, Franz; Li, Hong; Katz, Jonathan P; Silberg, Debra G; Suh, Eun Ran; Lynch, John P

    2008-01-01

    The Caudal-related homeobox genes Cdx1 and Cdx2 are intestine-specific transcription factors that regulate differentiation of intestinal cell types. Previously, we have shown Cdx1 to be antiproliferative and to promote cell differentiation. However, other studies have suggested that Cdx1 may be an oncogene. To test for oncogenic behavior, we used the murine villin promoter to ectopically express Cdx1 in the small intestinal villi and colonic surface epithelium. No changes in intestinal architecture, cell differentiation, or lineage selection were observed with expression of the transgene. Classic oncogenes enhance proliferation and induce tumors when ectopically expressed. However, the Cdx1 transgene neither altered intestinal proliferation nor induced spontaneous intestinal tumors. In a murine model for colitis-associated cancer, the Cdx1 transgene decreased, rather than increased, the number of adenomas that developed. In the polyps, the expression of the endogenous and the transgenic Cdx1 proteins was largely absent, whereas endogenous Villin expression was retained. This suggests that transgene silencing was specific and not due to a general Villin inactivation. In conclusion, neither the ectopic expression of Cdx1 was associated with changes in intestinal cell proliferation or differentiation nor was there increased intestinal cancer susceptibility. Our results therefore suggest that Cdx1 is not an oncogene in normal intestinal epithelium. PMID:18231635

  4. Polyamine-modulated c-Myc expression in normal intestinal epithelial cells regulates p21Cip1 transcription through a proximal promoter region

    PubMed Central

    Liu, Lan; Guo, Xin; Rao, Jaladanki N.; Zou, Tongtong; Marasa, Bernard S.; Chen, Jie; Greenspon, Jose; Casero, Robert A.; Wang, Jian-Ying

    2006-01-01

    Maintenance of intestinal mucosal epithelial integrity requires cellular polyamines that regulate expression of various genes involved in cell proliferation, growth arrest and apoptosis. Our previous studies have shown that polyamines are essential for expression of the c-myc gene and that polyamine-induced c-Myc plays a critical role in stimulation of normal IEC (intestinal epithelial cell) proliferation, but the exact downstream targets of induced c-Myc are still unclear. The p21Cip1 protein is a major player in cell cycle control, which is primarily regulated at the transcriptional level. The current study was designed to determine whether induced c-Myc stimulates normal IEC proliferation by repressing p21Cip1 transcription following up-regulation of polyamines. Overexpression of the ODC (ornithine decarboxylase) gene increased levels of cellular polyamines, induced c-Myc expression and inhibited p21Cip1 transcription, as indicated by repression of p21Cip1 promoter activity and a decrease in p21Cip1 protein levels. In contrast, depletion of cellular polyamines by inhibiting ODC enzyme activity with α-difluoromethylornithine decreased c-Myc, but increased p21Cip1 transcription. Ectopic expression of wild-type c-myc not only inhibited basal levels of p21Cip1 transcription in control cells, but also prevented increased p21Cip1 in polyamine-deficient cells. Experiments using different p21Cip1 promoter mutants showed that transcriptional repression of p21Cip1 by c-Myc was mediated through Miz-1- and Sp1-binding sites within the proximal region of the p21Cip1 promoter in normal IECs. These findings confirm that p21Cip1 is one of the direct mediators of induced c-Myc following increased polyamines and that p21Cip1 repression by c-Myc is implicated in stimulation of normal IEC proliferation. PMID:16706751

  5. Lesion Explorer: a video-guided, standardized protocol for accurate and reliable MRI-derived volumetrics in Alzheimer's disease and normal elderly.

    PubMed

    Ramirez, Joel; Scott, Christopher J M; McNeely, Alicia A; Berezuk, Courtney; Gao, Fuqiang; Szilagyi, Gregory M; Black, Sandra E

    2014-04-14

    Obtaining in vivo human brain tissue volumetrics from MRI is often complicated by various technical and biological issues. These challenges are exacerbated when significant brain atrophy and age-related white matter changes (e.g. Leukoaraiosis) are present. Lesion Explorer (LE) is an accurate and reliable neuroimaging pipeline specifically developed to address such issues commonly observed on MRI of Alzheimer's disease and normal elderly. The pipeline is a complex set of semi-automatic procedures which has been previously validated in a series of internal and external reliability tests(1,2). However, LE's accuracy and reliability is highly dependent on properly trained manual operators to execute commands, identify distinct anatomical landmarks, and manually edit/verify various computer-generated segmentation outputs. LE can be divided into 3 main components, each requiring a set of commands and manual operations: 1) Brain-Sizer, 2) SABRE, and 3) Lesion-Seg. Brain-Sizer's manual operations involve editing of the automatic skull-stripped total intracranial vault (TIV) extraction mask, designation of ventricular cerebrospinal fluid (vCSF), and removal of subtentorial structures. The SABRE component requires checking of image alignment along the anterior and posterior commissure (ACPC) plane, and identification of several anatomical landmarks required for regional parcellation. Finally, the Lesion-Seg component involves manual checking of the automatic lesion segmentation of subcortical hyperintensities (SH) for false positive errors. While on-site training of the LE pipeline is preferable, readily available visual teaching tools with interactive training images are a viable alternative. Developed to ensure a high degree of accuracy and reliability, the following is a step-by-step, video-guided, standardized protocol for LE's manual procedures.

  6. Cellular localization of tumor necrosis factor (TNF)-alpha transcripts in normal bowel and in necrotizing enterocolitis. TNF gene expression by Paneth cells, intestinal eosinophils, and macrophages.

    PubMed Central

    Tan, X.; Hsueh, W.; Gonzalez-Crussi, F.

    1993-01-01

    Tumor necrosis factor-alpha (TNF) has been shown to induce intestinal necrosis in animals. Moreover, plasma TNF levels are elevated in patients with necrotizing enterocolitis. Thus, it is possible that TNF plays a role in the pathogenesis of NEC. In the present study we used in situ hybridization (with human TNF riboprobes) to localize TNF transcripts in the intestinal tissues from normal biopsies and NEC patients. We found that in normal intestine a small amount of TNF mRNA was present only in Paneth cells. In contrast, in the acute stage of NEC, a high amount of TNF transcripts was detected in Paneth cells as well as in infiltrating eosinophils. In one case that showed infiltrating macrophages, TNF mRNA was also detected in these cells. Resident macrophages in the lamina propria and other inflammatory cells were negative for TNF transcripts. Our results suggest that: 1) Paneth cells are the major source of TNF transcripts in normal intestine, and 2) there is a marked increase in TNF mRNA formation in Paneth cells, as well as in infiltrating eosinophils and macrophages in patients with NEC. TNF-containing cells may play an important role in the pathophysiology of NEC. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:8506954

  7. Human collagen Krox up-regulates type I collagen expression in normal and scleroderma fibroblasts through interaction with Sp1 and Sp3 transcription factors.

    PubMed

    Kypriotou, Magdalini; Beauchef, Gallic; Chadjichristos, Christos; Widom, Russell; Renard, Emmanuelle; Jimenez, Sergio A; Korn, Joseph; Maquart, François-Xavier; Oddos, Thierry; Von Stetten, Otto; Pujol, Jean-Pierre; Galéra, Philippe

    2007-11-02

    Despite several investigations, the transcriptional mechanisms that regulate the expression of both type I collagen genes (COL1A1 and COL1A2) in either physiological or pathological situations, such as scleroderma, are not completely known. We have investigated the role of hc-Krox transcription factor on type I collagen expression by human dermal fibroblasts. hc-Krox exerted a stimulating effect on type I collagen protein synthesis and enhanced the corresponding mRNA steady-state levels of COL1A1 and COL1A2 in foreskin fibroblasts (FF), adult normal fibroblasts (ANF), and scleroderma fibroblasts (SF). Forced hc-Krox expression was found to up-regulate COL1A1 transcription through a -112/-61-bp sequence in FF, ANF, and SF. Knockdown of hc-Krox by short interfering RNA and decoy strategies confirmed the transactivating effect of hc-Krox and decreased substantially COL1A1 transcription levels in all fibro-blast types. The -112/-61-bp sequence bound specifically hc-Krox but also Sp1 and CBF. Attempts to elucidate the potential interactions between hc-Krox, Sp1, and Sp3 revealed that all of them co-immunoprecipitate from FF cellular extracts when a c-Krox antibody was used and bind to the COL1A1 promoter in chromatin immunoprecipitation assays. Moreover, hc-Krox DNA binding activity to its COL1A1-responsive element is increased in SF, cells producing higher amounts of type I collagen compared with ANF and FF. These data suggest that the regulation of COL1A1 gene transcription in human dermal fibroblasts involves a complex machinery that implicates at least three transcription proteins, hc-Krox, Sp1, and Sp3, which could act in concert to up-regulate COL1A1 transcriptional activity and provide evidence for a pro-fibrotic role of hc-Krox.

  8. Insulin-like growth factor-binding protein-7 (IGFBP7) transcript: A-to-I editing events in normal and cancerous human keratinocytes.

    PubMed

    Hochberg, Malka; Gilead, Leon; Markel, Gal; Nemlich, Yael; Feiler, Yulia; Enk, Claes David; Denichenko, Polina; Karni, Rotem; Ingber, Arieh

    2013-08-01

    Non-melanoma skin cancers (NMSC) are the most common malignancies in caucasians worldwide. Insulin-like growth factor-binding protein-7 (IGFBP7) was suggested to function as a tumor suppressor gene in several cancers, and to play a role in the proliferation of keratinocytes. A-to-I RNA editing is a post-transcriptional mechanism frequently used to expand and diversify transcriptome and proteome repertoire in eukaryotic cells. A-to-I RNA editing can alter codons, substitute amino acids and affect protein sequence, structure, and function. Two editing sites were identified within the IGFBP7 transcript. To evaluate the expression and editing of IGFBP7 mRNA in NMSC compared to normal epidermis. We examined the expression and mRNA editing level of IGFBP7 in 22 basal cell carcinoma (BCC), 15 squamous cell carcinoma (SCC), and 18 normal epidermis samples that were surgically removed from patients by the Mohs Micrographic Surgery procedure. We studied the effect of IGFBP7 editing on an immortalized HaCaT keratinocyte cell model. IGFBP7 mRNA is over expressed in BCC and SCC compared to normal epidermis. Moreover, the IGFBP7 transcript is highly edited in normal epidermis, but its editing is significantly reduced in BCC and SCC. The edited form of IGFBP7 can inhibit proliferation and induce senescence in cultured keratinocytes. This study describes for the first time A-to-I editing in the coding sequence of a tumor suppressor gene in humans, and suggests that IGFBP7 editing serves as a fine-tuning mechanism to maintain the equilibrium between proliferation and senescence in normal skin.

  9. Variations of transcript profiles between sea otters Enhydra lutris from Prince William Sound, Alaska, and clinically normal reference otters

    USGS Publications Warehouse

    Miles, A. Keith; Bowen, Lizabeth; Ballachey, Brenda E.; Bodkin, James L.; Murray, M.; Estes, J.L.; Keister, Robin A.; Stott, J.L.

    2012-01-01

    Development of blood leukocyte gene transcript profiles has the potential to expand condition assessments beyond those currently available to evaluate wildlife health, including sea otters Enhydra lutris, both individually and as populations. The 10 genes targeted in our study represent multiple physiological systems that play a role in immuno-modulation, inflammation, cell protection, tumor suppression, cellular stress-response, xenobiotic metabolizing enzymes, and antioxidant enzymes. These genes can be modified by biological, physical, or anthropogenic impacts and consequently provide information on the general type of stressors present in a given environment. We compared gene transcript profiles of sea otters sampled in 2008 among areas within Prince William Sound impacted to varying degrees by the 1989 ‘Exxon Valdez’ oil spill with those of captive and wild reference sea otters. Profiles of sea otters from Prince William Sound showed elevated transcription in genes associated with tumor formation, cell death, organic exposure, inflammation, and viral exposure when compared to the reference sea otter group, indicating possible recent and chronic exposure to organic contaminants. Sea otters from historically designated oiled areas within Prince William Sound 19 yr after the oil spill had higher transcription of genes associated with tumor formation, cell death, heat shock, and inflammation than those from areas designated as less impacted by the spill.

  10. Functional domains of the BACE1 and BACE2 promoters and mechanisms of transcriptional suppression of the BACE2 promoter in normal neuronal cells.

    PubMed

    Lahiri, Debomoy K; Maloney, Bryan; Ge, Yuan-Wen

    2006-01-01

    The beta-amyloid (Abeta) protein present in the neuritic plaques of Alzheimer's disease is cleaved from Abeta precursor protein (APP) by beta- and gamma-secretases. Following identification of beta-APP cleaving enzyme (BACE1) as the beta-secretase, a homologous beta-secretase 2 (BACE2) was described. Our goal is to characterize the regulatory region of the BACE genes. We compare functional domains within the BACE1 and BACE2 regulatory regions. Both BACE genes lack canonical TATAand CAAT boxes, but they contain distinguishing transcription start sites and transcription factor-binding sites. The BACE1 sequence contains more repetitive elements than does BACE2 (no elements). Regulatory domains do not overlap strongly between the two promoter regions. The BACE1upstream sequence contains both negative and positive domains, separated from the transcription seat by a long neutral domain. The corresponding BACE2sequence consists of a weakly positive domain directly upstream of a strongly positive domain, near a functionally active domain. DNA-protein interaction was corroborated by functional data. In primary rat cortical cultures, BACE1-driven reporter protein's expression was twice that of BACE2- driven reporter. The BACE2 gene promoter relatively reduced function in neuronal cells compared with BACE1. The BACE1 gene might operate through a single transcriptional control site. BACE2 operates through dual transcriptional control sites. Two (or more) regulatory pathways might control transcription in BACE2. Thus, BACE2 is partially suppressed in normal neuronal cells and likely to be a highly regulated gene expressed in a particularly tissue-specific fashion.

  11. Stem cells and the role of ETS transcription factors in the differentiation hierarchy of normal and malignant prostate epithelium.

    PubMed

    Archer, Leanne K; Frame, Fiona M; Maitland, Norman J

    2017-02-01

    Prostate cancer is the most common cancer of men in the UK and accounts for a quarter of all new cases. Although treatment of localised cancer can be successful, there is no cure for patients presenting with invasive prostate cancer and there are less treatment options. They are generally treated with androgen-ablation therapies but eventually the tumours become hormone resistant and patients develop castration-resistant prostate cancer (CRPC) for which there are no further successful or curative treatments. This highlights the need for new treatment strategies. In order to prevent prostate cancer recurrence and treatment resistance, all the cell populations in a heterogeneous prostate tumour must be targeted, including the rare cancer stem cell (CSC) population. The ETS transcription factor family members are now recognised as a common feature in multiple cancers including prostate cancer; with aberrant expression, loss of tumour suppressor function, inactivating mutations and the formation of fusion genes observed. Most notably, the TMPRSS2-ERG gene fusion is present in approximately 50% of prostate cancers and in prostate CSCs. However, the role of other ETS transcription factors in prostate cancer is less well understood. This review will describe the prostate epithelial cell hierarchy and discuss the evidence behind prostate CSCs and their inherent resistance to conventional cancer therapies. The known and proposed roles of the ETS family of transcription factors in prostate epithelial cell differentiation and regulation of the CSC phenotype will be discussed, as well as how they might be targeted for therapy.

  12. A novel bioinformatics pipeline for identification and characterization of fusion transcripts in breast cancer and normal cell lines.

    PubMed

    Asmann, Yan W; Hossain, Asif; Necela, Brian M; Middha, Sumit; Kalari, Krishna R; Sun, Zhifu; Chai, High-Seng; Williamson, David W; Radisky, Derek; Schroth, Gary P; Kocher, Jean-Pierre A; Perez, Edith A; Thompson, E Aubrey

    2011-08-01

    SnowShoes-FTD, developed for fusion transcript detection in paired-end mRNA-Seq data, employs multiple steps of false positive filtering to nominate fusion transcripts with near 100% confidence. Unique features include: (i) identification of multiple fusion isoforms from two gene partners; (ii) prediction of genomic rearrangements; (iii) identification of exon fusion boundaries; (iv) generation of a 5'-3' fusion spanning sequence for PCR validation; and (v) prediction of the protein sequences, including frame shift and amino acid insertions. We applied SnowShoes-FTD to identify 50 fusion candidates in 22 breast cancer and 9 non-transformed cell lines. Five additional fusion candidates with two isoforms were confirmed. In all, 30 of 55 fusion candidates had in-frame protein products. No fusion transcripts were detected in non-transformed cells. Consideration of the possible functions of a subset of predicted fusion proteins suggests several potentially important functions in transformation, including a possible new mechanism for overexpression of ERBB2 in a HER-positive cell line. The source code of SnowShoes-FTD is provided in two formats: one configured to run on the Sun Grid Engine for parallelization, and the other formatted to run on a single LINUX node. Executables in PERL are available for download from our web site: http://mayoresearch.mayo.edu/mayo/research/biostat/stand-alone-packages.cfm.

  13. Normalization with Corresponding Naïve Tissue Minimizes Bias Caused by Commercial Reverse Transcription Kits on Quantitative Real-Time PCR Results

    PubMed Central

    Garcia-Bardon, Andreas

    2016-01-01

    Real-time reverse transcription polymerase chain reaction (PCR) is the gold standard for expression analysis. Designed to improve reproducibility and sensitivity, commercial kits are commonly used for the critical step of cDNA synthesis. The present study was designed to determine the impact of these kits. mRNA from mouse brains were pooled to create serial dilutions ranging from 0.0625 μg to 2 μg, which were transcribed into cDNA using four different commercial reverse-transcription kits. Next, we transcribed mRNA from brain tissue after acute brain injury and naïve mice into cDNA for qPCR. Depending on tested genes, some kits failed to show linear results in dilution series and revealed strong variations in cDNA yield. Absolute expression data in naïve and trauma settings varied substantially between these kits. Normalization with a housekeeping gene failed to reduce kit-dependent variations, whereas normalization eliminated differences when naïve samples from the same region were used. The study shows strong evidence that choice of commercial cDNA synthesis kit has a major impact on PCR results and, consequently, on comparability between studies. Additionally, it provides a solution to overcome this limitation by normalization with data from naïve samples. This simple step helps to compare mRNA expression data between different studies and groups. PMID:27898720

  14. FGFR4 Gly388Arg polymorphism may affect the clinical stage of patients with lung cancer by modulating the transcriptional profile of normal lung.

    PubMed

    Falvella, Felicia S; Frullanti, Elisa; Galvan, Antonella; Spinola, Monica; Noci, Sara; De Cecco, Loris; Nosotti, Mario; Santambrogio, Luigi; Incarbone, Matteo; Alloisio, Marco; Calabrò, Elisa; Pastorino, Ugo; Skaug, Vidar; Haugen, Aage; Taioli, Emanuela; Dragani, Tommaso A

    2009-06-15

    The association of the fibroblast growth factor receptor 4 (FGFR4) Gly388Arg polymorphism with clinical stage and overall survival in a series of 541 Italian lung adenocarcinoma (ADCA) patients indicated a significantly decreased survival in patients carrying the rare Arg388 allele as compared to that in Gly/Gly homozygous patients [hazard ratio (HR) = 1.5; 95% confidence interval (CI) 1.1-1.9], with the decrease related to the association of the same polymorphism with clinical stage (HR = 1.8, 95% CI 1.3-2.6). By contrast, no significant association was detected in small series of either Norwegian lung ADCA patients or Italian lung squamous cell carcinoma (SQCC) patients. Single nucleotide polymorphisms of known FGFR4 ligands expressed in lung (FGF9, FGF18 and FGF19) were not associated with clinical stage or survival and showed no interaction with FGFR4. Analysis of gene expression profile in normal lungs according to FGFR4 genotype indicated a specific transcript pattern associated with the allele carrier status, suggesting a functional role for the FGFR4 polymorphism already detectable in normal lung. These findings confirm the significant association of the FGFR4 Gly388Arg polymorphism with clinical stage and overall survival in an Italian lung ADCA population and demonstrate a FGFR4 genotype-dependent transcriptional profile present in normal lung tissue.

  15. Morphometric and autoradiographic analysis of protein biosynthesis and transcription in sympathetic neurons of normal and partially desympathized rats

    SciTech Connect

    Chuchkova, N.N.; Morozov, I.A.; Yarygin, V.N.

    1986-06-01

    The authors undertake a differential morphometric analysis of the ultrastructural components for the protein-synthesizing apparatus: the rough endoplasmic reticulum and the ribosomes, polysomes, and monoribosomes in the cytoplasm of sympathetic nerve cells, connected with it, in rats. Tritium-UTP was applied to sections through a ganglion 8 micro thick and fixed in alcohol and acetone. Autoradiographhic analysis indicated differences in the level of template activity of the chromatin in the control and desympathized animals aged 6 months. The increase in the intensity of transcription was most marked for nucleolar chromatin.

  16. Identification and evaluation of reference genes for accurate gene expression normalization of fresh and frozen-thawed spermatozoa of water buffalo (Bubalus bubalis).

    PubMed

    Ashish, Shende; Bhure, S K; Harikrishna, Pillai; Ramteke, S S; Muhammed Kutty, V H; Shruthi, N; Ravi Kumar, G V P P S; Manish, Mahawar; Ghosh, S K; Mihir, Sarkar

    2017-04-01

    The quantitative real time PCR (qRT-PCR) has become an important tool for gene-expression analysis for a selected number of genes in life science. Although large dynamic range, sensitivity and reproducibility of qRT-PCR is good, the reliability majorly depend on the selection of proper reference genes (RGs) employed for normalization. Although, RGs expression has been reported to vary considerably within same cell type with different experimental treatments. No systematic study has been conducted to identify and evaluate the appropriate RGs in spermatozoa of domestic animals. Therefore, this study was conducted to analyze suitable stable RGs in fresh and frozen-thawed spermatozoa. We have assessed 13 candidate RGs (BACT, RPS18s, RPS15A, ATP5F1, HMBS, ATP2B4, RPL13, EEF2, TBP, EIF2B2, MDH1, B2M and GLUT5) of different functions and pathways using five algorithms. Regardless of the approach, the ranking of the most and the least candidate RGs remained almost same. The comprehensive ranking by RefFinder showed GLUT5, ATP2B4 and B2M, MDH1 as the top two stable and least stable RGs, respectively. The expression levels of four heat shock proteins (HSP) were employed as a target gene to evaluate RGs efficiency for normalization. The results demonstrated an exponential difference in expression levels of the four HSP genes upon normalization of the data with the most stable and the least stable RGs. Our study, provides a convenient RGs for normalization of gene-expression of key metabolic pathways effected during freezing and thawing of spermatozoa of buffalo and other closely related bovines.

  17. Three redundant brassinosteroid early response genes encode putative bHLH transcription factors required for normal growth.

    PubMed Central

    Friedrichsen, Danielle M; Nemhauser, Jennifer; Muramitsu, Takamichi; Maloof, Julin N; Alonso, José; Ecker, Joseph R; Furuya, Masaki; Chory, Joanne

    2002-01-01

    Brassinosteroids (BRs) are a class of polyhydroxylated steroids that are important regulators of plant growth and development. We have identified three closely related basic helix-loop-helix (bHLH) transcription factors, BEE1, BEE2, and BEE3, as products of early response genes required for full BR response. Comparison of the phenotypes of plants that overexpress BEE1 with bee1 bee2 bee3 triple-knockout mutant plants suggests that BEE1, BEE2, and BEE3 are functionally redundant positive regulators of BR signaling. Expression of BEE1, BEE2, and BEE3 is also regulated by other hormones, notably abscisic acid (ABA), a known antagonist of BR signaling. Reduced ABA response in plants overexpressing BEE1 suggests that BEE proteins may function as signaling intermediates in multiple pathways. PMID:12454087

  18. SPT5, an essential gene important for normal transcription in Saccharomyces cerevisiae, encodes an acidic nuclear protein with a carboxy-terminal repeat.

    PubMed Central

    Swanson, M S; Malone, E A; Winston, F

    1991-01-01

    Mutations in the SPT5 gene of Saccharomyces cerevisiae were isolated previously as suppressors of delta insertion mutations at HIS4 and LYS2. In this study we have shown that spt5 mutations suppress the his4-912 delta and lys2-128 delta alleles by altering transcription. We cloned the SPT5 gene and found that either an increase or a decrease in the copy number of the wild-type SPT5 gene caused an Spt- phenotype. Construction and analysis of an spt5 null mutation demonstrated that SPT5 is essential for growth, suggesting that SPT5 may be required for normal transcription of a large number of genes. The SPT5 DNA sequence was determined; it predicted a 116-kDa protein with an extremely acidic amino terminus and a novel six-amino-acid repeat at the carboxy terminus (consensus = S-T/A-W-G-G-A/Q). By indirect immunofluorescence microscopy we showed that a bifunctional SPT5-beta-galactosidase protein was located in the yeast nucleus. This molecular analysis of the SPT5 gene revealed a number of interesting similarities to the previously characterized SPT6 gene of S. cerevisiae. These results suggest that SPT5 and SPT6 act in a related fashion to influence essential transcriptional processes in S. cerevisiae. Images PMID:1840633

  19. Evaluation of reference genes for accurate normalization of gene expression for real time-quantitative PCR in Pyrus pyrifolia using different tissue samples and seasonal conditions.

    PubMed

    Imai, Tsuyoshi; Ubi, Benjamin E; Saito, Takanori; Moriguchi, Takaya

    2014-01-01

    We have evaluated suitable reference genes for real time (RT)-quantitative PCR (qPCR) analysis in Japanese pear (Pyrus pyrifolia). We tested most frequently used genes in the literature such as β-Tubulin, Histone H3, Actin, Elongation factor-1α, Glyceraldehyde-3-phosphate dehydrogenase, together with newly added genes Annexin, SAND and TIP41. A total of 17 primer combinations for these eight genes were evaluated using cDNAs synthesized from 16 tissue samples from four groups, namely: flower bud, flower organ, fruit flesh and fruit skin. Gene expression stabilities were analyzed using geNorm and NormFinder software packages or by ΔCt method. geNorm analysis indicated three best performing genes as being sufficient for reliable normalization of RT-qPCR data. Suitable reference genes were different among sample groups, suggesting the importance of validation of gene expression stability of reference genes in the samples of interest. Ranking of stability was basically similar between geNorm and NormFinder, suggesting usefulness of these programs based on different algorithms. ΔCt method suggested somewhat different results in some groups such as flower organ or fruit skin; though the overall results were in good correlation with geNorm or NormFinder. Gene expression of two cold-inducible genes PpCBF2 and PpCBF4 were quantified using the three most and the three least stable reference genes suggested by geNorm. Although normalized quantities were different between them, the relative quantities within a group of samples were similar even when the least stable reference genes were used. Our data suggested that using the geometric mean value of three reference genes for normalization is quite a reliable approach to evaluating gene expression by RT-qPCR. We propose that the initial evaluation of gene expression stability by ΔCt method, and subsequent evaluation by geNorm or NormFinder for limited number of superior gene candidates will be a practical way of finding out

  20. A diagnostic one-step real-time reverse transcription polymerase chain reaction method for accurate detection of influenza virus type A

    PubMed Central

    Behzadi, Mohammad Amin; Alborzi, Abdolvahab

    2016-01-01

    Introduction Influenza A is known as a public health concern worldwide. In this study, a novel one-step real-time reverse transcription polymerase chain reaction (rtRT-PCR) assay was designed and optimized for the detection of influenza A viruses. Material and methods The primers and probe were designed based on the analysis of 90 matrix nucleotide sequence data of influenza type A subtypes from the GenBank database of the National Center for Biotechnology Information (NCBI). The influenza virus A/Tehran/5652/2010 (H1N1 pdm09) was used as a reference. The rtRT-PCR assay was optimized, compared with that of the World Health Organization (WHO), and its analytical sensitivity, specificity and reproducibility were evaluated. In total, 64 nasopharyngeal swabs from patients with influenza-like illness (ILI) and 41 samples without ILI symptoms were tested for the virus, using conventional cell culture, direct immunofluorescence antibody (DFA) methods, and one-step rtRT-PCR with the designed primer set and probe and the WHO’s. Results The optimized assay results were similar to the WHO’s. The optimized assay results were similar to WHO’s, with non-significant differences for 10–103 copies of viral RNA/reaction (p > 0.05). It detected 10 copies of viral RNA/reaction with high reproducibility and no cross reactivity with other respiratory viruses. A specific cytopathic effect was observed in 6/64 (9.37%) of the ILI group using conventional culture and DFA staining methods; however, it was not seen in non-ILI. Also, the results of our assay and the WHO’s were similar to those of viral isolation and DFA staining. Conclusions Given the high specificity, sensitivity and reproducibility of this novel assay, it can serve as a reliable diagnostic tool for the detection of influenza A viruses in clinical specimens and lab experiments. PMID:27904520

  1. Haploinsufficiency in tumor predisposition syndromes: altered genomic transcription in morphologically normal cells heterozygous for VHL or TSC mutation.

    PubMed

    Peri, Suraj; Caretti, Elena; Tricarico, Rossella; Devarajan, Karthik; Cheung, Mitchell; Sementino, Eleonora; Menges, Craig W; Nicolas, Emmanuelle; Vanderveer, Lisa A; Howard, Sharon; Conrad, Peggy; Crowell, James A; Campbell, Kerry S; Ross, Eric A; Godwin, Andrew K; Yeung, Anthony T; Clapper, Margie L; Uzzo, Robert G; Henske, Elizabeth P; Ricketts, Christopher J; Vocke, Cathy D; Linehan, W Marston; Testa, Joseph R; Bellacosa, Alfonso; Kopelovich, Levy; Knudson, Alfred G

    2016-09-22

    Tumor suppressor genes and their effector pathways have been identified for many dominantly heritable cancers, enabling efforts to intervene early in the course of disease. Our approach on the subject of early intervention was to investigate gene expression patterns of morphologically normal "one-hit" cells before they become hemizygous or homozygous for the inherited mutant gene which is usually required for tumor formation. Here, we studied histologically non-transformed renal epithelial cells from patients with inherited disorders that predispose to renal tumors, including von Hippel-Lindau (VHL) disease and Tuberous Sclerosis (TSC). As controls, we studied histologically normal cells from non-cancerous renal epithelium of patients with sporadic clear cell renal cell carcinoma (ccRCC). Gene expression analyses of VHLmut/wt or TSC1/2mut/wt versus wild-type (WT) cells revealed transcriptomic alterations previously implicated in the transition to precancerous renal lesions. For example, the gene expression changes in VHLmut/wt cells were consistent with activation of the hypoxia response, associated, in part, with the "Warburg effect". Knockdown of any remaining VHL mRNA using shRNA induced secondary expression changes, such as activation of NFκB and interferon pathways, that are fundamentally important in the development of RCC. We posit that this is a general pattern of hereditary cancer predisposition, wherein haploinsufficiency for VHL or TSC1/2, or potentially other tumor susceptibility genes, is sufficient to promote development of early lesions, while cancer results from inactivation of the remaining normal allele. The gene expression changes identified here are related to the metabolic basis of renal cancer and may constitute suitable targets for early intervention.

  2. The duplication mutation of Quebec platelet disorder dysregulates PLAU, but not C10orf55, selectively increasing production of normal PLAU transcripts by megakaryocytes but not granulocytes

    PubMed Central

    Soomro, Asim; Waye, John S.; Paterson, Andrew D.; Rivard, Georges E.; Wilson, Michael D.

    2017-01-01

    Quebec Platelet disorder (QPD) is a unique bleeding disorder that markedly increases urokinase plasminogen activator (uPA) in megakaryocytes and platelets but not in plasma or urine. The cause is tandem duplication of a 78 kb region of chromosome 10 containing PLAU (the uPA gene) and C10orf55, a gene of unknown function. QPD increases uPA in platelets and megakaryocytes >100 fold, far more than expected for a gene duplication. To investigate the tissue-specific effect that PLAU duplication has on gene expression and transcript structure in QPD, we tested if QPD leads to: 1) overexpression of normal or unique PLAU transcripts; 2) increased uPA in leukocytes; 3) altered levels of C10orf55 mRNA and/or protein in megakaryocytes and leukocytes; and 4) global changes in megakaryocyte gene expression. Primary cells and cultured megakaryocytes from donors were prepared for quantitative reverse polymerase chain reaction analyses, RNA-seq and protein expression analyses. Rapidly isolated blood leukocytes from QPD subjects showed only a 3.9 fold increase in PLAU transcript levels, in keeping with the normal to minimally increased uPA in affinity purified, QPD leukocytes. All subjects had more uPA in granulocytes than monocytes and minimal uPA in lymphocytes. QPD leukocytes expressed PLAU alleles in proportions consistent with an extra copy of PLAU on the disease chromosome, unlike QPD megakaryocytes. QPD PLAU transcripts were consistent with reference gene models, with a much higher proportion of reads originating from the disease chromosome in megakaryocytes than granulocytes. QPD and control megakaryocytes contained minimal reads for C10orf55, and C10orf55 protein was not increased in QPD megakaryocytes or platelets. Finally, our QPD megakaryocyte transcriptome analysis revealed a global down regulation of the interferon type 1 pathway. We suggest that the low endogenous levels of uPA in blood are actively regulated, and that the regulatory mechanisms are disrupted in QPD

  3. Normal Tissue Complication Probability Estimation by the Lyman-Kutcher-Burman Method Does Not Accurately Predict Spinal Cord Tolerance to Stereotactic Radiosurgery

    SciTech Connect

    Daly, Megan E.; Luxton, Gary; Choi, Clara Y.H.; Gibbs, Iris C.; Chang, Steven D.; Adler, John R.; Soltys, Scott G.

    2012-04-01

    Purpose: To determine whether normal tissue complication probability (NTCP) analyses of the human spinal cord by use of the Lyman-Kutcher-Burman (LKB) model, supplemented by linear-quadratic modeling to account for the effect of fractionation, predict the risk of myelopathy from stereotactic radiosurgery (SRS). Methods and Materials: From November 2001 to July 2008, 24 spinal hemangioblastomas in 17 patients were treated with SRS. Of the tumors, 17 received 1 fraction with a median dose of 20 Gy (range, 18-30 Gy) and 7 received 20 to 25 Gy in 2 or 3 sessions, with cord maximum doses of 22.7 Gy (range, 17.8-30.9 Gy) and 22.0 Gy (range, 20.2-26.6 Gy), respectively. By use of conventional values for {alpha}/{beta}, volume parameter n, 50% complication probability dose TD{sub 50}, and inverse slope parameter m, a computationally simplified implementation of the LKB model was used to calculate the biologically equivalent uniform dose and NTCP for each treatment. Exploratory calculations were performed with alternate values of {alpha}/{beta} and n. Results: In this study 1 case (4%) of myelopathy occurred. The LKB model using radiobiological parameters from Emami and the logistic model with parameters from Schultheiss overestimated complication rates, predicting 13 complications (54%) and 18 complications (75%), respectively. An increase in the volume parameter (n), to assume greater parallel organization, improved the predictive value of the models. Maximum-likelihood LKB fitting of {alpha}/{beta} and n yielded better predictions (0.7 complications), with n = 0.023 and {alpha}/{beta} = 17.8 Gy. Conclusions: The spinal cord tolerance to the dosimetry of SRS is higher than predicted by the LKB model using any set of accepted parameters. Only a high {alpha}/{beta} value in the LKB model and only a large volume effect in the logistic model with Schultheiss data could explain the low number of complications observed. This finding emphasizes that radiobiological models

  4. Expression of the brain transcription factor OTX1 occurs in a subset of normal germinal-center B cells and in aggressive Non-Hodgkin Lymphoma.

    PubMed

    Omodei, Daniela; Acampora, Dario; Russo, Filippo; De Filippi, Rosaria; Severino, Valeria; Di Francia, Raffaele; Frigeri, Ferdinando; Mancuso, Pietro; De Chiara, Anna; Pinto, Antonio; Casola, Stefano; Simeone, Antonio

    2009-12-01

    The roles in brain development. Previous studies have shown the association between OTX2 and OTX1 with anaplastic and desmoplastic medulloblastomas, respectively. Here, we investigated OTX1 and OTX2 expression in Non-Hodgkin Lymphoma (NHL) and multiple myeloma. A combination of semiquantitative RT-PCR, Western blot, and immunohistochemical analyses was used to measure OTX1 and OTX2 levels in normal lymphoid tissues and in 184 tumor specimens representative of various forms of NHL and multiple myeloma. OTX1 expression was activated in 94% of diffuse large B-cell lymphomas, in all Burkitt lymphomas, and in 90% of high-grade follicular lymphomas. OTX1 was undetectable in precursor-B lymphoblastic lymphoma, chronic lymphocytic leukemia, and in most marginal zone and mantle cell lymphomas and multiple myeloma. OTX2 was undetectable in all analyzed malignancies. Analysis of OTX1 expression in normal lymphoid tissues identified a subset of resting germinal center (GC) B cells lacking PAX5 and BCL6 and expressing cytoplasmic IgG and syndecan. About 50% of OTX1(+) GC B cells co-expressed CD10 and CD20. This study identifies OTX1 as a molecular marker for high-grade GC-derived NHL and suggests an involvement of this transcription factor in B-cell lymphomagenesis. Furthermore, OTX1 expression in a subset of normal GC B cells carrying plasma cell markers suggests its possible contribution to terminal B-cell differentiation.

  5. Expression of the Brain Transcription Factor OTX1 Occurs in a Subset of Normal Germinal-Center B Cells and in Aggressive Non-Hodgkin Lymphoma

    PubMed Central

    Omodei, Daniela; Acampora, Dario; Russo, Filippo; De Filippi, Rosaria; Severino, Valeria; Di Francia, Raffaele; Frigeri, Ferdinando; Mancuso, Pietro; De Chiara, Anna; Pinto, Antonio; Casola, Stefano; Simeone, Antonio

    2009-01-01

    The roles in brain development. Previous studies have shown the association between OTX2 and OTX1 with anaplastic and desmoplastic medulloblastomas, respectively. Here, we investigated OTX1 and OTX2 expression in Non-Hodgkin Lymphoma (NHL) and multiple myeloma. A combination of semiquantitative RT-PCR, Western blot, and immunohistochemical analyses was used to measure OTX1 and OTX2 levels in normal lymphoid tissues and in 184 tumor specimens representative of various forms of NHL and multiple myeloma. OTX1 expression was activated in 94% of diffuse large B-cell lymphomas, in all Burkitt lymphomas, and in 90% of high-grade follicular lymphomas. OTX1 was undetectable in precursor-B lymphoblastic lymphoma, chronic lymphocytic leukemia, and in most marginal zone and mantle cell lymphomas and multiple myeloma. OTX2 was undetectable in all analyzed malignancies. Analysis of OTX1 expression in normal lymphoid tissues identified a subset of resting germinal center (GC) B cells lacking PAX5 and BCL6 and expressing cytoplasmic IgG and syndecan. About 50% of OTX1+ GC B cells co-expressed CD10 and CD20. This study identifies OTX1 as a molecular marker for high-grade GC-derived NHL and suggests an involvement of this transcription factor in B-cell lymphomagenesis. Furthermore, OTX1 expression in a subset of normal GC B cells carrying plasma cell markers suggests its possible contribution to terminal B-cell differentiation. PMID:19893048

  6. TCP transcription factor, BRANCH ANGLE DEFECTIVE 1 (BAD1), is required for normal tassel branch angle formation in maize

    PubMed Central

    Bai, Fang; Reinheimer, Renata; Durantini, Diego; Kellogg, Elizabeth A.; Schmidt, Robert J.

    2012-01-01

    In grass inflorescences, a structure called the “pulvinus” is found between the inflorescence main stem and lateral branches. The size of the pulvinus affects the angle of the lateral branches that emerge from the main axis and therefore has a large impact on inflorescence architecture. Through EMS mutagenesis we have identified three complementation groups of recessive mutants in maize having defects in pulvinus formation. All mutants showed extremely acute tassel branch angles accompanied by a significant reduction in the size of the pulvinus compared with normal plants. Two of the complementation groups correspond to mutations in the previously identified genes, RAMOSA2 (RA2) and LIGULELESS1 (LG1). Mutants corresponding to a third group were cloned using mapped-based approaches and found to encode a new member of the plant-specific TCP (TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL NUCLEAR ANTIGEN FACTOR) family of DNA-binding proteins, BRANCH ANGLE DEFECTIVE 1 (BAD1). BAD1 is expressed in the developing pulvinus as well as in other developing tissues, including the tassels and juvenile leaves. Both molecular and genetics studies show that RA2 is upstream of BAD1, whereas LG1 may function in a separate pathway. Our findings demonstrate that BAD1 is a TCP class II gene that functions to promote cell proliferation in a lateral organ, the pulvinus, and influences inflorescence architecture by impacting the angle of lateral branch emergence. PMID:22773815

  7. Novel Nine-Exon AR Transcripts (Exon 1/Exon 1b/Exons 2–8) in Normal and Cancerous Breast and Prostate Cells

    PubMed Central

    Hu, Dong Gui; McKinnon, Ross A.; Hulin, Julie-Ann; Mackenzie, Peter I.; Meech, Robyn

    2016-01-01

    Nearly 20 different transcripts of the human androgen receptor (AR) are reported with two currently listed as Refseq isoforms in the NCBI database. Isoform 1 encodes wild-type AR (type 1 AR) and isoform 2 encodes the variant AR45 (type 2 AR). Both variants contain eight exons: they share common exons 2–8 but differ in exon 1 with the canonical exon 1 in isoform 1 and the variant exon 1b in isoform 2. Splicing of exon 1 or exon 1b is reported to be mutually exclusive. In this study, we identified a novel exon 1b (1b/TAG) that contains an additional TAG trinucleotide upstream of exon 1b. Moreover, we identified AR transcripts in both normal and cancerous breast and prostate cells that contained either exon 1b or 1b/TAG spliced between the canonical exon 1 and exon 2, generating nine-exon AR transcripts that we have named isoforms 3a and 3b. The proteins encoded by these new AR variants could regulate androgen-responsive reporters in breast and prostate cancer cells under androgen-depleted conditions. Analysis of type 3 AR-GFP fusion proteins showed partial nuclear localization in PC3 cells under androgen-depleted conditions, supporting androgen-independent activation of the AR. Type 3 AR proteins inhibited androgen-induced growth of LNCaP cells. Microarray analysis identified a small set of type 3a AR target genes in LNCaP cells, including genes known to modulate growth and proliferation of prostate cancer (PCGEM1, PEG3, EPHA3, and EFNB2) or other types of human cancers (TOX3, ST8SIA4, and SLITRK3), and genes that are diagnostic/prognostic biomarkers of prostate cancer (GRINA3, and BCHE). PMID:28035996

  8. Novel Nine-Exon AR Transcripts (Exon 1/Exon 1b/Exons 2-8) in Normal and Cancerous Breast and Prostate Cells.

    PubMed

    Hu, Dong Gui; McKinnon, Ross A; Hulin, Julie-Ann; Mackenzie, Peter I; Meech, Robyn

    2016-12-27

    Nearly 20 different transcripts of the human androgen receptor (AR) are reported with two currently listed as Refseq isoforms in the NCBI database. Isoform 1 encodes wild-type AR (type 1 AR) and isoform 2 encodes the variant AR45 (type 2 AR). Both variants contain eight exons: they share common exons 2-8 but differ in exon 1 with the canonical exon 1 in isoform 1 and the variant exon 1b in isoform 2. Splicing of exon 1 or exon 1b is reported to be mutually exclusive. In this study, we identified a novel exon 1b (1b/TAG) that contains an additional TAG trinucleotide upstream of exon 1b. Moreover, we identified AR transcripts in both normal and cancerous breast and prostate cells that contained either exon 1b or 1b/TAG spliced between the canonical exon 1 and exon 2, generating nine-exon AR transcripts that we have named isoforms 3a and 3b. The proteins encoded by these new AR variants could regulate androgen-responsive reporters in breast and prostate cancer cells under androgen-depleted conditions. Analysis of type 3 AR-GFP fusion proteins showed partial nuclear localization in PC3 cells under androgen-depleted conditions, supporting androgen-independent activation of the AR. Type 3 AR proteins inhibited androgen-induced growth of LNCaP cells. Microarray analysis identified a small set of type 3a AR target genes in LNCaP cells, including genes known to modulate growth and proliferation of prostate cancer (PCGEM1, PEG3, EPHA3, and EFNB2) or other types of human cancers (TOX3, ST8SIA4, and SLITRK3), and genes that are diagnostic/prognostic biomarkers of prostate cancer (GRINA3, and BCHE).

  9. Microdissected double-minute DNA detects variable patterns of chromosomal localizations and multiple abundantly expressed transcripts in normal and leukemic cells

    SciTech Connect

    Sen, S.; Zhou, Hongyi; Stass, S.A.; Sen, P. ); Mulac-Jericevic, B.; Pirrotta, V. )

    1994-02-01

    Double-minute (dm) chromosomes are cytogenetically resolvable DNA amplification-mediating acentric extrachromosomal structures that are commonly seen in primary tumors, tumor cell lines, and drug-resistant cells grown in vitro. Selective isolation of dm DNAs with standard molecular biological techniques is difficult, and thus, detailed studies to elucidate their structure, site of chromosomal origin, and chromosomal reintegration patterns have been limited. In those instances in which a gene has been localized on dms, characterization of the remainder of the DNA, which far exceeds the size of the gene identified, has remained inconclusive. dms seen in the acute myeloid leukemia cell line HL-60 have been shown to harbor the c-myc protooncogene. In this paper, the authors report the successful isolation of the dm-specific DNAs from these cells by the microdissection/polymerase chain reaction technique and demonstrate that the dm DNAs derived from a single discrete normal chromosome segment 8q24.1-q24.2 reintegrate at various specific locations in the leukemic cells. The microdissected dm DNA detects multiple abundantly expressed transcripts distinct from c-myc mRNA on Northern blots. By devising a [open quotes]transcript selection[close quotes] strategy, they cloned the partial genomic sequence of a gene from the microdissected DNA that encodes two of these RNAs. This strategy will be generally applicable for rapid cloning of unknown amplified genes harbored on dms. With DNA from 20 microdissected dms, they constructed a genomic library of about 20,000 recombinant microclones with an average insert size of about 450 bp. The microclones should help in isolating corresponding yeast artificial chromosome clones for high-resolution physical mapping of dms in HL-60 cells. Furthermore, application of the microdissection technique appears to be an extremely feasible approach to characterization of dms in other cell types. 42 refs., 6 figs., 1 tab.

  10. Transcription profiling using RNA-Seq demonstrates expression differences in the body walls of juvenile albino and normal sea cucumbers Apostichopus japonicus

    NASA Astrophysics Data System (ADS)

    Ma, Deyou; Yang, Hongsheng; Sun, Lina; Chen, Muyan

    2014-01-01

    Sea cucumbers Apostichopus japonicus are one of the most important aquaculture species in China. Their normal body color is black to fit their surroundings. Wild albinos are rare and hard to breed. To understand the differences between albino and normal (control) sea cucumbers at the transcriptional level, we sequenced the transcriptomes in their body-wall tissues using RNA-Seq high-throughput sequencing. Approximately 4.876 million (M) and 4.884 M 200-nucleotide-long cDNA reads were produced in the cDNA libraries derived from the body walls of albino and control samples, respectively. A total of 9 561 (46.89%) putative genes were identified from among the RNA-Seq reads in both libraries. After filtering, 837 significantly differentially regulated genes were identified in the albino library compared with in the control library, and 3.6% of the differentially expressed genes (DEGs) were found to have changed those more than five-fold. The expression levels of 10 DEGs were checked by real-time PCR and the results were in full accord with the RNA-Seq expression trends, although the amplitude of the differences in expression levels was lower in all cases. A series of pathways were significantly enriched for the DEGs. These pathways were closely related to phagocytosis, the complement and coagulation cascades, apoptosis-related diseases, cytokine-cytokine receptor interaction, and cell adhesion. The differences in gene expression and enriched pathways between the albino and control sea cucumbers offer control targets for cultivating excellent albino A. japonicus strains in the future.

  11. Transcriptional profiles of benzo(a)pyrene exposure in normal human mammary epithelial cells in the absence or presence of chlorophyllin.

    PubMed

    John, Kaarthik; Keshava, Channa; Richardson, Diana L; Weston, Ainsley; Nath, Joginder

    2008-04-02

    Benzo(a)pyrene (BP) exposure causes alterations in gene expression in normal human mammary epithelial cells (NHMECs). This study used Affymetrix Hu-Gene133A arrays, with 14,500 genes represented, to evaluate modulation of BP-induced gene expression by chlorophyllin in six NHMEC strains derived from different donors. A major goal was to seek potential biomarkers of carcinogen exposure and how they behave in the presence of a chemopreventive agent. NHMECs (passage 6 and 70% confluence) were exposed for 24h to either vehicle control, or BP, or chlorophyllin followed by BP and chlorophyllin together. BP exposure resulted in approximately 3-fold altered expression of 49 genes in at least one of the six NHMEC strains. When cells were exposed to chlorophyllin pre-treatment followed by BP plus chlorophyllin, expression of 125 genes was similarly altered. Genes in the functional categories of xenobiotic metabolism, cell signaling, cell motility, cell proliferation, cellular transcription, metabolism, cell cycle control, apoptosis and DNA repair were identified. Only CYP1B1 and ALDH1A3 were consistently up-regulated by approximately 3-fold in most of the cell strains (at least 4) when exposed to BP. Cluster analysis identified a suite of 13 genes induced by BP where induction was mitigated in the presence of chlorophyllin. Additionally, cluster analysis identified a suite of 16 genes down-regulated by BP where induction was partially restored in the presence of chlorophyllin.

  12. Identification of stem cell transcriptional programs normally expressed in embryonic and neural stem cells in alloreactive CD8+ T cells mediating graft-versus-host disease

    PubMed Central

    Kato, Koji; Cui, Shuaiying; Kuick, Rork; Mineishi, Shin; Hexner, Elizabeth; Ferrara, James LM; Emerson, Stephen G.; Zhang, Yi

    2010-01-01

    A hallmark of graft-versus-host-disease (GVHD), a life-threatening complication after allogeneic hematopoietic stem cell transplantation, is the cytopathic injury of host tissues mediated by persistent alloreactive effector T cells (TE). However, the mechanisms that regulate the persistence of alloreactive TE during GVHD remain largely unknown. Using mouse GVHD models, we demonstrate that alloreactive CD8+ TE rapidly diminished in vivo when adoptively transferred into irradiated secondary congenic recipient mice. In contrast, although alloreactive CD8+ TE underwent massive apoptosis upon chronic exposure to alloantigens, they proliferated in vivo in secondary allogeneic recipients, persisted and caused severe GVHD. Thus, the continuous proliferation of alloreactive CD8+ TE, which is mediated by alloantigenic stimuli rather than homeostatic factors, is critical to maintaining their persistence. Gene expression profile analysis revealed that while alloreactive CD8+ TE increased the expression of genes associated with cell death, they activated a group of stem cell genes normally expressed in embryonic and neural stem cells. Most of these stem cell genes are associated with cell cycle regulation, DNA replication, chromatin modification and transcription. One of these genes, Ezh2, which encodes a chromatin modifying enzyme, was abundantly expressed in CD8+ TE. Silencing Ezh2 significantly reduced the proliferation of alloantigen-activated CD8+ T cells. Thus, these findings identify that a group of stem cell genes could play important roles in sustaining terminally differentiated alloreactive CD8+ TE and may be therapeutic targets for controlling GVHD. PMID:20116439

  13. Differentiation of the mRNA transcripts originating from the alpha 1- and alpha 2-globin loci in normals and alpha-thalassemics.

    PubMed

    Liebhaber, S A; Kan, Y W

    1981-08-01

    The alpha-globin polypeptide is encoded by two adjacent genes, alpha 1 and alpha 2. In the normal diploid state (alpha alpha/alpha alpha) all four alpha-globin genes are expressed. Loss or dysfunction of one or more of these genes leads to deficient alpha-globin production and results in alpha-thalassemia. We present a technique to differentially assess the steady-state levels of the alpha 1- and alpha-2-globin messenger RNA (mRNA) transcripts and thus delineate the relative level of expression of the two alpha-globin loci in a variety of alpha-thalassemia states. Only alpha 1 mRNA was produced in the alpha-thalassemia-2 haplotype (-alpha) (one of the two alpha-globin genes deleted from chromosome 16). This confirms previous gene mapping data which demonstrate deletion of the alpha 2 gene. The triple alpha-globin gene haplotype (alpha alpha alpha) is the reciprocal of the alpha-thalassemia-2 haplotype and thus contains an extra alpha 2-globin gene. RNA from this haplotype contained a greater than normal level of alpha 2-relative to alpha 1-globin mRNA. This data implies that the extra alpha 2 gene in the triple alpha-globin haplotype is functional. We detected a relative instability of the alpha 2-globin mRNA encoding the alpha-globin structural mutant Constant Spring. This instability may contribute to the low level of expression of the alpha-Constant Spring protein. In a Chinese patient with nondeletion hemoglobin-H disease (- -/alpha alpha T) (both alpha-globin genes are present but not fully functional) a normal ratio was maintained between the levels of alpha 1- and alpha 2-globin mRNA, implying that mRNA production from both alpha-globin genes is suppressed in a balanced manner. These observations extended previous findings concerning the structural rearrangements in the deletion types of alpha-thalassemia and the pathophysiology of two nondeletion variants.

  14. Accurate quantification of sphingosine-1-phosphate in normal and Fabry disease plasma, cells and tissues by LC-MS/MS with (13)C-encoded natural S1P as internal standard.

    PubMed

    Mirzaian, Mina; Wisse, Patrick; Ferraz, Maria J; Marques, André R A; Gabriel, Tanit L; van Roomen, Cindy P A A; Ottenhoff, Roelof; van Eijk, Marco; Codée, Jeroen D C; van der Marel, Gijsbert A; Overkleeft, Herman S; Aerts, Johannes M

    2016-08-01

    We developed a mass spectrometric procedure to quantify sphingosine-1-phosphate (S1P) in biological materials. The use of newly synthesized (13)C5 C18-S1P and commercial C17-S1P as internal standards rendered very similar results with respect to linearity, limit of detection and limit of quantitation. Caution is warranted with determination of plasma S1P levels. Earlier it was reported that S1P is elevated in plasma of Fabry disease patients. We investigated this with the improved quantification. No clear conclusion could be drawn for patient plasma samples given the lack of uniformity of blood collection and plasma preparation. To still obtain insight, plasma and tissues were identically collected from α-galactosidase A deficient Fabry mice and matched control animals. No significant difference was observed in plasma S1P levels. A significant 2.3 fold increase was observed in kidney of Fabry mice, but not in liver and heart. Comparative analysis of S1P in cultured fibroblasts from normal subjects and classically affected Fabry disease males revealed no significant difference. In conclusion, accurate quantification of S1P in biological materials is feasible by mass spectrometry using the internal standards (13)C5 C18-S1P or C17-S1P. Significant local increases of S1P in the kidney might occur in Fabry disease as suggested by the mouse model.

  15. All-trans retinoic acid converts E2F into a transcriptional suppressor and inhibits the growth of normal human bronchial epithelial cells through a retinoic acid receptor- dependent signaling pathway.

    PubMed Central

    Lee, H Y; Dohi, D F; Kim, Y H; Walsh, G L; Consoli, U; Andreeff, M; Dawson, M I; Hong, W K; Kurie, J M

    1998-01-01

    Retinoids, including retinol and retinoic acid derivatives, maintain the normal growth and differentiation of human bronchial epithelial (HBE) cells and are under investigation as agents for lung cancer prevention. In this study, we examined the biologic effects of retinoids on normal HBE cells and the molecular mechanisms of retinoid actions. At a dose of 10(-6) M, all-trans retinoic acid (t-RA) suppressed the proliferation of normal HBE cells, which accumulated in the G0 phase. No evidence of programmed cell death was observed. The class of retinoid nuclear receptor that mediated the growth arrest was explored. Normal HBE cell growth was suppressed by a retinoid that selectively activates retinoic acid receptors but not by one that activates retinoid X receptors. The E2F transcription factor has demonstrated a role in G0 entry through transcriptional suppression of genes that induce cell cycle progression. To investigate the role of E2F in retinoid signaling, transient transfection assays were performed using reporter plasmids containing E2F-binding sites. Findings from these experiments suggested that t-RA treatment converted E2F into a transcriptional suppressor. Supporting this possibility, t-RA inhibited the expression of the E2F target genes B-myb, cyclin A, and cyclin E. Further, t-RA increased the levels of nuclear E2F-4, p107, and p130 and enhanced the binding of E2F-4 to p107, which have been associated with the conversion of E2F into a transcriptional suppressor in other cells. These findings point to retinoic acid receptor- and E2F-dependent pathways as potential mediators of retinoid-induced growth arrest in normal HBE cells and have implications for the use of retinoids in clinical trials on the prevention of lung cancer. PMID:9486971

  16. Correlation between CYP1A1 transcript, protein level, enzyme activity and DNA adduct formation in normal human mammary epithelial cell strains exposed to benzo[a]pyrene

    PubMed Central

    Divi, Rao L.; Einem Lindeman, Tracey L.; Shockley, Marie E.; Keshava, Channa; Weston, Ainsley; Poirier, Miriam C.

    2014-01-01

    The polycyclic aromatic hydrocarbon (PAH) benzo(a)pyrene (BP) is thought to bind covalently to DNA, through metabolism by cytochrome P450 1A1 (CYP1A1) and CYP1B1, and other enzymes, to form r7, t8, t9-trihydroxy-c-10-(N 2-deoxyguanosyl)-7,8,9,10-tetrahydro-benzo[a]-pyrene (BPdG). Evaluation of RNA expression data, to understand the contribution of different metabolic enzymes to BPdG formation, is typically presented as fold-change observed upon BP exposure, leaving the actual number of RNA transcripts unknown. Here, we have quantified RNA copies/ng cDNA (RNA cpn) for CYP1A1 and CYP1B1, as well as NAD(P)H:quinone oxidoreductase 1 (NQO1), which may reduce formation of BPdG adducts, using primary normal human mammary epithelial cell (NHMEC) strains, and the MCF-7 breast cancer cell line. In unexposed NHMECs, basal RNA cpn values were 58–836 for CYP1A1, 336–5587 for CYP1B1 and 5943–40112 for NQO1. In cells exposed to 4.0 µM BP for 12h, RNA cpn values were 251–13234 for CYP1A1, 4133–57078 for CYP1B1 and 4456–55887 for NQO1. There were 3.5 (mean, range 0.2–15.8) BPdG adducts/108 nucleotides in the NHMECs (n = 16), and 790 in the MCF-7s. In the NHMECs, BP-induced CYP1A1 RNA cpn was highly associated with BPdG (P = 0.002), but CYP1B1 and NQO1 were not. Western blots of four NHMEC strains, chosen for different levels of BPdG adducts, showed a linear correlation between BPdG and CYP1A1, but not CYP1B1 or NQO1. Ethoxyresorufin-O-deethylase (EROD) activity, which measures CYP1A1 and CYP1B1 together, correlated with BPdG, but NQO1 activity did not. Despite more numerous levels of CYP1B1 and NQO1 RNA cpn in unexposed and BP-exposed NHMECs and MCF-7cells, BPdG formation was only correlated with induction of CYP1A1 RNA cpn. The higher level of BPdG in MCF-7 cells, compared to NHMECs, may have been due to a much increased induction of CYP1A1 and EROD. Overall, BPdG correlation was observed with CYP1A1 protein and CYP1A1/1B1 enzyme activity, but not with CYP1B1 or NQO

  17. Expression of the p66Shc protein adaptor is regulated by the activator of transcription STAT4 in normal and chronic lymphocytic leukemia B cells

    PubMed Central

    Cattaneo, Francesca; Patrussi, Laura; Capitani, Nagaja; Frezzato, Federica; D'Elios, Mario Milco; Trentin, Livio; Semenzato, Gianpietro; Baldari, Cosima T.

