Comparative genomics of metabolic capacities of regulons controlled by cis-regulatory RNA motifs in bacteria.

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Sun, Eric I; Leyn, Semen A; Kazanov, Marat D; Saier, Milton H; Novichkov, Pavel S; Rodionov, Dmitry A

2013-09-02

In silico comparative genomics approaches have been efficiently used for functional prediction and reconstruction of metabolic and regulatory networks. Riboswitches are metabolite-sensing structures often found in bacterial mRNA leaders controlling gene expression on transcriptional or translational levels.An increasing number of riboswitches and other cis-regulatory RNAs have been recently classified into numerous RNA families in the Rfam database. High conservation of these RNA motifs provides a unique advantage for their genomic identification and comparative analysis. A comparative genomics approach implemented in the RegPredict tool was used for reconstruction and functional annotation of regulons controlled by RNAs from 43 Rfam families in diverse taxonomic groups of Bacteria. The inferred regulons include ~5200 cis-regulatory RNAs and more than 12000 target genes in 255 microbial genomes. All predicted RNA-regulated genes were classified into specific and overall functional categories. Analysis of taxonomic distribution of these categories allowed us to establish major functional preferences for each analyzed cis-regulatory RNA motif family. Overall, most RNA motif regulons showed predictable functional content in accordance with their experimentally established effector ligands. Our results suggest that some RNA motifs (including thiamin pyrophosphate and cobalamin riboswitches that control the cofactor metabolism) are widespread and likely originated from the last common ancestor of all bacteria. However, many more analyzed RNA motifs are restricted to a narrow taxonomic group of bacteria and likely represent more recent evolutionary innovations. The reconstructed regulatory networks for major known RNA motifs substantially expand the existing knowledge of transcriptional regulation in bacteria. The inferred regulons can be used for genetic experiments, functional annotations of genes, metabolic reconstruction and evolutionary analysis. The obtained genome

On the Concept of Cis-regulatory Information: From Sequence Motifs to Logic Functions

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Tarpine, Ryan; Istrail, Sorin

The regulatory genome is about the “system level organization of the core genomic regulatory apparatus, and how this is the locus of causality underlying the twin phenomena of animal development and animal evolution” (E.H. Davidson. The Regulatory Genome: Gene Regulatory Networks in Development and Evolution, Academic Press, 2006). Information processing in the regulatory genome is done through regulatory states, defined as sets of transcription factors (sequence-specific DNA binding proteins which determine gene expression) that are expressed and active at the same time. The core information processing machinery consists of modular DNA sequence elements, called cis-modules, that interact with transcription factors. The cis-modules “read” the information contained in the regulatory state of the cell through transcription factor binding, “process” it, and directly or indirectly communicate with the basal transcription apparatus to determine gene expression. This endowment of each gene with the information-receiving capacity through their cis-regulatory modules is essential for the response to every possible regulatory state to which it might be exposed during all phases of the life cycle and in all cell types. We present here a set of challenges addressed by our CYRENE research project aimed at studying the cis-regulatory code of the regulatory genome. The CYRENE Project is devoted to (1) the construction of a database, the cis-Lexicon, containing comprehensive information across species about experimentally validated cis-regulatory modules; and (2) the software development of a next-generation genome browser, the cis-Browser, specialized for the regulatory genome. The presentation is anchored on three main computational challenges: the Gene Naming Problem, the Consensus Sequence Bottleneck Problem, and the Logic Function Inference Problem.

Patterns of cis regulatory variation in diverse human populations.

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Barbara E Stranger

Full Text Available The genetic basis of gene expression variation has long been studied with the aim to understand the landscape of regulatory variants, but also more recently to assist in the interpretation and elucidation of disease signals. To date, many studies have looked in specific tissues and population-based samples, but there has been limited assessment of the degree of inter-population variability in regulatory variation. We analyzed genome-wide gene expression in lymphoblastoid cell lines from a total of 726 individuals from 8 global populations from the HapMap3 project and correlated gene expression levels with HapMap3 SNPs located in cis to the genes. We describe the influence of ancestry on gene expression levels within and between these diverse human populations and uncover a non-negligible impact on global patterns of gene expression. We further dissect the specific functional pathways differentiated between populations. We also identify 5,691 expression quantitative trait loci (eQTLs after controlling for both non-genetic factors and population admixture and observe that half of the cis-eQTLs are replicated in one or more of the populations. We highlight patterns of eQTL-sharing between populations, which are partially determined by population genetic relatedness, and discover significant sharing of eQTL effects between Asians, European-admixed, and African subpopulations. Specifically, we observe that both the effect size and the direction of effect for eQTLs are highly conserved across populations. We observe an increasing proximity of eQTLs toward the transcription start site as sharing of eQTLs among populations increases, highlighting that variants close to TSS have stronger effects and therefore are more likely to be detected across a wider panel of populations. Together these results offer a unique picture and resource of the degree of differentiation among human populations in functional regulatory variation and provide an estimate for

Alignment and prediction of cis-regulatory modules based on a probabilistic model of evolution.

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Xin He

2009-03-01

Full Text Available Cross-species comparison has emerged as a powerful paradigm for predicting cis-regulatory modules (CRMs and understanding their evolution. The comparison requires reliable sequence alignment, which remains a challenging task for less conserved noncoding sequences. Furthermore, the existing models of DNA sequence evolution generally do not explicitly treat the special properties of CRM sequences. To address these limitations, we propose a model of CRM evolution that captures different modes of evolution of functional transcription factor binding sites (TFBSs and the background sequences. A particularly novel aspect of our work is a probabilistic model of gains and losses of TFBSs, a process being recognized as an important part of regulatory sequence evolution. We present a computational framework that uses this model to solve the problems of CRM alignment and prediction. Our alignment method is similar to existing methods of statistical alignment but uses the conserved binding sites to improve alignment. Our CRM prediction method deals with the inherent uncertainties of binding site annotations and sequence alignment in a probabilistic framework. In simulated as well as real data, we demonstrate that our program is able to improve both alignment and prediction of CRM sequences over several state-of-the-art methods. Finally, we used alignments produced by our program to study binding site conservation in genome-wide binding data of key transcription factors in the Drosophila blastoderm, with two intriguing results: (i the factor-bound sequences are under strong evolutionary constraints even if their neighboring genes are not expressed in the blastoderm and (ii binding sites in distal bound sequences (relative to transcription start sites tend to be more conserved than those in proximal regions. Our approach is implemented as software, EMMA (Evolutionary Model-based cis-regulatory Module Analysis, ready to be applied in a broad biological context.

Functional evolution of cis-regulatory modules at a homeotic gene in Drosophila.

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Margaret C W Ho

2009-11-01

Full Text Available It is a long-held belief in evolutionary biology that the rate of molecular evolution for a given DNA sequence is inversely related to the level of functional constraint. This belief holds true for the protein-coding homeotic (Hox genes originally discovered in Drosophila melanogaster. Expression of the Hox genes in Drosophila embryos is essential for body patterning and is controlled by an extensive array of cis-regulatory modules (CRMs. How the regulatory modules functionally evolve in different species is not clear. A comparison of the CRMs for the Abdominal-B gene from different Drosophila species reveals relatively low levels of overall sequence conservation. However, embryonic enhancer CRMs from other Drosophila species direct transgenic reporter gene expression in the same spatial and temporal patterns during development as their D. melanogaster orthologs. Bioinformatic analysis reveals the presence of short conserved sequences within defined CRMs, representing gap and pair-rule transcription factor binding sites. One predicted binding site for the gap transcription factor KRUPPEL in the IAB5 CRM was found to be altered in Superabdominal (Sab mutations. In Sab mutant flies, the third abdominal segment is transformed into a copy of the fifth abdominal segment. A model for KRUPPEL-mediated repression at this binding site is presented. These findings challenge our current understanding of the relationship between sequence evolution at the molecular level and functional activity of a CRM. While the overall sequence conservation at Drosophila CRMs is not distinctive from neighboring genomic regions, functionally critical transcription factor binding sites within embryonic enhancer CRMs are highly conserved. These results have implications for understanding mechanisms of gene expression during embryonic development, enhancer function, and the molecular evolution of eukaryotic regulatory modules.

Mapping cis-Regulatory Domains in the Human Genome UsingMulti-Species Conservation of Synteny

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Ahituv, Nadav; Prabhakar, Shyam; Poulin, Francis; Rubin, EdwardM.; Couronne, Olivier

2005-06-13

Our inability to associate distant regulatory elements with the genes that they regulate has largely precluded their examination for sequence alterations contributing to human disease. One major obstacle is the large genomic space surrounding targeted genes in which such elements could potentially reside. In order to delineate gene regulatory boundaries we used whole-genome human-mouse-chicken (HMC) and human-mouse-frog (HMF) multiple alignments to compile conserved blocks of synteny (CBS), under the hypothesis that these blocks have been kept intact throughout evolution at least in part by the requirement of regulatory elements to stay linked to the genes that they regulate. A total of 2,116 and 1,942 CBS>200 kb were assembled for HMC and HMF respectively, encompassing 1.53 and 0.86 Gb of human sequence. To support the existence of complex long-range regulatory domains within these CBS we analyzed the prevalence and distribution of chromosomal aberrations leading to position effects (disruption of a genes regulatory environment), observing a clear bias not only for mapping onto CBS but also for longer CBS size. Our results provide a genome wide data set characterizing the regulatory domains of genes and the conserved regulatory elements within them.

cisMEP: an integrated repository of genomic epigenetic profiles and cis-regulatory modules in Drosophila.

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Yang, Tzu-Hsien; Wang, Chung-Ching; Hung, Po-Cheng; Wu, Wei-Sheng

2014-01-01

Cis-regulatory modules (CRMs), or the DNA sequences required for regulating gene expression, play the central role in biological researches on transcriptional regulation in metazoan species. Nowadays, the systematic understanding of CRMs still mainly resorts to computational methods due to the time-consuming and small-scale nature of experimental methods. But the accuracy and reliability of different CRM prediction tools are still unclear. Without comparative cross-analysis of the results and combinatorial consideration with extra experimental information, there is no easy way to assess the confidence of the predicted CRMs. This limits the genome-wide understanding of CRMs. It is known that transcription factor binding and epigenetic profiles tend to determine functions of CRMs in gene transcriptional regulation. Thus integration of the genome-wide epigenetic profiles with systematically predicted CRMs can greatly help researchers evaluate and decipher the prediction confidence and possible transcriptional regulatory functions of these potential CRMs. However, these data are still fragmentary in the literatures. Here we performed the computational genome-wide screening for potential CRMs using different prediction tools and constructed the pioneer database, cisMEP (cis-regulatory module epigenetic profile database), to integrate these computationally identified CRMs with genomic epigenetic profile data. cisMEP collects the literature-curated TFBS location data and nine genres of epigenetic data for assessing the confidence of these potential CRMs and deciphering the possible CRM functionality. cisMEP aims to provide a user-friendly interface for researchers to assess the confidence of different potential CRMs and to understand the functions of CRMs through experimentally-identified epigenetic profiles. The deposited potential CRMs and experimental epigenetic profiles for confidence assessment provide experimentally testable hypotheses for the molecular mechanisms

Changes in cis-regulatory elements of a key floral regulator are associated with divergence of inflorescence architectures.

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Kusters, Elske; Della Pina, Serena; Castel, Rob; Souer, Erik; Koes, Ronald

2015-08-15

Higher plant species diverged extensively with regard to the moment (flowering time) and position (inflorescence architecture) at which flowers are formed. This seems largely caused by variation in the expression patterns of conserved genes that specify floral meristem identity (FMI), rather than changes in the encoded proteins. Here, we report a functional comparison of the promoters of homologous FMI genes from Arabidopsis, petunia, tomato and Antirrhinum. Analysis of promoter-reporter constructs in petunia and Arabidopsis, as well as complementation experiments, showed that the divergent expression of leafy (LFY) and the petunia homolog aberrant leaf and flower (ALF) results from alterations in the upstream regulatory network rather than cis-regulatory changes. The divergent expression of unusual floral organs (UFO) from Arabidopsis, and the petunia homolog double top (DOT), however, is caused by the loss or gain of cis-regulatory promoter elements, which respond to trans-acting factors that are expressed in similar patterns in both species. Introduction of pUFO:UFO causes no obvious defects in Arabidopsis, but in petunia it causes the precocious and ectopic formation of flowers. This provides an example of how a change in a cis-regulatory region can account for a change in the plant body plan. © 2015. Published by The Company of Biologists Ltd.

Cis-regulatory element based targeted gene finding: genome-wide identification of abscisic acid- and abiotic stress-responsive genes in Arabidopsis thaliana.

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Zhang, Weixiong; Ruan, Jianhua; Ho, Tuan-Hua David; You, Youngsook; Yu, Taotao; Quatrano, Ralph S

2005-07-15

A fundamental problem of computational genomics is identifying the genes that respond to certain endogenous cues and environmental stimuli. This problem can be referred to as targeted gene finding. Since gene regulation is mainly determined by the binding of transcription factors and cis-regulatory DNA sequences, most existing gene annotation methods, which exploit the conservation of open reading frames, are not effective in finding target genes. A viable approach to targeted gene finding is to exploit the cis-regulatory elements that are known to be responsible for the transcription of target genes. Given such cis-elements, putative target genes whose promoters contain the elements can be identified. As a case study, we apply the above approach to predict the genes in model plant Arabidopsis thaliana which are inducible by a phytohormone, abscisic acid (ABA), and abiotic stress, such as drought, cold and salinity. We first construct and analyze two ABA specific cis-elements, ABA-responsive element (ABRE) and its coupling element (CE), in A.thaliana, based on their conservation in rice and other cereal plants. We then use the ABRE-CE module to identify putative ABA-responsive genes in A.thaliana. Based on RT-PCR verification and the results from literature, this method has an accuracy rate of 67.5% for the top 40 predictions. The cis-element based targeted gene finding approach is expected to be widely applicable since a large number of cis-elements in many species are available.

Pathogenic adaptation of intracellular bacteria by rewiring a cis-regulatory input function.

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Osborne, Suzanne E; Walthers, Don; Tomljenovic, Ana M; Mulder, David T; Silphaduang, Uma; Duong, Nancy; Lowden, Michael J; Wickham, Mark E; Waller, Ross F; Kenney, Linda J; Coombes, Brian K

2009-03-10

The acquisition of DNA by horizontal gene transfer enables bacteria to adapt to previously unexploited ecological niches. Although horizontal gene transfer and mutation of protein-coding sequences are well-recognized forms of pathogen evolution, the evolutionary significance of cis-regulatory mutations in creating phenotypic diversity through altered transcriptional outputs is not known. We show the significance of regulatory mutation for pathogen evolution by mapping and then rewiring a cis-regulatory module controlling a gene required for murine typhoid. Acquisition of a binding site for the Salmonella pathogenicity island-2 regulator, SsrB, enabled the srfN gene, ancestral to the Salmonella genus, to play a role in pathoadaptation of S. typhimurium to a host animal. We identified the evolved cis-regulatory module and quantified the fitness gain that this regulatory output accrues for the bacterium using competitive infections of host animals. Our findings highlight a mechanism of pathogen evolution involving regulatory mutation that is selected because of the fitness advantage the new regulatory output provides the incipient clones.

Identification of a cis-regulatory element by transient analysis of co-ordinately regulated genes

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Allan Andrew C

2008-07-01

Full Text Available Abstract Background Transcription factors (TFs co-ordinately regulate target genes that are dispersed throughout the genome. This co-ordinate regulation is achieved, in part, through the interaction of transcription factors with conserved cis-regulatory motifs that are in close proximity to the target genes. While much is known about the families of transcription factors that regulate gene expression in plants, there are few well characterised cis-regulatory motifs. In Arabidopsis, over-expression of the MYB transcription factor PAP1 (PRODUCTION OF ANTHOCYANIN PIGMENT 1 leads to transgenic plants with elevated anthocyanin levels due to the co-ordinated up-regulation of genes in the anthocyanin biosynthetic pathway. In addition to the anthocyanin biosynthetic genes, there are a number of un-associated genes that also change in expression level. This may be a direct or indirect consequence of the over-expression of PAP1. Results Oligo array analysis of PAP1 over-expression Arabidopsis plants identified genes co-ordinately up-regulated in response to the elevated expression of this transcription factor. Transient assays on the promoter regions of 33 of these up-regulated genes identified eight promoter fragments that were transactivated by PAP1. Bioinformatic analysis on these promoters revealed a common cis-regulatory motif that we showed is required for PAP1 dependent transactivation. Conclusion Co-ordinated gene regulation by individual transcription factors is a complex collection of both direct and indirect effects. Transient transactivation assays provide a rapid method to identify direct target genes from indirect target genes. Bioinformatic analysis of the promoters of these direct target genes is able to locate motifs that are common to this sub-set of promoters, which is impossible to identify with the larger set of direct and indirect target genes. While this type of analysis does not prove a direct interaction between protein and DNA

An in vivo cis-regulatory screen at the type 2 diabetes associated TCF7L2 locus identifies multiple tissue-specific enhancers.

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Daniel Savic

Full Text Available Genome-wide association studies (GWAS have repeatedly shown an association between non-coding variants in the TCF7L2 locus and risk for type 2 diabetes (T2D, implicating a role for cis-regulatory variation within this locus in disease etiology. Supporting this hypothesis, we previously localized complex regulatory activity to the TCF7L2 T2D-associated interval using an in vivo bacterial artificial chromosome (BAC enhancer-trapping reporter strategy. To follow-up on this broad initial survey of the TCF7L2 regulatory landscape, we performed a fine-mapping enhancer scan using in vivo mouse transgenic reporter assays. We functionally interrogated approximately 50% of the sequences within the T2D-associated interval, utilizing sequence conservation within this 92-kb interval to determine the regulatory potential of all evolutionary conserved sequences that exhibited conservation to the non-eutherian mammal opossum. Included in this study was a detailed functional interrogation of sequences spanning both protective and risk alleles of single nucleotide polymorphism (SNP rs7903146, which has exhibited allele-specific enhancer function in pancreatic beta cells. Using these assays, we identified nine segments regulating various aspects of the TCF7L2 expression profile and that constitute nearly 70% of the sequences tested. These results highlight the regulatory complexity of this interval and support the notion that a TCF7L2 cis-regulatory disruption leads to T2D predisposition.

Comparative genome sequencing of Drosophila pseudoobscura: Chromosomal, gene, and cis-element evolution

DEFF Research Database (Denmark)

Richards, Stephen; Liu, Yue; Bettencourt, Brian R.

2005-01-01

years (Myr) since the pseudoobscura/melanogaster divergence. Genes expressed in the testes had higher amino acid sequence divergence than the genome-wide average, consistent with the rapid evolution of sex-specific proteins. Cis-regulatory sequences are more conserved than random and nearby sequences......We have sequenced the genome of a second Drosophila species, Drosophila pseudoobscura, and compared this to the genome sequence of Drosophila melanogaster, a primary model organism. Throughout evolution the vast majority of Drosophila genes have remained on the same chromosome arm, but within each...... between the species-but the difference is slight, suggesting that the evolution of cis-regulatory elements is flexible. Overall, a pattern of repeat-mediated chromosomal rearrangement, and high coadaptation of both male genes and cis-regulatory sequences emerges as important themes of genome divergence...

Identification of putative cis-regulatory elements in Cryptosporidium parvum by de novo pattern finding

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Kissinger Jessica C

2007-01-01

Full Text Available Abstract Background Cryptosporidium parvum is a unicellular eukaryote in the phylum Apicomplexa. It is an obligate intracellular parasite that causes diarrhea and is a significant AIDS-related pathogen. Cryptosporidium parvum is not amenable to long-term laboratory cultivation or classical molecular genetic analysis. The parasite exhibits a complex life cycle, a broad host range, and fundamental mechanisms of gene regulation remain unknown. We have used data from the recently sequenced genome of this organism to uncover clues about gene regulation in C. parvum. We have applied two pattern finding algorithms MEME and AlignACE to identify conserved, over-represented motifs in the 5' upstream regions of genes in C. parvum. To support our findings, we have established comparative real-time -PCR expression profiles for the groups of genes examined computationally. Results We find that groups of genes that share a function or belong to a common pathway share upstream motifs. Different motifs are conserved upstream of different groups of genes. Comparative real-time PCR studies show co-expression of genes within each group (in sub-sets during the life cycle of the parasite, suggesting co-regulation of these genes may be driven by the use of conserved upstream motifs. Conclusion This is one of the first attempts to characterize cis-regulatory elements in the absence of any previously characterized elements and with very limited expression data (seven genes only. Using de novo pattern finding algorithms, we have identified specific DNA motifs that are conserved upstream of genes belonging to the same metabolic pathway or gene family. We have demonstrated the co-expression of these genes (often in subsets using comparative real-time-PCR experiments thus establishing evidence for these conserved motifs as putative cis-regulatory elements. Given the lack of prior information concerning expression patterns and organization of promoters in C. parvum we

In silico analysis of cis-acting regulatory elements in 5' regulatory regions of sucrose transporter gene families in rice (Oryza sativa Japonica) and Arabidopsis thaliana.

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Ibraheem, Omodele; Botha, Christiaan E J; Bradley, Graeme

2010-12-01

The regulation of gene expression involves a multifarious regulatory system. Each gene contains a unique combination of cis-acting regulatory sequence elements in the 5' regulatory region that determines its temporal and spatial expression. Cis-acting regulatory elements are essential transcriptional gene regulatory units; they control many biological processes and stress responses. Thus a full understanding of the transcriptional gene regulation system will depend on successful functional analyses of cis-acting elements. Cis-acting regulatory elements present within the 5' regulatory region of the sucrose transporter gene families in rice (Oryza sativa Japonica cultivar-group) and Arabidopsis thaliana, were identified using a bioinformatics approach. The possible cis-acting regulatory elements were predicted by scanning 1.5kbp of 5' regulatory regions of the sucrose transporter genes translational start sites, using Plant CARE, PLACE and Genomatix Matinspector professional databases. Several cis-acting regulatory elements that are associated with plant development, plant hormonal regulation and stress response were identified, and were present in varying frequencies within the 1.5kbp of 5' regulatory region, among which are; A-box, RY, CAT, Pyrimidine-box, Sucrose-box, ABRE, ARF, ERE, GARE, Me-JA, ARE, DRE, GA-motif, GATA, GT-1, MYC, MYB, W-box, and I-box. This result reveals the probable cis-acting regulatory elements that possibly are involved in the expression and regulation of sucrose transporter gene families in rice and Arabidopsis thaliana during cellular development or environmental stress conditions. Copyright © 2010 Elsevier Ltd. All rights reserved.

Validation of Skeletal Muscle cis-Regulatory Module Predictions Reveals Nucleotide Composition Bias in Functional Enhancers

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Kwon, Andrew T.; Chou, Alice Yi; Arenillas, David J.; Wasserman, Wyeth W.

2011-01-01

We performed a genome-wide scan for muscle-specific cis-regulatory modules (CRMs) using three computational prediction programs. Based on the predictions, 339 candidate CRMs were tested in cell culture with NIH3T3 fibroblasts and C2C12 myoblasts for capacity to direct selective reporter gene expression to differentiated C2C12 myotubes. A subset of 19 CRMs validated as functional in the assay. The rate of predictive success reveals striking limitations of computational regulatory sequence analysis methods for CRM discovery. Motif-based methods performed no better than predictions based only on sequence conservation. Analysis of the properties of the functional sequences relative to inactive sequences identifies nucleotide sequence composition can be an important characteristic to incorporate in future methods for improved predictive specificity. Muscle-related TFBSs predicted within the functional sequences display greater sequence conservation than non-TFBS flanking regions. Comparison with recent MyoD and histone modification ChIP-Seq data supports the validity of the functional regions. PMID:22144875

Validation of skeletal muscle cis-regulatory module predictions reveals nucleotide composition bias in functional enhancers.

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Andrew T Kwon

2011-12-01

Full Text Available We performed a genome-wide scan for muscle-specific cis-regulatory modules (CRMs using three computational prediction programs. Based on the predictions, 339 candidate CRMs were tested in cell culture with NIH3T3 fibroblasts and C2C12 myoblasts for capacity to direct selective reporter gene expression to differentiated C2C12 myotubes. A subset of 19 CRMs validated as functional in the assay. The rate of predictive success reveals striking limitations of computational regulatory sequence analysis methods for CRM discovery. Motif-based methods performed no better than predictions based only on sequence conservation. Analysis of the properties of the functional sequences relative to inactive sequences identifies nucleotide sequence composition can be an important characteristic to incorporate in future methods for improved predictive specificity. Muscle-related TFBSs predicted within the functional sequences display greater sequence conservation than non-TFBS flanking regions. Comparison with recent MyoD and histone modification ChIP-Seq data supports the validity of the functional regions.

Creating and validating cis-regulatory maps of tissue-specific gene expression regulation

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O'Connor, Timothy R.; Bailey, Timothy L.

2014-01-01

Predicting which genomic regions control the transcription of a given gene is a challenge. We present a novel computational approach for creating and validating maps that associate genomic regions (cis-regulatory modules–CRMs) with genes. The method infers regulatory relationships that explain gene expression observed in a test tissue using widely available genomic data for ‘other’ tissues. To predict the regulatory targets of a CRM, we use cross-tissue correlation between histone modifications present at the CRM and expression at genes within 1 Mbp of it. To validate cis-regulatory maps, we show that they yield more accurate models of gene expression than carefully constructed control maps. These gene expression models predict observed gene expression from transcription factor binding in the CRMs linked to that gene. We show that our maps are able to identify long-range regulatory interactions and improve substantially over maps linking genes and CRMs based on either the control maps or a ‘nearest neighbor’ heuristic. Our results also show that it is essential to include CRMs predicted in multiple tissues during map-building, that H3K27ac is the most informative histone modification, and that CAGE is the most informative measure of gene expression for creating cis-regulatory maps. PMID:25200088

Coevolution within a transcriptional network by compensatory trans and cis mutations

KAUST Repository

Kuo, D.

2010-10-26

Transcriptional networks have been shown to evolve very rapidly, prompting questions as to how such changes arise and are tolerated. Recent comparisons of transcriptional networks across species have implicated variations in the cis-acting DNA sequences near genes as the main cause of divergence. What is less clear is how these changes interact with trans-acting changes occurring elsewhere in the genetic circuit. Here, we report the discovery of a system of compensatory trans and cis mutations in the yeast AP-1 transcriptional network that allows for conserved transcriptional regulation despite continued genetic change. We pinpoint a single species, the fungal pathogen Candida glabrata, in which a trans mutation has occurred very recently in a single AP-1 family member, distinguishing it from its Saccharomyces ortholog. Comparison of chromatin immunoprecipitation profiles between Candida and Saccharomyces shows that, despite their different DNA-binding domains, the AP-1 orthologs regulate a conserved block of genes. This conservation is enabled by concomitant changes in the cis-regulatory motifs upstream of each gene. Thus, both trans and cis mutations have perturbed the yeast AP-1 regulatory system in such a way as to compensate for one another. This demonstrates an example of “coevolution” between a DNA-binding transcription factor and its cis-regulatory site, reminiscent of the coevolution of protein binding partners.

Genetic mapping uncovers cis-regulatory landscape of RNA editing.

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Ramaswami, Gokul; Deng, Patricia; Zhang, Rui; Anna Carbone, Mary; Mackay, Trudy F C; Li, Jin Billy

2015-09-16

Adenosine-to-inosine (A-to-I) RNA editing, catalysed by ADAR enzymes conserved in metazoans, plays an important role in neurological functions. Although the fine-tuning mechanism provided by A-to-I RNA editing is important, the underlying rules governing ADAR substrate recognition are not well understood. We apply a quantitative trait loci (QTL) mapping approach to identify genetic variants associated with variability in RNA editing. With very accurate measurement of RNA editing levels at 789 sites in 131 Drosophila melanogaster strains, here we identify 545 editing QTLs (edQTLs) associated with differences in RNA editing. We demonstrate that many edQTLs can act through changes in the local secondary structure for edited dsRNAs. Furthermore, we find that edQTLs located outside of the edited dsRNA duplex are enriched in secondary structure, suggesting that distal dsRNA structure beyond the editing site duplex affects RNA editing efficiency. Our work will facilitate the understanding of the cis-regulatory code of RNA editing.

Organization of cis-acting regulatory elements in osmotic- and cold-stress-responsive promoters.

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Yamaguchi-Shinozaki, Kazuko; Shinozaki, Kazuo

2005-02-01

cis-Acting regulatory elements are important molecular switches involved in the transcriptional regulation of a dynamic network of gene activities controlling various biological processes, including abiotic stress responses, hormone responses and developmental processes. In particular, understanding regulatory gene networks in stress response cascades depends on successful functional analyses of cis-acting elements. The ever-improving accuracy of transcriptome expression profiling has led to the identification of various combinations of cis-acting elements in the promoter regions of stress-inducible genes involved in stress and hormone responses. Here we discuss major cis-acting elements, such as the ABA-responsive element (ABRE) and the dehydration-responsive element/C-repeat (DRE/CRT), that are a vital part of ABA-dependent and ABA-independent gene expression in osmotic and cold stress responses.

A saturation screen for cis-acting regulatory DNA in the Hox genes of Ciona intestinalis

Energy Technology Data Exchange (ETDEWEB)

Keys, David N.; Lee, Byung-in; Di Gregorio, Anna; Harafuji, Naoe; Detter, Chris; Wang, Mei; Kahsai, Orsalem; Ahn, Sylvia; Arellano, Andre; Zhang, Quin; Trong, Stephan; Doyle, Sharon A.; Satoh, Noriyuki; Satou, Yutaka; Saiga, Hidetoshi; Christian, Allen; Rokhsar, Dan; Hawkins, Trevor L.; Levine, Mike; Richardson, Paul

2005-01-05

A screen for the systematic identification of cis-regulatory elements within large (>100 kb) genomic domains containing Hox genes was performed by using the basal chordate Ciona intestinalis. Randomly generated DNA fragments from bacterial artificial chromosomes containing two clusters of Hox genes were inserted into a vector upstream of a minimal promoter and lacZ reporter gene. A total of 222 resultant fusion genes were separately electroporated into fertilized eggs, and their regulatory activities were monitored in larvae. In sum, 21 separable cis-regulatory elements were found. These include eight Hox linked domains that drive expression in nested anterior-posterior domains of ectodermally derived tissues. In addition to vertebrate-like CNS regulation, the discovery of cis-regulatory domains that drive epidermal transcription suggests that C. intestinalis has arthropod-like Hox patterning in the epidermis.

Comparative genome sequencing of drosophila pseudoobscura: Chromosomal, gene and cis-element evolution

Energy Technology Data Exchange (ETDEWEB)

Richards, Stephen; Liu, Yue; Bettencourt, Brian R.; Hradecky, Pavel; Letovsky, Stan; Nielsen, Rasmus; Thornton, Kevin; Todd, Melissa J.; Chen, Rui; Meisel, Richard P.; Couronne, Olivier; Hua, Sujun; Smith, Mark A.; Bussemaker, Harmen J.; van Batenburg, Marinus F.; Howells, Sally L.; Scherer, Steven E.; Sodergren, Erica; Matthews, Beverly B.; Crosby, Madeline A.; Schroeder, Andrew J.; Ortiz-Barrientos, Daniel; Rives, Catherine M.; Metzker, Michael L.; Muzny, Donna M.; Scott, Graham; Steffen, David; Wheeler, David A.; Worley, Kim C.; Havlak, Paul; Durbin, K. James; Egan, Amy; Gill, Rachel; Hume, Jennifer; Morgan, Margaret B.; Miner, George; Hamilton, Cerissa; Huang, Yanmei; Waldron, Lenee; Verduzco, Daniel; Blankenburg, Kerstin P.; Dubchak, Inna; Noor, Mohamed A.F.; Anderson, Wyatt; White, Kevin P.; Clark, Andrew G.; Schaeffer, Stephen W.; Gelbart, William; Weinstock, George M.; Gibbs, Richard A.

2004-04-01

The genome sequence of a second fruit fly, D. pseudoobscura, presents an opportunity for comparative analysis of a primary model organism D. melanogaster. The vast majority of Drosophila genes have remained on the same arm, but within each arm gene order has been extensively reshuffled leading to the identification of approximately 1300 syntenic blocks. A repetitive sequence is found in the D. pseudoobscura genome at many junctions between adjacent syntenic blocks. Analysis of this novel repetitive element family suggests that recombination between offset elements may have given rise to many paracentric inversions, thereby contributing to the shuffling of gene order in the D. pseudoobscura lineage. Based on sequence similarity and synteny, 10,516 putative orthologs have been identified as a core gene set conserved over 35 My since divergence. Genes expressed in the testes had higher amino acid sequence divergence than the genome wide average consistent with the rapid evolution of sex-specific proteins. Cis-regulatory sequences are more conserved than control sequences between the species but the difference is slight, suggesting that the evolution of cis-regulatory elements is flexible. Overall, a picture of repeat mediated chromosomal rearrangement, and high co-adaptation of both male genes and cis-regulatory sequences emerges as important themes of genome divergence between these species of Drosophila.

Bounded search for de novo identification of degenerate cis-regulatory elements

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Khetani Radhika S

2006-05-01

Full Text Available Abstract Background The identification of statistically overrepresented sequences in the upstream regions of coregulated genes should theoretically permit the identification of potential cis-regulatory elements. However, in practice many cis-regulatory elements are highly degenerate, precluding the use of an exhaustive word-counting strategy for their identification. While numerous methods exist for inferring base distributions using a position weight matrix, recent studies suggest that the independence assumptions inherent in the model, as well as the inability to reach a global optimum, limit this approach. Results In this paper, we report PRISM, a degenerate motif finder that leverages the relationship between the statistical significance of a set of binding sites and that of the individual binding sites. PRISM first identifies overrepresented, non-degenerate consensus motifs, then iteratively relaxes each one into a high-scoring degenerate motif. This approach requires no tunable parameters, thereby lending itself to unbiased performance comparisons. We therefore compare PRISM's performance against nine popular motif finders on 28 well-characterized S. cerevisiae regulons. PRISM consistently outperforms all other programs. Finally, we use PRISM to predict the binding sites of uncharacterized regulons. Our results support a proposed mechanism of action for the yeast cell-cycle transcription factor Stb1, whose binding site has not been determined experimentally. Conclusion The relationship between statistical measures of the binding sites and the set as a whole leads to a simple means of identifying the diverse range of cis-regulatory elements to which a protein binds. This approach leverages the advantages of word-counting, in that position dependencies are implicitly accounted for and local optima are more easily avoided. While we sacrifice guaranteed optimality to prevent the exponential blowup of exhaustive search, we prove that the error

Identification of two evolutionarily conserved 5' cis-elements involved in regulating spatiotemporal expression of Nolz-1 during mouse embryogenesis.

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Sunny Li-Yun Chang

Full Text Available Proper development of vertebrate embryos depends not only on the crucial funtions of key evolutionarily conserved transcriptional regulators, but also on the precisely spatiotemporal expression of these transcriptional regulators. The mouse Nolz-1/Znf503/Zfp503 gene is a mammalian member of the conserved zinc-finger containing NET family. The expression pattern of Nolz-1 in mouse embryos is highly correlated with that of its homologues in different species. To study the spatiotemporal regulation of Nolz-1, we first identified two evolutionarily conserved cis-elements, UREA and UREB, in 5' upstream regions of mouse Nolz-1 locus. We then generated UREA-LacZ and UREB-LacZ transgenic reporter mice to characterize the putative enhancer activity of UREA and UREB. The results indicated that both UREA and UREB contained tissue-specific enhancer activity for directing LacZ expression in selective tissue organs during mouse embryogensis. UREA directed LacZ expression preferentially in selective regions of developing central nervous system, including the forebrain, hindbrain and spinal cord, whereas UREB directed LacZ expression mainly in other developing tissue organs such as the Nolz-1 expressing branchial arches and its derivatives, the apical ectodermal ridge of limb buds and the urogenital tissues. Both UREA and UREB directed strong LacZ expression in the lateral plate mesoderm where endogenous Nolz-1 was also expressed. Despite that the LacZ expression pattern did not full recapitulated the endogenous Nolz-1 expression and some mismatched expression patterns were observed, co-expression of LacZ and Nolz-1 did occur in many cells of selective tissue organs, such as in the ventrolateral cortex and ventral spinal cord of UREA-LacZ embryos, and the urogenital tubes of UREB-LacZ embryos. Taken together, our study suggests that UREA and UREB may function as evolutionarily conserved cis-regulatory elements that coordinate with other cis-elements to regulate

[Analysis of cis-regulatory element distribution in gene promoters of Gossypium raimondii and Arabidopsis thaliana].

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Sun, Gao-Fei; He, Shou-Pu; Du, Xiong-Ming

2013-10-01

Cotton genomic studies have boomed since the release of Gossypium raimondii draft genome. In this study, cis-regulatory element (CRE) in 1 kb length sequence upstream 5' UTR of annotated genes were selected and scanned in the Arabidopsis thaliana (At) and Gossypium raimondii (Gr) genomes, based on the database of PLACE (Plant cis-acting Regulatory DNA Elements). According to the definition of this study, 44 (12.3%) and 57 (15.5%) CREs presented "peak-like" distribution in the 1 kb selected sequences of both genomes, respectively. Thirty-four of them were peak-like distributed in both genomes, which could be further categorized into 4 types based on their core sequences. The coincidence of TATABOX peak position and their actual position ((-) -30 bp) indicated that the position of a common CRE was conservative in different genes, which suggested that the peak position of these CREs was their possible actual position of transcription factors. The position of a common CRE was also different between the two genomes due to stronger length variation of 5' UTR in Gr than At. Furthermore, most of the peak-like CREs were located in the region of -110 bp-0 bp, which suggested that concentrated distribution might be conductive to the interaction of transcription factors, and then regulate the gene expression in downstream.

Characterization of Cer-1 cis-regulatory region during early Xenopus development.

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Silva, Ana Cristina; Filipe, Mário; Steinbeisser, Herbert; Belo, José António

2011-05-01

Cerberus-related molecules are well-known Wnt, Nodal, and BMP inhibitors that have been implicated in different processes including anterior–posterior patterning and left–right asymmetry. In both mouse and frog, two Cerberus-related genes have been isolated, mCer-1 and mCer-2, and Xcer and Xcoco, respectively. Until now, little is known about the mechanisms involved in their transcriptional regulation. Here, we report a heterologous analysis of the mouse Cerberus-1 gene upstream regulatory regions, responsible for its expression in the visceral endodermal cells. Our analysis showed that the consensus sequences for a TATA, CAAT, or GC boxes were absent but a TGTGG sequence was present at position -172 to -168 bp, relative to the ATG. Using a series of deletion constructs and transient expression in Xenopus embryos, we found that a fragment of 1.4 kb of Cer-1 promoter sequence could reproduce the endogenous expression pattern of Xenopus cerberus. A 0.7-kb mcer-1 upstream region was able to drive reporter expression to the involuting mesendodermal cells, while further deletions abolished reporter gene expression. Our results suggest that although no sequence similarity was found between mouse and Xenopus cerberus cis-regulatory regions, the signaling cascades regulating cerberus expression, during gastrulation, is conserved.

A method for selecting cis-acting regulatory sequences that respond to small molecule effectors

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Allas Ülar

2010-08-01

Full Text Available Abstract Background Several cis-acting regulatory sequences functioning at the level of mRNA or nascent peptide and specifically influencing transcription or translation have been described. These regulatory elements often respond to specific chemicals. Results We have developed a method that allows us to select cis-acting regulatory sequences that respond to diverse chemicals. The method is based on the β-lactamase gene containing a random sequence inserted into the beginning of the ORF. Several rounds of selection are used to isolate sequences that suppress β-lactamase expression in response to the compound under study. We have isolated sequences that respond to erythromycin, troleandomycin, chloramphenicol, meta-toluate and homoserine lactone. By introducing synonymous and non-synonymous mutations we have shown that at least in the case of erythromycin the sequences act at the peptide level. We have also tested the cross-activities of the constructs and found that in most cases the sequences respond most strongly to the compound on which they were isolated. Conclusions Several selected peptides showed ligand-specific changes in amino acid frequencies, but no consensus motif could be identified. This is consistent with previous observations on natural cis-acting peptides, showing that it is often impossible to demonstrate a consensus. Applying the currently developed method on a larger scale, by selecting and comparing an extended set of sequences, might allow the sequence rules underlying the activity of cis-acting regulatory peptides to be identified.

Using reporter gene assays to identify cis regulatory differences between humans and chimpanzees.

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Chabot, Adrien; Shrit, Ralla A; Blekhman, Ran; Gilad, Yoav

2007-08-01

Most phenotypic differences between human and chimpanzee are likely to result from differences in gene regulation, rather than changes to protein-coding regions. To date, however, only a handful of human-chimpanzee nucleotide differences leading to changes in gene regulation have been identified. To hone in on differences in regulatory elements between human and chimpanzee, we focused on 10 genes that were previously found to be differentially expressed between the two species. We then designed reporter gene assays for the putative human and chimpanzee promoters of the 10 genes. Of seven promoters that we found to be active in human liver cell lines, human and chimpanzee promoters had significantly different activity in four cases, three of which recapitulated the gene expression difference seen in the microarray experiment. For these three genes, we were therefore able to demonstrate that a change in cis influences expression differences between humans and chimpanzees. Moreover, using site-directed mutagenesis on one construct, the promoter for the DDA3 gene, we were able to identify three nucleotides that together lead to a cis regulatory difference between the species. High-throughput application of this approach can provide a map of regulatory element differences between humans and our close evolutionary relatives.

Nomadic enhancers: tissue-specific cis-regulatory elements of yellow have divergent genomic positions among Drosophila species.

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Gizem Kalay

2010-11-01

Full Text Available cis-regulatory DNA sequences known as enhancers control gene expression in space and time. They are central to metazoan development and are often responsible for changes in gene regulation that contribute to phenotypic evolution. Here, we examine the sequence, function, and genomic location of enhancers controlling tissue- and cell-type specific expression of the yellow gene in six Drosophila species. yellow is required for the production of dark pigment, and its expression has evolved largely in concert with divergent pigment patterns. Using Drosophila melanogaster as a transgenic host, we examined the expression of reporter genes in which either 5' intergenic or intronic sequences of yellow from each species controlled the expression of Green Fluorescent Protein. Surprisingly, we found that sequences controlling expression in the wing veins, as well as sequences controlling expression in epidermal cells of the abdomen, thorax, and wing, were located in different genomic regions in different species. By contrast, sequences controlling expression in bristle-associated cells were located in the intron of all species. Differences in the precise pattern of spatial expression within the developing epidermis of D. melanogaster transformants usually correlated with adult pigmentation in the species from which the cis-regulatory sequences were derived, which is consistent with cis-regulatory evolution affecting yellow expression playing a central role in Drosophila pigmentation divergence. Sequence comparisons among species favored a model in which sequential nucleotide substitutions were responsible for the observed changes in cis-regulatory architecture. Taken together, these data demonstrate frequent changes in yellow cis-regulatory architecture among Drosophila species. Similar analyses of other genes, combining in vivo functional tests of enhancer activity with in silico comparative genomics, are needed to determine whether the pattern of

Combinatorial Cis-regulation in Saccharomyces Species

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Aaron T. Spivak

2016-03-01

Full Text Available Transcriptional control of gene expression requires interactions between the cis-regulatory elements (CREs controlling gene promoters. We developed a sensitive computational method to identify CRE combinations with conserved spacing that does not require genome alignments. When applied to seven sensu stricto and sensu lato Saccharomyces species, 80% of the predicted interactions displayed some evidence of combinatorial transcriptional behavior in several existing datasets including: (1 chromatin immunoprecipitation data for colocalization of transcription factors, (2 gene expression data for coexpression of predicted regulatory targets, and (3 gene ontology databases for common pathway membership of predicted regulatory targets. We tested several predicted CRE interactions with chromatin immunoprecipitation experiments in a wild-type strain and strains in which a predicted cofactor was deleted. Our experiments confirmed that transcription factor (TF occupancy at the promoters of the CRE combination target genes depends on the predicted cofactor while occupancy of other promoters is independent of the predicted cofactor. Our method has the additional advantage of identifying regulatory differences between species. By analyzing the S. cerevisiae and S. bayanus genomes, we identified differences in combinatorial cis-regulation between the species and showed that the predicted changes in gene regulation explain several of the species-specific differences seen in gene expression datasets. In some instances, the same CRE combinations appear to regulate genes involved in distinct biological processes in the two different species. The results of this research demonstrate that (1 combinatorial cis-regulation can be inferred by multi-genome analysis and (2 combinatorial cis-regulation can explain differences in gene expression between species.

Characterization of Putative cis-Regulatory Elements in Genes Preferentially Expressed in Arabidopsis Male Meiocytes

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Junhua Li

2014-01-01

Full Text Available Meiosis is essential for plant reproduction because it is the process during which homologous chromosome pairing, synapsis, and meiotic recombination occur. The meiotic transcriptome is difficult to investigate because of the size of meiocytes and the confines of anther lobes. The recent development of isolation techniques has enabled the characterization of transcriptional profiles in male meiocytes of Arabidopsis. Gene expression in male meiocytes shows unique features. The direct interaction of transcription factors (TFs with DNA regulatory sequences forms the basis for the specificity of transcriptional regulation. Here, we identified putative cis-regulatory elements (CREs associated with male meiocyte-expressed genes using in silico tools. The upstream regions (1 kb of the top 50 genes preferentially expressed in Arabidopsis meiocytes possessed conserved motifs. These motifs are putative binding sites of TFs, some of which share common functions, such as roles in cell division. In combination with cell-type-specific analysis, our findings could be a substantial aid for the identification and experimental verification of the protein-DNA interactions for the specific TFs that drive gene expression in meiocytes.

Dynamic SPR monitoring of yeast nuclear protein binding to a cis-regulatory element

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Mao, Grace; Brody, James P.

2007-01-01

Gene expression is controlled by protein complexes binding to short specific sequences of DNA, called cis-regulatory elements. Expression of most eukaryotic genes is controlled by dozens of these elements. Comprehensive identification and monitoring of these elements is a major goal of genomics. In pursuit of this goal, we are developing a surface plasmon resonance (SPR) based assay to identify and monitor cis-regulatory elements. To test whether we could reliably monitor protein binding to a regulatory element, we immobilized a 16 bp region of Saccharomyces cerevisiae chromosome 5 onto a gold surface. This 16 bp region of DNA is known to bind several proteins and thought to control expression of the gene RNR1, which varies through the cell cycle. We synchronized yeast cell cultures, and then sampled these cultures at a regular interval. These samples were processed to purify nuclear lysate, which was then exposed to the sensor. We found that nuclear protein binds this particular element of DNA at a significantly higher rate (as compared to unsynchronized cells) during G1 phase. Other time points show levels of DNA-nuclear protein binding similar to the unsynchronized control. We also measured the apparent association complex of the binding to be 0.014 s -1 . We conclude that (1) SPR-based assays can monitor DNA-nuclear protein binding and that (2) for this particular cis-regulatory element, maximum DNA-nuclear protein binding occurs during G1 phase

Evolutionary conservation of regulatory elements in vertebrate HOX gene clusters

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Santini, Simona; Boore, Jeffrey L.; Meyer, Axel

2003-12-31

Due to their high degree of conservation, comparisons of DNA sequences among evolutionarily distantly-related genomes permit to identify functional regions in noncoding DNA. Hox genes are optimal candidate sequences for comparative genome analyses, because they are extremely conserved in vertebrates and occur in clusters. We aligned (Pipmaker) the nucleotide sequences of HoxA clusters of tilapia, pufferfish, striped bass, zebrafish, horn shark, human and mouse (over 500 million years of evolutionary distance). We identified several highly conserved intergenic sequences, likely to be important in gene regulation. Only a few of these putative regulatory elements have been previously described as being involved in the regulation of Hox genes, while several others are new elements that might have regulatory functions. The majority of these newly identified putative regulatory elements contain short fragments that are almost completely conserved and are identical to known binding sites for regulatory proteins (Transfac). The conserved intergenic regions located between the most rostrally expressed genes in the developing embryo are longer and better retained through evolution. We document that presumed regulatory sequences are retained differentially in either A or A clusters resulting from a genome duplication in the fish lineage. This observation supports both the hypothesis that the conserved elements are involved in gene regulation and the Duplication-Deletion-Complementation model.

CRX ChIP-seq reveals the cis-regulatory architecture of mouse photoreceptors

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J.C. Corbo (Joseph); K.A. Lawrence (Karen); M. Karlstetter (Marcus); C.A. Myers (Connie); M. Abdelaziz (Musa); W. Dirkes (William); K. Weigelt (Karin); M. Seifert (Martin); V. Benes (Vladimir); L.G. Fritsche (Lars); B.H.F. Weber (Bernhard); T. Langmann (Thomas)

2010-01-01

textabstractApproximately 98% of mammalian DNA is noncoding, yet we understand relatively little about the function of this enigmatic portion of the genome. The cis-regulatory elements that control gene expression reside in noncoding regions and can be identified by mapping the binding sites of

PlantCARE, a plant cis-acting regulatory element database

OpenAIRE

Rombauts, Stephane; Déhais, Patrice; Van Montagu, Marc; Rouzé, Pierre

1999-01-01

PlantCARE is a database of plant cis- acting regulatory elements, enhancers and repressors. Besides the transcription motifs found on a sequence, it also offers a link to the EMBL entry that contains the full gene sequence as well as a description of the conditions in which a motif becomes functional. The information on these sites is given by matrices, consensus and individual site sequences on particular genes, depending on the available information. PlantCARE is a relational database avail...

Retinal Expression of the Drosophila eyes absent Gene Is Controlled by Several Cooperatively Acting Cis-regulatory Elements

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Neuman, Sarah D.; Bashirullah, Arash; Kumar, Justin P.