    2016-01-01

    p66Shc attenuates mitogenic, prosurvival and chemotactic signaling and promotes apoptosis in lymphocytes. Consistently, p66Shc deficiency contributes to the survival and trafficking abnormalities of chronic lymphocytic leukemia (CLL) B cells. The mechanism of p66shc silencing in CLL B cells is methylation-independent, at variance with other cancer cell types. Here we identify STAT4 as a novel transcriptional regulator of p66Shc in B cells. Chromatin immunoprecipitation and reporter gene assays showed that STAT4 binds to and activates the p66shc promoter. Silencing or overexpression of STAT4 resulted in a co-modulation of p66Shc. IL-12-dependent STAT4 activation caused a coordinate increase in STAT4 and p66Shc expression, which correlated with enhanced B cell apoptosis. Treatment with the STAT4 inhibitor lisofylline reverted partly this effect, suggesting that STAT4 phosphorylation is not essential for but enhances p66shc transcription. Additionally, we demonstrate that CLL B lymphocytes have a STAT4 expression defect which partly accounts for their p66Shc deficiency, as supported by reconstitution experiments. Finally, we show that p66Shc participates in a positive feedback loop to promote STAT4 expression. These results provide new insights into the mechanism of p66Shc expression in B cells and its defect in CLL, identifying the STAT4/IL-12 pathway as a potential therapeutic target in this neoplasia. PMID:27494881

  18. Mouse homolog of Saccharomyces cerevisiae spo11 is induced in normal mu(+)B-cells by stimuli that cause germline C(H) transcription and subsequent class switch recombination.

    PubMed

    Tokuyama, H; Tokuyama, Y

    2000-05-25

    The first step of Ig heavy chain class switch recombination (CSR) is considered to be DNA double strand break (DSB) formation in the two switch (S) regions (S(mu) and downstream S(H)), although the underlying mechanism is unknown. Recently, it has been demonstrated that at least Spo11, a homolog of the novel type II topoisomerase (topo VI) that catalyzes DSB formation, is involved in the initiation of meiotic recombination of Saccaromyces cerevisiae. In the present study, we examined whether the mouse homolog of Spo11 is induced in normal mouse mu(+)B-cells by stimuli that cause an early step of CSR, germline C(H) transcription, and subsequent CSR. Two CSR systems were used: IgA CSR induced by all-trans retinoic acid, IL-5, and LPS, and IgG1 CSR induced by IL-4 and LPS. Germline transcript and mouse Spo11 expression were analyzed by RT-PCR. In both systems, first germline transcripts were clearly detected on day 2 and then Spo11 was detected on day 3, increasing thereafter with time. The time course of changes in Spo11 expression coincided with that of CSR. Spo11 seems to be induced by CSR-inducing stimuli, regardless of the direction of CSR. These results suggested that mouse Spo11 might participate in the initiation step of CSR.

  19. Berry Phenolic Compounds Increase Expression of Hepatocyte Nuclear Factor-1α (HNF-1α) in Caco-2 and Normal Colon Cells Due to High Affinities with Transcription and Dimerization Domains of HNF-1α.

    PubMed

    Real Hernandez, Luis M; Fan, Junfeng; Johnson, Michelle H; Gonzalez de Mejia, Elvira

    2015-01-01

    Hepatocyte nuclear factor-1α (HNF-1α) is found in the kidneys, spleen, thymus, testis, skin, and throughout the digestive organs. It has been found to promote the transcription of various proteins involved in the management of type II diabetes, including dipeptidyl peptidase-IV (DPP-IV). Phenolic compounds from berries and citrus fruits are known to inhibit DPP-IV, but have not been tested for their interactions with wild-type HNF-1α. By studying the interactions of compounds from berries and citrus fruits have with HNF-1α, pre-transcriptional mechanisms that inhibit the expression of proteins such as DPP-IV may be elucidated. In this study, the interactions of berry phenolic compounds and citrus flavonoids with the dimerization and transcriptional domains of HNF-1α were characterized using the molecular docking program AutoDock Vina. The anthocyanin delphinidin-3-O-arabinoside had the highest binding affinity for the dimerization domain as a homodimer (-7.2 kcal/mol) and transcription domain (-8.3 kcal/mol) of HNF-1α. Anthocyanins and anthocyanidins had relatively higher affinities than resveratrol and citrus flavonoids for both, the transcription domain and the dimerization domain as a homodimer. The flavonoid flavone had the highest affinity for a single unit of the dimerization domain (-6.5 kcal/mol). Nuclear expression of HNF-1α was measured in Caco-2 and human normal colon cells treated with blueberry and blackberry anthocyanin extracts. All extracts tested increased significantly (P < 0.05) the nuclear expression of HNF-1α in Caco-2 cells by 85.2 to 260% compared to a control. The extracts tested increased significantly (P < 0.02) the nuclear expression of HNF-1α in normal colon cells by 48.6 to 243%. It was confirmed that delphinidin-3-O-glucoside increased by 3-fold nuclear HNF-1α expression in Caco-2 cells (P < 0.05). Anthocyanins significantly increased nuclear HNF-1α expression, suggesting that these compounds might regulate the genes HNF-1

  20. Berry Phenolic Compounds Increase Expression of Hepatocyte Nuclear Factor-1α (HNF-1α) in Caco-2 and Normal Colon Cells Due to High Affinities with Transcription and Dimerization Domains of HNF-1α

    PubMed Central

    Real Hernandez, Luis M.; Fan, Junfeng; Johnson, Michelle H.; Gonzalez de Mejia, Elvira

    2015-01-01

    Hepatocyte nuclear factor-1α (HNF-1α) is found in the kidneys, spleen, thymus, testis, skin, and throughout the digestive organs. It has been found to promote the transcription of various proteins involved in the management of type II diabetes, including dipeptidyl peptidase-IV (DPP-IV). Phenolic compounds from berries and citrus fruits are known to inhibit DPP-IV, but have not been tested for their interactions with wild-type HNF-1α. By studying the interactions of compounds from berries and citrus fruits have with HNF-1α, pre-transcriptional mechanisms that inhibit the expression of proteins such as DPP-IV may be elucidated. In this study, the interactions of berry phenolic compounds and citrus flavonoids with the dimerization and transcriptional domains of HNF-1α were characterized using the molecular docking program AutoDock Vina. The anthocyanin delphinidin-3-O-arabinoside had the highest binding affinity for the dimerization domain as a homodimer (-7.2 kcal/mol) and transcription domain (-8.3 kcal/mol) of HNF-1α. Anthocyanins and anthocyanidins had relatively higher affinities than resveratrol and citrus flavonoids for both, the transcription domain and the dimerization domain as a homodimer. The flavonoid flavone had the highest affinity for a single unit of the dimerization domain (-6.5 kcal/mol). Nuclear expression of HNF-1α was measured in Caco-2 and human normal colon cells treated with blueberry and blackberry anthocyanin extracts. All extracts tested increased significantly (P < 0.05) the nuclear expression of HNF-1α in Caco-2 cells by 85.2 to 260% compared to a control. The extracts tested increased significantly (P < 0.02) the nuclear expression of HNF-1α in normal colon cells by 48.6 to 243%. It was confirmed that delphinidin-3-O-glucoside increased by 3-fold nuclear HNF-1α expression in Caco-2 cells (P < 0.05). Anthocyanins significantly increased nuclear HNF-1α expression, suggesting that these compounds might regulate the genes HNF-1

  1. Establishment and characterization of A novel Philadelphia-chromosome positive chronic myeloid leukemia cell line, TCC-S, expressing P210 and P190 BCR/ABL transcripts but missing normal ABL gene.

    PubMed

    Van, Phan Nguyen Thanh; Xinh, Phan Thi; Kano, Yasuhiko; Tokunaga, Katsushi; Sato, Yuko

    2005-03-01

    A novel Philadelphia-chromosome positive (Ph+) cell line, TCC-S, has been established from a patient with Ph+ chronic myeloid leukemia (CML) in the blastic crisis. TCC-S cells were shown to express both P210 and P190 BCR/ABL transcripts by reverse transcriptase-polymerase chain reaction (PCR), although quantitative-PCR revealed that TCC-S cells mainly expressed P210 BCR/ABL transcript. Karyotype analysis revealed several triploid clones which constantly harbored two der(9)del(9) (p12)t(9;22) (q34;qll)s and two del(9) (q21)s. The der(9)del(9) (p12)t(9;22) (q34;q11) is rarely found in other CML cell lines. Moreover, to the best of our knowledge, del(9) (q21) resulting in missing of a restrict region including normal ABL gene has not been found among CML cell lines previously described. Thus, TCC-S cells with only BCR/ABL gene and no normal ABL gene may be a useful tool for functional study of ABL in Ph+ CML.

  2. Caulobacter crescentus CdnL is a non-essential RNA polymerase-binding protein whose depletion impairs normal growth and rRNA transcription.

    PubMed

    Gallego-García, Aránzazu; Iniesta, Antonio A; González, Diego; Collier, Justine; Padmanabhan, S; Elías-Arnanz, Montserrat

    2017-02-24

    CdnL is an essential RNA polymerase (RNAP)-binding activator of rRNA transcription in mycobacteria and myxobacteria but reportedly not in Bacillus. Whether its function and mode of action are conserved in other bacteria thus remains unclear. Because virtually all alphaproteobacteria have a CdnL homolog and none of these have been characterized, we studied the homolog (CdnLCc) of the model alphaproteobacterium Caulobacter crescentus. We show that CdnLCc is not essential for viability but that its absence or depletion causes slow growth and cell filamentation. CdnLCc is degraded in vivo in a manner dependent on its C-terminus, yet excess CdnLCc resulting from its stabilization did not adversely affect growth. We find that CdnLCc interacts with itself and with the RNAP β subunit, and localizes to at least one rRNA promoter in vivo, whose activity diminishes upon depletion of CdnLCc. Interestingly, cells expressing CdnLCc mutants unable to interact with the RNAP were cold-sensitive, suggesting that CdnLCc interaction with RNAP is especially required at lower than standard growth temperatures in C. crescentus. Our study indicates that despite limited sequence similarities and regulatory differences compared to its myco/myxobacterial homologs, CdnLCc may share similar biological functions, since it affects rRNA synthesis, probably by stabilizing open promoter-RNAP complexes.

  3. Caulobacter crescentus CdnL is a non-essential RNA polymerase-binding protein whose depletion impairs normal growth and rRNA transcription

    PubMed Central

    Gallego-García, Aránzazu; Iniesta, Antonio A.; González, Diego; Collier, Justine; Padmanabhan, S.; Elías-Arnanz, Montserrat

    2017-01-01

    CdnL is an essential RNA polymerase (RNAP)-binding activator of rRNA transcription in mycobacteria and myxobacteria but reportedly not in Bacillus. Whether its function and mode of action are conserved in other bacteria thus remains unclear. Because virtually all alphaproteobacteria have a CdnL homolog and none of these have been characterized, we studied the homolog (CdnLCc) of the model alphaproteobacterium Caulobacter crescentus. We show that CdnLCc is not essential for viability but that its absence or depletion causes slow growth and cell filamentation. CdnLCc is degraded in vivo in a manner dependent on its C-terminus, yet excess CdnLCc resulting from its stabilization did not adversely affect growth. We find that CdnLCc interacts with itself and with the RNAP β subunit, and localizes to at least one rRNA promoter in vivo, whose activity diminishes upon depletion of CdnLCc. Interestingly, cells expressing CdnLCc mutants unable to interact with the RNAP were cold-sensitive, suggesting that CdnLCc interaction with RNAP is especially required at lower than standard growth temperatures in C. crescentus. Our study indicates that despite limited sequence similarities and regulatory differences compared to its myco/myxobacterial homologs, CdnLCc may share similar biological functions, since it affects rRNA synthesis, probably by stabilizing open promoter-RNAP complexes. PMID:28233804

  4. p210(BCR-ABL) reprograms transformed and normal human megakaryocytic progenitor cells into erythroid cells and suppresses FLI-1 transcription.

    PubMed

    Buet, D; Raslova, H; Geay, J-F; Jarrier, P; Lazar, V; Turhan, A; Morlé, F; Vainchenker, W; Louache, F

    2007-05-01

    The BCR-ABL oncoprotein exhibits deregulated protein tyrosine kinase activity and is implicated in the pathogenesis of Philadelphia chromosome (Ph)-positive human leukemias. Here, we report that ectopic expression of p210(BCR-ABL) in the megakaryoblastic Mo7e cell line and in primary human CD34(+) progenitors trigger erythroid differentiation at the expense of megakaryocyte (MK) differentiation. Clonal culture of purified CD41(+)CD42(-) cells, a population highly enriched in MK progenitors, combined with the conditional expression of p210(BCR-ABL) tyrosine kinase activity by imatinib identified a true lineage reprogramming. In both Mo7e or CD41(+)CD42(-) cells transduced with p210(BCR-ABL), lineage switching was associated with a downregulation of the friend leukemia Integration 1 (FLI-1) transcription factor. Re-expression of FLI-1 in p210(BCR-ABL)-transduced Mo7e cells rescued the megakaryoblastic phenotype. Altogether, these results demonstrate that alteration of signal transduction via p210(BCR-ABL) reprograms MK cells into erythroid cells by a downregulation of FLI-1. In addition, our findings underscore the role of kinases in lineage choice and infidelity in pathology and suggest that downregulation of FLI-1 may have important implications in CML pathogenesis.

  5. Transcription Regulation in Archaea

    PubMed Central

    Gehring, Alexandra M.; Walker, Julie E.

    2016-01-01

    The known diversity of metabolic strategies and physiological adaptations of archaeal species to extreme environments is extraordinary. Accurate and responsive mechanisms to ensure that gene expression patterns match the needs of the cell necessitate regulatory strategies that control the activities and output of the archaeal transcription apparatus. Archaea are reliant on a single RNA polymerase for all transcription, and many of the known regulatory mechanisms employed for archaeal transcription mimic strategies also employed for eukaryotic and bacterial species. Novel mechanisms of transcription regulation have become apparent by increasingly sophisticated in vivo and in vitro investigations of archaeal species. This review emphasizes recent progress in understanding archaeal transcription regulatory mechanisms and highlights insights gained from studies of the influence of archaeal chromatin on transcription. PMID:27137495

  6. Laminin alpha 5, a major transcript of normal and malignant rat liver epithelial cells, is differentially expressed in developing and adult liver.

    PubMed

    Seebacher, T; Medina, J L; Bade, E G

    1997-11-25

    The laminin family of extracellular matrix glycoproteins plays a major role in cell migration and differentiation and in tumor cell invasion. As previously shown, the laminin deposited by normal and malignant rat liver epithelial cells in their extracellular matrix (ECM) and into their ECM migration tracks does not contain a typical (EHS-like) alpha 1 heavy chain. By RT-PCR screening we have now identified two alpha chains among a total of five additional laminin chains produced by these cells. Three of the newly identified chains were not previously known for the rat. Their sequences have been deposited in the EMBL nucleotide sequence data bank. The alpha 5 chain now identified is expressed at comparably high levels by both the normal and the malignant liver epithelial cells. The chain is also expressed in fetal liver together with the alpha 2 and beta 2 chains, but it is only vestigially expressed in the mature organ as shown by RT-PCR. These results suggest for alpha 5 a role in development and production of the chain by only a small subset of cells in adult liver. At the level of detection used, no changes were observed in regenerating liver after partial hepatectomy. In addition to the alpha 5 chain, the cultured cells express the beta 1 and beta 2 light chains, indicating the expression of more than one laminin isoform by the same cell line. The expression of the alpha 5 chain and of the other new non-EHS isoform chains was also analyzed in various tissues. The malignant liver epithelial cells, but not their nontumorigenic parental cells, also express, in addition to the alpha 5 chain the alpha 2 chain, which is expressed at high level by the NBT II bladder carcinoma cell line, suggesting a relationship with malignancy.

  7. Selection of internal reference genes for normalization of reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis in the rumen epithelium

    PubMed Central

    Die, Jose V.; Rowland, Lisa J.; Li, Robert; Oh, Sunghee; Li, Congjun; Connor, Erin E.; Ranilla, Maria-Jose

    2017-01-01

    The rumen is lined on the luminal side by a stratified squamous epithelium that is responsible for not only absorption, but also transport, extensive short-chain fatty acid (SCFA) metabolism and protection. Butyrate has been demonstrated to initiate the differentiation of the tissue following introduction of solid feed to the weaning neonate as well as affecting the metabolism of other nutrients and absorption of nutrients in in vitro experiments. The objective of the present study was to validate expression stability of eight putative reference genes bovine rumen, considering the intrinsic heterogeneity of bovine rumen with regard to different luminal characteristics due to direct infusion of butyrate to double the intra-ruminal content of the rumen liquor. Our focus was on identifying stable reference genes which are suitable to normalize real-time RT-qPCR experiments from rumen samples collected from clinical assays, irrespective of localization within the organ and the across physiological state. The most stably expressed genes included: ACTB, UXT, DBNDD2, RPS9, DDX54 and HMBS. Their high stability values suggest these reference genes will facilitate better evaluation of variation of across an array of conditions including: localization within the rumen, differences among cattle fed an array of rations, as well as response to development in the weaning animal. Moreover, we anticipate these reference genes may be useful for expression studies in other ruminants. PMID:28234977

  8. The correlation of TRPM1 (Melastatin) mRNA expression with microphthalmia-associated transcription factor (MITF) and other melanogenesis-related proteins in normal and pathological skin, hair follicles and melanocytic nevi

    PubMed Central

    Lu, Song; Slominski, Andrzej; Yang, Sung-Eun; Sheehan, Christine; Ross, Jeffrey; Carlson, J. Andrew

    2010-01-01

    Background Melastatin (TRPM1), a.k.a. transient receptor potential cation channel, subfamily M, member 1 (TRPM-1) regulates melanocyte differentiation and proliferation. TRPM1 is transcriptionally regulated by the essential melanocyte transcription factor MITF (microphthalmia-associated transcription factor). For the most part, MITF expression is preserved during melanoma progression, while TRPM1 mRNA expression decreases or is completely lost. The loss of TRPM1 is associated with melanomas that are more aggressive. Objective To assess the relationship between TRPM1 mRNA expression and the expression of MITF and nine other markers of melanocytes and melanin-related proteins by immunohistochemistry in normal skin, scars, hair follicles and ordinary melanocytic nevi. Methods Samples of normal skin (n = 102; from tumor excisions and plastic procedures), scars (n = 5; from re-excision specimens) and compound melanocytic nevi (n = 4) were evaluated for the presence of TRPM1 mRNA transcripts as detected by chromogenic in situ hybridization (CISH). Immunohistochemical techniques were used to detect melanin-related proteins including: MITF, S100 protein, Mart-1, tyrosinase, Mel5, HMB45, tyrosinase-related protein-1 (TRP1), TRP2 and α-melanocyte stimulating hormone (αMSH). The labeling index (LI) was defined as the number of intraepidermal cells expressing mRNA or protein per one hundred basal keratinocytes. Results A wide range of LI was found for all markers (0–33 positive cells/100 keratinocytes). When these LI were compared, no significant differences in the expression of MITF, S100, Mart1, tyrosinase proteins and TRPM1 mRNA were identified. The LI for TRPM1 mRNA expression ranged from 74% of that for MITF to 86% for tyrosinase. The LI for TRP-1, TRP-2 and Mel5 was similar to that of TRPM1, while HMB-45 had a significantly lower LI than all other markers. TRPM1 mRNA correlated most tightly with MITF and tyrosinase expression (r = 0.81 and 0.68, respectively, both p

  9. Acute intermittent porphyria: identification and expression of exonic mutations in the hydroxymethylbilane synthase gene. An initiation codon missense mutation in the housekeeping transcript causes "variant acute intermittent porphyria" with normal expression of the erythroid-specific enzyme.

    PubMed Central

    Chen, C H; Astrin, K H; Lee, G; Anderson, K E; Desnick, R J

    1994-01-01

    Acute intermittent porphyria (AIP), an autosomal dominant inborn error, results from the half-normal activity of the heme biosynthetic enzyme, hydroxymethylbilane synthase (EC 4.3.1.8). Diagnosis of AIP heterozygotes is essential to prevent acute, life-threatening neurologic attacks by avoiding various precipitating factors. Since biochemical diagnosis is problematic, the identification of hydroxymethylbilane synthase mutations has facilitated the detection of AIP heterozygotes. Molecular analyses of unrelated AIP patients revealed six exonic mutations: an initiating methionine to isoleucine substitution (M1I) in a patient with variant AIP, which precluded translation of the housekeeping, but not the erythroid-specific isozyme; four missense mutations in classical AIP patients, V93F, R116W, R201W, C247F; and a nonsense mutation W283X in a classical AIP patient, which truncated the housekeeping and erythroid-specific isozymes. Each mutation was confirmed in genomic DNA from family members. The W283X lesion was found in another unrelated AIP family. Expression of each mutation in Escherichia coli revealed that R201W, C247F, and W283X had residual activity. In vitro transcription/translation studies indicated that the M1I allele produced only the erythroid-specific enzyme, while the other mutant alleles encoded both isozymes. These mutations provide insight into the molecular pathology of classic and variant AIP and facilitate molecular diagnosis in AIP families. Images PMID:7962538

  10. Accurate Finite Difference Algorithms

    NASA Technical Reports Server (NTRS)

    Goodrich, John W.

    1996-01-01

    Two families of finite difference algorithms for computational aeroacoustics are presented and compared. All of the algorithms are single step explicit methods, they have the same order of accuracy in both space and time, with examples up to eleventh order, and they have multidimensional extensions. One of the algorithm families has spectral like high resolution. Propagation with high order and high resolution algorithms can produce accurate results after O(10(exp 6)) periods of propagation with eight grid points per wavelength.

  11. Accurate monotone cubic interpolation

    NASA Technical Reports Server (NTRS)

    Huynh, Hung T.

    1991-01-01

    Monotone piecewise cubic interpolants are simple and effective. They are generally third-order accurate, except near strict local extrema where accuracy degenerates to second-order due to the monotonicity constraint. Algorithms for piecewise cubic interpolants, which preserve monotonicity as well as uniform third and fourth-order accuracy are presented. The gain of accuracy is obtained by relaxing the monotonicity constraint in a geometric framework in which the median function plays a crucial role.

  12. Accurate Guitar Tuning by Cochlear Implant Musicians

    PubMed Central

    Lu, Thomas; Huang, Juan; Zeng, Fan-Gang

    2014-01-01

    Modern cochlear implant (CI) users understand speech but find difficulty in music appreciation due to poor pitch perception. Still, some deaf musicians continue to perform with their CI. Here we show unexpected results that CI musicians can reliably tune a guitar by CI alone and, under controlled conditions, match simultaneously presented tones to <0.5 Hz. One subject had normal contralateral hearing and produced more accurate tuning with CI than his normal ear. To understand these counterintuitive findings, we presented tones sequentially and found that tuning error was larger at ∼30 Hz for both subjects. A third subject, a non-musician CI user with normal contralateral hearing, showed similar trends in performance between CI and normal hearing ears but with less precision. This difference, along with electric analysis, showed that accurate tuning was achieved by listening to beats rather than discriminating pitch, effectively turning a spectral task into a temporal discrimination task. PMID:24651081

  13. Accurate guitar tuning by cochlear implant musicians.

    PubMed

    Lu, Thomas; Huang, Juan; Zeng, Fan-Gang

    2014-01-01

    Modern cochlear implant (CI) users understand speech but find difficulty in music appreciation due to poor pitch perception. Still, some deaf musicians continue to perform with their CI. Here we show unexpected results that CI musicians can reliably tune a guitar by CI alone and, under controlled conditions, match simultaneously presented tones to <0.5 Hz. One subject had normal contralateral hearing and produced more accurate tuning with CI than his normal ear. To understand these counterintuitive findings, we presented tones sequentially and found that tuning error was larger at ∼ 30 Hz for both subjects. A third subject, a non-musician CI user with normal contralateral hearing, showed similar trends in performance between CI and normal hearing ears but with less precision. This difference, along with electric analysis, showed that accurate tuning was achieved by listening to beats rather than discriminating pitch, effectively turning a spectral task into a temporal discrimination task.

  14. On numerically accurate finite element

    NASA Technical Reports Server (NTRS)

    Nagtegaal, J. C.; Parks, D. M.; Rice, J. R.

    1974-01-01

    A general criterion for testing a mesh with topologically similar repeat units is given, and the analysis shows that only a few conventional element types and arrangements are, or can be made suitable for computations in the fully plastic range. Further, a new variational principle, which can easily and simply be incorporated into an existing finite element program, is presented. This allows accurate computations to be made even for element designs that would not normally be suitable. Numerical results are given for three plane strain problems, namely pure bending of a beam, a thick-walled tube under pressure, and a deep double edge cracked tensile specimen. The effects of various element designs and of the new variational procedure are illustrated. Elastic-plastic computation at finite strain are discussed.

  15. Transcription and cancer.

    PubMed Central

    Cox, P. M.; Goding, C. R.

    1991-01-01

    The normal growth, development and function of an organism requires precise and co-ordinated control of gene expression. A major part of this control is exerted by regulating messenger RNA (mRNA) production and involves complex interactions between an array of transcriptionally active proteins and specific regulatory DNA sequences. The combination of such proteins and DNA sequences is specific for given gene or group of genes in a particular cell type and the proteins regulating the same gene may vary between cell types. In addition the expression or activity of these regulatory proteins may be modified depending on the state of differentiation of a cell or in response to an external stimulus. Thus, the differentiation of embryonic cells into diverse tissues is achieved and the mature structure and function of the organism is maintained. This review focusses on the role of perturbations of these transcriptional controls in neoplasia. Deregulation of transcription may result in the failure to express genes responsible for cellular differentiation, or alternatively, in the transcription of genes involved in cell division, through the inappropriate expression or activation of positively acting transcription factors and nuclear oncogenes. Whether the biochemical abnormalities that lead to the disordered growth and differentiation of a malignant tumour affect cell surface receptors, membrane or cytoplasmic signalling proteins or nuclear transcription factors, the end result is the inappropriate expression of some genes and failure to express others. Current research is starting to elucidate which of the elements of this complicated system are important in neoplasia. PMID:1645561

  16. Accurate quantum chemical calculations

    NASA Technical Reports Server (NTRS)

    Bauschlicher, Charles W., Jr.; Langhoff, Stephen R.; Taylor, Peter R.

    1989-01-01

    An important goal of quantum chemical calculations is to provide an understanding of chemical bonding and molecular electronic structure. A second goal, the prediction of energy differences to chemical accuracy, has been much harder to attain. First, the computational resources required to achieve such accuracy are very large, and second, it is not straightforward to demonstrate that an apparently accurate result, in terms of agreement with experiment, does not result from a cancellation of errors. Recent advances in electronic structure methodology, coupled with the power of vector supercomputers, have made it possible to solve a number of electronic structure problems exactly using the full configuration interaction (FCI) method within a subspace of the complete Hilbert space. These exact results can be used to benchmark approximate techniques that are applicable to a wider range of chemical and physical problems. The methodology of many-electron quantum chemistry is reviewed. Methods are considered in detail for performing FCI calculations. The application of FCI methods to several three-electron problems in molecular physics are discussed. A number of benchmark applications of FCI wave functions are described. Atomic basis sets and the development of improved methods for handling very large basis sets are discussed: these are then applied to a number of chemical and spectroscopic problems; to transition metals; and to problems involving potential energy surfaces. Although the experiences described give considerable grounds for optimism about the general ability to perform accurate calculations, there are several problems that have proved less tractable, at least with current computer resources, and these and possible solutions are discussed.

  17. BIOACCESSIBILITY TESTS ACCURATELY ESTIMATE ...

    EPA Pesticide Factsheets

    Hazards of soil-borne Pb to wild birds may be more accurately quantified if the bioavailability of that Pb is known. To better understand the bioavailability of Pb to birds, we measured blood Pb concentrations in Japanese quail (Coturnix japonica) fed diets containing Pb-contaminated soils. Relative bioavailabilities were expressed by comparison with blood Pb concentrations in quail fed a Pb acetate reference diet. Diets containing soil from five Pb-contaminated Superfund sites had relative bioavailabilities from 33%-63%, with a mean of about 50%. Treatment of two of the soils with P significantly reduced the bioavailability of Pb. The bioaccessibility of the Pb in the test soils was then measured in six in vitro tests and regressed on bioavailability. They were: the “Relative Bioavailability Leaching Procedure” (RBALP) at pH 1.5, the same test conducted at pH 2.5, the “Ohio State University In vitro Gastrointestinal” method (OSU IVG), the “Urban Soil Bioaccessible Lead Test”, the modified “Physiologically Based Extraction Test” and the “Waterfowl Physiologically Based Extraction Test.” All regressions had positive slopes. Based on criteria of slope and coefficient of determination, the RBALP pH 2.5 and OSU IVG tests performed very well. Speciation by X-ray absorption spectroscopy demonstrated that, on average, most of the Pb in the sampled soils was sorbed to minerals (30%), bound to organic matter 24%, or present as Pb sulfate 18%. Ad

  18. Accurate spectral color measurements

    NASA Astrophysics Data System (ADS)

    Hiltunen, Jouni; Jaeaeskelaeinen, Timo; Parkkinen, Jussi P. S.

    1999-08-01

    Surface color measurement is of importance in a very wide range of industrial applications including paint, paper, printing, photography, textiles, plastics and so on. For a demanding color measurements spectral approach is often needed. One can measure a color spectrum with a spectrophotometer using calibrated standard samples as a reference. Because it is impossible to define absolute color values of a sample, we always work with approximations. The human eye can perceive color difference as small as 0.5 CIELAB units and thus distinguish millions of colors. This 0.5 unit difference should be a goal for the precise color measurements. This limit is not a problem if we only want to measure the color difference of two samples, but if we want to know in a same time exact color coordinate values accuracy problems arise. The values of two instruments can be astonishingly different. The accuracy of the instrument used in color measurement may depend on various errors such as photometric non-linearity, wavelength error, integrating sphere dark level error, integrating sphere error in both specular included and specular excluded modes. Thus the correction formulas should be used to get more accurate results. Another question is how many channels i.e. wavelengths we are using to measure a spectrum. It is obvious that the sampling interval should be short to get more precise results. Furthermore, the result we get is always compromise of measuring time, conditions and cost. Sometimes we have to use portable syste or the shape and the size of samples makes it impossible to use sensitive equipment. In this study a small set of calibrated color tiles measured with the Perkin Elmer Lamda 18 and the Minolta CM-2002 spectrophotometers are compared. In the paper we explain the typical error sources of spectral color measurements, and show which are the accuracy demands a good colorimeter should have.

  19. Mapping yeast transcriptional networks.

    PubMed

    Hughes, Timothy R; de Boer, Carl G

    2013-09-01

    The term "transcriptional network" refers to the mechanism(s) that underlies coordinated expression of genes, typically involving transcription factors (TFs) binding to the promoters of multiple genes, and individual genes controlled by multiple TFs. A multitude of studies in the last two decades have aimed to map and characterize transcriptional networks in the yeast Saccharomyces cerevisiae. We review the methodologies and accomplishments of these studies, as well as challenges we now face. For most yeast TFs, data have been collected on their sequence preferences, in vivo promoter occupancy, and gene expression profiles in deletion mutants. These systematic studies have led to the identification of new regulators of numerous cellular functions and shed light on the overall organization of yeast gene regulation. However, many yeast TFs appear to be inactive under standard laboratory growth conditions, and many of the available data were collected using techniques that have since been improved. Perhaps as a consequence, comprehensive and accurate mapping among TF sequence preferences, promoter binding, and gene expression remains an open challenge. We propose that the time is ripe for renewed systematic efforts toward a complete mapping of yeast transcriptional regulatory mechanisms.

  20. An Accurate, Simplified Model Intrabeam Scattering

    SciTech Connect

    Bane, Karl LF

    2002-05-23

    Beginning with the general Bjorken-Mtingwa solution for intrabeam scattering (IBS) we derive an accurate, greatly simplified model of IBS, valid for high energy beams in normal storage ring lattices. In addition, we show that, under the same conditions, a modified version of Piwinski's IBS formulation (where {eta}{sub x,y}{sup 2}/{beta}{sub x,y} has been replaced by {Eta}{sub x,y}) asymptotically approaches the result of Bjorken-Mtingwa.

  1. An accurate registration technique for distorted images

    NASA Technical Reports Server (NTRS)

    Delapena, Michele; Shaw, Richard A.; Linde, Peter; Dravins, Dainis

    1990-01-01

    Accurate registration of International Ultraviolet Explorer (IUE) images is crucial because the variability of the geometrical distortions that are introduced by the SEC-Vidicon cameras ensures that raw science images are never perfectly aligned with the Intensity Transfer Functions (ITFs) (i.e., graded floodlamp exposures that are used to linearize and normalize the camera response). A technique for precisely registering IUE images which uses a cross correlation of the fixed pattern that exists in all raw IUE images is described.

  2. Triple-layer dissection of the lung adenocarcinoma transcriptome – regulation at the gene, transcript, and exon levels

    PubMed Central

    Hsu, Min-Kung; Wu, I-Ching; Cheng, Ching-Chia; Su, Jen-Liang; Hsieh, Chang-Huain; Lin, Yeong-Shin; Chen, Feng-Chi

    2015-01-01

    Lung adenocarcinoma is one of the most deadly human diseases. However, the molecular mechanisms underlying this disease, particularly RNA splicing, have remained underexplored. Here, we report a triple-level (gene-, transcript-, and exon-level) analysis of lung adenocarcinoma transcriptomes from 77 paired tumor and normal tissues, as well as an analysis pipeline to overcome genetic variability for accurate differentiation between tumor and normal tissues. We report three major results. First, more than 5,000 differentially expressed transcripts/exonic regions occur repeatedly in lung adenocarcinoma patients. These transcripts/exonic regions are enriched in nicotine metabolism and ribosomal functions in addition to the pathways enriched for differentially expressed genes (cell cycle, extracellular matrix receptor interaction, and axon guidance). Second, classification models based on rationally selected transcripts or exonic regions can reach accuracies of 0.93 to 1.00 in differentiating tumor from normal tissues. Of the 28 selected exonic regions, 26 regions correspond to alternative exons located in such regulators as tumor suppressor (GDF10), signal receptor (LYVE1), vascular-specific regulator (RASIP1), ubiquitination mediator (RNF5), and transcriptional repressor (TRIM27). Third, classification systems based on 13 to 14 differentially expressed genes yield accuracies near 100%. Genes selected by both detection methods include C16orf59, DAP3, ETV4, GABARAPL1, PPAR, RADIL, RSPO1, SERTM1, SRPK1, ST6GALNAC6, and TNXB. Our findings imply a multilayered lung adenocarcinoma regulome in which transcript-/exon-level regulation may be dissociated from gene-level regulation. Our described method may be used to identify potentially important genes/transcripts/exonic regions for the tumorigenesis of lung adenocarcinoma and to construct accurate tumor vs. normal classification systems for this disease. PMID:26356813

  3. Normal anatomy of the skull base.

    PubMed

    Lustrin, E S; Robertson, R L; Tilak, S

    1994-08-01

    CT and MR imaging increasingly are being used for the evaluation of the skull base. New innovative techniques have revolutionized radiologic understanding of normal skull base anatomy. Thus, normal anatomic relationships with radiographic correlation are vital for accurate pathologic assessment.

  4. Isolation and characterization of transcription fidelity mutants.

    PubMed

    Strathern, Jeffrey N; Jin, Ding Jun; Court, Donald L; Kashlev, Mikhail

    2012-07-01

    Accurate transcription is an essential step in maintaining genetic information. Error-prone transcription has been proposed to contribute to cancer, aging, adaptive mutagenesis, and mutagenic evolution of retroviruses and retrotransposons. The mechanisms controlling transcription fidelity and the biological consequences of transcription errors are poorly understood. Because of the transient nature of mRNAs and the lack of reliable experimental systems, the identification and characterization of defects that increase transcription errors have been particularly challenging. In this review we describe novel genetic screens for the isolation of fidelity mutants in both Saccharomyces cerevisiae and Escherichia coli RNA polymerases. We obtained and characterized two distinct classes of mutants altering NTP misincorporation and transcription slippage both in vivo and in vitro. Our study not only validates the genetic schemes for the isolation of RNA polymerase mutants that alter fidelity, but also sheds light on the mechanism of transcription accuracy. This article is part of a Special Issue entitled: Chromatin in time and space.

  5. Metabolic labeling and recovery of nascent RNA to accurately quantify mRNA stability.

    PubMed

    Russo, Joseph; Heck, Adam M; Wilusz, Jeffrey; Wilusz, Carol J

    2017-02-20

    Changes in the rate of mRNA decay are closely coordinated with transcriptional changes and together these events have profound effects on gene expression during development and disease. Traditional approaches to assess mRNA decay have relied on inhibition of transcription, which can alter mRNA decay rates and confound interpretation. More recently, metabolic labeling combined with chemical modification and fractionation of labeled RNAs has allowed the isolation of nascent transcripts and the subsequent calculation of mRNA decay rates. This approach has been widely adopted for measuring mRNA half-lives on a global scale, but has proven challenging to use for analysis of single genes. We present a series of normalization and quality assurance steps to be used in combination with 4-thiouridine pulse labeling of cultured eukaryotic cells. Importantly, we demonstrate how the relative amount of 4sU-labeled nascent RNA influences accurate quantification. The approach described facilitates reproducible measurement of individual mRNA half-lives using 4-thiouridine and could be adapted for use with other nucleoside analogs.

  6. Writing an Independently Composed Sentence by Spanish-Speaking Children With and Without Poor Transcription Skills: A Writing-Level Match Design.

    PubMed

    García, Eduardo; Crespo, Patricia; Bermúdez, Ivana

    2016-03-10

    The main objective of this research was to analyze the impact of transcription skills of Spanish writers when writing an independently composed sentence within a writing-level design. The free-writing sentence task from the Early Grade Writing Assessment (Jiménez, in press) was used to examine the production, accuracy, speed, syntactic complexity, quality, and fluency of children with poor transcription skills (PTS). The results showed that there were significant differences between children with PTS and peers who had good transcription skills. The PTS group members were less accurate, slower, and less fluent or even dysfluent. Furthermore, their sentences were less complex and contained lower quality content. These results suggest that transcription skills play a crucial role in early written expression in Spanish, and poor transcription abilities hamper the acquisition and normal development of sentence composition.

  7. Multivariate normality

    NASA Technical Reports Server (NTRS)

    Crutcher, H. L.; Falls, L. W.

    1976-01-01

    Sets of experimentally determined or routinely observed data provide information about the past, present and, hopefully, future sets of similarly produced data. An infinite set of statistical models exists which may be used to describe the data sets. The normal distribution is one model. If it serves at all, it serves well. If a data set, or a transformation of the set, representative of a larger population can be described by the normal distribution, then valid statistical inferences can be drawn. There are several tests which may be applied to a data set to determine whether the univariate normal model adequately describes the set. The chi-square test based on Pearson's work in the late nineteenth and early twentieth centuries is often used. Like all tests, it has some weaknesses which are discussed in elementary texts. Extension of the chi-square test to the multivariate normal model is provided. Tables and graphs permit easier application of the test in the higher dimensions. Several examples, using recorded data, illustrate the procedures. Tests of maximum absolute differences, mean sum of squares of residuals, runs and changes of sign are included in these tests. Dimensions one through five with selected sample sizes 11 to 101 are used to illustrate the statistical tests developed.

  8. A unified model for yeast transcript definition.

    PubMed

    de Boer, Carl G; van Bakel, Harm; Tsui, Kyle; Li, Joyce; Morris, Quaid D; Nislow, Corey; Greenblatt, Jack F; Hughes, Timothy R

    2014-01-01

    Identifying genes in the genomic context is central to a cell's ability to interpret the genome. Yet, in general, the signals used to define eukaryotic genes are poorly described. Here, we derived simple classifiers that identify where transcription will initiate and terminate using nucleic acid sequence features detectable by the yeast cell, which we integrate into a Unified Model (UM) that models transcription as a whole. The cis-elements that denote where transcription initiates function primarily through nucleosome depletion, and, using a synthetic promoter system, we show that most of these elements are sufficient to initiate transcription in vivo. Hrp1 binding sites are the major characteristic of terminators; these binding sites are often clustered in terminator regions and can terminate transcription bidirectionally. The UM predicts global transcript structure by modeling transcription of the genome using a hidden Markov model whose emissions are the outputs of the initiation and termination classifiers. We validated the novel predictions of the UM with available RNA-seq data and tested it further by directly comparing the transcript structure predicted by the model to the transcription generated by the cell for synthetic DNA segments of random design. We show that the UM identifies transcription start sites more accurately than the initiation classifier alone, indicating that the relative arrangement of promoter and terminator elements influences their function. Our model presents a concrete description of how the cell defines transcript units, explains the existence of nongenic transcripts, and provides insight into genome evolution.

  9. A unified model for yeast transcript definition

    PubMed Central

    de Boer, Carl G.; van Bakel, Harm; Tsui, Kyle; Li, Joyce; Morris, Quaid D.; Nislow, Corey; Greenblatt, Jack F.; Hughes, Timothy R.

    2014-01-01

    Identifying genes in the genomic context is central to a cell's ability to interpret the genome. Yet, in general, the signals used to define eukaryotic genes are poorly described. Here, we derived simple classifiers that identify where transcription will initiate and terminate using nucleic acid sequence features detectable by the yeast cell, which we integrate into a Unified Model (UM) that models transcription as a whole. The cis-elements that denote where transcription initiates function primarily through nucleosome depletion, and, using a synthetic promoter system, we show that most of these elements are sufficient to initiate transcription in vivo. Hrp1 binding sites are the major characteristic of terminators; these binding sites are often clustered in terminator regions and can terminate transcription bidirectionally. The UM predicts global transcript structure by modeling transcription of the genome using a hidden Markov model whose emissions are the outputs of the initiation and termination classifiers. We validated the novel predictions of the UM with available RNA-seq data and tested it further by directly comparing the transcript structure predicted by the model to the transcription generated by the cell for synthetic DNA segments of random design. We show that the UM identifies transcription start sites more accurately than the initiation classifier alone, indicating that the relative arrangement of promoter and terminator elements influences their function. Our model presents a concrete description of how the cell defines transcript units, explains the existence of nongenic transcripts, and provides insight into genome evolution. PMID:24170600

  10. Obtaining accurate translations from expressed sequence tags.

    PubMed

    Wasmuth, James; Blaxter, Mark

    2009-01-01

    The genomes of an increasing number of species are being investigated through the generation of expressed sequence tags (ESTs). However, ESTs are prone to sequencing errors and typically define incomplete transcripts, making downstream annotation difficult. Annotation would be greatly improved with robust polypeptide translations. Many current solutions for EST translation require a large number of full-length gene sequences for training purposes, a resource that is not available for the majority of EST projects. As part of our ongoing EST programs investigating these "neglected" genomes, we have developed a polypeptide prediction pipeline, prot4EST. It incorporates freely available software to produce final translations that are more accurate than those derived from any single method. We describe how this integrated approach goes a long way to overcoming the deficit in training data.

  11. WMAP normalization of inflationary cosmologies

    SciTech Connect

    Liddle, Andrew R.; Parkinson, David; Mukherjee, Pia; Leach, Samuel M.

    2006-10-15

    We use the three-year WMAP observations to determine the normalization of the matter power spectrum in inflationary cosmologies. In this context, the quantity of interest is not the normalization marginalized over all parameters, but rather the normalization as a function of the inflationary parameters n{sub S} and r with marginalization over the remaining cosmological parameters. We compute this normalization and provide an accurate fitting function. The statistical uncertainty in the normalization is 3%, roughly half that achieved by COBE. We use the k-l relation for the standard cosmological model to identify the pivot scale for the WMAP normalization. We also quote the inflationary energy scale corresponding to the WMAP normalization.

  12. Influence of CGG-repeat length upon FMR1 transcription and translation

    SciTech Connect

    Warren, S.T.; Zhang, F.; Lokey, L.K.

    1994-09-01

    Fragile X syndrome is the result of the expansion of a 5{prime} untranslated CGG-repeat in the FMR1 gene. In penetrant individuals the repeat is typically >230 triplets and is abnormally methylated and transcriptionally silent while premutation alleles generally contain 60-200 repeats. Consistent with the lack of penetrance among premutation carriers, FMR1 transcription and translation were found similar among normal and premutation cell lines, suggesting little influence of CGG-repeat length of <110 repeats on FMRP expression. However, similar studies of a mildly affected mosaic male with a predominantly hypomethylated allele exhibiting a mode of 300 repeats indicates an influence of larger repeats upon FMRP translation. Lymphoblasts and fibroblasts from this individual revealed normal transcription but {approximately}30% of normal FMRP levels. From an explant fibroblast culture, individual colonies were isolated and clones containing 57, 168, 182, 207, 266 and 285 hypomethylated repeats were identified. Quantitative RNase protection revealed normal steady state levels of FMR1 mRNA in these clonal cultures and RT-PCR of the repeat showed accurate transcription through the repeats. FMRP levels, quantitated using monoclonal antibody, were normal up to 182 repeats and reduced in clones containing larger repeat lengths with very little FMRP detected in the 285 repeat cells. Sucrose fractionation of ribosomes followed by RNase protection showed FMR1 mRNA associated with polyribosomes in control cells while transcripts containing 266 repeats localized predominantly to the 40-80S subunit fraction. These data indicate an inability of the ribosome to scan through lengthy trinucleotide repeats. This has implications regarding the molecular mechanisms of fragile X syndrome and other disorders due to trinucleotide repeat expansions.

  13. Accurate Evaluation of Quantum Integrals

    NASA Technical Reports Server (NTRS)

    Galant, D. C.; Goorvitch, D.; Witteborn, Fred C. (Technical Monitor)

    1995-01-01

    Combining an appropriate finite difference method with Richardson's extrapolation results in a simple, highly accurate numerical method for solving a Schrodinger's equation. Important results are that error estimates are provided, and that one can extrapolate expectation values rather than the wavefunctions to obtain highly accurate expectation values. We discuss the eigenvalues, the error growth in repeated Richardson's extrapolation, and show that the expectation values calculated on a crude mesh can be extrapolated to obtain expectation values of high accuracy.

  14. Genome-wide identification and characterization of reference genes with different transcript abundances for Streptomyces coelicolor

    PubMed Central

    Li, Shanshan; Wang, Weishan; Li, Xiao; Fan, Keqiang; Yang, Keqian

    2015-01-01

    The lack of reliable reference genes (RGs) in the genus Streptomyces hampers effort to obtain the precise data of transcript levels. To address this issue, we aimed to identify reliable RGs in the model organism Streptomyces coelicolor. A pool of potential RGs containing 1,471 genes was first identified by determining the intersection of genes with stable transcript levels from four time-series transcriptome microarray datasets of S. coelicolor M145 cultivated in different conditions. Then, following a strict rational selection scheme including homology analysis, disturbance analysis, function analysis and transcript abundance analysis, 13 candidates were selected from the 1,471 genes. Based on real-time quantitative reverse transcription PCR assays, SCO0710, SCO6185, SCO1544, SCO3183 and SCO4758 were identified as the top five genes with the most stable transcript levels among the 13 candidates. Further analyses showed these five genes also maintained stable transcript levels in different S. coelicolor strains, as well as in Streptomyces avermitilis MA-4680 and Streptomyces clavuligerus NRRL 3585, suggesting they could fulfill the requirements of accurate data normalization in streptomycetes. Moreover, the systematic strategy employed in this work could be used for reference in other microorganism to select reliable RGs. PMID:26527303

  15. Normal-reflection image

    SciTech Connect

    Huang, L.; Fehler, Michael C.

    2003-01-01

    Common-angle wave-equation migration using the double-square-root is generally less accurate than the common-shot migration because the wavefield continuation equation for thc former involves additional approximations compared to that for the latter. We present a common-angle wave-equation migration that has the same accuracy as common-shot wave-equation migration. An image obtained from common-angle migration is a four- to five-dimensional output volume for 3D cases. We propose a normal-reflection imaging condition for common-angle migration to produce a 3D output volume for 3D migration. The image is closely related to the normal-reflection coefficients at interfaces. This imaging condition will allow amplitude-preserving migration to generate an image with clear physical meaning.

  16. Transcriptomic profiles differentiate normal rectal epithelium and adenocarcinoma.

    PubMed

    Hogan, J; Dejulius, K; Liu, X; Coffey, J C; Kalady, M F

    2015-05-01

    Adenocarcinoma is a histologic diagnosis based on subjective findings. Transcriptional profiles have been used to differentiate normal tissue from disease and could provide a means of identifying malignancy. The goal of this study was to generate and test transcriptomic profiles that differentiate normal from adenocarcinomatous rectum. Comparisons were made between cDNA microarrays derived from normal epithelium and rectal adenocarcinoma. Results were filtered according to standard deviation to retain only highly dysregulated genes. Genes differentially expressed between cancer and normal tissue on two-groups t test (P < 0.05, Bonferroni P value adjustment) were further analyzed. Genes were rank ordered in terms of descending fold change. For each comparison (tumor versus normal epithelium), those 5 genes with the greatest positive fold change were grouped in a classifier. Five separate tests were applied to evaluate the discriminatory capacity of each classifier. Genetic classifiers derived comparing normal epithelium with malignant rectal epithelium from pooled stages had a mean sensitivity and specificity of 99.6% and 98.2%, respectively. The classifiers derived from comparing normal and stage I cancer had comparable mean sensitivities and specificities (97% and 98%, respectively). Areas under the summary receiver-operator characteristic curves for each classifier were 0.981 and 0.972, respectively. One gene was common to both classifiers. Classifiers were tested in an independent Gene Expression Omnibus-derived dataset. Both classifiers retained their predictive properties. Transcriptomic profiles comprising as few as 5 genes are highly accurate in differentiating normal from adenocarcinomatous rectal epithelium, including early-stage disease.

  17. Prognostic Importance of MN1 Transcript Levels, and Biologic Insights From MN1-Associated Gene and MicroRNA Expression Signatures in Cytogenetically Normal Acute Myeloid Leukemia: A Cancer and Leukemia Group B Study

    PubMed Central

    Langer, Christian; Marcucci, Guido; Holland, Kelsi B.; Radmacher, Michael D.; Maharry, Kati; Paschka, Peter; Whitman, Susan P.; Mrózek, Krzysztof; Baldus, Claudia D.; Vij, Ravi; Powell, Bayard L.; Carroll, Andrew J.; Kolitz, Jonathan E.; Caligiuri, Michael A.; Larson, Richard A.; Bloomfield, Clara D.

    2009-01-01

    Purpose To determine the prognostic importance of the meningioma 1 (MN1) gene expression levels in the context of other predictive molecular markers, and to derive MN1 associated gene– and microRNA–expression profiles in cytogenetically normal acute myeloid leukemia (CN-AML). Patients and Methods MN1 expression was measured in 119 untreated primary CN-AML adults younger than 60 years by real-time reverse-transcriptase polymerase chain reaction. Patients were also tested for FLT3, NPM1, CEBPA, and WT1 mutations, MLL partial tandem duplications, and BAALC and ERG expression. Gene- and microRNA-expression profiles were attained by performing genome-wide microarray assays. Patients were intensively treated on two first-line Cancer and Leukemia Group B clinical trials. Results Higher MN1 expression associated with NPM1 wild-type (P < .001), increased BAALC expression (P = .004), and less extramedullary involvement (P = .01). In multivariable analyses, higher MN1 expression associated with a lower complete remission rate (P = .005) after adjustment for WBC; shorter disease-free survival (P = .01) after adjustment for WT1 mutations, FLT3 internal tandem duplications (FLT3-ITD), and high ERG expression; and shorter survival (P = .04) after adjustment for WT1 and NPM1 mutations, FLT3-ITD, and WBC. Gene- and microRNA-expression profiles suggested that high MN1 expressers share features with high BAALC expressers and patients with wild-type NPM1. Higher MN1 expression also appears to be associated with genes and microRNAs that are active in aberrant macrophage/monocytoid function and differentiation. Conclusion MN1 expression independently predicts outcome in CN-AML patients. The MN1 gene- and microRNA-expression signatures suggest biologic features that could be exploited as therapeutic targets. PMID:19451432

  18. DBD: a transcription factor prediction database.

    PubMed

    Kummerfeld, Sarah K; Teichmann, Sarah A

    2006-01-01

    Regulation of gene expression influences almost all biological processes in an organism; sequence-specific DNA-binding transcription factors are critical to this control. For most genomes, the repertoire of transcription factors is only partially known. Hitherto transcription factor identification has been largely based on genome annotation pipelines that use pairwise sequence comparisons, which detect only those factors similar to known genes, or on functional classification schemes that amalgamate many types of proteins into the category of 'transcription factor'. Using a novel transcription factor identification method, the DBD transcription factor database fills this void, providing genome-wide transcription factor predictions for organisms from across the tree of life. The prediction method behind DBD identifies sequence-specific DNA-binding transcription factors through homology using profile hidden Markov models (HMMs) of domains. Thus, it is limited to factors that are homologus to those HMMs. The collection of HMMs is taken from two existing databases (Pfam and SUPERFAMILY), and is limited to models that exclusively detect transcription factors that specifically recognize DNA sequences. It does not include basal transcription factors or chromatin-associated proteins, for instance. Based on comparison with experimentally verified annotation, the prediction procedure is between 95% and 99% accurate. Between one quarter and one-half of our genome-wide predicted transcription factors represent previously uncharacterized proteins. The DBD (www.transcriptionfactor.org) consists of predicted transcription factor repertoires for 150 completely sequenced genomes, their domain assignments and the hand curated list of DNA-binding domain HMMs. Users can browse, search or download the predictions by genome, domain family or sequence identifier, view families of transcription factors based on domain architecture and receive predictions for a protein sequence.

  19. Swinger RNAs with sharp switches between regular transcription and transcription systematically exchanging ribonucleotides: Case studies.

    PubMed

    Seligmann, Hervé

    2015-09-01

    During RNA transcription, DNA nucleotides A,C,G, T are usually matched by ribonucleotides A, C, G and U. However occasionally, this rule does not apply: transcript-DNA homologies are detectable only assuming systematic exchanges between ribonucleotides. Nine symmetric (X ↔ Y, e.g. A ↔ C) and fourteen asymmetric (X ↔ Y ↔ Z, e.g. A ↔ C ↔ G) exchanges exist, called swinger transcriptions. Putatively, polymerases occasionally stabilize in unspecified swinger conformations, possibly similar to transient conformations causing punctual misinsertions. This predicts chimeric transcripts, part regular, part swinger-transformed, reflecting polymerases switching to swinger polymerization conformation(s). Four chimeric Genbank transcripts (three from human mitochondrion and one murine cytosolic) are described here: (a) the 5' and 3' extremities reflect regular polymerization, the intervening sequence exchanges systematically between ribonucleotides (swinger rule G ↔ U, transcript (1), with sharp switches between regular and swinger sequences; (b) the 5' half is 'normal', the 3' half systematically exchanges ribonucleotides (swinger rule C ↔ G, transcript (2), with an intercalated sequence lacking homology; (c) the 3' extremity fits A ↔ G exchanges (10% of transcript length), the 5' half follows regular transcription; the intervening region seems a mix of regular and A ↔ G transcriptions (transcript 3); (d) murine cytosolic transcript 4 switches to A ↔ U + C ↔ G, and is fused with A ↔ U + C ↔ G swinger transformed precursor rRNA. In (c), each concomitant transcript 5' and 3' extremities match opposite genome strands. Transcripts 3 and 4 combine transcript fusions with partial swinger transcriptions. Occasional (usually sharp) switches between regular and swinger transcriptions reveal greater coding potential than detected until now, suggest stable polymerase swinger conformations.

  20. Ellipsoidal-mirror reflectometer accurately measures infrared reflectance of materials

    NASA Technical Reports Server (NTRS)

    Dunn, S. T.; Richmond, J. C.

    1967-01-01

    Reflectometer accurately measures the reflectance of specimens in the infrared beyond 2.5 microns and under geometric conditions approximating normal irradiation and hemispherical viewing. It includes an ellipsoidal mirror, a specially coated averaging sphere associated with a detector for minimizing spatial and angular sensitivity, and an incident flux chopper.

  1. Assessment of reference genes for reliable analysis of gene transcription by RT-qPCR in ovine leukocytes.

    PubMed

    Mahakapuge, T A N; Scheerlinck, J-P Y; Rojas, C A Alvarez; Every, A L; Hagen, J

    2016-03-01

    With the availability of genetic sequencing data, quantitative reverse transcription PCR (RT-qPCR) is increasingly being used for the quantification of gene transcription across species. Too often there is little regard to the selection of reference genes and the impact that a poor choice has on data interpretation. Indeed, RT-qPCR provides a snapshot of relative gene transcription at a given time-point, and hence is highly dependent on the stability of the transcription of the reference gene(s). Using ovine efferent lymph cells and peripheral blood mono-nuclear cells (PBMCs), the two most frequently used leukocytes in immunological studies, we have compared the stability of transcription of the most commonly used ovine reference genes: YWHAZ, RPL-13A, PGK1, B2M, GAPDH, HPRT, SDHA and ACTB. Using established algorithms for reference gene normalization "geNorm" and "Norm Finder", PGK1, GAPDH and YWHAZ were deemed the most stably transcribed genes for efferent leukocytes and PGK1, YWHAZ and SDHA were optimal in PBMCs. These genes should therefore be considered for accurate and reproducible RT-qPCR data analysis of gene transcription in sheep.

  2. Transcriptional enhancers: Transcription, function and flexibility.

    PubMed

    Melamed, Philippa; Yosefzon, Yahav; Rudnizky, Sergei; Pnueli, Lilach

    2016-01-01

    Active transcriptional enhancers are often transcribed to eRNAs, whose changing levels mirror those of the target gene mRNA. We discuss some of the reported functions of these eRNAs and their likely diversity to allow utilization of distinct cis regulatory regions to enhance transcription in diverse developmental and cellular contexts.

  3. CrossNorm: a novel normalization strategy for microarray data in cancers

    PubMed Central

    Cheng, Lixin; Lo, Leung-Yau; Tang, Nelson L. S.; Wang, Dong; Leung, Kwong-Sak

    2016-01-01

    Normalization is essential to get rid of biases in microarray data for their accurate analysis. Existing normalization methods for microarray gene expression data commonly assume a similar global expression pattern among samples being studied. However, scenarios of global shifts in gene expressions are dominant in cancers, making the assumption invalid. To alleviate the problem, here we propose and develop a novel normalization strategy, Cross Normalization (CrossNorm), for microarray data with unbalanced transcript levels among samples. Conventional procedures, such as RMA and LOESS, arbitrarily flatten the difference between case and control groups leading to biased gene expression estimates. Noticeably, applying these methods under the strategy of CrossNorm, which makes use of the overall statistics of the original signals, the results showed significantly improved robustness and accuracy in estimating transcript level dynamics for a series of publicly available datasets, including titration experiment, simulated data, spike-in data and several real-life microarray datasets across various types of cancers. The results have important implications for the past and the future cancer studies based on microarray samples with non-negligible difference. Moreover, the strategy can also be applied to other sorts of high-throughput data as long as the experiments have global expression variations between conditions. PMID:26732145

  4. Nuclear gadgets in mitochondrial DNA replication and transcription.

    PubMed

    Clayton, D A

    1991-03-01

    In mammalian mitochondrial DNA, activation of the light-strand promoter mediates both priming of leading-strand replication and initiation of light-strand transcription. Accurate and efficient transcription requires at least two proteins: mitochondrial RNA polymerase and a separable transcription factor that can function across species boundaries. Subsequently, primer RNAs are cleaved by a site-specific ribonucleoprotein endoribonuclease that recognizes short, highly conserved sequence elements in the RNA substrate.

  5. Transcription in archaea

    NASA Technical Reports Server (NTRS)

    Kyrpides, N. C.; Ouzounis, C. A.; Woese, C. R. (Principal Investigator)

    1999-01-01

    Using the sequences of all the known transcription-associated proteins from Bacteria and Eucarya (a total of 4,147), we have identified their homologous counterparts in the four complete archaeal genomes. Through extensive sequence comparisons, we establish the presence of 280 predicted transcription factors or transcription-associated proteins in the four archaeal genomes, of which 168 have homologs only in Bacteria, 51 have homologs only in Eucarya, and the remaining 61 have homologs in both phylogenetic domains. Although bacterial and eukaryotic transcription have very few factors in common, each exclusively shares a significantly greater number with the Archaea, especially the Bacteria. This last fact contrasts with the obvious close relationship between the archaeal and eukaryotic transcription mechanisms per se, and in particular, basic transcription initiation. We interpret these results to mean that the archaeal transcription system has retained more ancestral characteristics than have the transcription mechanisms in either of the other two domains.

  6. Accurate ab Initio Spin Densities.

    PubMed

    Boguslawski, Katharina; Marti, Konrad H; Legeza, Ors; Reiher, Markus

    2012-06-12

    We present an approach for the calculation of spin density distributions for molecules that require very large active spaces for a qualitatively correct description of their electronic structure. Our approach is based on the density-matrix renormalization group (DMRG) algorithm to calculate the spin density matrix elements as a basic quantity for the spatially resolved spin density distribution. The spin density matrix elements are directly determined from the second-quantized elementary operators optimized by the DMRG algorithm. As an analytic convergence criterion for the spin density distribution, we employ our recently developed sampling-reconstruction scheme [J. Chem. Phys.2011, 134, 224101] to build an accurate complete-active-space configuration-interaction (CASCI) wave function from the optimized matrix product states. The spin density matrix elements can then also be determined as an expectation value employing the reconstructed wave function expansion. Furthermore, the explicit reconstruction of a CASCI-type wave function provides insight into chemically interesting features of the molecule under study such as the distribution of α and β electrons in terms of Slater determinants, CI coefficients, and natural orbitals. The methodology is applied to an iron nitrosyl complex which we have identified as a challenging system for standard approaches [J. Chem. Theory Comput.2011, 7, 2740].