2016-01-01

The eyes absent (eya) gene of the fruit fly, Drosophila melanogaster, is a member of an evolutionarily conserved gene regulatory network that controls eye formation in all seeing animals. The loss of eya leads to the complete elimination of the compound eye while forced expression of eya in non-retinal tissues is sufficient to induce ectopic eye formation. Within the developing retina eya is expressed in a dynamic pattern and is involved in tissue specification/determination, cell proliferation, apoptosis, and cell fate choice. In this report we explore the mechanisms by which eya expression is spatially and temporally governed in the developing eye. We demonstrate that multiple cis-regulatory elements function cooperatively to control eya transcription and that spacing between a pair of enhancer elements is important for maintaining correct gene expression. Lastly, we show that the loss of eya expression in sine oculis (so) mutants is the result of massive cell death and a progressive homeotic transformation of retinal progenitor cells into head epidermis. PMID:27930646

Cis-regulatory elements in the primate brain: from functional specialization to neurodegeneration

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Vermunt, Marit W.

2017-01-01

Over the last decade, the noncoding part of the genome has been shown to harbour thousands of cis-regulatory elements, such as enhancers, that activate well-defined gene expression programs. Here, we charted active enhancers in a multiplicity of human brain regions to understand the role of

Plasticity of the cis-regulatory input function of a gene.

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Avraham E Mayo

2006-04-01

Full Text Available The transcription rate of a gene is often controlled by several regulators that bind specific sites in the gene's cis-regulatory region. The combined effect of these regulators is described by a cis-regulatory input function. What determines the form of an input function, and how variable is it with respect to mutations? To address this, we employ the well-characterized lac operon of Escherichia coli, which has an elaborate input function, intermediate between Boolean AND-gate and OR-gate logic. We mapped in detail the input function of 12 variants of the lac promoter, each with different point mutations in the regulator binding sites, by means of accurate expression measurements from living cells. We find that even a few mutations can significantly change the input function, resulting in functions that resemble Pure AND gates, OR gates, or single-input switches. Other types of gates were not found. The variant input functions can be described in a unified manner by a mathematical model. The model also lets us predict which functions cannot be reached by point mutations. The input function that we studied thus appears to be plastic, in the sense that many of the mutations do not ruin the regulation completely but rather result in new ways to integrate the inputs.

Human developmental enhancers conserved between deuterostomes and protostomes.

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Shoa L Clarke

Full Text Available The identification of homologies, whether morphological, molecular, or genetic, is fundamental to our understanding of common biological principles. Homologies bridging the great divide between deuterostomes and protostomes have served as the basis for current models of animal evolution and development. It is now appreciated that these two clades share a common developmental toolkit consisting of conserved transcription factors and signaling pathways. These patterning genes sometimes show common expression patterns and genetic interactions, suggesting the existence of similar or even conserved regulatory apparatus. However, previous studies have found no regulatory sequence conserved between deuterostomes and protostomes. Here we describe the first such enhancers, which we call bilaterian conserved regulatory elements (Bicores. Bicores show conservation of sequence and gene synteny. Sequence conservation of Bicores reflects conserved patterns of transcription factor binding sites. We predict that Bicores act as response elements to signaling pathways, and we show that Bicores are developmental enhancers that drive expression of transcriptional repressors in the vertebrate central nervous system. Although the small number of identified Bicores suggests extensive rewiring of cis-regulation between the protostome and deuterostome clades, additional Bicores may be revealed as our understanding of cis-regulatory logic and sample of bilaterian genomes continue to grow.

Network-directed cis-mediator analysis of normal prostate tissue expression profiles reveals downstream regulatory associations of prostate cancer susceptibility loci.

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Larson, Nicholas B; McDonnell, Shannon K; Fogarty, Zach; Larson, Melissa C; Cheville, John; Riska, Shaun; Baheti, Saurabh; Weber, Alexandra M; Nair, Asha A; Wang, Liang; O'Brien, Daniel; Davila, Jaime; Schaid, Daniel J; Thibodeau, Stephen N

2017-10-17

Large-scale genome-wide association studies have identified multiple single-nucleotide polymorphisms associated with risk of prostate cancer. Many of these genetic variants are presumed to be regulatory in nature; however, follow-up expression quantitative trait loci (eQTL) association studies have to-date been restricted largely to cis -acting associations due to study limitations. While trans -eQTL scans suffer from high testing dimensionality, recent evidence indicates most trans -eQTL associations are mediated by cis -regulated genes, such as transcription factors. Leveraging a data-driven gene co-expression network, we conducted a comprehensive cis -mediator analysis using RNA-Seq data from 471 normal prostate tissue samples to identify downstream regulatory associations of previously identified prostate cancer risk variants. We discovered multiple trans -eQTL associations that were significantly mediated by cis -regulated transcripts, four of which involved risk locus 17q12, proximal transcription factor HNF1B , and target trans -genes with known HNF response elements ( MIA2 , SRC , SEMA6A , KIF12 ). We additionally identified evidence of cis -acting down-regulation of MSMB via rs10993994 corresponding to reduced co-expression of NDRG1 . The majority of these cis -mediator relationships demonstrated trans -eQTL replicability in 87 prostate tissue samples from the Gene-Tissue Expression Project. These findings provide further biological context to known risk loci and outline new hypotheses for investigation into the etiology of prostate cancer.

In silico modeling of epigenetic-induced changes in photoreceptor cis-regulatory elements.

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Hossain, Reafa A; Dunham, Nicholas R; Enke, Raymond A; Berndsen, Christopher E

2018-01-01

DNA methylation is a well-characterized epigenetic repressor of mRNA transcription in many plant and vertebrate systems. However, the mechanism of this repression is not fully understood. The process of transcription is controlled by proteins that regulate recruitment and activity of RNA polymerase by binding to specific cis-regulatory sequences. Cone-rod homeobox (CRX) is a well-characterized mammalian transcription factor that controls photoreceptor cell-specific gene expression. Although much is known about the functions and DNA binding specificity of CRX, little is known about how DNA methylation modulates CRX binding affinity to genomic cis-regulatory elements. We used bisulfite pyrosequencing of human ocular tissues to measure DNA methylation levels of the regulatory regions of RHO , PDE6B, PAX6 , and LINE1 retrotransposon repeats. To describe the molecular mechanism of repression, we used molecular modeling to illustrate the effect of DNA methylation on human RHO regulatory sequences. In this study, we demonstrate an inverse correlation between DNA methylation in regulatory regions adjacent to the human RHO and PDE6B genes and their subsequent transcription in human ocular tissues. Docking of CRX to the DNA models shows that CRX interacts with the grooves of these sequences, suggesting changes in groove structure could regulate binding. Molecular dynamics simulations of the RHO promoter and enhancer regions show changes in the flexibility and groove width upon epigenetic modification. Models also demonstrate changes in the local dynamics of CRX binding sites within RHO regulatory sequences which may account for the repression of CRX-dependent transcription. Collectively, these data demonstrate epigenetic regulation of CRX binding sites in human retinal tissue and provide insight into the mechanism of this mode of epigenetic regulation to be tested in future experiments.

A New Algorithm for Identifying Cis-Regulatory Modules Based on Hidden Markov Model

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Haitao Guo

2017-01-01

Full Text Available The discovery of cis-regulatory modules (CRMs is the key to understanding mechanisms of transcription regulation. Since CRMs have specific regulatory structures that are the basis for the regulation of gene expression, how to model the regulatory structure of CRMs has a considerable impact on the performance of CRM identification. The paper proposes a CRM discovery algorithm called ComSPS. ComSPS builds a regulatory structure model of CRMs based on HMM by exploring the rules of CRM transcriptional grammar that governs the internal motif site arrangement of CRMs. We test ComSPS on three benchmark datasets and compare it with five existing methods. Experimental results show that ComSPS performs better than them.

A New Algorithm for Identifying Cis-Regulatory Modules Based on Hidden Markov Model

Science.gov (United States)

2017-01-01

The discovery of cis-regulatory modules (CRMs) is the key to understanding mechanisms of transcription regulation. Since CRMs have specific regulatory structures that are the basis for the regulation of gene expression, how to model the regulatory structure of CRMs has a considerable impact on the performance of CRM identification. The paper proposes a CRM discovery algorithm called ComSPS. ComSPS builds a regulatory structure model of CRMs based on HMM by exploring the rules of CRM transcriptional grammar that governs the internal motif site arrangement of CRMs. We test ComSPS on three benchmark datasets and compare it with five existing methods. Experimental results show that ComSPS performs better than them. PMID:28497059

Using hexamers to predict cis-regulatory motifs in Drosophila

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Kibler Dennis

2005-10-01

Full Text Available Abstract Background Cis-regulatory modules (CRMs are short stretches of DNA that help regulate gene expression in higher eukaryotes. They have been found up to 1 megabase away from the genes they regulate and can be located upstream, downstream, and even within their target genes. Due to the difficulty of finding CRMs using biological and computational techniques, even well-studied regulatory systems may contain CRMs that have not yet been discovered. Results We present a simple, efficient method (HexDiff based only on hexamer frequencies of known CRMs and non-CRM sequence to predict novel CRMs in regulatory systems. On a data set of 16 gap and pair-rule genes containing 52 known CRMs, predictions made by HexDiff had a higher correlation with the known CRMs than several existing CRM prediction algorithms: Ahab, Cluster Buster, MSCAN, MCAST, and LWF. After combining the results of the different algorithms, 10 putative CRMs were identified and are strong candidates for future study. The hexamers used by HexDiff to distinguish between CRMs and non-CRM sequence were also analyzed and were shown to be enriched in regulatory elements. Conclusion HexDiff provides an efficient and effective means for finding new CRMs based on known CRMs, rather than known binding sites.

Prediction of tissue-specific cis-regulatory modules using Bayesian networks and regression trees

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Chen Xiaoyu

2007-12-01

Full Text Available Abstract Background In vertebrates, a large part of gene transcriptional regulation is operated by cis-regulatory modules. These modules are believed to be regulating much of the tissue-specificity of gene expression. Results We develop a Bayesian network approach for identifying cis-regulatory modules likely to regulate tissue-specific expression. The network integrates predicted transcription factor binding site information, transcription factor expression data, and target gene expression data. At its core is a regression tree modeling the effect of combinations of transcription factors bound to a module. A new unsupervised EM-like algorithm is developed to learn the parameters of the network, including the regression tree structure. Conclusion Our approach is shown to accurately identify known human liver and erythroid-specific modules. When applied to the prediction of tissue-specific modules in 10 different tissues, the network predicts a number of important transcription factor combinations whose concerted binding is associated to specific expression.

Accelerated Evolution of Conserved Noncoding Sequences in theHuman Genome

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Prambhakar, Shyam; Noonan, James P.; Paabo, Svante; Rubin, EdwardM.

2006-07-06

Genomic comparisons between human and distant, non-primatemammals are commonly used to identify cis-regulatory elements based onconstrained sequence evolution. However, these methods fail to detect"cryptic" functional elements, which are too weakly conserved amongmammals to distinguish from nonfunctional DNA. To address this problem,we explored the potential of deep intra-primate sequence comparisons. Wesequenced the orthologs of 558 kb of human genomic sequence, coveringmultiple loci involved in cholesterol homeostasis, in 6 nonhumanprimates. Our analysis identified 6 noncoding DNA elements displayingsignificant conservation among primates, but undetectable in more distantcomparisons. In vitro and in vivo tests revealed that at least three ofthese 6 elements have regulatory function. Notably, the mouse orthologsof these three functional human sequences had regulatory activity despitetheir lack of significant sequence conservation, indicating that they arecryptic ancestral cis-regulatory elements. These regulatory elementscould still be detected in a smaller set of three primate speciesincluding human, rhesus and marmoset. Since the human and rhesus genomesequences are already available, and the marmoset genome is activelybeing sequenced, the primate-specific conservation analysis describedhere can be applied in the near future on a whole-genome scale, tocomplement the annotation provided by more distant speciescomparisons.

Genes involved in complex adaptive processes tend to have highly conserved upstream regions in mammalian genomes

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Kohane Isaac

2005-11-01

Full Text Available Abstract Background Recent advances in genome sequencing suggest a remarkable conservation in gene content of mammalian organisms. The similarity in gene repertoire present in different organisms has increased interest in studying regulatory mechanisms of gene expression aimed at elucidating the differences in phenotypes. In particular, a proximal promoter region contains a large number of regulatory elements that control the expression of its downstream gene. Although many studies have focused on identification of these elements, a broader picture on the complexity of transcriptional regulation of different biological processes has not been addressed in mammals. The regulatory complexity may strongly correlate with gene function, as different evolutionary forces must act on the regulatory systems under different biological conditions. We investigate this hypothesis by comparing the conservation of promoters upstream of genes classified in different functional categories. Results By conducting a rank correlation analysis between functional annotation and upstream sequence alignment scores obtained by human-mouse and human-dog comparison, we found a significantly greater conservation of the upstream sequence of genes involved in development, cell communication, neural functions and signaling processes than those involved in more basic processes shared with unicellular organisms such as metabolism and ribosomal function. This observation persists after controlling for G+C content. Considering conservation as a functional signature, we hypothesize a higher density of cis-regulatory elements upstream of genes participating in complex and adaptive processes. Conclusion We identified a class of functions that are associated with either high or low promoter conservation in mammals. We detected a significant tendency that points to complex and adaptive processes were associated with higher promoter conservation, despite the fact that they have emerged

BET Bromodomain Inhibition Releases the Mediator Complex from Select cis-Regulatory Elements.

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Bhagwat, Anand S; Roe, Jae-Seok; Mok, Beverly Y L; Hohmann, Anja F; Shi, Junwei; Vakoc, Christopher R

2016-04-19

The bromodomain and extraterminal (BET) protein BRD4 can physically interact with the Mediator complex, but the relevance of this association to the therapeutic effects of BET inhibitors in cancer is unclear. Here, we show that BET inhibition causes a rapid release of Mediator from a subset of cis-regulatory elements in the genome of acute myeloid leukemia (AML) cells. These sites of Mediator eviction were highly correlated with transcriptional suppression of neighboring genes, which are enriched for targets of the transcription factor MYB and for functions related to leukemogenesis. A shRNA screen of Mediator in AML cells identified the MED12, MED13, MED23, and MED24 subunits as performing a similar regulatory function to BRD4 in this context, including a shared role in sustaining a block in myeloid maturation. These findings suggest that the interaction between BRD4 and Mediator has functional importance for gene-specific transcriptional activation and for AML maintenance. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

RNA-ID, a highly sensitive and robust method to identify cis-regulatory sequences using superfolder GFP and a fluorescence-based assay.

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Dean, Kimberly M; Grayhack, Elizabeth J

2012-12-01

We have developed a robust and sensitive method, called RNA-ID, to screen for cis-regulatory sequences in RNA using fluorescence-activated cell sorting (FACS) of yeast cells bearing a reporter in which expression of both superfolder green fluorescent protein (GFP) and yeast codon-optimized mCherry red fluorescent protein (RFP) is driven by the bidirectional GAL1,10 promoter. This method recapitulates previously reported progressive inhibition of translation mediated by increasing numbers of CGA codon pairs, and restoration of expression by introduction of a tRNA with an anticodon that base pairs exactly with the CGA codon. This method also reproduces effects of paromomycin and context on stop codon read-through. Five key features of this method contribute to its effectiveness as a selection for regulatory sequences: The system exhibits greater than a 250-fold dynamic range, a quantitative and dose-dependent response to known inhibitory sequences, exquisite resolution that allows nearly complete physical separation of distinct populations, and a reproducible signal between different cells transformed with the identical reporter, all of which are coupled with simple methods involving ligation-independent cloning, to create large libraries. Moreover, we provide evidence that there are sequences within a 9-nt library that cause reduced GFP fluorescence, suggesting that there are novel cis-regulatory sequences to be found even in this short sequence space. This method is widely applicable to the study of both RNA-mediated and codon-mediated effects on expression.

Seed storage protein gene promoters contain conserved DNA motifs in Brassicaceae, Fabaceae and Poaceae

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Fauteux, François; Strömvik, Martina V

2009-01-01

Background Accurate computational identification of cis-regulatory motifs is difficult, particularly in eukaryotic promoters, which typically contain multiple short and degenerate DNA sequences bound by several interacting factors. Enrichment in combinations of rare motifs in the promoter sequence of functionally or evolutionarily related genes among several species is an indicator of conserved transcriptional regulatory mechanisms. This provides a basis for the computational identification of cis-regulatory motifs. Results We have used a discriminative seeding DNA motif discovery algorithm for an in-depth analysis of 54 seed storage protein (SSP) gene promoters from three plant families, namely Brassicaceae (mustards), Fabaceae (legumes) and Poaceae (grasses) using backgrounds based on complete sets of promoters from a representative species in each family, namely Arabidopsis (Arabidopsis thaliana (L.) Heynh.), soybean (Glycine max (L.) Merr.) and rice (Oryza sativa L.) respectively. We have identified three conserved motifs (two RY-like and one ACGT-like) in Brassicaceae and Fabaceae SSP gene promoters that are similar to experimentally characterized seed-specific cis-regulatory elements. Fabaceae SSP gene promoter sequences are also enriched in a novel, seed-specific E2Fb-like motif. Conserved motifs identified in Poaceae SSP gene promoters include a GCN4-like motif, two prolamin-box-like motifs and an Skn-1-like motif. Evidence of the presence of a variant of the TATA-box is found in the SSP gene promoters from the three plant families. Motifs discovered in SSP gene promoters were used to score whole-genome sets of promoters from Arabidopsis, soybean and rice. The highest-scoring promoters are associated with genes coding for different subunits or precursors of seed storage proteins. Conclusion Seed storage protein gene promoter motifs are conserved in diverse species, and different plant families are characterized by a distinct combination of conserved motifs

Seed storage protein gene promoters contain conserved DNA motifs in Brassicaceae, Fabaceae and Poaceae

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Fauteux François

2009-10-01

Full Text Available Abstract Background Accurate computational identification of cis-regulatory motifs is difficult, particularly in eukaryotic promoters, which typically contain multiple short and degenerate DNA sequences bound by several interacting factors. Enrichment in combinations of rare motifs in the promoter sequence of functionally or evolutionarily related genes among several species is an indicator of conserved transcriptional regulatory mechanisms. This provides a basis for the computational identification of cis-regulatory motifs. Results We have used a discriminative seeding DNA motif discovery algorithm for an in-depth analysis of 54 seed storage protein (SSP gene promoters from three plant families, namely Brassicaceae (mustards, Fabaceae (legumes and Poaceae (grasses using backgrounds based on complete sets of promoters from a representative species in each family, namely Arabidopsis (Arabidopsis thaliana (L. Heynh., soybean (Glycine max (L. Merr. and rice (Oryza sativa L. respectively. We have identified three conserved motifs (two RY-like and one ACGT-like in Brassicaceae and Fabaceae SSP gene promoters that are similar to experimentally characterized seed-specific cis-regulatory elements. Fabaceae SSP gene promoter sequences are also enriched in a novel, seed-specific E2Fb-like motif. Conserved motifs identified in Poaceae SSP gene promoters include a GCN4-like motif, two prolamin-box-like motifs and an Skn-1-like motif. Evidence of the presence of a variant of the TATA-box is found in the SSP gene promoters from the three plant families. Motifs discovered in SSP gene promoters were used to score whole-genome sets of promoters from Arabidopsis, soybean and rice. The highest-scoring promoters are associated with genes coding for different subunits or precursors of seed storage proteins. Conclusion Seed storage protein gene promoter motifs are conserved in diverse species, and different plant families are characterized by a distinct combination

PReMod: a database of genome-wide mammalian cis-regulatory module predictions.

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Ferretti, Vincent; Poitras, Christian; Bergeron, Dominique; Coulombe, Benoit; Robert, François; Blanchette, Mathieu

2007-01-01

We describe PReMod, a new database of genome-wide cis-regulatory module (CRM) predictions for both the human and the mouse genomes. The prediction algorithm, described previously in Blanchette et al. (2006) Genome Res., 16, 656-668, exploits the fact that many known CRMs are made of clusters of phylogenetically conserved and repeated transcription factors (TF) binding sites. Contrary to other existing databases, PReMod is not restricted to modules located proximal to genes, but in fact mostly contains distal predicted CRMs (pCRMs). Through its web interface, PReMod allows users to (i) identify pCRMs around a gene of interest; (ii) identify pCRMs that have binding sites for a given TF (or a set of TFs) or (iii) download the entire dataset for local analyses. Queries can also be refined by filtering for specific chromosomal regions, for specific regions relative to genes or for the presence of CpG islands. The output includes information about the binding sites predicted within the selected pCRMs, and a graphical display of their distribution within the pCRMs. It also provides a visual depiction of the chromosomal context of the selected pCRMs in terms of neighboring pCRMs and genes, all of which are linked to the UCSC Genome Browser and the NCBI. PReMod: http://genomequebec.mcgill.ca/PReMod.

The lncRNA Malat1 Is Dispensable for Mouse Development but Its Transcription Plays a cis-Regulatory Role in the Adult

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Bin Zhang

2012-07-01

Full Text Available Genome-wide studies have identified thousands of long noncoding RNAs (lncRNAs lacking protein-coding capacity. However, most lncRNAs are expressed at a very low level, and in most cases there is no genetic evidence to support their in vivo function. Malat1 (metastasis associated lung adenocarcinoma transcript 1 is among the most abundant and highly conserved lncRNAs, and it exhibits an uncommon 3′-end processing mechanism. In addition, its specific nuclear localization, developmental regulation, and dysregulation in cancer are suggestive of it having a critical biological function. We have characterized a Malat1 loss-of-function genetic model that indicates that Malat1 is not essential for mouse pre- and postnatal development. Furthermore, depletion of Malat1 does not affect global gene expression, splicing factor level and phosphorylation status, or alternative pre-mRNA splicing. However, among a small number of genes that were dysregulated in adult Malat1 knockout mice, many were Malat1 neighboring genes, thus indicating a potential cis-regulatory role of Malat1 gene transcription.

Close Sequence Comparisons are Sufficient to Identify Humancis-Regulatory Elements

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Prabhakar, Shyam; Poulin, Francis; Shoukry, Malak; Afzal, Veena; Rubin, Edward M.; Couronne, Olivier; Pennacchio, Len A.

2005-12-01

Cross-species DNA sequence comparison is the primary method used to identify functional noncoding elements in human and other large genomes. However, little is known about the relative merits of evolutionarily close and distant sequence comparisons, due to the lack of a universal metric for sequence conservation, and also the paucity of empirically defined benchmark sets of cis-regulatory elements. To address this problem, we developed a general-purpose algorithm (Gumby) that detects slowly-evolving regions in primate, mammalian and more distant comparisons without requiring adjustment of parameters, and ranks conserved elements by P-value using Karlin-Altschul statistics. We benchmarked Gumby predictions against previously identified cis-regulatory elements at diverse genomic loci, and also tested numerous extremely conserved human-rodent sequences for transcriptional enhancer activity using reporter-gene assays in transgenic mice. Human regulatory elements were identified with acceptable sensitivity and specificity by comparison with 1-5 other eutherian mammals or 6 other simian primates. More distant comparisons (marsupial, avian, amphibian and fish) failed to identify many of the empirically defined functional noncoding elements. We derived an intuitive relationship between ancient and recent noncoding sequence conservation from whole genome comparative analysis, which explains some of these findings. Lastly, we determined that, in addition to strength of conservation, genomic location and/or density of surrounding conserved elements must also be considered in selecting candidate enhancers for testing at embryonic time points.

Multiple cis-regulatory elements are involved in the complex regulation of the sieve element-specific MtSEO-F1 promoter from Medicago truncatula.

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Bucsenez, M; Rüping, B; Behrens, S; Twyman, R M; Noll, G A; Prüfer, D

2012-09-01

The sieve element occlusion (SEO) gene family includes several members that are expressed specifically in immature sieve elements (SEs) in the developing phloem of dicotyledonous plants. To determine how this restricted expression profile is achieved, we analysed the SE-specific Medicago truncatula SEO-F1 promoter (PMtSEO-F1) by constructing deletion, substitution and hybrid constructs and testing them in transgenic tobacco plants using green fluorescent protein as a reporter. This revealed four promoter regions, each containing cis-regulatory elements that activate transcription in SEs. One of these segments also contained sufficient information to suppress PMtSEO-F1 transcription in the phloem companion cells (CCs). Subsequent in silico analysis revealed several candidate cis-regulatory elements that PMtSEO-F1 shares with other SEO promoters. These putative sieve element boxes (PSE boxes) are promising candidates for cis-regulatory elements controlling the SE-specific expression of PMtSEO-F1. © 2012 German Botanical Society and The Royal Botanical Society of the Netherlands.

Cis-regulatory control of the nuclear receptor Coup-TF gene in the sea urchin Paracentrotus lividus embryo.