  7. Nuclear sequestration of COL1A1 mRNA transcript associated with type I osteogenesis imperfecta (OI)

    SciTech Connect

    Primorac, D.; Stover, M.L.; McKinstry, M.B.

    1994-09-01

    Previously we identified an OI type I patient with a splice donor mutation that resulted in intron 26 retention instead of exon skipping and sequestration of normal levels of the mutant transcript in the nuclear compartment. Intron retention was consistent with the exon definition hypothesis for splice site selection since the size of the exon-intron-exon unit was less than 300 bp. Furthermore, the retained intron contained in-frame stop codons which is thought to cause the mutant RNA to remain within the nucleus rather than appearing in the cytoplasm. To test these hypotheses, genomic fragments containing the normal sequence or the donor mutation were cloned into a collagen minigene and expressed in stably tansfected NIH 3T3 cells. None of the modifications to the normal intron altered the level of RNA that accumulated in the cytoplasm, as expected. However none of the modifications to the mutant intron allowed accumulation of normal levels of mRNA in the cytoplasm. Moreover, in contrast to our findings in the patient`s cells only low levels of mutant transcript were found in the nucleus; a fraction of the transcript did appear in the cytoplasm which had spliced the mutant donor site correctly. Nuclear run-on experiments demonstrated equal levels of transcription from each transgene. Expression of another donor mutation known to cause in-frame exon skipping in OI type IV was accurately reproduced in the minigene in transfected 3T3 cells. Our experience suggests that either mechanism can lead to formation of a null allele possibly related to the type of splicing events surrounding the potential stop codons. Understanding the rules governing inactivation of a collagen RNA transcript may be important in designing a strategy to inactivate a dominate negative mutation associated with the more severe forms of OI.

  8. Quantitatively predictable control of Drosophila transcriptional enhancers in vivo with engineered transcription factors.

    PubMed

    Crocker, Justin; Ilsley, Garth R; Stern, David L

    2016-03-01

    Genes are regulated by transcription factors that bind to regions of genomic DNA called enhancers. Considerable effort is focused on identifying transcription factor binding sites, with the goal of predicting gene expression from DNA sequence. Despite this effort, general, predictive models of enhancer function are currently lacking. Here we combine quantitative models of enhancer function with manipulations using engineered transcription factors to examine the extent to which enhancer function can be controlled in a quantitatively predictable manner. Our models, which incorporate few free parameters, can accurately predict the contributions of ectopic transcription factor inputs. These models allow the predictable 'tuning' of enhancers, providing a framework for the quantitative control of enhancers with engineered transcription factors.

  9. Evaluation of Appropriate Reference Genes for Gene Expression Normalization during Watermelon Fruit Development.

    PubMed

    Kong, Qiusheng; Yuan, Jingxian; Gao, Lingyun; Zhao, Liqiang; Cheng, Fei; Huang, Yuan; Bie, Zhilong

    2015-01-01

    Gene expression analysis in watermelon (Citrullus lanatus) fruit has drawn considerable attention with the availability of genome sequences to understand the regulatory mechanism of fruit development and to improve its quality. Real-time quantitative reverse-transcription PCR (qRT-PCR) is a routine technique for gene expression analysis. However, appropriate reference genes for transcript normalization in watermelon fruits have not been well characterized. The aim of this study was to evaluate the appropriateness of 12 genes for their potential use as reference genes in watermelon fruits. Expression variations of these genes were measured in 48 samples obtained from 12 successive developmental stages of parthenocarpic and fertilized fruits of two watermelon genotypes by using qRT-PCR analysis. Considering the effects of genotype, fruit setting method, and developmental stage, geNorm determined clathrin adaptor complex subunit (ClCAC), β-actin (ClACT), and alpha tubulin 5 (ClTUA5) as the multiple reference genes in watermelon fruit. Furthermore, ClCAC alone or together with SAND family protein (ClSAND) was ranked as the single or two best reference genes by NormFinder. By using the top-ranked reference genes to normalize the transcript abundance of phytoene synthase (ClPSY1), a good correlation between lycopene accumulation and ClPSY1 expression pattern was observed in ripening watermelon fruit. These validated reference genes will facilitate the accurate measurement of gene expression in the studies on watermelon fruit biology.

  10. Accurate stress resultants equations for laminated composite deep thick shells

    SciTech Connect

    Qatu, M.S.

    1995-11-01

    This paper derives accurate equations for the normal and shear force as well as bending and twisting moment resultants for laminated composite deep, thick shells. The stress resultant equations for laminated composite thick shells are shown to be different from those of plates. This is due to the fact the stresses over the thickness of the shell have to be integrated on a trapezoidal-like shell element to obtain the stress resultants. Numerical results are obtained and showed that accurate stress resultants are needed for laminated composite deep thick shells, especially if the curvature is not spherical.

  11. Short-Patch Reverse Transcription in Escherichia Coli

    PubMed Central

    Thaler, D. S.; Tombline, G.; Zahn, K.

    1995-01-01

    Chimeras of RNA and DNA have distinctive physical and biological properties. Chimeric oligonucleotides that contained one, two or three ribonucleotides whose phosphodiester backbone was covalently continuous with DNA were synthesized. Site-directed mutagenesis was used to assess genetic information transfer from the ribonucleotide positions. Transfer was scored by the formation or reversion of an ochre site that also corresponded to a restriction cleavage site. This allowed physical as well as genetic assay of mutational events. Bases attached to the ribonucleotides were able to accurately direct the synthesis of progeny DNA. The results suggest that in vivo DNA polymerases utilize a ``running start'' on a DNA backbone to continue across a covalent backbone junction into a region of ribonucleotides and then back again onto a normal DNA backbone. The phenomenon is designated short-patch reverse transcription (SPRT) by analogy to short-patch mismatch correction and reverse transcription as the term is generally used. The possibility is considered that SPRT contributes to an unrecognized pathway of mutagenesis. PMID:7545627

  12. Library preparation for highly accurate population sequencing of RNA viruses

    PubMed Central

    Acevedo, Ashley; Andino, Raul

    2015-01-01

    Circular resequencing (CirSeq) is a novel technique for efficient and highly accurate next-generation sequencing (NGS) of RNA virus populations. The foundation of this approach is the circularization of fragmented viral RNAs, which are then redundantly encoded into tandem repeats by ‘rolling-circle’ reverse transcription. When sequenced, the redundant copies within each read are aligned to derive a consensus sequence of their initial RNA template. This process yields sequencing data with error rates far below the variant frequencies observed for RNA viruses, facilitating ultra-rare variant detection and accurate measurement of low-frequency variants. Although library preparation takes ~5 d, the high-quality data generated by CirSeq simplifies downstream data analysis, making this approach substantially more tractable for experimentalists. PMID:24967624

  13. 38 CFR 4.46 - Accurate measurement.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 38 Pensions, Bonuses, and Veterans' Relief 1 2013-07-01 2013-07-01 false Accurate measurement. 4... RATING DISABILITIES Disability Ratings The Musculoskeletal System § 4.46 Accurate measurement. Accurate measurement of the length of stumps, excursion of joints, dimensions and location of scars with respect...

  14. 38 CFR 4.46 - Accurate measurement.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 38 Pensions, Bonuses, and Veterans' Relief 1 2012-07-01 2012-07-01 false Accurate measurement. 4... RATING DISABILITIES Disability Ratings The Musculoskeletal System § 4.46 Accurate measurement. Accurate measurement of the length of stumps, excursion of joints, dimensions and location of scars with respect...

  15. 38 CFR 4.46 - Accurate measurement.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 38 Pensions, Bonuses, and Veterans' Relief 1 2010-07-01 2010-07-01 false Accurate measurement. 4... RATING DISABILITIES Disability Ratings The Musculoskeletal System § 4.46 Accurate measurement. Accurate measurement of the length of stumps, excursion of joints, dimensions and location of scars with respect...

  16. 38 CFR 4.46 - Accurate measurement.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 38 Pensions, Bonuses, and Veterans' Relief 1 2014-07-01 2014-07-01 false Accurate measurement. 4... RATING DISABILITIES Disability Ratings The Musculoskeletal System § 4.46 Accurate measurement. Accurate measurement of the length of stumps, excursion of joints, dimensions and location of scars with respect...

  17. 38 CFR 4.46 - Accurate measurement.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 38 Pensions, Bonuses, and Veterans' Relief 1 2011-07-01 2011-07-01 false Accurate measurement. 4... RATING DISABILITIES Disability Ratings The Musculoskeletal System § 4.46 Accurate measurement. Accurate measurement of the length of stumps, excursion of joints, dimensions and location of scars with respect...

  18. Thyrotropin controls transcription of the thyroglobulin gene.

    PubMed

    Van Heuverswyn, B; Streydio, C; Brocas, H; Refetoff, S; Dumont, J; Vassart, G

    1984-10-01

    The availability of rat thyroglobulin cDNA clones was exploited to study the regulation of thyroglobulin gene transcription by thyrotropin (TSH). Groups of rats were subjected to treatments leading to reduction or increase in the rat serum TSH (rTSH) levels. Thyroid gland nuclei were isolated, incubated in vitro in the presence of 32P-labeled uridine triphosphate, and thyroglobulin transcripts were quantitated by hybridization to immobilized rat thyroglobulin cDNA clones. Transcription of the thyroglobulin gene was found to be very active in thyroid nuclei from control animals. It represented about 10% of total RNA polymerase II activity. Chronic hyperstimulation of the thyroid glands with endogenous rTSH was achieved in rats treated with the goitrogen propylthiouracil. No significant increase of thyroglobulin gene transcription could be measured in thyroid nuclei from these animals. On the contrary, a dramatic decrease in thyroglobulin gene transcription was observed in those animals in which endogenous rTSH levels had been suppressed by hypophysectomy or by the administration of triiodothyronine. Injection of exogenous bovine TSH in such animals readily restored transcriptional activity of the gene. Our results identify transcription as an important regulatory step involved in TSH action. They suggest that normal TSH levels induce close to maximal expression of the thyroglobulin gene but that continuous presence of TSH is required in order to maintain the gene in an activated state.

  19. Nascent RNA sequencing reveals distinct features in plant transcription

    PubMed Central

    Hetzel, Jonathan; Duttke, Sascha H.; Benner, Christopher; Chory, Joanne

    2016-01-01

    Transcriptional regulation of gene expression is a major mechanism used by plants to confer phenotypic plasticity, and yet compared with other eukaryotes or bacteria, little is known about the design principles. We generated an extensive catalog of nascent and steady-state transcripts in Arabidopsis thaliana seedlings using global nuclear run-on sequencing (GRO-seq), 5′GRO-seq, and RNA-seq and reanalyzed published maize data to capture characteristics of plant transcription. De novo annotation of nascent transcripts accurately mapped start sites and unstable transcripts. Examining the promoters of coding and noncoding transcripts identified comparable chromatin signatures, a conserved “TGT” core promoter motif and unreported transcription factor-binding sites. Mapping of engaged RNA polymerases showed a lack of enhancer RNAs, promoter-proximal pausing, and divergent transcription in Arabidopsis seedlings and maize, which are commonly present in yeast and humans. In contrast, Arabidopsis and maize genes accumulate RNA polymerases in proximity of the polyadenylation site, a trend that coincided with longer genes and CpG hypomethylation. Lack of promoter-proximal pausing and a higher correlation of nascent and steady-state transcripts indicate Arabidopsis may regulate transcription predominantly at the level of initiation. Our findings provide insight into plant transcription and eukaryotic gene expression as a whole. PMID:27729530

  20. Normalization of human RNA-seq experiments using chimpanzee RNA as a spike-in standard.

    PubMed

    Yu, Hannah; Hahn, Yoonsoo; Park, Sang-Ryoul; Chung, Sun-Ku; Jeong, Sangkyun; Yang, Inchul

    2016-08-24

    Normalization of human RNA-seq experiments employing chimpanzee RNA as a spike-in standard is reported. Human and chimpanzee RNAs exhibit single nucleotide variations (SNVs) in average 210-bp intervals. Spike-in chimpanzee RNA would behave the same as the human counterparts during the whole NGS procedures owing to the high sequence similarity. After discrimination of species origins of the NGS reads based on SNVs, the chimpanzee reads were used to read-by-read normalize biases and variations of human reads. By this approach, as many as 10,119 transcripts were simultaneously normalized for the entire NGS procedures leading to accurate and reproducible quantification of differential gene expression. In addition, incomparable data sets from different in-process degradations or from different library preparation methods were made well comparable by the normalization. Based on these results, we expect that the normalization approaches using near neighbor genomes as internal standards could be employed as a standard protocol, which will improve both accuracy and comparability of NGS results across different sample batches, laboratories and NGS platforms.

  1. Nonspecific hybridization scaling of microarray expression estimates: a physicochemical approach for chip-to-chip normalization.

    PubMed

    Binder, Hans; Brücker, Jan; Burden, Conrad J

    2009-03-05

    The problem of inferring accurate quantitative estimates of transcript abundances from gene expression microarray data is addressed. Particular attention is paid to correcting chip-to-chip variations arising mainly as a result of unwanted nonspecific background hybridization to give transcript abundances measured in a common scale. This study verifies and generalizes a model of the mutual dependence between nonspecific background hybridization and the sensitivity of the specific signal using an approach based on the physical chemistry of surface hybridization. We have analyzed GeneChip oligonucleotide microarray data taken from a set of five benchmark experiments including dilution, Latin Square, and "Golden spike" designs. Our analysis concentrates on the important effect of changes in the unwanted nonspecific background inherent in the technology due to changes in total RNA target concentration and/or composition. We find that incremental changes in nonspecific background entail opposite sign incremental changes in the effective specific binding constant. This effect, which we refer to as the "up-down" effect, results from the subtle interplay of competing interactions between the probes and specific and nonspecific targets at the chip surface and in bulk solution. We propose special rules for proper normalization of expression values considering the specifics of the up-down effect. Particularly for normalization one has to level the expression values of invariant expressed probes. Existing heuristic normalization techniques which do not exclude absent probes, level intensities instead of expression values, and/or use low variance criteria for identifying invariant sets of probes lead to biased results. Strengths and pitfalls of selected normalization methods are discussed. We also find that the extent of the up-down effect is modified if RNA targets are replaced by DNA targets, in that microarray sensitivity and specificity are improved via a decrease in

  2. Comprehensive Selection of Reference Genes for Gene Expression Normalization in Sugarcane by Real Time Quantitative RT-PCR

    PubMed Central

    Ling, Hui; Wu, Qibin; Guo, Jinlong; Xu, Liping; Que, Youxiong

    2014-01-01

    The increasingly used real time quantitative reverse transcription-PCR (qRT-PCR) method for gene expression analysis requires one or several reference gene(s) acting as normalization factor(s). In order to facilitate gene expression studies in sugarcane (Saccharum officinarum), a non-model plant with limited genome information, the stability of 13 candidate reference genes was evaluated. The geNorm, NormFinder and deltaCt methods were used for selecting stably expressed internal controls across different tissues and under various experimental treatments. These results revealed that, among these 13 candidate reference genes, GAPDH, eEF-1a and eIF-4α were the most stable and suitable for use as normalization factors across all various experimental samples. In addition, APRT could be a candidate for examining the relationship between gene copy number and transcript levels in sugarcane tissue samples. According to the results evaluated by geNorm, combining CUL and eEF-1α in hormone treatment experiments; CAC and CUL in abiotic stress tests; GAPDH, eEF-1a and CUL in all treatment samples plus CAC, CUL, APRT and TIPS-41 in cultivar tissues as groups for normalization would lead to more accurate and reliable expression quantification in sugarcane. This is the first systematic validation of reference genes for quantification of transcript expression profiles in sugarcane. This study should provide useful information for selecting reference genes for more accurate quantification of gene expression in sugarcane and other plant species. PMID:24823940

  3. SWItching on the transcriptional circuitry in melanoma.

    PubMed

    Vinod Saladi, Srinivas; Marathe, Himangi; de la Serna, Ivana L

    2010-08-16

    Melanoma is an aggressive malignancy that is resistant to current therapy, and the most lethal of all human skin cancers. It is characterized by several genetic alterations that lead to changes in gene expression and tumorigenesis by triggering alterations in the normal transcriptional circuitry. Transformation and tumor progression are thought to be promoted by a complex interplay between the accumulation of genetic alterations and epigenetic changes. In this review, we discuss recent studies that have implicated SWI/SNF chromatin remodeling enzymes as epigenetic regulators of a transcriptional circuit that operates within the context the genetic alterations that frequently occur in melanoma.

  4. ASTP Onboard Voice Transcription

    NASA Technical Reports Server (NTRS)

    1975-01-01

    The transcription is presented of the Apollo-Soyuz Test Project voice communications as recorded on the command module data storage equipment. Data from this recorder are telemetered (dumped) to Space Tracking and Data Network sites for retransmission to the Johnson Space Center. The transcript is divided into three columns -- time, speaker, and text. The Greenwich mean time column consists of three two-digit numbers representing hours, minutes, and seconds (e.g., 22 34 14) for the Julian dates shown at the top of the page on which a new day begins. The speaker column indicates the source of a transmission; the text column contains the verbatim transcript of the communications.

  5. A comprehensive library of fluorescent transcriptional reporters for Escherichia coli.

    PubMed

    Zaslaver, Alon; Bren, Anat; Ronen, Michal; Itzkovitz, Shalev; Kikoin, Ilya; Shavit, Seagull; Liebermeister, Wolfram; Surette, Michael G; Alon, Uri

    2006-08-01

    E. coli is widely used for systems biology research; there exists a need, however, for tools that can be used to accurately and comprehensively measure expression dynamics in individual living cells. To address this we present a library of transcriptional fusions of gfp to each of about 2,000 different promoters in E. coli K12, covering the great majority of the promoters in the organism. Each promoter fusion is expressed from a low-copy plasmid. We demonstrate that this library can be used to obtain highly accurate dynamic measurements of promoter activity on a genomic scale, in a glucose-lactose diauxic shift experiment. The library allowed detection of about 80 previously uncharacterized transcription units in E. coli, including putative internal promoters within previously known operons, such as the lac operon. This library can serve as a tool for accurate, high-resolution analysis of transcription networks in living E. coli cells.

  6. Inhibition of host cell RNA polymerase III-mediated transcription by poliovirus: Inactivation of specific transcription factors

    SciTech Connect

    Fradkin, L.G.; Yoshinaga, S.K.; Berk, A.J.; Dasgupta, A.

    1987-11-01

    The inhibition of transcription by RNA polymerase III in poliovirus-infected cells was studied. Experiments utilizing two different cell lines showed that the initiation step of transcription by RNA polymerase III was impaired by infection of these cells with the virus. The observed inhibition of transcription was not due to shut-off of host cell protein synthesis by poliovirus. Among four distinct components required for accurate transcription in vitro from cloned DNA templates, activities of RNA polymerase III and transcription factor TFIIIA were not significantly affected by virus infection. The activity of transcription factor TFIIIC, the limiting component required for transcription of RNA polymerase III genes, was severely inhibited in infected cells, whereas that of transcription factor TFIIIB was inhibited to a lesser extent. The sequence-specific DNA-binding of TFIIIC to the adenovirus VA1 gene internal promoted, however, was not altered by infection of cells with the virus. The authors conclude that (i) at least two transcription factors, TFIIIB and TFIIIC, are inhibited by infection of cells with poliovirtus, (ii) inactivation of TFIIIC does not involve destruction of its DNA-binding domain, and (iii) sequence-specific DNA binding by TFIIIC may be necessary but is not sufficient for the formation of productive transcription complexes.

  7. PPARγ, an important gene related to lipid metabolism and immunity in Megalobrama amblycephala: cloning, characterization and transcription analysis by GeNorm.

    PubMed

    Li, Su; Gul, Yasmeen; Wang, Weimin; Qian, Xueqiao; Zhao, Yuhua

    2013-01-10

    In order to be able to modulate and improve the function of PPARγ and decrease further some metabolic diseases of M. amblycephala, we have cloned and identified the full-length cDNA of PPARγ in M. amblycephala and examined its transcription patterns at different embryo developmental stages and in different tissues of adult and immature fish. We also accurately normalized seven reference genes by GeNorm and calculated their gene transcription normalization factors. The full-length of PPARγ was 1968 bp, consisting of 218 bp 5'-untranslated region, 1,533 bp open reading frame encoding 510 amino acids residues and 217 bp 3'-untranslated region. M. amblycephala PPARγ peptide was predicted to consist of 4 conserved domains, i.e. N-terminal domain, DNA-binding domain, ligand binding domain and flexible hinge region. PPARγ mRNAs were detected in all studied tissues of adult and immature fish including adipose tissue, gill, heart, liver, spleen, kidney, white muscle, intestine, brain and gonad. In adult fish, PPARγ transcription in liver was highest, followed by gills and it was lowest in female gonads. Moreover, the differences among liver, gill, intestine/brain, spleen/white muscle, kidney and female gonads were greatly significant (p<0.01). The transcription of PPARγ in male gonads was significantly higher than in female gonads (p<0.01). In immature fish, the transcription of PPARγ was highest in intestines followed by adipose tissue, and it was lowest in hearts and white muscles. A great difference was observed (p<0.01) in the transcription of PPARγ among adipose tissue, intestines, liver and heart/white muscles. At different embryo developmental stages, PPARγ transcription in unfertilized spermatozoa was greatly higher than in unfertilized ovum (p<0.01) and it was highest among different embryo developmental stages. The transcription of PPARγ increased gradually during 2 cells stage and 32 cells stage and then decreased until gastrula stage at which it was

  8. DNA supercoiling during transcription

    PubMed Central

    Ma, Jie; Wang, Michelle D.

    2017-01-01

    The twin-supercoiled-domain model describes how transcription can drive DNA supercoiling, and how DNA supercoiling, in turn plays an important role in regulating gene transcription. In vivo and in vitro experiments have disclosed many details of the complex interactions in this relationship, and recently new insights have been gained with the help of genome-wide DNA supercoiling mapping techniques and single molecule methods. This review summarizes the general mechanisms of the interplay between DNA supercoiling and transcription, considers the biological implications, and focuses on recent important discoveries and technical advances in this field. We highlight the significant impact of DNA supercoiling in transcription, but also more broadly in all processes operating on DNA.

  9. Groundtruth approach to accurate quantitation of fluorescence microarrays

    SciTech Connect

    Mascio-Kegelmeyer, L; Tomascik-Cheeseman, L; Burnett, M S; van Hummelen, P; Wyrobek, A J

    2000-12-01

    To more accurately measure fluorescent signals from microarrays, we calibrated our acquisition and analysis systems by using groundtruth samples comprised of known quantities of red and green gene-specific DNA probes hybridized to cDNA targets. We imaged the slides with a full-field, white light CCD imager and analyzed them with our custom analysis software. Here we compare, for multiple genes, results obtained with and without preprocessing (alignment, color crosstalk compensation, dark field subtraction, and integration time). We also evaluate the accuracy of various image processing and analysis techniques (background subtraction, segmentation, quantitation and normalization). This methodology calibrates and validates our system for accurate quantitative measurement of microarrays. Specifically, we show that preprocessing the images produces results significantly closer to the known ground-truth for these samples.

  10. Advocating for Normal Birth With Normal Clothes

    PubMed Central

    Waller-Wise, Renece

    2007-01-01

    Childbirth educators need to be aware that the clothes they wear when teaching classes send a nonverbal message to class participants. Regardless of who wears the clothing or what is worn, clothes send a message; thus, both the advantages and disadvantages related to clothing choice should be considered. Ultimately, the message should reflect the values of supporting normal birth. For childbirth educators who are allowed to choose their own apparel to wear in their classes, street clothes may be the benchmark for which to strive. This article discusses the many nonverbal messages that clothes convey and provides support for the choice of street clothes as the dress for the professional childbirth educator; thus, “normal clothes to promote normal birth.” PMID:18408807

  11. Transcriptional Regulation of Telomerase Reverse Transcriptase (TERT) by MYC

    PubMed Central

    Khattar, Ekta; Tergaonkar, Vinay

    2017-01-01

    Telomerase elongates telomeres and is crucial for maintaining genomic stability. While stem cells and cancer cells display high telomerase activity, normal somatic cells lack telomerase activity primarily due to transcriptional repression of telomerase reverse transcriptase (TERT), the catalytic component of telomerase. Transcription factor binding, chromatin status as well as epigenetic modifications at the TERT promoter regulates TERT transcription. Myc is an important transcriptional regulator of TERT that directly controls its expression by promoter binding and associating with other transcription factors. In this review, we discuss the current understanding of the molecular mechanisms behind regulation of TERT transcription by Myc. We also discuss future perspectives in investigating the regulation of Myc at TERT promoter during cancer development. PMID:28184371

  12. Mill profiler machines soft materials accurately

    NASA Technical Reports Server (NTRS)

    Rauschl, J. A.

    1966-01-01

    Mill profiler machines bevels, slots, and grooves in soft materials, such as styrofoam phenolic-filled cores, to any desired thickness. A single operator can accurately control cutting depths in contour or straight line work.

  13. Accurate energy levels for singly ionized platinum (Pt II)

    NASA Technical Reports Server (NTRS)

    Reader, Joseph; Acquista, Nicolo; Sansonetti, Craig J.; Engleman, Rolf, Jr.

    1988-01-01

    New observations of the spectrum of Pt II have been made with hollow-cathode lamps. The region from 1032 to 4101 A was observed photographically with a 10.7-m normal-incidence spectrograph. The region from 2245 to 5223 A was observed with a Fourier-transform spectrometer. Wavelength measurements were made for 558 lines. The uncertainties vary from 0.0005 to 0.004 A. From these measurements and three parity-forbidden transitions in the infrared, accurate values were determined for 28 even and 72 odd energy levels of Pt II.

  14. beta-Glucuronidase is an optimal normalization control gene for molecular monitoring of chronic myelogenous leukemia.

    PubMed

    Lee, Joong Won; Chen, Qiaofang; Knowles, Daniel M; Cesarman, Ethel; Wang, Y Lynn

    2006-07-01

    Quantitative monitoring of breakpoint cluster region (BCR)-Abelson kinase (ABL) transcripts has become indispensable in the clinical care of patients with chronic myelogenous leukemia. Because quantity and quality of RNA in clinical samples are highly variable, a suitable internal normalization control is required for accurate BCR-ABL quantification. However, few studies have examined suitability of the control genes using criteria relevant to residual disease testing. In this study, we evaluated a number of control genes with the application of several novel criteria, including control gene performance on serial patient sample testing and in a residual disease model. We also examined expression of the control genes in BCR-ABL-positive K562 cells in response to Gleevec treatment. We found that beta-glucuronidase is the best control gene among those studied. Importantly, ABL, a widely used control gene, generates misleading BCR-ABL changes that potentially affect the clinical management of chronic myelogenous leukemia patients.

  15. A compendium of nucleosome and transcript profiles reveals determinants of chromatin architecture and transcription.

    PubMed

    van Bakel, Harm; Tsui, Kyle; Gebbia, Marinella; Mnaimneh, Sanie; Hughes, Timothy R; Nislow, Corey

    2013-05-01

    Nucleosomes in all eukaryotes examined to date adopt a characteristic architecture within genes and play fundamental roles in regulating transcription, yet the identity and precise roles of many of the trans-acting factors responsible for the establishment and maintenance of this organization remain to be identified. We profiled a compendium of 50 yeast strains carrying conditional alleles or complete deletions of genes involved in transcriptional regulation, histone biology, and chromatin remodeling, as well as compounds that target transcription and histone deacetylases, to assess their respective roles in nucleosome positioning and transcription. We find that nucleosome patterning in genes is affected by many factors, including the CAF-1 complex, Spt10, and Spt21, in addition to previously reported remodeler ATPases and histone chaperones. Disruption of these factors or reductions in histone levels led genic nucleosomes to assume positions more consistent with their intrinsic sequence preferences, with pronounced and specific shifts of the +1 nucleosome relative to the transcription start site. These shifts of +1 nucleosomes appear to have functional consequences, as several affected genes in Ino80 mutants exhibited altered expression responses. Our parallel expression profiling compendium revealed extensive transcription changes in intergenic and antisense regions, most of which occur in regions with altered nucleosome occupancy and positioning. We show that the nucleosome-excluding transcription factors Reb1, Abf1, Tbf1, and Rsc3 suppress cryptic transcripts at their target promoters, while a combined analysis of nucleosome and expression profiles identified 36 novel transcripts that are normally repressed by Tup1/Cyc8. Our data confirm and extend the roles of chromatin remodelers and chaperones as major determinants of genic nucleosome positioning, and these data provide a valuable resource for future studies.

  16. A Compendium of Nucleosome and Transcript Profiles Reveals Determinants of Chromatin Architecture and Transcription

    PubMed Central

    van Bakel, Harm; Tsui, Kyle; Gebbia, Marinella; Mnaimneh, Sanie; Hughes, Timothy R.; Nislow, Corey

    2013-01-01

    Nucleosomes in all eukaryotes examined to date adopt a characteristic architecture within genes and play fundamental roles in regulating transcription, yet the identity and precise roles of many of the trans-acting factors responsible for the establishment and maintenance of this organization remain to be identified. We profiled a compendium of 50 yeast strains carrying conditional alleles or complete deletions of genes involved in transcriptional regulation, histone biology, and chromatin remodeling, as well as compounds that target transcription and histone deacetylases, to assess their respective roles in nucleosome positioning and transcription. We find that nucleosome patterning in genes is affected by many factors, including the CAF-1 complex, Spt10, and Spt21, in addition to previously reported remodeler ATPases and histone chaperones. Disruption of these factors or reductions in histone levels led genic nucleosomes to assume positions more consistent with their intrinsic sequence preferences, with pronounced and specific shifts of the +1 nucleosome relative to the transcription start site. These shifts of +1 nucleosomes appear to have functional consequences, as several affected genes in Ino80 mutants exhibited altered expression responses. Our parallel expression profiling compendium revealed extensive transcription changes in intergenic and antisense regions, most of which occur in regions with altered nucleosome occupancy and positioning. We show that the nucleosome-excluding transcription factors Reb1, Abf1, Tbf1, and Rsc3 suppress cryptic transcripts at their target promoters, while a combined analysis of nucleosome and expression profiles identified 36 novel transcripts that are normally repressed by Tup1/Cyc8. Our data confirm and extend the roles of chromatin remodelers and chaperones as major determinants of genic nucleosome positioning, and these data provide a valuable resource for future studies. PMID:23658529

  17. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    NASA Astrophysics Data System (ADS)

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-10-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors.

  18. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells.

    PubMed

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C; Côté, Maxime C; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-10-14

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors.

  19. FOXA and master transcription factors recruit Mediator and Cohesin to the core transcriptional regulatory circuitry of cancer cells

    PubMed Central

    Fournier, Michèle; Bourriquen, Gaëlle; Lamaze, Fabien C.; Côté, Maxime C.; Fournier, Éric; Joly-Beauparlant, Charles; Caron, Vicky; Gobeil, Stéphane; Droit, Arnaud; Bilodeau, Steve

    2016-01-01

    Controlling the transcriptional program is essential to maintain the identity and the biological functions of a cell. The Mediator and Cohesin complexes have been established as central cofactors controlling the transcriptional program in normal cells. However, the distribution, recruitment and importance of these complexes in cancer cells have not been fully investigated. Here we show that FOXA and master transcription factors are part of the core transcriptional regulatory circuitry of cancer cells and are essential to recruit M ediator and Cohesin. Indeed, Mediator and Cohesin occupied the enhancer and promoter regions of actively transcribed genes and maintained the proliferation and colony forming potential. Through integration of publically available ChIP-Seq datasets, we predicted the core transcriptional regulatory circuitry of each cancer cell. Unexpectedly, for all cells investigated, the pioneer transcription factors FOXA1 and/or FOXA2 were identified in addition to cell-specific master transcription factors. Loss of both types of transcription factors phenocopied the loss of Mediator and Cohesin. Lastly, the master and pioneer transcription factors were essential to recruit Mediator and Cohesin to regulatory regions of actively transcribed genes. Our study proposes that maintenance of the cancer cell state is dependent on recruitment of Mediator and Cohesin through FOXA and master transcription factors. PMID:27739523

  20. Regulation of Transcript Elongation

    PubMed Central

    Belogurov, Georgiy A.; Artsimovitch, Irina

    2015-01-01

    Bacteria lack subcellular compartments and harbor a single RNA polymerase that synthesizes both structural and protein-coding RNAs, which are cotranscriptionally processed by distinct pathways. Nascent rRNAs fold into elaborate secondary structures and associate with ribosomal proteins, whereas nascent mRNAs are translated by ribosomes. During elongation, nucleic acid signals and regulatory proteins modulate concurrent RNA-processing events, instruct RNA polymerase where to pause and terminate transcription, or act as roadblocks to the moving enzyme. Communications among complexes that carry out transcription, translation, repair, and other cellular processes ensure timely execution of the gene expression program and survival under conditions of stress. This network is maintained by auxiliary proteins that act as bridges between RNA polymerase, ribosome, and repair enzymes, blurring boundaries between separate information-processing steps and making assignments of unique regulatory functions meaningless. Understanding the regulation of transcript elongation thus requires genome-wide approaches, which confirm known and reveal new regulatory connections. PMID:26132790

  1. The transcription factor encyclopedia.

    PubMed

    Yusuf, Dimas; Butland, Stefanie L; Swanson, Magdalena I; Bolotin, Eugene; Ticoll, Amy; Cheung, Warren A; Zhang, Xiao Yu Cindy; Dickman, Christopher T D; Fulton, Debra L; Lim, Jonathan S; Schnabl, Jake M; Ramos, Oscar H P; Vasseur-Cognet, Mireille; de Leeuw, Charles N; Simpson, Elizabeth M; Ryffel, Gerhart U; Lam, Eric W-F; Kist, Ralf; Wilson, Miranda S C; Marco-Ferreres, Raquel; Brosens, Jan J; Beccari, Leonardo L; Bovolenta, Paola; Benayoun, Bérénice A; Monteiro, Lara J; Schwenen, Helma D C; Grontved, Lars; Wederell, Elizabeth; Mandrup, Susanne; Veitia, Reiner A; Chakravarthy, Harini; Hoodless, Pamela A; Mancarelli, M Michela; Torbett, Bruce E; Banham, Alison H; Reddy, Sekhar P; Cullum, Rebecca L; Liedtke, Michaela; Tschan, Mario P; Vaz, Michelle; Rizzino, Angie; Zannini, Mariastella; Frietze, Seth; Farnham, Peggy J; Eijkelenboom, Astrid; Brown, Philip J; Laperrière, David; Leprince, Dominique; de Cristofaro, Tiziana; Prince, Kelly L; Putker, Marrit; del Peso, Luis; Camenisch, Gieri; Wenger, Roland H; Mikula, Michal; Rozendaal, Marieke; Mader, Sylvie; Ostrowski, Jerzy; Rhodes, Simon J; Van Rechem, Capucine; Boulay, Gaylor; Olechnowicz, Sam W Z; Breslin, Mary B; Lan, Michael S; Nanan, Kyster K; Wegner, Michael; Hou, Juan; Mullen, Rachel D; Colvin, Stephanie C; Noy, Peter John; Webb, Carol F; Witek, Matthew E; Ferrell, Scott; Daniel, Juliet M; Park, Jason; Waldman, Scott A; Peet, Daniel J; Taggart, Michael; Jayaraman, Padma-Sheela; Karrich, Julien J; Blom, Bianca; Vesuna, Farhad; O'Geen, Henriette; Sun, Yunfu; Gronostajski, Richard M; Woodcroft, Mark W; Hough, Margaret R; Chen, Edwin; Europe-Finner, G Nicholas; Karolczak-Bayatti, Magdalena; Bailey, Jarrod; Hankinson, Oliver; Raman, Venu; LeBrun, David P; Biswal, Shyam; Harvey, Christopher J; DeBruyne, Jason P; Hogenesch, John B; Hevner, Robert F; Héligon, Christophe; Luo, Xin M; Blank, Marissa Cathleen; Millen, Kathleen Joyce; Sharlin, David S; Forrest, Douglas; Dahlman-Wright, Karin; Zhao, Chunyan; Mishima, Yuriko; Sinha, Satrajit; Chakrabarti, Rumela; Portales-Casamar, Elodie; Sladek, Frances M; Bradley, Philip H; Wasserman, Wyeth W

    2012-01-01

    Here we present the Transcription Factor Encyclopedia (TFe), a new web-based compendium of mini review articles on transcription factors (TFs) that is founded on the principles of open access and collaboration. Our consortium of over 100 researchers has collectively contributed over 130 mini review articles on pertinent human, mouse and rat TFs. Notable features of the TFe website include a high-quality PDF generator and web API for programmatic data retrieval. TFe aims to rapidly educate scientists about the TFs they encounter through the delivery of succinct summaries written and vetted by experts in the field. TFe is available at http://www.cisreg.ca/tfe.

  2. The Transcription Factor Encyclopedia

    PubMed Central

    2012-01-01

    Here we present the Transcription Factor Encyclopedia (TFe), a new web-based compendium of mini review articles on transcription factors (TFs) that is founded on the principles of open access and collaboration. Our consortium of over 100 researchers has collectively contributed over 130 mini review articles on pertinent human, mouse and rat TFs. Notable features of the TFe website include a high-quality PDF generator and web API for programmatic data retrieval. TFe aims to rapidly educate scientists about the TFs they encounter through the delivery of succinct summaries written and vetted by experts in the field. TFe is available at http://www.cisreg.ca/tfe. PMID:22458515

  3. Simple, rapid and accurate molecular diagnosis of acute promyelocytic leukemia by loop mediated amplification technology.

    PubMed

    Spinelli, Orietta; Rambaldi, Alessandro; Rigo, Francesca; Zanghì, Pamela; D'Agostini, Elena; Amicarelli, Giulia; Colotta, Francesco; Divona, Mariadomenica; Ciardi, Claudia; Coco, Francesco Lo; Minnucci, Giulia

    2015-01-01

    The diagnostic work-up of acute promyelocytic leukemia (APL) includes the cytogenetic demonstration of the t(15;17) translocation and/or the PML-RARA chimeric transcript by RQ-PCR or RT-PCR. This latter assays provide suitable results in 3-6 hours. We describe here two new, rapid and specific assays that detect PML-RARA transcripts, based on the RT-QLAMP (Reverse Transcription-Quenching Loop-mediated Isothermal Amplification) technology in which RNA retrotranscription and cDNA amplification are carried out in a single tube with one enzyme at one temperature, in fluorescence and real time format. A single tube triplex assay detects bcr1 and bcr3 PML-RARA transcripts along with GUS housekeeping gene. A single tube duplex assay detects bcr2 and GUSB. In 73 APL cases, these assays detected in 16 minutes bcr1, bcr2 and bcr3 transcripts. All 81 non-APL samples were negative by RT-QLAMP for chimeric transcripts whereas GUSB was detectable. In 11 APL patients in which RT-PCR yielded equivocal breakpoint type results, RT-QLAMP assays unequivocally and accurately defined the breakpoint type (as confirmed by sequencing). Furthermore, RT-QLAMP could amplify two bcr2 transcripts with particularly extended PML exon 6 deletions not amplified by RQ-PCR. RT-QLAMP reproducible sensitivity is 10(-3) for bcr1 and bcr3 and 10(-)2 for bcr2 thus making this assay particularly attractive at diagnosis and leaving RQ-PCR for the molecular monitoring of minimal residual disease during the follow up. In conclusion, PML-RARA RT-QLAMP compared to RT-PCR or RQ-PCR is a valid improvement to perform rapid, simple and accurate molecular diagnosis of APL.

  4. Simple, rapid and accurate molecular diagnosis of acute promyelocytic leukemia by loop mediated amplification technology

    PubMed Central

    Spinelli, Orietta; Rambaldi, Alessandro; Rigo, Francesca; Zanghì, Pamela; D'Agostini, Elena; Amicarelli, Giulia; Colotta, Francesco; Divona, Mariadomenica; Ciardi, Claudia; Coco, Francesco Lo; Minnucci, Giulia

    2015-01-01

    The diagnostic work-up of acute promyelocytic leukemia (APL) includes the cytogenetic demonstration of the t(15;17) translocation and/or the PML-RARA chimeric transcript by RQ-PCR or RT-PCR. This latter assays provide suitable results in 3-6 hours. We describe here two new, rapid and specific assays that detect PML-RARA transcripts, based on the RT-QLAMP (Reverse Transcription-Quenching Loop-mediated Isothermal Amplification) technology in which RNA retrotranscription and cDNA amplification are carried out in a single tube with one enzyme at one temperature, in fluorescence and real time format. A single tube triplex assay detects bcr1 and bcr3 PML-RARA transcripts along with GUS housekeeping gene. A single tube duplex assay detects bcr2 and GUSB. In 73 APL cases, these assays detected in 16 minutes bcr1, bcr2 and bcr3 transcripts. All 81 non-APL samples were negative by RT-QLAMP for chimeric transcripts whereas GUSB was detectable. In 11 APL patients in which RT-PCR yielded equivocal breakpoint type results, RT-QLAMP assays unequivocally and accurately defined the breakpoint type (as confirmed by sequencing). Furthermore, RT-QLAMP could amplify two bcr2 transcripts with particularly extended PML exon 6 deletions not amplified by RQ-PCR. RT-QLAMP reproducible sensitivity is 10−3 for bcr1 and bcr3 and 10−2 for bcr2 thus making this assay particularly attractive at diagnosis and leaving RQ-PCR for the molecular monitoring of minimal residual disease during the follow up. In conclusion, PML-RARA RT-QLAMP compared to RT-PCR or RQ-PCR is a valid improvement to perform rapid, simple and accurate molecular diagnosis of APL. PMID:25815362

  5. CT Scan Method Accurately Assesses Humeral Head Retroversion

    PubMed Central

    Boileau, P.; Mazzoleni, N.; Walch, G.; Urien, J. P.

    2008-01-01

    Humeral head retroversion is not well described with the literature controversial regarding accuracy of measurement methods and ranges of normal values. We therefore determined normal humeral head retroversion and assessed the measurement methods. We measured retroversion in 65 cadaveric humeri, including 52 paired specimens, using four methods: radiographic, computed tomography (CT) scan, computer-assisted, and direct methods. We also assessed the distance between the humeral head central axis and the bicipital groove. CT scan methods accurately measure humeral head retroversion, while radiographic methods do not. The retroversion with respect to the transepicondylar axis was 17.9° and 21.5° with respect to the trochlear tangent axis. The difference between the right and left humeri was 8.9°. The distance between the central axis of the humeral head and the bicipital groove was 7.0 mm and was consistent between right and left humeri. Humeral head retroversion may be most accurately obtained using the patient’s own anatomic landmarks or, if not, identifiable retroversion as measured by those landmarks on contralateral side or the bicipital groove. PMID:18264854

  6. Accurate modelling of unsteady flows in collapsible tubes.

    PubMed

    Marchandise, Emilie; Flaud, Patrice

    2010-01-01

    The context of this paper is the development of a general and efficient numerical haemodynamic tool to help clinicians and researchers in understanding of physiological flow phenomena. We propose an accurate one-dimensional Runge-Kutta discontinuous Galerkin (RK-DG) method coupled with lumped parameter models for the boundary conditions. The suggested model has already been successfully applied to haemodynamics in arteries and is now extended for the flow in collapsible tubes such as veins. The main difference with cardiovascular simulations is that the flow may become supercritical and elastic jumps may appear with the numerical consequence that scheme may not remain monotone if no limiting procedure is introduced. We show that our second-order RK-DG method equipped with an approximate Roe's Riemann solver and a slope-limiting procedure allows us to capture elastic jumps accurately. Moreover, this paper demonstrates that the complex physics associated with such flows is more accurately modelled than with traditional methods such as finite difference methods or finite volumes. We present various benchmark problems that show the flexibility and applicability of the numerical method. Our solutions are compared with analytical solutions when they are available and with solutions obtained using other numerical methods. Finally, to illustrate the clinical interest, we study the emptying process in a calf vein squeezed by contracting skeletal muscle in a normal and pathological subject. We compare our results with experimental simulations and discuss the sensitivity to parameters of our model.

  7. Normalized medical information visualization.

    PubMed

    Sánchez-de-Madariaga, Ricardo; Muñoz, Adolfo; Somolinos, Roberto; Castro, Antonio; Velázquez, Iker; Moreno, Oscar; García-Pacheco, José L; Pascual, Mario; Salvador, Carlos H

    2015-01-01

    A new mark-up programming language is introduced in order to facilitate and improve the visualization of ISO/EN 13606 dual model-based normalized medical information. This is the first time that visualization of normalized medical information is addressed and the programming language is intended to be used by medical non-IT professionals.

  8. Normal Shock Vortex Interaction

    DTIC Science & Technology

    2003-03-01

    Figure 9: Breakdown map for normal-shock vortex-interaction. References [1] O. Thomer, W. Schroder and M. Meinke , Numerical Simulation of Normal...and Oblique-Shock Vortex Interaction, ZAMM Band 80, Sub. 1, pp. 181-184, 2000. [2] O. Thomer, E. Krause, W. Schroder and M. Meinke , Computational

  9. Transcription of mitochondrial DNA.

    PubMed

    Tabak, H F; Grivell, L A; Borst, P

    1983-01-01

    While mitochondrial DNA (mtDNA) is the simplest DNA in nature, coding for rRNAs and tRNAs, results of DNA sequence, and transcript analysis have demonstrated that both the synthesis and processing of mitochondrial RNAs involve remarkably intricate events. At one extreme, genes in animal mtDNAs are tightly packed, both DNA strands are completely transcribed (symmetric transcription), and the appearance of specific mRNAs is entirely dependent on processing at sites signalled by the sequences of the tRNAs, which abut virtually every gene. At the other extreme, gene organization in yeast (Saccharomyces) is anything but compact, with long stretches of AT-rich DNA interspaced between coding sequences and no obvious logic to the order of genes. Transcription is asymmetric and several RNAs are initiated de novo. Nevertheless, extensive RNA processing occurs due largely to the presence of split genes. RNA splicing is complex, is controlled by both mitochondrial and nuclear genes, and in some cases is accompanied by the formation of RNAs that behave as covalently closed circles. The present article reviews current knowledge of mitochondrial transcription and RNA processing in relation to possible mechanisms for the regulation of mitochondrial gene expression.

  10. Fungal CSL transcription factors

    PubMed Central

    Převorovský, Martin; Půta, František; Folk, Petr

    2007-01-01

    Background The CSL (CBF1/RBP-Jκ/Suppressor of Hairless/LAG-1) transcription factor family members are well-known components of the transmembrane receptor Notch signaling pathway, which plays a critical role in metazoan development. They function as context-dependent activators or repressors of transcription of their responsive genes, the promoters of which harbor the GTG(G/A)GAA consensus elements. Recently, several studies described Notch-independent activities of the CSL proteins. Results We have identified putative CSL genes in several fungal species, showing that this family is not confined to metazoans. We have analyzed their sequence conservation and identified the presence of well-defined domains typical of genuine CSL proteins. Furthermore, we have shown that the candidate fungal protein sequences contain highly conserved regions known to be required for sequence-specific DNA binding in their metazoan counterparts. The phylogenetic analysis of the newly identified fungal CSL proteins revealed the existence of two distinct classes, both of which are present in all the species studied. Conclusion Our findings support the evolutionary origin of the CSL transcription factor family in the last common ancestor of fungi and metazoans. We hypothesize that the ancestral CSL function involved DNA binding and Notch-independent regulation of transcription and that this function may still be shared, to a certain degree, by the present CSL family members from both fungi and metazoans. PMID:17629904

  11. Focus on Refugees. Transcript.

    ERIC Educational Resources Information Center

    Brandel, Sarah; And Others

    This is the transcript of the "Focus on Refugees," proqram conducted by the Overseas Development Council. Remarks from the following participants are included: (1) Sarah Brandel, Associate Fellow at the Overseas Development Council; (2) Gary Perkins, Chief of Mission of the Washington Office of the United Nations High Commissioner for Refugees…

  12. Narrative Transcription Accuracy and Reliability in Two Languages

    ERIC Educational Resources Information Center

    Heilmann, John; Miller, Jon F.; Iglesias, Aquiles; Fabiano-Smith, Leah; Nockerts, Ann; Andriacchi, Karen Digney

    2008-01-01

    A study was conducted to examine the issues of accuracy and reliability for transcription and analysis of oral narratives from Spanish-English bilingual children. Findings reveal that oral narrative data from ELL children can be accurately transcribed and the narrative measures are stable over time.

  13. A new method to synthesize competitor RNAs for accurate analyses by competitive RT-PCR.

    PubMed

    Ishibashi, O

    1997-12-03

    A method to synthesize competitor RNAs as internal standards for competitive RT-PCR is improved by using the long accurate PCR (LA-PCR) technique. Competitor templates synthesized by the new method are almost the same in length, and possibly in secondary structure, as target mRNAs to be quantified except that they include the short deletion within the segments to be amplified. This allows the reverse transcription to be achieved with almost the same efficiency from both target mRNAs and competitor RNAs. Therefore, more accurate quantification can be accomplished by using such competitor RNAs.

  14. Divisive normalization in olfactory population codes

    PubMed Central

    Olsen, Shawn R; Bhandawat, Vikas; Wilson, Rachel Irene

    2010-01-01

    In many regions of the visual system, the activity of a neuron is normalized by the activity of other neurons in the same region. Here we show that a similar normalization occurs during olfactory processing in the Drosophila antennal lobe. We exploit the orderly anatomy of this circuit to independently manipulate feedforward and lateral input to second-order projection neurons (PNs). Lateral inhibition increases the level of feedforward input needed to drive PNs to saturation, and this normalization scales with the total activity of the olfactory receptor neuron (ORN) population. Increasing total ORN activity also makes PN responses more transient. Strikingly, a model with just two variables (feedforward and total ORN activity) accurately predicts PN odor responses. Finally, we show that discrimination by a linear decoder is facilitated by two complementary transformations: the saturating transformation intrinsic to each processing channel boosts weak signals, while normalization helps equalize responses to different stimuli. PMID:20435004

  15. Accurate pointing of tungsten welding electrodes

    NASA Technical Reports Server (NTRS)

    Ziegelmeier, P.

    1971-01-01

    Thoriated-tungsten is pointed accurately and quickly by using sodium nitrite. Point produced is smooth and no effort is necessary to hold the tungsten rod concentric. The chemically produced point can be used several times longer than ground points. This method reduces time and cost of preparing tungsten electrodes.

  16. Immunohistochemical characteristics of normal canine eyes.

    PubMed

    Labelle, P; Reilly, C M; Naydan, D K; Labelle, A L

    2012-09-01

    Immunohistochemistry is widely utilized in diagnostic laboratories to study neoplastic and nonneoplastic diseases. Knowledge of the immunohistochemical characteristics of normal tissue is essential for interpretation of immunoreactivity in pathologic conditions. In this study, immunohistochemistry was performed with a broad panel of diagnostically relevant antibodies on 4 normal canine globes--namely, vimentin, pan-cytokeratin (AE1/AE3), cytokeratin 7, cytokeratin 8/18, cytokeratin 20, α-smooth muscle actin, muscle specific actin, desmin, Melan-A, microphthalmia transcription factor, S-100, glial fibrillary acidic protein, triple neurofilaments, neuron-specific enolase, chromogranin A, synaptophysin, laminin and CD31. Results include cytokeratin immunoreactivity limited to the conjunctival epithelium, corneal epithelium, and retinal pigment epithelium; distinct patterns of immunopositivity of muscle markers; and widespread immunoreactivity for vimentin and most neural/neuroendocrine markers. These findings in normal eyes provide the basis for interpretation of ocular immunohistochemistry in dogs. Published immunophenotypes of primary ocular neoplasms are also reviewed.

  17. Reference gene selection for quantitative real-time PCR normalization in Quercus suber.

    PubMed

    Marum, Liliana; Miguel, Andreia; Ricardo, Cândido P; Miguel, Célia

    2012-01-01

    The use of reverse transcription quantitative PCR technology to assess gene expression levels requires an accurate normalization of data in order to avoid misinterpretation of experimental results and erroneous analyses. Despite being the focus of several transcriptomics projects, oaks, and particularly cork oak (Quercus suber), have not been investigated regarding the identification of reference genes suitable for the normalization of real-time quantitative PCR data. In this study, ten candidate reference genes (Act, CACs, EF-1α, GAPDH, His3, PsaH, Sand, PP2A, ß-Tub and Ubq) were evaluated to determine the most stable internal reference for quantitative PCR normalization in cork oak. The transcript abundance of these genes was analysed in several tissues of cork oak, including leaves, reproduction cork, and periderm from branches at different developmental stages (1-, 2-, and 3-year old) or collected in different dates (active growth period versus dormancy). The three statistical methods (geNorm, NormFinder, and CV method) used in the evaluation of the most suitable combination of reference genes identified Act and CACs as the most stable candidates when all the samples were analysed together, while ß-Tub and PsaH showed the lowest expression stability. However, when different tissues, developmental stages, and collection dates were analysed separately, the reference genes exhibited some variation in their expression levels. In this study, and for the first time, we have identified and validated reference genes in cork oak that can be used for quantification of target gene expression in different tissues and experimental conditions and will be useful as a starting point for gene expression studies in other oaks.

  18. The AACRAO 2003 Academic Record and Transcript Guide. AACRAO Professional Development & Education Series.

    ERIC Educational Resources Information Center

    American Association of Collegiate Registrars and Admissions Officers, Washington, DC.

    This guide is a source of information on a wide range of issues involving student records and transcripts. It focuses on the necessity of reconciling the need to provide accurate information promptly to various constituencies and the need to safeguard privacy. Recommendations are provided for database and transcript elements, and current issues…

  19. Normal Modes Expose Active Sites in Enzymes

    PubMed Central

    Glantz-Gashai, Yitav; Samson, Abraham O.

    2016-01-01

    Accurate prediction of active sites is an important tool in bioinformatics. Here we present an improved structure based technique to expose active sites that is based on large changes of solvent accessibility accompanying normal mode dynamics. The technique which detects EXPOsure of active SITes through normal modEs is named EXPOSITE. The technique is trained using a small 133 enzyme dataset and tested using a large 845 enzyme dataset, both with known active site residues. EXPOSITE is also tested in a benchmark protein ligand dataset (PLD) comprising 48 proteins with and without bound ligands. EXPOSITE is shown to successfully locate the active site in most instances, and is found to be more accurate than other structure-based techniques. Interestingly, in several instances, the active site does not correspond to the largest pocket. EXPOSITE is advantageous due to its high precision and paves the way for structure based prediction of active site in enzymes. PMID:28002427

  20. SVD identifies transcript length distribution functions from DNA microarray data and reveals evolutionary forces globally affecting GBM metabolism.

    PubMed

    Bertagnolli, Nicolas M; Drake, Justin A; Tennessen, Jason M; Alter, Orly

    2013-01-01

    To search for evolutionary forces that might act upon transcript length, we use the singular value decomposition (SVD) to identify the length distribution functions of sets and subsets of human and yeast transcripts from profiles of mRNA abundance levels across gel electrophoresis migration distances that were previously measured by DNA microarrays. We show that the SVD identifies the transcript length distribution functions as "asymmetric generalized coherent states" from the DNA microarray data and with no a-priori assumptions. Comparing subsets of human and yeast transcripts of the same gene ontology annotations, we find that in both disparate eukaryotes, transcripts involved in protein synthesis or mitochondrial metabolism are significantly shorter than typical, and in particular, significantly shorter than those involved in glucose metabolism. Comparing the subsets of human transcripts that are overexpressed in glioblastoma multiforme (GBM) or normal brain tissue samples from The Cancer Genome Atlas, we find that GBM maintains normal brain overexpression of significantly short transcripts, enriched in transcripts that are involved in protein synthesis or mitochondrial metabolism, but suppresses normal overexpression of significantly longer transcripts, enriched in transcripts that are involved in glucose metabolism and brain activity. These global relations among transcript length, cellular metabolism and tumor development suggest a previously unrecognized physical mode for tumor and normal cells to differentially regulate metabolism in a transcript length-dependent manner. The identified distribution functions support a previous hypothesis from mathematical modeling of evolutionary forces that act upon transcript length in the manner of the restoring force of the harmonic oscillator.

  1. Normality in Analytical Psychology

    PubMed Central

    Myers, Steve

    2013-01-01

    Although C.G. Jung’s interest in normality wavered throughout his career, it was one of the areas he identified in later life as worthy of further research. He began his career using a definition of normality which would have been the target of Foucault’s criticism, had Foucault chosen to review Jung’s work. However, Jung then evolved his thinking to a standpoint that was more aligned to Foucault’s own. Thereafter, the post Jungian concept of normality has remained relatively undeveloped by comparison with psychoanalysis and mainstream psychology. Jung’s disjecta membra on the subject suggest that, in contemporary analytical psychology, too much focus is placed on the process of individuation to the neglect of applications that consider collective processes. Also, there is potential for useful research and development into the nature of conflict between individuals and societies, and how normal people typically develop in relation to the spectrum between individuation and collectivity. PMID:25379262

  2. Normal Pressure Hydrocephalus

    MedlinePlus

    ... techniques and neuroimaging, and finding improved treatments and preventions. Information from the National Library of Medicine’s MedlinePlus Normal Pressure Hydrocephalus × What research is being ...

  3. Normal Functioning Family

    MedlinePlus

    ... Spread the Word Shop AAP Find a Pediatrician Family Life Medical Home Family Dynamics Adoption & Foster Care ... Español Text Size Email Print Share Normal Functioning Family Page Content Article Body Is there any way ...

  4. Feedback about More Accurate versus Less Accurate Trials: Differential Effects on Self-Confidence and Activation

    ERIC Educational Resources Information Center

    Badami, Rokhsareh; VaezMousavi, Mohammad; Wulf, Gabriele; Namazizadeh, Mahdi

    2012-01-01

    One purpose of the present study was to examine whether self-confidence or anxiety would be differentially affected by feedback from more accurate rather than less accurate trials. The second purpose was to determine whether arousal variations (activation) would predict performance. On Day 1, participants performed a golf putting task under one of…

  5. Cosmic shear covariance: the log-normal approximation

    NASA Astrophysics Data System (ADS)

    Hilbert, S.; Hartlap, J.; Schneider, P.

    2011-12-01

    Context. Accurate estimates of the errors on the cosmological parameters inferred from cosmic shear surveys require accurate estimates of the covariance of the cosmic shear correlation functions. Aims: We seek approximations to the cosmic shear covariance that are as easy to use as the common approximations based on normal (Gaussian) statistics, but yield more accurate covariance matrices and parameter errors. Methods: We derive expressions for the cosmic shear covariance under the assumption that the underlying convergence field follows log-normal statistics. We also derive a simplified version of this log-normal approximation by only retaining the most important terms beyond normal statistics. We use numerical simulations of weak lensing to study how well the normal, log-normal, and simplified log-normal approximations as well as empirical corrections to the normal approximation proposed in the literature reproduce shear covariances for cosmic shear surveys. We also investigate the resulting confidence regions for cosmological parameters inferred from such surveys. Results: We find that the normal approximation substantially underestimates the cosmic shear covariances and the inferred parameter confidence regions, in particular for surveys with small fields of view and large galaxy densities, but also for very wide surveys. In contrast, the log-normal approximation yields more realistic covariances and confidence regions, but also requires evaluating slightly more complicated expressions. However, the simplified log-normal approximation, although as simple as the normal approximation, yields confidence regions that are almost as accurate as those obtained from the log-normal approximation. The empirical corrections to the normal approximation do not yield more accurate covariances and confidence regions than the (simplified) log-normal approximation. Moreover, they fail to produce positive-semidefinite data covariance matrices in certain cases, rendering them

  6. A method for normalization of X-ray absorption spectra

    SciTech Connect

    Weng, T.-C.; Waldo, G.S.; Penner-Hahn, J.E.

    2010-07-20

    Accurate normalization of X-ray absorption data is essential for quantitative analysis of near-edge features. A method, implemented as the program MBACK, to normalize X-ray absorption data to tabulated mass absorption coefficients is described. Comparison of conventional normalization methods with MBACK demonstrates that the new normalization method is not sensitive to the shape of the background function, thus allowing accurate comparison of data collected in transmission mode with data collected using fluorescence ion chambers or solid-state fluorescence detectors. The new method is shown to have better reliability and consistency and smaller errors than conventional normalization methods. The sensitivity of the new normalization method is illustrated by analysis of data collected during an equilibrium titration.