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Lamprini G Kalampoki

Full Text Available Coup-TF, an orphan member of the nuclear receptor super family, has a fundamental role in the development of metazoan embryos. The study of the gene's regulatory circuit in the sea urchin embryo will facilitate the placement of this transcription factor in the well-studied embryonic Gene Regulatory Network (GRN. The Paracentrotus lividus Coup-TF gene (PlCoup-TF is expressed throughout embryonic development preferentially in the oral ectoderm of the gastrula and the ciliary band of the pluteus stage. Two overlapping λ genomic clones, containing three exons and upstream sequences of PlCoup-TF, were isolated from a genomic library. The transcription initiation site was determined and 5' deletions and individual segments of a 1930 bp upstream region were placed ahead of a GFP reporter cassette and injected into fertilized P.lividus eggs. Module a (-532 to -232, was necessary and sufficient to confer ciliary band expression to the reporter. Comparison of P.lividus and Strongylocentrotus purpuratus upstream Coup-TF sequences, revealed considerable conservation, but none within module a. 5' and internal deletions into module a, defined a smaller region that confers ciliary band specific expression. Putative regulatory cis-acting elements (RE1, RE2 and RE3 within module a, were specifically bound by proteins in sea urchin embryonic nuclear extracts. Site-specific mutagenesis of these elements resulted in loss of reporter activity (RE1 or ectopic expression (RE2, RE3. It is proposed that sea urchin transcription factors, which bind these three regulatory sites, are necessary for spatial and quantitative regulation of the PlCoup-TF gene at pluteus stage sea urchin embryos. These findings lead to the future identification of these factors and to the hierarchical positioning of PlCoup-TF within the embryonic GRN.

Conserved Transcriptional Regulatory Programs Underlying Rice and Barley Germination

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Lin, Li; Tian, Shulan; Kaeppler, Shawn; Liu, Zongrang; An, Yong-Qiang (Charles)

2014-01-01

Germination is a biological process important to plant development and agricultural production. Barley and rice diverged 50 million years ago, but share a similar germination process. To gain insight into the conservation of their underlying gene regulatory programs, we compared transcriptomes of barley and rice at start, middle and end points of germination, and revealed that germination regulated barley and rice genes (BRs) diverged significantly in expression patterns and/or protein sequences. However, BRs with higher protein sequence similarity tended to have more conserved expression patterns. We identified and characterized 316 sets of conserved barley and rice genes (cBRs) with high similarity in both protein sequences and expression patterns, and provided a comprehensive depiction of the transcriptional regulatory program conserved in barley and rice germination at gene, pathway and systems levels. The cBRs encoded proteins involved in a variety of biological pathways and had a wide range of expression patterns. The cBRs encoding key regulatory components in signaling pathways often had diverse expression patterns. Early germination up-regulation of cell wall metabolic pathway and peroxidases, and late germination up-regulation of chromatin structure and remodeling pathways were conserved in both barley and rice. Protein sequence and expression pattern of a gene change quickly if it is not subjected to a functional constraint. Preserving germination-regulated expression patterns and protein sequences of those cBRs for 50 million years strongly suggests that the cBRs are functionally significant and equivalent in germination, and contribute to the ancient characteristics of germination preserved in barley and rice. The functional significance and equivalence of the cBR genes predicted here can serve as a foundation to further characterize their biological functions and facilitate bridging rice and barley germination research with greater confidence. PMID

Direct activation of a notochord cis-regulatory module by Brachyury and FoxA in the ascidian Ciona intestinalis.

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Passamaneck, Yale J; Katikala, Lavanya; Perrone, Lorena; Dunn, Matthew P; Oda-Ishii, Izumi; Di Gregorio, Anna

2009-11-01

The notochord is a defining feature of the chordate body plan. Experiments in ascidian, frog and mouse embryos have shown that co-expression of Brachyury and FoxA class transcription factors is required for notochord development. However, studies on the cis-regulatory sequences mediating the synergistic effects of these transcription factors are complicated by the limited knowledge of notochord genes and cis-regulatory modules (CRMs) that are directly targeted by both. We have identified an easily testable model for such investigations in a 155-bp notochord-specific CRM from the ascidian Ciona intestinalis. This CRM contains functional binding sites for both Ciona Brachyury (Ci-Bra) and FoxA (Ci-FoxA-a). By combining point mutation analysis and misexpression experiments, we demonstrate that binding of both transcription factors to this CRM is necessary and sufficient to activate transcription. To gain insights into the cis-regulatory criteria controlling its activity, we investigated the organization of the transcription factor binding sites within the 155-bp CRM. The 155-bp sequence contains two Ci-Bra binding sites with identical core sequences but opposite orientations, only one of which is required for enhancer activity. Changes in both orientation and spacing of these sites substantially affect the activity of the CRM, as clusters of identical sites found in the Ciona genome with different arrangements are unable to activate transcription in notochord cells. This work presents the first evidence of a synergistic interaction between Brachyury and FoxA in the activation of an individual notochord CRM, and highlights the importance of transcription factor binding site arrangement for its function.

Characterization of a putative cis-regulatory element that controls transcriptional activity of the pig uroplakin II gene promoter

International Nuclear Information System (INIS)

Kwon, Deug-Nam; Park, Mi-Ryung; Park, Jong-Yi; Cho, Ssang-Goo; Park, Chankyu; Oh, Jae-Wook; Song, Hyuk; Kim, Jae-Hwan; Kim, Jin-Hoi

2011-01-01

Highlights: → The sequences of -604 to -84 bp of the pUPII promoter contained the region of a putative negative cis-regulatory element. → The core promoter was located in the 5F-1. → Transcription factor HNF4 can directly bind in the pUPII core promoter region, which plays a critical role in controlling promoter activity. → These features of the pUPII promoter are fundamental to development of a target-specific vector. -- Abstract: Uroplakin II (UPII) is a one of the integral membrane proteins synthesized as a major differentiation product of mammalian urothelium. UPII gene expression is bladder specific and differentiation dependent, but little is known about its transcription response elements and molecular mechanism. To identify the cis-regulatory elements in the pig UPII (pUPII) gene promoter region, we constructed pUPII 5' upstream region deletion mutants and demonstrated that each of the deletion mutants participates in controlling the expression of the pUPII gene in human bladder carcinoma RT4 cells. We also identified a new core promoter region and putative negative cis-regulatory element within a minimal promoter region. In addition, we showed that hepatocyte nuclear factor 4 (HNF4) can directly bind in the pUPII core promoter (5F-1) region, which plays a critical role in controlling promoter activity. Transient cotransfection experiments showed that HNF4 positively regulates pUPII gene promoter activity. Thus, the binding element and its binding protein, HNF4 transcription factor, may be involved in the mechanism that specifically regulates pUPII gene transcription.

The cis-regulatory element CCACGTGG is involved in ABA and water-stress responses of the maize gene rab28.

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Pla, M; Vilardell, J; Guiltinan, M J; Marcotte, W R; Niogret, M F; Quatrano, R S; Pagès, M

1993-01-01

The maize gene rab28 has been identified as ABA-inducible in embryos and vegetative tissues. It is also induced by water stress in young leaves. The proximal promoter region contains the conserved cis-acting element CCACGTGG (ABRE) reported for ABA induction in other plant genes. Transient expression assays in rice protoplasts indicate that a 134 bp fragment (-194 to -60 containing the ABRE) fused to a truncated cauliflower mosaic virus promoter (35S) is sufficient to confer ABA-responsiveness upon the GUS reporter gene. Gel retardation experiments indicate that nuclear proteins from tissues in which the rab28 gene is expressed can interact specifically with this 134 bp DNA fragment. Nuclear protein extracts from embryo and water-stressed leaves generate specific complexes of different electrophoretic mobility which are stable in the presence of detergent and high salt. However, by DMS footprinting the same guanine-specific contacts with the ABRE in both the embryo and leaf binding activities were detected. These results indicate that the rab28 promoter sequence CCACGTGG is a functional ABA-responsive element, and suggest that distinct regulatory factors with apparent similar affinity for the ABRE sequence may be involved in the hormone action during embryo development and in vegetative tissues subjected to osmotic stress.

Sequence conservation and combinatorial complexity of Drosophila neural precursor cell enhancers

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Kuzin Alexander

2008-08-01

Full Text Available Abstract Background The presence of highly conserved sequences within cis-regulatory regions can serve as a valuable starting point for elucidating the basis of enhancer function. This study focuses on regulation of gene expression during the early events of Drosophila neural development. We describe the use of EvoPrinter and cis-Decoder, a suite of interrelated phylogenetic footprinting and alignment programs, to characterize highly conserved sequences that are shared among co-regulating enhancers. Results Analysis of in vivo characterized enhancers that drive neural precursor gene expression has revealed that they contain clusters of highly conserved sequence blocks (CSBs made up of shorter shared sequence elements which are present in different combinations and orientations within the different co-regulating enhancers; these elements contain either known consensus transcription factor binding sites or consist of novel sequences that have not been functionally characterized. The CSBs of co-regulated enhancers share a large number of sequence elements, suggesting that a diverse repertoire of transcription factors may interact in a highly combinatorial fashion to coordinately regulate gene expression. We have used information gained from our comparative analysis to discover an enhancer that directs expression of the nervy gene in neural precursor cells of the CNS and PNS. Conclusion The combined use EvoPrinter and cis-Decoder has yielded important insights into the combinatorial appearance of fundamental sequence elements required for neural enhancer function. Each of the 30 enhancers examined conformed to a pattern of highly conserved blocks of sequences containing shared constituent elements. These data establish a basis for further analysis and understanding of neural enhancer function.

Deciphering Cis-Regulatory Element Mediated Combinatorial Regulation in Rice under Blast Infected Condition.

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Arindam Deb

Full Text Available Combinations of cis-regulatory elements (CREs present at the promoters facilitate the binding of several transcription factors (TFs, thereby altering the consequent gene expressions. Due to the eminent complexity of the regulatory mechanism, the combinatorics of CRE-mediated transcriptional regulation has been elusive. In this work, we have developed a new methodology that quantifies the co-occurrence tendencies of CREs present in a set of promoter sequences; these co-occurrence scores are filtered in three consecutive steps to test their statistical significance; and the significantly co-occurring CRE pairs are presented as networks. These networks of co-occurring CREs are further transformed to derive higher order of regulatory combinatorics. We have further applied this methodology on the differentially up-regulated gene-sets of rice tissues under fungal (Magnaporthe infected conditions to demonstrate how it helps to understand the CRE-mediated combinatorial gene regulation. Our analysis includes a wide spectrum of biologically important results. The CRE pairs having a strong tendency to co-occur often exhibit very similar joint distribution patterns at the promoters of rice. We couple the network approach with experimental results of plant gene regulation and defense mechanisms and find evidences of auto and cross regulation among TF families, cross-talk among multiple hormone signaling pathways, similarities and dissimilarities in regulatory combinatorics between different tissues, etc. Our analyses have pointed a highly distributed nature of the combinatorial gene regulation facilitating an efficient alteration in response to fungal attack. All together, our proposed methodology could be an important approach in understanding the combinatorial gene regulation. It can be further applied to unravel the tissue and/or condition specific combinatorial gene regulation in other eukaryotic systems with the availability of annotated genomic

The MTP1 promoters from Arabidopsis halleri reveal cis-regulating elements for the evolution of metal tolerance.

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Fasani, Elisa; DalCorso, Giovanni; Varotto, Claudio; Li, Mingai; Visioli, Giovanna; Mattarozzi, Monica; Furini, Antonella

2017-06-01

In the hyperaccumulator Arabidopsis halleri, the zinc (Zn) vacuolar transporter MTP1 is a key component of hypertolerance. Because protein sequences and functions are highly conserved between A. halleri and Arabidopsis thaliana, Zn tolerance in A. halleri may reflect the constitutively higher MTP1 expression compared with A. thaliana, based on copy number expansion and different cis regulation. Three MTP1 promoters were characterized in A. halleri ecotype I16. The comparison with the A. thaliana MTP1 promoter revealed different expression profiles correlated with specific cis-acting regulatory elements. The MTP1 5' untranslated region, highly conserved among A. thaliana, Arabidopsis lyrata and A. halleri, contains a dimer of MYB-binding motifs in the A. halleri promoters absent in the A. thaliana and A. lyrata sequences. Site-directed mutagenesis of these motifs revealed their role for expression in trichomes. A. thaliana mtp1 transgenic lines expressing AtMTP1 controlled by the native A. halleri promoter were more Zn-tolerant than lines carrying mutations on MYB-binding motifs. Differences in Zn tolerance were associated with different distribution of Zn among plant organs and in trichomes. The different cis-acting elements in the MTP1 promoters of A. halleri, particularly the MYB-binding sites, are probably involved in the evolution of Zn tolerance. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Structural and functional analysis of mouse Msx1 gene promoter: sequence conservation with human MSX1 promoter points at potential regulatory elements.

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Gonzalez, S M; Ferland, L H; Robert, B; Abdelhay, E

1998-06-01

Vertebrate Msx genes are related to one of the most divergent homeobox genes of Drosophila, the muscle segment homeobox (msh) gene, and are expressed in a well-defined pattern at sites of tissue interactions. This pattern of expression is conserved in vertebrates as diverse as quail, zebrafish, and mouse in a range of sites including neural crest, appendages, and craniofacial structures. In the present work, we performed structural and functional analyses in order to identify potential cis-acting elements that may be regulating Msx1 gene expression. To this end, a 4.9-kb segment of the 5'-flanking region was sequenced and analyzed for transcription-factor binding sites. Four regions showing a high concentration of these sites were identified. Transfection assays with fragments of regulatory sequences driving the expression of the bacterial lacZ reporter gene showed that a region of 4 kb upstream of the transcription start site contains positive and negative elements responsible for controlling gene expression. Interestingly, a fragment of 130 bp seems to contain the minimal elements necessary for gene expression, as its removal completely abolishes gene expression in cultured cells. These results are reinforced by comparison of this region with the human Msx1 gene promoter, which shows extensive conservation, including many consensus binding sites, suggesting a regulatory role for them.

Spatially conserved regulatory elements identified within human and mouse Cd247 gene using high-throughput sequencing data from the ENCODE project

DEFF Research Database (Denmark)

Pundhir, Sachin; Hannibal, Tine Dahlbæk; Bang-Berthelsen, Claus Heiner

2014-01-01

. In this study, we have utilized the wealth of high-throughput sequencing data produced during the Encyclopedia of DNA Elements (ENCODE) project to identify spatially conserved regulatory elements within the Cd247 gene from human and mouse. We show the presence of two transcription factor binding sites...

PlantPAN: Plant promoter analysis navigator, for identifying combinatorial cis-regulatory elements with distance constraint in plant gene groups

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Huang Hsien-Da

2008-11-01

Full Text Available Abstract Background The elucidation of transcriptional regulation in plant genes is important area of research for plant scientists, following the mapping of various plant genomes, such as A. thaliana, O. sativa and Z. mays. A variety of bioinformatic servers or databases of plant promoters have been established, although most have been focused only on annotating transcription factor binding sites in a single gene and have neglected some important regulatory elements (tandem repeats and CpG/CpNpG islands in promoter regions. Additionally, the combinatorial interaction of transcription factors (TFs is important in regulating the gene group that is associated with the same expression pattern. Therefore, a tool for detecting the co-regulation of transcription factors in a group of gene promoters is required. Results This study develops a database-assisted system, PlantPAN (Plant Promoter Analysis Navigator, for recognizing combinatorial cis-regulatory elements with a distance constraint in sets of plant genes. The system collects the plant transcription factor binding profiles from PLACE, TRANSFAC (public release 7.0, AGRIS, and JASPER databases and allows users to input a group of gene IDs or promoter sequences, enabling the co-occurrence of combinatorial transcription factor binding sites (TFBSs within a defined distance (20 bp to 200 bp to be identified. Furthermore, the new resource enables other regulatory features in a plant promoter, such as CpG/CpNpG islands and tandem repeats, to be displayed. The regulatory elements in the conserved regions of the promoters across homologous genes are detected and presented. Conclusion In addition to providing a user-friendly input/output interface, PlantPAN has numerous advantages in the analysis of a plant promoter. Several case studies have established the effectiveness of PlantPAN. This novel analytical resource is now freely available at http://PlantPAN.mbc.nctu.edu.tw.

Multiple Functional Variants in cis Modulate PDYN Expression.

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Babbitt, Courtney C; Silverman, Jesse S; Haygood, Ralph; Reininga, Jennifer M; Rockman, Matthew V; Wray, Gregory A

2010-02-01

Understanding genetic variation and its functional consequences within cis-regulatory regions remains an important challenge in human genetics and evolution. Here, we present a fine-scale functional analysis of segregating variation within the cis-regulatory region of prodynorphin, a gene that encodes an endogenous opioid precursor with roles in cognition and disease. In order to characterize the functional consequences of segregating variation in cis in a region under balancing selection in different human populations, we examined associations between specific polymorphisms and gene expression in vivo and in vitro. We identified five polymorphisms within the 5' flanking region that affect transcript abundance: a 68-bp repeat recognized in prior studies, as well as two microsatellites and two single nucleotide polymorphisms not previously implicated as functional variants. The impact of these variants on transcription differs by brain region, sex, and cell type, implying interactions between cis genotype and the differentiated state of cells. The effects of individual variants on expression level are not additive in some combinations, implying epistatic interactions between nearby variants. These data reveal an unexpectedly complex relationship between segregating genetic variation and its expression-trait consequences and highlights the importance of close functional scrutiny of natural genetic variation within even relatively well-studied cis-regulatory regions.

Changes in Cis-regulatory Elements during Morphological Evolution

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Yu-Lee Paul

2012-10-01

Full Text Available How have animals evolved new body designs (morphological evolution? This requires explanations both for simple morphological changes, such as differences in pigmentation and hair patterns between different Drosophila populations and species, and also for more complex changes, such as differences in the forelimbs of mice and bats, and the necks of amphibians and reptiles. The genetic changes and pathways involved in these evolutionary steps require identification. Many, though not all, of these events occur by changes in cis-regulatory (enhancer elements within developmental genes. Enhancers are modular, each affecting expression in only one or a few tissues. Therefore it is possible to add, remove or alter an enhancer without producing changes in multiple tissues, and thereby avoid widespread (pleiotropic deleterious effects. Ideally, for a given step in morphological evolution it is necessary to identify (i the change in phenotype, (ii the changes in gene expression, (iii the DNA region, enhancer or otherwise, affected, (iv the mutation involved, (v the nature of the transcription or other factors that bind to this site. In practice these data are incomplete for most of the published studies upon morphological evolution. Here, the investigations are categorized according to how far these analyses have proceeded.

CORECLUST: identification of the conserved CRM grammar together with prediction of gene regulation.

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Nikulova, Anna A; Favorov, Alexander V; Sutormin, Roman A; Makeev, Vsevolod J; Mironov, Andrey A

2012-07-01

Identification of transcriptional regulatory regions and tracing their internal organization are important for understanding the eukaryotic cell machinery. Cis-regulatory modules (CRMs) of higher eukaryotes are believed to possess a regulatory 'grammar', or preferred arrangement of binding sites, that is crucial for proper regulation and thus tends to be evolutionarily conserved. Here, we present a method CORECLUST (COnservative REgulatory CLUster STructure) that predicts CRMs based on a set of positional weight matrices. Given regulatory regions of orthologous and/or co-regulated genes, CORECLUST constructs a CRM model by revealing the conserved rules that describe the relative location of binding sites. The constructed model may be consequently used for the genome-wide prediction of similar CRMs, and thus detection of co-regulated genes, and for the investigation of the regulatory grammar of the system. Compared with related methods, CORECLUST shows better performance at identification of CRMs conferring muscle-specific gene expression in vertebrates and early-developmental CRMs in Drosophila.

Shared regulatory sites are abundant in the human genome and shed light on genome evolution and disease pleiotropy.

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Tong, Pin; Monahan, Jack; Prendergast, James G D

2017-03-01

Large-scale gene expression datasets are providing an increasing understanding of the location of cis-eQTLs in the human genome and their role in disease. However, little is currently known regarding the extent of regulatory site-sharing between genes. This is despite it having potentially wide-ranging implications, from the determination of the way in which genetic variants may shape multiple phenotypes to the understanding of the evolution of human gene order. By first identifying the location of non-redundant cis-eQTLs, we show that regulatory site-sharing is a relatively common phenomenon in the human genome, with over 10% of non-redundant regulatory variants linked to the expression of multiple nearby genes. We show that these shared, local regulatory sites are linked to high levels of chromatin looping between the regulatory sites and their associated genes. In addition, these co-regulated gene modules are found to be strongly conserved across mammalian species, suggesting that shared regulatory sites have played an important role in shaping human gene order. The association of these shared cis-eQTLs with multiple genes means they also appear to be unusually important in understanding the genetics of human phenotypes and pleiotropy, with shared regulatory sites more often linked to multiple human phenotypes than other regulatory variants. This study shows that regulatory site-sharing is likely an underappreciated aspect of gene regulation and has important implications for the understanding of various biological phenomena, including how the two and three dimensional structures of the genome have been shaped and the potential causes of disease pleiotropy outside coding regions.

A HLA class I cis-regulatory element whose activity can be modulated by hormones.

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Sim, B C; Hui, K M

1994-12-01

To elucidate the basis of the down-regulation in major histocompatibility complex (MHC) class I gene expression and to identify possible DNA-binding regulatory elements that have the potential to interact with class I MHC genes, we have studied the transcriptional regulation of class I HLA genes in human breast carcinoma cells. A 9 base pair (bp) negative cis-regulatory element (NRE) has been identified using band-shift assays employing DNA sequences derived from the 5'-flanking region of HLA class I genes. This 9-bp element, GTCATGGCG, located within exon I of the HLA class I gene, can potently inhibit the expression of a heterologous thymidine kinase (TK) gene promoter and the HLA enhancer element. Furthermore, this regulatory element can exert its suppressive function in either the sense or anti-sense orientation. More interestingly, NRE can suppress dexamethasone-mediated gene activation in the context of the reported glucocorticoid-responsive element (GRE) in MCF-7 cells but has no influence on the estrogen-mediated transcriptional activation of MCF-7 cells in the context of the reported estrogen-responsive element (ERE). Furthermore, the presence of such a regulatory element within the HLA class I gene whose activity can be modulated by hormones correlates well with our observation that the level of HLA class I gene expression can be down-regulated by hormones in human breast carcinoma cells. Such interactions between negative regulatory elements and specific hormone trans-activators are novel and suggest a versatile form of transcriptional control.

PlantCARE, a database of plant cis-acting regulatory elements and a portal to tools for in silico analysis of promoter sequences

OpenAIRE

Lescot, Magali; Déhais, Patrice; Thijs, Gert; Marchal, Kathleen; Moreau, Yves; Van de Peer, Yves; Rouzé, Pierre; Rombauts, Stephane

2002-01-01

PlantCARE is a database of plant cis-acting regulatory elements, enhancers and repressors. Regulatory elements are represented by positional matrices, consensus sequences and individual sites on particular promoter sequences. Links to the EMBL, TRANSFAC and MEDLINE databases are provided when available. Data about the transcription sites are extracted mainly from the literature, supplemented with an increasing number of in silico predicted data. Apart from a general description for specific t...

An integrative and applicable phylogenetic footprinting framework for cis-regulatory motifs identification in prokaryotic genomes.

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Liu, Bingqiang; Zhang, Hanyuan; Zhou, Chuan; Li, Guojun; Fennell, Anne; Wang, Guanghui; Kang, Yu; Liu, Qi; Ma, Qin

2016-08-09

Phylogenetic footprinting is an important computational technique for identifying cis-regulatory motifs in orthologous regulatory regions from multiple genomes, as motifs tend to evolve slower than their surrounding non-functional sequences. Its application, however, has several difficulties for optimizing the selection of orthologous data and reducing the false positives in motif prediction. Here we present an integrative phylogenetic footprinting framework for accurate motif predictions in prokaryotic genomes (MP(3)). The framework includes a new orthologous data preparation procedure, an additional promoter scoring and pruning method and an integration of six existing motif finding algorithms as basic motif search engines. Specifically, we collected orthologous genes from available prokaryotic genomes and built the orthologous regulatory regions based on sequence similarity of promoter regions. This procedure made full use of the large-scale genomic data and taxonomy information and filtered out the promoters with limited contribution to produce a high quality orthologous promoter set. The promoter scoring and pruning is implemented through motif voting by a set of complementary predicting tools that mine as many motif candidates as possible and simultaneously eliminate the effect of random noise. We have applied the framework to Escherichia coli k12 genome and evaluated the prediction performance through comparison with seven existing programs. This evaluation was systematically carried out at the nucleotide and binding site level, and the results showed that MP(3) consistently outperformed other popular motif finding tools. We have integrated MP(3) into our motif identification and analysis server DMINDA, allowing users to efficiently identify and analyze motifs in 2,072 completely sequenced prokaryotic genomes. The performance evaluation indicated that MP(3) is effective for predicting regulatory motifs in prokaryotic genomes. Its application may enhance

Comparative evaluation of capillary electrophoresis and high-performance liquid chromatography for the separation of cis-cis, cis-trans, and trans-trans isomers of atracurium besylate.

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de Moraes, M de L; Polakiewicz, B; Mattua, M F; Tavares, M F

1998-01-01

Atracurium besylate is a highly selective nondepolarizing neuromuscular blocking agent routinely used during anesthetic procedures. The commercial presentation of this drug is a mixture of positional isomers, cis-cis, cis-trans, and trans-trans. Reversed-phase high-performance liquid chromatography has been the technique of choice for the analysis of atracurium besylate formulations at the quality control laboratory of Núcleo de Desenvolvimento Cristália (São Paulo, Brazil), a local pharmaceutical company. HPLC analysis is usually conducted under gradient elution using acetonitrile/0.1 M phosphate buffer eluent mixture as mobile phase and an octadecyl silica (ODS)-packed column. The complete elution of the three isomers takes about 1 hr. In this work, an alternative capillary electrophoresis methodology was developed. The complete resolution of all three isomers was accomplished in about 13 min (+20 kV/72 cm, 211 nm direct detection) using a 60-mM phosphate buffer solution (pH 4) containing 20 mM beta-cyclodextrin and 4 M urea. The isomer ratio was found to be 59.1% cis-cis, 35.9% cis-trans, and 5.02% trans-trans (expected ratio: 59:35:6). Laudanosine, a major metabolite of atracurium besylate, was identified in two commercially available formulations, Tracur (Núcleo de Desenvolvimento Cristália) and Tracrium (Glaxo Wellcome, S.A., Rio de Janeiro, Brazil). Its concentration increases considerably during storage of the product, even if the product is stored at low temperatures.

RNA expression in a cartilaginous fish cell line reveals ancient 3′ noncoding regions highly conserved in vertebrates

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Forest, David; Nishikawa, Ryuhei; Kobayashi, Hiroshi; Parton, Angela; Bayne, Christopher J.; Barnes, David W.

2007-01-01

We have established a cartilaginous fish cell line [Squalus acanthias embryo cell line (SAE)], a mesenchymal stem cell line derived from the embryo of an elasmobranch, the spiny dogfish shark S. acanthias. Elasmobranchs (sharks and rays) first appeared >400 million years ago, and existing species provide useful models for comparative vertebrate cell biology, physiology, and genomics. Comparative vertebrate genomics among evolutionarily distant organisms can provide sequence conservation information that facilitates identification of critical coding and noncoding regions. Although these genomic analyses are informative, experimental verification of functions of genomic sequences depends heavily on cell culture approaches. Using ESTs defining mRNAs derived from the SAE cell line, we identified lengthy and highly conserved gene-specific nucleotide sequences in the noncoding 3′ UTRs of eight genes involved in the regulation of cell growth and proliferation. Conserved noncoding 3′ mRNA regions detected by using the shark nucleotide sequences as a starting point were found in a range of other vertebrate orders, including bony fish, birds, amphibians, and mammals. Nucleotide identity of shark and human in these regions was remarkably well conserved. Our results indicate that highly conserved gene sequences dating from the appearance of jawed vertebrates and representing potential cis-regulatory elements can be identified through the use of cartilaginous fish as a baseline. Because the expression of genes in the SAE cell line was prerequisite for their identification, this cartilaginous fish culture system also provides a physiologically valid tool to test functional hypotheses on the role of these ancient conserved sequences in comparative cell biology. PMID:17227856

PAA, WSH, and CIS Overview Self-Study #47656

Energy Technology Data Exchange (ETDEWEB)

Schroeder, Rachel Anne [Los Alamos National Lab. (LANL), Los Alamos, NM (United States)

2017-09-14

This course presents an overview of the Department of Energy’s (DOE’s) regulatory requirements relevant to the Price-Anderson Amendments Act (PAAA, also referred to as nuclear safety), worker safety and health (WSH), and classified information security (CIS) that are enforceable under the DOE enforcement program; describes the DOE enforcement process; and provides an overview of Los Alamos National Laboratory’s (LANL’s) internal compliance program relative to these DOE regulatory requirements. The LANL PAAA Program is responsible for maintaining LANL’s internal compliance program, which ensures the prompt identification, screening, and reporting of noncompliances to DOE regulatory requirements pertaining to nuclear safety, WSH, and CIS to build the strongest mitigation position for the Laboratory with respect to civil or other penalties.

Lmx1b-targeted cis-regulatory modules involved in limb dorsalization.

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Haro, Endika; Watson, Billy A; Feenstra, Jennifer M; Tegeler, Luke; Pira, Charmaine U; Mohan, Subburaman; Oberg, Kerby C

2017-06-01

Lmx1b is a homeodomain transcription factor responsible for limb dorsalization. Despite striking double-ventral (loss-of-function) and double-dorsal (gain-of-function) limb phenotypes, no direct gene targets in the limb have been confirmed. To determine direct targets, we performed a chromatin immunoprecipitation against Lmx1b in mouse limbs at embryonic day 12.5 followed by next-generation sequencing (ChIP-seq). Nearly 84% ( n =617) of the Lmx1b-bound genomic intervals (LBIs) identified overlap with chromatin regulatory marks indicative of potential cis -regulatory modules (PCRMs). In addition, 73 LBIs mapped to CRMs that are known to be active during limb development. We compared Lmx1b-bound PCRMs with genes regulated by Lmx1b and found 292 PCRMs within 1 Mb of 254 Lmx1b-regulated genes. Gene ontological analysis suggests that Lmx1b targets extracellular matrix production, bone/joint formation, axonal guidance, vascular development, cell proliferation and cell movement. We validated the functional activity of a PCRM associated with joint-related Gdf5 that provides a mechanism for Lmx1b-mediated joint modification and a PCRM associated with Lmx1b that suggests a role in autoregulation. This is the first report to describe genome-wide Lmx1b binding during limb development, directly linking Lmx1b to targets that accomplish limb dorsalization. © 2017. Published by The Company of Biologists Ltd.

Thermodynamic state ensemble models of cis-regulation.

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Marc S Sherman

Full Text Available A major goal in computational biology is to develop models that accurately predict a gene's expression from its surrounding regulatory DNA. Here we present one class of such models, thermodynamic state ensemble models. We describe the biochemical derivation of the thermodynamic framework in simple terms, and lay out the mathematical components that comprise each model. These components include (1 the possible states of a promoter, where a state is defined as a particular arrangement of transcription factors bound to a DNA promoter, (2 the binding constants that describe the affinity of the protein-protein and protein-DNA interactions that occur in each state, and (3 whether each state is capable of transcribing. Using these components, we demonstrate how to compute a cis-regulatory function that encodes the probability of a promoter being active. Our intention is to provide enough detail so that readers with little background in thermodynamics can compose their own cis-regulatory functions. To facilitate this goal, we also describe a matrix form of the model that can be easily coded in any programming language. This formalism has great flexibility, which we show by illustrating how phenomena such as competition between transcription factors and cooperativity are readily incorporated into these models. Using this framework, we also demonstrate that Michaelis-like functions, another class of cis-regulatory models, are a subset of the thermodynamic framework with specific assumptions. By recasting Michaelis-like functions as thermodynamic functions, we emphasize the relationship between these models and delineate the specific circumstances representable by each approach. Application of thermodynamic state ensemble models is likely to be an important tool in unraveling the physical basis of combinatorial cis-regulation and in generating formalisms that accurately predict gene expression from DNA sequence.

Phylogeny based discovery of regulatory elements

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Cohen Barak A

2006-05-01

Full Text Available Abstract Background Algorithms that locate evolutionarily conserved sequences have become powerful tools for finding functional DNA elements, including transcription factor binding sites; however, most methods do not take advantage of an explicit model for the constrained evolution of functional DNA sequences. Results We developed a probabilistic framework that combines an HKY85 model, which assigns probabilities to different base substitutions between species, and weight matrix models of transcription factor binding sites, which describe the probabilities of observing particular nucleotides at specific positions in the binding site. The method incorporates the phylogenies of the species under consideration and takes into account the position specific variation of transcription factor binding sites. Using our framework we assessed the suitability of alignments of genomic sequences from commonly used species as substrates for comparative genomic approaches to regulatory motif finding. We then applied this technique to Saccharomyces cerevisiae and related species by examining all possible six base pair DNA sequences (hexamers and identifying sequences that are conserved in a significant number of promoters. By combining similar conserved hexamers we reconstructed known cis-regulatory motifs and made predictions of previously unidentified motifs. We tested one prediction experimentally, finding it to be a regulatory element involved in the transcriptional response to glucose. Conclusion The experimental validation of a regulatory element prediction missed by other large-scale motif finding studies demonstrates that our approach is a useful addition to the current suite of tools for finding regulatory motifs.

Transformation of Migration Flows Between the Russian Far East and CIS and non-CIS States

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Motrich E. L.

2010-06-01

Full Text Available Basic trends in the migration processes in the Russian Far East are shown. Special emphasis is placed on the transformation of migration interactions with CIS and non-CIS countries both at the level of the region as a whole, and at the level of the Far Eastern territories of the Russian Federation. An extent of using foreign labor in different periods of the Russian Far East socio-economic development and the regulatory support of this process are shown. Prospects for attracting and utilizing foreign labor are stated

Quantitative statistical analysis of cis-regulatory sequences in ABA/VP1- and CBF/DREB1-regulated genes of Arabidopsis.

Science.gov (United States)

Suzuki, Masaharu; Ketterling, Matthew G; McCarty, Donald R

2005-09-01

We have developed a simple quantitative computational approach for objective analysis of cis-regulatory sequences in promoters of coregulated genes. The program, designated MotifFinder, identifies oligo sequences that are overrepresented in promoters of coregulated genes. We used this approach to analyze promoter sequences of Viviparous1 (VP1)/abscisic acid (ABA)-regulated genes and cold-regulated genes, respectively, of Arabidopsis (Arabidopsis thaliana). We detected significantly enriched sequences in up-regulated genes but not in down-regulated genes. This result suggests that gene activation but not repression is mediated by specific and common sequence elements in promoters. The enriched motifs include several known cis-regulatory sequences as well as previously unidentified motifs. With respect to known cis-elements, we dissected the flanking nucleotides of the core sequences of Sph element, ABA response elements (ABREs), and the C repeat/dehydration-responsive element. This analysis identified the motif variants that may correlate with qualitative and quantitative differences in gene expression. While both VP1 and cold responses are mediated in part by ABA signaling via ABREs, these responses correlate with unique ABRE variants distinguished by nucleotides flanking the ACGT core. ABRE and Sph motifs are tightly associated uniquely in the coregulated set of genes showing a strict dependence on VP1 and ABA signaling. Finally, analysis of distribution of the enriched sequences revealed a striking concentration of enriched motifs in a proximal 200-base region of VP1/ABA and cold-regulated promoters. Overall, each class of coregulated genes possesses a discrete set of the enriched motifs with unique distributions in their promoters that may account for the specificity of gene regulation.

Identification of conserved regulatory elements by comparative genome analysis

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Jareborg Niclas

2003-05-01

Full Text Available Abstract Background For genes that have been successfully delineated within the human genome sequence, most regulatory sequences remain to be elucidated. The annotation and interpretation process requires additional data resources and significant improvements in computational methods for the detection of regulatory regions. One approach of growing popularity is based on the preferential conservation of functional sequences over the course of evolution by selective pressure, termed 'phylogenetic footprinting'. Mutations are more likely to be disruptive if they appear in functional sites, resulting in a measurable difference in evolution rates between functional and non-functional genomic segments. Results We have devised a flexible suite of methods for the identification and visualization of conserved transcription-factor-binding sites. The system reports those putative transcription-factor-binding sites that are both situated in conserved regions and located as pairs of sites in equivalent positions in alignments between two orthologous sequences. An underlying collection of metazoan transcription-factor-binding profiles was assembled to facilitate the study. This approach results in a significant improvement in the detection of transcription-factor-binding sites because of an increased signal-to-noise ratio, as demonstrated with two sets of promoter sequences. The method is implemented as a graphical web application, ConSite, which is at the disposal of the scientific community at http://www.phylofoot.org/. Conclusions Phylogenetic footprinting dramatically improves the predictive selectivity of bioinformatic approaches to the analysis of promoter sequences. ConSite delivers unparalleled performance using a novel database of high-quality binding models for metazoan transcription factors. With a dynamic interface, this bioinformatics tool provides broad access to promoter analysis with phylogenetic footprinting.

Expression of the central growth regulator BIG BROTHER is regulated by multiple cis-elements

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Breuninger Holger

2012-03-01

Full Text Available Abstract Background Much of the organismal variation we observe in nature is due to differences in organ size. The observation that even closely related species can show large, stably inherited differences in organ size indicates a strong genetic component to the control of organ size. Despite recent progress in identifying factors controlling organ growth in plants, our overall understanding of this process remains limited, partly because the individual factors have not yet been connected into larger regulatory pathways or networks. To begin addressing this aim, we have studied the upstream regulation of expression of BIG BROTHER (BB, a central growth-control gene in Arabidopsis thaliana that prevents overgrowth of organs. Final organ size and BB expression levels are tightly correlated, implying the need for precise control of its expression. BB expression mirrors proliferative activity, yet the gene functions to limit proliferation, suggesting that it acts in an incoherent feedforward loop downstream of growth activators to prevent over-proliferation. Results To investigate the upstream regulation of BB we combined a promoter deletion analysis with a phylogenetic footprinting approach. We were able to narrow down important, highly conserved, cis-regulatory elements within the BB promoter. Promoter sequences of other Brassicaceae species were able to partially complement the A. thaliana bb-1 mutant, suggesting that at least within the Brassicaceae family the regulatory pathways are conserved. Conclusions This work underlines the complexity involved in precise quantitative control of gene expression and lays the foundation for identifying important upstream regulators that determine BB expression levels and thus final organ size.

Mapping of cis-regulatory sites in the promoter of testis-specific stellate genes of Drosophila melanogaster.

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Olenkina, O M; Egorova, K S; Aravin, A A; Naumova, N M; Gvozdev, V A; Olenina, L V

2012-11-01

Tandem Stellate genes organized into two clusters in heterochromatin and euchromatin of the X-chromosome are part of the Ste-Su(Ste) genetic system required for maintenance of male fertility and reproduction of Drosophila melanogaster. Stellate genes encode a regulatory subunit of protein kinase CK2 and are the main targets of germline-specific piRNA-silencing; their derepression leads to appearance of protein crystals in spermatocytes, meiotic disturbances, and male sterility. A short promoter region of 134 bp appears to be sufficient for testis-specific transcription of Stellate, and it contains three closely located cis-regulatory elements called E-boxes. By using reporter analysis, we confirmed a strong functionality of the E-boxes in the Stellate promoter for in vivo transcription. Using selective mutagenesis, we have shown that the presence of the central E-box 2 is preferable to maintain a high-level testis-specific transcription of the reporter gene under the Stellate promoter. The Stellate promoter provides transcription even in heterochromatin, and corresponding mRNAs are translated with the generation of full-size protein products in case of disturbances in the piRNA-silencing process. We have also shown for the first time that the activity of the Stellate promoter is determined by chromatin context of the X-chromosome in male germinal cells, and it increases at about twofold when relocating in autosomes.

cis-Acting Complex-Trait-Associated lincRNA Expression Correlates with Modulation of Chromosomal Architecture

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Jennifer Yihong Tan

2017-02-01

Full Text Available Summary: Intergenic long noncoding RNAs (lincRNAs are the largest class of transcripts in the human genome. Although many have recently been linked to complex human traits, the underlying mechanisms for most of these transcripts remain undetermined. We investigated the regulatory roles of a high-confidence and reproducible set of 69 trait-relevant lincRNAs (TR-lincRNAs in human lymphoblastoid cells whose biological relevance is supported by their evolutionary conservation during recent human history and genetic interactions with other trait-associated loci. Their enrichment in enhancer-like chromatin signatures, interactions with nearby trait-relevant protein-coding loci, and preferential location at topologically associated domain (TAD boundaries provide evidence that TR-lincRNAs likely regulate proximal trait-relevant gene expression in cis by modulating local chromosomal architecture. This is consistent with the positive and significant correlation found between TR-lincRNA abundance and intra-TAD DNA-DNA contacts. Our results provide insights into the molecular mode of action by which TR-lincRNAs contribute to complex human traits. : Tan et al. identify and characterize 69 human complex trait/disease-associated lincRNAs in LCLs. They show that these loci are often associated with cis-regulation of gene expression and tend to be localized at TAD boundaries, suggesting that these lincRNAs may influence chromosomal architecture. Keywords: intergenic long noncoding RNA, lincRNA, GWAS, expression quantitative trait loci, eQTL, complex trait and disease, enhancer, cis-regulation, topologically associated domains, TAD

PROSPECT improves cis-acting regulatory element prediction by integrating expression profile data with consensus pattern searches

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Fujibuchi, Wataru; Anderson, John S. J.; Landsman, David

2001-01-01

Consensus pattern and matrix-based searches designed to predict cis-acting transcriptional regulatory sequences have historically been subject to large numbers of false positives. We sought to decrease false positives by incorporating expression profile data into a consensus pattern-based search method. We have systematically analyzed the expression phenotypes of over 6000 yeast genes, across 121 expression profile experiments, and correlated them with the distribution of 14 known regulatory elements over sequences upstream of the genes. Our method is based on a metric we term probabilistic element assessment (PEA), which is a ranking of potential sites based on sequence similarity in the upstream regions of genes with similar expression phenotypes. For eight of the 14 known elements that we examined, our method had a much higher selectivity than a naïve consensus pattern search. Based on our analysis, we have developed a web-based tool called PROSPECT, which allows consensus pattern-based searching of gene clusters obtained from microarray data. PMID:11574681

Microevolution of cis-regulatory elements: an example from the pair-rule segmentation gene fushi tarazu in the Drosophila melanogaster subgroup.

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Mohammed Bakkali

Full Text Available The importance of non-coding DNAs that control transcription is ever noticeable, but the characterization and analysis of the evolution of such DNAs presents challenges not found in the analysis of coding sequences. In this study of the cis-regulatory elements of the pair rule segmentation gene fushi tarazu (ftz I report the DNA sequences of ftz's zebra element (promoter and a region containing the proximal enhancer from a total of 45 fly lines belonging to several populations of the species Drosophila melanogaster, D. simulans, D. sechellia, D. mauritiana, D. yakuba, D. teissieri, D. orena and D. erecta. Both elements evolve at slower rate than ftz synonymous sites, thus reflecting their functional importance. The promoter evolves more slowly than the average for ftz's coding sequence while, on average, the enhancer evolves more rapidly, suggesting more functional constraint and effective purifying selection on the former. Comparative analysis of the number and nature of base substitutions failed to detect significant evidence for positive/adaptive selection in transcription-factor-binding sites. These seem to evolve at similar rates to regions not known to bind transcription factors. Although this result reflects the evolutionary flexibility of the transcription factor binding sites, it also suggests a complex and still not completely understood nature of even the characterized cis-regulatory sequences. The latter seem to contain more functional parts than those currently identified, some of which probably transcription factor binding. This study illustrates ways in which functional assignments of sequences within cis-acting sequences can be used in the search for adaptive evolution, but also highlights difficulties in how such functional assignment and analysis can be carried out.

Functional conservation of the Drosophila gooseberry gene and its evolutionary alleles.

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Wei Liu

Full Text Available The Drosophila Pax gene gooseberry (gsb is required for development of the larval cuticle and CNS, survival to adulthood, and male fertility. These functions can be rescued in gsb mutants by two gsb evolutionary alleles, gsb-Prd and gsb-Pax3, which express the Drosophila Paired and mouse Pax3 proteins under the control of gooseberry cis-regulatory region. Therefore, both Paired and Pax3 proteins have conserved all the Gsb functions that are required for survival of embryos to fertile adults, despite the divergent primary sequences in their C-terminal halves. As gsb-Prd and gsb-Pax3 uncover a gsb function involved in male fertility, construction of evolutionary alleles may provide a powerful strategy to dissect hitherto unknown gene functions. Our results provide further evidence for the essential role of cis-regulatory regions in the functional diversification of duplicated genes during evolution.

Viscometric study of high-cis polybutadiene in toluene solution

International Nuclear Information System (INIS)

Mwllo, Ivana L.; Coutinho, Fernanda M.B.; Delpech, Marcia C.; Albino, Fernanda F.M.

2006-01-01

Viscometric measurements, in toluene solution at 30 deg C, were performed with high-cis polybutadiene synthesized by neodymium based catalyst. Six different equations were used to calculate intrinsic viscosities and viscosimetric constant values: Huggins, Kraemer, Martin and Schulz-Blaschke by graphic extrapolation, and Solomon-Ciuta, Deb-Chanterjee and again Schulz- Blaschke, through a single point determination. The molecular weight of the polymers was also determined applying Mark-Houwink-Sakurada equation using the values of intrinsic viscosity obtained by the six equations. The values of intrinsic viscosity and viscosity-average molecular weight obtained by the two methods were compared in order to verify the validity of the single point determination for high-cis polybutadiene. (author)

Identification of Cis-Acting Promoter Elements in Cold- and Dehydration-Induced Transcriptional Pathways in Arabidopsis, Rice, and Soybean

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Maruyama, Kyonoshin; Todaka, Daisuke; Mizoi, Junya; Yoshida, Takuya; Kidokoro, Satoshi; Matsukura, Satoko; Takasaki, Hironori; Sakurai, Tetsuya; Yamamoto, Yoshiharu Y.; Yoshiwara, Kyouko; Kojima, Mikiko; Sakakibara, Hitoshi; Shinozaki, Kazuo; Yamaguchi-Shinozaki, Kazuko

2012-01-01

The genomes of three plants, Arabidopsis (Arabidopsis thaliana), rice (Oryza sativa), and soybean (Glycine max), have been sequenced, and their many genes and promoters have been predicted. In Arabidopsis, cis-acting promoter elements involved in cold- and dehydration-responsive gene expression have been extensively analysed; however, the characteristics of such cis-acting promoter sequences in cold- and dehydration-inducible genes of rice and soybean remain to be clarified. In this study, we performed microarray analyses using the three species, and compared characteristics of identified cold- and dehydration-inducible genes. Transcription profiles of the cold- and dehydration-responsive genes were similar among these three species, showing representative upregulated (dehydrin/LEA) and downregulated (photosynthesis-related) genes. All (46 = 4096) hexamer sequences in the promoters of the three species were investigated, revealing the frequency of conserved sequences in cold- and dehydration-inducible promoters. A core sequence of the abscisic acid-responsive element (ABRE) was the most conserved in dehydration-inducible promoters of all three species, suggesting that transcriptional regulation for dehydration-inducible genes is similar among these three species, with the ABRE-dependent transcriptional pathway. In contrast, for cold-inducible promoters, the conserved hexamer sequences were diversified among these three species, suggesting the existence of diverse transcriptional regulatory pathways for cold-inducible genes among the species. PMID:22184637

CONDOR: a database resource of developmentally associated conserved non-coding elements

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Smith Sarah

2007-08-01

Full Text Available Abstract Background Comparative genomics is currently one of the most popular approaches to study the regulatory architecture of vertebrate genomes. Fish-mammal genomic comparisons have proved powerful in identifying conserved non-coding elements likely to be distal cis-regulatory modules such as enhancers, silencers or insulators that control the expression of genes involved in the regulation of early development. The scientific community is showing increasing interest in characterizing the function, evolution and language of these sequences. Despite this, there remains little in the way of user-friendly access to a large dataset of such elements in conjunction with the analysis and the visualization tools needed to study them. Description Here we present CONDOR (COnserved Non-coDing Orthologous Regions available at: http://condor.fugu.biology.qmul.ac.uk. In an interactive and intuitive way the website displays data on > 6800 non-coding elements associated with over 120 early developmental genes and conserved across vertebrates. The database regularly incorporates results of ongoing in vivo zebrafish enhancer assays of the CNEs carried out in-house, which currently number ~100. Included and highlighted within this set are elements derived from duplication events both at the origin of vertebrates and more recently in the teleost lineage, thus providing valuable data for studying the divergence of regulatory roles between paralogs. CONDOR therefore provides a number of tools and facilities to allow scientists to progress in their own studies on the function and evolution of developmental cis-regulation. Conclusion By providing access to data with an approachable graphics interface, the CONDOR database presents a rich resource for further studies into the regulation and evolution of genes involved in early development.

Selective constraints in experimentally defined primate regulatory regions.

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Daniel J Gaffney

2008-08-01

Full Text Available Changes in gene regulation may be important in evolution. However, the evolutionary properties of regulatory mutations are currently poorly understood. This is partly the result of an incomplete annotation of functional regulatory DNA in many species. For example, transcription factor binding sites (TFBSs, a major component of eukaryotic regulatory architecture, are typically short, degenerate, and therefore difficult to differentiate from randomly occurring, nonfunctional sequences. Furthermore, although sites such as TFBSs can be computationally predicted using evolutionary conservation as a criterion, estimates of the true level of selective constraint (defined as the fraction of strongly deleterious mutations occurring at a locus in regulatory regions will, by definition, be upwardly biased in datasets that are a priori evolutionarily conserved. Here we investigate the fitness effects of regulatory mutations using two complementary datasets of human TFBSs that are likely to be relatively free of ascertainment bias with respect to evolutionary conservation but, importantly, are supported by experimental data. The first is a collection of almost >2,100 human TFBSs drawn from the literature in the TRANSFAC database, and the second is derived from several recent high-throughput chromatin immunoprecipitation coupled with genomic microarray (ChIP-chip analyses. We also define a set of putative cis-regulatory modules (pCRMs by spatially clustering multiple TFBSs that regulate the same gene. We find that a relatively high proportion ( approximately 37% of mutations at TFBSs are strongly deleterious, similar to that at a 2-fold degenerate protein-coding site. However, constraint is significantly reduced in human and chimpanzee pCRMS and ChIP-chip sequences, relative to macaques. We estimate that the fraction of regulatory mutations that have been driven to fixation by positive selection in humans is not significantly different from zero. We also find

Two negative cis-regulatory regions involved in fruit-specific promoter activity from watermelon (Citrullus vulgaris S.).