  7. Genetic and epigenetic control of RKIP transcription.

    PubMed

    Datar, Ila; Tegegne, Hanna; Qin, Kevin; Al-Mulla, Fahd; Bitar, Milad S; Trumbly, Robert J; Yeung, Kam C

    2014-01-01

    Raf kinase inhibitory protein (RKIP) is known to modulate key signaling cascades and regulate normal physiological processes such as cellular proliferation, differentiation, and apoptosis. The expression of RKIP is found to be downregulated in several cancer metastases and the repressed RKIP expression can be reactivated on treatment with chemotherapeutic agents. RKIP is a proven tumor metastasis suppressor gene and investigating the mechanisms of transcriptional regulation of RKIP is therefore of immense clinical importance. In this review, we discuss the basal expression of RKIP in various tissues and the genetic aspects of the RKIP chromosomal locus including the structure of the RKIP promoter as well as gene regulatory elements such as enhancers. We also review the genetic and epigenetic modulation of RKIP transcription through EZH2, a component of the polycomb repressive complex 2 (PRC2) and sequence specific transcription factors (TFs) BACH1 and Snail. Emerging experimental evidence supports a unifying model in which both these TFs repress RKIP transcription in cancers by recruiting the EZH2 containing repressive complex to the proximal RKIP promoter. Finally, we review the known mechanisms employed by different types of chemotherapeutic agents to activate RKIP expression in cancer cells.

  8. Transcriptional control of cardiac fibroblast plasticity.

    PubMed

    Lighthouse, Janet K; Small, Eric M

    2016-02-01

    Cardiac fibroblasts help maintain the normal architecture of the healthy heart and are responsible for scar formation and the healing response to pathological insults. Various genetic, biomechanical, or humoral factors stimulate fibroblasts to become contractile smooth muscle-like cells called myofibroblasts that secrete large amounts of extracellular matrix. Unfortunately, unchecked myofibroblast activation in heart disease leads to pathological fibrosis, which is a major risk factor for the development of cardiac arrhythmias and heart failure. A better understanding of the molecular mechanisms that control fibroblast plasticity and myofibroblast activation is essential to develop novel strategies to specifically target pathological cardiac fibrosis without disrupting the adaptive healing response. This review highlights the major transcriptional mediators of fibroblast origin and function in development and disease. The contribution of the fetal epicardial gene program will be discussed in the context of fibroblast origin in development and following injury, primarily focusing on Tcf21 and C/EBP. We will also highlight the major transcriptional regulatory axes that control fibroblast plasticity in the adult heart, including transforming growth factor β (TGFβ)/Smad signaling, the Rho/myocardin-related transcription factor (MRTF)/serum response factor (SRF) axis, and Calcineurin/transient receptor potential channel (TRP)/nuclear factor of activated T-Cell (NFAT) signaling. Finally, we will discuss recent strategies to divert the fibroblast transcriptional program in an effort to promote cardiomyocyte regeneration. This article is a part of a Special Issue entitled "Fibrosis and Myocardial Remodeling".

  9. Machine Transcription--Practically Speaking.

    ERIC Educational Resources Information Center

    Clippinger, Dorinda A.

    1984-01-01

    Draws transcription teaching principles from Gagne's theories about learning. Recommends 12-16 weeks of instruction, pre-transcription development of related skills, frequent feedback, and use of teaching materials that are arranged to take advantage of learning cycles. (SK)

  10. New model accurately predicts reformate composition

    SciTech Connect

    Ancheyta-Juarez, J.; Aguilar-Rodriguez, E. )

    1994-01-31

    Although naphtha reforming is a well-known process, the evolution of catalyst formulation, as well as new trends in gasoline specifications, have led to rapid evolution of the process, including: reactor design, regeneration mode, and operating conditions. Mathematical modeling of the reforming process is an increasingly important tool. It is fundamental to the proper design of new reactors and revamp of existing ones. Modeling can be used to optimize operating conditions, analyze the effects of process variables, and enhance unit performance. Instituto Mexicano del Petroleo has developed a model of the catalytic reforming process that accurately predicts reformate composition at the higher-severity conditions at which new reformers are being designed. The new AA model is more accurate than previous proposals because it takes into account the effects of temperature and pressure on the rate constants of each chemical reaction.

  11. Accurate colorimetric feedback for RGB LED clusters

    NASA Astrophysics Data System (ADS)

    Man, Kwong; Ashdown, Ian

    2006-08-01

    We present an empirical model of LED emission spectra that is applicable to both InGaN and AlInGaP high-flux LEDs, and which accurately predicts their relative spectral power distributions over a wide range of LED junction temperatures. We further demonstrate with laboratory measurements that changes in LED spectral power distribution with temperature can be accurately predicted with first- or second-order equations. This provides the basis for a real-time colorimetric feedback system for RGB LED clusters that can maintain the chromaticity of white light at constant intensity to within +/-0.003 Δuv over a range of 45 degrees Celsius, and to within 0.01 Δuv when dimmed over an intensity range of 10:1.

  12. On accurate determination of contact angle

    NASA Technical Reports Server (NTRS)

    Concus, P.; Finn, R.

    1992-01-01

    Methods are proposed that exploit a microgravity environment to obtain highly accurate measurement of contact angle. These methods, which are based on our earlier mathematical results, do not require detailed measurement of a liquid free-surface, as they incorporate discontinuous or nearly-discontinuous behavior of the liquid bulk in certain container geometries. Physical testing is planned in the forthcoming IML-2 space flight and in related preparatory ground-based experiments.

  13. Validation of reference transcripts in strawberry (Fragaria spp.).

    PubMed

    Clancy, Maureen A; Rosli, Hernan G; Chamala, Srikar; Barbazuk, W Brad; Civello, P Marcos; Folta, Kevin M

    2013-12-01

    Contemporary methods to assay gene expression depend on a stable set of reference transcripts for accurate quantitation. A lack of well-tested reference genes slows progress in characterizing gene expression in high-value specialty crops. In this study, a set of strawberry (Fragaria spp.) constitutively expressed reference genes has been identified by merging digital gene expression data with expression profiling. Constitutive reference candidates were validated using quantitative PCR and hybridization. Several transcripts have been identified that show improved stability across tissues relative to traditional reference transcripts. Results are similar between commercial octoploid strawberry and the diploid model. Our findings also show that while some never-before-used references are appropriate for most applications, even the most stable reference transcripts require careful assessment across the diverse tissues and fruit developmental states before being adopted as controls.

  14. Untangling the brain's neuroinflammatory and neurodegenerative transcriptional responses

    PubMed Central

    Srinivasan, Karpagam; Friedman, Brad A.; Larson, Jessica L.; Lauffer, Benjamin E.; Goldstein, Leonard D.; Appling, Laurie L.; Borneo, Jovencio; Poon, Chungkee; Ho, Terence; Cai, Fang; Steiner, Pascal; van der Brug, Marcel P.; Modrusan, Zora; Kaminker, Joshua S.; Hansen, David V.

    2016-01-01

    A common approach to understanding neurodegenerative disease is comparing gene expression in diseased versus healthy tissues. We illustrate that expression profiles derived from whole tissue RNA highly reflect the degenerating tissues' altered cellular composition, not necessarily transcriptional regulation. To accurately understand transcriptional changes that accompany neuropathology, we acutely purify neurons, astrocytes and microglia from single adult mouse brains and analyse their transcriptomes by RNA sequencing. Using peripheral endotoxemia to establish the method, we reveal highly specific transcriptional responses and altered RNA processing in each cell type, with Tnfr1 required for the astrocytic response. Extending the method to an Alzheimer's disease model, we confirm that transcriptomic changes observed in whole tissue are driven primarily by cell type composition, not transcriptional regulation, and identify hundreds of cell type-specific changes undetected in whole tissue RNA. Applying similar methods to additional models and patient tissues will transform our understanding of aberrant gene expression in neurological disease. PMID:27097852

  15. Apparent diffusion coefficient normalization of normal liver

    PubMed Central

    Zhu, Jie; Zhang, Jie; Gao, Jia-Yin; Li, Jin-Ning; Yang, Da-Wei; Chen, Min; Zhou, Cheng; Yang, Zheng-Han

    2017-01-01

    Abstract Apparent diffusion coefficient (ADC) measurement in diffusion-weighted imaging (DWI) has been reported to be a helpful biomarker for detection and characterization of lesion. In view of the importance of ADC measurement reproducibility, the aim of this study was to probe the variability of the healthy hepatic ADC values measured at 3 MR scanners from different vendors and with different field strengths, and to investigate the reproducibility of normalized ADC (nADC) value with the spleen as the reference organ. Thirty enrolled healthy volunteers received DWI with GE 1.5T, Siemens 1.5T, and Philips 3.0T magnetic resonance (MR) systems on liver and spleen (session 1) and were imaged again after 10 to 14 days using only GE 1.5T MR and Philips 3.0T MR systems (session 2). Interscan agreement and reproducibility of ADC measurements of liver and the calculated nADC values (ADCliver/ADCspleen) were statistically evaluated between 2 sessions. In session 1, ADC and nADC values of liver were evaluated for the scanner-related variability by 2-way analysis of variance and intraclass correlation coefficients (ICCs). Coefficients of variation (CVs) of ADCs and nADCs of liver were calculated for both 1.5 and 3.0-T MR system. Interscan agreement and reproducibility of ADC measurements of liver and related nADCs between 2 sessions were found to be satisfactory with ICC values of 0.773 to 0.905. In session 1, the liver nADCs obtained from different scanners were consistent (P = 0.112) without any significant difference in multiple comparison (P = 0.117 to >0.99) by using 2-way analysis of variance with post-hoc analysis of Bonferroni method, although the liver ADCs varied significantly (P < 0.001). nADCs measured by 3 scanners were in good interscanner agreements with ICCs of 0.685 to 0.776. The mean CV of nADCs of both 1.5T MR scanners (9.6%) was similar to that of 3.0T MR scanner (8.9%). ADCs measured at 3 MR scanners with different field strengths and vendors

  16. The WRKY transcription factor family in Brachypodium distachyon

    PubMed Central

    2012-01-01

    Background A complete assembled genome sequence of wheat is not yet available. Therefore, model plant systems for wheat are very valuable. Brachypodium distachyon (Brachypodium) is such a system. The WRKY family of transcription factors is one of the most important families of plant transcriptional regulators with members regulating important agronomic traits. Studies of WRKY transcription factors in Brachypodium and wheat therefore promise to lead to new strategies for wheat improvement. Results We have identified and manually curated the WRKY transcription factor family from Brachypodium using a pipeline designed to identify all potential WRKY genes. 86 WRKY transcription factors were found, a total higher than all other current databases. We therefore propose that our numbering system (BdWRKY1-BdWRKY86) becomes the standard nomenclature. In the JGI v1.0 assembly of Brachypodium with the MIPS/JGI v1.0 annotation, nine of the transcription factors have no gene model and eleven gene models are probably incorrectly predicted. In total, twenty WRKY transcription factors (23.3%) do not appear to have accurate gene models. To facilitate use of our data, we have produced The Database of Brachypodium distachyon WRKY Transcription Factors. Each WRKY transcription factor has a gene page that includes predicted protein domains from MEME analyses. These conserved protein domains reflect possible input and output domains in signaling. The database also contains a BLAST search function where a large dataset of WRKY transcription factors, published genes, and an extensive set of wheat ESTs can be searched. We also produced a phylogram containing the WRKY transcription factor families from Brachypodium, rice, Arabidopsis, soybean, and Physcomitrella patens, together with published WRKY transcription factors from wheat. This phylogenetic tree provides evidence for orthologues, co-orthologues, and paralogues of Brachypodium WRKY transcription factors. Conclusions The description

  17. Non-transcriptional regulatory processes shape transcriptional network dynamics.

    PubMed

    Ray, J Christian J; Tabor, Jeffrey J; Igoshin, Oleg A

    2011-10-11

    Information about the extra- or intracellular environment is often captured as biochemical signals that propagate through regulatory networks. These signals eventually drive phenotypic changes, typically by altering gene expression programmes in the cell. Reconstruction of transcriptional regulatory networks has given a compelling picture of bacterial physiology, but transcriptional network maps alone often fail to describe phenotypes. Cellular response dynamics are ultimately determined by interactions between transcriptional and non-transcriptional networks, with dramatic implications for physiology and evolution. Here, we provide an overview of non-transcriptional interactions that can affect the performance of natural and synthetic bacterial regulatory networks.

  18. High Frequency QRS ECG Accurately Detects Cardiomyopathy

    NASA Technical Reports Server (NTRS)

    Schlegel, Todd T.; Arenare, Brian; Poulin, Gregory; Moser, Daniel R.; Delgado, Reynolds

    2005-01-01

    High frequency (HF, 150-250 Hz) analysis over the entire QRS interval of the ECG is more sensitive than conventional ECG for detecting myocardial ischemia. However, the accuracy of HF QRS ECG for detecting cardiomyopathy is unknown. We obtained simultaneous resting conventional and HF QRS 12-lead ECGs in 66 patients with cardiomyopathy (EF = 23.2 plus or minus 6.l%, mean plus or minus SD) and in 66 age- and gender-matched healthy controls using PC-based ECG software recently developed at NASA. The single most accurate ECG parameter for detecting cardiomyopathy was an HF QRS morphological score that takes into consideration the total number and severity of reduced amplitude zones (RAZs) present plus the clustering of RAZs together in contiguous leads. This RAZ score had an area under the receiver operator curve (ROC) of 0.91, and was 88% sensitive, 82% specific and 85% accurate for identifying cardiomyopathy at optimum score cut-off of 140 points. Although conventional ECG parameters such as the QRS and QTc intervals were also significantly longer in patients than controls (P less than 0.001, BBBs excluded), these conventional parameters were less accurate (area under the ROC = 0.77 and 0.77, respectively) than HF QRS morphological parameters for identifying underlying cardiomyopathy. The total amplitude of the HF QRS complexes, as measured by summed root mean square voltages (RMSVs), also differed between patients and controls (33.8 plus or minus 11.5 vs. 41.5 plus or minus 13.6 mV, respectively, P less than 0.003), but this parameter was even less accurate in distinguishing the two groups (area under ROC = 0.67) than the HF QRS morphologic and conventional ECG parameters. Diagnostic accuracy was optimal (86%) when the RAZ score from the HF QRS ECG and the QTc interval from the conventional ECG were used simultaneously with cut-offs of greater than or equal to 40 points and greater than or equal to 445 ms, respectively. In conclusion 12-lead HF QRS ECG employing

  19. Transcriptional regulation of epithelial-mesenchymal transition.

    PubMed

    Teng, Yingqi; Zeisberg, Michael; Kalluri, Raghu

    2007-02-01

    It has become increasingly obvious that the notion of a terminally differentiated cell is likely a simplified concept. Epithelial-mesenchymal transition (EMT), during which epithelial cells assume a mesenchymal phenotype, is a key event occurring during normal development and pathological processes. Multiple extracellular stimuli and transcriptional regulators can trigger EMT, but how such distinct signaling pathways orchestrate the complex cellular events that facilitate EMT is not well understood. In this issue of the JCI, Venkov et al. report on their examination of fibroblasts resulting from EMT and describe a novel protein-DNA complex that is essential for transcription of fibroblast-specific protein 1 (FSP1) and sufficient to induce early EMT events (see the related article beginning on page 482). Collectively, their results suggest that this complex is an important regulator of the EMT transcriptome.

  20. Transcriptional regulation of epithelial-mesenchymal transition

    PubMed Central

    Teng, Yingqi; Zeisberg, Michael; Kalluri, Raghu

    2007-01-01

    It has become increasingly obvious that the notion of a terminally differentiated cell is likely a simplified concept. Epithelial-mesenchymal transition (EMT), during which epithelial cells assume a mesenchymal phenotype, is a key event occurring during normal development and pathological processes. Multiple extracellular stimuli and transcriptional regulators can trigger EMT, but how such distinct signaling pathways orchestrate the complex cellular events that facilitate EMT is not well understood. In this issue of the JCI, Venkov et al. report on their examination of fibroblasts resulting from EMT and describe a novel protein-DNA complex that is essential for transcription of fibroblast-specific protein 1 (FSP1) and sufficient to induce early EMT events (see the related article beginning on page 482). Collectively, their results suggest that this complex is an important regulator of the EMT transcriptome. PMID:17273552

  1. Jade data transcription system final report

    SciTech Connect

    Eaton, R.; Iskra, M.; McLean, J. . Advanced Technology Div.)

    1990-07-25

    The OWL sensor, which is used in conjunction with the Jade program, generates a tremendous volume of data during normal field operations. Historically, the dissemination of this data to analysts has been slowed by difficulties in transcribing to a widely readable media and format. TRW, under contract from Lawrence Livermore National Laboratory, was tasked by Defense Advanced Research Projects Agency (DARPA) with finding an improved method of transcribing the Jade experimental data. During the period of performance on this contract TRW helped to guide the development and operation of an improved transcription system. This final report summarizes the work performed, and provides a written record of information which may be helpful to future users of the newly developed data transcription system. 4 figs.

  2. Transcriptional Targeting in Cancer Gene Therapy

    PubMed Central

    2003-01-01

    Cancer gene therapy has been one of the most exciting areas of therapeutic research in the past decade. In this review, we discuss strategies to restrict transcription of transgenes to tumour cells. A range of promoters which are tissue-specific, tumour-specific, or inducible by exogenous agents are presented. Transcriptional targeting should prevent normal tissue toxicities associated with other cancer treatments, such as radiation and chemotherapy. In addition, the specificity of these strategies should provide improved targeting of metastatic tumours following systemic gene delivery. Rapid progress in the ability to specifically control transgenes will allow systemic gene delivery for cancer therapy to become a real possibility in the near future. PMID:12721516

  3. Mechanism of transcription initiation by the yeast mitochondrial RNA polymerase.

    PubMed

    Deshpande, Aishwarya P; Patel, Smita S

    2012-01-01

    Mitochondria are the major supplier of cellular energy in the form of ATP. Defects in normal ATP production due to dysfunctions in mitochondrial gene expression are responsible for many mitochondrial and aging related disorders. Mitochondria carry their own DNA genome which is transcribed by relatively simple transcriptional machinery consisting of the mitochondrial RNAP (mtRNAP) and one or more transcription factors. The mtRNAPs are remarkably similar in sequence and structure to single-subunit bacteriophage T7 RNAP but they require accessory transcription factors for promoter-specific initiation. Comparison of the mechanisms of T7 RNAP and mtRNAP provides a framework to better understand how mtRNAP and the transcription factors work together to facilitate promoter selection, DNA melting, initiating nucleotide binding, and promoter clearance. This review focuses primarily on the mechanistic characterization of transcription initiation by the yeast Saccharomyces cerevisiae mtRNAP (Rpo41) and its transcription factor (Mtf1) drawing insights from the homologous T7 and the human mitochondrial transcription systems. We discuss regulatory mechanisms of mitochondrial transcription and the idea that the mtRNAP acts as the in vivo ATP "sensor" to regulate gene expression. This article is part of a Special Issue entitled: Mitochondrial Gene Expression.

  4. Nascent transcription affected by RNA polymerase IV in Zea mays.

    PubMed

    Erhard, Karl F; Talbot, Joy-El R B; Deans, Natalie C; McClish, Allison E; Hollick, Jay B

    2015-04-01

    All eukaryotes use three DNA-dependent RNA polymerases (RNAPs) to create cellular RNAs from DNA templates. Plants have additional RNAPs related to Pol II, but their evolutionary role(s) remain largely unknown. Zea mays (maize) RNA polymerase D1 (RPD1), the largest subunit of RNA polymerase IV (Pol IV), is required for normal plant development, paramutation, transcriptional repression of certain transposable elements (TEs), and transcriptional regulation of specific alleles. Here, we define the nascent transcriptomes of rpd1 mutant and wild-type (WT) seedlings using global run-on sequencing (GRO-seq) to identify the broader targets of RPD1-based regulation. Comparisons of WT and rpd1 mutant GRO-seq profiles indicate that Pol IV globally affects transcription at both transcriptional start sites and immediately downstream of polyadenylation addition sites. We found no evidence of divergent transcription from gene promoters as seen in mammalian GRO-seq profiles. Statistical comparisons identify genes and TEs whose transcription is affected by RPD1. Most examples of significant increases in genic antisense transcription appear to be initiated by 3'-proximal long terminal repeat retrotransposons. These results indicate that maize Pol IV specifies Pol II-based transcriptional regulation for specific regions of the maize genome including genes having developmental significance.

  5. Quantifying surface normal estimation

    NASA Astrophysics Data System (ADS)

    Reid, Robert B.; Oxley, Mark E.; Eismann, Michael T.; Goda, Matthew E.

    2006-05-01

    An inverse algorithm for surface normal estimation from thermal polarimetric imagery was developed and used to quantify the requirements on a priori information. Building on existing knowledge that calculates the degree of linear polarization (DOLP) and the angle of polarization (AOP) for a given surface normal in a forward model (from an object's characteristics to calculation of the DOLP and AOP), this research quantifies the impact of a priori information with the development of an inverse algorithm to estimate surface normals from thermal polarimetric emissions in long-wave infrared (LWIR). The inverse algorithm assumes a polarized infrared focal plane array capturing LWIR intensity images which are then converted to Stokes vectors. Next, the DOLP and AOP are calculated from the Stokes vectors. Last, the viewing angles, θ v, to the surface normals are estimated assuming perfect material information about the imaged scene. A sensitivity analysis is presented to quantitatively describe the a priori information's impact on the amount of error in the estimation of surface normals, and a bound is determined given perfect information about an object. Simulations explored the impact of surface roughness (σ) and the real component (n) of a dielectric's complex index of refraction across a range of viewing angles (θ v) for a given wavelength of observation.

  6. Are Children "Normal"?

    PubMed

    Black, Dan A; Kolesnikova, Natalia; Sanders, Seth G; Taylor, Lowell J

    2013-03-01

    We examine Becker's (1960) contention that children are "normal." For the cross section of non-Hispanic white married couples in the U.S., we show that when we restrict comparisons to similarly-educated women living in similarly-expensive locations, completed fertility is positively correlated with the husband's income. The empirical evidence is consistent with children being "normal." In an effort to show causal effects, we analyze the localized impact on fertility of the mid-1970s increase in world energy prices - an exogenous shock that substantially increased men's incomes in the Appalachian coal-mining region. Empirical evidence for that population indicates that fertility increases in men's income.

  7. CT of Normal Developmental and Variant Anatomy of the Pediatric Skull: Distinguishing Trauma from Normality.

    PubMed

    Idriz, Sanjin; Patel, Jaymin H; Ameli Renani, Seyed; Allan, Rosemary; Vlahos, Ioannis

    2015-01-01

    The use of computed tomography (CT) in clinical practice has been increasing rapidly, with the number of CT examinations performed in adults and children rising by 10% per year in England. Because the radiology community strives to reduce the radiation dose associated with pediatric examinations, external factors, including guidelines for pediatric head injury, are raising expectations for use of cranial CT in the pediatric population. Thus, radiologists are increasingly likely to encounter pediatric head CT examinations in daily practice. The variable appearance of cranial sutures at different ages can be confusing for inexperienced readers of radiologic images. The evolution of multidetector CT with thin-section acquisition increases the clarity of some of these sutures, which may be misinterpreted as fractures. Familiarity with the normal anatomy of the pediatric skull, how it changes with age, and normal variants can assist in translating the increased resolution of multidetector CT into more accurate detection of fractures and confident determination of normality, thereby reducing prolonged hospitalization of children with normal developmental structures that have been misinterpreted as fractures. More important, the potential morbidity and mortality related to false-negative interpretation of fractures as normal sutures may be avoided. The authors describe the normal anatomy of all standard pediatric sutures, common variants, and sutural mimics, thereby providing an accurate and safe framework for CT evaluation of skull trauma in pediatric patients.

  8. Accurate upwind methods for the Euler equations

    NASA Technical Reports Server (NTRS)

    Huynh, Hung T.

    1993-01-01

    A new class of piecewise linear methods for the numerical solution of the one-dimensional Euler equations of gas dynamics is presented. These methods are uniformly second-order accurate, and can be considered as extensions of Godunov's scheme. With an appropriate definition of monotonicity preservation for the case of linear convection, it can be shown that they preserve monotonicity. Similar to Van Leer's MUSCL scheme, they consist of two key steps: a reconstruction step followed by an upwind step. For the reconstruction step, a monotonicity constraint that preserves uniform second-order accuracy is introduced. Computational efficiency is enhanced by devising a criterion that detects the 'smooth' part of the data where the constraint is redundant. The concept and coding of the constraint are simplified by the use of the median function. A slope steepening technique, which has no effect at smooth regions and can resolve a contact discontinuity in four cells, is described. As for the upwind step, existing and new methods are applied in a manner slightly different from those in the literature. These methods are derived by approximating the Euler equations via linearization and diagonalization. At a 'smooth' interface, Harten, Lax, and Van Leer's one intermediate state model is employed. A modification for this model that can resolve contact discontinuities is presented. Near a discontinuity, either this modified model or a more accurate one, namely, Roe's flux-difference splitting. is used. The current presentation of Roe's method, via the conceptually simple flux-vector splitting, not only establishes a connection between the two splittings, but also leads to an admissibility correction with no conditional statement, and an efficient approximation to Osher's approximate Riemann solver. These reconstruction and upwind steps result in schemes that are uniformly second-order accurate and economical at smooth regions, and yield high resolution at discontinuities.

  9. Accurate measurement of unsteady state fluid temperature

    NASA Astrophysics Data System (ADS)

    Jaremkiewicz, Magdalena

    2017-03-01

    In this paper, two accurate methods for determining the transient fluid temperature were presented. Measurements were conducted for boiling water since its temperature is known. At the beginning the thermometers are at the ambient temperature and next they are immediately immersed into saturated water. The measurements were carried out with two thermometers of different construction but with the same housing outer diameter equal to 15 mm. One of them is a K-type industrial thermometer widely available commercially. The temperature indicated by the thermometer was corrected considering the thermometers as the first or second order inertia devices. The new design of a thermometer was proposed and also used to measure the temperature of boiling water. Its characteristic feature is a cylinder-shaped housing with the sheath thermocouple located in its center. The temperature of the fluid was determined based on measurements taken in the axis of the solid cylindrical element (housing) using the inverse space marching method. Measurements of the transient temperature of the air flowing through the wind tunnel using the same thermometers were also carried out. The proposed measurement technique provides more accurate results compared with measurements using industrial thermometers in conjunction with simple temperature correction using the inertial thermometer model of the first or second order. By comparing the results, it was demonstrated that the new thermometer allows obtaining the fluid temperature much faster and with higher accuracy in comparison to the industrial thermometer. Accurate measurements of the fast changing fluid temperature are possible due to the low inertia thermometer and fast space marching method applied for solving the inverse heat conduction problem.

  10. The first accurate description of an aurora

    NASA Astrophysics Data System (ADS)

    Schröder, Wilfried

    2006-12-01

    As technology has advanced, the scientific study of auroral phenomena has increased by leaps and bounds. A look back at the earliest descriptions of aurorae offers an interesting look into how medieval scholars viewed the subjects that we study.Although there are earlier fragmentary references in the literature, the first accurate description of the aurora borealis appears to be that published by the German Catholic scholar Konrad von Megenberg (1309-1374) in his book Das Buch der Natur (The Book of Nature). The book was written between 1349 and 1350.

  11. Determining accurate distances to nearby galaxies

    NASA Astrophysics Data System (ADS)

    Bonanos, Alceste Zoe

    2005-11-01

    Determining accurate distances to nearby or distant galaxies is a very simple conceptually, yet complicated in practice, task. Presently, distances to nearby galaxies are only known to an accuracy of 10-15%. The current anchor galaxy of the extragalactic distance scale is the Large Magellanic Cloud, which has large (10-15%) systematic uncertainties associated with it, because of its morphology, its non-uniform reddening and the unknown metallicity dependence of the Cepheid period-luminosity relation. This work aims to determine accurate distances to some nearby galaxies, and subsequently help reduce the error in the extragalactic distance scale and the Hubble constant H 0 . In particular, this work presents the first distance determination of the DIRECT Project to M33 with detached eclipsing binaries. DIRECT aims to obtain a new anchor galaxy for the extragalactic distance scale by measuring direct, accurate (to 5%) distances to two Local Group galaxies, M31 and M33, with detached eclipsing binaries. It involves a massive variability survey of these galaxies and subsequent photometric and spectroscopic follow-up of the detached binaries discovered. In this work, I also present a catalog of variable stars discovered in one of the DIRECT fields, M31Y, which includes 41 eclipsing binaries. Additionally, we derive the distance to the Draco Dwarf Spheroidal galaxy, with ~100 RR Lyrae found in our first CCD variability study of this galaxy. A "hybrid" method of discovering Cepheids with ground-based telescopes is described next. It involves applying the image subtraction technique on the images obtained from ground-based telescopes and then following them up with the Hubble Space Telescope to derive Cepheid period-luminosity distances. By re-analyzing ESO Very Large Telescope data on M83 (NGC 5236), we demonstrate that this method is much more powerful for detecting variability, especially in crowded fields. I finally present photometry for the Wolf-Rayet binary WR 20a

  12. New law requires 'medically accurate' lesson plans.

    PubMed

    1999-09-17

    The California Legislature has passed a bill requiring all textbooks and materials used to teach about AIDS be medically accurate and objective. Statements made within the curriculum must be supported by research conducted in compliance with scientific methods, and published in peer-reviewed journals. Some of the current lesson plans were found to contain scientifically unsupported and biased information. In addition, the bill requires material to be "free of racial, ethnic, or gender biases." The legislation is supported by a wide range of interests, but opposed by the California Right to Life Education Fund, because they believe it discredits abstinence-only material.

  13. Nucleotides of transcription factor binding sites exert interdependent effects on the binding affinities of transcription factors

    PubMed Central

    Bulyk, Martha L.; Johnson, Philip L. F.; Church, George M.

    2002-01-01

    We can determine the effects of many possible sequence variations in transcription factor binding sites using microarray binding experiments. Analysis of wild-type and mutant Zif268 (Egr1) zinc fingers bound to microarrays containing all possible central 3 bp triplet binding sites indicates that the nucleotides of transcription factor binding sites cannot be treated independently. This indicates that the current practice of characterizing transcription factor binding sites by mutating individual positions of binding sites one base pair at a time does not provide a true picture of the sequence specificity. Similarly, current bioinformatic practices using either just a consensus sequence, or even mononucleotide frequency weight matrices to provide more complete descriptions of transcription factor binding sites, are not accurate in depicting the true binding site specificities, since these methods rely upon the assumption that the nucleotides of binding sites exert independent effects on binding affinity. Our results stress the importance of complete reference tables of all possible binding sites for comparing protein binding preferences for various DNA sequences. We also show results suggesting that microarray binding data using particular subsets of all possible binding sites can be used to extrapolate the relative binding affinities of all possible full-length binding sites, given a known binding site for use as a starting sequence for site preference refinement. PMID:11861919

  14. Normals to a Parabola

    ERIC Educational Resources Information Center

    Srinivasan, V. K.

    2013-01-01

    Given a parabola in the standard form y[superscript 2] = 4ax, corresponding to three points on the parabola, such that the normals at these three points P, Q, R concur at a point M = (h, k), the equation of the circumscribing circle through the three points P, Q, and R provides a tremendous opportunity to illustrate "The Art of Algebraic…

  15. Transfection of mouse ribosomal DNA into rat cells: faithful transcription and processing.

    PubMed Central

    Vance, V B; Thompson, E A; Bowman, L H

    1985-01-01

    Truncated mouse ribosomal DNA (rDNA) genes were stably incorporated into rat HTC-5 cells by DNA-mediated cell transfection techniques. The mouse rDNA genes were accurately transcribed in these rat cells indicating that there is no absolute species specificity of rDNA transcription between mouse and rat. No more than 170 nucleotides of the 5' nontranscribed spacer was required for the accurate initiation of mouse rDNA transcription in rat cells. Further, the mouse transcripts were accurately cleaved at the 5' end of the 18S rRNA sequence, even though these transcripts contained neither the 3' end of mouse 18S rRNA nor any other downstream mouse sequences. Thus, cleavage at the 5' end of 18S rRNA is not dependent on long range interactions involving these downstream sequences. Images PMID:2997749

  16. Accurate taxonomic assignment of short pyrosequencing reads.

    PubMed

    Clemente, José C; Jansson, Jesper; Valiente, Gabriel

    2010-01-01

    Ambiguities in the taxonomy dependent assignment of pyrosequencing reads are usually resolved by mapping each read to the lowest common ancestor in a reference taxonomy of all those sequences that match the read. This conservative approach has the drawback of mapping a read to a possibly large clade that may also contain many sequences not matching the read. A more accurate taxonomic assignment of short reads can be made by mapping each read to the node in the reference taxonomy that provides the best precision and recall. We show that given a suffix array for the sequences in the reference taxonomy, a short read can be mapped to the node of the reference taxonomy with the best combined value of precision and recall in time linear in the size of the taxonomy subtree rooted at the lowest common ancestor of the matching sequences. An accurate taxonomic assignment of short reads can thus be made with about the same efficiency as when mapping each read to the lowest common ancestor of all matching sequences in a reference taxonomy. We demonstrate the effectiveness of our approach on several metagenomic datasets of marine and gut microbiota.

  17. Accurate shear measurement with faint sources

    SciTech Connect

    Zhang, Jun; Foucaud, Sebastien; Luo, Wentao E-mail: walt@shao.ac.cn

    2015-01-01

    For cosmic shear to become an accurate cosmological probe, systematic errors in the shear measurement method must be unambiguously identified and corrected for. Previous work of this series has demonstrated that cosmic shears can be measured accurately in Fourier space in the presence of background noise and finite pixel size, without assumptions on the morphologies of galaxy and PSF. The remaining major source of error is source Poisson noise, due to the finiteness of source photon number. This problem is particularly important for faint galaxies in space-based weak lensing measurements, and for ground-based images of short exposure times. In this work, we propose a simple and rigorous way of removing the shear bias from the source Poisson noise. Our noise treatment can be generalized for images made of multiple exposures through MultiDrizzle. This is demonstrated with the SDSS and COSMOS/ACS data. With a large ensemble of mock galaxy images of unrestricted morphologies, we show that our shear measurement method can achieve sub-percent level accuracy even for images of signal-to-noise ratio less than 5 in general, making it the most promising technique for cosmic shear measurement in the ongoing and upcoming large scale galaxy surveys.

  18. Accurate pose estimation for forensic identification

    NASA Astrophysics Data System (ADS)

    Merckx, Gert; Hermans, Jeroen; Vandermeulen, Dirk

    2010-04-01

    In forensic authentication, one aims to identify the perpetrator among a series of suspects or distractors. A fundamental problem in any recognition system that aims for identification of subjects in a natural scene is the lack of constrains on viewing and imaging conditions. In forensic applications, identification proves even more challenging, since most surveillance footage is of abysmal quality. In this context, robust methods for pose estimation are paramount. In this paper we will therefore present a new pose estimation strategy for very low quality footage. Our approach uses 3D-2D registration of a textured 3D face model with the surveillance image to obtain accurate far field pose alignment. Starting from an inaccurate initial estimate, the technique uses novel similarity measures based on the monogenic signal to guide a pose optimization process. We will illustrate the descriptive strength of the introduced similarity measures by using them directly as a recognition metric. Through validation, using both real and synthetic surveillance footage, our pose estimation method is shown to be accurate, and robust to lighting changes and image degradation.

  19. Sparse and accurate high resolution SAR imaging

    NASA Astrophysics Data System (ADS)

    Vu, Duc; Zhao, Kexin; Rowe, William; Li, Jian

    2012-05-01

    We investigate the usage of an adaptive method, the Iterative Adaptive Approach (IAA), in combination with a maximum a posteriori (MAP) estimate to reconstruct high resolution SAR images that are both sparse and accurate. IAA is a nonparametric weighted least squares algorithm that is robust and user parameter-free. IAA has been shown to reconstruct SAR images with excellent side lobes suppression and high resolution enhancement. We first reconstruct the SAR images using IAA, and then we enforce sparsity by using MAP with a sparsity inducing prior. By coupling these two methods, we can produce a sparse and accurate high resolution image that are conducive for feature extractions and target classification applications. In addition, we show how IAA can be made computationally efficient without sacrificing accuracies, a desirable property for SAR applications where the size of the problems is quite large. We demonstrate the success of our approach using the Air Force Research Lab's "Gotcha Volumetric SAR Data Set Version 1.0" challenge dataset. Via the widely used FFT, individual vehicles contained in the scene are barely recognizable due to the poor resolution and high side lobe nature of FFT. However with our approach clear edges, boundaries, and textures of the vehicles are obtained.

  20. Accurate basis set truncation for wavefunction embedding

    NASA Astrophysics Data System (ADS)

    Barnes, Taylor A.; Goodpaster, Jason D.; Manby, Frederick R.; Miller, Thomas F.

    2013-07-01

    Density functional theory (DFT) provides a formally exact framework for performing embedded subsystem electronic structure calculations, including DFT-in-DFT and wavefunction theory-in-DFT descriptions. In the interest of efficiency, it is desirable to truncate the atomic orbital basis set in which the subsystem calculation is performed, thus avoiding high-order scaling with respect to the size of the MO virtual space. In this study, we extend a recently introduced projection-based embedding method [F. R. Manby, M. Stella, J. D. Goodpaster, and T. F. Miller III, J. Chem. Theory Comput. 8, 2564 (2012)], 10.1021/ct300544e to allow for the systematic and accurate truncation of the embedded subsystem basis set. The approach is applied to both covalently and non-covalently bound test cases, including water clusters and polypeptide chains, and it is demonstrated that errors associated with basis set truncation are controllable to well within chemical accuracy. Furthermore, we show that this approach allows for switching between accurate projection-based embedding and DFT embedding with approximate kinetic energy (KE) functionals; in this sense, the approach provides a means of systematically improving upon the use of approximate KE functionals in DFT embedding.

  1. Statokinesigram normalization method.

    PubMed

    de Oliveira, José Magalhães

    2017-02-01

    Stabilometry is a technique that aims to study the body sway of human subjects, employing a force platform. The signal obtained from this technique refers to the position of the foot base ground-reaction vector, known as the center of pressure (CoP). The parameters calculated from the signal are used to quantify the displacement of the CoP over time; there is a large variability, both between and within subjects, which prevents the definition of normative values. The intersubject variability is related to differences between subjects in terms of their anthropometry, in conjunction with their muscle activation patterns (biomechanics); and the intrasubject variability can be caused by a learning effect or fatigue. Age and foot placement on the platform are also known to influence variability. Normalization is the main method used to decrease this variability and to bring distributions of adjusted values into alignment. In 1996, O'Malley proposed three normalization techniques to eliminate the effect of age and anthropometric factors from temporal-distance parameters of gait. These techniques were adopted to normalize the stabilometric signal by some authors. This paper proposes a new method of normalization of stabilometric signals to be applied in balance studies. The method was applied to a data set collected in a previous study, and the results of normalized and nonnormalized signals were compared. The results showed that the new method, if used in a well-designed experiment, can eliminate undesirable correlations between the analyzed parameters and the subjects' characteristics and show only the experimental conditions' effects.

  2. Transcriptional Landscape of Glomerular Parietal Epithelial Cells

    PubMed Central

    Gharib, Sina A.; Pippin, Jeffrey W.; Ohse, Takamoto; Pickering, Scott G.; Krofft, Ronald D.; Shankland, Stuart J.

    2014-01-01

    Very little is known about the function of glomerular parietal epithelial cells (PECs). In this study, we performed genome-wide expression analysis on PEC-enriched capsulated vs. PEC-deprived decapsulated rat glomeruli to determine the transcriptional state of PECs under normal conditions. We identified hundreds of differentially expressed genes that mapped to distinct biologic modules including development, tight junction, ion transport, and metabolic processes. Since developmental programs were highly enriched in PECs, we characterized several of their candidate members at the protein level. Collectively, our findings confirm that PECs are multifaceted cells and help define their diverse functional repertoire. PMID:25127402

  3. Accurate sperm morphology assessment predicts sperm function.

    PubMed

    Abu Hassan Abu, D; Franken, D R; Hoffman, B; Henkel, R

    2012-05-01

    Sperm morphology has been associated with in vitro as well as in vivo fertilisation. The study aimed to evaluate the possible relation between the percentage of spermatozoa with normal morphology and the following sperm functional assays: (i) zona-induced acrosome reaction (ZIAR); (ii) DNA integrity; (iii) chromatin condensation; (iv) sperm apoptosis; and (v) fertilisation rates. Regression analysis was employed to calculate the association between morphology and different functional tests. Normal sperm morphology correlated significantly with the percentages of live acrosome-reacted spermatozoa in the ZIAR (r = 0.518; P < 0.0001; n = 92), DNA integrity (r = -0.515; P = 0.0018; n = 34), CMA(3) -positive spermatozoa (r = -0.745; P < 0.0001; n = 92), sperm apoptosis (r = -0.395; P = 0.0206; n = 34) and necrosis (r = -0.545; P = 0.0009; n = 34). Negative correlations existed between for the acrosome reaction, and DNA integrity, while negative associations were recorded with the percentages of CMA(3) -positive spermatozoa, apoptotic and necrotic spermatozoa. Sperm morphology is related to sperm dysfunction such as poor chromatin condensation, acrosome reaction and DNA integrity. Negative and significant correlations existed between normal sperm morphology and chromatin condensation, the percentage of spermatozoa with abnormal DNA and spermatozoa with apoptotic activity. The authors do not regard sperm morphology as the only test for the diagnosis of male fertility, but sperm morphology can serve as a valuable indicator of underlying dysfunction.

  4. Transcriptional regulation of the FSH receptor: new perspectives

    PubMed Central

    Hermann, Brian P.; Heckert, Leslie L.

    2013-01-01

    The cell-surface receptor for the gonadotropin follicle-stimulating hormone (FSH) is expressed exclusively on Sertoli cells of the testis and granulosa cells of the ovary. FSH signal transduction through its receptor (Fshr) is critical for the timing and maintenance of normal gametogenesis in the mammalian gonad. In the 13 years since the gene encoding Fshr was first cloned, the mechanisms controlling its transcription have been extensively examined, but a clear understanding of what drives its unique cell-specificity remains elusive. Current knowledge of basal Fshr transcription highlights the role of an E-box in the proximal promoter which is bound by the basic helix-loop-helix transcription factors upstream stimulatory factor 1 (Usf1) and Usf2. Recent studies utilizing knockout mice and chromatin immunoprecipitation validated the importance of Usf to Fshr transcription and demonstrated a sexually dimorphic requirement for the Usf proteins to maintain normal Fshr expression. Studies have also shown that the promoter region itself is insufficient for appropriate Fshr expression in transgenic mice, indicating Fshr transcription depends on regulatory elements that lie outside of the promoter. Identification of such elements has been propelled by recent availability of genome sequence data, which facilitated studies using comparative genomics, DNase I hypersensitivity mapping, and transgenic analysis with large fragments of DNA. This review will focus on the current understanding of transcriptional regulatory processes that control expression of rat Fshr, including recent advances from our laboratory. PMID:17084019

  5. Selection of Reference Genes for Gene Expression Normalization in Peucedanum praeruptorum Dunn under Abiotic Stresses, Hormone Treatments and Different Tissues.

    PubMed

    Zhao, Yucheng; Luo, Jun; Xu, Sheng; Wang, Wei; Liu, Tingting; Han, Chao; Chen, Yijun; Kong, Lingyi

    2016-01-01

    Peucedanum praeruptorum Dunn is one of the main traditional Chinese medicines producing coumarins and plenty of literatures are focused on the biosynthesis of coumarins. Quantitative real-time reverse transcription PCR (qRT-PCR) is a widely used method in studying the biosynthesis pathway and the selection of reference genes plays a crucial role in accurate normalization. To facilitate biosynthesis study of coumarins, twelve candidate reference genes were selected from the transcriptome database of P. praeruptorum according to previous studies. Then, BestKeeper, geNoFrm and NormFinder were used for selecting stably expressed reference genes in different tissues and under various stress treatments. The results indicated that, among the twelve candidate reference genes, the SAND family protein (SAND), actin 2 (ACT2), ubiquitin-conjugating enzyme 9 (UBC9), protein phosphatase 2A gene (PP2A) and polypyrimidine tract-binding protein (PTBP1) were the most stable reference genes under different experimental treatments, while glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and tubulin beta-6 (TUB6) were the least stable genes. In addition, the suitability of SAND, TIP41-like protein (TIP41), UBC9, ACT2, TUB6 and their combination as reference genes were confirmed by normalizing the expression of 1-aminocyclopropane-1-carboxylate oxidase (ACO) in different treatments. This work is the first survey of the stability of reference genes in P. praeruptorum and provides guidelines to obtain more accurate qRT-PCR results in P. praeruptorum and other plant species.

  6. Selection of Reference Genes for Gene Expression Normalization in Peucedanum praeruptorum Dunn under Abiotic Stresses, Hormone Treatments and Different Tissues

    PubMed Central

    Zhao, Yucheng; Luo, Jun; Xu, Sheng; Wang, Wei; Liu, Tingting; Han, Chao; Chen, Yijun; Kong, Lingyi

    2016-01-01

    Peucedanum praeruptorum Dunn is one of the main traditional Chinese medicines producing coumarins and plenty of literatures are focused on the biosynthesis of coumarins. Quantitative real-time reverse transcription PCR (qRT-PCR) is a widely used method in studying the biosynthesis pathway and the selection of reference genes plays a crucial role in accurate normalization. To facilitate biosynthesis study of coumarins, twelve candidate reference genes were selected from the transcriptome database of P. praeruptorum according to previous studies. Then, BestKeeper, geNoFrm and NormFinder were used for selecting stably expressed reference genes in different tissues and under various stress treatments. The results indicated that, among the twelve candidate reference genes, the SAND family protein (SAND), actin 2 (ACT2), ubiquitin-conjugating enzyme 9 (UBC9), protein phosphatase 2A gene (PP2A) and polypyrimidine tract-binding protein (PTBP1) were the most stable reference genes under different experimental treatments, while glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and tubulin beta-6 (TUB6) were the least stable genes. In addition, the suitability of SAND, TIP41-like protein (TIP41), UBC9, ACT2, TUB6 and their combination as reference genes were confirmed by normalizing the expression of 1-aminocyclopropane-1-carboxylate oxidase (ACO) in different treatments. This work is the first survey of the stability of reference genes in P. praeruptorum and provides guidelines to obtain more accurate qRT-PCR results in P. praeruptorum and other plant species. PMID:27022972

  7. Babbling Development of Hearing-Impaired and Normally Hearing Subjects.

    ERIC Educational Resources Information Center

    Stoel-Gammon, Carol; Otomo, Kiyoshi

    1986-01-01

    Phonetic transcriptions of babbling samples from 11 normally hearing subjects, age 4-18 months, were compared with samples for 11 hearing-impaired students, age 4-28 months. Findings suggested both qualitative and quantitative differences in the babbling of the two groups. (Author/CL)

  8. A Complete Set of Nascent Transcription Rates for Yeast Genes

    PubMed Central

    Pelechano, Vicent; Chávez, Sebastián; Pérez-Ortín, José E.

    2010-01-01

    The amount of mRNA in a cell is the result of two opposite reactions: transcription and mRNA degradation. These reactions are governed by kinetics laws, and the most regulated step for many genes is the transcription rate. The transcription rate, which is assumed to be exercised mainly at the RNA polymerase recruitment level, can be calculated using the RNA polymerase densities determined either by run-on or immunoprecipitation using specific antibodies. The yeast Saccharomyces cerevisiae is the ideal model organism to generate a complete set of nascent transcription rates that will prove useful for many gene regulation studies. By combining genomic data from both the GRO (Genomic Run-on) and the RNA pol ChIP-on-chip methods we generated a new, more accurate nascent transcription rate dataset. By comparing this dataset with the indirect ones obtained from the mRNA stabilities and mRNA amount datasets, we are able to obtain biological information about posttranscriptional regulation processes and a genomic snapshot of the location of the active transcriptional machinery. We have obtained nascent transcription rates for 4,670 yeast genes. The median RNA polymerase II density in the genes is 0.078 molecules/kb, which corresponds to an average of 0.096 molecules/gene. Most genes have transcription rates of between 2 and 30 mRNAs/hour and less than 1% of yeast genes have >1 RNA polymerase molecule/gene. Histone and ribosomal protein genes are the highest transcribed groups of genes and other than these exceptions the transcription of genes is an infrequent phenomenon in a yeast cell. PMID:21103382

  9. Apparatus for accurately measuring high temperatures

    DOEpatents

    Smith, D.D.

    The present invention is a thermometer used for measuring furnace temperatures in the range of about 1800/sup 0/ to 2700/sup 0/C. The thermometer comprises a broadband multicolor thermal radiation sensor positioned to be in optical alignment with the end of a blackbody sight tube extending into the furnace. A valve-shutter arrangement is positioned between the radiation sensor and the sight tube and a chamber for containing a charge of high pressure gas is positioned between the valve-shutter arrangement and the radiation sensor. A momentary opening of the valve shutter arrangement allows a pulse of the high gas to purge the sight tube of air-borne thermal radiation contaminants which permits the radiation sensor to accurately measure the thermal radiation emanating from the end of the sight tube.

  10. Apparatus for accurately measuring high temperatures

    DOEpatents

    Smith, Douglas D.

    1985-01-01

    The present invention is a thermometer used for measuring furnace temperaes in the range of about 1800.degree. to 2700.degree. C. The thermometer comprises a broadband multicolor thermal radiation sensor positioned to be in optical alignment with the end of a blackbody sight tube extending into the furnace. A valve-shutter arrangement is positioned between the radiation sensor and the sight tube and a chamber for containing a charge of high pressure gas is positioned between the valve-shutter arrangement and the radiation sensor. A momentary opening of the valve shutter arrangement allows a pulse of the high gas to purge the sight tube of air-borne thermal radiation contaminants which permits the radiation sensor to accurately measure the thermal radiation emanating from the end of the sight tube.

  11. LSM: perceptually accurate line segment merging

    NASA Astrophysics Data System (ADS)

    Hamid, Naila; Khan, Nazar

    2016-11-01

    Existing line segment detectors tend to break up perceptually distinct line segments into multiple segments. We propose an algorithm for merging such broken segments to recover the original perceptually accurate line segments. The algorithm proceeds by grouping line segments on the basis of angular and spatial proximity. Then those line segment pairs within each group that satisfy unique, adaptive mergeability criteria are successively merged to form a single line segment. This process is repeated until no more line segments can be merged. We also propose a method for quantitative comparison of line segment detection algorithms. Results on the York Urban dataset show that our merged line segments are closer to human-marked ground-truth line segments compared to state-of-the-art line segment detection algorithms.

  12. Highly accurate articulated coordinate measuring machine

    DOEpatents

    Bieg, Lothar F.; Jokiel, Jr., Bernhard; Ensz, Mark T.; Watson, Robert D.

    2003-12-30

    Disclosed is a highly accurate articulated coordinate measuring machine, comprising a revolute joint, comprising a circular encoder wheel, having an axis of rotation; a plurality of marks disposed around at least a portion of the circumference of the encoder wheel; bearing means for supporting the encoder wheel, while permitting free rotation of the encoder wheel about the wheel's axis of rotation; and a sensor, rigidly attached to the bearing means, for detecting the motion of at least some of the marks as the encoder wheel rotates; a probe arm, having a proximal end rigidly attached to the encoder wheel, and having a distal end with a probe tip attached thereto; and coordinate processing means, operatively connected to the sensor, for converting the output of the sensor into a set of cylindrical coordinates representing the position of the probe tip relative to a reference cylindrical coordinate system.

  13. Practical aspects of spatially high accurate methods

    NASA Technical Reports Server (NTRS)

    Godfrey, Andrew G.; Mitchell, Curtis R.; Walters, Robert W.

    1992-01-01

    The computational qualities of high order spatially accurate methods for the finite volume solution of the Euler equations are presented. Two dimensional essentially non-oscillatory (ENO), k-exact, and 'dimension by dimension' ENO reconstruction operators are discussed and compared in terms of reconstruction and solution accuracy, computational cost and oscillatory behavior in supersonic flows with shocks. Inherent steady state convergence difficulties are demonstrated for adaptive stencil algorithms. An exact solution to the heat equation is used to determine reconstruction error, and the computational intensity is reflected in operation counts. Standard MUSCL differencing is included for comparison. Numerical experiments presented include the Ringleb flow for numerical accuracy and a shock reflection problem. A vortex-shock interaction demonstrates the ability of the ENO scheme to excel in simulating unsteady high-frequency flow physics.

  14. Toward Accurate and Quantitative Comparative Metagenomics

    PubMed Central

    Nayfach, Stephen; Pollard, Katherine S.

    2016-01-01

    Shotgun metagenomics and computational analysis are used to compare the taxonomic and functional profiles of microbial communities. Leveraging this approach to understand roles of microbes in human biology and other environments requires quantitative data summaries whose values are comparable across samples and studies. Comparability is currently hampered by the use of abundance statistics that do not estimate a meaningful parameter of the microbial community and biases introduced by experimental protocols and data-cleaning approaches. Addressing these challenges, along with improving study design, data access, metadata standardization, and analysis tools, will enable accurate comparative metagenomics. We envision a future in which microbiome studies are replicable and new metagenomes are easily and rapidly integrated with existing data. Only then can the potential of metagenomics for predictive ecological modeling, well-powered association studies, and effective microbiome medicine be fully realized. PMID:27565341

  15. Micron Accurate Absolute Ranging System: Range Extension

    NASA Technical Reports Server (NTRS)

    Smalley, Larry L.; Smith, Kely L.

    1999-01-01

    The purpose of this research is to investigate Fresnel diffraction as a means of obtaining absolute distance measurements with micron or greater accuracy. It is believed that such a system would prove useful to the Next Generation Space Telescope (NGST) as a non-intrusive, non-contact measuring system for use with secondary concentrator station-keeping systems. The present research attempts to validate past experiments and develop ways to apply the phenomena of Fresnel diffraction to micron accurate measurement. This report discusses past research on the phenomena, and the basis of the use Fresnel diffraction distance metrology. The apparatus used in the recent investigations, experimental procedures used, preliminary results are discussed in detail. Continued research and equipment requirements on the extension of the effective range of the Fresnel diffraction systems is also described.

  16. Accurate radio positions with the Tidbinbilla interferometer

    NASA Technical Reports Server (NTRS)

    Batty, M. J.; Gulkis, S.; Jauncey, D. L.; Rayner, P. T.

    1979-01-01

    The Tidbinbilla interferometer (Batty et al., 1977) is designed specifically to provide accurate radio position measurements of compact radio sources in the Southern Hemisphere with high sensitivity. The interferometer uses the 26-m and 64-m antennas of the Deep Space Network at Tidbinbilla, near Canberra. The two antennas are separated by 200 m on a north-south baseline. By utilizing the existing antennas and the low-noise traveling-wave masers at 2.29 GHz, it has been possible to produce a high-sensitivity instrument with a minimum of capital expenditure. The north-south baseline ensures that a good range of UV coverage is obtained, so that sources lying in the declination range between about -80 and +30 deg may be observed with nearly orthogonal projected baselines of no less than about 1000 lambda. The instrument also provides high-accuracy flux density measurements for compact radio sources.

  17. Magnetic ranging tool accurately guides replacement well

    SciTech Connect

    Lane, J.B.; Wesson, J.P. )

    1992-12-21

    This paper reports on magnetic ranging surveys and directional drilling technology which accurately guided a replacement well bore to intersect a leaking gas storage well with casing damage. The second well bore was then used to pump cement into the original leaking casing shoe. The repair well bore kicked off from the surface hole, bypassed casing damage in the middle of the well, and intersected the damaged well near the casing shoe. The repair well was subsequently completed in the gas storage zone near the original well bore, salvaging the valuable bottom hole location in the reservoir. This method would prevent the loss of storage gas, and it would prevent a potential underground blowout that could permanently damage the integrity of the storage field.

  18. Idiopathic Normal Pressure Hydrocephalus

    PubMed Central

    Nassar, Basant R.; Lippa, Carol F.

    2016-01-01

    Idiopathic normal pressure hydrocephalus (iNPH) is a potentially reversible neurodegenerative disease commonly characterized by a triad of dementia, gait, and urinary disturbance. Advancements in diagnosis and treatment have aided in properly identifying and improving symptoms in patients. However, a large proportion of iNPH patients remain either undiagnosed or misdiagnosed. Using PubMed search engine of keywords “normal pressure hydrocephalus,” “diagnosis,” “shunt treatment,” “biomarkers,” “gait disturbances,” “cognitive function,” “neuropsychology,” “imaging,” and “pathogenesis,” articles were obtained for this review. The majority of the articles were retrieved from the past 10 years. The purpose of this review article is to aid general practitioners in further understanding current findings on the pathogenesis, diagnosis, and treatment of iNPH. PMID:28138494

  19. Studies of normal hearing.

    PubMed

    Catlin, F I

    1984-01-01

    Auditory function changes continually from birth to old age. A variety of methods to assess hearing have evolved since the invention of the audiometer. Types of measurement include: electrical response in the central nervous system, cochlear acuity and speech responses. While some of these tests correlate fairly well with each other, their ability to represent overall hearing function is questionable. Other attempts to improve the assessment of hearing have been made in the area of self-appraisal, but these, too, have significant limitations. Most self-report and peer appraisal questionnaires have been established by studies of hearing-impaired populations. Norms for these techniques in normal-hearing populations need to be established. There is still room for valid tests of everyday communication. What we have in measurement procedures does not achieve this goal. Research studies of today will hopefully produce better definition of normal auditory function.

  20. The high cost of accurate knowledge.

    PubMed

    Sutcliffe, Kathleen M; Weber, Klaus

    2003-05-01

    Many business thinkers believe it's the role of senior managers to scan the external environment to monitor contingencies and constraints, and to use that precise knowledge to modify the company's strategy and design. As these thinkers see it, managers need accurate and abundant information to carry out that role. According to that logic, it makes sense to invest heavily in systems for collecting and organizing competitive information. Another school of pundits contends that, since today's complex information often isn't precise anyway, it's not worth going overboard with such investments. In other words, it's not the accuracy and abundance of information that should matter most to top executives--rather, it's how that information is interpreted. After all, the role of senior managers isn't just to make decisions; it's to set direction and motivate others in the face of ambiguities and conflicting demands. Top executives must interpret information and communicate those interpretations--they must manage meaning more than they must manage information. So which of these competing views is the right one? Research conducted by academics Sutcliffe and Weber found that how accurate senior executives are about their competitive environments is indeed less important for strategy and corresponding organizational changes than the way in which they interpret information about their environments. Investments in shaping those interpretations, therefore, may create a more durable competitive advantage than investments in obtaining and organizing more information. And what kinds of interpretations are most closely linked with high performance? Their research suggests that high performers respond positively to opportunities, yet they aren't overconfident in their abilities to take advantage of those opportunities.

  1. Neuroethics beyond Normal.

    PubMed

    Shook, John R; Giordano, James

    2016-01-01

    An integrated and principled neuroethics offers ethical guidelines able to transcend conventional and medical reliance on normality standards. Elsewhere we have proposed four principles for wise guidance on human transformations. Principles like these are already urgently needed, as bio- and cyberenhancements are rapidly emerging. Context matters. Neither "treatments" nor "enhancements" are objectively identifiable apart from performance expectations, social contexts, and civic orders. Lessons learned from disability studies about enablement and inclusion suggest a fresh way to categorize modifications to the body and its performance. The term "enhancement" should be broken apart to permit recognition of enablements and augmentations, and kinds of radical augmentation for specialized performance. Augmentations affecting the self, self-worth, and self-identity of persons require heightened ethical scrutiny. Reversibility becomes the core problem, not the easy answer, as augmented persons may not cooperate with either decommissioning or displacement into unaccommodating societies. We conclude by indicating how our four principles of self-creativity, nonobsolescence, empowerment, and citizenship establish a neuroethics beyond normal that is better prepared for a future in which humans and their societies are going so far beyond normal.