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Yin, Tao; Wu, Hanying; Zhang, Shanglong; Lu, Hongyu; Zhang, Lingxiao; Xu, Yong; Chen, Daming; Liu, Jingmei

2009-01-01

A 1.8 kb 5'-flanking region of the large subunit of ADP-glucose pyrophosphorylase, isolated from watermelon (Citrullus vulgaris S.), has fruit-specific promoter activity in transgenic tomato plants. Two negative regulatory regions, from -986 to -959 and from -472 to -424, were identified in this promoter region by fine deletion analyses. Removal of both regions led to constitutive expression in epidermal cells. Gain-of-function experiments showed that these two regions were sufficient to inhibit RFP (red fluorescent protein) expression in transformed epidermal cells when fused to the cauliflower mosaic virus (CaMV) 35S minimal promoter. Gel mobility shift experiments demonstrated the presence of leaf nuclear factors that interact with these two elements. A TCCAAAA motif was identified in these two regions, as well as one in the reverse orientation, which was confirmed to be a novel specific cis-element. A quantitative beta-glucuronidase (GUS) activity assay of stable transgenic tomato plants showed that the activities of chimeric promoters harbouring only one of the two cis-elements, or both, were approximately 10-fold higher in fruits than in leaves. These data confirm that the TCCAAAA motif functions as a fruit-specific element by inhibiting gene expression in leaves.

A New Approach to Sequence Analysis Exemplified by Identification of cis-Elements in Abscisic Acid Inducible Promoters

DEFF Research Database (Denmark)

Busk, Peter Kamp; Hallin, Peter Fischer; Salomon, Jesper

-regulatory elements. We have developed a method for identifying short, conserved motifs in biological sequences such as proteins, DNA and RNA5. This method was used for analysis of approximately 2000 Arabidopsis thaliana promoters that have been shown by DNA array analysis to be induced by abscisic acid6....... These promoters were compared to 28000 promoters that are not induced by abscisic acid. The analysis identified previously described ABA-inducible promoter elements such as ABRE, CE3 and CRT1 but also new cis-elements were found. Furthermore, the list of DNA elements could be used to predict ABA...

MicroRNA signature of cis-platin resistant vs. cis-platin sensitive ovarian cancer cell lines

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Kumar Smriti

2011-09-01

Full Text Available Abstract Background Ovarian cancer is the leading cause of death from gynecologic cancer in women worldwide. According to the National Cancer Institute, ovarian cancer has the highest mortality rate among all the reproductive cancers in women. Advanced stage diagnosis and chemo/radio-resistance is a major obstacle in treating advanced ovarian cancer. The most commonly employed chemotherapeutic drug for ovarian cancer treatment is cis-platin. As with most chemotherapeutic drugs, many patients eventually become resistant to cis-platin and therefore, diminishing its effect. The efficacy of current treatments may be improved by increasing the sensitivity of cancer cells to chemo/radiation therapies. Methods The present study is focused on identifying the differential expression of regulatory microRNAs (miRNAs between cis-platin sensitive (A2780, and cis-platin resistant (A2780/CP70 cell lines. Cell proliferation assays were conducted to test the sensitivity of the two cell lines to cis-platin. Differential expression patterns of miRNA between cis-platin sensitive and cis-platin resistant cell lines were analyzed using novel LNA technology. Results Our results revealed changes in expression of 11 miRNAs out of 1,500 miRNAs analyzed. Out of the 11 miRNAs identified, 5 were up-regulated in the A2780/CP70 cell line and 6 were down regulated as compared to cis-platin sensitive A2780 cells. Our microRNA data was further validated by quantitative real-time PCR for these selected miRNAs. Ingenuity Pathway Analysis (IPA and Kyoto Encyclopedia of Genes and Genomes (KEGG analysis was performed for the selected miRNAs and their putative targets to identify the potential pathways and networks involved in cis-platin resistance. Conclusions Our data clearly showed the differential expression of 11 miRNAs in cis-platin resistant cells, which could potentially target many important pathways including MAPK, TGF-β signaling, actin cytoskeleton, ubiquitin mediated

Speeding cis-trans regulation discovery by phylogenomic analyses coupled with screenings of an arrayed library of Arabidopsis transcription factors.

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Gabriel Castrillo

Full Text Available Transcriptional regulation is an important mechanism underlying gene expression and has played a crucial role in evolution. The number, position and interactions between cis-elements and transcription factors (TFs determine the expression pattern of a gene. To identify functionally relevant cis-elements in gene promoters, a phylogenetic shadowing approach with a lipase gene (LIP1 was used. As a proof of concept, in silico analyses of several Brassicaceae LIP1 promoters identified a highly conserved sequence (LIP1 element that is sufficient to drive strong expression of a reporter gene in planta. A collection of ca. 1,200 Arabidopsis thaliana TF open reading frames (ORFs was arrayed in a 96-well format (RR library and a convenient mating based yeast one hybrid (Y1H screening procedure was established. We constructed an episomal plasmid (pTUY1H to clone the LIP1 element and used it as bait for Y1H screenings. A novel interaction with an HD-ZIP (AtML1 TF was identified and abolished by a 2 bp mutation in the LIP1 element. A role of this interaction in transcriptional regulation was confirmed in planta. In addition, we validated our strategy by reproducing the previously reported interaction between a MYB-CC (PHR1 TF, a central regulator of phosphate starvation responses, with a conserved promoter fragment (IPS1 element containing its cognate binding sequence. Finally, we established that the LIP1 and IPS1 elements were differentially bound by HD-ZIP and MYB-CC family members in agreement with their genetic redundancy in planta. In conclusion, combining in silico analyses of orthologous gene promoters with Y1H screening of the RR library represents a powerful approach to decipher cis- and trans-regulatory codes.

Properties of non-coding DNA and identification of putative cis-regulatory elements in Theileria parva

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Guo Xiang

2008-12-01

regulatory motifs in other species. These results suggest that these two motifs are likely to represent transcription factor binding sites in Theileria. Conclusion Theileria genomes are highly compact, with selection seemingly favoring short introns and intergenic regions. Three over-represented sequence motifs were independently identified in intergenic regions of both Theileria species, and the evidence suggests that at least two of them play a role in transcriptional control in T. parva. These are prime candidates for experimental validation of transcription factor binding sites in this single-celled eukaryotic parasite. Sequences similar to two of these Theileria motifs are conserved in Plasmodium hinting at the possibility of common regulatory machinery across the phylum Apicomplexa.

Fanconi anemia core complex gene promoters harbor conserved transcription regulatory elements.

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Meier, Daniel; Schindler, Detlev

2011-01-01

The Fanconi anemia (FA) gene family is a recent addition to the complex network of proteins that respond to and repair certain types of DNA damage in the human genome. Since little is known about the regulation of this novel group of genes at the DNA level, we characterized the promoters of the eight genes (FANCA, B, C, E, F, G, L and M) that compose the FA core complex. The promoters of these genes show the characteristic attributes of housekeeping genes, such as a high GC content and CpG islands, a lack of TATA boxes and a low conservation. The promoters functioned in a monodirectional way and were, in their most active regions, comparable in strength to the SV40 promoter in our reporter plasmids. They were also marked by a distinctive transcriptional start site (TSS). In the 5' region of each promoter, we identified a region that was able to negatively regulate the promoter activity in HeLa and HEK 293 cells in isolation. The central and 3' regions of the promoter sequences harbor binding sites for several common and rare transcription factors, including STAT, SMAD, E2F, AP1 and YY1, which indicates that there may be cross-connections to several established regulatory pathways. Electrophoretic mobility shift assays and siRNA experiments confirmed the shared regulatory responses between the prominent members of the TGF-β and JAK/STAT pathways and members of the FA core complex. Although the promoters are not well conserved, they share region and sequence specific regulatory motifs and transcription factor binding sites (TBFs), and we identified a bi-partite nature to these promoters. These results support a hypothesis based on the co-evolution of the FA core complex genes that was expanded to include their promoters.

Fanconi anemia core complex gene promoters harbor conserved transcription regulatory elements.

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Daniel Meier

Full Text Available The Fanconi anemia (FA gene family is a recent addition to the complex network of proteins that respond to and repair certain types of DNA damage in the human genome. Since little is known about the regulation of this novel group of genes at the DNA level, we characterized the promoters of the eight genes (FANCA, B, C, E, F, G, L and M that compose the FA core complex. The promoters of these genes show the characteristic attributes of housekeeping genes, such as a high GC content and CpG islands, a lack of TATA boxes and a low conservation. The promoters functioned in a monodirectional way and were, in their most active regions, comparable in strength to the SV40 promoter in our reporter plasmids. They were also marked by a distinctive transcriptional start site (TSS. In the 5' region of each promoter, we identified a region that was able to negatively regulate the promoter activity in HeLa and HEK 293 cells in isolation. The central and 3' regions of the promoter sequences harbor binding sites for several common and rare transcription factors, including STAT, SMAD, E2F, AP1 and YY1, which indicates that there may be cross-connections to several established regulatory pathways. Electrophoretic mobility shift assays and siRNA experiments confirmed the shared regulatory responses between the prominent members of the TGF-β and JAK/STAT pathways and members of the FA core complex. Although the promoters are not well conserved, they share region and sequence specific regulatory motifs and transcription factor binding sites (TBFs, and we identified a bi-partite nature to these promoters. These results support a hypothesis based on the co-evolution of the FA core complex genes that was expanded to include their promoters.

Identification of putative regulatory upstream ORFs in the yeast genome using heuristics and evolutionary conservation

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Bilsland Elizabeth

2007-08-01

Full Text Available Abstract Background The translational efficiency of an mRNA can be modulated by upstream open reading frames (uORFs present in certain genes. A uORF can attenuate translation of the main ORF by interfering with translational reinitiation at the main start codon. uORFs also occur by chance in the genome, in which case they do not have a regulatory role. Since the sequence determinants for functional uORFs are not understood, it is difficult to discriminate functional from spurious uORFs by sequence analysis. Results We have used comparative genomics to identify novel uORFs in yeast with a high likelihood of having a translational regulatory role. We examined uORFs, previously shown to play a role in regulation of translation in Saccharomyces cerevisiae, for evolutionary conservation within seven Saccharomyces species. Inspection of the set of conserved uORFs yielded the following three characteristics useful for discrimination of functional from spurious uORFs: a length between 4 and 6 codons, a distance from the start of the main ORF between 50 and 150 nucleotides, and finally a lack of overlap with, and clear separation from, neighbouring uORFs. These derived rules are inherently associated with uORFs with properties similar to the GCN4 locus, and may not detect most uORFs of other types. uORFs with high scores based on these rules showed a much higher evolutionary conservation than randomly selected uORFs. In a genome-wide scan in S. cerevisiae, we found 34 conserved uORFs from 32 genes that we predict to be functional; subsequent analysis showed the majority of these to be located within transcripts. A total of 252 genes were found containing conserved uORFs with properties indicative of a functional role; all but 7 are novel. Functional content analysis of this set identified an overrepresentation of genes involved in transcriptional control and development. Conclusion Evolutionary conservation of uORFs in yeasts can be traced up to 100

Diverse activities of viral cis-acting RNA regulatory elements revealed using multicolor, long-term, single-cell imaging.

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Pocock, Ginger M; Zimdars, Laraine L; Yuan, Ming; Eliceiri, Kevin W; Ahlquist, Paul; Sherer, Nathan M

2017-02-01

Cis-acting RNA structural elements govern crucial aspects of viral gene expression. How these structures and other posttranscriptional signals affect RNA trafficking and translation in the context of single cells is poorly understood. Herein we describe a multicolor, long-term (>24 h) imaging strategy for measuring integrated aspects of viral RNA regulatory control in individual cells. We apply this strategy to demonstrate differential mRNA trafficking behaviors governed by RNA elements derived from three retroviruses (HIV-1, murine leukemia virus, and Mason-Pfizer monkey virus), two hepadnaviruses (hepatitis B virus and woodchuck hepatitis virus), and an intron-retaining transcript encoded by the cellular NXF1 gene. Striking behaviors include "burst" RNA nuclear export dynamics regulated by HIV-1's Rev response element and the viral Rev protein; transient aggregations of RNAs into discrete foci at or near the nuclear membrane triggered by multiple elements; and a novel, pulsiform RNA export activity regulated by the hepadnaviral posttranscriptional regulatory element. We incorporate single-cell tracking and a data-mining algorithm into our approach to obtain RNA element-specific, high-resolution gene expression signatures. Together these imaging assays constitute a tractable, systems-based platform for studying otherwise difficult to access spatiotemporal features of viral and cellular gene regulation. © 2017 Pocock et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

Identification of direct regulatory targets of the transcription factor Sox10 based on function and conservation

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Lee Sanghyuk

2008-09-01

Full Text Available Abstract Background Sox10, a member of the Sry-related HMG-Box gene family, is a critical transcription factor for several important cell lineages, most notably the neural crest stem cells and the derivative peripheral glial cells and melanocytes. Thus far, only a handful of direct target genes are known for this transcription factor limiting our understanding of the biological network it governs. Results We describe identification of multiple direct regulatory target genes of Sox10 through a procedure based on function and conservation. By combining RNA interference technique and DNA microarray technology, we have identified a set of genes that show significant down-regulation upon introduction of Sox10 specific siRNA into Schwannoma cells. Subsequent comparative genomics analyses led to potential binding sites for Sox10 protein conserved across several mammalian species within the genomic region proximal to these genes. Multiple sites belonging to 4 different genes (proteolipid protein, Sox10, extracellular superoxide dismutase, and pleiotrophin were shown to directly interact with Sox10 by chromatin immunoprecipitation assay. We further confirmed the direct regulation through the identified cis-element for one of the genes, extracellular superoxide dismutase, using electrophoretic mobility shift assay and reporter assay. Conclusion In sum, the process of combining differential expression profiling and comparative genomics successfully led to further defining the role of Sox10, a critical transcription factor for the development of peripheral glia. Our strategy utilizing relatively accessible techniques and tools should be applicable to studying the function of other transcription factors.

Investment in the CEE/CIS region

International Nuclear Information System (INIS)

Lemierre, J.

2002-01-01

The energy investments in the Central and Eastern European region and the Commonwealth of Independent States (CIS) region are discussed in this Keynote Address. The message is addressed to regulators and governments. The restructuring of old industries to save energy is highlighted. The regulatory system must undergo a substantial reform. Another message is placed for investors in the energy field. (R.P.)

In silico discovery of transcription regulatory elements in Plasmodium falciparum

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Le Roch Karine G

2008-02-01

Full Text Available Abstract Background With the sequence of the Plasmodium falciparum genome and several global mRNA and protein life cycle expression profiling projects now completed, elucidating the underlying networks of transcriptional control important for the progression of the parasite life cycle is highly pertinent to the development of new anti-malarials. To date, relatively little is known regarding the specific mechanisms the parasite employs to regulate gene expression at the mRNA level, with studies of the P. falciparum genome sequence having revealed few cis-regulatory elements and associated transcription factors. Although it is possible the parasite may evoke mechanisms of transcriptional control drastically different from those used by other eukaryotic organisms, the extreme AT-rich nature of P. falciparum intergenic regions (~90% AT presents significant challenges to in silico cis-regulatory element discovery. Results We have developed an algorithm called Gene Enrichment Motif Searching (GEMS that uses a hypergeometric-based scoring function and a position-weight matrix optimization routine to identify with high-confidence regulatory elements in the nucleotide-biased and repeat sequence-rich P. falciparum genome. When applied to promoter regions of genes contained within 21 co-expression gene clusters generated from P. falciparum life cycle microarray data using the semi-supervised clustering algorithm Ontology-based Pattern Identification, GEMS identified 34 putative cis-regulatory elements associated with a variety of parasite processes including sexual development, cell invasion, antigenic variation and protein biosynthesis. Among these candidates were novel motifs, as well as many of the elements for which biological experimental evidence already exists in the Plasmodium literature. To provide evidence for the biological relevance of a cell invasion-related element predicted by GEMS, reporter gene and electrophoretic mobility shift assays

Nutritional control of gene expression in Drosophila larvae via TOR, Myc and a novel cis-regulatory element

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Grewal Savraj S

2010-01-01

Full Text Available Abstract Background Nutrient availability is a key determinant of eukaryotic cell growth. In unicellular organisms many signaling and transcriptional networks link nutrient availability to the expression of metabolic genes required for growth. However, less is known about the corresponding mechanisms that operate in metazoans. We used gene expression profiling to explore this issue in developing Drosophila larvae. Results We found that starvation for dietary amino acids (AA's leads to dynamic changes in transcript levels of many metabolic genes. The conserved insulin/PI3K and TOR signaling pathways mediate nutrition-dependent growth in Drosophila and other animals. We found that many AA starvation-responsive transcripts were also altered in TOR mutants. In contrast, although PI3K overexpression induced robust changes in the expression of many metabolic genes, these changes showed limited overlap with the AA starvation expression profile. We did however identify a strong overlap between genes regulated by the transcription factor, Myc, and AA starvation-responsive genes, particularly those involved in ribosome biogenesis, protein synthesis and mitochondrial function. The consensus Myc DNA binding site is enriched in promoters of these AA starvation genes, and we found that Myc overexpression could bypass dietary AA to induce expression of these genes. We also identified another sequence motif (Motif 1 enriched in the promoters of AA starvation-responsive genes. We showed that Motif 1 was both necessary and sufficient to mediate transcriptional responses to dietary AA in larvae. Conclusions Our data suggest that many of the transcriptional effects of amino acids are mediated via signaling through the TOR pathway in Drosophila larvae. We also find that these transcriptional effects are mediated through at least two mechanisms: via the transcription factor Myc, and via the Motif 1 cis-regulatory element. These studies begin to elucidate a nutrient

Asymmetrical distribution of non-conserved regulatory sequences at PHOX2B is reflected at the ENCODE loci and illuminates a possible genome-wide trend

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McCallion Andrew S

2009-01-01

Full Text Available Abstract Background Transcriptional regulatory elements are central to development and interspecific phenotypic variation. Current regulatory element prediction tools rely heavily upon conservation for prediction of putative elements. Recent in vitro observations from the ENCODE project combined with in vivo analyses at the zebrafish phox2b locus suggests that a significant fraction of regulatory elements may fall below commonly applied metrics of conservation. We propose to explore these observations in vivo at the human PHOX2B locus, and also evaluate the potential evidence for genome-wide applicability of these observations through a novel analysis of extant data. Results Transposon-based transgenic analysis utilizing a tiling path proximal to human PHOX2B in zebrafish recapitulates the observations at the zebrafish phox2b locus of both conserved and non-conserved regulatory elements. Analysis of human sequences conserved with previously identified zebrafish phox2b regulatory elements demonstrates that the orthologous sequences exhibit overlapping regulatory control. Additionally, analysis of non-conserved sequences scattered over 135 kb 5' to PHOX2B, provides evidence of non-conserved regulatory elements positively biased with close proximity to the gene. Furthermore, we provide a novel analysis of data from the ENCODE project, finding a non-uniform distribution of regulatory elements consistent with our in vivo observations at PHOX2B. These observations remain largely unchanged when one accounts for the sequence repeat content of the assayed intervals, when the intervals are sub-classified by biological role (developmental versus non-developmental, or by gene density (gene desert versus non-gene desert. Conclusion While regulatory elements frequently display evidence of evolutionary conservation, a fraction appears to be undetected by current metrics of conservation. In vivo observations at the PHOX2B locus, supported by our analyses of in

Cis-regulatory RNA elements that regulate specialized ribosome activity.

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Xue, Shifeng; Barna, Maria

2015-01-01

Recent evidence has shown that the ribosome itself can play a highly regulatory role in the specialized translation of specific subpools of mRNAs, in particular at the level of ribosomal proteins (RP). However, the mechanism(s) by which this selection takes place has remained poorly understood. In our recent study, we discovered a combination of unique RNA elements in the 5'UTRs of mRNAs that allows for such control by the ribosome. These mRNAs contain a Translation Inhibitory Element (TIE) that inhibits general cap-dependent translation, and an Internal Ribosome Entry Site (IRES) that relies on a specific RP for activation. The unique combination of an inhibitor of general translation and an activator of specialized translation is key to ribosome-mediated control of gene expression. Here we discuss how these RNA regulatory elements provide a new level of control to protein expression and their implications for gene expression, organismal development and evolution.

Preaxial polydactyly/triphalangeal thumb is associated with changed transcription factor-binding affinity in a family with a novel point mutation in the long-range cis-regulatory element ZRS

DEFF Research Database (Denmark)

Farooq, Muhammad; Troelsen, Jesper T; Boyd, Mette

2010-01-01

A cis-regulatory sequence also known as zone of polarizing activity (ZPA) regulatory sequence (ZRS) located in intron 5 of LMBR1 is essential for expression of sonic hedgehog (SHH) in the developing posterior limb bud mesenchyme. Even though many point mutations causing preaxial duplication defects...... demonstrated a marked difference between wild-type and the mutant probe, which uniquely bound one or several transcription factors extracted from Caco-2 cells. This finding supports a model in which ectopic anterior SHH expression in the developing limb results from abnormal binding of one or more...

Identifying Cis-Regulatory Changes Involved in the Evolution of Aerobic Fermentation in Yeasts

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Lin, Zhenguo; Wang, Tzi-Yuan; Tsai, Bing-Shi; Wu, Fang-Ting; Yu, Fu-Jung; Tseng, Yu-Jung; Sung, Huang-Mo; Li, Wen-Hsiung

2013-01-01

Gene regulation change has long been recognized as an important mechanism for phenotypic evolution. We used the evolution of yeast aerobic fermentation as a model to explore how gene regulation has evolved and how this process has contributed to phenotypic evolution and adaptation. Most eukaryotes fully oxidize glucose to CO2 and H2O in mitochondria to maximize energy yield, whereas some yeasts, such as Saccharomyces cerevisiae and its relatives, predominantly ferment glucose into ethanol even in the presence of oxygen, a phenomenon known as aerobic fermentation. We examined the genome-wide gene expression levels among 12 different yeasts and found that a group of genes involved in the mitochondrial respiration process showed the largest reduction in gene expression level during the evolution of aerobic fermentation. Our analysis revealed that the downregulation of these genes was significantly associated with massive loss of binding motifs of Cbf1p in the fermentative yeasts. Our experimental assays confirmed the binding of Cbf1p to the predicted motif and the activator role of Cbf1p. In summary, our study laid a foundation to unravel the long-time mystery about the genetic basis of evolution of aerobic fermentation, providing new insights into understanding the role of cis-regulatory changes in phenotypic evolution. PMID:23650209

Integrative modeling of eQTLs and cis-regulatory elements suggests mechanisms underlying cell type specificity of eQTLs.

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Christopher D Brown

Full Text Available Genetic variants in cis-regulatory elements or trans-acting regulators frequently influence the quantity and spatiotemporal distribution of gene transcription. Recent interest in expression quantitative trait locus (eQTL mapping has paralleled the adoption of genome-wide association studies (GWAS for the analysis of complex traits and disease in humans. Under the hypothesis that many GWAS associations tag non-coding SNPs with small effects, and that these SNPs exert phenotypic control by modifying gene expression, it has become common to interpret GWAS associations using eQTL data. To fully exploit the mechanistic interpretability of eQTL-GWAS comparisons, an improved understanding of the genetic architecture and causal mechanisms of cell type specificity of eQTLs is required. We address this need by performing an eQTL analysis in three parts: first we identified eQTLs from eleven studies on seven cell types; then we integrated eQTL data with cis-regulatory element (CRE data from the ENCODE project; finally we built a set of classifiers to predict the cell type specificity of eQTLs. The cell type specificity of eQTLs is associated with eQTL SNP overlap with hundreds of cell type specific CRE classes, including enhancer, promoter, and repressive chromatin marks, regions of open chromatin, and many classes of DNA binding proteins. These associations provide insight into the molecular mechanisms generating the cell type specificity of eQTLs and the mode of regulation of corresponding eQTLs. Using a random forest classifier with cell specific CRE-SNP overlap as features, we demonstrate the feasibility of predicting the cell type specificity of eQTLs. We then demonstrate that CREs from a trait-associated cell type can be used to annotate GWAS associations in the absence of eQTL data for that cell type. We anticipate that such integrative, predictive modeling of cell specificity will improve our ability to understand the mechanistic basis of human

Evolution of Enzymatic Activities in the Enolase Superfamily: Stereochemically Distinct Mechanisms in Two Families of cis,cis-Muconate Lactonizing Enzymes

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Sakai, A.; Fedorov, A; Fedorov, E; Schnoes, A; Glasner, M; Burley, S; Babbitt, P; Almo, S; Gerlt, J

2009-01-01

The mechanistically diverse enolase superfamily is a paradigm for elucidating Nature's strategies for divergent evolution of enzyme function. Each of the different reactions catalyzed by members of the superfamily is initiated by abstraction of the a-proton of a carboxylate substrate that is coordinated to an essential Mg2+. The muconate lactonizing enzyme (MLE) from Pseudomonas putida, a member of a family that catalyzes the syn-cycloisomerization of cis,cis-muconate to (4S)-muconolactone in the e-ketoadipate pathway, has provided critical insights into the structural bases for evolution of function within the superfamily. A second, divergent family of homologous MLEs that catalyzes anti-cycloisomerization has been identified. Structures of members of both families liganded with the common (4S)-muconolactone product (syn, Pseudomonas fluorescens, gi 70731221; anti, Mycobacterium smegmatis, gi 118470554) document that the conserved Lys at the end of the second e-strand in the (e/a)7e-barrel domain serves as the acid catalyst in both reactions. The different stereochemical courses (syn and anti) result from different structural strategies for determining substrate specificity: although the distal carboxylate group of the cis,cis-muconate substrate attacks the same face of the proximal double bond, opposite faces of the resulting enolate anion intermediate are presented to the conserved Lys acid catalyst. The discovery of two families of homologous, but stereochemically distinct, MLEs likely provides an example of 'pseudoconvergent' evolution of the same function from different homologous progenitors within the enolase superfamily, in which different spatial arrangements of active site functional groups and substrate specificity determinants support catalysis of the same reaction.