  2. Accurate detection of differential RNA processing

    PubMed Central

    Drewe, Philipp; Stegle, Oliver; Hartmann, Lisa; Kahles, André; Bohnert, Regina; Wachter, Andreas; Borgwardt, Karsten; Rätsch, Gunnar

    2013-01-01

    Deep transcriptome sequencing (RNA-Seq) has become a vital tool for studying the state of cells in the context of varying environments, genotypes and other factors. RNA-Seq profiling data enable identification of novel isoforms, quantification of known isoforms and detection of changes in transcriptional or RNA-processing activity. Existing approaches to detect differential isoform abundance between samples either require a complete isoform annotation or fall short in providing statistically robust and calibrated significance estimates. Here, we propose a suite of statistical tests to address these open needs: a parametric test that uses known isoform annotations to detect changes in relative isoform abundance and a non-parametric test that detects differential read coverages and can be applied when isoform annotations are not available. Both methods account for the discrete nature of read counts and the inherent biological variability. We demonstrate that these tests compare favorably to previous methods, both in terms of accuracy and statistical calibrations. We use these techniques to analyze RNA-Seq libraries from Arabidopsis thaliana and Drosophila melanogaster. The identified differential RNA processing events were consistent with RT–qPCR measurements and previous studies. The proposed toolkit is available from http://bioweb.me/rdiff and enables in-depth analyses of transcriptomes, with or without available isoform annotation. PMID:23585274

  3. A non-proteolytic function of ubiquitin in transcription repression

    PubMed Central

    Ndoja, Ada; Yao, Tingting

    2014-01-01

    Regulation of transcription is vitally important for maintaining normal cellular homeostasis and is also the basis for cellular differentiation, morphogenesis and the adaptability of any organism. Transcription activators, which orchestrate time and locus-specific assembly of complex transcription machinery, act as key players in these processes. One way in which these activators are controlled is by the covalent attachment of the conserved protein, ubiquitin (Ub), which can serve as either a proteolytic or non-proteolytic signal. For a subset of the activators, polyubiquitination-dependent degradation of the activator controls its abundance. In these cases transcription activation can require protein synthesis as well as internal or external stimulus. In contrast, other activators have been reported to undergo mono- or oligoubiquitination that does not lead to protein degradation. The mechanisms by which monoubiquitination of transcription activators affect their activities have been poorly understood. In a recent study, we demonstrated that monoubiquitination of some transcription activators can inhibit transcription by recruiting the AAA+ ATPase Cdc48 (also known in metazoan organisms as p97 or valosin-contain protein, VCP), which then extracts the ubiquitinated activator from DNA. PMID:28357251

  4. Approaching system equilibrium with accurate or not accurate feedback information in a two-route system

    NASA Astrophysics Data System (ADS)

    Zhao, Xiao-mei; Xie, Dong-fan; Li, Qi

    2015-02-01

    With the development of intelligent transport system, advanced information feedback strategies have been developed to reduce traffic congestion and enhance the capacity. However, previous strategies provide accurate information to travelers and our simulation results show that accurate information brings negative effects, especially in delay case. Because travelers prefer to the best condition route with accurate information, and delayed information cannot reflect current traffic condition but past. Then travelers make wrong routing decisions, causing the decrease of the capacity and the increase of oscillations and the system deviating from the equilibrium. To avoid the negative effect, bounded rationality is taken into account by introducing a boundedly rational threshold BR. When difference between two routes is less than the BR, routes have equal probability to be chosen. The bounded rationality is helpful to improve the efficiency in terms of capacity, oscillation and the gap deviating from the system equilibrium.

  5. Post-transcriptional regulation of ornithine decarboxylase

    PubMed Central

    Nowotarski, Shannon L.; Origanti, Sofia; Shantz, Lisa M.

    2013-01-01

    Activity of the polyamine biosynthetic enzyme ornithine decarboxylase (ODC), and intracellular levels of ODC protein are controlled very tightly. Numerous studies have described ODC regulation at the levels of transcription, translation and protein degradation in normal cells, and dysregulation of these processes in response to oncogenic stimuli. Although post-transcriptional regulation of ODC has been well-documented, the RNA binding proteins (RBPs) that interact with ODC mRNA and control synthesis of the ODC protein have not been defined. Using Ras-transformed rat intestinal epithelial cells (Ras12V cells) as a model, we have begun identifying the RBPs that associate with the ODC transcript. Binding of RBPs could potentially regulate ODC synthesis by either changing mRNA stability or rate of mRNA translation. Techniques for measuring RBP binding and translation initiation are described here. Targeting control of ODC translation or mRNA decay could be a valuable method of limiting polyamine accumulation and subsequent tumor development in a variety of cancers. PMID:21318880

  6. Higher order accurate partial implicitization: An unconditionally stable fourth-order-accurate explicit numerical technique

    NASA Technical Reports Server (NTRS)

    Graves, R. A., Jr.

    1975-01-01

    The previously obtained second-order-accurate partial implicitization numerical technique used in the solution of fluid dynamic problems was modified with little complication to achieve fourth-order accuracy. The Von Neumann stability analysis demonstrated the unconditional linear stability of the technique. The order of the truncation error was deduced from the Taylor series expansions of the linearized difference equations and was verified by numerical solutions to Burger's equation. For comparison, results were also obtained for Burger's equation using a second-order-accurate partial-implicitization scheme, as well as the fourth-order scheme of Kreiss.

  7. The Werner Syndrome Protein Is Involved in RNA Polymerase II Transcription

    PubMed Central

    Balajee, Adayabalam S.; Machwe, Amrita; May, Alfred; Gray, Matthew D.; Oshima, Junko; Martin, George M.; Nehlin, Jan O.; Brosh, Robert; Orren, David K.; Bohr, Vilhelm A.

    1999-01-01

    Werner syndrome (WS) is a human progeroid syndrome characterized by the early onset of a large number of clinical features associated with the normal aging process. The complex molecular and cellular phenotypes of WS involve characteristic features of genomic instability and accelerated replicative senescence. The gene involved (WRN) was recently cloned, and its gene product (WRNp) was biochemically characterized as a helicase. Helicases play important roles in a variety of DNA transactions, including DNA replication, transcription, repair, and recombination. We have assessed the role of the WRN gene in transcription by analyzing the efficiency of basal transcription in WS lymphoblastoid cell lines that carry homozygous WRN mutations. Transcription was measured in permeabilized cells by [3H]UTP incorporation and in vitro by using a plasmid template containing the RNA polymerase II (RNA pol II)–dependent adenovirus major late promoter. With both of these approaches, we find that the transcription efficiency in different WS cell lines is reduced to 40–60% of the transcription in cells from normal individuals. This defect can be complemented by the addition of normal cell extracts to the chromatin of WS cells. Addition of purified wild-type WRNp but not mutated WRNp to the in vitro transcription assay markedly stimulates RNA pol II–dependent transcription carried out by nuclear extracts. A nonhelicase domain (a direct repeat of 27 amino acids) also appears to have a role in transcription enhancement, as revealed by a yeast hybrid–protein reporter assay. This is further supported by the lack of stimulation of transcription when mutant WRNp lacking this domain was added to the in vitro assay. We have thus used several approaches to show a role for WRNp in RNA pol II transcription, possibly as a transcriptional activator. A deficit in either global or regional transcription in WS cells may be a primary molecular defect responsible for the WS clinical phenotype

  8. Transcriptional and post-transcriptional regulation of the ionizing radiation response by ATM and p53

    PubMed Central

    Venkata Narayanan, Ishwarya; Paulsen, Michelle T.; Bedi, Karan; Berg, Nathan; Ljungman, Emily A.; Francia, Sofia; Veloso, Artur; Magnuson, Brian; di Fagagna, Fabrizio d’Adda; Wilson, Thomas E.; Ljungman, Mats

    2017-01-01

    In response to ionizing radiation (IR), cells activate a DNA damage response (DDR) pathway to re-program gene expression. Previous studies using total cellular RNA analyses have shown that the stress kinase ATM and the transcription factor p53 are integral components required for induction of IR-induced gene expression. These studies did not distinguish between changes in RNA synthesis and RNA turnover and did not address the role of enhancer elements in DDR-mediated transcriptional regulation. To determine the contribution of synthesis and degradation of RNA and monitor the activity of enhancer elements following exposure to IR, we used the recently developed Bru-seq, BruChase-seq and BruUV-seq techniques. Our results show that ATM and p53 regulate both RNA synthesis and stability as well as enhancer element activity following exposure to IR. Importantly, many genes in the p53-signaling pathway were coordinately up-regulated by both increased synthesis and RNA stability while down-regulated genes were suppressed either by reduced synthesis or stability. Our study is the first of its kind that independently assessed the effects of ionizing radiation on transcription and post-transcriptional regulation in normal human cells. PMID:28256581

  9. Drosophila OVO regulates ovarian tumor transcription by binding unusually near the transcription start site.

    PubMed

    Lü, J; Oliver, B

    2001-05-01

    Evolutionarily conserved ovo loci encode developmentally regulated, sequence-specific, DNA-binding, C(2)H(2)-zinc-finger proteins required in the germline and epidermal cells of flies and mice. The direct targets of OVO activity are not known. Genetic experiments suggest that ovo acts in the same regulatory network as ovarian tumor (otu), but the relative position of these genes in the pathway is controversial. Three OVO-binding sites exist in a compact regulatory region that controls germline expression of the otu gene. Interestingly, the strongest OVO-binding site is very near the otu transcription start, where basal transcriptional complexes must function. Loss-of-function, gain-of-function and promoter swapping constructs demonstrate that OVO binding near the transcription start site is required for OVO-dependent otu transcription in vivo. These data unambiguously identify otu as a direct OVO target gene and raise the tantalizing possibility that an OVO site, at the location normally occupied by basal components, functions as part of a specialized core promoter.

  10. Does a pneumotach accurately characterize voice function?

    NASA Astrophysics Data System (ADS)

    Walters, Gage; Krane, Michael

    2016-11-01

    A study is presented which addresses how a pneumotach might adversely affect clinical measurements of voice function. A pneumotach is a device, typically a mask, worn over the mouth, in order to measure time-varying glottal volume flow. By measuring the time-varying difference in pressure across a known aerodynamic resistance element in the mask, the glottal volume flow waveform is estimated. Because it adds aerodynamic resistance to the vocal system, there is some concern that using a pneumotach may not accurately portray the behavior of the voice. To test this hypothesis, experiments were performed in a simplified airway model with the principal dimensions of an adult human upper airway. A compliant constriction, fabricated from silicone rubber, modeled the vocal folds. Variations of transglottal pressure, time-averaged volume flow, model vocal fold vibration amplitude, and radiated sound with subglottal pressure were performed, with and without the pneumotach in place, and differences noted. Acknowledge support of NIH Grant 2R01DC005642-10A1.

  11. Accurate thermoplasmonic simulation of metallic nanoparticles

    NASA Astrophysics Data System (ADS)

    Yu, Da-Miao; Liu, Yan-Nan; Tian, Fa-Lin; Pan, Xiao-Min; Sheng, Xin-Qing

    2017-01-01

    Thermoplasmonics leads to enhanced heat generation due to the localized surface plasmon resonances. The measurement of heat generation is fundamentally a complicated task, which necessitates the development of theoretical simulation techniques. In this paper, an efficient and accurate numerical scheme is proposed for applications with complex metallic nanostructures. Light absorption and temperature increase are, respectively, obtained by solving the volume integral equation (VIE) and the steady-state heat diffusion equation through the method of moments (MoM). Previously, methods based on surface integral equations (SIEs) were utilized to obtain light absorption. However, computing light absorption from the equivalent current is as expensive as O(NsNv), where Ns and Nv, respectively, denote the number of surface and volumetric unknowns. Our approach reduces the cost to O(Nv) by using VIE. The accuracy, efficiency and capability of the proposed scheme are validated by multiple simulations. The simulations show that our proposed method is more efficient than the approach based on SIEs under comparable accuracy, especially for the case where many incidents are of interest. The simulations also indicate that the temperature profile can be tuned by several factors, such as the geometry configuration of array, beam direction, and light wavelength.

  12. Accurate method for computing correlated color temperature.

    PubMed

    Li, Changjun; Cui, Guihua; Melgosa, Manuel; Ruan, Xiukai; Zhang, Yaoju; Ma, Long; Xiao, Kaida; Luo, M Ronnier

    2016-06-27

    For the correlated color temperature (CCT) of a light source to be estimated, a nonlinear optimization problem must be solved. In all previous methods available to compute CCT, the objective function has only been approximated, and their predictions have achieved limited accuracy. For example, different unacceptable CCT values have been predicted for light sources located on the same isotemperature line. In this paper, we propose to compute CCT using the Newton method, which requires the first and second derivatives of the objective function. Following the current recommendation by the International Commission on Illumination (CIE) for the computation of tristimulus values (summations at 1 nm steps from 360 nm to 830 nm), the objective function and its first and second derivatives are explicitly given and used in our computations. Comprehensive tests demonstrate that the proposed method, together with an initial estimation of CCT using Robertson's method [J. Opt. Soc. Am. 58, 1528-1535 (1968)], gives highly accurate predictions below 0.0012 K for light sources with CCTs ranging from 500 K to 106 K.

  13. Accurate Theoretical Thermochemistry for Fluoroethyl Radicals.

    PubMed

    Ganyecz, Ádám; Kállay, Mihály; Csontos, József

    2017-02-09

    An accurate coupled-cluster (CC) based model chemistry was applied to calculate reliable thermochemical quantities for hydrofluorocarbon derivatives including radicals 1-fluoroethyl (CH3-CHF), 1,1-difluoroethyl (CH3-CF2), 2-fluoroethyl (CH2F-CH2), 1,2-difluoroethyl (CH2F-CHF), 2,2-difluoroethyl (CHF2-CH2), 2,2,2-trifluoroethyl (CF3-CH2), 1,2,2,2-tetrafluoroethyl (CF3-CHF), and pentafluoroethyl (CF3-CF2). The model chemistry used contains iterative triple and perturbative quadruple excitations in CC theory, as well as scalar relativistic and diagonal Born-Oppenheimer corrections. To obtain heat of formation values with better than chemical accuracy perturbative quadruple excitations and scalar relativistic corrections were inevitable. Their contributions to the heats of formation steadily increase with the number of fluorine atoms in the radical reaching 10 kJ/mol for CF3-CF2. When discrepancies were found between the experimental and our values it was always possible to resolve the issue by recalculating the experimental result with currently recommended auxiliary data. For each radical studied here this study delivers the best heat of formation as well as entropy data.

  14. Accurate, reliable prototype earth horizon sensor head

    NASA Technical Reports Server (NTRS)

    Schwarz, F.; Cohen, H.

    1973-01-01

    The design and performance is described of an accurate and reliable prototype earth sensor head (ARPESH). The ARPESH employs a detection logic 'locator' concept and horizon sensor mechanization which should lead to high accuracy horizon sensing that is minimally degraded by spatial or temporal variations in sensing attitude from a satellite in orbit around the earth at altitudes in the 500 km environ 1,2. An accuracy of horizon location to within 0.7 km has been predicted, independent of meteorological conditions. This corresponds to an error of 0.015 deg-at 500 km altitude. Laboratory evaluation of the sensor indicates that this accuracy is achieved. First, the basic operating principles of ARPESH are described; next, detailed design and construction data is presented and then performance of the sensor under laboratory conditions in which the sensor is installed in a simulator that permits it to scan over a blackbody source against background representing the earth space interface for various equivalent plant temperatures.

  15. Accurate methods for large molecular systems.

    PubMed

    Gordon, Mark S; Mullin, Jonathan M; Pruitt, Spencer R; Roskop, Luke B; Slipchenko, Lyudmila V; Boatz, Jerry A

    2009-07-23

    Three exciting new methods that address the accurate prediction of processes and properties of large molecular systems are discussed. The systematic fragmentation method (SFM) and the fragment molecular orbital (FMO) method both decompose a large molecular system (e.g., protein, liquid, zeolite) into small subunits (fragments) in very different ways that are designed to both retain the high accuracy of the chosen quantum mechanical level of theory while greatly reducing the demands on computational time and resources. Each of these methods is inherently scalable and is therefore eminently capable of taking advantage of massively parallel computer hardware while retaining the accuracy of the corresponding electronic structure method from which it is derived. The effective fragment potential (EFP) method is a sophisticated approach for the prediction of nonbonded and intermolecular interactions. Therefore, the EFP method provides a way to further reduce the computational effort while retaining accuracy by treating the far-field interactions in place of the full electronic structure method. The performance of the methods is demonstrated using applications to several systems, including benzene dimer, small organic species, pieces of the alpha helix, water, and ionic liquids.

  16. Accurate equilibrium structures for piperidine and cyclohexane.

    PubMed

    Demaison, Jean; Craig, Norman C; Groner, Peter; Écija, Patricia; Cocinero, Emilio J; Lesarri, Alberto; Rudolph, Heinz Dieter

    2015-03-05

    Extended and improved microwave (MW) measurements are reported for the isotopologues of piperidine. New ground state (GS) rotational constants are fitted to MW transitions with quartic centrifugal distortion constants taken from ab initio calculations. Predicate values for the geometric parameters of piperidine and cyclohexane are found from a high level of ab initio theory including adjustments for basis set dependence and for correlation of the core electrons. Equilibrium rotational constants are obtained from GS rotational constants corrected for vibration-rotation interactions and electronic contributions. Equilibrium structures for piperidine and cyclohexane are fitted by the mixed estimation method. In this method, structural parameters are fitted concurrently to predicate parameters (with appropriate uncertainties) and moments of inertia (with uncertainties). The new structures are regarded as being accurate to 0.001 Å and 0.2°. Comparisons are made between bond parameters in equatorial piperidine and cyclohexane. Another interesting result of this study is that a structure determination is an effective way to check the accuracy of the ground state experimental rotational constants.

  17. Accurate lineshape spectroscopy and the Boltzmann constant

    PubMed Central

    Truong, G.-W.; Anstie, J. D.; May, E. F.; Stace, T. M.; Luiten, A. N.

    2015-01-01

    Spectroscopy has an illustrious history delivering serendipitous discoveries and providing a stringent testbed for new physical predictions, including applications from trace materials detection, to understanding the atmospheres of stars and planets, and even constraining cosmological models. Reaching fundamental-noise limits permits optimal extraction of spectroscopic information from an absorption measurement. Here, we demonstrate a quantum-limited spectrometer that delivers high-precision measurements of the absorption lineshape. These measurements yield a very accurate measurement of the excited-state (6P1/2) hyperfine splitting in Cs, and reveals a breakdown in the well-known Voigt spectral profile. We develop a theoretical model that accounts for this breakdown, explaining the observations to within the shot-noise limit. Our model enables us to infer the thermal velocity dispersion of the Cs vapour with an uncertainty of 35 p.p.m. within an hour. This allows us to determine a value for Boltzmann's constant with a precision of 6 p.p.m., and an uncertainty of 71 p.p.m. PMID:26465085

  18. Accurate upper body rehabilitation system using kinect.

    PubMed

    Sinha, Sanjana; Bhowmick, Brojeshwar; Chakravarty, Kingshuk; Sinha, Aniruddha; Das, Abhijit

    2016-08-01

    The growing importance of Kinect as a tool for clinical assessment and rehabilitation is due to its portability, low cost and markerless system for human motion capture. However, the accuracy of Kinect in measuring three-dimensional body joint center locations often fails to meet clinical standards of accuracy when compared to marker-based motion capture systems such as Vicon. The length of the body segment connecting any two joints, measured as the distance between three-dimensional Kinect skeleton joint coordinates, has been observed to vary with time. The orientation of the line connecting adjoining Kinect skeletal coordinates has also been seen to differ from the actual orientation of the physical body segment. Hence we have proposed an optimization method that utilizes Kinect Depth and RGB information to search for the joint center location that satisfies constraints on body segment length and as well as orientation. An experimental study have been carried out on ten healthy participants performing upper body range of motion exercises. The results report 72% reduction in body segment length variance and 2° improvement in Range of Motion (ROM) angle hence enabling to more accurate measurements for upper limb exercises.

  19. Noninvasive hemoglobin monitoring: how accurate is enough?

    PubMed

    Rice, Mark J; Gravenstein, Nikolaus; Morey, Timothy E

    2013-10-01

    Evaluating the accuracy of medical devices has traditionally been a blend of statistical analyses, at times without contextualizing the clinical application. There have been a number of recent publications on the accuracy of a continuous noninvasive hemoglobin measurement device, the Masimo Radical-7 Pulse Co-oximeter, focusing on the traditional statistical metrics of bias and precision. In this review, which contains material presented at the Innovations and Applications of Monitoring Perfusion, Oxygenation, and Ventilation (IAMPOV) Symposium at Yale University in 2012, we critically investigated these metrics as applied to the new technology, exploring what is required of a noninvasive hemoglobin monitor and whether the conventional statistics adequately answer our questions about clinical accuracy. We discuss the glucose error grid, well known in the glucose monitoring literature, and describe an analogous version for hemoglobin monitoring. This hemoglobin error grid can be used to evaluate the required clinical accuracy (±g/dL) of a hemoglobin measurement device to provide more conclusive evidence on whether to transfuse an individual patient. The important decision to transfuse a patient usually requires both an accurate hemoglobin measurement and a physiologic reason to elect transfusion. It is our opinion that the published accuracy data of the Masimo Radical-7 is not good enough to make the transfusion decision.

  20. Accurate, reproducible measurement of blood pressure.

    PubMed Central

    Campbell, N R; Chockalingam, A; Fodor, J G; McKay, D W

    1990-01-01

    The diagnosis of mild hypertension and the treatment of hypertension require accurate measurement of blood pressure. Blood pressure readings are altered by various factors that influence the patient, the techniques used and the accuracy of the sphygmomanometer. The variability of readings can be reduced if informed patients prepare in advance by emptying their bladder and bowel, by avoiding over-the-counter vasoactive drugs the day of measurement and by avoiding exposure to cold, caffeine consumption, smoking and physical exertion within half an hour before measurement. The use of standardized techniques to measure blood pressure will help to avoid large systematic errors. Poor technique can account for differences in readings of more than 15 mm Hg and ultimately misdiagnosis. Most of the recommended procedures are simple and, when routinely incorporated into clinical practice, require little additional time. The equipment must be appropriate and in good condition. Physicians should have a suitable selection of cuff sizes readily available; the use of the correct cuff size is essential to minimize systematic errors in blood pressure measurement. Semiannual calibration of aneroid sphygmomanometers and annual inspection of mercury sphygmomanometers and blood pressure cuffs are recommended. We review the methods recommended for measuring blood pressure and discuss the factors known to produce large differences in blood pressure readings. PMID:2192791

  1. Fast and accurate exhaled breath ammonia measurement.

    PubMed

    Solga, Steven F; Mudalel, Matthew L; Spacek, Lisa A; Risby, Terence H

    2014-06-11

    This exhaled breath ammonia method uses a fast and highly sensitive spectroscopic method known as quartz enhanced photoacoustic spectroscopy (QEPAS) that uses a quantum cascade based laser. The monitor is coupled to a sampler that measures mouth pressure and carbon dioxide. The system is temperature controlled and specifically designed to address the reactivity of this compound. The sampler provides immediate feedback to the subject and the technician on the quality of the breath effort. Together with the quick response time of the monitor, this system is capable of accurately measuring exhaled breath ammonia representative of deep lung systemic levels. Because the system is easy to use and produces real time results, it has enabled experiments to identify factors that influence measurements. For example, mouth rinse and oral pH reproducibly and significantly affect results and therefore must be controlled. Temperature and mode of breathing are other examples. As our understanding of these factors evolves, error is reduced, and clinical studies become more meaningful. This system is very reliable and individual measurements are inexpensive. The sampler is relatively inexpensive and quite portable, but the monitor is neither. This limits options for some clinical studies and provides rational for future innovations.

  2. Accurate Fission Data for Nuclear Safety

    NASA Astrophysics Data System (ADS)

    Solders, A.; Gorelov, D.; Jokinen, A.; Kolhinen, V. S.; Lantz, M.; Mattera, A.; Penttilä, H.; Pomp, S.; Rakopoulos, V.; Rinta-Antila, S.

    2014-05-01

    The Accurate fission data for nuclear safety (AlFONS) project aims at high precision measurements of fission yields, using the renewed IGISOL mass separator facility in combination with a new high current light ion cyclotron at the University of Jyväskylä. The 30 MeV proton beam will be used to create fast and thermal neutron spectra for the study of neutron induced fission yields. Thanks to a series of mass separating elements, culminating with the JYFLTRAP Penning trap, it is possible to achieve a mass resolving power in the order of a few hundred thousands. In this paper we present the experimental setup and the design of a neutron converter target for IGISOL. The goal is to have a flexible design. For studies of exotic nuclei far from stability a high neutron flux (1012 neutrons/s) at energies 1 - 30 MeV is desired while for reactor applications neutron spectra that resembles those of thermal and fast nuclear reactors are preferred. It is also desirable to be able to produce (semi-)monoenergetic neutrons for benchmarking and to study the energy dependence of fission yields. The scientific program is extensive and is planed to start in 2013 with a measurement of isomeric yield ratios of proton induced fission in uranium. This will be followed by studies of independent yields of thermal and fast neutron induced fission of various actinides.

  3. Clusterin expression in normal mucosa and colorectal cancer.

    PubMed

    Andersen, Claus Lindbjerg; Schepeler, Troels; Thorsen, Kasper; Birkenkamp-Demtröder, Karin; Mansilla, Francisco; Aaltonen, Lauri A; Laurberg, Søren; Ørntoft, Torben Falck

    2007-06-01

    The gene Clusterin is a target for cancer therapy in clinical trials. The indication for intervention is up-regulated Clusterin expression. Clusterin has been reported to be deregulated in multiple cancer types, including colorectal cancer (CRC). However, for CRC the studies have disagreed on whether Clusterin is up- or down-regulated by neoplastic cells. In the present study we sought to clarify the expression and distribution of Clusterin mRNAs and proteins in normal and neoplastic colorectal tissue through laser microdissection, variant-specific real time RT-PCR, immunohistochemistry, immunofluorescence, Western blotting, and array-based transcriptional profiling. At the transcript level we demonstrated the expression of two novel Clusterin transcripts in addition to the known transcript, and at the protein level we demonstrated two Clusterin isoforms. Our analysis of normal epithelial cells revealed that among these, Clusterin was only expressed by rare neuroendocrine subtype. Furthermore our analysis showed that in the normal mucosa the majority of the observed Clusterin protein originated from the stromal compartment. In tumors we found that Clusterin was de novo synthesized by non-neuroendocrine cancer cells in approximately 25% of cases. Moreover we found that the overall Clusterin level in tumors often appeared to be lower than in normal mucosa due to the stromal compartment often being suppressed in tumors. Although Clusterin in normal neuroendocrine cells showed a basal localization, the localization in cancer cells was often apical and in some cases associated with apical secretion. Collectively our results indicate that Clusterin expression is very complex. We conclude that Clusterin expression is associated with neuroendocrine differentiation in normal epithelia and that the Clusterin observed in neoplastic cells is de novo synthesized. The cases with de novo synthesized Clusterin define a distinct subgroup of CRC that may be of clinical importance as

  4. Assigning numbers to the arrows: Parameterizing a gene regulation network by using accurate expression kinetics

    NASA Astrophysics Data System (ADS)

    Ronen, Michal; Rosenberg, Revital; Shraiman, Boris I.; Alon, Uri

    2002-08-01

    A basic challenge in systems biology is to understand the dynamical behavior of gene regulation networks. Current approaches aim at determining the network structure based on genomic-scale data. However, the network connectivity alone is not sufficient to define its dynamics; one needs to also specify the kinetic parameters for the regulation reactions. Here, we ask whether effective kinetic parameters can be assigned to a transcriptional network based on expression data. We present a combined experimental and theoretical approach based on accurate high temporal-resolution measurement of promoter activities from living cells by using green fluorescent protein (GFP) reporter plasmids. We present algorithms that use these data to assign effective kinetic parameters within a mathematical model of the network. To demonstrate this, we employ a well defined network, the SOS DNA repair system of Escherichia coli. We find a strikingly detailed temporal program of expression that correlates with the functional role of the SOS genes and is driven by a hierarchy of effective kinetic parameter strengths for the various promoters. The calculated parameters can be used to determine the kinetics of all SOS genes given the expression profile of just one representative, allowing a significant reduction in complexity. The concentration profile of the master SOS transcriptional repressor can be calculated, demonstrating that relative protein levels may be determined from purely transcriptional data. This finding opens the possibility of assigning kinetic parameters to transcriptional networks on a genomic scale.

  5. Maternal recall of infant feeding events is accurate.

    PubMed Central

    Launer, L J; Forman, M R; Hundt, G L; Sarov, B; Chang, D; Berendes, H W; Naggan, L

    1992-01-01

    STUDY OBJECTIVE--Retrospective infant feeding data are important to the study of child and adult health patterns. The accuracy of maternal recall of past infant feeding events was examined and specifically the infant's age when breast feeding was stopped and formula feeding and solid foods were introduced. DESIGN AND SETTING--The sample consisted of Bedouin Arab women (n = 318) living in the Negev in Israel who were a part of a larger cohort participating in a prospective study of infant health and who were delivered of their infants between July 1 and December 15, 1981. Data from interviews conducted 12 and 18 months postpartum were compared to the standard data collected six months postpartum. MAIN RESULTS--As length of recall increased there was a small increase in the mean difference, and its standard deviation, between the standard and recalled age when breast feeding was stopped and formula feeding and solid foods were started. Recall on formula feeding was less accurate than recall on solid foods and breast feeding. In particular, among those 61% reporting formula use at the six month interview, 51% did not recall introducing formula when interviewed at 18 months. The odds ratio (95% CI) of stunting versus normal length for age for formula fed versus breast fed infants based on recall data (OR = 2.07; 95% CI 0.82-5.22) differed only slightly from those based on the standard data (OR = 2.21; 95% CI 0.77-6.37). The accuracy of a mother's recall varied with her child's nutritional status at the time of the interview, but not with other sociodemographic, infant, or interviewer characteristics. CONCLUSIONS--Retrospective infant feeding data based on maternal recall of events up to 18 months in the past can be used with confidence in epidemiological studies. However, data on formula feeding may not be as accurate as data on breast feeding and solid food feeding, and accuracy may decrease as length of recall increases. PMID:1645071

  6. Molecular mechanisms of ETS transcription factor-mediated tumorigenesis.

    PubMed

    Kar, Adwitiya; Gutierrez-Hartmann, Arthur

    2013-01-01

    The E26 transformation-specific (ETS) family of transcription factors is critical for development, differentiation, proliferation and also has a role in apoptosis and tissue remodeling. Changes in expression of ETS proteins therefore have a significant impact on normal physiology of the cell. Transcriptional consequences of ETS protein deregulation by overexpression, gene fusion, and modulation by RAS/MAPK signaling are linked to alterations in normal cell functions, and lead to unlimited increased proliferation, sustained angiogenesis, invasion and metastasis. Existing data show that ETS proteins control pathways in epithelial cells as well as stromal compartments, and the crosstalk between the two is essential for normal development and cancer. In this review, we have focused on ETS factors with a known contribution in cancer development. Instead of focusing on a prototype, we address cancer associated ETS proteins and have highlighted the diverse mechanisms by which they affect carcinogenesis. Finally, we discuss strategies for ETS factor targeting as a potential means for cancer therapeutics.

  7. Molecular mechanisms of ETS transcription factor mediated tumorigenesis

    PubMed Central

    Kar, Adwitiya; Gutierrez-Hartmann, Arthur

    2014-01-01

    The ETS family of transcription factors is critical for development, differentiation, proliferation and also has a role in apoptosis and tissue remodeling. Changes in expression of ETS proteins therefore have a significant impact on normal physiology of the cell. Transcriptional consequences of ETS protein deregulation by overexpression, gene fusion, and modulation by RAS/MAPK signaling are linked to alterations in normal cell functions, and lead to unlimited increased proliferation, sustained angiogenesis, invasion and metastasis. Existing data show that ETS proteins control pathways in epithelial cells as well as stromal compartments, and the crosstalk between the two is essential for normal development and cancer. In this review we have focused on ETS factors with a known contribution in cancer development. Instead of focusing on a prototype, we address cancer associated ETS proteins and have highlighted the diverse mechanisms by which they affect carcinogenesis. Finally, we discuss strategies for ETS factor targeting as a potential means for cancer therapeutics. PMID:24066765

  8. Coordination of a transcriptional switch by HMGI(Y) acetylation.

    PubMed

    Munshi, N; Agalioti, T; Lomvardas, S; Merika, M; Chen, G; Thanos, D

    2001-08-10

    Dynamic control of interferon-beta (IFN-beta) gene expression requires the regulated assembly and disassembly of the enhanceosome, a higher-order nucleoprotein complex formed in response to virus infection. The enhanceosome activates transcription by recruiting the histone acetyltransferase proteins CREB binding protein (CBP) and p300/CBP-associated factors (PCAF)/GCN5, which, in addition to modifying histones, acetylate HMGI(Y), the architectural component required for enhanceosome assembly. We show that the accurate execution of the IFN-beta transcriptional switch depends on the ordered acetylation of the high-mobility group I protein HMGI(Y) by PCAF/GCN5 and CBP, which acetylate HMGI(Y) at distinct lysine residues on endogenous promoters. Whereas acetylation of HMGI(Y) by CBP at lysine-65 destabilizes the enhanceosome, acetylation of HMGI(Y) by PCAF/GCN5 at lysine-71 potentiates transcription by stabilizing the enhanceosome and preventing acetylation by CBP.

  9. Heat shock factor-4 (HSF-4a) represses basal transcription through interaction with TFIIF.

    PubMed

    Frejtag, W; Zhang, Y; Dai, R; Anderson, M G; Mivechi, N F

    2001-05-04

    The heat shock transcription factors (HSFs) regulate the expression of heat shock proteins (hsps), which are critical for normal cellular proliferation and differentiation. One of the HSFs, HSF-4, contains two alternative splice variants, one of which possesses transcriptional repressor properties in vivo. This repressor isoform inhibits basal transcription of hsps 27 and 90 in tissue culture cells. The molecular mechanisms of HSF-4a isoform-mediated transcriptional repression is unknown. Here, we present evidence that HSF-4a inhibits basal transcription in vivo when it is artificially targeted to basal promoters via the DNA-binding domain of the yeast transcription factor, GAL4. By using a highly purified, reconstituted in vitro transcription system, we show that HSF-4a represses basal transcription at an early step during preinitiation complex assembly, as pre-assembled preinitiation complexes are refractory to the inhibitory effect on transcription. This repression occurs by the HSF-4a isoform, but not by the HSF-4b isoform, which we show is capable of activating transcription from a heat shock element-driven promoter in vitro. The repression of basal transcription by HSF-4a occurs through interaction with the basal transcription factor TFIIF. TFIIF interacts with a segment of HSF-4a that is required for the trimerization of HSF-4a, and deletion of this segment no longer inhibits basal transcription. These studies suggest that HSF-4a inhibits basal transcription both in vivo and in vitro. Furthermore, this is the first report identifying an interaction between a transcriptional repressor with the basal transcription factor TFIIF.

  10. Adaptation with transcriptional regulation

    NASA Astrophysics Data System (ADS)

    Shi, Wenjia; Ma, Wenzhe; Xiong, Liyang; Zhang, Mingyue; Tang, Chao

    2017-02-01

    Biochemical adaptation is one of the basic functions that are widely implemented in biological systems for a variety of purposes such as signal sensing, stress response and homeostasis. The adaptation time scales span from milliseconds to days, involving different regulatory machineries in different processes. The adaptive networks with enzymatic regulation (ERNs) have been investigated in detail. But it remains unclear if and how other forms of regulation will impact the network topology and other features of the function. Here, we systematically studied three-node transcriptional regulatory networks (TRNs), with three different types of gene regulation logics. We found that the topologies of adaptive gene regulatory networks can still be grouped into two general classes: negative feedback loop (NFBL) and incoherent feed-forward loop (IFFL), but with some distinct topological features comparing to the enzymatic networks. Specifically, an auto-activation loop on the buffer node is necessary for the NFBL class. For IFFL class, the control node can be either a proportional node or an inversely-proportional node. Furthermore, the tunability of adaptive behavior differs between TRNs and ERNs. Our findings highlight the role of regulation forms in network topology, implementation and dynamics.

  11. Adaptation with transcriptional regulation.

    PubMed

    Shi, Wenjia; Ma, Wenzhe; Xiong, Liyang; Zhang, Mingyue; Tang, Chao

    2017-02-24

    Biochemical adaptation is one of the basic functions that are widely implemented in biological systems for a variety of purposes such as signal sensing, stress response and homeostasis. The adaptation time scales span from milliseconds to days, involving different regulatory machineries in different processes. The adaptive networks with enzymatic regulation (ERNs) have been investigated in detail. But it remains unclear if and how other forms of regulation will impact the network topology and other features of the function. Here, we systematically studied three-node transcriptional regulatory networks (TRNs), with three different types of gene regulation logics. We found that the topologies of adaptive gene regulatory networks can still be grouped into two general classes: negative feedback loop (NFBL) and incoherent feed-forward loop (IFFL), but with some distinct topological features comparing to the enzymatic networks. Specifically, an auto-activation loop on the buffer node is necessary for the NFBL class. For IFFL class, the control node can be either a proportional node or an inversely-proportional node. Furthermore, the tunability of adaptive behavior differs between TRNs and ERNs. Our findings highlight the role of regulation forms in network topology, implementation and dynamics.

  12. Adaptation with transcriptional regulation

    PubMed Central

    Shi, Wenjia; Ma, Wenzhe; Xiong, Liyang; Zhang, Mingyue; Tang, Chao

    2017-01-01

    Biochemical adaptation is one of the basic functions that are widely implemented in biological systems for a variety of purposes such as signal sensing, stress response and homeostasis. The adaptation time scales span from milliseconds to days, involving different regulatory machineries in different processes. The adaptive networks with enzymatic regulation (ERNs) have been investigated in detail. But it remains unclear if and how other forms of regulation will impact the network topology and other features of the function. Here, we systematically studied three-node transcriptional regulatory networks (TRNs), with three different types of gene regulation logics. We found that the topologies of adaptive gene regulatory networks can still be grouped into two general classes: negative feedback loop (NFBL) and incoherent feed-forward loop (IFFL), but with some distinct topological features comparing to the enzymatic networks. Specifically, an auto-activation loop on the buffer node is necessary for the NFBL class. For IFFL class, the control node can be either a proportional node or an inversely-proportional node. Furthermore, the tunability of adaptive behavior differs between TRNs and ERNs. Our findings highlight the role of regulation forms in network topology, implementation and dynamics. PMID:28233824

  13. Accurate orbit propagation with planetary close encounters

    NASA Astrophysics Data System (ADS)

    Baù, Giulio; Milani Comparetti, Andrea; Guerra, Francesca

    2015-08-01

    We tackle the problem of accurately propagating the motion of those small bodies that undergo close approaches with a planet. The literature is lacking on this topic and the reliability of the numerical results is not sufficiently discussed. The high-frequency components of the perturbation generated by a close encounter makes the propagation particularly challenging both from the point of view of the dynamical stability of the formulation and the numerical stability of the integrator. In our approach a fixed step-size and order multistep integrator is combined with a regularized formulation of the perturbed two-body problem. When the propagated object enters the region of influence of a celestial body, the latter becomes the new primary body of attraction. Moreover, the formulation and the step-size will also be changed if necessary. We present: 1) the restarter procedure applied to the multistep integrator whenever the primary body is changed; 2) new analytical formulae for setting the step-size (given the order of the multistep, formulation and initial osculating orbit) in order to control the accumulation of the local truncation error and guarantee the numerical stability during the propagation; 3) a new definition of the region of influence in the phase space. We test the propagator with some real asteroids subject to the gravitational attraction of the planets, the Yarkovsky and relativistic perturbations. Our goal is to show that the proposed approach improves the performance of both the propagator implemented in the OrbFit software package (which is currently used by the NEODyS service) and of the propagator represented by a variable step-size and order multistep method combined with Cowell's formulation (i.e. direct integration of position and velocity in either the physical or a fictitious time).

  14. Important Nearby Galaxies without Accurate Distances

    NASA Astrophysics Data System (ADS)

    McQuinn, Kristen

    2014-10-01

    The Spitzer Infrared Nearby Galaxies Survey (SINGS) and its offspring programs (e.g., THINGS, HERACLES, KINGFISH) have resulted in a fundamental change in our view of star formation and the ISM in galaxies, and together they represent the most complete multi-wavelength data set yet assembled for a large sample of nearby galaxies. These great investments of observing time have been dedicated to the goal of understanding the interstellar medium, the star formation process, and, more generally, galactic evolution at the present epoch. Nearby galaxies provide the basis for which we interpret the distant universe, and the SINGS sample represents the best studied nearby galaxies.Accurate distances are fundamental to interpreting observations of galaxies. Surprisingly, many of the SINGS spiral galaxies have numerous distance estimates resulting in confusion. We can rectify this situation for 8 of the SINGS spiral galaxies within 10 Mpc at a very low cost through measurements of the tip of the red giant branch. The proposed observations will provide an accuracy of better than 0.1 in distance modulus. Our sample includes such well known galaxies as M51 (the Whirlpool), M63 (the Sunflower), M104 (the Sombrero), and M74 (the archetypal grand design spiral).We are also proposing coordinated parallel WFC3 UV observations of the central regions of the galaxies, rich with high-mass UV-bright stars. As a secondary science goal we will compare the resolved UV stellar populations with integrated UV emission measurements used in calibrating star formation rates. Our observations will complement the growing HST UV atlas of high resolution images of nearby galaxies.

  15. Accurate paleointensities - the multi-method approach

    NASA Astrophysics Data System (ADS)

    de Groot, Lennart

    2016-04-01

    The accuracy of models describing rapid changes in the geomagnetic field over the past millennia critically depends on the availability of reliable paleointensity estimates. Over the past decade methods to derive paleointensities from lavas (the only recorder of the geomagnetic field that is available all over the globe and through geologic times) have seen significant improvements and various alternative techniques were proposed. The 'classical' Thellier-style approach was optimized and selection criteria were defined in the 'Standard Paleointensity Definitions' (Paterson et al, 2014). The Multispecimen approach was validated and the importance of additional tests and criteria to assess Multispecimen results must be emphasized. Recently, a non-heating, relative paleointensity technique was proposed -the pseudo-Thellier protocol- which shows great potential in both accuracy and efficiency, but currently lacks a solid theoretical underpinning. Here I present work using all three of the aforementioned paleointensity methods on suites of young lavas taken from the volcanic islands of Hawaii, La Palma, Gran Canaria, Tenerife, and Terceira. Many of the sampled cooling units are <100 years old, the actual field strength at the time of cooling is therefore reasonably well known. Rather intuitively, flows that produce coherent results from two or more different paleointensity methods yield the most accurate estimates of the paleofield. Furthermore, the results for some flows pass the selection criteria for one method, but fail in other techniques. Scrutinizing and combing all acceptable results yielded reliable paleointensity estimates for 60-70% of all sampled cooling units - an exceptionally high success rate. This 'multi-method paleointensity approach' therefore has high potential to provide the much-needed paleointensities to improve geomagnetic field models for the Holocene.

  16. Accurate glucose detection in a small etalon

    NASA Astrophysics Data System (ADS)

    Martini, Joerg; Kuebler, Sebastian; Recht, Michael; Torres, Francisco; Roe, Jeffrey; Kiesel, Peter; Bruce, Richard

    2010-02-01

    We are developing a continuous glucose monitor for subcutaneous long-term implantation. This detector contains a double chamber Fabry-Perot-etalon that measures the differential refractive index (RI) between a reference and a measurement chamber at 850 nm. The etalon chambers have wavelength dependent transmission maxima which dependent linearly on the RI of their contents. An RI difference of ▵n=1.5.10-6 changes the spectral position of a transmission maximum by 1pm in our measurement. By sweeping the wavelength of a single-mode Vertical-Cavity-Surface-Emitting-Laser (VCSEL) linearly in time and detecting the maximum transmission peaks of the etalon we are able to measure the RI of a liquid. We have demonstrated accuracy of ▵n=+/-3.5.10-6 over a ▵n-range of 0 to 1.75.10-4 and an accuracy of 2% over a ▵nrange of 1.75.10-4 to 9.8.10-4. The accuracy is primarily limited by the reference measurement. The RI difference between the etalon chambers is made specific to glucose by the competitive, reversible release of Concanavalin A (ConA) from an immobilized dextran matrix. The matrix and ConA bound to it, is positioned outside the optical detection path. ConA is released from the matrix by reacting with glucose and diffuses into the optical path to change the RI in the etalon. Factors such as temperature affect the RI in measurement and detection chamber equally but do not affect the differential measurement. A typical standard deviation in RI is +/-1.4.10-6 over the range 32°C to 42°C. The detector enables an accurate glucose specific concentration measurement.

  17. Accurate Biomass Estimation via Bayesian Adaptive Sampling

    NASA Astrophysics Data System (ADS)

    Wheeler, K.; Knuth, K.; Castle, P.

    2005-12-01

    and IKONOS imagery and the 3-D volume estimates. The combination of these then allow for a rapid and hopefully very accurate estimation of biomass.

  18. How flatbed scanners upset accurate film dosimetry

    NASA Astrophysics Data System (ADS)

    van Battum, L. J.; Huizenga, H.; Verdaasdonk, R. M.; Heukelom, S.

    2016-01-01

    Film is an excellent dosimeter for verification of dose distributions due to its high spatial resolution. Irradiated film can be digitized with low-cost, transmission, flatbed scanners. However, a disadvantage is their lateral scan effect (LSE): a scanner readout change over its lateral scan axis. Although anisotropic light scattering was presented as the origin of the LSE, this paper presents an alternative cause. Hereto, LSE for two flatbed scanners (Epson 1680 Expression Pro and Epson 10000XL), and Gafchromic film (EBT, EBT2, EBT3) was investigated, focused on three effects: cross talk, optical path length and polarization. Cross talk was examined using triangular sheets of various optical densities. The optical path length effect was studied using absorptive and reflective neutral density filters with well-defined optical characteristics (OD range 0.2-2.0). Linear polarizer sheets were used to investigate light polarization on the CCD signal in absence and presence of (un)irradiated Gafchromic film. Film dose values ranged between 0.2 to 9 Gy, i.e. an optical density range between 0.25 to 1.1. Measurements were performed in the scanner’s transmission mode, with red-green-blue channels. LSE was found to depend on scanner construction and film type. Its magnitude depends on dose: for 9 Gy increasing up to 14% at maximum lateral position. Cross talk was only significant in high contrast regions, up to 2% for very small fields. The optical path length effect introduced by film on the scanner causes 3% for pixels in the extreme lateral position. Light polarization due to film and the scanner’s optical mirror system is the main contributor, different in magnitude for the red, green and blue channel. We concluded that any Gafchromic EBT type film scanned with a flatbed scanner will face these optical effects. Accurate dosimetry requires correction of LSE, therefore, determination of the LSE per color channel and dose delivered to the film.

  19. Towards Accurate Application Characterization for Exascale (APEX)

    SciTech Connect

    Hammond, Simon David

    2015-09-01

    Sandia National Laboratories has been engaged in hardware and software codesign activities for a number of years, indeed, it might be argued that prototyping of clusters as far back as the CPLANT machines and many large capability resources including ASCI Red and RedStorm were examples of codesigned solutions. As the research supporting our codesign activities has moved closer to investigating on-node runtime behavior a nature hunger has grown for detailed analysis of both hardware and algorithm performance from the perspective of low-level operations. The Application Characterization for Exascale (APEX) LDRD was a project concieved of addressing some of these concerns. Primarily the research was to intended to focus on generating accurate and reproducible low-level performance metrics using tools that could scale to production-class code bases. Along side this research was an advocacy and analysis role associated with evaluating tools for production use, working with leading industry vendors to develop and refine solutions required by our code teams and to directly engage with production code developers to form a context for the application analysis and a bridge to the research community within Sandia. On each of these accounts significant progress has been made, particularly, as this report will cover, in the low-level analysis of operations for important classes of algorithms. This report summarizes the development of a collection of tools under the APEX research program and leaves to other SAND and L2 milestone reports the description of codesign progress with Sandia’s production users/developers.

  20. Normal osteoid tissue

    PubMed Central

    Raina, Vinita

    1972-01-01

    The results of a histological study of normal osteoid tissue in man, the monkey, the dog, and the rat, using thin microtome sections of plastic-embedded undecalcified bone, are described. Osteoid tissue covers the entire bone surface, except for areas of active resorption, although the thickness of the layer of osteoid tissue varies at different sites and in different species of animals. The histological features of osteoid tissue, apart from its amount, are the same in the different species studied. Distinct bands or zones are recognizable in some layers of osteoid tissue, particularly those of greatest thickness, and their significance is discussed. Some of the histological features of the calcification front are described. Images PMID:4111820

  1. Normal Untreated Jurkat Cells

    NASA Technical Reports Server (NTRS)

    2004-01-01

    Biomedical research offers hope for a variety of medical problems, from diabetes to the replacement of damaged bone and tissues. Bioreactors, which are used to grow cells and tissue cultures, play a major role in such research and production efforts. The objective of the research was to define a way to differentiate between effects due to microgravity and those due to possible stress from non-optimal spaceflight conditions. These Jurkat cells, a human acute T-cell leukemia was obtained to evaluate three types of potential experimental stressors: a) Temperature elevation; b) Serum starvation; and c) Centrifugal force. The data from previous spaceflight experiments showed that actin filaments and cell shape are significantly different for the control. These normal cells serve as the baseline for future spaceflight experiments.

  2. Pornography, normalization, and empowerment.

    PubMed

    Weinberg, Martin S; Williams, Colin J; Kleiner, Sibyl; Irizarry, Yasmiyn

    2010-12-01

    Opponents and proponents of erotic representations (referred to hereafter as "pornography") have described the effects of pornography from their perspective. Little, however, has been done in the way of research to investigate these claims from the consumer's point of view. This especially has been so regarding the positive impact of such consumption on a person's sex life. Using a study group of 245 college students, we examined this question in a framework of scripting theory. We wanted to see whether viewing pornography appeared to expand sexual horizons through normalization and facilitate a willingness to explore new sexual behaviors and sexual relationships through empowerment. The data supported this viewpoint and further showed the effects to be mediated by gender and sexual preference identity. They suggested, however, that established scripts were extended rather than abandoned. We conclude with connections between our findings and the widespread viewing of pornography in contemporary society.

  3. Bidirectional transcription of trinucleotide repeats: roles for excision repair

    PubMed Central

    Budworth, Helen; McMurray, Cynthia T.

    2013-01-01

    SUMMARY Genomic instability at repetitive DNA regions in cells of the nervous system leads to a number of neurodegenerative and neuromuscular diseases, including those with an expanded trinucleotide repeat (TNR) tract at or nearby an expressed gene. Expansion causes disease when a particular base sequence is repeated beyond the normal range, interfering with the expression or properties of a gene product. Disease severity and onset depend on the number of repeats. As the length of the repeat tract grows, so does the size of the successive expansions and the likelihood of another unstable event. In fragile X syndrome, for example, CGG repeat instability and pathogenesis are not typically observed below tracts of roughly 50 repeats, but occur frequently at or above 55 repeats, and are virtually certain above 100–300 repeats. Recent evidence points to bidirectional transcription as a new aspect of TNR instability and pathophysiology. Bidirectional transcription of TNR genes produces novel proteins and/or regulatory RNAs that influence both toxicity and epigenetic changes in TNR promoters. Bidirectional transcription of the TNR tract appears to influence aspects of its stability, gene processing, splicing, gene silencing, and chemical modification of DNAs. Paradoxically, however, some of the same effects are observed on both the expanded TNR gene and on its normal gene counterpart. In this review, we discuss the possible normal and abnormal effects of bidirectional transcription on trinucleotide repeat instability, the role of DNA repair in causing, preventing, or maintaining methylation, and chromatin environment of TNR genes. PMID:23669397

  4. Highly accurate fast lung CT registration

    NASA Astrophysics Data System (ADS)

    Rühaak, Jan; Heldmann, Stefan; Kipshagen, Till; Fischer, Bernd

    2013-03-01

    Lung registration in thoracic CT scans has received much attention in the medical imaging community. Possible applications range from follow-up analysis, motion correction for radiation therapy, monitoring of air flow and pulmonary function to lung elasticity analysis. In a clinical environment, runtime is always a critical issue, ruling out quite a few excellent registration approaches. In this paper, a highly efficient variational lung registration method based on minimizing the normalized gradient fields distance measure with curvature regularization is presented. The method ensures diffeomorphic deformations by an additional volume regularization. Supplemental user knowledge, like a segmentation of the lungs, may be incorporated as well. The accuracy of our method was evaluated on 40 test cases from clinical routine. In the EMPIRE10 lung registration challenge, our scheme ranks third, with respect to various validation criteria, out of 28 algorithms with an average landmark distance of 0.72 mm. The average runtime is about 1:50 min on a standard PC, making it by far the fastest approach of the top-ranking algorithms. Additionally, the ten publicly available DIR-Lab inhale-exhale scan pairs were registered to subvoxel accuracy at computation times of only 20 seconds. Our method thus combines very attractive runtimes with state-of-the-art accuracy in a unique way.

  5. Transcription factor binding energy vs. biological function

    NASA Astrophysics Data System (ADS)

    Djordjevic, M.; Grotewold, E.

    2007-03-01

    Transcription factors (TFs) are proteins that bind to DNA and regulate expression of genes. Identification of transcription factor binding sites within the regulatory segments of genomic DNA is an important step towards understanding of gene regulatory networks. Recent theoretical advances that we developed [1,2], allow us to infer TF-DNA interaction parameters from in-vitro selection experiments [3]. We use more than 6000 binding sequences [3], assembled under controlled conditions, to obtain protein-DNA interaction parameters for a mammalian TF with up to now unprecedented accuracy. Can one accurately identify biologically functional TF binding sites (i.e. the binding sites that regulate gene expression), even with the best possible protein-DNA interaction parameters? To address this issue we i) compare our prediction of protein binding with gene expression data, ii) use evolutionary comparison between related mammalian genomes. Our results strongly suggest that in a genome there exists a large number of randomly occurring high energy binding sites that are not biologically functional. [1] M Djordjevic, submitted to Biomol. Eng. [2] M. Djordjevic and A. M. Sengupta, Phys. Biol. 3: 13, 2006. [3] E. Roulet et al., Nature Biotech. 20: 831, 2002.

  6. ETS transcription factors in hematopoietic stem cell development.

    PubMed

    Ciau-Uitz, Aldo; Wang, Lu; Patient, Roger; Liu, Feng

    2013-12-01

    Hematopoietic stem cells (HSCs) are essential for the maintenance of the hematopoietic system. However, these cells cannot be maintained or created in vitro, and very little is known about their generation during embryogenesis. Many transcription factors and signaling pathways play essential roles at various stages of HSC development. Members of the ETS ('E twenty-six') family of transcription factors are recognized as key regulators within the gene regulatory networks governing hematopoiesis, including the ontogeny of HSCs. Remarkably, although all ETS transcription factors bind the same DNA consensus sequence and overlapping tissue expression is observed, individual ETS transcription factors play unique roles in the development of HSCs. Also, these transcription factors are recurrently used throughout development and their functions are context-dependent, increasing the challenge of studying their mechanism of action. Critically, ETS factors also play roles under pathological conditions, such as leukemia and, therefore, deciphering their mechanism of action will not only enhance our knowledge of normal hematopoiesis, but also inform protocols for their creation in vitro from pluripotent stem cells and the design of new therapeutic approaches for the treatment of malignant blood cell diseases. In this review, we summarize the key findings on the roles of ETS transcription factors in HSC development and discuss novel mechanisms by which they could control hematopoiesis.

  7. The SCL gene is formed from a transcriptionally complex locus.

    PubMed Central

    Aplan, P D; Begley, C G; Bertness, V; Nussmeier, M; Ezquerra, A; Coligan, J; Kirsch, I R

    1990-01-01

    We describe the structural organization of the human SCL gene, a helix-loop-helix family member which we believe plays a fundamental role in hematopoietic differentiation. The SCL locus is composed of eight exons distributed over 16 kb. SCL shows a pattern of expression quite restricted to early hematopoietic tissues, although in malignant states expression of the gene may be somewhat extended into later developmental stages. A detailed analysis of the transcript(s) arising from the SCL locus revealed that (i) the 5' noncoding portion of the SCL transcript, which resides within a CpG island, has a complex pattern of alternative exon utilization as well as two distinct transcription initiation sites; (ii) the 5' portions of the SCL transcript contain features that suggest a possible regulatory role for these segments; (iii) the pattern of utilization of the 5' exons is cell lineage dependent; and (iv) all of the currently studied chromosomal aberrations that affect the SCL locus either structurally or functionally eliminate the normal 5' transcription initiation sites. These data suggest that the SCL gene, and specifically its 5' region, may be a target for regulatory interactions during early hematopoietic development. Images PMID:2247063

  8. The Role of Co-transcriptional Histone Methylations

    PubMed Central

    Buratowski, Stephen; Kim, TaeSoo

    2011-01-01

    The C-terminal domain (CTD) of the RNA polymerase II subunit Rpb1 undergoes dynamic phosphorylation, with different phosphorylation sites predominating at different stages of transcription. Our lab studies how various mRNA processing and chromatin-modifying enzymes interact with the phosphorylated CTD to efficiently produce mRNAs. The H3K36 methyltransferase Set2 interacts with CTD carrying phosphorylations characteristic of downstream elongation complexes, and the resulting co-transcriptional H3K36 methylation targets the Rpd3S histone deacetylase to downstream transcribed regions. Although positively correlated with gene activity, this pathway actually inhibits transcription elongation as well as initiation from cryptic promoters within genes. During early elongation, CTD serine 5 phosphorylation helps recruit the H3K4 methyltransferase complex containing Set1. Within 5' transcribed regions, co-transcriptional H3K4 dimethylation (H3K4me2) by Set1 recruits the deacetylase complex Set3C. Finally, H3K4 trimethylation at the most promoter-proximal nucleosomes is thought to stimulate transcription by promoting histone acetylation by complexes containing the ING/Yng PHD finger proteins. Surprisingly, the Rpd3L histone deacetylase complex, normally a transcription repressor, may also recognize H3K4me3. Together, the cotranscriptional histone methylations appear to primarily function to distinguish active promoter regions, which are marked by high levels of acetylation and nucleosome turnover, from the deacetylated, downstream transcribed regions of genes. PMID:21447819

  9. Evolutionary Profiling of the Transcriptional and Post-Transcriptional Wiring of the Circadian Clock in Normal and Perturbed Conditions

    DTIC Science & Technology

    2014-08-14

    activity of eight different species of drosophila from different parts of the world, that have adapted to different environmental conditions in three...different temperatures and compared the results to the well-studied Drosophila melanogaster. We kept the flies in 12 hours light 12 hours dark cycles and...how the circadian cycle adapts to environmental stress. More specifically: As expected, Drosophila melanogaster showed two peaks of activity: one in

  10. Transcription of Trypanosoma brucei maxicircles

    SciTech Connect

    Michelotti, E.F.; Hajduk, S.L.

    1987-05-01

    Trypanosoma brucei is a protozoan parasite which developmentally regulates mitochondrial activity. In the mammal T. brucei produces ATP entirely by glycolysis while cytochrome mediated respiration resumes in the life-stage in the midgut of the insect vector. Using quantitative S1 nuclease protection assays two types of regulation of the steady state levels of the mitochondrial transcripts were found. Transcription of cytochrome b, cytochrome oxidase, and the rRNA genes is repressed in early bloodstream developmental stages, undergoes dramatic activation in later bloodstream stages, and finally a lesser activation in the insect developmental stage. Transcription of NADH dehydrogenase genes, however, is unregulated. Mitochondrial transcripts with a 5' triphosphate terminus, representing the site of transcription initiation, were capped using guanylyl transferase. The in vitro capped RNA hybridized to only one of eight mitochondrial restriction fragments on a Southern blot, however, hybridization of Southern blots with RNA from ..cap alpha..-/sup 32/P-UTP pulsed mitochondria labelled all restriction fragments equally. These results suggest that each DNA strand has a single promoter which directs the transcription of a full-length RNA which is subsequently processed. Different mitochondrial genes, despite being expressed on the same precursor RNA molecule, are independently regulated by both transcription initiation and RNA processing.

  11. Accurate Reading with Sequential Presentation of Single Letters

    PubMed Central

    Price, Nicholas S. C.; Edwards, Gemma L.

    2012-01-01

    Rapid, accurate reading is possible when isolated, single words from a sentence are sequentially presented at a fixed spatial location. We investigated if reading of words and sentences is possible when single letters are rapidly presented at the fovea under user-controlled or automatically controlled rates. When tested with complete sentences, trained participants achieved reading rates of over 60 wpm and accuracies of over 90% with the single letter reading (SLR) method and naive participants achieved average reading rates over 30 wpm with greater than 90% accuracy. Accuracy declined as individual letters were presented for shorter periods of time, even when the overall reading rate was maintained by increasing the duration of spaces between words. Words in the lexicon that occur more frequently were identified with higher accuracy and more quickly, demonstrating that trained participants have lexical access. In combination, our data strongly suggest that comprehension is possible and that SLR is a practicable form of reading under conditions in which normal scanning of text is not possible, or for scenarios with limited spatial and temporal resolution such as patients with low vision or prostheses. PMID:23115548

  12. Accurate reading with sequential presentation of single letters.

    PubMed

    Price, Nicholas S C; Edwards, Gemma L

    2012-01-01

    Rapid, accurate reading is possible when isolated, single words from a sentence are sequentially presented at a fixed spatial location. We investigated if reading of words and sentences is possible when single letters are rapidly presented at the fovea under user-controlled or automatically controlled rates. When tested with complete sentences, trained participants achieved reading rates of over 60 wpm and accuracies of over 90% with the single letter reading (SLR) method and naive participants achieved average reading rates over 30 wpm with greater than 90% accuracy. Accuracy declined as individual letters were presented for shorter periods of time, even when the overall reading rate was maintained by increasing the duration of spaces between words. Words in the lexicon that occur more frequently were identified with higher accuracy and more quickly, demonstrating that trained participants have lexical access. In combination, our data strongly suggest that comprehension is possible and that SLR is a practicable form of reading under conditions in which normal scanning of text is not possible, or for scenarios with limited spatial and temporal resolution such as patients with low vision or prostheses.

  13. AthaMap, integrating transcriptional and post-transcriptional data

    PubMed Central

    Bülow, Lorenz; Engelmann, Stefan; Schindler, Martin; Hehl, Reinhard

    2009-01-01

    The AthaMap database generates a map of predicted transcription factor binding sites (TFBS) for the whole Arabidopsis thaliana genome. AthaMap has now been extended to include data on post-transcriptional regulation. A total of 403 173 genomic positions of small RNAs have been mapped in the A. thaliana genome. These identify 5772 putative post-transcriptionally regulated target genes. AthaMap tools have been modified to improve the identification of common TFBS in co-regulated genes by subtracting post-transcriptionally regulated genes from such analyses. Furthermore, AthaMap was updated to the TAIR7 genome annotation, a graphic display of gene analysis results was implemented, and the TFBS data content was increased. AthaMap is freely available at http://www.athamap.de/. PMID:18842622

  14. Transcription regulation mechanisms of bacteriophages

    PubMed Central

    Yang, Haiquan; Ma, Yingfang; Wang, Yitian; Yang, Haixia; Shen, Wei; Chen, Xianzhong

    2014-01-01

    Phage diversity significantly contributes to ecology and evolution of new bacterial species through horizontal gene transfer. Therefore, it is essential to understand the mechanisms underlying phage-host interactions. After initial infection, the phage utilizes the transcriptional machinery of the host to direct the expression of its own genes. This review presents a view on the transcriptional regulation mechanisms of bacteriophages, and its contribution to phage diversity and classification. Through this review, we aim to broaden the understanding of phage-host interactions while providing a reference source for researchers studying the regulation of phage transcription. PMID:25482231

  15. [Normal aging and cognition].

    PubMed

    Ska, Bernadette; Joanette, Yves

    2006-03-01

    It is now well documented that normal aging modifies the cognitive functioning and most observations suggest that cognition evolves in the direction of deterioration. The more frequently impaired functions are memory, attention and visual-spatial abilities. On the other hand, some abilities seem to increase, such as vocabulary. Considering the aging effect on cognition, questions remain regarding directionality, universality and reversibility. A great variability in aged related impacts is observed among subjects and among cognitive domains. Some individuals evolved more rapidly than others. Some cognitive functions are more affected by aging than others. General and specific factors are hypothesized to explain the aged related cognitive decline. Among them, educational level, health, cognitive style, life style, personality, are likely to modulate the aged related cognitive evolution by influencing attentional resources and cerebral plasticity. Cognitive resources are essential to develop adaptative strategies. During the life span, resources are activated and increased by learning and training. Considering the role of cognitive resources, successful aging is dependent on several conditions : absence of disease leading to a loss of autonomy, maintenance of cognitive and physical activities, and active and social engaged lifestyle.

  16. 77 FR 3800 - Accurate NDE & Inspection, LLC; Confirmatory Order

    Federal Register 2010, 2011, 2012, 2013, 2014

    2012-01-25

    ... COMMISSION Accurate NDE & Inspection, LLC; Confirmatory Order In the Matter of Accurate NDE & Docket: 150... request ADR with the NRC in an attempt to resolve issues associated with this matter. In response, on August 9, 2011, Accurate NDE requested ADR to resolve this matter with the NRC. On September 28,...

  17. Using the international normalized ratio to standardize prothrombin time.

    PubMed

    Stern, R; Karlis, V; Kinney, L; Glickman, R

    1997-08-01

    The international normalized ratio, or INR, was introduced in 1983 by the World Health Organization, or WHO, Committee on Biological Standards to more accurately assess patients receiving anticoagulation therapy. The INR mandates the universal standardization of prothrombin time. This article describes the method used to calculate INR, as well as its clinical relevance to the practice of dentistry.

  18. Anticipatory Imagery Ability in Normal and Language-Disabled Children.

    ERIC Educational Resources Information Center

    Savich, Patricia A.

    1984-01-01

    five spatial tasks were administered to two groups of seven and one-half to nine and one-half year olds: 18 language-disabled and 18 children with normal language development. The language-disabled were less accurate on all tasks which involved anticipation or prediction of mental rotations, movements, or other transformations. (Author/CL)

  19. Coupling transcription and alternative splicing.

    PubMed

    Kornblihtt, Alberto R

    2007-01-01

    Alternative splicing regulation not only depends on the interaction of splicing factors with splicing enhancers and silencers in the pre-mRNA, but also on the coupling between transcription and splicing. This coupling is possible because splicing is often cotranscriptional and promoter identity and occupation may affect alternative splicing. We discuss here the different mechanisms by which transcription regulates alternative splicing. These include the recruitment of splicing factors to the transcribing polymerase and "kinetic coupling", which involves changes in the rate of transcriptional elongation that in turn affect the timing in which splice sites are presented to the splicing machinery. The recruitment mechanism may depend on the particular features of the carboxyl terminal domain of RNA polymerase II, whereas kinetic coupling seems to be linked to how changes in chromatin structure and other factors affect transcription elongation.

  20. RNA-guided transcriptional regulation

    DOEpatents

    Church, George M.; Mali, Prashant G.; Esvelt, Kevin M.

    2016-02-23

    Methods of modulating expression of a target nucleic acid in a cell are provided including introducing into the cell a first foreign nucleic acid encoding one or more RNAs complementary to DNA, wherein the DNA includes the target nucleic acid, introducing into the cell a second foreign nucleic acid encoding a nuclease-null Cas9 protein that binds to the DNA and is guided by the one or more RNAs, introducing into the cell a third foreign nucleic acid encoding a transcriptional regulator protein or domain, wherein the one or more RNAs, the nuclease-null Cas9 protein, and the transcriptional regulator protein or domain are expressed, wherein the one or more RNAs, the nuclease-null Cas9 protein and the transcriptional regulator protein or domain co-localize to the DNA and wherein the transcriptional regulator protein or domain regulates expression of the target nucleic acid.

  1. Nucleolar localization of myc transcripts.

    PubMed Central

    Bond, V C; Wold, B

    1993-01-01

    In situ hybridization has revealed a striking subnuclear distribution of c-myc RNA transcripts. A major fraction of the sense-strand nuclear c-myc transcripts was localized to the nucleoli. myc intron 1-containing RNAs were noticeably absent from nucleoli, accumulating instead in the nucleoplasm. The localization of myc RNA to nucleoli was shown to be common to a number of diverse cell types, including primary Sertoli cells and several cell lines. Furthermore, nucleolar localization was not restricted to c-myc and N-myc and myoD transcripts also displayed this phenomenon. In contrast, gamma-actin or lactate dehydrogenase transcripts did not display nucleolar localization. These observations suggest a new role for the nucleolus in transport and/or turnover of potential mRNAs. Images PMID:7684491

  2. Normal Conducting CLIC Technology

    SciTech Connect

    Jensen, Erk

    2006-01-03

    The CLIC (Compact Linear Collider) multi-lateral study group based at CERN is studying the technology for an electron-positron linear collider with a centre-of-mass energy up to 5 TeV. In contrast to the International Linear Collider (ILC) study which has chosen to use super-conducting cavities with accelerating gradients in the range of 30-40 MV/m to obtain centre-of-mass collision energies of 0.5-1 TeV, the CLIC study aims to use a normal-conducting system based on two-beam technology with gradients of 150 MV/m. It is generally accepted that this change in technology is not only necessary but the only viable choice for a cost-effective multi-TeV collider. The CLIC study group is studying the technology issues of such a machine, and is in particular developing state-of-the-art 30 GHz molybdenum-iris accelerating structures and power extraction and transfer structures (PETS). The accelerating structure has a new geometry which includes fully-profiled RF surfaces optimised to minimize surface fields, and hybrid damping using both iris slots and radial waveguides. A newly-developed structure-optimisation procedure has been used to simultaneously balance surface fields, power flow, short and long-range transverse wakefields, RF-to-beam efficiency and the ratio of luminosity to input power. The slotted irises allow a simple structure fabrication by high-precision high-speed 3D milling of just four pieces, and an even easier bolted assembly in a vacuum chamber.

  3. Normal Conducting CLIC Technology

    NASA Astrophysics Data System (ADS)

    Jensen, Erk

    2006-01-01

    The CLIC (Compact Linear Collider) multi-lateral study group based at CERN is studying the technology for an electron-positron linear collider with a centre-of-mass energy up to 5 TeV. In contrast to the International Linear Collider (ILC) study which has chosen to use super-conducting cavities with accelerating gradients in the range of 30-40 MV/m to obtain centre-of-mass collision energies of 0.5-1 TeV, the CLIC study aims to use a normal-conducting system based on two-beam technology with gradients of 150 MV/m. It is generally accepted that this change in technology is not only necessary but the only viable choice for a cost-effective multi-TeV collider. The CLIC study group is studying the technology issues of such a machine, and is in particular developing state-of-the-art 30 GHz molybdenum-iris accelerating structures and power extraction and transfer structures (PETS). The accelerating structure has a new geometry which includes fully-profiled RF surfaces optimised to minimize surface fields, and hybrid damping using both iris slots and radial waveguides. A newly-developed structure-optimisation procedure has been used to simultaneously balance surface fields, power flow, short and long-range transverse wakefields, RF-to-beam efficiency and the ratio of luminosity to input power. The slotted irises allow a simple structure fabrication by high-precision high-speed 3D milling of just four pieces, and an even easier bolted assembly in a vacuum chamber.

  4. The transcriptional landscape of the mammalian genome.

    PubMed

    Carninci, P; Kasukawa, T; Katayama, S; Gough, J; Frith, M C; Maeda, N; Oyama, R; Ravasi, T; Lenhard, B; Wells, C; Kodzius, R; Shimokawa, K; Bajic, V B; Brenner, S E; Batalov, S; Forrest, A R R; Zavolan, M; Davis, M J; Wilming, L G; Aidinis, V; Allen, J E; Ambesi-Impiombato, A; Apweiler, R; Aturaliya, R N; Bailey, T L; Bansal, M; Baxter, L; Beisel, K W; Bersano, T; Bono, H; Chalk, A M; Chiu, K P; Choudhary, V; Christoffels, A; Clutterbuck, D R; Crowe, M L; Dalla, E; Dalrymple, B P; de Bono, B; Della Gatta, G; di Bernardo, D; Down, T; Engstrom, P; Fagiolini, M; Faulkner, G; Fletcher, C F; Fukushima, T; Furuno, M; Futaki, S; Gariboldi, M; Georgii-Hemming, P; Gingeras, T R; Gojobori, T; Green, R E; Gustincich, S; Harbers, M; Hayashi, Y; Hensch, T K; Hirokawa, N; Hill, D; Huminiecki, L; Iacono, M; Ikeo, K; Iwama, A; Ishikawa, T; Jakt, M; Kanapin, A; Katoh, M; Kawasawa, Y; Kelso, J; Kitamura, H; Kitano, H; Kollias, G; Krishnan, S P T; Kruger, A; Kummerfeld, S K; Kurochkin, I V; Lareau, L F; Lazarevic, D; Lipovich, L; Liu, J; Liuni, S; McWilliam, S; Madan Babu, M; Madera, M; Marchionni, L; Matsuda, H; Matsuzawa, S; Miki, H; Mignone, F; Miyake, S; Morris, K; Mottagui-Tabar, S; Mulder, N; Nakano, N; Nakauchi, H; Ng, P; Nilsson, R; Nishiguchi, S; Nishikawa, S; Nori, F; Ohara, O; Okazaki, Y; Orlando, V; Pang, K C; Pavan, W J; Pavesi, G; Pesole, G; Petrovsky, N; Piazza, S; Reed, J; Reid, J F; Ring, B Z; Ringwald, M; Rost, B; Ruan, Y; Salzberg, S L; Sandelin, A; Schneider, C; Schönbach, C; Sekiguchi, K; Semple, C A M; Seno, S; Sessa, L; Sheng, Y; Shibata, Y; Shimada, H; Shimada, K; Silva, D; Sinclair, B; Sperling, S; Stupka, E; Sugiura, K; Sultana, R; Takenaka, Y; Taki, K; Tammoja, K; Tan, S L; Tang, S; Taylor, M S; Tegner, J; Teichmann, S A; Ueda, H R; van Nimwegen, E; Verardo, R; Wei, C L; Yagi, K; Yamanishi, H; Zabarovsky, E; Zhu, S; Zimmer, A; Hide, W; Bult, C; Grimmond, S M; Teasdale, R D; Liu, E T; Brusic, V; Quackenbush, J; Wahlestedt, C; Mattick, J S; Hume, D A; Kai, C; Sasaki, D; Tomaru, Y; Fukuda, S; Kanamori-Katayama, M; Suzuki, M; Aoki, J; Arakawa, T; Iida, J; Imamura, K; Itoh, M; Kato, T; Kawaji, H; Kawagashira, N; Kawashima, T; Kojima, M; Kondo, S; Konno, H; Nakano, K; Ninomiya, N; Nishio, T; Okada, M; Plessy, C; Shibata, K; Shiraki, T; Suzuki, S; Tagami, M; Waki, K; Watahiki, A; Okamura-Oho, Y; Suzuki, H; Kawai, J; Hayashizaki, Y

    2005-09-02

    This study describes comprehensive polling of transcription start and termination sites and analysis of previously unidentified full-length complementary DNAs derived from the mouse genome. We identify the 5' and 3' boundaries of 181,047 transcripts with extensive variation in transcripts arising from alternative promoter usage, splicing, and polyadenylation. There are 16,247 new mouse protein-coding transcripts, including 5154 encoding previously unidentified proteins. Genomic mapping of the transcriptome reveals transcriptional forests, with overlapping transcription on both strands, separated by deserts in which few transcripts are observed. The data provide a comprehensive platform for the comparative analysis of mammalian transcriptional regulation in differentiation and development.

  5. Transcription factor repertoire of homeostatic eosinophilopoiesis

    PubMed Central

    Bouffi, Carine; Kartashov, Andrey V.; Schollaert, Kaila L.; Chen, Xiaoting; Bacon, W. Clark; Weirauch, Matthew T.; Barski, Artem; Fulkerson, Patricia C.

    2015-01-01

    The production of mature eosinophils is a tightly orchestrated process with the aim to sustain normal eosinophil levels in tissues while also maintaining low numbers of these complex and sensitive cells in the blood. To identify regulators of homeostatic eosinophilopoiesis in mice, we took a global approach to identify genome-wide transcriptome and epigenome changes that occur during homeostasis at critical developmental stages, including eosinophil-lineage commitment and lineage maturation. Our analyses revealed a markedly greater number of transcriptome alterations associated with eosinophil maturation (1199 genes) than with eosinophil-lineage commitment (490 genes), highlighting the greater transcriptional investment necessary for differentiation. Eosinophil progenitors (EoPs) were noted to express high levels of granule proteins and contain granules with an ultrastructure distinct from that of mature resting eosinophils. Our analyses also delineated a 976-gene eosinophil-lineage transcriptome that included a repertoire of 56 transcription factors, many of which have never previously been associated with eosinophils. EoPs and eosinophils, but not granulocyte-monocyte progenitors (GMPs) or neutrophils, expressed Helios and Aiolos, members of the Ikaros family of transcription factors, which regulate gene expression via modulation of chromatin structure and DNA accessibility. Epigenetic studies revealed a distinct distribution of active chromatin marks between genes induced with lineage commitment and genes induced with cell maturation during eosinophil development. In addition, Aiolos and Helios binding sites were significantly enriched in genes expressed by EoPs and eosinophils with active chromatin, highlighting a potential novel role for Helios and Aiolos in regulating gene expression during eosinophil development. PMID:26268651

  6. An analytic model for accurate spring constant calibration of rectangular atomic force microscope cantilevers.

    PubMed

    Li, Rui; Ye, Hongfei; Zhang, Weisheng; Ma, Guojun; Su, Yewang

    2015-10-29

    Spring constant calibration of the atomic force microscope (AFM) cantilever is of fundamental importance for quantifying the force between the AFM cantilever tip and the sample. The calibration within the framework of thin plate theory undoubtedly has a higher accuracy and broader scope than that within the well-established beam theory. However, thin plate theory-based accurate analytic determination of the constant has been perceived as an extremely difficult issue. In this paper, we implement the thin plate theory-based analytic modeling for the static behavior of rectangular AFM cantilevers, which reveals that the three-dimensional effect and Poisson effect play important roles in accurate determination of the spring constants. A quantitative scaling law is found that the normalized spring constant depends only on the Poisson's ratio, normalized dimension and normalized load coordinate. Both the literature and our refined finite element model validate the present results. The developed model is expected to serve as the benchmark for accurate calibration of rectangular AFM cantilevers.

  7. Polyallelic structural variants can provide accurate, highly informative genetic markers focused on diagnosis and therapeutic targets: Accuracy vs. Precision.

    PubMed

    Roses, A D

    2016-02-01

    Structural variants (SVs) include all insertions, deletions, and rearrangements in the genome, with several common types of nucleotide repeats including single sequence repeats, short tandem repeats, and insertion-deletion length variants. Polyallelic SVs provide highly informative markers for association studies with well-phenotyped cohorts. SVs can influence gene regulation by affecting epigenetics, transcription, splicing, and/or translation. Accurate assays of polyallelic SV loci are required to define the range and allele frequency of variable length alleles.

  8. A Novel Method for Accurate Operon Predictions in All SequencedProkaryotes

    SciTech Connect

    Price, Morgan N.; Huang, Katherine H.; Alm, Eric J.; Arkin, Adam P.

    2004-12-01

    We combine comparative genomic measures and the distance separating adjacent genes to predict operons in 124 completely sequenced prokaryotic genomes. Our method automatically tailors itself to each genome using sequence information alone, and thus can be applied to any prokaryote. For Escherichia coli K12 and Bacillus subtilis, our method is 85 and 83% accurate, respectively, which is similar to the accuracy of methods that use the same features but are trained on experimentally characterized transcripts. In Halobacterium NRC-1 and in Helicobacterpylori, our method correctly infers that genes in operons are separated by shorter distances than they are in E.coli, and its predictions using distance alone are more accurate than distance-only predictions trained on a database of E.coli transcripts. We use microarray data from sixphylogenetically diverse prokaryotes to show that combining intergenic distance with comparative genomic measures further improves accuracy and that our method is broadly effective. Finally, we survey operon structure across 124 genomes, and find several surprises: H.pylori has many operons, contrary to previous reports; Bacillus anthracis has an unusual number of pseudogenes within conserved operons; and Synechocystis PCC6803 has many operons even though it has unusually wide spacings between conserved adjacent genes.

  9. CLASS2: accurate and efficient splice variant annotation from RNA-seq reads

    PubMed Central

    Song, Li; Sabunciyan, Sarven; Florea, Liliana

    2016-01-01

    Next generation sequencing of cellular RNA is making it possible to characterize genes and alternative splicing in unprecedented detail. However, designing bioinformatics tools to accurately capture splicing variation has proven difficult. Current programs can find major isoforms of a gene but miss lower abundance variants, or are sensitive but imprecise. CLASS2 is a novel open source tool for accurate genome-guided transcriptome assembly from RNA-seq reads based on the model of splice graph. An extension of our program CLASS, CLASS2 jointly optimizes read patterns and the number of supporting reads to score and prioritize transcripts, implemented in a novel, scalable and efficient dynamic programming algorithm. When compared against reference programs, CLASS2 had the best overall accuracy and could detect up to twice as many splicing events with precision similar to the best reference program. Notably, it was the only tool to produce consistently reliable transcript models for a wide range of applications and sequencing strategies, including ribosomal RNA-depleted samples. Lightweight and multi-threaded, CLASS2 requires <3GB RAM and can analyze a 350 million read set within hours, and can be widely applied to transcriptomics studies ranging from clinical RNA sequencing, to alternative splicing analyses, and to the annotation of new genomes. PMID:26975657

  10. Peroxide-Sensing Transcriptional Regulators in Bacteria

    PubMed Central

    Mongkolsuk, Skorn

    2012-01-01

    The ability to maintain intracellular concentrations of toxic reactive oxygen species (ROS) within safe limits is essential for all aerobic life forms. In bacteria, as well as other organisms, ROS are produced during the normal course of aerobic metabolism, necessitating the constitutive expression of ROS scavenging systems. However, bacteria can also experience transient high-level exposure to ROS derived either from external sources, such as the host defense response, or as a secondary effect of other seemingly unrelated environmental stresses. Consequently, transcriptional regulators have evolved to sense the levels of ROS and coordinate the appropriate oxidative stress response. Three well-studied examples of these are the peroxide responsive regulators OxyR, PerR, and OhrR. OxyR and PerR are sensors of primarily H2O2, while OhrR senses organic peroxide (ROOH) and sodium hypochlorite (NaOCl). OxyR and OhrR sense oxidants by means of the reversible oxidation of specific cysteine residues. In contrast, PerR senses H2O2 via the Fe-catalyzed oxidation of histidine residues. These transcription regulators also influence complex biological phenomena, such as biofilm formation, the evasion of host immune responses, and antibiotic resistance via the direct regulation of specific proteins. PMID:22797754

  11. Peroxide-sensing transcriptional regulators in bacteria.

    PubMed

    Dubbs, James M; Mongkolsuk, Skorn

    2012-10-01

    The ability to maintain intracellular concentrations of toxic reactive oxygen species (ROS) within safe limits is essential for all aerobic life forms. In bacteria, as well as other organisms, ROS are produced during the normal course of aerobic metabolism, necessitating the constitutive expression of ROS scavenging systems. However, bacteria can also experience transient high-level exposure to ROS derived either from external sources, such as the host defense response, or as a secondary effect of other seemingly unrelated environmental stresses. Consequently, transcriptional regulators have evolved to sense the levels of ROS and coordinate the appropriate oxidative stress response. Three well-studied examples of these are the peroxide responsive regulators OxyR, PerR, and OhrR. OxyR and PerR are sensors of primarily H(2)O(2), while OhrR senses organic peroxide (ROOH) and sodium hypochlorite (NaOCl). OxyR and OhrR sense oxidants by means of the reversible oxidation of specific cysteine residues. In contrast, PerR senses H(2)O(2) via the Fe-catalyzed oxidation of histidine residues. These transcription regulators also influence complex biological phenomena, such as biofilm formation, the evasion of host immune responses, and antibiotic resistance via the direct regulation of specific proteins.

  12. From tissue mechanics to transcription factors

    PubMed Central

    Janmey, Paul A.; Wells, Rebecca G.; Assoian, Richard K.; McCulloch, Christopher A.

    2015-01-01

    Changes in tissue stiffness are frequently associated with diseases such as cancer, fibrosis, and atherosclerosis. Several recent studies suggest that, in addition to resulting from pathology, mechanical changes may play a role akin to soluble factors in causing the progression of disease, and similar mechanical control might be essential for normal tissue development and homeostasis. Many cell types alter their structure and function in response to exogenous forces or as a function of the mechanical properties of the materials to which they adhere. This review summarizes recent progress in identifying intracellular signaling pathways, and especially transcriptional programs, that are differentially activated when cells adhere to materials with different mechanical properties or when they are subject to tension arising from external forces. Several cytoplasmic or cytoskeletal signaling pathways involving small GTPases, focal adhesion kinase and transforming growth factor beta as well as the transcriptional regulators MRTF-A, NFκB, and Yap/Taz have emerged as important mediators of mechanical signaling. PMID:23969122

  13. Transcriptional regulation by post-transcriptional modification--role of phosphorylation in Sp1 transcriptional activity.

    PubMed

    Chu, Shijian

    2012-10-15

    Sp1 is a ubiquitously expressed transcription factor involved in the regulation of a large number of genes including housekeeping genes as well as actively regulated genes. Although Sp1 was discovered nearly three decades ago, its functional diversity is still not completely understood. One of the ways that make Sp1 versatile in transcriptional regulation is its post-transcriptional modification, which alters Sp1 structure in different cells and at different times. Compared to other types of modifications of the Sp1 protein, phosphorylation has been studied far more extensively. This review focuses on the inducers, pathways, enzymes, and biological effects of Sp1 phosphorylation. Recent data are beginning to reveal the biological significance and universal presence of Sp1 phosphorylation-related cell/molecular responses. Studies in this field provide a quick glance at how a simple chemical modification of a transcription factor could produce significant functional diversity of the protein.

  14. Sequence-specific thermodynamic properties of nucleic acids influence both transcriptional pausing and backtracking in yeast

    PubMed Central

    2017-01-01

    RNA Polymerase II pauses and backtracks during transcription, with many consequences for gene expression and cellular physiology. Here, we show that the energy required to melt double-stranded nucleic acids in the transcription bubble predicts pausing in Saccharomyces cerevisiae far more accurately than nucleosome roadblocks do. In addition, the same energy difference also determines when the RNA polymerase backtracks instead of continuing to move forward. This data-driven model corroborates—in a genome wide and quantitative manner—previous evidence that sequence-dependent thermodynamic features of nucleic acids influence both transcriptional pausing and backtracking. PMID:28301878

  15. DNA display of folded RNA libraries enabling RNA-SELEX without reverse transcription.

    PubMed

    MacPherson, I S; Temme, J S; Krauss, I J

    2017-03-02

    A method for the physical attachment of folded RNA libraries to their encoding DNA is presented as a way to circumvent the reverse transcription step during systematic evolution of RNA ligands by exponential enrichment (RNA-SELEX). A DNA library is modified with one isodC base to stall T7 polymerase and a 5' "capture strand" which anneals to the nascent RNA transcript. This method is validated in a selection of RNA aptamers against human α-thrombin with dissociation constants in the low nanomolar range. This method will be useful in the discovery of RNA aptamers and ribozymes containing base modifications that make them resistant to accurate reverse transcription.

  16. The use of normal forms for analysing nonlinear mechanical vibrations

    PubMed Central

    Neild, Simon A.; Champneys, Alan R.; Wagg, David J.; Hill, Thomas L.; Cammarano, Andrea

    2015-01-01

    A historical introduction is given of the theory of normal forms for simplifying nonlinear dynamical systems close to resonances or bifurcation points. The specific focus is on mechanical vibration problems, described by finite degree-of-freedom second-order-in-time differential equations. A recent variant of the normal form method, that respects the specific structure of such models, is recalled. It is shown how this method can be placed within the context of the general theory of normal forms provided the damping and forcing terms are treated as unfolding parameters. The approach is contrasted to the alternative theory of nonlinear normal modes (NNMs) which is argued to be problematic in the presence of damping. The efficacy of the normal form method is illustrated on a model of the vibration of a taut cable, which is geometrically nonlinear. It is shown how the method is able to accurately predict NNM shapes and their bifurcations. PMID:26303917

  17. Snail Family Transcription Factors Are Implicated in Thyroid Carcinogenesis

    PubMed Central

    Hardy, Robert G.; Vicente-Dueñas, Carolina; González-Herrero, Ines; Anderson, Catriona; Flores, Teresa; Hughes, Sharon; Tselepis, Chris; Ross, James A.; Sánchez-García, Isidro

    2007-01-01

    E-Cadherin (CDH1) expression is reduced in thyroid carcinomas by primarily unknown mechanisms. In several tissues, SNAIL (SNAI1) and SLUG (SNAI2) induce epithelial-mesenchymal transition by altering target gene transcription, including CDH1 repression, but these transcription factors have not been studied in thyroid carcinoma. Recently, our group has provided direct evidence that ectopic SNAI1 expression induces epithelial and mesenchymal mouse tumors. SNAI1, SNAI2, and CDH1 expression were analyzed in thyroid-derived cell lines and samples of human follicular and papillary thyroid carcinoma by reverse transcriptase-polymerase chain reaction, Western blotting, and immunohistochemistry. The effect of SNAI1 expression on CDH1 transcription was analyzed by reverse transcriptase-polymerase chain reaction and Western blotting in ori-3 cells. Thyroid carcinoma development was analyzed in CombitTA-Snail mice, in which SNAI1 levels are up-regulated. SNAI1 and SNAI2 were not expressed in cells derived from normal thyroid tissue, or in normal human thyroid samples, but were highly expressed in cell lines derived from thyroid carcinomas, in human thyroid carcinoma samples, and their metastases. SNAI1 expression in ori-3 cells repressed CDH1 transcription. Combi-TA mice developed papillary thyroid carcinomas, the incidence of which was increased by concomitant radiotherapy. In conclusion, SNAI1 and SNAI2 are ectopically expressed in thyroid carcinomas, and aberrant expression in mice is associated with papillary carcinoma development. PMID:17724139

  18. Ultrasound appearance of the normal Lisfranc ligament.

    PubMed

    Kaicker, Jatin; Zajac, Mercedes; Shergill, Ravi; Choudur, Hema N

    2016-12-01

    This study aims to prospectively evaluate the ultrasound appearance of the normal Lisfranc's ligament in 50 patients (100 ft) with no prior or current ligament injury. Fifty normal asymptomatic patients between the ages of 18 and 80 years were assessed. Three key features were recorded: ultrasound appearance, thickness, and length of the Lisfranc's ligament. Patients excluded from this study included pediatric patients and those with history of injury or symptoms related to the foot. The mean right- and left-sided ligament (RT) thickness were 0.096 (0.021) and 0.104 (0.023), respectively. The mean right- and left-sided ligament RT length was 0.54 (0.11) and 0.57 (0.11), respectively. The appearance of the ligament was similar in all patients with a central thin band of hypoechogenicity lined by hyperechoic lines on either side. Understanding the normal appearance, thickness, and length of the Lisfranc's ligament in a large sample is imperative to diagnose abnormal appearances of this ligament including sprains and tears by ultrasound. Ultrasound, with its easy accessibility, can be used in the emergency department to rapidly exclude injury of the ligament. Increased understanding and awareness of the Lisfranc's ligament on ultrasound can allow for more accurate diagnosis and treatment.

  19. ATF2, a paradigm of the multifaceted regulation of transcription factors in biology and disease.

    PubMed

    Watson, Gregory; Ronai, Ze'ev; Lau, Eric

    2017-02-15

    Stringent transcriptional regulation is crucial for normal cellular biology and organismal development. Perturbations in the proper regulation of transcription factors can result in numerous pathologies, including cancer. Thus, understanding how transcription factors are regulated and how they are dysregulated in disease states is key to the therapeutic targeting of these factors and/or the pathways that they regulate. Activating transcription factor 2 (ATF2) has been studied in a number of developmental and pathological conditions. Recent findings have shed light on the transcriptional, post-transcriptional, and post-translational regulatory mechanisms that influence ATF2 function, and thus, the transcriptional programs coordinated by ATF2. Given our current knowledge of its multiple levels of regulation and function, ATF2 represents a paradigm for the mechanistic complexity that can regulate transcription factor function. Thus, increasing our understanding of the regulation and function of ATF2 will provide insights into fundamental regulatory mechanisms that influence how cells integrate extracellular and intracellular signals into a genomic response through transcription factors. Characterization of ATF2 dysfunction in the context of pathological conditions, particularly in cancer biology and response to therapy, will be important in understanding how pathways controlled by ATF2 or other transcription factors might be therapeutically exploited. In this review, we provide an overview of the currently known upstream regulators and downstream targets of ATF2.

  20. Screening with MRI for Accurate and Rapid Stroke Treatment

    PubMed Central

    Shah, Shreyansh; Luby, Marie; Poole, Karen; Morella, Teresa; Keller, Elizabeth; Benson, Richard T.; Lynch, John K.; Nadareishvili, Zurab

    2015-01-01

    Objective: The objective of this study was to demonstrate the feasibility of timely multimodal MRI screening before thrombolysis in acute stroke patients. Methods: Quality improvement processes were initiated in 2013 to reduce door-to-needle (DTN) time at the 2 hospitals where the NIH stroke team provides clinical care. Acute ischemic stroke (AIS) patients who received IV tissue plasminogen activator (tPA) ≤4.5 hours from last known normal were identified. Demographic and clinical characteristics and timing metrics were analyzed comparing the time periods before, during, and after the quality improvement processes. Results: There were 157 patients treated with IV tPA for AIS during 2012–2013, of whom 135 (86%) were screened with MRI. DTN time was significantly reduced by 40% during this period from a median of 93 minutes in the first half of 2012 to 55 minutes in the last half of 2013 (p < 0.0001) with a significant 4-fold increase in the proportion of treated patients with DTN time ≤60 minutes from 13.0% to 61.5%, respectively (p < 0.00001). Improvement in DTN time was associated with reduced door-to-MRI time, and there were no differences in demographic or clinical characteristics (p = 0.21–0.76). Conclusions: It is feasible and practical to consistently and rapidly deliver IV tPA to AIS patients within national benchmark times using MRI as the routine screening modality. The processes used in the SMART (Screening with MRI for Accurate and Rapid Stroke Treatment) Study to reduce DTN time have the potential to be widely applicable to other hospitals. PMID:25972494

  1. Robust, accurate and fast automatic segmentation of the spinal cord.

    PubMed

    De Leener, Benjamin; Kadoury, Samuel; Cohen-Adad, Julien

    2014-09-01

    Spinal cord segmentation provides measures of atrophy and facilitates group analysis via inter-subject correspondence. Automatizing this procedure enables studies with large throughput and minimizes user bias. Although several automatic segmentation methods exist, they are often restricted in terms of image contrast and field-of-view. This paper presents a new automatic segmentation method (PropSeg) optimized for robustness, accuracy and speed. The algorithm is based on the propagation of a deformable model and is divided into three parts: firstly, an initialization step detects the spinal cord position and orientation using a circular Hough transform on multiple axial slices rostral and caudal to the starting plane and builds an initial elliptical tubular mesh. Secondly, a low-resolution deformable model is propagated along the spinal cord. To deal with highly variable contrast levels between the spinal cord and the cerebrospinal fluid, the deformation is coupled with a local contrast-to-noise adaptation at each iteration. Thirdly, a refinement process and a global deformation are applied on the propagated mesh to provide an accurate segmentation of the spinal cord. Validation was performed in 15 healthy subjects and two patients with spinal cord injury, using T1- and T2-weighted images of the entire spinal cord and on multiecho T2*-weighted images. Our method was compared against manual segmentation and against an active surface method. Results show high precision for all the MR sequences. Dice coefficients were 0.9 for the T1- and T2-weighted cohorts and 0.86 for the T2*-weighted images. The proposed method runs in less than 1min on a normal computer and can be used to quantify morphological features such as cross-sectional area along the whole spinal cord.

  2. Transcriptional Signatures in Huntington's Disease

    PubMed Central

    2007-01-01

    While selective neuronal death has been an influential theme in Huntington's disease (HD), there is now a preponderance of evidence that significant neuronal dysfunction precedes frank neuronal death. The best evidence for neuronal dysfunction is the observation that gene expression is altered in HD brain, suggesting that transcriptional dysregulation is a central mechanism. Studies of altered gene expression began with careful observations of post-mortem human HD brain and subsequently were accelerated by the development of transgenic mouse models. The application of DNA microarray technology has spurred tremendous progress with respect to the altered transcriptional processes that occur in HD, through gene expression studies of both transgenic mouse models as well as cellular models of HD. Gene expression profiles are remarkably comparable across these models, bolstering the idea that transcriptional signatures reflect an essential feature of disease pathogenesis. Finally, gene expression studies have been applied to human HD, thus not only validating the approach of using model systems, but also solidifying the idea that altered transcription is a key mechanism in HD pathogenesis. In the future, gene expression profiling will be used as a readout in clinical trials aimed at correcting transcriptional dysregulation in Huntington's disease. PMID:17467140

  3. Kinetic Modelling of Transcription Elongation

    NASA Astrophysics Data System (ADS)

    O'Maoileidigh, Daibhid; Tadigotla, Vasisht; Sengupta, Anirvan; Epshtein, Vitaly; Ebright, Richard; Nudler, Evgeny; Ruckenstein, Andrei

    2006-03-01

    Transcription is the first step in gene expression and it is at this stage that most of genetic regulation occurs. The enzyme RNA polymerase (RNAP) walks along DNA creating an RNA transcript at a highly non-uniform rate. We discuss how many non-intuitive features of the system may be experimentally and physically motivated and present first a model, which agrees qualitatively with a host of experimental evidence. We also examine intrinsic pauses where it is thought that the RNAP will move backwards along the DNA template without changing the length of the RNA transcript. We describe a simplified kinetic scheme for the recovery of intrinsic pauses with the same degree of predictive power as our thermodynamic model (presented separately). The separation of timescales between the movement of the RNAP and global changes in the RNA secondary structure is seen to be crucial for the function of RNAP. This is essentially a model of a Brownian ratchet where RNAP executes a 1D random walk in a sequence dependent potential over a range determined by the co-transcriptional RNA fold for each transcript length

  4. Transcriptional gene silencing in humans

    PubMed Central

    Weinberg, Marc S.; Morris, Kevin V.

    2016-01-01

    It has been over a decade since the first observation that small non-coding RNAs can functionally modulate epigenetic states in human cells to achieve functional transcriptional gene silencing (TGS). TGS is mechanistically distinct from the RNA interference (RNAi) gene-silencing pathway. TGS can result in long-term stable epigenetic modifications to gene expression that can be passed on to daughter cells during cell division, whereas RNAi does not. Early studies of TGS have been largely overlooked, overshadowed by subsequent discoveries of small RNA-directed post-TGS and RNAi. A reappraisal of early work has been brought about by recent findings in human cells where endogenous long non-coding RNAs function to regulate the epigenome. There are distinct and common overlaps between the proteins involved in small and long non-coding RNA transcriptional regulatory mechanisms, suggesting that the early studies using small non-coding RNAs to modulate transcription were making use of a previously unrecognized endogenous mechanism of RNA-directed gene regulation. Here we review how non-coding RNA plays a role in regulation of transcription and epigenetic gene silencing in human cells by revisiting these earlier studies and the mechanistic insights gained to date. We also provide a list of mammalian genes that have been shown to be transcriptionally regulated by non-coding RNAs. Lastly, we explore how TGS may serve as the basis for development of future therapeutic agents. PMID:27060137

  5. A Robust Method for Transcript Quantification with RNA-Seq Data

    PubMed Central

    Huang, Yan; Hu, Yin; Jones, Corbin D.; MacLeod, James N.; Chiang, Derek Y.; Liu, Yufeng; Prins, Jan F.

    2013-01-01

    Abstract The advent of high throughput RNA-seq technology allows deep sampling of the transcriptome, making it possible to characterize both the diversity and the abundance of transcript isoforms. Accurate abundance estimation or transcript quantification of isoforms is critical for downstream differential analysis (e.g., healthy vs. diseased cells) but remains a challenging problem for several reasons. First, while various types of algorithms have been developed for abundance estimation, short reads often do not uniquely identify the transcript isoforms from which they were sampled. As a result, the quantification problem may not be identifiable, i.e., lacks a unique transcript solution even if the read maps uniquely to the reference genome. In this article, we develop a general linear model for transcript quantification that leverages reads spanning multiple splice junctions to ameliorate identifiability. Second, RNA-seq reads sampled from the transcriptome exhibit unknown position-specific and sequence-specific biases. We extend our method to simultaneously learn bias parameters during transcript quantification to improve accuracy. Third, transcript quantification is often provided with a candidate set of isoforms, not all of which are likely to be significantly expressed in a given tissue type or condition. By resolving the linear system with LASSO, our approach can infer an accurate set of dominantly expressed transcripts while existing methods tend to assign positive expression to every candidate isoform. Using simulated RNA-seq datasets, our method demonstrated better quantification accuracy and the inference of dominant set of transcripts than existing methods. The application of our method on real data experimentally demonstrated that transcript quantification is effective for differential analysis of transcriptomes. PMID:23461570

  6. 46 CFR 502.165 - Official transcript.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... incremental cost of transcription above the regular copy transcription cost borne by the Commission, in... full cost of transcription being borne by the Commission. (B) In the event a request for daily copy is... of transcription over and above that borne by the Commission, i.e., the incremental cost between...

  7. 46 CFR 502.165 - Official transcript.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... incremental cost of transcription above the regular copy transcription cost borne by the Commission, in... full cost of transcription being borne by the Commission. (B) In the event a request for daily copy is... of transcription over and above that borne by the Commission, i.e., the incremental cost between...

  8. 46 CFR 502.165 - Official transcript.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... incremental cost of transcription above the regular copy transcription cost borne by the Commission, in... full cost of transcription being borne by the Commission. (B) In the event a request for daily copy is... of transcription over and above that borne by the Commission, i.e., the incremental cost between...

  9. 46 CFR 502.165 - Official transcript.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... incremental cost of transcription above the regular copy transcription cost borne by the Commission, in... full cost of transcription being borne by the Commission. (B) In the event a request for daily copy is... of transcription over and above that borne by the Commission, i.e., the incremental cost between...

  10. 46 CFR 502.165 - Official transcript.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... incremental cost of transcription above the regular copy transcription cost borne by the Commission, in... full cost of transcription being borne by the Commission. (B) In the event a request for daily copy is... of transcription over and above that borne by the Commission, i.e., the incremental cost between...

  11. Transcription and splicing: when the twain meet.

    PubMed

    Brody, Yehuda; Shav-Tal, Yaron

    2011-01-01

    Splicing can occur co-transcriptionally. What happens when the splicing reaction lags after the completed transcriptional process? We found that elongation rates are independent of ongoing splicing on the examined genes and suggest that when transcription has completed but splicing has not, the splicing machinery is retained at the site of transcription, independently of the polymerase.

  12. Pervasive transcription: detecting functional RNAs in bacteria.

    PubMed

    Lybecker, Meghan; Bilusic, Ivana; Raghavan, Rahul

    2014-01-01

    Pervasive, or genome-wide, transcription has been reported in all domains of life. In bacteria, most pervasive transcription occurs antisense to protein-coding transcripts, although recently a new class of pervasive RNAs was identified that originates from within annotated genes. Initially considered to be non-functional transcriptional noise, pervasive transcription is increasingly being recognized as important in regulating gene expression. The function of pervasive transcription is an extensively debated question in the field of transcriptomics and regulatory RNA biology. Here, we highlight the most recent contributions addressing the purpose of pervasive transcription in bacteria and discuss their implications.

  13. Longitudinal evaluation of leukocyte transcripts in killer whales (Orcinus Orca)

    USGS Publications Warehouse

    Sitt, Tatjana; Bowen, Lizabeth; Lee, Chia-Shan; Blanchard, Myra; McBain, James; Dold, Christopher; Stott, Jeffrey L.

    2016-01-01

    Early identification of illness and/or presence of environmental and/or social stressors in free-ranging and domestic cetaceans is a priority for marine mammal health care professionals. Incorporation of leukocyte gene transcript analysis into the diagnostic tool kit has the potential to augment classical diagnostics based upon ease of sample storage and shipment, inducible nature and well-defined roles of transcription and associated downstream actions. Development of biomarkers that could serve to identify “insults” and potentially differentiate disease etiology would be of great diagnostic value. To this end, a modest number of peripheral blood leukocyte gene transcripts were selected for application to a domestic killer whale population with a focus on broad representation of inducible immunologically relevant genes. Normalized leukocyte transcript values, longitudinally acquired from 232 blood samples derived from 26 clinically healthy whales, were not visibly influenced temporally nor by sex or the specific Park in which they resided. Stability in leukocyte transcript number during periods of health enhances their potential use in diagnostics through identification of outliers. Transcript levels of two cytokine genes, IL-4 and IL-17, were highly variable within the group as compared to the other transcripts. IL-4 transcripts were typically absent. Analysis of transcript levels on the other genes of interest, on an individual animal basis, identified more outliers than were visible when analyzed in the context of the entire population. The majority of outliers (9 samples) were low, though elevated transcripts were identified for IL-17 from 2 animals and one each for Cox-2 and IL-10. The low number of outliers was not unexpected as sample selection was intentionally directed towards animals that were clinically healthy at the time of collection. Outliers may reflect animals experiencing subclinical disease that is transient and self-limiting. The

  14. Longitudinal evaluation of leukocyte transcripts in killer whales (Orcinus Orca).

    PubMed

    Sitt, Tatjana; Bowen, Lizabeth; Lee, Chia-Shan; Blanchard, Myra T; McBain, James; Dold, Christopher; Stott, Jeffrey L

    2016-07-01

    Early identification of illness and/or presence of environmental and/or social stressors in free-ranging and domestic cetaceans is a priority for marine mammal health care professionals. Incorporation of leukocyte gene transcript analysis into the diagnostic tool kit has the potential to augment classical diagnostics based upon ease of sample storage and shipment, inducible nature and well-defined roles of transcription and associated downstream actions. Development of biomarkers that could serve to identify "insults" and potentially differentiate disease etiology would be of great diagnostic value. To this end, a modest number of peripheral blood leukocyte gene transcripts were selected for application to a domestic killer whale population with a focus on broad representation of inducible immunologically relevant genes. Normalized leukocyte transcript values, longitudinally acquired from 232 blood samples derived from 26 clinically healthy whales, were not visibly influenced temporally nor by sex or the specific Park in which they resided. Stability in leukocyte transcript number during periods of health enhances their potential use in diagnostics through identification of outliers. Transcript levels of two cytokine genes, IL-4 and IL-17, were highly variable within the group as compared to the other transcripts. IL-4 transcripts were typically absent. Analysis of transcript levels on the other genes of interest, on an individual animal basis, identified more outliers than were visible when analyzed in the context of the entire population. The majority of outliers (9 samples) were low, though elevated transcripts were identified for IL-17 from 2 animals and one each for Cox-2 and IL-10. The low number of outliers was not unexpected as sample selection was intentionally directed towards animals that were clinically healthy at the time of collection. Outliers may reflect animals experiencing subclinical disease that is transient and self-limiting. The immunologic

  15. Subventricular zone microglia transcriptional networks.

    PubMed

    Starossom, Sarah C; Imitola, Jaime; Wang, Yue; Cao, Li; Khoury, Samia J

    2011-07-01

    Microglia play an important role in inflammatory diseases of the central nervous system. There is evidence of microglial diversity with distinct phenotypes exhibiting either neuroprotection and repair or neurotoxicity. However the precise molecular mechanisms underlying this diversity are still unknown. Using a model of experimental autoimmune encephalomyelitis (EAE) we performed transcriptional profiling of isolated subventricular zone microglia from the acute and chronic disease phases of EAE. We found that microglia exhibit disease phase specific gene expression signatures, that correspond to unique gene ontology functions and genomic networks. Our data demonstrate for the first time, distinct transcriptional networks of microglia activation in vivo, that suggests a role as mediators of injury or repair.

  16. Chromatin and Transcription in Yeast

    PubMed Central

    Rando, Oliver J.; Winston, Fred

    2012-01-01

    Understanding the mechanisms by which chromatin structure controls eukaryotic transcription has been an intense area of investigation for the past 25 years. Many of the key discoveries that created the foundation for this field came from studies of Saccharomyces cerevisiae, including the discovery of the role of chromatin in transcriptional silencing, as well as the discovery of chromatin-remodeling factors and histone modification activities. Since that time, studies in yeast have continued to contribute in leading ways. This review article summarizes the large body of yeast studies in this field. PMID:22345607

  17. Uncovering ancient transcription systems with a novel evolutionary indicator

    PubMed Central

    Adachi, Naruhiko; Senda, Toshiya; Horikoshi, Masami

    2016-01-01

    TBP and TFIIB are evolutionarily conserved transcription initiation factors in archaea and eukaryotes. Information about their ancestral genes would be expected to provide insight into the origin of the RNA polymerase II-type transcription apparatus. In obtaining such information, the nucleotide sequences of current genes of both archaea and eukaryotes should be included in the analysis. However, the present methods of evolutionary analysis require that a subset of the genes should be excluded as an outer group. To overcome this limitation, we propose an innovative concept for evolutionary analysis that does not require an outer group. This approach utilizes the similarity in intramolecular direct repeats present in TBP and TFIIB as an evolutionary measure revealing the degree of similarity between the present offspring genes and their ancestors. Information on the properties of the ancestors and the order of emergence of TBP and TFIIB was also revealed. These findings imply that, for evolutionarily early transcription systems billions of years ago, interaction of RNA polymerase II with transcription initiation factors and the regulation of its enzymatic activity was required prior to the accurate positioning of the enzyme. Our approach provides a new way to discuss mechanistic and system evolution in a quantitative manner. PMID:27307191

  18. Nuclear mRNA quality control in yeast is mediated by Nrd1 co-transcriptional recruitment, as revealed by the targeting of Rho-induced aberrant transcripts

    PubMed Central

    Honorine, Romy; Mosrin-Huaman, Christine; Hervouet-Coste, Nadège; Libri, Domenico; Rahmouni, A. Rachid

    2011-01-01

    The production of mature export-competent transcripts is under the surveillance of quality control steps where aberrant mRNP molecules resulting from inappropriate or inefficient processing and packaging reactions are subject to exosome-mediated degradation. Previously, we have shown that the heterologous expression of bacterial Rho factor in yeast interferes in normal mRNP biogenesis leading to the production of full-length yet aberrant transcripts that are degraded by the nuclear exosome with ensuing growth defect. Here, we took advantage of this new tool to investigate the molecular mechanisms by which an integrated system recognizes aberrancies at each step of mRNP biogenesis and targets the defective molecules for destruction. We show that the targeting and degradation of Rho-induced aberrant transcripts is associated with a large increase of Nrd1 recruitment to the transcription complex via its CID and RRM domains and a concomitant enrichment of exosome component Rrp6 association. The targeting and degradation of the aberrant transcripts is suppressed by the overproduction of Pcf11 or its isolated CID domain, through a competition with Nrd1 for recruitment by the transcription complex. Altogether, our results support a model in which a stimulation of Nrd1 co-transcriptional recruitment coordinates the recognition and removal of aberrant transcripts by promoting the attachment of the nuclear mRNA degradation machinery. PMID:21113025

  19. Transcriptional map of chromosome region 6q16-->q21.

    PubMed

    Karayianni, E; Magnanini, C; Orphanos, V; Negrini, M; Maniatis, G M; Spathas, D H; Barbanti-Brodano, G; Morelli, C

    1999-01-01

    We present the transcription map of chromosome region 6q16-->q21 by mapping fifteen known genes within this region. Five genes lay in the subregion containing a tumor suppressor gene, eight genes are located in the subregion harboring a senescence gene, and two genes are distal to the latter region. The precise location of the genes was obtained using a previously described translocation and deletion mouse/human hybrid panel. An even more accurate definition was possible for the genes spanning the senescence gene region, since a previously described YAC contig with its restriction map was available. From this transcription map it is possible to derive a large region of synteny with mouse chromosome 10.

  20. Development of an efficient in vitro transcription system for bloodstream form Trypanosoma brucei reveals life cycle-independent functionality of class I transcription factor A.

    PubMed

    Park, Sung Hee; Nguyen, Tu N; Günzl, Arthur

    2012-01-01

    Trypanosomatid parasites possess extremely divergent transcription factors whose identification typically relied on biochemical, structural and functional analyses because they could not be identified by standard sequence analysis. For example, subunits of the Trypanosoma brucei mediator and class I transcription factor A (CITFA) have no sequence resemblance to putative counterparts in higher eukaryotes. Therefore, homologous in vitro transcription systems have been crucial in evaluating the transcriptional roles of T. brucei proteins but so far such systems have been restricted to the insect-stage, procyclic form (PF) of the parasite. Here, we report the development of a homologous system for the mammalian-infective, bloodstream form (BF) of T. brucei which supports accurately initiated transcription from three different RNA polymerase (pol) I promoters as well as from the RNA pol II-recruiting spliced leader RNA gene promoter. The system is based on a small scale extract preparation procedure which accommodates the low cell densities obtainable in BF culture. BF and PF systems behave surprisingly similar and we show that the CITFA complex purified from procyclic extract is fully functional in the BF system indicating that the transcriptional machinery in general is equivalent in both life cycle stages. A notable difference, however, was observed with the RNA pol I-recruiting GPEET procyclin promoter whose reduced promoter strength and increased sensitivity to manganese ions in the BF system suggests the presence of a specific transcriptional activator in the PF system.

  1. Transcription factor-based biosensor

    DOEpatents

    Dietrich, Jeffrey A; Keasling, Jay D

    2013-10-08

    The present invention provides for a system comprising a BmoR transcription factor, a .sigma..sup.54-RNA polymerase, and a pBMO promoter operatively linked to a reporter gene, wherein the pBMO promoter is capable of expression of the reporter gene with an activated form of the BmoR and the .sigma..sup.54-RNA polymerase.

  2. Transcriptional networks in plant immunity.

    PubMed

    Tsuda, Kenichi; Somssich, Imre E

    2015-05-01

    Next to numerous abiotic stresses, plants are constantly exposed to a variety of pathogens within their environment. Thus, their ability to survive and prosper during the course of evolution was strongly dependent on adapting efficient strategies to perceive and to respond to such potential threats. It is therefore not surprising that modern plants have a highly sophisticated immune repertoire consisting of diverse signal perception and intracellular signaling pathways. This signaling network is intricate and deeply interconnected, probably reflecting the diverse lifestyles and infection strategies used by the multitude of invading phytopathogens. Moreover it allows signal communication between developmental and defense programs thereby ensuring that plant growth and fitness are not significantly retarded. How plants integrate and prioritize the incoming signals and how this information is transduced to enable appropriate immune responses is currently a major research area. An important finding has been that pathogen-triggered cellular responses involve massive transcriptional reprogramming within the host. Additional key observations emerging from such studies are that transcription factors (TFs) are often sites of signal convergence and that signal-regulated TFs act in concert with other context-specific TFs and transcriptional co-regulators to establish sensory transcription regulatory networks required for plant immunity.

  3. Regulating transcription traffic around DSBs.

    PubMed

    Plosky, Brian S

    2015-05-07

    If a double-strand break (DSB) occurs and either a DNA polymerase or RNA polymerase is coming along, how do we save the train? In this issue of Molecular Cell, Ui et al. (2015) describe a connection between an elongation factor and a repressive complex to prevent transcription in proximity to a DSB.

  4. Transcription factors in alkaloid biosynthesis.

    PubMed

    Yamada, Yasuyuki; Sato, Fumihiko

    2013-01-01

    Higher plants produce a large variety of low-molecular weight secondary compounds. Among them, nitrogen-containing alkaloids are the most biologically active and are often used pharmaceutically. Whereas alkaloid chemistry has been intensively investigated, alkaloid biosynthesis, including the relevant biosynthetic enzymes, genes and their regulation, and especially transcription factors, is largely unknown, as only a limited number of plant species produce certain types of alkaloids and they are difficult to study. Recently, however, several groups have succeeded in isolating the transcription factors that are involved in the biosynthesis of several types of alkaloids, including bHLH, ERF, and WRKY. Most of them show Jasmonate (JA) responsiveness, which suggests that the JA signaling cascade plays an important role in alkaloid biosynthesis. Here, we summarize the types and functions of transcription factors that have been isolated in alkaloid biosynthesis, and characterize their similarities and differences compared to those in other secondary metabolite pathways, such as phenylpropanoid and terpenoid biosyntheses. The evolution of this biosynthetic pathway and regulatory network, as well as the application of these transcription factors to metabolic engineering, is discussed.

  5. Normal matter storage of antiprotons

    SciTech Connect

    Campbell, L.J.

    1987-01-01

    Various simple issues connected with the possible storage of anti p in relative proximity to normal matter are discussed. Although equilibrium storage looks to be impossible, condensed matter systems are sufficiently rich and controllable that nonequilibrium storage is well worth pursuing. Experiments to elucidate the anti p interactions with normal matter are suggested. 32 refs.

  6. Normalizing Catastrophe: An Educational Response

    ERIC Educational Resources Information Center

    Jickling, Bob

    2013-01-01

    Processes of normalizing assumptions and values have been the subjects of theoretical framing and critique for several decades now. Critique has often been tied to issues of environmental sustainability and social justice. Now, in an era of global warming, there is a rising concern that the results of normalizing of present values could be…

  7. A Simple yet Accurate Method for Students to Determine Asteroid Rotation Periods from Fragmented Light Curve Data

    ERIC Educational Resources Information Center

    Beare, R. A.

    2008-01-01

    Professional astronomers use specialized software not normally available to students to determine the rotation periods of asteroids from fragmented light curve data. This paper describes a simple yet accurate method based on Microsoft Excel[R] that enables students to find periods in asteroid light curve and other discontinuous time series data of…

  8. Synaptonemal Complex Protein 3 Transcript Analysis in Breast Cancer

    PubMed Central

    MOBASHERI, Maryam Beigom; SHIRKOOHI, Reza; MODARRESSI, Mohammad Hossein

    2016-01-01

    Background: Breast cancer is the most frequent cancer in women. Cancer/Testis antigens are immunogenic proteins ectopically expressed in human neoplasms. Synaptonemal complex protein 3 (SYCP3) belongs to cancer/testis genes family involved in meiotic events and spermatogenesis. The aim of this study was to express analysis of SYCP3 in breast cancer and validate it as a breast cancer biomarker. Methods: Expression of SYCP3 transcripts in 47 breast tumors, 6 breast cancer cell lines (MCF7, SKBR3, T47D, BT474, MDA-MB-231 and MDA-MB 468), 5 normal breast and 2 testis tissues was studied by Real Time RT-PCR reaction. The reference genes phosphoglucomutase 1 and hypoxanthine guanine phosphoribosyl transferase were used as reactions normalizers. The software tool REST 2009 was applied for statistical analysis of the data. The research was conducted from Apr 2014 to August 2015 in Faculty of Medicine, Tehran University of Medical Sciences, Tehran, Iran. Results: All of the studied breast cancer cell lines showed very high levels of SYCP3 overexpression in comparison to normal breast (P=0.001) and even to normal testis (P=0.001), except for MCF7 cell line. Breast tumors showed moderately increasing in transcript changes in comparison to normal breast. Conclusion: SYCP3 is a known testis-specific gene, but interestingly five out of six studied breast cancer of cell lines showed higher expression levels of SYCP3 in comparison to normal testis and normal breast tissues. SYCP3 has critical role in cell division with known interaction with the tumor suppressor genes, BRCA1 and BRCA2, which are critical genes in breast cancer. PMID:28053928

  9. A predictable and accurate technique with elastomeric impression materials.

    PubMed

    Barghi, N; Ontiveros, J C

    1999-08-01

    A method for obtaining more predictable and accurate final impressions with polyvinylsiloxane impression materials in conjunction with stock trays is proposed and tested. Heavy impression material is used in advance for construction of a modified custom tray, while extra-light material is used for obtaining a more accurate final impression.

  10. Tube dimpling tool assures accurate dip-brazed joints

    NASA Technical Reports Server (NTRS)

    Beuyukian, C. S.; Heisman, R. M.

    1968-01-01

    Portable, hand-held dimpling tool assures accurate brazed joints between tubes of different diameters. Prior to brazing, the tool performs precise dimpling and nipple forming and also provides control and accurate measuring of the height of nipples and depth of dimples so formed.

  11. Managing incontinence: women's normalizing strategies.

    PubMed

    Skoner, M M; Haylor, M J

    1993-01-01

    Women's strategies for managing urinary incontinence were examined in a grounded-theory study. The women's basic social concern was dealing with incontinence in a manner that enabled them to feel normal. Feeling normal meant being able to do what they wanted to do and needed to do to have a normal life-style as they perceived it. This goal was accomplished by normalizing incontinence and its management. Normalization was achieved by directing its course through self-management, accounting for it in terms of personal history and life experiences, and delaying medical counsel. These strategies are described. The findings provide fresh insights about women's response to incontinence and their practice of self-managing its consequences.