Evolution of Enzymatic Activities in the Enolase Superfamily: Stereochemically Distinct Mechanisms in Two Families of cis,cis-Muconate Lactonizing Enzymes†

Science.gov (United States)

Sakai, Ayano; Fedorov, Alexander A.; Fedorov, Elena V.; Schnoes, Alexandra M.; Glasner, Margaret E.; Brown, Shoshana; Rutter, Marc E.; Bain, Kevin; Chang, Shawn; Gheyi, Tarun; Sauder, J. Michael; Burley, Stephen K.; Babbitt, Patricia C.; Almo, Steven C.; Gerlt, John A.

2009-01-01

The mechanistically diverse enolase superfamily is a paradigm for elucidating Nature’s strategies for divergent evolution of enzyme function. Each of the different reactions catalyzed by members of the superfamily is initiated by abstraction of the α-proton of a carboxylate substrate that is coordinated to an essential Mg2+. The muconate lactonizing enzyme (MLE) from Pseudomonas putida, a member of a family that catalyzes the syn-cycloisomerization of cis,cis-muconate to (4S)-muconolactone in the β-ketoadipate pathway, has provided critical insights into the structural bases for evolution of function within the superfamily. A second, divergent family of homologues MLEs that catalyzes anti-cycloisomerization has been identified. Structures of members of both families liganded with the common (4S)-muconolactone product (syn, Pseudomonas fluorescens, GI:70731221; anti, Mycobacterium smegmatis, GI:118470554) document that the conserved Lys at the end of the second β-strand in the (β/α)7β-barrel domain serves as the acid catalyst in both reactions. The different stereochemical courses (syn and anti) result from different structural strategies for determining substrate specificity: although the distal carboxylate group of the cis,cis-muconate substrate attacks the same face of the proximal double bond, opposite faces of the resulting enolate anion intermediate are presented to the conserved Lys acid catalyst. The discovery of two families of homologous, but stereochemically distinct, MLEs likely provides an example of “pseudoconvergent” evolution of the same function from different homologous progenitors within the enolase superfamily, in which different spatial arrangements of active site functional groups and substrate specificity determinants support catalysis of the same reaction. PMID:19220063

Conserving Coherence and Storing Energy during Internal Conversion: Photoinduced Dynamics of cis- and trans-Azobenzene Radical Cations

KAUST Repository

Munkerup, Kristin

2017-10-24

Light harvesting via energy storage in azobenzene has been a key topic for decades, and the process of energy distribution over the molecular degrees of freedom following photoexcitation remains to be understood. Dynamics of a photoexcited system can exhibit high degrees of non-ergodicity when it is driven by just a few degrees of freedom. Typically, an internal conversion leads to the loss of such localization of dynamics, as the intramolecular energy becomes statistically redistributed over all molecular degrees of freedom. Here, we present a unique case where the excitation energy remains localized even subsequent to internal conversion. Strong-field ionization is used to prepare cis- and trans-azobenzene radical cations on the D1 surface with little excess energy, at the equilibrium neutral geometry. These D1 ions are preferably formed because in this case D1 and D0 switch place in the presence of the strong laser field. The post-ionization dynamics is dictated by the potential energy landscape. The D1 surface is steep downhill along the cis/trans isomerization coordinate and towards a common minimum shared by the two isomers in the region of D1/D0 conical intersection. Coherent cis/trans torsional motion along this coordinate is manifested in the ion transients by a cosine modulation. In this scenario, D0 becomes populated with molecules that are energized mainly along the cis-trans isomerization coordinate, with the kinetic energy above the cis-trans inter-conversion barrier. These activated azobenzene molecules easily cycle back and forth along the D0 surface, and give rise to several periods of modulated signal before coherence is lost. This persistent localization of the internal energy during internal conversion is provided by the steep downhill potential energy surface, small initial internal energy content, and a strong hole-lone pair interaction that drives the molecule along the cis-trans isomerization coordinate to facilitate the transition between

WeederH: an algorithm for finding conserved regulatory motifs and regions in homologous sequences

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Pesole Graziano

2007-02-01

Full Text Available Abstract Background This work addresses the problem of detecting conserved transcription factor binding sites and in general regulatory regions through the analysis of sequences from homologous genes, an approach that is becoming more and more widely used given the ever increasing amount of genomic data available. Results We present an algorithm that identifies conserved transcription factor binding sites in a given sequence by comparing it to one or more homologs, adapting a framework we previously introduced for the discovery of sites in sequences from co-regulated genes. Differently from the most commonly used methods, the approach we present does not need or compute an alignment of the sequences investigated, nor resorts to descriptors of the binding specificity of known transcription factors. The main novel idea we introduce is a relative measure of conservation, assuming that true functional elements should present a higher level of conservation with respect to the rest of the sequence surrounding them. We present tests where we applied the algorithm to the identification of conserved annotated sites in homologous promoters, as well as in distal regions like enhancers. Conclusion Results of the tests show how the algorithm can provide fast and reliable predictions of conserved transcription factor binding sites regulating the transcription of a gene, with better performances than other available methods for the same task. We also show examples on how the algorithm can be successfully employed when promoter annotations of the genes investigated are missing, or when regulatory sites and regions are located far away from the genes.

Predicting tissue specific cis-regulatory modules in the human genome using pairs of co-occurring motifs

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Girgis Hani Z

2012-02-01

Full Text Available Abstract Background Researchers seeking to unlock the genetic basis of human physiology and diseases have been studying gene transcription regulation. The temporal and spatial patterns of gene expression are controlled by mainly non-coding elements known as cis-regulatory modules (CRMs and epigenetic factors. CRMs modulating related genes share the regulatory signature which consists of transcription factor (TF binding sites (TFBSs. Identifying such CRMs is a challenging problem due to the prohibitive number of sequence sets that need to be analyzed. Results We formulated the challenge as a supervised classification problem even though experimentally validated CRMs were not required. Our efforts resulted in a software system named CrmMiner. The system mines for CRMs in the vicinity of related genes. CrmMiner requires two sets of sequences: a mixed set and a control set. Sequences in the vicinity of the related genes comprise the mixed set, whereas the control set includes random genomic sequences. CrmMiner assumes that a large percentage of the mixed set is made of background sequences that do not include CRMs. The system identifies pairs of closely located motifs representing vertebrate TFBSs that are enriched in the training mixed set consisting of 50% of the gene loci. In addition, CrmMiner selects a group of the enriched pairs to represent the tissue-specific regulatory signature. The mixed and the control sets are searched for candidate sequences that include any of the selected pairs. Next, an optimal Bayesian classifier is used to distinguish candidates found in the mixed set from their control counterparts. Our study proposes 62 tissue-specific regulatory signatures and putative CRMs for different human tissues and cell types. These signatures consist of assortments of ubiquitously expressed TFs and tissue-specific TFs. Under controlled settings, CrmMiner identified known CRMs in noisy sets up to 1:25 signal-to-noise ratio. CrmMiner was

A cis-regulatory sequence driving metabolic insecticide resistance in mosquitoes: functional characterisation and signatures of selection.

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Wilding, Craig S; Smith, Ian; Lynd, Amy; Yawson, Alexander Egyir; Weetman, David; Paine, Mark J I; Donnelly, Martin J

2012-09-01

Although cytochrome P450 (CYP450) enzymes are frequently up-regulated in mosquitoes resistant to insecticides, no regulatory motifs driving these expression differences with relevance to wild populations have been identified. Transposable elements (TEs) are often enriched upstream of those CYP450s involved in insecticide resistance, leading to the assumption that they contribute regulatory motifs that directly underlie the resistance phenotype. A partial CuRE1 (Culex Repetitive Element 1) transposable element is found directly upstream of CYP9M10, a cytochrome P450 implicated previously in larval resistance to permethrin in the ISOP450 strain of Culex quinquefasciatus, but is absent from the equivalent genomic region of a susceptible strain. Via expression of CYP9M10 in Escherichia coli we have now demonstrated time- and NADPH-dependant permethrin metabolism, prerequisites for confirmation of a role in metabolic resistance, and through qPCR shown that CYP9M10 is >20-fold over-expressed in ISOP450 compared to a susceptible strain. In a fluorescent reporter assay the region upstream of CYP9M10 from ISOP450 drove 10× expression compared to the equivalent region (lacking CuRE1) from the susceptible strain. Close correspondence with the gene expression fold-change implicates the upstream region including CuRE1 as a cis-regulatory element involved in resistance. Only a single CuRE1 bearing allele, identical to the CuRE1 bearing allele in the resistant strain, is found throughout Sub-Saharan Africa, in contrast to the diversity encountered in non-CuRE1 alleles. This suggests a single origin and subsequent spread due to selective advantage. CuRE1 is detectable using a simple diagnostic. When applied to C. quinquefasciatus larvae from Ghana we have demonstrated a significant association with permethrin resistance in multiple field sites (mean Odds Ratio = 3.86) suggesting this marker has relevance to natural populations of vector mosquitoes. However, when CuRE1 was excised

Computational exploration of cis-regulatory modules in rhythmic expression data using the "Exploration of Distinctive CREs and CRMs" (EDCC) and "CRM Network Generator" (CNG) programs.

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Bekiaris, Pavlos Stephanos; Tekath, Tobias; Staiger, Dorothee; Danisman, Selahattin

2018-01-01

Understanding the effect of cis-regulatory elements (CRE) and clusters of CREs, which are called cis-regulatory modules (CRM), in eukaryotic gene expression is a challenge of computational biology. We developed two programs that allow simple, fast and reliable analysis of candidate CREs and CRMs that may affect specific gene expression and that determine positional features between individual CREs within a CRM. The first program, "Exploration of Distinctive CREs and CRMs" (EDCC), correlates candidate CREs and CRMs with specific gene expression patterns. For pairs of CREs, EDCC also determines positional preferences of the single CREs in relation to each other and to the transcriptional start site. The second program, "CRM Network Generator" (CNG), prioritizes these positional preferences using a neural network and thus allows unbiased rating of the positional preferences that were determined by EDCC. We tested these programs with data from a microarray study of circadian gene expression in Arabidopsis thaliana. Analyzing more than 1.5 million pairwise CRE combinations, we found 22 candidate combinations, of which several contained known clock promoter elements together with elements that had not been identified as relevant to circadian gene expression before. CNG analysis further identified positional preferences of these CRE pairs, hinting at positional information that may be relevant for circadian gene expression. Future wet lab experiments will have to determine which of these combinations confer daytime specific circadian gene expression.

High-throughput sequencing, characterization and detection of new and conserved cucumber miRNAs.

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Germán Martínez

Full Text Available Micro RNAS (miRNAs are a class of endogenous small non coding RNAs involved in the post-transcriptional regulation of gene expression. In plants, a great number of conserved and specific miRNAs, mainly arising from model species, have been identified to date. However less is known about the diversity of these regulatory RNAs in vegetal species with agricultural and/or horticultural importance. Here we report a combined approach of bioinformatics prediction, high-throughput sequencing data and molecular methods to analyze miRNAs populations in cucumber (Cucumis sativus plants. A set of 19 conserved and 6 known but non-conserved miRNA families were found in our cucumber small RNA dataset. We also identified 7 (3 with their miRNA* strand not previously described miRNAs, candidates to be cucumber-specific. To validate their description these new C. sativus miRNAs were detected by northern blot hybridization. Additionally, potential targets for most conserved and new miRNAs were identified in cucumber genome.In summary, in this study we have identified, by first time, conserved, known non-conserved and new miRNAs arising from an agronomically important species such as C. sativus. The detection of this complex population of regulatory small RNAs suggests that similarly to that observe in other plant species, cucumber miRNAs may possibly play an important role in diverse biological and metabolic processes.

Unveiling combinatorial regulation through the combination of ChIP information and in silico cis-regulatory module detection

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Sun, Hong; Guns, Tias; Fierro, Ana Carolina; Thorrez, Lieven; Nijssen, Siegfried; Marchal, Kathleen

2012-01-01

Computationally retrieving biologically relevant cis-regulatory modules (CRMs) is not straightforward. Because of the large number of candidates and the imperfection of the screening methods, many spurious CRMs are detected that are as high scoring as the biologically true ones. Using ChIP-information allows not only to reduce the regions in which the binding sites of the assayed transcription factor (TF) should be located, but also allows restricting the valid CRMs to those that contain the assayed TF (here referred to as applying CRM detection in a query-based mode). In this study, we show that exploiting ChIP-information in a query-based way makes in silico CRM detection a much more feasible endeavor. To be able to handle the large datasets, the query-based setting and other specificities proper to CRM detection on ChIP-Seq based data, we developed a novel powerful CRM detection method ‘CPModule’. By applying it on a well-studied ChIP-Seq data set involved in self-renewal of mouse embryonic stem cells, we demonstrate how our tool can recover combinatorial regulation of five known TFs that are key in the self-renewal of mouse embryonic stem cells. Additionally, we make a number of new predictions on combinatorial regulation of these five key TFs with other TFs documented in TRANSFAC. PMID:22422841

Distinct cis regulatory elements govern the expression of TAG1 in embryonic sensory ganglia and spinal cord.

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Yoav Hadas

Full Text Available Cell fate commitment of spinal progenitor neurons is initiated by long-range, midline-derived, morphogens that regulate an array of transcription factors that, in turn, act sequentially or in parallel to control neuronal differentiation. Included among these are transcription factors that regulate the expression of receptors for guidance cues, thereby determining axonal trajectories. The Ig/FNIII superfamily molecules TAG1/Axonin1/CNTN2 (TAG1 and Neurofascin (Nfasc are co-expressed in numerous neuronal cell types in the CNS and PNS - for example motor, DRG and interneurons - both promote neurite outgrowth and both are required for the architecture and function of nodes of Ranvier. The genes encoding TAG1 and Nfasc are adjacent in the genome, an arrangement which is evolutionarily conserved. To study the transcriptional network that governs TAG1 and Nfasc expression in spinal motor and commissural neurons, we set out to identify cis elements that regulate their expression. Two evolutionarily conserved DNA modules, one located between the Nfasc and TAG1 genes and the second directly 5' to the first exon and encompassing the first intron of TAG1, were identified that direct complementary expression to the CNS and PNS, respectively, of the embryonic hindbrain and spinal cord. Sequential deletions and point mutations of the CNS enhancer element revealed a 130bp element containing three conserved E-boxes required for motor neuron expression. In combination, these two elements appear to recapitulate a major part of the pattern of TAG1 expression in the embryonic nervous system.

Computational methods to dissect cis-regulatory transcriptional ...

Indian Academy of Sciences (India)

The formation of diverse cell types from an invariant set of genes is governed by biochemical and molecular processes that regulate gene activity. A complete understanding of the regulatory mechanisms of gene expression is the major function of genomics. Computational genomics is a rapidly emerging area for ...

Properties of Sequence Conservation in Upstream Regulatory and Protein Coding Sequences among Paralogs in Arabidopsis thaliana

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Richardson, Dale N.; Wiehe, Thomas

Whole genome duplication (WGD) has catalyzed the formation of new species, genes with novel functions, altered expression patterns, complexified signaling pathways and has provided organisms a level of genetic robustness. We studied the long-term evolution and interrelationships of 5’ upstream regulatory sequences (URSs), protein coding sequences (CDSs) and expression correlations (EC) of duplicated gene pairs in Arabidopsis. Three distinct methods revealed significant evolutionary conservation between paralogous URSs and were highly correlated with microarray-based expression correlation of the respective gene pairs. Positional information on exact matches between sequences unveiled the contribution of micro-chromosomal rearrangements on expression divergence. A three-way rank analysis of URS similarity, CDS divergence and EC uncovered specific gene functional biases. Transcription factor activity was associated with gene pairs exhibiting conserved URSs and divergent CDSs, whereas a broad array of metabolic enzymes was found to be associated with gene pairs showing diverged URSs but conserved CDSs.

XcisClique: analysis of regulatory bicliques

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Grene Ruth

2006-04-01

Full Text Available Abstract Background Modeling of cis-elements or regulatory motifs in promoter (upstream regions of genes is a challenging computational problem. In this work, set of regulatory motifs simultaneously present in the promoters of a set of genes is modeled as a biclique in a suitably defined bipartite graph. A biologically meaningful co-occurrence of multiple cis-elements in a gene promoter is assessed by the combined analysis of genomic and gene expression data. Greater statistical significance is associated with a set of genes that shares a common set of regulatory motifs, while simultaneously exhibiting highly correlated gene expression under given experimental conditions. Methods XcisClique, the system developed in this work, is a comprehensive infrastructure that associates annotated genome and gene expression data, models known cis-elements as regular expressions, identifies maximal bicliques in a bipartite gene-motif graph; and ranks bicliques based on their computed statistical significance. Significance is a function of the probability of occurrence of those motifs in a biclique (a hypergeometric distribution, and on the new sum of absolute values statistic (SAV that uses Spearman correlations of gene expression vectors. SAV is a statistic well-suited for this purpose as described in the discussion. Results XcisClique identifies new motif and gene combinations that might indicate as yet unidentified involvement of sets of genes in biological functions and processes. It currently supports Arabidopsis thaliana and can be adapted to other organisms, assuming the existence of annotated genomic sequences, suitable gene expression data, and identified regulatory motifs. A subset of Xcis Clique functionalities, including the motif visualization component MotifSee, source code, and supplementary material are available at https://bioinformatics.cs.vt.edu/xcisclique/.

Transcriptional Regulatory Network Analysis of MYB Transcription Factor Family Genes in Rice

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Shuchi eSmita

2015-12-01

Full Text Available MYB transcription factor (TF is one of the largest TF families and regulates defense responses to various stresses, hormone signaling as well as many metabolic and developmental processes in plants. Understanding these regulatory hierarchies of gene expression networks in response to developmental and environmental cues is a major challenge due to the complex interactions between the genetic elements. Correlation analyses are useful to unravel co-regulated gene pairs governing biological process as well as identification of new candidate hub genes in response to these complex processes. High throughput expression profiling data are highly useful for construction of co-expression networks. In the present study, we utilized transcriptome data for comprehensive regulatory network studies of MYB TFs by top down and guide gene approaches. More than 50% of OsMYBs were strongly correlated under fifty experimental conditions with 51 hub genes via top down approach. Further, clusters were identified using Markov Clustering (MCL. To maximize the clustering performance, parameter evaluation of the MCL inflation score (I was performed in terms of enriched GO categories by measuring F-score. Comparison of co-expressed cluster and clads analyzed from phylogenetic analysis signifies their evolutionarily conserved co-regulatory role. We utilized compendium of known interaction and biological role with Gene Ontology enrichment analysis to hypothesize function of coexpressed OsMYBs. In the other part, the transcriptional regulatory network analysis by guide gene approach revealed 40 putative targets of 26 OsMYB TF hubs with high correlation value utilizing 815 microarray data. The putative targets with MYB-binding cis-elements enrichment in their promoter region, functional co-occurrence as well as nuclear localization supports our finding. Specially, enrichment of MYB binding regions involved in drought-inducibility implying their regulatory role in drought

Phylogenetic analysis reveals conservation and diversification of micro RNA166 genes among diverse plant species.

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Barik, Suvakanta; SarkarDas, Shabari; Singh, Archita; Gautam, Vibhav; Kumar, Pramod; Majee, Manoj; Sarkar, Ananda K

2014-01-01

Similar to the majority of the microRNAs, mature miR166s are derived from multiple members of MIR166 genes (precursors) and regulate various aspects of plant development by negatively regulating their target genes (Class III HD-ZIP). The evolutionary conservation or functional diversification of miRNA166 family members remains elusive. Here, we show the phylogenetic relationships among MIR166 precursor and mature sequences from three diverse model plant species. Despite strong conservation, some mature miR166 sequences, such as ppt-miR166m, have undergone sequence variation. Critical sequence variation in ppt-miR166m has led to functional diversification, as it targets non-HD-ZIPIII gene transcript (s). MIR166 precursor sequences have diverged in a lineage specific manner, and both precursors and mature osa-miR166i/j are highly conserved. Interestingly, polycistronic MIR166s were present in Physcomitrella and Oryza but not in Arabidopsis. The nature of cis-regulatory motifs on the upstream promoter sequences of MIR166 genes indicates their possible contribution to the functional variation observed among miR166 species. Copyright © 2013 Elsevier Inc. All rights reserved.

5' Region of the human interleukin 4 gene: structure and potential regulatory elements

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Eder, A; Krafft-Czepa, H; Krammer, P H

1988-01-25

The lymphokine Interleukin 4 (IL-4) is secreted by antigen or mitogen activated T lymphocytes. IL-4 stimulates activation and differentiation of B lymphocytes and growth of T lymphocytes and mast cells. The authors isolated the human IL-4 gene from a lambda EMBL3 genomic library. As a probe they used a synthetic oligonucleotide spanning position 40 to 79 of the published IL-4 cDNA sequence. The 5' promoter region contains several sequence elements which may have a cis-acting regulatory function for IL-4 gene expression. These elements include a TATA-box, three CCAAT-elements (two are on the non-coding strand) and an octamer motif. A comparison of the 5' flanking region of the human murine IL-4 gene (4) shows that the region between position -306 and +44 is highly conserved (83% homology).

Meta-analysis of breast cancer microarray studies in conjunction with conserved cis-elements suggest patterns for coordinate regulation

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Lundberg Cathryn

2008-01-01

Full Text Available Abstract Background Gene expression measurements from breast cancer (BrCa tumors are established clinical predictive tools to identify tumor subtypes, identify patients showing poor/good prognosis, and identify patients likely to have disease recurrence. However, diverse breast cancer datasets in conjunction with diagnostic clinical arrays show little overlap in the sets of genes identified. One approach to identify a set of consistently dysregulated candidate genes in these tumors is to employ meta-analysis of multiple independent microarray datasets. This allows one to compare expression data from a diverse collection of breast tumor array datasets generated on either cDNA or oligonucleotide arrays. Results We gathered expression data from 9 published microarray studies examining estrogen receptor positive (ER+ and estrogen receptor negative (ER- BrCa tumor cases from the Oncomine database. We performed a meta-analysis and identified genes that were universally up or down regulated with respect to ER+ versus ER- tumor status. We surveyed both the proximal promoter and 3' untranslated regions (3'UTR of our top-ranking genes in each expression group to test whether common sequence elements may contribute to the observed expression patterns. Utilizing a combination of known transcription factor binding sites (TFBS, evolutionarily conserved mammalian promoter and 3'UTR motifs, and microRNA (miRNA seed sequences, we identified numerous motifs that were disproportionately represented between the two gene classes suggesting a common regulatory network for the observed gene expression patterns. Conclusion Some of the genes we identified distinguish key transcripts previously seen in array studies, while others are newly defined. Many of the genes identified as overexpressed in ER- tumors were previously identified as expression markers for neoplastic transformation in multiple human cancers. Moreover, our motif analysis identified a collection of

Highly conserved non-coding elements on either side of SOX9 associated with Pierre Robin sequence.