  12. Voice recognition software versus a traditional transcription service for physician charting in the ED.

    PubMed

    Zick, R G; Olsen, J

    2001-07-01

    This study was conducted to compare real-time voice recognition software to a traditional transcription service. Two emergency department (ED) physicians dictated 47 charts using a voice dictation software program and a traditional transcription service. Accuracy, word per minute dictation time and turnaround time were calculated from the data. The transcription service used in our study was more accurate than the voice recognition program with an accuracy of 99.7 percent versus 98.5 percent for the voice recognition program. The average number of corrections per chart was 2.5 for the voice recognition program and 1.2 for the traditional transcription service. Turnaround time was much better using the computer voice recognition program with an average turnaround time of 3.65 minutes versus a turnaround time of 39.6 minutes for the traditionally transcribed charts. The charts dictated using the voice recognition program were considerably less costly than the manually transcribed charts. In summary, computer voice recognition is nearly as accurate as traditional transcription, it has a much shorter turnaround time and is less expensive than traditional transcription. We recommend its use as a tool for physician charting in the ED.

  13. Investigating transcription reinitiation through in vitro approaches.

    PubMed

    Dieci, Giorgio; Fermi, Beatrice; Bosio, Maria Cristina

    2014-01-01

    By influencing the number of RNA molecules repeatedly synthesized from the same gene, the control of transcription reinitiation has the potential to shape the transcriptome. Transcription reinitiation mechanisms have been mainly addressed in vitro, through approaches based on both crude and reconstituted systems. These studies support the notion that transcription reinitiation and its regulation rely on dedicated networks of molecular interactions within transcription machineries. At the same time, comparison with in vivo transcription rates suggests that additional mechanisms, factors and conditions must exist in the nucleus, whose biochemical elucidation is a fascinating challenge for future in vitro transcription studies.

  14. RNA polymerase II transcription: structure and mechanism.

    PubMed

    Liu, Xin; Bushnell, David A; Kornberg, Roger D

    2013-01-01

    A minimal RNA polymerase II (pol II) transcription system comprises the polymerase and five general transcription factors (GTFs) TFIIB, -D, -E, -F, and -H. The addition of Mediator enables a response to regulatory factors. The GTFs are required for promoter recognition and the initiation of transcription. Following initiation, pol II alone is capable of RNA transcript elongation and of proofreading. Structural studies reviewed here reveal roles of GTFs in the initiation process and shed light on the transcription elongation mechanism. This article is part of a Special Issue entitled: RNA Polymerase II Transcript Elongation.

  15. Fast and accurate line scanner based on white light interferometry

    NASA Astrophysics Data System (ADS)

    Lambelet, Patrick; Moosburger, Rudolf

    2013-04-01

    White-light interferometry is a highly accurate technology for 3D measurements. The principle is widely utilized in surface metrology instruments but rarely adopted for in-line inspection systems. The main challenges for rolling out inspection systems based on white-light interferometry to the production floor are its sensitivity to environmental vibrations and relatively long measurement times: a large quantity of data needs to be acquired and processed in order to obtain a single topographic measurement. Heliotis developed a smart-pixel CMOS camera (lock-in camera) which is specially suited for white-light interferometry. The demodulation of the interference signal is treated at the level of the pixel which typically reduces the acquisition data by one orders of magnitude. Along with the high bandwidth of the dedicated lock-in camera, vertical scan-speeds of more than 40mm/s are reachable. The high scan speed allows for the realization of inspection systems that are rugged against external vibrations as present on the production floor. For many industrial applications such as the inspection of wafer-bumps, surface of mechanical parts and solar-panel, large areas need to be measured. In this case either the instrument or the sample are displaced laterally and several measurements are stitched together. The cycle time of such a system is mostly limited by the stepping time for multiple lateral displacements. A line-scanner based on white light interferometry would eliminate most of the stepping time while maintaining robustness and accuracy. A. Olszak proposed a simple geometry to realize such a lateral scanning interferometer. We demonstrate that such inclined interferometers can benefit significantly from the fast in-pixel demodulation capabilities of the lock-in camera. One drawback of an inclined observation perspective is that its application is limited to objects with scattering surfaces. We therefore propose an alternate geometry where the incident light is

  16. PyVCI: A flexible open-source code for calculating accurate molecular infrared spectra

    NASA Astrophysics Data System (ADS)

    Sibaev, Marat; Crittenden, Deborah L.

    2016-06-01

    The PyVCI program package is a general purpose open-source code for simulating accurate molecular spectra, based upon force field expansions of the potential energy surface in normal mode coordinates. It includes harmonic normal coordinate analysis and vibrational configuration interaction (VCI) algorithms, implemented primarily in Python for accessibility but with time-consuming routines written in C. Coriolis coupling terms may be optionally included in the vibrational Hamiltonian. Non-negligible VCI matrix elements are stored in sparse matrix format to alleviate the diagonalization problem. CPU and memory requirements may be further controlled by algorithmic choices and/or numerical screening procedures, and recommended values are established by benchmarking using a test set of 44 molecules for which accurate analytical potential energy surfaces are available. Force fields in normal mode coordinates are obtained from the PyPES library of high quality analytical potential energy surfaces (to 6th order) or by numerical differentiation of analytic second derivatives generated using the GAMESS quantum chemical program package (to 4th order).

  17. Treatment of experimental colitis in mice with LMP-420, an inhibitor of TNF transcription

    PubMed Central

    Hale, Laura P; Cianciolo, George

    2008-01-01

    Background LMP-420 is a boronic acid-containing purine nucleoside analogue that transcriptionally inhibits TNF production but is non-cytotoxic to TNF-producing cells. Methods This study investigated the efficacy of LMP-420 as an anti-inflammatory agent in acute and chronic colitis induced by oral administration of dextran sulfate sodium (DSS) to mice and in chronic colitis following piroxicam administration to IL-10-deficient mice. The severity of colon inflammation was assessed histologically. TNF levels were measured by enzyme immunoassay. Results Administration of DSS for 7 days resulted in severe acute colitis that was associated with a marked increase in stool and colon tissue TNF levels. Initiation of therapy with intraperitoneal (i.p.) LMP-420 on day 4 of DSS exposure decreased colonic TNF to near normal levels on day 7. However, neither i.p. nor oral treatment with LMP-420 affected the development or severity of acute DSS colitis. Initiation of LMP-420 therapy after 3 cycles of DSS administration to establish chronic colitis also had no effect on the severity of chronic colitis. Analysis of colonic TNF combined with longitudinal analysis of TNF and TNF receptor (TNF-RII) levels in stool during the development of chronic DSS colitis demonstrated that the initially elevated colonic TNF levels returned to normal despite intense on-going inflammation in mice with chronic colitis. RAG-2-/- mice deficient in T and B cells also developed severe ongoing colitis in response to 3 cycles of DSS, but showed marked differences vs. wild type mice in stool TNF and TNF-RII in response to DSS exposure. Systemic and oral LMP-420 treatment for 16 days decreased colonic TNF levels in IL-10-deficient mice with chronic colitis, with a trend to decreased histologic inflammation for oral LMP-420. Conclusion These studies demonstrate that short-term treatment with a transcriptional inhibitor of TNF production can decrease systemic and local colonic levels of TNF but may not

  18. Reproduction of Hip Offset and Leg Length in Navigated Total Hip Arthroplasty: How Accurate Are We?

    PubMed

    Ellapparadja, Pregash; Mahajan, Vivek; Deakin, Angela H; Deep, Kamal

    2015-06-01

    This study assesses how accurately we can restore hip offset and leg length in navigated total hip arthroplasty (THA). 152 consecutive patients with navigated THA formed the study group. The contra-lateral hip formed control for measuring hip offset and leg length. All radiological measurements were made using Orthoview digital software. In the normal hip offset group, the mean is 75.73 (SD- 8.61). In the reconstructed hip offset group, the mean is 75.35 (SD - 7.48). 95.39% had hip offset within 6 mm of opposite side while 96.04% had leg length restored within 6 mm of contra-lateral side. Equivalence test revealed that the two groups of hip offsets were essentially the same. We conclude that computer navigation can successfully reproduce hip offset and leg length accurately.

  19. Interplay between Transcription Factors and the Epigenome: Insight from the Role of RUNX1 in Leukemia

    PubMed Central

    Brettingham-Moore, Kate H.; Taberlay, Phillippa C.; Holloway, Adele F.

    2015-01-01

    The genome has the ability to respond in a precise and co-ordinated manner to cellular signals. It achieves this through the concerted actions of transcription factors and the chromatin platform, which are targets of the signaling pathways. Our understanding of the molecular mechanisms through which transcription factors and the chromatin landscape each control gene activity has expanded dramatically over recent years, and attention has now turned to understanding the complex, multifaceted interplay between these regulatory layers in normal and disease states. It has become apparent that transcription factors as well as the components and modifiers of the epigenetic machinery are frequent targets of genomic alterations in cancer cells. Through the study of these factors, we can gain unique insight into the dynamic interplay between transcription factors and the epigenome, and how their dysregulation leads to aberrant gene expression programs in cancer. Here, we will highlight how these factors normally co-operate to establish and maintain the transcriptional and epigenetic landscape of cells, and how this is reprogramed in cancer, focusing on the RUNX1 transcription factor and oncogenic derivative RUNX1–ETO in leukemia as paradigms of transcriptional and epigenetic reprograming. PMID:26483790

  20. Problems in publishing accurate color in IEEE journals.

    PubMed

    Vrhel, Michael J; Trussell, H J

    2002-01-01

    To demonstrate the performance of color image processing algorithms, it is desirable to be able to accurately display color images in archival publications. In poster presentations, the authors have substantial control of the printing process, although little control of the illumination. For journal publication, the authors must rely on professional intermediaries (printers) to accurately reproduce their results. Our previous work describes requirements for accurately rendering images using your own equipment. This paper discusses the problems of dealing with intermediaries and offers suggestions for improved communication and rendering.

  1. Fabricating an Accurate Implant Master Cast: A Technique Report.

    PubMed

    Balshi, Thomas J; Wolfinger, Glenn J; Alfano, Stephen G; Cacovean, Jeannine N; Balshi, Stephen F

    2015-12-01

    The technique for fabricating an accurate implant master cast following the 12-week healing period after Teeth in a Day® dental implant surgery is detailed. The clinical, functional, and esthetic details captured during the final master impression are vital to creating an accurate master cast. This technique uses the properties of the all-acrylic resin interim prosthesis to capture these details. This impression captures the relationship between the remodeled soft tissue and the interim prosthesis. This provides the laboratory technician with an accurate orientation of the implant replicas in the master cast with which a passive fitting restoration can be fabricated.

  2. The Transcriptional Cascade in the Heat Stress Response of Arabidopsis Is Strictly Regulated at the Level of Transcription Factor Expression.

    PubMed

    Ohama, Naohiko; Kusakabe, Kazuya; Mizoi, Junya; Zhao, Huimei; Kidokoro, Satoshi; Koizumi, Shinya; Takahashi, Fuminori; Ishida, Tetsuya; Yanagisawa, Shuichi; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

    2016-01-01

    Group A1 heat shock transcription factors (HsfA1s) are the master regulators of the heat stress response (HSR) in plants. Upon heat shock, HsfA1s trigger a transcriptional cascade that is composed of many transcription factors. Despite the importance of HsfA1s and their downstream transcriptional cascade in the acquisition of thermotolerance in plants, the molecular basis of their activation remains poorly understood. Here, domain analysis of HsfA1d, one of several HsfA1s in Arabidopsis thaliana, demonstrated that the central region of HsfA1d is a key regulatory domain that represses HsfA1d transactivation activity through interaction with HEAT SHOCK PROTEIN70 (HSP70) and HSP90. We designated this region as the temperature-dependent repression (TDR) domain. We found that HSP70 dissociates from HsfA1d in response to heat shock and that the dissociation is likely regulated by an as yet unknown activation mechanism, such as HsfA1d phosphorylation. Overexpression of constitutively active HsfA1d that lacked the TDR domain induced expression of heat shock proteins in the absence of heat stress, thereby conferring potent thermotolerance on the overexpressors. However, transcriptome analysis of the overexpressors demonstrated that the constitutively active HsfA1d could not trigger the complete transcriptional cascade under normal conditions, thereby indicating that other factors are necessary to fully induce the HSR. These complex regulatory mechanisms related to the transcriptional cascade may enable plants to respond resiliently to various heat stress conditions.

  3. Is My Child's Appetite Normal?

    MedlinePlus

    ... HEALTH 17 Nutrition Newsletters for Parents of Young Children, USDA, Food and Nutrition Service Is My Child’s Appetite Normal? ... HEALTH 17 Nutrition Newsletters for Parents of Young Children, USDA, Food and Nutrition Service

  4. Diverse Topics Advance Normal Birth

    PubMed Central

    Humenick, Sharron S.

    2006-01-01

    The editor of The Journal of Perinatal Education describes the contents of this issue, which offer a broad range of resources, research, and inspiration for childbirth educators in their efforts to promote normal birth.

  5. Rethinking transcription coupled DNA repair.

    PubMed

    Kamarthapu, Venu; Nudler, Evgeny

    2015-04-01

    Nucleotide excision repair (NER) is an evolutionarily conserved, multistep process that can detect a wide variety of DNA lesions. Transcription coupled repair (TCR) is a subpathway of NER that repairs the transcribed DNA strand faster than the rest of the genome. RNA polymerase (RNAP) stalled at DNA lesions mediates the recruitment of NER enzymes to the damage site. In this review we focus on a newly identified bacterial TCR pathway in which the NER enzyme UvrD, in conjunction with NusA, plays a major role in initiating the repair process. We discuss the tradeoff between the new and conventional models of TCR, how and when each pathway operates to repair DNA damage, and the necessity of pervasive transcription in maintaining genome integrity.

  6. Chromatin Dynamics of Circadian Transcription

    PubMed Central

    Aguilar-Arnal, Lorena; Sassone-Corsi, Paolo

    2015-01-01

    The molecular circadian clock orchestrates the daily cyclical expression of thousands of genes. Disruption of this transcriptional program leads to a variety of pathologies, including insomnia, depression and metabolic disorders. Circadian rhythms in gene expression rely on specific chromatin transitions which are ultimately coordinated by the molecular clock. As a consequence, a highly plastic and dynamic circadian epigenome can be delineated across different tissues and cell types. Intriguingly, genome topology appears to coordinate cyclic transcription at circadian interactomes, in which circadian genes are in physical contact within the cell nucleus in a time-specific manner. Moreover, the clock machinery shows functional interplays with key metabolic regulators, thereby connecting the circadian epigenome to cellular metabolism. Unraveling the molecular aspects of such interplays is likely to reveal new therapeutic strategies towards the treatment of metabolic disorders. PMID:27014564

  7. Beam normal spin asymmetries: Theory

    SciTech Connect

    M. Vanderhaeghen

    2007-06-01

    The beam normal spin asymmetry in elastic electron-nucleon scattering is discussed. This beam normal spin asymmetry depends on the imaginary part of two-photon exchange processes between electron and nucleon, and measures the non-forward structure functions of the nucleon. After briefly reviewing the theoretical formalism, we discuss calculations in the threshold region, in the resonance region, as well as in the diffractive region, corresponding with high energy and forward angles.

  8. A proximal activator of transcription in epithelial-mesenchymal transition

    PubMed Central

    Venkov, Christo D.; Link, Andrew J.; Jennings, Jennifer L.; Plieth, David; Inoue, Tsutomu; Nagai, Kojiro; Xu, Carol; Dimitrova, Yoana N.; Rauscher, Frank J.; Neilson, Eric G.

    2007-01-01

    Epithelial-mesenchymal transition (EMT) is an important mechanism for phenotypic conversion in normal development and disease states such as tissue fibrosis and metastasis. While this conversion of epithelia is under tight transcriptional control, few of the key transcriptional proteins are known. Fibroblasts produced by EMT express a gene encoding fibroblast-specific protein 1 (FSP1), which is regulated by a proximal cis-acting promoter element called fibroblast transcription site–1 (FTS-1). In mass spectrometry, chromatin immunoprecipitation, and siRNA studies, we used FTS-1 as a unique probe for mediators of EMT and identified a complex of 2 proteins, CArG box–binding factor–A (CBF-A) and KRAB-associated protein 1 (KAP-1), that bind this site. Epithelial cells engineered to conditionally express recombinant CBF-A (rCBF-A) activate the transcription of FSP1 and undergo EMT. The FTS-1 response element also exists in the promoters modulating a broader EMT transcriptome, including Twist, and Snail, as well as E-cadherin, β-catenin, ZO 1, vimentin, α1(I) collagen, and α–smooth muscle actin, and the induction of rCBF-A appropriately alters their expression as well. We believe formation of the CBF-A/KAP-1/FTS-1 complex is sufficient for the induction of FSP1 and a novel proximal activator of EMT. PMID:17273560

  9. A proximal activator of transcription in epithelial-mesenchymal transition.

    PubMed

    Venkov, Christo D; Link, Andrew J; Jennings, Jennifer L; Plieth, David; Inoue, Tsutomu; Nagai, Kojiro; Xu, Carol; Dimitrova, Yoana N; Rauscher, Frank J; Neilson, Eric G

    2007-02-01

    Epithelial-mesenchymal transition (EMT) is an important mechanism for phenotypic conversion in normal development and disease states such as tissue fibrosis and metastasis. While this conversion of epithelia is under tight transcriptional control, few of the key transcriptional proteins are known. Fibroblasts produced by EMT express a gene encoding fibroblast-specific protein 1 (FSP1), which is regulated by a proximal cis-acting promoter element called fibroblast transcription site-1 (FTS-1). In mass spectrometry, chromatin immunoprecipitation, and siRNA studies, we used FTS-1 as a unique probe for mediators of EMT and identified a complex of 2 proteins, CArG box-binding factor-A (CBF-A) and KRAB-associated protein 1 (KAP-1), that bind this site. Epithelial cells engineered to conditionally express recombinant CBF-A (rCBF-A) activate the transcription of FSP1 and undergo EMT. The FTS-1 response element also exists in the promoters modulating a broader EMT transcriptome, including Twist, and Snail, as well as E-cadherin, beta-catenin, ZO 1, vimentin, alpha1(I) collagen, and alpha-smooth muscle actin, and the induction of rCBF-A appropriately alters their expression as well. We believe formation of the CBF-A/KAP-1/FTS-1 complex is sufficient for the induction of FSP1 and a novel proximal activator of EMT.

  10. Transcriptional response to dietary restriction in Drosophila melanogaster

    PubMed Central

    Ding, Feifei; Gil, M. Pilar; Franklin, Michael; Ferreira, Jonathan; Tatar, Marc; Helfand, Stephen L.; Neretti, Nicola

    2014-01-01

    Dietary restriction (DR) extends lifespan in a wide variety of organisms. Although several genes and pathways associated with this longevity response have been identified, the specific mechanism through which DR extends lifespan is not fully understood. We have recently developed a novel methodology to screen for transcriptional changes in response to acutely imposed DR upon adult Drosophila melanogaster and identified groups of genes that switch their transcriptional patterns from a normal diet pattern to a restricted diet pattern, or ‘switching genes’. In this current report we extend our transcriptional data analysis with Gene Set Enrichment Analysis to generate a pathway-centered perspective. The pattern of temporal behavior in response to the diet switch is strikingly similar within and across pathways associated with mRNA processing and protein translation. Furthermore, most genes within these pathways display an initial spike in activity within 6 to 8 hours from the diet switch, followed by a coordinated, partial down-regulation after 24 hours. We propose this represents a stereotypical response to DR, which ultimately leads to a mild but widespread inhibition of transcriptional and translational activity. Inhibition of the protein synthesis pathway has been observed in DR in other studies and has been shown to extend lifespan in several model organisms. PMID:24819200

  11. Yorkie promotes transcription by recruiting a Histone methyltransferase complex

    PubMed Central

    Oh, Hyangyee; Slattery, Matthew; Ma, Lijia; White, Kevin P.; Mann, Richard S.

    2014-01-01

    SUMMARY Hippo signaling limits organ growth by inhibiting the transcriptional coactivator Yorkie. Despite the key role of Yorkie in both normal and oncogenic growth, the mechanism by which it activates transcription has not been defined. We report that Yorkie binding to chromatin correlates with histone H3K4 methylation, and is sufficient to locally increase it. We show that Yorkie can recruit a histone methyltransferase complex, through binding between WW domains of Yorkie and PPxY sequence motifs of NcoA6, a subunit of the Trithorax-related (Trr) methyltransferase complex. Cell culture and in vivo assays establish that this recruitment of NcoA6 contributes to Yorkie’s ability to activate transcription. Mammalian NcoA6, a subunit of Trr-homologous methyltransferase complexes, can similarly interact with Yorkie’s mammalian homologue YAP. Our results implicate direct recruitment of a histone methyltransferase complex as central to transcriptional activation by Yorkie, linking the control of cell proliferation by Hippo signaling to chromatin modification. PMID:25017066

  12. Drugging the Undruggable: Transcription Therapy for Cancer

    PubMed Central

    Yan, Chunhong; Higgins, Paul J.

    2012-01-01

    Transcriptional regulation is often the convergence point of oncogenic signaling. It is not surprising, therefore, that aberrant gene expression is a hallmark of cancer. Transformed cells often develop a dependency on such a reprogramming highlighting the therapeutic potential of rectifying cancer-associated transcriptional abnormalities in malignant cells. Although transcription is traditionally considered as undruggable, agents have been developed that target various levels of transcriptional regulation including DNA binding by transcription factors, protein-protein interactions, and epigenetic alterations. Some of these agents have been approved for clinical use or entered clinical trials. While artificial transcription factors have been developed that can theoretically modulate expression of any given gene, the emergence of reliable reporter assays greatly facilitate the search for transcription-targeted agents. This review provides a comprehensive overview of these developments, and discusses various strategies applicable for developing transcription-targeted therapeutic agents. PMID:23147197

  13. Improved Methods for Teaching Machine Transcription.

    ERIC Educational Resources Information Center

    Smith, Clara J.

    1980-01-01

    The increased use of machine transcription in business and industry demands that business educators attract and train more highly skilled machine transcriptionists. Realistic production measurement and appropriate vocabulary should be taught to link machine transcription to word processing. (Author)

  14. Sry is a transcriptional activator.

    PubMed

    Dubin, R A; Ostrer, H

    1994-09-01

    The SRY gene functions as a genetic switch in gonadal ridge initiating testis determination. The mouse Sry and human SRY open reading frames (ORFs) share a conserved DNA-binding domain (the HMG-box) yet exhibit no additional homology outside this region. As judged by the accumulation of lacZ-SRY hybrid proteins in the nucleus, both the human and mouse SRY ORFs contain a nuclear localization signal. The mouse Sry HMG-box domain selectively binds the sequence NACAAT in vitro when challenged with a random pool of oligonucleotides and binds AACAAT with the highest affinity. When put under the control of a heterologous promotor, the mouse Sry gene activated transcription of a reporter gene containing multiple copies of the AACAAT binding site. Activation was likewise observed for a GAL4-responsive reporter gene, when the mouse Sry gene was linked to the DNA-binding domain of GAL4. Using this system, the activation function was mapped to a glutamine/histidine-rich domain. In addition, LexA-mouse Sry fusion genes activated a LexA-responsive reporter gene in yeast. In contrast, a GAL4-human SRY fusion gene did not cause transcriptional activation. These studies suggest that both the human and the mouse SRY ORFs encode nuclear, DNA-binding proteins and that the mouse Sry ORF can function as a transcriptional activator with separable DNA-binding and activator domains.

  15. Transcription termination maintains chromosome integrity.

    PubMed

    Washburn, Robert S; Gottesman, Max E

    2011-01-11

    DNA replication fork movement is impeded by collisions with transcription elongation complexes (TEC). We propose that a critical function of transcription termination factors is to prevent TEC from blocking DNA replication and inducing replication fork arrest, one consequence of which is DNA double-strand breaks. We show that inhibition of Rho-dependent transcription termination by bicyclomycin in Escherichia coli induced double-strand breaks. Cells deleted for Rho-cofactors nusA and nusG were hypersensitive to bicyclomycin, and had extensive chromosome fragmentation even in the absence of the drug. An RNA polymerase mutation that destabilizes TEC (rpoB*35) increased bicyclomycin resistance >40-fold. Double-strand break formation depended on DNA replication, and can be explained by replication fork collapse. Deleting recombination genes required for replication fork repair (recB and ruvC) increased sensitivity to bicyclomycin, as did loss of the replication fork reloading helicases rep and priA. We propose that Rho responds to a translocating replisome by releasing obstructing TEC.

  16. Linking Smads and transcriptional activation.

    PubMed

    Inman, Gareth J

    2005-02-15

    TGF-beta1 (transforming growth factor-beta1) is the prototypical member of a large family of pleiotropic cytokines that regulate diverse biological processes during development and adult tissue homoeostasis. TGF-beta signals via membrane bound serine/threonine kinase receptors which transmit their signals via the intracellular signalling molecules Smad2, Smad3 and Smad4. These Smads contain conserved MH1 and MH2 domains separated by a flexible linker domain. Smad2 and Smad3 act as kinase substrates for the receptors, and, following phosphorylation, they form complexes with Smad4 and translocate to the nucleus. These Smad complexes regulate gene expression and ultimately determine the biological response to TGF-beta. In this issue of the Biochemical Journal, Wang et al. have shown that, like Smad4, the linker domain of Smad3 contains a Smad transcriptional activation domain. This is capable of recruiting the p300 transcriptional co-activator and is required for Smad3-dependent transcriptional activation. This study raises interesting questions about the nature and regulation of Smad-regulated gene activation and elevates the status of the linker domain to rival that of the much-lauded MH1 and MH2 domains.

  17. Production of the 2400 kb Duchenne muscular dystrophy (DMD) gene transcript; transcription time and cotranscriptional splicing

    SciTech Connect

    Tennyson, C.N.; Worton, R.G.

    1994-09-01

    The largest known gene in any organism is the human DMD gene which has 79 exons that span 2400 kb. The extreme nature of the DMD gene raises questions concerning the time required for transcription and whether splicing begins before transcription is complete. DMD gene transcription is induced as cultured human myoblasts differentiate to form multinucleated myotubes, providing a system for studying the kinetics of transcription and splicing. Using quantitative RT-PCR, transcript accumulation was monitored from four different regions within the gene following induction of expression. By comparing the accumulation of transcripts from the 5{prime} and 3{prime} ends of the gene we have shown that approximately 12 hours are required to transcribe 1770 kb of the gene, extrapolating to a time of 16 hours for the transcription unit expressed in muscle. Comparison of accumulation profiles for spliced and total transcript demonstrated that transcripts are spliced at the 5{prime} end before transcription is complete, providing strong evidence for cotranscriptional splicing of DMD gene transcripts. Finally, the rate of transcript accumulation was reduced at the 3{prime} end of the gene relative to the 5{prime} end, perhaps due to premature termination of transcription complexes as they traverse this enormous transcription unit. The lag between transcription initiation and the appearance of complete transcripts could be important in limiting transcript production in dividing cells and to the timing of mRNA appearance in differentiating muscle.

  18. Switching on sex: transcriptional regulation of the testis-determining gene Sry.

    PubMed

    Larney, Christian; Bailey, Timothy L; Koopman, Peter

    2014-06-01

    Mammalian sex determination hinges on the development of ovaries or testes, with testis fate being triggered by the expression of the transcription factor sex-determining region Y (Sry). Reduced or delayed Sry expression impairs testis development, highlighting the importance of its accurate spatiotemporal regulation and implying a potential role for SRY dysregulation in human intersex disorders. Several epigenetic modifiers, transcription factors and kinases are implicated in regulating Sry transcription, but it remains unclear whether or how this farrago of factors acts co-ordinately. Here we review our current understanding of Sry regulation and provide a model that assembles all known regulators into three modules, each converging on a single transcription factor that binds to the Sry promoter. We also discuss potential future avenues for discovering the cis-elements and trans-factors required for Sry regulation.

  19. Optimized rapid amplification of cDNA ends (RACE) for mapping bacterial mRNA transcripts.

    PubMed

    Tillett, D; Burns, B P; Neilan, B A

    2000-03-01

    A simple, efficient and sensitive RACE-based procedure was developed for the determination of unknown 5' regions from bacterial cDNA. A number of critical modifications were made to the standard RACE method, including the optimization of the RNA extraction, reverse transcription and PCR conditions. This procedure was used to accurately determine the site of transcript initiation and structure of the promoter region of the Helicobacter pylori aspartate carbamoyltransferase gene (pyrB). The technique avoids many of the difficulties associated with established bacterial transcript mapping protocols and can be performed in two days starting with less than 1 microgram of total RNA. The modifications described here have significant potential for the identification of transcript start sites of bacterial genes and non-polyadenylated eukaryotic RNA.

  20. Controlling Hay Fever Symptoms with Accurate Pollen Counts

    MedlinePlus

    ... Library ▸ Hay fever and pollen counts Share | Controlling Hay Fever Symptoms with Accurate Pollen Counts This article has ... Pongdee, MD, FAAAAI Seasonal allergic rhinitis known as hay fever is caused by pollen carried in the air ...

  1. Digital system accurately controls velocity of electromechanical drive

    NASA Technical Reports Server (NTRS)

    Nichols, G. B.

    1965-01-01

    Digital circuit accurately regulates electromechanical drive mechanism velocity. The gain and phase characteristics of digital circuits are relatively unimportant. Control accuracy depends only on the stability of the input signal frequency.

  2. Mutual interdependence of splicing and transcription elongation.

    PubMed

    Brzyżek, Grzegorz; Świeżewski, Szymon

    2015-01-01

    Transcription and splicing are intrinsically linked, as splicing needs a pre-mRNA substrate to commence. The more nuanced view is that the rate of transcription contributes to splicing regulation. On the other hand there is accumulating evidence that splicing has an active role in controlling transcription elongation by DNA-dependent RNA polymerase II (RNAP II). We briefly review those mechanisms and propose a unifying model where splicing controls transcription elongation to provide an optimal timing for successive rounds of splicing.

  3. Transcriptional Regulation by Hypoxia Inducible Factors

    PubMed Central

    Espinosa, Joaquín M.

    2015-01-01

    The cellular response to oxygen deprivation is governed largely by a family of transcription factors known as Hypoxia Inducible Factors (HIFs). This review focuses on the molecular mechanisms by which HIFs regulate the transcriptional apparatus to enable the cellular and organismal response to hypoxia. We discuss here how the various HIF polypeptides, their post-translational modifications, binding partners and transcriptional cofactors affect RNA polymerase II activity to drive context-dependent transcriptional programs during hypoxia. PMID:24099156

  4. Accurate tracking of high dynamic vehicles with translated GPS

    NASA Astrophysics Data System (ADS)

    Blankshain, Kenneth M.

    The GPS concept and the translator processing system (TPS) which were developed for accurate and cost-effective tracking of various types of high dynamic expendable vehicles are described. A technique used by the translator processing system (TPS) to accomplish very accurate high dynamic tracking is presented. Automatic frequency control and fast Fourier transform processes are combined to track 100 g acceleration and 100 g/s jerk with 1-sigma velocity measurement error less than 1 ft/sec.

  5. Accurate Alignment of Plasma Channels Based on Laser Centroid Oscillations

    SciTech Connect

    Gonsalves, Anthony; Nakamura, Kei; Lin, Chen; Osterhoff, Jens; Shiraishi, Satomi; Schroeder, Carl; Geddes, Cameron; Toth, Csaba; Esarey, Eric; Leemans, Wim

    2011-03-23

    A technique has been developed to accurately align a laser beam through a plasma channel by minimizing the shift in laser centroid and angle at the channel outptut. If only the shift in centroid or angle is measured, then accurate alignment is provided by minimizing laser centroid motion at the channel exit as the channel properties are scanned. The improvement in alignment accuracy provided by this technique is important for minimizing electron beam pointing errors in laser plasma accelerators.

  6. Interplay between DNA supercoiling and transcription elongation.

    PubMed

    Ma, Jie; Wang, Michelle

    2014-01-01

    Transcription-coupled DNA supercoiling has been shown to be an important regulator of transcription that is broadly present in the cell. Here we review experimental work which shows that RNA polymerase is a powerful torsional motor that can alter DNA topology and structure, and DNA supercoiling in turn directly affects transcription elongation.

  7. The great repression: chromatin and cryptic transcription.

    PubMed

    Hennig, Bianca P; Fischer, Tamás

    2013-01-01

    The eukaryotic chromatin structure is essential in correctly defining transcription units. Impairing this structure can activate cryptic promoters, and lead to the accumulation of aberrant RNA transcripts. Here we discuss critical pathways that are responsible for the repression of cryptic transcription and the maintenance of genome integrity.

  8. 40 CFR 1610.4 - Deposition Transcripts.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 32 2010-07-01 2010-07-01 false Deposition Transcripts. 1610.4 Section 1610.4 Protection of Environment CHEMICAL SAFETY AND HAZARD INVESTIGATION BOARD ADMINISTRATIVE INVESTIGATIONS § 1610.4 Deposition Transcripts. (a) Transcripts of depositions of witnesses compelled by...

  9. 18 CFR 1b.12 - Transcripts.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 18 Conservation of Power and Water Resources 1 2011-04-01 2011-04-01 false Transcripts. 1b.12 Section 1b.12 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES RULES RELATING TO INVESTIGATIONS § 1b.12 Transcripts. Transcripts, if any,...

  10. 18 CFR 1b.12 - Transcripts.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 18 Conservation of Power and Water Resources 1 2014-04-01 2014-04-01 false Transcripts. 1b.12 Section 1b.12 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES RULES RELATING TO INVESTIGATIONS § 1b.12 Transcripts. Transcripts, if any,...

  11. 18 CFR 1b.12 - Transcripts.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 18 Conservation of Power and Water Resources 1 2012-04-01 2012-04-01 false Transcripts. 1b.12 Section 1b.12 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES RULES RELATING TO INVESTIGATIONS § 1b.12 Transcripts. Transcripts, if any,...

  12. 18 CFR 1b.12 - Transcripts.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 18 Conservation of Power and Water Resources 1 2013-04-01 2013-04-01 false Transcripts. 1b.12 Section 1b.12 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES RULES RELATING TO INVESTIGATIONS § 1b.12 Transcripts. Transcripts, if any,...

  13. 18 CFR 1b.12 - Transcripts.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 18 Conservation of Power and Water Resources 1 2010-04-01 2010-04-01 false Transcripts. 1b.12 Section 1b.12 Conservation of Power and Water Resources FEDERAL ENERGY REGULATORY COMMISSION, DEPARTMENT OF ENERGY GENERAL RULES RULES RELATING TO INVESTIGATIONS § 1b.12 Transcripts. Transcripts, if any,...

  14. Single-site transcription rates through fitting of ensemble-averaged data from fluorescence recovery after photobleaching: a fat-tailed distribution.

    PubMed

    Rosenfeld, Liat; Kepten, Eldad; Yunger, Sharon; Shav-Tal, Yaron; Garini, Yuval

    2015-09-01

    The stochastic process of gene expression is commonly controlled at the level of RNA transcription. The synthesis of messenger RNA (mRNA) is a multistep process, performed by RNA polymerase II and controlled by many transcription factors. Although mRNA transcription is intensively studied, real-time in vivo dynamic rates of a single transcribing polymerase are still not available. A popular method for examining transcription kinetics is the fluorescence recovery after photobleaching (FRAP) approach followed by kinetic modeling. Such analysis has yielded a surprisingly broad range of transcription rates. As transcription depends on many variables such as the chromatin state, binding and unbinding of transcription factors, and cell phase, transcription rates are stochastic variables. Thus, the distribution of rates is expected to follow Poissonian statistics, which does not coincide with the wide range of transcription rate results. Here we present an approach for analyzing FRAP data for single-gene transcription. We find that the transcription dynamics of a single gene can be described with a constant rate for all transcribing polymerases, while cell population transcription rates follow a fat-tailed distribution. This distribution suggests a larger probability for extreme rates than would be implied by normal distribution. Our analysis supports experimental results of transcription from two different promoters, and it explains the puzzling observation of extreme average rate values of transcription.

  15. Single-site transcription rates through fitting of ensemble-averaged data from fluorescence recovery after photobleaching: A fat-tailed distribution

    NASA Astrophysics Data System (ADS)

    Rosenfeld, Liat; Kepten, Eldad; Yunger, Sharon; Shav-Tal, Yaron; Garini, Yuval

    2015-09-01

    The stochastic process of gene expression is commonly controlled at the level of RNA transcription. The synthesis of messenger RNA (mRNA) is a multistep process, performed by RNA polymerase II and controlled by many transcription factors. Although mRNA transcription is intensively studied, real-time in vivo dynamic rates of a single transcribing polymerase are still not available. A popular method for examining transcription kinetics is the fluorescence recovery after photobleaching (FRAP) approach followed by kinetic modeling. Such analysis has yielded a surprisingly broad range of transcription rates. As transcription depends on many variables such as the chromatin state, binding and unbinding of transcription factors, and cell phase, transcription rates are stochastic variables. Thus, the distribution of rates is expected to follow Poissonian statistics, which does not coincide with the wide range of transcription rate results. Here we present an approach for analyzing FRAP data for single-gene transcription. We find that the transcription dynamics of a single gene can be described with a constant rate for all transcribing polymerases, while cell population transcription rates follow a fat-tailed distribution. This distribution suggests a larger probability for extreme rates than would be implied by normal distribution. Our analysis supports experimental results of transcription from two different promoters, and it explains the puzzling observation of extreme average rate values of transcription.

  16. Cell proliferation in normal epidermis

    SciTech Connect

    Weinstein, G.D.; McCullough, J.L.; Ross, P.

    1984-06-01

    A detailed examination of cell proliferation kinetics in normal human epidermis is presented. Using tritiated thymidine with autoradiographic techniques, proliferative and differentiated cell kinetics are defined and interrelated. The proliferative compartment of normal epidermis has a cell cycle duration (Tc) of 311 h derived from 3 components: the germinative labeling index (LI), the duration of DNA synthesis (ts), and the growth fraction (GF). The germinative LI is 2.7% +/- 1.2 and ts is 14 h, the latter obtained from a composite fraction of labeled mitoses curve obtained from 11 normal subjects. The GF obtained from the literature and from human skin xenografts to nude mice is estimated to be 60%. Normal-appearing epidermis from patients with psoriasis appears to have a higher proliferation rate. The mean LI is 4.2% +/- 0.9, approximately 50% greater than in normal epidermis. Absolute cell kinetic values for this tissue, however, cannot yet be calculated for lack of other information on ts and GF. A kinetic model for epidermal cell renewal in normal epidermis is described that interrelates the rate of birth/entry, transit, and/or loss of keratinocytes in the 3 epidermal compartments: proliferative, viable differentiated (stratum malpighii), and stratum corneum. Expected kinetic homeostasis in the epidermis is confirmed by the very similar ''turnover'' rates in each of the compartments that are, respectively, 1246, 1417, and 1490 cells/day/mm2 surface area. The mean epidermal turnover time of the entire tissue is 39 days. The Tc of 311 h in normal cells in 8-fold longer than the psoriatic Tc of 36 h and is necessary for understanding the hyperproliferative pathophysiologic process in psoriasis.

  17. Evaluation of Suitable Reference Genes for Normalization of qPCR Gene Expression Studies in Brinjal (Solanum melongena L.) During Fruit Developmental Stages.

    PubMed

    Kanakachari, Mogilicherla; Solanke, Amolkumar U; Prabhakaran, Narayanasamy; Ahmad, Israr; Dhandapani, Gurusamy; Jayabalan, Narayanasamy; Kumar, Polumetla Ananda

    2016-02-01

    Brinjal/eggplant/aubergine is one of the major solanaceous vegetable crops. Recent availability of genome information greatly facilitates the fundamental research on brinjal. Gene expression patterns during different stages of fruit development can provide clues towards the understanding of its biological functions. Quantitative real-time PCR (qPCR) has become one of the most widely used methods for rapid and accurate quantification of gene expression. However, its success depends on the use of a suitable reference gene for data normalization. For qPCR analysis, a single reference gene is not universally suitable for all experiments. Therefore, reference gene validation is a crucial step. Suitable reference genes for qPCR analysis of brinjal fruit development have not been investigated so far. In this study, we have selected 21 candidate reference genes from the Brinjal (Solanum melongena) Plant Gene Indices database (compbio.dfci.harvard.edu/tgi/plant.html) and studied their expression profiles by qPCR during six different fruit developmental stages (0, 5, 10, 20, 30, and 50 days post anthesis) along with leaf samples of the Pusa Purple Long (PPL) variety. To evaluate the stability of gene expression, geNorm and NormFinder analytical softwares were used. geNorm identified SAND (SAND family protein) and TBP (TATA binding protein) as the best pairs of reference genes in brinjal fruit development. The results showed that for brinjal fruit development, individual or a combination of reference genes should be selected for data normalization. NormFinder identified Expressed gene (expressed sequence) as the best single reference gene in brinjal fruit development. In this study, we have identified and validated for the first time reference genes to provide accurate transcript normalization and quantification at various fruit developmental stages of brinjal which can also be useful for gene expression studies in other Solanaceae plant species.

  18. Fast Monte Carlo Electron-Photon Transport Method and Application in Accurate Radiotherapy

    NASA Astrophysics Data System (ADS)

    Hao, Lijuan; Sun, Guangyao; Zheng, Huaqing; Song, Jing; Chen, Zhenping; Li, Gui

    2014-06-01

    Monte Carlo (MC) method is the most accurate computational method for dose calculation, but its wide application on clinical accurate radiotherapy is hindered due to its poor speed of converging and long computation time. In the MC dose calculation research, the main task is to speed up computation while high precision is maintained. The purpose of this paper is to enhance the calculation speed of MC method for electron-photon transport with high precision and ultimately to reduce the accurate radiotherapy dose calculation time based on normal computer to the level of several hours, which meets the requirement of clinical dose verification. Based on the existing Super Monte Carlo Simulation Program (SuperMC), developed by FDS Team, a fast MC method for electron-photon coupled transport was presented with focus on two aspects: firstly, through simplifying and optimizing the physical model of the electron-photon transport, the calculation speed was increased with slightly reduction of calculation accuracy; secondly, using a variety of MC calculation acceleration methods, for example, taking use of obtained information in previous calculations to avoid repeat simulation of particles with identical history; applying proper variance reduction techniques to accelerate MC method convergence rate, etc. The fast MC method was tested by a lot of simple physical models and clinical cases included nasopharyngeal carcinoma, peripheral lung tumor, cervical carcinoma, etc. The result shows that the fast MC method for electron-photon transport was fast enough to meet the requirement of clinical accurate radiotherapy dose verification. Later, the method will be applied to the Accurate/Advanced Radiation Therapy System ARTS as a MC dose verification module.

  19. Roles of transcriptional network during the formation of lymphatic vessels.

    PubMed

    Watabe, Tetsuro

    2012-09-01

    The lymphatic vascular system, also known as the second vascular system in vertebrates, plays crucial roles in various physiological and pathological processes. It participates in the maintenance of normal tissue fluid balance, trafficking of the immune cells and absorption of fatty acids in the gut. Furthermore, lymphatic system is associated with the pathogenesis of a number of diseases, including lymphedema, inflammatory diseases and tumour metastasis. Lymphatic vessels are comprised of lymphatic endothelial cells (LECs), which are differentiated from blood vascular endothelial cells. This review highlights recent advances in our understanding of the transcriptional control of LEC fate determination and reflects on efforts to understand the roles of transcriptional networks during this discrete developmental process.

  20. Developmental-stage-dependent transcriptional response to leukaemic oncogene expression

    PubMed Central

    Regha, Kakkad; Assi, Salam A.; Tsoulaki, Olga; Gilmour, Jane; Lacaud, Georges; Bonifer, Constanze

    2015-01-01

    Acute myeloid leukaemia (AML) is characterized by a block in myeloid differentiation the stage of which is dependent on the nature of the transforming oncogene and the developmental stage of the oncogenic hit. This is also true for the t(8;21) translocation that gives rise to the RUNX1-ETO fusion protein and initiates the most common form of human AML. Here we study the differentiation of mouse embryonic stem cells expressing an inducible RUNX1-ETO gene into blood cells as a model, combined with genome-wide analyses of transcription factor binding and gene expression. RUNX1-ETO interferes with both the activating and repressive function of its normal counterpart, RUNX1, at early and late stages of blood cell development. However, the response of the transcriptional network to RUNX1-ETO expression is developmental stage specific, highlighting the molecular mechanisms determining specific target cell expansion after an oncogenic hit. PMID:26018585

  1. Transcriptional regulation of human small nuclear RNA genes

    PubMed Central

    Jawdekar, Gauri W.; Henry, R. William

    2009-01-01

    The products of human snRNA genes have been frequently described as performing housekeeping functions and their synthesis refractory to regulation. However, recent studies have emphasized that snRNA and other related non-coding RNA molecules control multiple facets of the central dogma, and their regulated expression is critical to cellular homeostasis during normal growth and in response to stress. Human snRNA genes contain compact and yet powerful promoters that are recognized by increasingly well-characterized transcription factors, thus providing a premier model system to study gene regulation. This review summarizes many recent advances deciphering the mechanism by which the transcription of human snRNA and related genes are regulated. PMID:18442490

  2. Transcription factor co-repressors in cancer biology: roles and targeting.

    PubMed

    Battaglia, Sebastiano; Maguire, Orla; Campbell, Moray J

    2010-06-01

    Normal transcription displays a high degree of flexibility over the choice, timing and magnitude of mRNA expression levels that tend to oscillate and cycle. These processes allow for combinatorial actions, feedback control and fine-tuning. A central role has emerged for the transcriptional co-repressor proteins such as NCOR1, NCOR2/SMRT, CoREST and CTBPs, to control the actions of many transcriptional factors, in large part, by recruitment and activation of a range of chromatin remodeling enzymes. Thus, co-repressors and chromatin remodeling factors are recruited to transcription factors at specific promoter/enhancer regions and execute changes in the chromatin structure. The specificity of this recruitment is controlled in a spatial-temporal manner. By playing a central role in transcriptional control, as they move and target transcription factors, co-repressors act as a key driver in the epigenetic economy of the nucleus. Co-repressor functions are selectively distorted in malignancy, by both loss and gain of function and contribute to the generation of transcriptional rigidity. Features of transcriptional rigidity apparent in cancer cells include the distorted signaling of nuclear receptors and the WNTs/beta-catenin axis. Understanding and predicting the consequences of altered co-repressor expression patterns in cancer cells has diagnostic and prognostic significance, and also have the capacity to be targeted through selective epigenetic therapies.

  3. Transcription factors relevant to auxin signalling coordinate broad-spectrum metabolic shifts including sulphur metabolism

    PubMed Central

    Falkenberg, Bettina; Witt, Isabell; Zanor, Maria Inés; Steinhauser, Dirk; Mueller-Roeber, Bernd; Hesse, Holger; Hoefgen, Rainer

    2008-01-01

    A systems approach has previously been used to follow the response behaviour of Arabidopsis thaliana plants upon sulphur limitation. A response network was reconstructed from a time series of transcript and metabolite profiles, integrating complex metabolic and transcript data in order to investigate a potential causal relationship. The resulting scale-free network allowed potential transcriptional regulators of sulphur metabolism to be identified. Here, three sulphur-starvation responsive transcription factors, IAA13, IAA28, and ARF-2 (ARF1-Binding Protein), all of which are related to auxin signalling, were selected for further investigation. IAA28 overexpressing and knock-down lines showed no major morphological changes, whereas IAA13- and ARF1-BP-overexpressing plants grew more slowly than the wild type. Steady-state metabolite levels and expression of pathway-relevant genes were monitored under normal and sulphate-depleted conditions. For all lines, changes in transcript and metabolite levels were observed, yet none of these changes could exclusively be linked to sulphur stress. Instead, up- or down-regulation of the transcription factors caused metabolic changes which in turn affected sulphur metabolism. Auxin-relevant transcription factors are thus part of a complex response pattern to nutrient starvation that serve as coordinators of the metabolic shifts driving sulphur homeostasis rather then as direct effectors of the sulphate assimilation pathway. This study provides the first evidence ever presented that correlates auxin-related transcriptional regulators with primary plant metabolism. PMID:18596113

  4. Increased expression of the Hutchinson-Gilford progeria syndrome truncated lamin A transcript during cell aging.

    PubMed

    Rodriguez, Sofia; Coppedè, Fabio; Sagelius, Hanna; Eriksson, Maria

    2009-07-01

    Most cases of the segmental progeroid syndrome, Hutchinson-Gilford progeria syndrome (HGPS), are caused by a de novo dominant mutation within a single codon of the LMNA gene. This mutation leads to the increased usage of an internal splice site that generates an alternative lamin A transcript with an internal deletion of 150 nucleotides, called lamin A Delta 150. The LMNA gene encodes two major proteins of the inner nuclear lamina, lamins A and C, but not much is known about their expression levels. Determination of the overall expression levels of the LMNA gene transcripts is an important step to further the understanding of the HGPS. In this study, we have performed absolute quantification of the lamins A, C and A Delta 150 transcripts in primary dermal fibroblasts from HGPS patients and unaffected age-matched and parent controls. We show that the lamin A Delta 150 transcript is present in unaffected controls but its expression is >160-fold lower than that in samples from HGPS patients. Analysis of transcript expression during in vitro aging shows that although the levels of lamin A and lamin C transcripts remain unchanged, the lamin A Delta 150 transcript increases in late passage cells from HGPS patients and parental controls. This study provides a new method for LMNA transcript analysis and insights into the expression of the LMNA gene in HGPS and normal cells.

  5. Transcription factor profiling shows new ways towards new treatment options of cutaneous T cell lymphomas.

    PubMed

    Döbbeling, Udo

    2007-06-01

    Most oncogenes encode activators of transcription factors or transcription factors themselves. Transcription factors that are induced by growth stimuli are, in contrast to transcription factors that regulate house keeping genes, tightly regulated and only active, when a stimulus (e.g. cytokines or other growth factors) is given. Examples of such transcription factors are members of the jun, fos, myc, NFkB and STAT gene families. In cancer cells this regulation is interrupted, resulting in constitutive activities of transcription factors that are normally silent. This in turn results in the increased expression of target genes that are necessary for growth and protection from apoptosis. Since inducible transcription factors are activated by specific pathways, the identification of unusual constitutively active transcription factors also identifies the involved signal transduction pathway. Inhibitors of the components of these pathways may be effective anti-cancer agents, as they interrupt the abnormal signalling and in cancer cells. We applied this strategy for two forms of cutaneous T cell lymphomas and identified several groups of agents that may be the prototypes of new drugs to fight these diseases.

  6. Transcription factors relevant to auxin signalling coordinate broad-spectrum metabolic shifts including sulphur metabolism.

    PubMed

    Falkenberg, Bettina; Witt, Isabell; Zanor, Maria Inés; Steinhauser, Dirk; Mueller-Roeber, Bernd; Hesse, Holger; Hoefgen, Rainer

    2008-01-01

    A systems approach has previously been used to follow the response behaviour of Arabidopsis thaliana plants upon sulphur limitation. A response network was reconstructed from a time series of transcript and metabolite profiles, integrating complex metabolic and transcript data in order to investigate a potential causal relationship. The resulting scale-free network allowed potential transcriptional regulators of sulphur metabolism to be identified. Here, three sulphur-starvation responsive transcription factors, IAA13, IAA28, and ARF-2 (ARF1-Binding Protein), all of which are related to auxin signalling, were selected for further investigation. IAA28 overexpressing and knock-down lines showed no major morphological changes, whereas IAA13- and ARF1-BP-overexpressing plants grew more slowly than the wild type. Steady-state metabolite levels and expression of pathway-relevant genes were monitored under normal and sulphate-depleted conditions. For all lines, changes in transcript and metabolite levels were observed, yet none of these changes could exclusively be linked to sulphur stress. Instead, up- or down-regulation of the transcription factors caused metabolic changes which in turn affected sulphur metabolism. Auxin-relevant transcription factors are thus part of a complex response pattern to nutrient starvation that serve as coordinators of the metabolic shifts driving sulphur homeostasis rather then as direct effectors of the sulphate assimilation pathway. This study provides the first evidence ever presented that correlates auxin-related transcriptional regulators with primary plant metabolism.

  7. Identification of differentially expressed sense and antisense transcript pairs in breast epithelial tissues

    PubMed Central

    Grigoriadis, Anita; Oliver, Gavin R; Tanney, Austin; Kendrick, Howard; Smalley, Matt J; Jat, Parmjit; Neville, A Munro

    2009-01-01

    Background More than 20% of human transcripts have naturally occurring antisense products (or natural antisense transcripts – NATs), some of which may play a key role in a range of human diseases. To date, several databases of in silico defined human sense-antisense (SAS) pairs have appeared, however no study has focused on differential expression of SAS pairs in breast tissue. We therefore investigated the expression levels of sense and antisense transcripts in normal and malignant human breast epithelia using the Affymetrix HG-U133 Plus 2.0 and Almac Diagnostics Breast Cancer DSA microarray technologies as well as massively parallel signature sequencing (MPSS) data. Results The expression of more than 2500 antisense transcripts were detected in normal breast duct luminal cells and in primary breast tumors substantially enriched for their epithelial cell content by DSA microarray. Expression of 431 NATs were confirmed by either of the other two technologies. A corresponding sense transcript could be identified on DSA for 257 antisense transcripts. Of these SAS pairs, 163 have not been previously reported. A positive correlation of differential expression between normal and malignant breast samples was observed for most SAS pairs. Orientation specific RT-QPCR of selected SAS pairs validated their expression in several breast cancer cell lines and solid breast tumours. Conclusion Disease-focused and antisense enriched microarray platforms (such as Breast Cancer DSA) confirm the assumption that antisense transcription in the human breast is more prevalent than previously anticipated. Expression of a proportion of these NATs has already been confirmed by other technologies while the true existence of the remaining ones has to be validated. Nevertheless, future studies will reveal whether the relative abundances of antisense and sense transcripts have regulatory influences on the translation of these mRNAs. PMID:19615061

  8. Normalization for Relative Quantification of mRNA and microRNA in Soybean Exposed to Various Abiotic Stresses

    PubMed Central

    Zhou, Yonggang; Chen, Huan; Dong, Yuanyuan; Wang, Nan; Li, Xiaowei; Jameel, Aysha; Yang, He; Zhang, Min; Chen, Kai; Wang, Fawei; Li, Haiyan

    2016-01-01

    Plant microRNAs are small non-coding, endogenic RNA molecule (containing 20–24 nucleotides) produced from miRNA precursors (pri-miRNA and pre-miRNA). Evidence suggests that up and down regulation of the miRNA targets the mRNA genes involved in resistance against biotic and abiotic stresses. Reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) is a powerful technique to analyze variations in mRNA levels. Normalizing the data using reference genes is essential for the analysis of reliable RT-qPCR data. In this study, two groups of candidate reference mRNAs and miRNAs in soybean leaves and roots treated with various abiotic stresses (PEG-simulated drought, salinity, alkalinity, salinity+alkalinity, and abscisic acid) were analyzed by RT-qPCR. We analyzed the most appropriate reference mRNA/miRNAs using the geNorm, NormFinder, and BestKeeper algorithms. According to the results, Act and EF1b were the most suitable reference mRNAs in leaf and root samples, for mRNA and miRNA precursor data normalization. The most suitable reference miRNAs found in leaf and root samples were 166a and 167a for mature miRNA data normalization. Hence the best combinations of reference mRNAs for mRNA and miRNA precursor data normalization were EF1a + Act or EF1b + Act in leaf samples, and EF1a + EF1b or 60s + EF1b in root samples. For mature miRNA data normalization, the most suitable combinations of reference miRNAs were 166a + 167d in leaf samples, and 171a + 156a or 167a + 171a in root samples. We identified potential reference mRNA/miRNAs for accurate RT-qPCR data normalization for mature miRNA, miRNA precursors, and their targeted mRNAs. Our results promote miRNA-based studies on soybean plants exposed to abiotic stress conditions. PMID:27176476

  9. Normal probability plots with confidence.

    PubMed

    Chantarangsi, Wanpen; Liu, Wei; Bretz, Frank; Kiatsupaibul, Seksan; Hayter, Anthony J; Wan, Fang

    2015-01-01

    Normal probability plots are widely used as a statistical tool for assessing whether an observed simple random sample is drawn from a normally distributed population. The users, however, have to judge subjectively, if no objective rule is provided, whether the plotted points fall close to a straight line. In this paper, we focus on how a normal probability plot can be augmented by intervals for all the points so that, if the population distribution is normal, then all the points should fall into the corresponding intervals simultaneously with probability 1-α. These simultaneous 1-α probability intervals provide therefore an objective mean to judge whether the plotted points fall close to the straight line: the plotted points fall close to the straight line if and only if all the points fall into the corresponding intervals. The powers of several normal probability plot based (graphical) tests and the most popular nongraphical Anderson-Darling and Shapiro-Wilk tests are compared by simulation. Based on this comparison, recommendations are given in Section 3 on which graphical tests should be used in what circumstances. An example is provided to illustrate the methods.

  10. The lack of a systematic validation of reference genes: a serious pitfall undervalued in reverse transcription-polymerase chain reaction (RT-PCR) analysis in plants.

    PubMed

    Gutierrez, Laurent; Mauriat, Mélanie; Guénin, Stéphanie; Pelloux, Jérôme; Lefebvre, Jean-François; Louvet, Romain; Rusterucci, Christine; Moritz, Thomas; Guerineau, François; Bellini, Catherine; Van Wuytswinkel, Olivier

    2008-08-01

    Reverse transcription-polymerase chain reaction (RT-PCR) approaches have been used in a large proportion of transcriptome analyses published to date. The accuracy of the results obtained by this method strongly depends on accurate transcript normalization using stably expressed genes, known as references. Statistical algorithms have been developed recently to help validate reference genes, and most studies of gene expression in mammals, yeast and bacteria now include such validation. Surprisingly, this important approach is under-utilized in plant studies, where putative housekeeping genes tend to be used as references without any appropriate validation. Using quantitative RT-PCR, the expression stability of several genes commonly used as references was tested in various tissues of Arabidopsis thaliana and hybrid aspen (Populus tremula x Populus tremuloides). It was found that the expression of most of these genes was unstable, indicating that their use as references is inappropriate. The major impact of the use of such inappropriate references on the results obtained by RT-PCR is demonstrated in this study. Using aspen as a model, evidence is presented indicating that no gene can act as a universal reference, implying the need for a systematic validation of reference genes. For the first time, the extent to which the lack of a systematic validation of reference genes is a stumbling block to the reliability of results obtained by RT-PCR in plants is clearly shown.

  11. Novel Transcription Factor Variants through RNA-Sequencing: The Importance of Being “Alternative”

    PubMed Central

    Scarpato, Margherita; Federico, Antonio; Ciccodicola, Alfredo; Costa, Valerio

    2015-01-01

    Alternative splicing is a pervasive mechanism of RNA maturation in higher eukaryotes, which increases proteomic diversity and biological complexity. It has a key regulatory role in several physiological and pathological states. The diffusion of Next Generation Sequencing, particularly of RNA-Sequencing, has exponentially empowered the identification of novel transcripts revealing that more than 95% of human genes undergo alternative splicing. The highest rate of alternative splicing occurs in transcription factors encoding genes, mostly in Krüppel-associated box domains of zinc finger proteins. Since these molecules are responsible for gene expression, alternative splicing is a crucial mechanism to “regulate the regulators”. Indeed, different transcription factors isoforms may have different or even opposite functions. In this work, through a targeted re-analysis of our previously published RNA-Sequencing datasets, we identified nine novel transcripts in seven transcription factors genes. In silico analysis, combined with RT-PCR, cloning and Sanger sequencing, allowed us to experimentally validate these new variants. Through computational approaches we also predicted their novel structural and functional properties. Our findings indicate that alternative splicing is a major determinant of transcription factor diversity, confirming that accurate analysis of RNA-Sequencing data can reliably lead to the identification of novel transcripts, with potentially new functions. PMID:25590302

  12. Conductivity Modulation in a gated Normal-CDW-Normal configuration

    NASA Astrophysics Data System (ADS)

    Biswas, Saumya; Lake, Roger

    There is considerable interest in switching by exploiting a voltage controlled phase transition, and one such phase is the charge density wave phase that occurs in a number of quasi one dimensional and two dimensional transition metal dichalcogenides. Voltage controlled switching of the charge density wave transition in 1T-TaS2 has recently been demonstrated. We consider a transistor geometry with normal metal contacts and a channel of CDW material. The interaction is modeled with a negative U Hubbard term. Normal-CDW-temperature-U phase diagrams show the regime of the CDW in the ideal lattice. The wavelength of the CDW in the transistor channel is determined by both the conditions of Fermi surface nesting and also the condition of commensurability with the channel length between the two normal leads. Moving the Fermi level of the channel first results in phase boundaries within the CDW as the conditions of commensurability and Fermi surface nesting become incompatible. Moving the Fermi level from half filling by few tens of meV causes a collapsing of the CDW gap and an effective CDW-normal transition, leaving vestiges of the CDW in the channel. The transition is accompanied by one to two orders of magnitude increase in the conductivity. This work is supported by the National Science Foundation (NSF) Grant No. 1124733 and the Semiconductor Research Corporation (SRC) Nanoelectronic Research Initiative as a part of the Nanoelectronics for 2020 and Beyond (NEB-2020) program.

  13. 3j Symbols: To Normalize or Not to Normalize?

    ERIC Educational Resources Information Center

    van Veenendaal, Michel

    2011-01-01

    The systematic use of alternative normalization constants for 3j symbols can lead to a more natural expression of quantities, such as vector products and spherical tensor operators. The redefined coupling constants directly equate tensor products to the inner and outer products without any additional square roots. The approach is extended to…

  14. Characterization of proopiomelanocortin transcripts in human nonpituitary tissues

    SciTech Connect

    Lacaze-Masmonteil, T.; De Keyzer, Y.; Luton, J.P.; Kahn, A.; Bertagna, X.

    1987-10-01

    Proopiomelanocortin (POMC), the precursor to adrenocorticotropic hormone and other related peptides, was originally identified in the corticotropic cell. Recent evidence shows that POMC products are also normally present in a variety of nonpituitary tissues. To investigate this phenomenon in humans the authors looked for the presence and characteristics of POMC transcripts in various adult tissues. Blot hybridization analysis of normal adrenal, thymus, and testis RNAs revealed a small RNA species approximately 400 nucleotides shorter than the 1200-nucleotide pituitary species. Primer extension and S1 nuclease mapping studies showed that this small RNA lacked exon 1 and exon 2 of the gene, and it corresponded to a set of at least six molecules starting 41 to 162 nucleotides downstream from the 5' end of exon 3. These RNAs appear to result from heterogeneous transcription initiation sites presumably under the control of GC box promoter sequences located in the 3' end of intron 2. They cannot encode a complete POMC molecule, and the only truncated POMC molecules that could be translated would lack a signal peptide necessary for membrane translocation and precursor processing. The use of highly sensitive S1 nuclease mapping techniques with uniformly labeled single-stranded DNA probes allowed the detection of a small but definite amount of the normal, 1200-nucleotide, mRNA species. It is suggested that it is this POMC mRNA that is responsible for the local production of all the POMC peptides.

  15. Contributions of in vitro transcription to the understanding of human RNA polymerase III transcription.

    PubMed

    Dumay-Odelot, Hélène; Durrieu-Gaillard, Stéphanie; El Ayoubi, Leyla; Parrot, Camila; Teichmann, Martin

    2014-01-01

    Human RNA polymerase III transcribes small untranslated RNAs that contribute to the regulation of essential cellular processes, including transcription, RNA processing and translation. Analysis of this transcription system by in vitro transcription techniques has largely contributed to the discovery of its transcription factors and to the understanding of the regulation of human RNA polymerase III transcription. Here we review some of the key steps that led to the identification of transcription factors and to the definition of minimal promoter sequences for human RNA polymerase III transcription.

  16. Contribution of transcription to animal early development.

    PubMed

    Wang, Jianbin; Davis, Richard E

    2014-01-01

    In mature gametes and during the oocyte-to-embryo transition, transcription is generally silenced and gene expression is post-transcriptionally regulated. However, we recently discovered that major transcription can occur immediately after fertilization, prior to pronuclear fusion, and in the first cell division of the oocyte-to-embryo transition in the nematode Ascaris suum. We postulate that the balance between transcriptional and post-transcriptional regulation during the oocyte-to-embryo transition may largely be determined by cell cycle length and thus the time available for the genome to be transcribed.

  17. Switch Transcripts in Immunoglobulin Class Switching

    NASA Astrophysics Data System (ADS)

    Lorenz, Matthias; Jung, Steffen; Radbruch, Andreas

    1995-03-01

    B cells can exchange gene segments for the constant region of the immunoglobulin heavy chain, altering the class and effector function of the antibodies that they produce. Class switching is directed to distinct classes by cytokines, which induce transcription of the targeted DNA sequences. These transcripts are processed, resulting in spliced "switch" transcripts. Switch recombination can be directed to immunoglobulin G1 (IgG1) by the heterologous human metallothionein II_A promoter in mutant mice. Induction of the structurally conserved, spliced switch transcripts is sufficient to target switch recombination to IgG1, whereas transcription alone is not.

  18. Mechanical stratigraphy and normal faulting

    NASA Astrophysics Data System (ADS)

    Ferrill, David A.; Morris, Alan P.; McGinnis, Ronald N.; Smart, Kevin J.; Wigginton, Sarah S.; Hill, Nicola J.

    2017-01-01

    Mechanical stratigraphy encompasses the mechanical properties, thicknesses, and interface properties of rock units. Although mechanical stratigraphy often relates directly to lithostratigraphy, lithologic description alone does not adequately describe mechanical behavior. Analyses of normal faults with displacements of millimeters to 10's of kilometers in mechanically layered rocks reveal that mechanical stratigraphy influences nucleation, failure mode, fault geometry, displacement gradient, displacement distribution, fault core and damage zone characteristics, and fault zone deformation processes. The relationship between normal faulting and mechanical stratigraphy can be used either to predict structural style using knowledge of mechanical stratigraphy, or conversely to interpret mechanical stratigraphy based on characterization of the structural style. This review paper explores a range of mechanical stratigraphic controls on normal faulting illustrated by natural and modeled examples.

  19. An accurate assay for HCV based on real-time fluorescence detection of isothermal RNA amplification.

    PubMed

    Wu, Xuping; Wang, Jianfang; Song, Jinyun; Li, Jiayan; Yang, Yongfeng

    2016-09-01

    Hepatitis C virus (HCV) is one of the common reasons of liver fibrosis and hepatocellular carcinoma (HCC). Early, rapid and accurate HCV RNA detection is important to prevent and control liver disease. A simultaneous amplification and testing (SAT) assay, which is based on isothermal amplification of RNA and real-time fluorescence detection, was designed to optimize routine HCV RNA detection. In this study, HCV RNA and an internal control (IC) were amplified and analyzed simultaneously by SAT assay and detection of fluorescence using routine real-time PCR equipment. The assay detected as few as 10 copies of HCV RNA transcripts. We tested 705 serum samples with SAT, among which 96.4% (680/705) showed consistent results compared with routine real-time PCR. About 92% (23/25) discordant samples were confirmed to be same results as SAT-HCV by using a second real-time PCR. The sensitivity and specificity of SAT-HCV assay were 99.6% (461/463) and 100% (242/242), respectively. In conclusion, the SAT assay is an accurate test with a high specificity and sensitivity which may increase the detection rate of HCV. It is therefore a promising tool to diagnose HCV infection.

  20. TOBFAC: the database of tobacco transcription factors

    PubMed Central

    Rushton, Paul J; Bokowiec, Marta T; Laudeman, Thomas W; Brannock, Jennifer F; Chen, Xianfeng; Timko, Michael P

    2008-01-01

    Background Regulation of gene expression at the level of transcription is a major control point in many biological processes. Transcription factors (TFs) can activate and/or repress the transcriptional rate of target genes and vascular plant genomes devote approximately 7% of their coding capacity to TFs. Global analysis of TFs has only been performed for three complete higher plant genomes – Arabidopsis (Arabidopsis thaliana), poplar (Populus trichocarpa) and rice (Oryza sativa). Presently, no large-scale analysis of TFs has been made from a member of the Solanaceae, one of the most important families of vascular plants. To fill this void, we have analysed tobacco (Nicotiana tabacum) TFs using a dataset of 1,159,022 gene-space sequence reads (GSRs) obtained by methylation filtering of the tobacco genome. An analytical pipeline was developed to isolate TF sequences from the GSR data set. This involved multiple (typically 10–15) independent searches with different versions of the TF family-defining domain(s) (normally the DNA-binding domain) followed by assembly into contigs and verification. Our analysis revealed that tobacco contains a minimum of 2,513 TFs representing all of the 64 well-characterised plant TF families. The number of TFs in tobacco is higher than previously reported for Arabidopsis and rice. Results TOBFAC: the database of tobacco transcription factors, is an integrative database that provides a portal to sequence and phylogeny data for the identified TFs, together with a large quantity of other data concerning TFs in tobacco. The database contains an individual page dedicated to each of the 64 TF families. These contain background information, domain architecture via Pfam links, a list of all sequences and an assessment of the minimum number of TFs in this family in tobacco. Downloadable phylogenetic trees of the major families are provided along with detailed information on the bioinformatic pipeline that was used to find all family members

  1. The regulation of mammalian mRNA transcription by lncRNAs: recent discoveries and current concepts.

    PubMed

    Kugel, Jennifer F; Goodrich, James A

    2013-02-01

    Transcription by RNA Pol II is a tightly controlled process that is critical to normal cellular metabolism. Understanding how transcriptional regulation is orchestrated has mainly involved identifying and characterizing proteins that function as transcription factors. During the past decade, however, an increasing number of lncRNAs have been identified as transcriptional regulators. This revelation has spurred new discoveries, novel techniques and paradigm shifts, which together are redefining our understanding of transcriptional control and broadening our view of RNA function. Here, we summarize recent discoveries concerning the role of lncRNAs as regulators of mammalian mRNA transcription, with a focus on key concepts that are guiding current research in the field.

  2. Social Support in Normal Aging

    PubMed Central

    Matthews, Anne Martin

    1984-01-01

    The role of social support in helping elderly people deal with stressful life events is quite complex. This complexity exists because it is difficult to define exactly what social support is, and because the experiences of `normal' aging vary. This article uses the example of adaptation to widowhood to examine the relationship between normal aging and sources, types, and patterns of social support. These factors influence the extent to which support lessens the impact of age-related stressful events. The physician has a role in primary social support, and also in facilitating the supportive functions of family and others. PMID:21279087

  3. Asymptotic Normality of Quadratic Estimators.

    PubMed

    Robins, James; Li, Lingling; Tchetgen, Eric; van der Vaart, Aad

    2016-12-01

    We prove conditional asymptotic normality of a class of quadratic U-statistics that are dominated by their degenerate second order part and have kernels that change with the number of observations. These statistics arise in the construction of estimators in high-dimensional semi- and non-parametric models, and in the construction of nonparametric confidence sets. This is illustrated by estimation of the integral of a square of a density or regression function, and estimation of the mean response with missing data. We show that estimators are asymptotically normal even in the case that the rate is slower than the square root of the observations.

  4. Normalized cDNA libraries

    DOEpatents

    Soares, M.B.; Efstratiadis, A.

    1997-06-10

    This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3{prime} noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to moderate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library. 4 figs.

  5. Normalized cDNA libraries

    DOEpatents

    Soares, Marcelo B.; Efstratiadis, Argiris

    1997-01-01

    This invention provides a method to normalize a directional cDNA library constructed in a vector that allows propagation in single-stranded circle form comprising: (a) propagating the directional cDNA library in single-stranded circles; (b) generating fragments complementary to the 3' noncoding sequence of the single-stranded circles in the library to produce partial duplexes; (c) purifying the partial duplexes; (d) melting and reassociating the purified partial duplexes to moderate Cot; and (e) purifying the unassociated single-stranded circles, thereby generating a normalized cDNA library.

  6. Mammalian transcription-coupled excision repair.

    PubMed

    Vermeulen, Wim; Fousteri, Maria

    2013-08-01

    Transcriptional arrest caused by DNA damage is detrimental for cells and organisms as it impinges on gene expression and thereby on cell growth and survival. To alleviate transcriptional arrest, cells trigger a transcription-dependent genome surveillance pathway, termed transcription-coupled nucleotide excision repair (TC-NER) that ensures rapid removal of such transcription-impeding DNA lesions and prevents persistent stalling of transcription. Defective TC-NER is causatively linked to Cockayne syndrome, a rare severe genetic disorder with multisystem abnormalities that results in patients' death in early adulthood. Here we review recent data on how damage-arrested transcription is actively coupled to TC-NER in mammals and discuss new emerging models concerning the role of TC-NER-specific factors in this process.

  7. Effects of elongation delay in transcription dynamics.

    PubMed

    Zhang, Xuan; Jin, Huiqin; Yang, Zhuoqin; Lei, Jinzhi

    2014-12-01

    In the transcription process, elongation delay is induced by the movement of RNA polymerases (RNAP) along the DNA sequence, and can result in changes in the transcription dynamics. This paper studies the transcription dynamics that involved the elongation delay and effects of cell division and DNA replication. The stochastic process of gene expression is modeled with delay chemical master equation with periodic coefficients, and is studied numerically through the stochastic simulation algorithm with delay. We show that the average transcription level approaches to a periodic dynamics over cell cycles at homeostasis, and the elongation delay can reduce the transcription level and increase the transcription noise. Moreover, the transcription elongation can induce bimodal distribution of mRNA levels that can be measured by the techniques of flow cytometry.

  8. Transcriptional Memory in the Drosophila Embryo.

    PubMed

    Ferraro, Teresa; Esposito, Emilia; Mancini, Laure; Ng, Sam; Lucas, Tanguy; Coppey, Mathieu; Dostatni, Nathalie; Walczak, Aleksandra M; Levine, Michael; Lagha, Mounia

    2016-01-25

    Transmission of active transcriptional states from mother to daughter cells has the potential to foster precision in the gene expression programs underlying development. Such transcriptional memory has been specifically proposed to promote rapid reactivation of complex gene expression profiles after successive mitoses in Drosophila development [1]. By monitoring transcription in living Drosophila embryos, we provide the first evidence for transcriptional memory in animal development. We specifically monitored the activities of stochastically expressed transgenes in order to distinguish active and inactive mother cells and the behaviors of their daughter nuclei after mitosis. Quantitative analyses reveal that there is a 4-fold higher probability for rapid reactivation after mitosis when the mother experienced transcription. Moreover, memory nuclei activate transcription twice as fast as neighboring inactive mothers, thus leading to augmented levels of gene expression. We propose that transcriptional memory is a mechanism of precision, which helps coordinate gene activity during embryogenesis.

  9. Transcriptional networks in leaf senescence.

    PubMed

    Schippers, Jos H M

    2015-10-01

    Plant senescence is a natural phenomenon known for the appearance of beautiful autumn colors and the ripening of cereals in the field. Senescence is a controlled process that plants utilize to remobilize nutrients from source leaves to developing tissues. While during the past decades, molecular components underlying the onset of senescence have been intensively studied, knowledge remains scarce on the age-dependent mechanisms that control the onset of senescence. Recent advances have uncovered transcriptional networks regulating the competence to senesce. Here, gene regulatory networks acting as internal timing mechanisms for the onset of senescence are highlighted, illustrating that early and late leaf developmental phases are highly connected.

  10. Targeting Transcription Factors in Cancer

    PubMed Central

    Bhagwat, Anand S.; Vakoc, Christopher R.

    2015-01-01

    Transcription factors (TFs) are commonly deregulated in the pathogenesis of human cancer and are a major class of cancer cell dependencies. Consequently, targeting of TFs can be highly effective in treating particular malignancies, as highlighted by the clinical efficacy of agents that target nuclear hormone receptors. In this review we discuss recent advances in our understanding of TFs as drug targets in oncology, with an emphasis on the emerging chemical approaches to modulate TF function. The remarkable diversity and potency of TFs as drivers of cell transformation justifies a continued pursuit of TFs as therapeutic targets for drug discovery. PMID:26645049

  11. Accurately measuring dynamic coefficient of friction in ultraform finishing

    NASA Astrophysics Data System (ADS)

    Briggs, Dennis; Echaves, Samantha; Pidgeon, Brendan; Travis, Nathan; Ellis, Jonathan D.

    2013-09-01

    UltraForm Finishing (UFF) is a deterministic sub-aperture computer numerically controlled grinding and polishing platform designed by OptiPro Systems. UFF is used to grind and polish a variety of optics from simple spherical to fully freeform, and numerous materials from glasses to optical ceramics. The UFF system consists of an abrasive belt around a compliant wheel that rotates and contacts the part to remove material. This work aims to accurately measure the dynamic coefficient of friction (μ), how it changes as a function of belt wear, and how this ultimately affects material removal rates. The coefficient of friction has been examined in terms of contact mechanics and Preston's equation to determine accurate material removal rates. By accurately predicting changes in μ, polishing iterations can be more accurately predicted, reducing the total number of iterations required to meet specifications. We have established an experimental apparatus that can accurately measure μ by measuring triaxial forces during translating loading conditions or while manufacturing the removal spots used to calculate material removal rates. Using this system, we will demonstrate μ measurements for UFF belts during different states of their lifecycle and assess the material removal function from spot diagrams as a function of wear. Ultimately, we will use this system for qualifying belt-wheel-material combinations to develop a spot-morphing model to better predict instantaneous material removal functions.

  12. Nonexposure Accurate Location K-Anonymity Algorithm in LBS

    PubMed Central

    2014-01-01

    This paper tackles location privacy protection in current location-based services (LBS) where mobile users have to report their exact location information to an LBS provider in order to obtain their desired services. Location cloaking has been proposed and well studied to protect user privacy. It blurs the user's accurate coordinate and replaces it with a well-shaped cloaked region. However, to obtain such an anonymous spatial region (ASR), nearly all existent cloaking algorithms require knowing the accurate locations of all users. Therefore, location cloaking without exposing the user's accurate location to any party is urgently needed. In this paper, we present such two nonexposure accurate location cloaking algorithms. They are designed for K-anonymity, and cloaking is performed based on the identifications (IDs) of the grid areas which were reported by all the users, instead of directly on their accurate coordinates. Experimental results show that our algorithms are more secure than the existent cloaking algorithms, need not have all the users reporting their locations all the time, and can generate smaller ASR. PMID:24605060

  13. Nonexposure accurate location K-anonymity algorithm in LBS.

    PubMed

    Jia, Jinying; Zhang, Fengli

    2014-01-01

    This paper tackles location privacy protection in current location-based services (LBS) where mobile users have to report their exact location information to an LBS provider in order to obtain their desired services. Location cloaking has been proposed and well studied to protect user privacy. It blurs the user's accurate coordinate and replaces it with a well-shaped cloaked region. However, to obtain such an anonymous spatial region (ASR), nearly all existent cloaking algorithms require knowing the accurate locations of all users. Therefore, location cloaking without exposing the user's accurate location to any party is urgently needed. In this paper, we present such two nonexposure accurate location cloaking algorithms. They are designed for K-anonymity, and cloaking is performed based on the identifications (IDs) of the grid areas which were reported by all the users, instead of directly on their accurate coordinates. Experimental results show that our algorithms are more secure than the existent cloaking algorithms, need not have all the users reporting their locations all the time, and can generate smaller ASR.

  14. Accurate and unambiguous tag-to-gene mapping in serial analysis of gene expression

    PubMed Central

    Malig, Rodrigo; Varela, Cristian; Agosin, Eduardo; Melo, Francisco

    2006-01-01

    Background In this study, we present a robust and reliable computational method for tag-to-gene assignment in serial analysis of gene expression (SAGE). The method relies on current genome information and annotation, incorporation of several new features, and key improvements over alternative methods, all of which are important to determine gene expression levels more accurately. The method provides a complete annotation of potential virtual SAGE tags within a genome, along with an estimation of their confidence for experimental observation that ranks tags that present multiple matches in the genome. Results We applied this method to the Saccharomyces cerevisiae genome, producing the most thorough and accurate annotation of potential virtual SAGE tags that is available today for this organism. The usefulness of this method is exemplified by the significant reduction of ambiguous cases in existing experimental SAGE data. In addition, we report new insights from the analysis of existing SAGE data. First, we found that experimental SAGE tags mapping onto introns, intron-exon boundaries, and non-coding RNA elements are observed in all available SAGE data. Second, a significant fraction of experimental SAGE tags was found to map onto genomic regions currently annotated as intergenic. Third, a significant number of existing experimental SAGE tags for yeast has been derived from truncated cDNAs, which are synthesized through oligo-d(T) priming to internal poly-(A) regions during reverse transcription. Conclusion We conclude that an accurate and unambiguous tag mapping process is essential to increase the quality and the amount of information that can be extracted from SAGE experiments. This is supported by the results obtained here and also by the large impact that the erroneous interpretation of these data could have on downstream applications. PMID:17083742

  15. An alternative transcription start site yields estrogen unresponsive Kiss1 mRNA transcripts in the hypothalamus of prepubertal female rats

    PubMed Central

    Castellano, Juan Manuel; Wright, Hollis; Ojeda, Sergio R.; Lomniczi, Alejandro

    2014-01-01

    The importance of the Kiss1 gene in the control of reproductive development is well documented. However, much less is known about the transcriptional regulation of Kiss1 expression in the hypothalamus. Critical for these studies is an accurate identification of the site(s) where Kiss1 transcription is initiated. Employing 5’-RACE PCR we detected a transcription start site (TSS1) used by the hypothalamus of rats, mice, nonhuman primates and humans to initiate Kiss1 transcription. In rodents, an exon 1 encoding 5’-untranslated sequences is followed by an alternatively spliced second exon, which encodes 5’-untranslated regions of two different lengths and contains the translation initiation codon (ATG). In nonhuman primates and humans exon 2 is not alternatively spliced. Surprisingly, in rat mediobasal hypothalamus (MBH), but not preoptic region (POA), an additional TSS (TSS2) located upstream from TSS1 generates an exon 1 longer (377 bp) than the TSS1-derived exon 1 (98 bp). The content of TSS1-derived transcripts increased at puberty in the POA and MBH of female rats. It also increased in the MBH after ovariectomy, and this change was prevented by estrogen. In contrast, no such changes in TSS2-derived transcript abundance were detected. Promoter assays showed that the proximal TSS1 promoter is much more active than the putative TSS2 promoter, and that only the TSS1 promoter is regulated by estrogen. These differences appear to be related to the presence of a TATA box and binding sites for transcription factors activating transcription and interacting with estrogen receptor alpha (ERα) in the TSS1, but not TSS2, promoter. PMID:24686008

  16. An information transmission model for transcription factor binding at regulatory DNA sites

    PubMed Central

    2012-01-01

    Background Computational identification of transcription factor binding sites (TFBSs) is a rapid, cost-efficient way to locate unknown regulatory elements. With increased potential for high-throughput genome sequencing, the availability of accurate computational methods for TFBS prediction has never been as important as it currently is. To date, identifying TFBSs with high sensitivity and specificity is still an open challenge, necessitating the development of novel models for predicting transcription factor-binding regulatory DNA elements. Results Based on the information theory, we propose a model for transcription factor binding of regulatory DNA sites. Our model incorporates position interdependencies in effective ways. The model computes the information transferred (TI) between the transcription factor and the TFBS during the binding process and uses TI as the criterion to determine whether the sequence motif is a possible TFBS. Based on this model, we developed a computational method to identify TFBSs. By theoretically proving and testing our model using both real and artificial data, we found that our model provides highly accurate predictive results. Conclusions In this study, we present a novel model for transcription factor binding regulatory DNA sites. The model can provide an increased ability to detect TFBSs. PMID:22672438

  17. Identification of reference genes suitable for normalization of RT-qPCR expression data in Saccharomyces cerevisiae during alcoholic fermentation.

    PubMed

    Vaudano, Enrico; Noti, Olta; Costantini, Antonella; Garcia-Moruno, Emilia

    2011-08-01

    Expression data from RT-qPCR (reverse transcription quantitative PCR) needs to be normalized to account for experimental variability among samples caused by differential yields of the transcripts in RNA extraction or in the reverse transcription. The most common method is to normalize against one or more reference genes (RG). We have selected RGs suitable for normalization of RT-qPCR raw data in Saccharomyces cerevisiae during alcoholic fermentation. The RGs were evaluated by three different statistical methods. The suitability of the selected RG sets was compared with ACT1, a commonly used non-validated single RG, by normalizing the expression of two target genes. Expression profiles of the target genes revealed the risk of misleading interpretation of expression data due to an unreliable RG.

  18. An efficient RNA isolation procedure and identification of reference genes for normalization of gene expression in blueberry.

    PubMed

    Vashisth, Tripti; Johnson, Lisa Klima; Malladi, Anish

    2011-12-01

    Application of transcriptomics approaches can greatly enhance our understanding of blueberry physiology. The success of transcriptomics approaches is dependent on the extraction of high-quality RNA which is complicated by the abundance of polyphenolics and polysaccharides in blueberry. Additionally, transcriptomics requires the accurate quantification of transcript abundance. Quantitative real-time polymerase chain reaction (qRT-PCR) is a robust method to determine transcript abundance. Normalization of gene expression using stably expressed reference genes is essential in qRT-PCR. An evaluation of the stability of expression of reference genes has not yet been reported in blueberry. The objectives of this study were to develop an effective procedure for extracting RNA from different organs and to evaluate potential reference genes for qRT-PCR analyses in blueberry. RNA of high quality and yield was extracted from eight and six organs of rabbiteye and southern highbush blueberry, respectively, using a modified cetyltrimethyl ammonium bromide-based method. The expression stability of 12 reference genes was evaluated. UBIQUITIN-CONJUGATING ENZYME (UBC28), RNA HELICASE-LIKE (RH8), CLATHRIN ADAPTER COMPLEXES MEDIUM SUBUNIT FAMILY PROTEIN (CACSa), and POLYUBIQUITIN (UBQ3b) were the most stably expressed genes across multiple organs in both blueberry species. Further, the expression stability of the reference genes in the branch abscission zone following treatment with fruit abscission-inducing compounds was analyzed. CACSa, RH8, and UBC28 were the most stably expressed genes in the abscission zone under abscission-inducing conditions. We suggest a preliminary evaluation of UBC28, CACSa, RH8, and UBQ3b to identify the most suitable reference genes for the experimental conditions under consideration in blueberry.

  19. Normalizing Catastrophe: Sustainability and Scientism

    ERIC Educational Resources Information Center

    Bonnett, Michael

    2013-01-01

    Making an adequate response to our deteriorating environmental situation is a matter of ever increasing urgency. It is argued that a central obstacle to achieving this is the way that scientism has become normalized in our thinking about environmental issues. This is taken to reflect on an underlying "metaphysics of mastery" that vitiates proper…

  20. Complete normal ordering 1: Foundations

    NASA Astrophysics Data System (ADS)

    Ellis, John; Mavromatos, Nick E.; Skliros, Dimitri P.

    2016-08-01

    We introduce a new prescription for quantising scalar field theories (in generic spacetime dimension and background) perturbatively around a true minimum of the full quantum effective action, which is to 'complete normal order' the bare action of interest. When the true vacuum of the theory is located at zero field value, the key property of this prescription is the automatic cancellation, to any finite order in perturbation theory, of all tadpole and, more generally, all 'cephalopod' Feynman diagrams. The latter are connected diagrams that can be disconnected into two pieces by cutting one internal vertex, with either one or both pieces free from external lines. In addition, this procedure of 'complete normal ordering' (which is an extension of the standard field theory definition of normal ordering) reduces by a substantial factor the number of Feynman diagrams to be calculated at any given loop order. We illustrate explicitly the complete normal ordering procedure and the cancellation of cephalopod diagrams in scalar field theories with non-derivative interactions, and by using a point splitting 'trick' we extend this result to theories with derivative interactions, such as those appearing as non-linear σ-models in the world-sheet formulation of string theory. We focus here on theories with trivial vacua, generalising the discussion to non-trivial vacua in a follow-up paper.

  1. Trojan dynamics well approximated by a new Hamiltonian normal form

    NASA Astrophysics Data System (ADS)

    Páez, Rocío Isabel; Locatelli, Ugo

    2015-10-01

    We revisit a classical perturbative approach to the Hamiltonian related to the motions of Trojan bodies, in the framework of the planar circular restricted three-body problem, by introducing a number of key new ideas in the formulation. In some sense, we adapt the approach of Garfinkel to the context of the normal form theory and its modern techniques. First, we make use of Delaunay variables for a physically accurate representation of the system. Therefore, we introduce a novel manipulation of the variables so as to respect the natural behaviour of the model. We develop a normalization procedure over the fast angle which exploits the fact that singularities in this model are essentially related to the slow angle. Thus, we produce a new normal form, i.e. an integrable approximation to the Hamiltonian. We emphasize some practical examples of the applicability of our normalizing scheme, e.g. the estimation of the stable libration region. Finally, we compare the level curves produced by our normal form with surfaces of section provided by the integration of the non-normalized Hamiltonian, with very good agreement. Further precision tests are also provided. In addition, we give a step-by-step description of the algorithm, allowing for extensions to more complicated models.

  2. Memory conformity affects inaccurate memories more than accurate memories.

    PubMed

    Wright, Daniel B; Villalba, Daniella K

    2012-01-01

    After controlling for initial confidence, inaccurate memories were shown to be more easily distorted than accurate memories. In two experiments groups of participants viewed 50 stimuli and were then presented with these stimuli plus 50 fillers. During this test phase participants reported their confidence that each stimulus was originally shown. This was followed by computer-generated responses from a bogus participant. After being exposed to this response participants again rated the confidence of their memory. The computer-generated responses systematically distorted participants' responses. Memory distortion depended on initial memory confidence, with uncertain memories being more malleable than confident memories. This effect was moderated by whether the participant's memory was initially accurate or inaccurate. Inaccurate memories were more malleable than accurate memories. The data were consistent with a model describing two types of memory (i.e., recollective and non-recollective memories), which differ in how susceptible these memories are to memory distortion.

  3. Accurate Fiber Length Measurement Using Time-of-Flight Technique

    NASA Astrophysics Data System (ADS)

    Terra, Osama; Hussein, Hatem

    2016-06-01

    Fiber artifacts of very well-measured length are required for the calibration of optical time domain reflectometers (OTDR). In this paper accurate length measurement of different fiber lengths using the time-of-flight technique is performed. A setup is proposed to measure accurately lengths from 1 to 40 km at 1,550 and 1,310 nm using high-speed electro-optic modulator and photodetector. This setup offers traceability to the SI unit of time, the second (and hence to meter by definition), by locking the time interval counter to the Global Positioning System (GPS)-disciplined quartz oscillator. Additionally, the length of a recirculating loop artifact is measured and compared with the measurement made for the same fiber by the National Physical Laboratory of United Kingdom (NPL). Finally, a method is proposed to relatively correct the fiber refractive index to allow accurate fiber length measurement.

  4. Differential equation based method for accurate approximations in optimization

    NASA Technical Reports Server (NTRS)

    Pritchard, Jocelyn I.; Adelman, Howard M.

    1990-01-01

    A method to efficiently and accurately approximate the effect of design changes on structural response is described. The key to this method is to interpret sensitivity equations as differential equations that may be solved explicitly for closed form approximations, hence, the method is denoted the Differential Equation Based (DEB) method. Approximations were developed for vibration frequencies, mode shapes and static displacements. The DEB approximation method was applied to a cantilever beam and results compared with the commonly-used linear Taylor series approximations and exact solutions. The test calculations involved perturbing the height, width, cross-sectional area, tip mass, and bending inertia of the beam. The DEB method proved to be very accurate, and in most cases, was more accurate than the linear Taylor series approximation. The method is applicable to simultaneous perturbation of several design variables. Also, the approximations may be used to calculate other system response quantities. For example, the approximations for displacements are used to approximate bending stresses.

  5. The utility of accurate mass and LC elution time information in the analysis of complex proteomes

    SciTech Connect

    Norbeck, Angela D.; Monroe, Matthew E.; Adkins, Joshua N.; Anderson, Kevin K.; Daly, Don S.; Smith, Richard D.

    2005-08-01

    Theoretical tryptic digests of all predicted proteins from the genomes of three organisms of varying complexity were evaluated for specificity and possible utility of combined peptide accurate mass and predicted LC normalized elution time (NET) information. The uniqueness of each peptide was evaluated using its combined mass (+/- 5 ppm and 1 ppm) and NET value (no constraint, +/- 0.05 and 0.01 on a 0-1 NET scale). The set of peptides both underestimates actual biological complexity due to the lack of specific modifications, and overestimates the expected complexity since many proteins will not be present in the sample or observable on the mass spectrometer because of dynamic range limitations. Once a peptide is identified from an LCMS/MS experiment, its mass and elution time is representative of a unique fingerprint for that peptide. The uniqueness of that fingerprint in comparison to that for the other peptides present is indicative of the ability to confidently identify that peptide based on accurate mass and NET measurements. These measurements can be made using HPLC coupled with high resolution MS in a high-throughput manner. Results show that for organisms with comparatively small proteomes, such as Deinococcus radiodurans, modest mass and elution time accuracies are generally adequate for peptide identifications. For more complex proteomes, increasingly accurate easurements are required. However, the majority of proteins should be uniquely identifiable by using LC-MS with mass accuracies within +/- 1 ppm and elution time easurements within +/- 0.01 NET.

  6. Whole muscle length-tension relationships are accurately modeled as scaled sarcomeres in rabbit hindlimb muscles.

    PubMed

    Winters, Taylor M; Takahashi, Mitsuhiko; Lieber, Richard L; Ward, Samuel R

    2011-01-04

    An a priori model of the whole active muscle length-tension relationship was constructed utilizing only myofilament length and serial sarcomere number for rabbit tibialis anterior (TA), extensor digitorum longus (EDL), and extensor digitorum II (EDII) muscles. Passive tension was modeled with a two-element Hill-type model. Experimental length-tension relations were then measured for each of these muscles and compared to predictions. The model was able to accurately capture the active-tension characteristics of experimentally-measured data for all muscles (ICC=0.88 ± 0.03). Despite their varied architecture, no differences in predicted versus experimental correlations were observed among muscles. In addition, the model demonstrated that excursion, quantified by full-width-at-half-maximum (FWHM) of the active length-tension relationship, scaled linearly (slope=0.68) with normalized muscle fiber length. Experimental and theoretical FWHM values agreed well with an intraclass correlation coefficient of 0.99 (p<0.001). In contrast to active tension, the passive tension model deviated from experimentally-measured values and thus, was not an accurate predictor of passive tension (ICC=0.70 ± 0.07). These data demonstrate that modeling muscle as a scaled sarcomere provides accurate active functional but not passive functional predictions for rabbit TA, EDL, and EDII muscles and call into question the need for more complex modeling assumptions often proposed.

  7. Modular composition of gene transcription networks.

    PubMed

    Gyorgy, Andras; Del Vecchio, Domitilla

    2014-03-01

    Predicting the dynamic behavior of a large network from that of the composing modules is a central problem in systems and synthetic biology. Yet, this predictive ability is still largely missing because modules display context-dependent behavior. One cause of context-dependence is retroactivity, a phenomenon similar to loading that influences in non-trivial ways the dynamic performance of a module upon connection to other modules. Here, we establish an analysis framework for gene transcription networks that explicitly accounts for retroactivity. Specifically, a module's key properties are encoded by three retroactivity matrices: internal, scaling, and mixing retroactivity. All of them have a physical interpretation and can be computed from macroscopic parameters (dissociation constants and promoter concentrations) and from the modules' topology. The internal retroactivity quantifies the effect of intramodular connections on an isolated module's dynamics. The scaling and mixing retroactivity establish how intermodular connections change the dynamics of connected modules. Based on these matrices and on the dynamics of modules in isolation, we can accurately predict how loading will affect the behavior of an arbitrary interconnection of modules. We illustrate implications of internal, scaling, and mixing retroactivity on the performance of recurrent network motifs, including negative autoregulation, combinatorial regulation, two-gene clocks, the toggle switch, and the single-input motif. We further provide a quantitative metric that determines how robust the dynamic behavior of a module is to interconnection with other modules. This metric can be employed both to evaluate the extent of modularity of natural networks and to establish concrete design guidelines to minimize retroactivity between modules in synthetic systems.

  8. 10 CFR 9.108 - Certification, transcripts, recordings and minutes.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ...). Copies of such transcript, or minutes, or a transcription of such recording disclosing the identity of... transcription as provided in § 9.14. The Secretary shall maintain a complete verbatim copy of the transcript,...

  9. Extracting Time-Accurate Acceleration Vectors From Nontrivial Accelerometer Arrangements.

    PubMed

    Franck, Jennifer A; Blume, Janet; Crisco, Joseph J; Franck, Christian

    2015-09-01

    Sports-related concussions are of significant concern in many impact sports, and their detection relies on accurate measurements of the head kinematics during impact. Among the most prevalent recording technologies are videography, and more recently, the use of single-axis accelerometers mounted in a helmet, such as the HIT system. Successful extraction of the linear and angular impact accelerations depends on an accurate analysis methodology governed by the equations of motion. Current algorithms are able to estimate the magnitude of acceleration and hit location, but make assumptions about the hit orientation and are often limited in the position and/or orientation of the accelerometers. The newly formulated algorithm presented in this manuscript accurately extracts the full linear and rotational acceleration vectors from a broad arrangement of six single-axis accelerometers directly from the governing set of kinematic equations. The new formulation linearizes the nonlinear centripetal acceleration term with a finite-difference approximation and provides a fast and accurate solution for all six components of acceleration over long time periods (>250 ms). The approximation of the nonlinear centripetal acceleration term provides an accurate computation of the rotational velocity as a function of time and allows for reconstruction of a multiple-impact signal. Furthermore, the algorithm determines the impact location and orientation and can distinguish between glancing, high rotational velocity impacts, or direct impacts through the center of mass. Results are shown for ten simulated impact locations on a headform geometry computed with three different accelerometer configurations in varying degrees of signal noise. Since the algorithm does not require simplifications of the actual impacted geometry, the impact vector, or a specific arrangement of accelerometer orientations, it can be easily applied to many impact investigations in which accurate kinematics need to

  10. Informational Requirements for Transcriptional Regulation

    PubMed Central

    O'Neill, Patrick K.; Forder, Robert

    2014-01-01

    Abstract Transcription factors (TFs) regulate transcription by binding to specific sites in promoter regions. Information theory provides a useful mathematical framework to analyze the binding motifs associated with TFs but imposes several assumptions that limit their applicability to specific regulatory scenarios. Explicit simulations of the co-evolution of TFs and their binding motifs allow the study of the evolution of regulatory networks with a high degree of realism. In this work we analyze the impact of differential regulatory demands on the information content of TF-binding motifs by means of evolutionary simulations. We generalize a predictive index based on information theory, and we validate its applicability to regulatory scenarios in which the TF binds significantly to the genomic background. Our results show a logarithmic dependence of the evolved information content on the occupancy of target sites and indicate that TFs may actively exploit pseudo-sites to modulate their occupancy of target sites. In regulatory networks with differentially regulated targets, we observe that information content in TF-binding motifs is dictated primarily by the fraction of total probability mass that the TF assigns to its target sites, and we provide a predictive index to estimate the amount of information associated with arbitrarily complex regulatory systems. We observe that complex regulatory patterns can exert additional demands on evolved information content, but, given a total occupancy for target sites, we do not find conclusive evidence that this effect is because of the range of required binding affinities. PMID:24689750

  11. Gene transcription and electromagnetic fields

    SciTech Connect

    Henderson, A.S.

    1992-01-01

    Our overall aim is to obtain sufficient information to allow us to ultimately determine whether ELF EM field exposure is an initiating factor in neoplastic transformation and/or if exposure can mimic characteristics of the second-step counterpart in neoplastic disease. This aim is based on our previous findings that levels of some transcripts are increased in cells exposed to EM fields. While the research is basic in nature, the ramifications have bearing on the general safety of exposure to EM fields in industrial and everyday life. A large array of diverse biological effects are reported to occur as the result of exposure to elf EM fields, suggesting that the cell response to EM fields is at a basic level, presumably initiated by molecular and/or biophysical events at the cell membrane. The hypothesized route is a signal transduction pathway involving membrane calcium fluxes. Information flow resulting from signal transduction can mediate the induction of regulatory factors in the cell, and directly affect how transcription is regulated.

  12. Must Kohn-Sham oscillator strengths be accurate at threshold?

    SciTech Connect

    Yang Zenghui; Burke, Kieron; Faassen, Meta van

    2009-09-21

    The exact ground-state Kohn-Sham (KS) potential for the helium atom is known from accurate wave function calculations of the ground-state density. The threshold for photoabsorption from this potential matches the physical system exactly. By carefully studying its absorption spectrum, we show the answer to the title question is no. To address this problem in detail, we generate a highly accurate simple fit of a two-electron spectrum near the threshold, and apply the method to both the experimental spectrum and that of the exact ground-state Kohn-Sham potential.

  13. Accurate torque-speed performance prediction for brushless dc motors

    NASA Astrophysics Data System (ADS)

    Gipper, Patrick D.

    Desirable characteristics of the brushless dc motor (BLDCM) have resulted in their application for electrohydrostatic (EH) and electromechanical (EM) actuation systems. But to effectively apply the BLDCM requires accurate prediction of performance. The minimum necessary performance characteristics are motor torque versus speed, peak and average supply current and efficiency. BLDCM nonlinear simulation software specifically adapted for torque-speed prediction is presented. The capability of the software to quickly and accurately predict performance has been verified on fractional to integral HP motor sizes, and is presented. Additionally, the capability of torque-speed prediction with commutation angle advance is demonstrated.

  14. Accurate upwind-monotone (nonoscillatory) methods for conservation laws

    NASA Technical Reports Server (NTRS)

    Huynh, Hung T.

    1992-01-01

    The well known MUSCL scheme of Van Leer is constructed using a piecewise linear approximation. The MUSCL scheme is second order accurate at the smooth part of the solution except at extrema where the accuracy degenerates to first order due to the monotonicity constraint. To construct accurate schemes which are free from oscillations, the author introduces the concept of upwind monotonicity. Several classes of schemes, which are upwind monotone and of uniform second or third order accuracy are then presented. Results for advection with constant speed are shown. It is also shown that the new scheme compares favorably with state of the art methods.

  15. In-line sensor for accurate rf power measurements

    NASA Astrophysics Data System (ADS)

    Gahan, D.; Hopkins, M. B.

    2005-10-01

    An in-line sensor has been constructed with 50Ω characteristic impedance to accurately measure rf power dissipated in a matched or unmatched load with a view to being implemented as a rf discharge diagnostic. The physical construction and calibration technique are presented. The design is a wide band, hybrid directional coupler/current-voltage sensor suitable for fundamental and harmonic power measurements. A comparison with a standard wattmeter using dummy load impedances shows that this in-line sensor is significantly more accurate in mismatched conditions.

  16. In-line sensor for accurate rf power measurements

    SciTech Connect

    Gahan, D.; Hopkins, M.B.

    2005-10-15

    An in-line sensor has been constructed with 50 {omega} characteristic impedance to accurately measure rf power dissipated in a matched or unmatched load with a view to being implemented as a rf discharge diagnostic. The physical construction and calibration technique are presented. The design is a wide band, hybrid directional coupler/current-voltage sensor suitable for fundamental and harmonic power measurements. A comparison with a standard wattmeter using dummy load impedances shows that this in-line sensor is significantly more accurate in mismatched conditions.

  17. Time-Accurate Numerical Simulations of Synthetic Jet Quiescent Air

    NASA Technical Reports Server (NTRS)

    Rupesh, K-A. B.; Ravi, B. R.; Mittal, R.; Raju, R.; Gallas, Q.; Cattafesta, L.

    2007-01-01

    The unsteady evolution of three-dimensional synthetic jet into quiescent air is studied by time-accurate numerical simulations using a second-order accurate mixed explicit-implicit fractional step scheme on Cartesian grids. Both two-dimensional and three-dimensional calculations of synthetic jet are carried out at a Reynolds number (based on average velocity during the discharge phase of the cycle V(sub j), and jet width d) of 750 and Stokes number of 17.02. The results obtained are assessed against PIV and hotwire measurements provided for the NASA LaRC workshop on CFD validation of synthetic jets.

  18. Expression of splice variants of mts1 gene in normal and neoplastic human tissues

    SciTech Connect

    Ambartsumyan, N.S. |; Grigorian, M.S.; Lukanidin, E.M.

    1995-09-01

    Data on cloning of cDNA corresponding to human mts1 gene transcripts are presented. By comparing nucleotide sequences of the genomic DNA clone and cDNA of mts1, it was shown that human osteosarcoma OHS cells contain two alternative splice variants of mts1 transcripts. Alternative splicing occurs in the 5{prime}-untranslated region of the mts1 pre-mRNA. Both splice variants, hu-mts1 and hu-mts1(var), demonstrate similar stability in the cells, and each contains one open reading frame for the MTS1 protein. However, the two types of transcripts are translated with different effectiveness. The level of transcription of mts1 splice variants in different normal and neoplastic tissues and cell lines varies significantly. The role of alternative splicing as the mechanism responsible for posttranscriptional regulation of mts1 gene expression is discussed. 31 refs., 5 figs.

  19. Reverse transcription quantitative real-time polymerase chain reaction reference genes in the spared nerve injury model of neuropathic pain: validation and literature search

    PubMed Central

    2013-01-01

    Background The reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) is a widely used, highly sensitive laboratory technique to rapidly and easily detect, identify and quantify gene expression. Reliable RT-qPCR data necessitates accurate normalization with validated control genes (reference genes) whose expression is constant in all studied conditions. This stability has to be demonstrated. We performed a literature search for studies using quantitative or semi-quantitative PCR in the rat spared nerve injury (SNI) model of neuropathic pain to verify whether any reference genes had previously been validated. We then analyzed the stability over time of 7 commonly used reference genes in the nervous system – specifically in the spinal cord dorsal horn and the dorsal root ganglion (DRG). These were: Actin beta (Actb), Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), ribosomal proteins 18S (18S), L13a (RPL13a) and L29 (RPL29), hypoxanthine phosphoribosyltransferase 1 (HPRT1) and hydroxymethylbilane synthase (HMBS). We compared the candidate genes and established a stability ranking using the geNorm algorithm. Finally, we assessed the number of reference genes necessary for accurate normalization in this neuropathic pain model. Results We found GAPDH, HMBS, Actb, HPRT1 and 18S cited as reference genes in literature on studies using the SNI model. Only HPRT1 and 18S had been once previously demonstrated as stable in RT-qPCR arrays. All the genes tested in this study, using the geNorm algorithm, presented gene stability values (M-value) acceptable enough for them to qualify as potential reference genes in both DRG and spinal cord. Using the coefficient of variation, 18S failed the 50% cut-off with a value of 61% in the DRG. The two most stable genes in the dorsal horn were RPL29 and RPL13a; in the DRG they were HPRT1 and Actb. Using a 0.15 cut-off for pairwise variations we found that any pair of stable reference gene was sufficient for the

  20. Statistical treatment of fluorescence in situ hybridization validation data to generate normal reference ranges using Excel functions.

    PubMed

    Ciolino, Allison L; Tang, Mary E; Bryant, Ron

    2009-07-01

    Fluorescent in situ hybridization has become an essential tool for diagnosing and monitoring hematological disease. Testing for minimal residual disease requires precise and accurate normal cut-offs. There is no consensus in the field on the correct method of establishing a normal reference range. We discuss and compare several proposed statistical methods to calculate normal reference ranges, including Gaussian statistics, the beta inverse function, and a binomial treatment of the data. We demonstrate that a binomial treatment of the data is an accurate and simple method to calculate a normal reference range.

  1. Mitotic Transcriptional Activation: Clearance of Actively Engaged Pol II via Transcriptional Elongation Control in Mitosis.

    PubMed

    Liang, Kaiwei; Woodfin, Ashley R; Slaughter, Brian D; Unruh, Jay R; Box, Andrew C; Rickels, Ryan A; Gao, Xin; Haug, Jeffrey S; Jaspersen, Sue L; Shilatifard, Ali

    2015-11-05

    Although it is established that some general transcription factors are inactivated at mitosis, many details of mitotic transcription inhibition (MTI) and its underlying mechanisms are largely unknown. We have identified mitotic transcriptional activation (MTA) as a key regulatory step to control transcription in mitosis for genes with transcriptionally engaged RNA polymerase II (Pol II) to activate and transcribe until the end of the gene to clear Pol II from mitotic chromatin, followed by global impairment of transcription reinitiation through MTI. Global nascent RNA sequencing and RNA fluorescence in situ hybridization demonstrate the existence of transcriptionally engaged Pol II in early mitosis. Both genetic and chemical inhibition of P-TEFb in mitosis lead to delays in the progression of cell division. Together, our study reveals a mechanism for MTA and MTI whereby transcriptionally engaged Pol II can progress into productive elongation and finish transcription to allow proper cellular division.

  2. Quantification of co-transcriptional splicing from RNA-Seq data.

    PubMed

    Herzel, Lydia; Neugebauer, Karla M

    2015-09-01

    During gene expression, protein-coding transcripts are shaped by multiple processing events: 5' end capping, pre-mRNA splicing, RNA editing, and 3' end cleavage and polyadenylation. These events are required to produce mature mRNA, which can be subsequently translated. Nearly all of these RNA processing steps occur during transcription, while the nascent RNA is still attached to the DNA template by RNA polymerase II (i.e. co-transcriptionally). Polyadenylation occurs after 3' end cleavage or post-transcriptionally. Pre-mRNA splicing - the removal of introns and ligation of exons - can be initiated and concluded co-transcriptionally, although this is not strictly required. Recently, a number of studies using global methods have shown that the majority of splicing is co-transcriptional, yet not all published studies agree in their conclusions. Short read sequencing of RNA (RNA-Seq) is the prevailing approach to measuring splicing levels in nascent RNA, mRNA or total RNA. Here, we compare four different strategies for analyzing and quantifying co-transcriptional splicing. To do so, we reanalyze two nascent RNA-Seq datasets of the same species, but different cell type and RNA isolation procedure. Average co-transcriptional splicing values calculated on a per intron basis are similar, independent of the strategy used. We emphasize the technical requirements for identifying co-transcriptional splicing events with high confidence, e.g. how to calculate co-transcriptional splicing from nascent RNA- versus mRNA-Seq data, the number of biological replicates needed, depletion of polyA+RNA, and appropriate normalization. Finally, we present guidelines for planning a nascent RNA-Seq experiment.

  3. The transcriptional programme of the androgen receptor (AR) in prostate cancer.

    PubMed

    Lamb, Alastair D; Massie, Charlie E; Neal, David E

    2014-03-01

    The androgen receptor (AR) is essential for normal prostate and prostate cancer cell growth. AR transcriptional activity is almost always maintained even in hormone relapsed prostate cancer (HRPC) in the absence of normal levels of circulating testosterone. Current molecular techniques, such as chromatin-immunoprecipitation sequencing (ChIP-seq), have permitted identification of direct AR-binding sites in cell lines and human tissue with a distinct coordinate network evident in HRPC. The effectiveness of novel agents, such as abiraterone acetate (suppresses adrenal androgens) or enzalutamide (MDV3100, potent AR antagonist), in treating advanced prostate cancer underlines the on-going critical role of the AR throughout all stages of the disease. Persistent AR activity in advanced disease regulates cell cycle activity, steroid biosynthesis and anabolic metabolism in conjunction with regulatory co-factors, such as the E2F family, c-Myc and signal transducer and activator of transcription (STAT) transcription factors. Further treatment approaches must target these other factors.

  4. Validation and Comparison of Reference Genes for qPCR Normalization of Celery (Apium graveolens) at Different Development Stages.

    PubMed

    Li, Meng-Yao; Wang, Feng; Jiang, Qian; Wang, Guan-Long; Tian, Chang; Xiong, Ai-Sheng

    2016-01-01

    A suitable reference gene is an important prerequisite for guarantying accurate and reliable results in qPCR analysis. Celery is one of the representative vegetable in Apiaceae and is widely cultivated and consumed in the world. However, no reports have been previously published concerning reference genes in celery. In this study, the expression stabilities of nine candidate reference genes in leaf blade and petiole at different development stages were evaluated using three statistics algorithms geNorm, NormFinder, and BestKeeper. Our results showed that TUB-B, TUB-A, and UBC were the most reference genes among all tested samples. GAPDH represented the maximum stability for most individual sample, while the UBQ displayed the minimum stability. To further validate the stability of reference genes, the expression pattern of AgAP2-2 was calculated by using the selected genes for normalization. In addition, the expression patterns of several development-related genes were studied using the selected reference gene. Our results will be beneficial for further studies on gene transcription in celery.

  5. Correcting Transcription Factor Gene Sets for Copy Number and Promoter Methylation Variations

    PubMed Central

    Rathi, Komal S.; Gaykalova, Daria A.; Hennesey, Patrick; Califano, Joseph A.; Ochs, Michael F.

    2014-01-01

    Gene set analysis provides a method to generate statistical inferences across sets of linked genes, primarily using high-throughput expression data. Common gene sets include biological pathways, operons, and targets of transcriptional regulators. In higher eukaryotes, especially when dealing with diseases with strong genetic and epigenetic components such as cancer, copy number loss and gene silencing through promoter methylation can eliminate the possibility that a gene is transcribed. This, in turn, can adversely affect the estimation of transcription factor or pathway activity from a set of target genes, since some of the targets may not be responsive to transcriptional regulation. Here we introduce a simple filtering approach that removes genes from consideration if they show copy number loss or promoter methylation and demonstrate the improvement in inference of transcription factor activity in a simulated data set based on the background expression observed in normal head and neck tissue. PMID:25195578

  6. Correcting transcription factor gene sets for copy number and promoter methylation variations.

    PubMed

    Rathi, Komal S; Gaykalova, Daria A; Hennessey, Patrick; Califano, Joseph A; Ochs, Michael F

    2014-09-01

    Gene set analysis provides a method to generate statistical inferences across sets of linked genes, primarily using high-throughput expression data. Common gene sets include biological pathways, operons, and targets of transcriptional regulators. In higher eukaryotes, especially when dealing with diseases with strong genetic and epigenetic components such as cancer, copy number loss and gene silencing through promoter methylation can eliminate the possibility that a gene is transcribed. This, in turn, can adversely affect the estimation of transcription factor or pathway activity from a set of target genes, as some of the targets may not be responsive to transcriptional regulation. Here we introduce a simple filtering approach that removes genes from consideration if they show copy number loss or promoter methylation, and demonstrate the improvement in inference of transcription factor activity in a simulated dataset based on the background expression observed in normal head and neck tissue.

  7. Trypanosoma brucei: Enrichment by UV of intergenic transcripts from the variable surface glycoprotein gene expression site

    SciTech Connect

    Coquelet, H.; Tebabi, P.; Pays, A.; Steinert, M.; Pays, E. )

    1989-09-01

    The expression site for the variable surface glycoprotein (VSG) gene AnTat 1.3A of Trypanosoma brucei is 45 kilobases long and encompasses seven expression site-associated genes (ESAGs). After UV irradiation, several large transcripts from the putative promoter region were strongly enriched. We report that one such major transcript starts near the poly(A) addition site of the first gene (ESAG 7), spans the intergenic region, and extends to the poly(A) addition site of the second gene (ESAG 6), thus bypassing the normal 3' splice site of the ESAG 6 mRNA. Since this transcript is spliced, we conclude that UV irradiation does not inhibit splicing but stabilizes unstable processing products. This demonstrates that at least some intergenic regions of the VSG gene expression site are continuously transcribed in accordance with a polycistronic transcription model.

  8. Dominance and interloci interactions in transcriptional activation cascades: models explaining compensatory mutations and inheritance patterns.

    PubMed

    Bost, Bruno; Veitia, Reiner A

    2014-01-01

    Mutations in human genes encoding transcription factors are often dominant because one active allele cannot ensure a normal phenotype (haploinsufficiency). In other instances, heterozygous mutations of two genes are required for a phenotype to appear (combined haploinsufficiency). Here, we explore with models (i) the basis of haploinsufficiency and combined haploinsufficiency owing to mutations in transcription activators, and (ii) how the effects of such mutations can be amplified or buffered by subsequent steps in a transcription cascade. We propose that the non-linear (sigmoidal) response of transcription to the concentration of activators can explain haploinsufficiency. We further show that the sigmoidal character of the output of a cascade increases with the number of steps involved, the settings of which will determine the buffering or enhancement of the effects of a decreased concentration of an upstream activator. This exploration provides insights into the bases of compensatory mutations and on interloci interactions underlying oligogenic inheritance patterns.

  9. BIOACCESSIBILITY TESTS ACCURATELY ESTIMATE BIOAVAILABILITY OF LEAD TO QUAIL

    EPA Science Inventory

    Hazards of soil-borne Pb to wild birds may be more accurately quantified if the bioavailability of that Pb is known. To better understand the bioavailability of Pb to birds, we measured blood Pb concentrations in Japanese quail (Coturnix japonica) fed diets containing Pb-contami...

  10. Device accurately measures and records low gas-flow rates

    NASA Technical Reports Server (NTRS)

    Branum, L. W.

    1966-01-01

    Free-floating piston in a vertical column accurately measures and records low gas-flow rates. The system may be calibrated, using an adjustable flow-rate gas supply, a low pressure gage, and a sequence recorder. From the calibration rates, a nomograph may be made for easy reduction. Temperature correction may be added for further accuracy.

  11. Ultrasonic system for accurate distance measurement in the air.

    PubMed

    Licznerski, Tomasz J; Jaroński, Jarosław; Kosz, Dariusz

    2011-12-01

    This paper presents a system that accurately measures the distance travelled by ultrasound waves through the air. The simple design of the system and its obtained accuracy provide a tool for non-contact distance measurements required in the laser's optical system that investigates the surface of the eyeball.

  12. A Self-Instructional Device for Conditioning Accurate Prosody.

    ERIC Educational Resources Information Center

    Buiten, Roger; Lane, Harlan

    1965-01-01

    A self-instructional device for conditioning accurate prosody in second-language learning is described in this article. The Speech Auto-Instructional Device (SAID) is electro-mechanical and performs three functions: SAID (1) presents to the student tape-recorded pattern sentences that are considered standards in prosodic performance; (2) processes…

  13. Monitoring circuit accurately measures movement of solenoid valve

    NASA Technical Reports Server (NTRS)

    Gillett, J. D.

    1966-01-01

    Solenoid operated valve in a control system powered by direct current issued to accurately measure the valve travel. This system is currently in operation with a 28-vdc power system used for control of fluids in liquid rocket motor test facilities.

  14. Instrument accurately measures small temperature changes on test surface

    NASA Technical Reports Server (NTRS)

    Harvey, W. D.; Miller, H. B.

    1966-01-01

    Calorimeter apparatus accurately measures very small temperature rises on a test surface subjected to aerodynamic heating. A continuous thin sheet of a sensing material is attached to a base support plate through which a series of holes of known diameter have been drilled for attaching thermocouples to the material.

  15. A Simple and Accurate Method for Measuring Enzyme Activity.

    ERIC Educational Resources Information Center

    Yip, Din-Yan

    1997-01-01

    Presents methods commonly used for investigating enzyme activity using catalase and presents a new method for measuring catalase activity that is more reliable and accurate. Provides results that are readily reproduced and quantified. Can also be used for investigations of enzyme properties such as the effects of temperature, pH, inhibitors,…

  16. Bioaccessibility tests accurately estimate bioavailability of lead to quail

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Hazards of soil-borne Pb to wild birds may be more accurately quantified if the bioavailability of that Pb is known. To better understand the bioavailability of Pb, we incorporated Pb-contaminated soils or Pb acetate into diets for Japanese quail (Coturnix japonica), fed the quail for 15 days, and ...

  17. Second-order accurate nonoscillatory schemes for scalar conservation laws

    NASA Technical Reports Server (NTRS)

    Huynh, Hung T.

    1989-01-01

    Explicit finite difference schemes for the computation of weak solutions of nonlinear scalar conservation laws is presented and analyzed. These schemes are uniformly second-order accurate and nonoscillatory in the sense that the number of extrema of the discrete solution is not increasing in time.

  18. Foresight begins with FMEA. Delivering accurate risk assessments.

    PubMed

    Passey, R D

    1999-03-01

    If sufficient factors are taken into account and two- or three-stage analysis is employed, failure mode and effect analysis represents an excellent technique for delivering accurate risk assessments for products and processes, and for relating them to legal liability. This article describes a format that facilitates easy interpretation.

  19. How Accurate Are Judgments of Intelligence by Strangers?

    ERIC Educational Resources Information Center

    Borkenau, Peter

    Whether judgments made by complete strangers as to the intelligence of subjects are accurate or merely illusory was studied in Germany. Target subjects were 50 female and 50 male adults recruited through a newspaper article. Eighteen judges, who did not know the subjects, were recruited from a university community. Videorecordings of the subjects,…

  20. Quantifying Accurate Calorie Estimation Using the "Think Aloud" Method

    ERIC Educational Resources Information Center

    Holmstrup, Michael E.; Stearns-Bruening, Kay; Rozelle, Jeffrey

    2013-01-01

    Objective: Clients often have limited time in a nutrition education setting. An improved understanding of the strategies used to accurately estimate calories may help to identify areas of focused instruction to improve nutrition knowledge. Methods: A "Think Aloud" exercise was recorded during the estimation of calories in a standard dinner meal…