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Benko, Sabina; Fantes, Judy A; Amiel, Jeanne; Kleinjan, Dirk-Jan; Thomas, Sophie; Ramsay, Jacqueline; Jamshidi, Negar; Essafi, Abdelkader; Heaney, Simon; Gordon, Christopher T; McBride, David; Golzio, Christelle; Fisher, Malcolm; Perry, Paul; Abadie, Véronique; Ayuso, Carmen; Holder-Espinasse, Muriel; Kilpatrick, Nicky; Lees, Melissa M; Picard, Arnaud; Temple, I Karen; Thomas, Paul; Vazquez, Marie-Paule; Vekemans, Michel; Roest Crollius, Hugues; Hastie, Nicholas D; Munnich, Arnold; Etchevers, Heather C; Pelet, Anna; Farlie, Peter G; Fitzpatrick, David R; Lyonnet, Stanislas

2009-03-01

Pierre Robin sequence (PRS) is an important subgroup of cleft palate. We report several lines of evidence for the existence of a 17q24 locus underlying PRS, including linkage analysis results, a clustering of translocation breakpoints 1.06-1.23 Mb upstream of SOX9, and microdeletions both approximately 1.5 Mb centromeric and approximately 1.5 Mb telomeric of SOX9. We have also identified a heterozygous point mutation in an evolutionarily conserved region of DNA with in vitro and in vivo features of a developmental enhancer. This enhancer is centromeric to the breakpoint cluster and maps within one of the microdeletion regions. The mutation abrogates the in vitro enhancer function and alters binding of the transcription factor MSX1 as compared to the wild-type sequence. In the developing mouse mandible, the 3-Mb region bounded by the microdeletions shows a regionally specific chromatin decompaction in cells expressing Sox9. Some cases of PRS may thus result from developmental misexpression of SOX9 due to disruption of very-long-range cis-regulatory elements.

Network perturbation by recurrent regulatory variants in cancer.

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Kiwon Jang

2017-03-01

Full Text Available Cancer driving genes have been identified as recurrently affected by variants that alter protein-coding sequences. However, a majority of cancer variants arise in noncoding regions, and some of them are thought to play a critical role through transcriptional perturbation. Here we identified putative transcriptional driver genes based on combinatorial variant recurrence in cis-regulatory regions. The identified genes showed high connectivity in the cancer type-specific transcription regulatory network, with high outdegree and many downstream genes, highlighting their causative role during tumorigenesis. In the protein interactome, the identified transcriptional drivers were not as highly connected as coding driver genes but appeared to form a network module centered on the coding drivers. The coding and regulatory variants associated via these interactions between the coding and transcriptional drivers showed exclusive and complementary occurrence patterns across tumor samples. Transcriptional cancer drivers may act through an extensive perturbation of the regulatory network and by altering protein network modules through interactions with coding driver genes.

Strand-specific RNA-seq reveals widespread occurrence of novel cis-natural antisense transcripts in rice

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Lu Tingting

2012-12-01

Full Text Available Abstract Background Cis-natural antisense transcripts (cis-NATs are RNAs transcribed from the antisense strand of a gene locus, and are complementary to the RNA transcribed from the sense strand. Common techniques including microarray approach and analysis of transcriptome databases are the major ways to globally identify cis-NATs in various eukaryotic organisms. Genome-wide in silico analysis has identified a large number of cis-NATs that may generate endogenous short interfering RNAs (nat-siRNAs, which participate in important biogenesis mechanisms for transcriptional and post-transcriptional regulation in rice. However, the transcriptomes are yet to be deeply sequenced to comprehensively investigate cis-NATs. Results We applied high-throughput strand-specific complementary DNA sequencing technology (ssRNA-seq to deeply sequence mRNA for assessing sense and antisense transcripts that were derived under salt, drought and cold stresses, and normal conditions, in the model plant rice (Oryza sativa. Combined with RAP-DB genome annotation (the Rice Annotation Project Database build-5 data set, 76,013 transcripts corresponding to 45,844 unique gene loci were assembled, in which 4873 gene loci were newly identified. Of 3819 putative rice cis-NATs, 2292 were detected as expressed and giving rise to small RNAs from their overlapping regions through integrated analysis of ssRNA-seq data and small RNA data. Among them, 503 cis-NATs seemed to be associated with specific conditions. The deep sequence data from isolated epidermal cells of rice seedlings further showed that 54.0% of cis-NATs were expressed simultaneously in a population of homogenous cells. Nearly 9.7% of rice transcripts were involved in one-to-one or many-to-many cis-NATs formation. Furthermore, only 17.4-34.7% of 223 many-to-many cis-NAT groups were all expressed and generated nat-siRNAs, indicating that only some cis-NAT groups may be involved in complex regulatory networks. Conclusions

Genomic dissection of conserved transcriptional regulation in intestinal epithelial cells.

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Colin R Lickwar

2017-08-01

Full Text Available The intestinal epithelium serves critical physiologic functions that are shared among all vertebrates. However, it is unknown how the transcriptional regulatory mechanisms underlying these functions have changed over the course of vertebrate evolution. We generated genome-wide mRNA and accessible chromatin data from adult intestinal epithelial cells (IECs in zebrafish, stickleback, mouse, and human species to determine if conserved IEC functions are achieved through common transcriptional regulation. We found evidence for substantial common regulation and conservation of gene expression regionally along the length of the intestine from fish to mammals and identified a core set of genes comprising a vertebrate IEC signature. We also identified transcriptional start sites and other putative regulatory regions that are differentially accessible in IECs in all 4 species. Although these sites rarely showed sequence conservation from fish to mammals, surprisingly, they drove highly conserved IEC expression in a zebrafish reporter assay. Common putative transcription factor binding sites (TFBS found at these sites in multiple species indicate that sequence conservation alone is insufficient to identify much of the functionally conserved IEC regulatory information. Among the rare, highly sequence-conserved, IEC-specific regulatory regions, we discovered an ancient enhancer upstream from her6/HES1 that is active in a distinct population of Notch-positive cells in the intestinal epithelium. Together, these results show how combining accessible chromatin and mRNA datasets with TFBS prediction and in vivo reporter assays can reveal tissue-specific regulatory information conserved across 420 million years of vertebrate evolution. We define an IEC transcriptional regulatory network that is shared between fish and mammals and establish an experimental platform for studying how evolutionarily distilled regulatory information commonly controls IEC development

Evidence for deep regulatory similarities in early developmental programs across highly diverged insects.

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Kazemian, Majid; Suryamohan, Kushal; Chen, Jia-Yu; Zhang, Yinan; Samee, Md Abul Hassan; Halfon, Marc S; Sinha, Saurabh

2014-09-01

Many genes familiar from Drosophila development, such as the so-called gap, pair-rule, and segment polarity genes, play important roles in the development of other insects and in many cases appear to be deployed in a similar fashion, despite the fact that Drosophila-like "long germband" development is highly derived and confined to a subset of insect families. Whether or not these similarities extend to the regulatory level is unknown. Identification of regulatory regions beyond the well-studied Drosophila has been challenging as even within the Diptera (flies, including mosquitoes) regulatory sequences have diverged past the point of recognition by standard alignment methods. Here, we demonstrate that methods we previously developed for computational cis-regulatory module (CRM) discovery in Drosophila can be used effectively in highly diverged (250-350 Myr) insect species including Anopheles gambiae, Tribolium castaneum, Apis mellifera, and Nasonia vitripennis. In Drosophila, we have successfully used small sets of known CRMs as "training data" to guide the search for other CRMs with related function. We show here that although species-specific CRM training data do not exist, training sets from Drosophila can facilitate CRM discovery in diverged insects. We validate in vivo over a dozen new CRMs, roughly doubling the number of known CRMs in the four non-Drosophila species. Given the growing wealth of Drosophila CRM annotation, these results suggest that extensive regulatory sequence annotation will be possible in newly sequenced insects without recourse to costly and labor-intensive genome-scale experiments. We develop a new method, Regulus, which computes a probabilistic score of similarity based on binding site composition (despite the absence of nucleotide-level sequence alignment), and demonstrate similarity between functionally related CRMs from orthologous loci. Our work represents an important step toward being able to trace the evolutionary history of gene

Evidence for Deep Regulatory Similarities in Early Developmental Programs across Highly Diverged Insects

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Zhang, Yinan; Samee, Md. Abul Hassan; Halfon, Marc S.; Sinha, Saurabh

2014-01-01

Many genes familiar from Drosophila development, such as the so-called gap, pair-rule, and segment polarity genes, play important roles in the development of other insects and in many cases appear to be deployed in a similar fashion, despite the fact that Drosophila-like “long germband” development is highly derived and confined to a subset of insect families. Whether or not these similarities extend to the regulatory level is unknown. Identification of regulatory regions beyond the well-studied Drosophila has been challenging as even within the Diptera (flies, including mosquitoes) regulatory sequences have diverged past the point of recognition by standard alignment methods. Here, we demonstrate that methods we previously developed for computational cis-regulatory module (CRM) discovery in Drosophila can be used effectively in highly diverged (250–350 Myr) insect species including Anopheles gambiae, Tribolium castaneum, Apis mellifera, and Nasonia vitripennis. In Drosophila, we have successfully used small sets of known CRMs as “training data” to guide the search for other CRMs with related function. We show here that although species-specific CRM training data do not exist, training sets from Drosophila can facilitate CRM discovery in diverged insects. We validate in vivo over a dozen new CRMs, roughly doubling the number of known CRMs in the four non-Drosophila species. Given the growing wealth of Drosophila CRM annotation, these results suggest that extensive regulatory sequence annotation will be possible in newly sequenced insects without recourse to costly and labor-intensive genome-scale experiments. We develop a new method, Regulus, which computes a probabilistic score of similarity based on binding site composition (despite the absence of nucleotide-level sequence alignment), and demonstrate similarity between functionally related CRMs from orthologous loci. Our work represents an important step toward being able to trace the evolutionary

Analysis of tomato plasma membrane H(+)-ATPase gene family suggests a mycorrhiza-mediated regulatory mechanism conserved in diverse plant species.

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Liu, Junli; Liu, Jianjian; Chen, Aiqun; Ji, Minjie; Chen, Jiadong; Yang, Xiaofeng; Gu, Mian; Qu, Hongye; Xu, Guohua

2016-10-01

In plants, the plasma membrane H(+)-ATPase (HA) is considered to play a crucial role in regulating plant growth and respoding to environment stresses. Multiple paralogous genes encoding different isozymes of HA have been identified and characterized in several model plants, while limited information of the HA gene family is available to date for tomato. Here, we describe the molecular and expression features of eight HA-encoding genes (SlHA1-8) from tomato. All these genes are interrupted by multiple introns with conserved positions. SlHA1, 2, and 4 were widely expressed in all tissues, while SlHA5, 6, and 7 were almost only expressed in flowers. SlHA8, the transcripts of which were barely detectable under normal or nutrient-/salt-stress growth conditions, was strongly activated in arbuscular mycorrhizal (AM) fungal-colonized roots. Extreme lack of SlHA8 expression in M161, a mutant defective to AM fungal colonization, provided genetic evidence towards the dependence of its expression on AM symbiosis. A 1521-bp SlHA8 promoter could direct the GUS reporter expression specifically in colonized cells of transgenic tobacco, soybean, and rice mycorrhizal roots. Promoter deletion assay revealed a 223-bp promoter fragment of SlHA8 containing a variant of AM-specific cis-element MYCS (vMYCS) sufficient to confer the AM-induced activity. Targeted deletion of this motif in the corresponding promoter region causes complete abolishment of GUS staining in mycorrhizal roots. Together, these results lend cogent evidence towards the evolutionary conservation of a potential regulatory mechanism mediating the activation of AM-responsive HA genes in diverse mycorrhizal plant species.

Characterization of STAT5B phosphorylation correlating with expression of cytokine-inducible SH2-containing protein (CIS).

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Cooper, John C; Boustead, Jared N; Yu, Chao-Lan

2006-06-01

Cytokine-inducible SH2-containing protein (CIS) is the first identified member of genes encoding for the suppressor of cytokine signaling (SOCS). CIS is also a well-known target gene of signal transducer and activator of transcription 5 (STAT5) pathways, providing normal negative feedback control of signaling by cytokines and growth factors. Three other SOCS genes, SOCS1, SOCS2, and SOCS3, can be silenced by DNA hypermethylation in human cancers, suggesting a potential mechanism for constitutive STAT activation. However, it is not known whether CIS expression is similarly perturbed in tumor cells. We report here the absence of CIS expression in T lymphoma LSTRA that overexpresses the Lck protein tyrosine kinase and exhibits elevated STAT5 activity. Pervanadate-induced CIS expression and STAT5 binding to the CIS promoter in vivo over a short time course implies that mechanisms other than DNA hypermethylation may contribute to defective CIS expression in LSTRA cells. Comparison with cytokine-dependent BaF3 cells stimulated with interleukin-3 (IL-3) further reveals that CIS induction correlates with specific STAT5b post-translational modifications. It exhibits as the slowest migrating form through SDS-polyacrylamide gel electrophoresis (SDS-PAGE) analysis. This distinctly modified STAT5b is the predominant form that binds to the consensus STAT5 sites in the CIS promoter and accumulates in the nucleus. In vitro phosphatase assays and phosphoamino acid analysis suggest the involvement of phosphorylation on residues other than the highly conserved tyrosine and serine sites in this distinct STAT5b mobility shift. All together, our results provide a novel link between incomplete STAT5b phosphorylation and defective SOCS gene expression in cancer cells.

Evolution of cichlid vision via trans-regulatory divergence

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O’Quin Kelly E

2012-12-01

Full Text Available Abstract Background Phenotypic evolution may occur through mutations that affect either the structure or expression of protein-coding genes. Although the evolution of color vision has historically been attributed to structural mutations within the opsin genes, recent research has shown that opsin regulatory mutations can also tune photoreceptor sensitivity and color vision. Visual sensitivity in African cichlid fishes varies as a result of the differential expression of seven opsin genes. We crossed cichlid species that express different opsin gene sets and scanned their genome for expression Quantitative Trait Loci (eQTL responsible for these differences. Our results shed light on the role that different structural, cis-, and trans-regulatory mutations play in the evolution of color vision. Results We identified 11 eQTL that contribute to the divergent expression of five opsin genes. On three linkage groups, several eQTL formed regulatory “hotspots” associated with the expression of multiple opsins. Importantly, however, the majority of the eQTL we identified (8/11 or 73% occur on linkage groups located trans to the opsin genes, suggesting that cichlid color vision has evolved primarily via trans-regulatory divergence. By modeling the impact of just two of these trans-regulatory eQTL, we show that opsin regulatory mutations can alter cichlid photoreceptor sensitivity and color vision at least as much as opsin structural mutations can. Conclusions Combined with previous work, we demonstrate that the evolution of cichlid color vision results from the interplay of structural, cis-, and especially trans-regulatory loci. Although there are numerous examples of structural and cis-regulatory mutations that contribute to phenotypic evolution, our results suggest that trans-regulatory mutations could contribute to phenotypic divergence more commonly than previously expected, especially in systems like color vision, where compensatory changes in the

Characterization of noncoding regulatory DNA in the human genome.

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Elkon, Ran; Agami, Reuven

2017-08-08

Genetic variants associated with common diseases are usually located in noncoding parts of the human genome. Delineation of the full repertoire of functional noncoding elements, together with efficient methods for probing their biological roles, is therefore of crucial importance. Over the past decade, DNA accessibility and various epigenetic modifications have been associated with regulatory functions. Mapping these features across the genome has enabled researchers to begin to document the full complement of putative regulatory elements. High-throughput reporter assays to probe the functions of regulatory regions have also been developed but these methods separate putative regulatory elements from the chromosome so that any effects of chromatin context and long-range regulatory interactions are lost. Definitive assignment of function(s) to putative cis-regulatory elements requires perturbation of these elements. Genome-editing technologies are now transforming our ability to perturb regulatory elements across entire genomes. Interpretation of high-throughput genetic screens that incorporate genome editors might enable the construction of an unbiased map of functional noncoding elements in the human genome.

Genome-wide identification and quantification of cis- and trans-regulated genes responding to Marek’s disease virus infection via analysis of allele-specific expression

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Sean eMaceachern

2012-01-01

Full Text Available Marek’s disease (MD is a commercially important neoplastic disease of chickens caused by Marek’s disease virus (MDV, an oncogenic alphaherpesvirus. Selecting for increased genetic resistance to MD is a control strategy that can augment vaccinal control measures. To identify high-confidence candidate MD resistance genes, we conducted a genome-wide screen for allele-specific expression (ASE amongst F1 progeny of two inbred chicken lines that differ in MD resistance. High throughput sequencing was used to profile transcriptomes from pools of uninfected and infected individuals at 4 days post-infection to identify any genes showing ASE in response to MDV infection. RNA sequencing identified 22,655 single nucleotide polymorphisms (SNPs of which 5,360 in 3,773 genes exhibited significant allelic imbalance. Illumina GoldenGate assays were subsequently used to quantify regulatory variation controlled at the gene (cis and elsewhere in the genome (trans by examining differences in expression between F1 individuals and artificial F1 RNA pools over 6 time periods in 1,536 of the most significant SNPs identified by RNA sequencing. Allelic imbalance as a result of cis-regulatory changes was confirmed in 861 of the 1,233 GoldenGate assays successfully examined. Furthermore we have identified 7 genes that display trans-regulation only in infected animals and approximately 500 SNP that show a complex interaction between cis- and trans-regulatory changes. Our results indicate ASE analyses are a powerful approach to identify regulatory variation responsible for differences in transcript abundance in genes underlying complex traits. And the genes with SNPs exhibiting ASE provide a strong foundation to further investigate the causative polymorphisms and genetic mechanisms for MD resistance. Finally, the methods used here for identifying specific genes and SNPs may have practical implications for applying marker-assisted selection to complex traits that are

Mouse transgenesis identifies conserved functional enhancers and cis-regulatory motif in the vertebrate LIM homeobox gene Lhx2 locus.

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Alison P Lee

Full Text Available The vertebrate Lhx2 is a member of the LIM homeobox family of transcription factors. It is essential for the normal development of the forebrain, eye, olfactory system and liver as well for the differentiation of lymphoid cells. However, despite the highly restricted spatio-temporal expression pattern of Lhx2, nothing is known about its transcriptional regulation. In mammals and chicken, Crb2, Dennd1a and Lhx2 constitute a conserved linkage block, while the intervening Dennd1a is lost in the fugu Lhx2 locus. To identify functional enhancers of Lhx2, we predicted conserved noncoding elements (CNEs in the human, mouse and fugu Crb2-Lhx2 loci and assayed their function in transgenic mouse at E11.5. Four of the eight CNE constructs tested functioned as tissue-specific enhancers in specific regions of the central nervous system and the dorsal root ganglia (DRG, recapitulating partial and overlapping expression patterns of Lhx2 and Crb2 genes. There was considerable overlap in the expression domains of the CNEs, which suggests that the CNEs are either redundant enhancers or regulating different genes in the locus. Using a large set of CNEs (810 CNEs associated with transcription factor-encoding genes that express predominantly in the central nervous system, we predicted four over-represented 8-mer motifs that are likely to be associated with expression in the central nervous system. Mutation of one of them in a CNE that drove reporter expression in the neural tube and DRG abolished expression in both domains indicating that this motif is essential for expression in these domains. The failure of the four functional enhancers to recapitulate the complete expression pattern of Lhx2 at E11.5 indicates that there must be other Lhx2 enhancers that are either located outside the region investigated or divergent in mammals and fishes. Other approaches such as sequence comparison between multiple mammals are required to identify and characterize such enhancers.

Molecular evidence for increased regulatory conservation during metamorphosis, and against deleterious cascading effects of hybrid breakdown in Drosophila

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Artieri Carlo G

2010-03-01

circuits during morphogenesis may lead to it being more refractory to divergence of underlying gene regulatory mechanisms - more than that suggested by the conservation of gene expression levels between species during earlier stages. This provides support for a 'developmental hourglass' model of divergence of gene expression in Drosophila resulting in a highly conserved pupal stage.

Conserved-peptide upstream open reading frames (CPuORFs are associated with regulatory genes in angiosperms

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Richard A Jorgensen

2012-08-01

Full Text Available Upstream open reading frames (uORFs are common in eukaryotic transcripts, but those that encode conserved peptides (CPuORFs occur in less than 1% of transcripts. The peptides encoded by three plant CPuORF families are known to control translation of the downstream ORF in response to a small signal molecule (sucrose, polyamines and phosphocholine. In flowering plants, transcription factors are statistically over-represented among genes that possess CPuORFs, and in general it appeared that many CPuORF genes also had other regulatory functions, though the significance of this suggestion was uncertain (Hayden and Jorgensen, 2007. Five years later the literature provides much more information on the functions of many CPuORF genes. Here we reassess the functions of 27 known CPuORF gene families and find that 22 of these families play a variety of different regulatory roles, from transcriptional control to protein turnover, and from small signal molecules to signal transduction kinases. Clearly then, there is indeed a strong association of CPuORFs with regulatory genes. In addition, 16 of these families play key roles in a variety of different biological processes. Most strikingly, the core sucrose response network includes three different CPuORFs, creating the potential for sophisticated balancing of the network in response to three different molecular inputs. We propose that the function of most CPuORFs is to modulate translation of a downstream major ORF (mORF in response to a signal molecule recognized by the conserved peptide and that because the mORFs of CPuORF genes generally encode regulatory proteins, many of them centrally important in the biology of plants, CPuORFs play key roles in balancing such regulatory networks.

Cis-trans photoisomerization of abscisic acid

International Nuclear Information System (INIS)

Brabham, D.E.; Biggs, R.H.

1981-01-01

An important regulator of numerous physiological processes in higher plants is abscisic acid (ABA), which is photoisomerized from the more biologically active cis isomer to the nearly inactive trans isomer by natural sunlight. It is possible that this photoisomerization is a UV control mechanism in functions regulated by ABA. The quantum yields of both the cis to trans and trans to cis photoisomerizations were measured under various conditions of pH and oxygen concentration at room temperature. The yield for photoisomerization of cis-ABA ranged from 0.25 at pH 3.0 to 0.11 at pH 7.0. Oxygen partially quenched the process. The quantum yield varied only slightly with wavelength. The quantum yield of photolysis of cis-ABA was reported for pH 3.0 as 0.06. This yield also varied slightly with wavelength and was relatively insensitive to oxygen. This relatively high yield explains the loss of potency of ABA during UV irradiation. Phosphorescence of cis- and trans-ABA was observed in methanol at 77 K. Onset of the emission was at 350 nm. The emission spectra were the same for both isomers. From these results a mechanism of UV action on plants based on the photoisomerization of the inactive trans-ABA to the biologically active cis isomer is proposed. (author)

Eucalyptus ESTs involved in the production of 9-cis epoxycarotenoid dioxygenase, a regulatory enzyme of abscisic acid production

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Iraê A. Guerrini

2005-01-01

Full Text Available Abscisic acid (ABA regulates stress responses in plants, and genomic tools can help us to understand the mechanisms involved in that process. FAPESP, a Brazilian research foundation, in association with four private forestry companies, has established the FORESTs database (https://forests.esalq.usp.br. A search was carried out in the Eucalyptus expressed sequence tag database to find ESTs involved with 9-cis epoxycarotenoid dioxygenase (NCED, the regulatory enzyme for ABA biosynthesis, using the basic local BLAST alignment tool. We found four clusters (EGEZLV2206B11.g, EGJMWD2252H08.g, EGBFRT3107F10.g, and EGEQFB1200H10.g, which represent similar sequences of the gene that produces NCED. Data showed that the EGBFRT3107F10.g cluster was similar to the maize (Zea mays NCED enzyme, while EGEZLV2206B11.g and EGJMWD2252H08.g clusters were similar to the avocado (Persea americana NCED enzyme. All Eucalyptus clusters were expressed in several tissues, especially in flower buds, where ABA has a special participation during the floral development process.

Genome-wide decoding of hierarchical modular structure of transcriptional regulation by cis-element and expression clustering.

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Leyfer, Dmitriy; Weng, Zhiping

2005-09-01

A holistic approach to the study of cellular processes is identifying both gene-expression changes and regulatory elements promoting such changes. Cellular regulatory processes can be viewed as transcriptional modules (TMs), groups of coexpressed genes regulated by groups of transcription factors (TFs). We set out to devise a method that would identify TMs while avoiding arbitrary thresholds on TM sizes and number. Assuming that gene expression is determined by TFs that bind to the gene's promoter, clustering of genes based on TF binding sites (cis-elements) should create gene groups similar to those obtained by gene expression clustering. Intersections between the expression and cis-element-based gene clusters reveal TMs. Statistical significance assigned to each TM allows identification of regulatory units of any size. Our method correctly identifies the number and sizes of TMs on simulated datasets. We demonstrate that yeast experimental TMs are biologically relevant by comparing them with MIPS and GO categories. Our modules are in statistically significant agreement with TMs from other research groups. This work suggests that there is no preferential division of biological processes into regulatory units; each degree of partitioning exhibits a slice of biological network revealing hierarchical modular organization of transcriptional regulation.

Highly conserved non-coding sequences are associated with vertebrate development.

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Adam Woolfe

2005-01-01

Full Text Available In addition to protein coding sequence, the human genome contains a significant amount of regulatory DNA, the identification of which is proving somewhat recalcitrant to both in silico and functional methods. An approach that has been used with some success is comparative sequence analysis, whereby equivalent genomic regions from different organisms are compared in order to identify both similarities and differences. In general, similarities in sequence between highly divergent organisms imply functional constraint. We have used a whole-genome comparison between humans and the pufferfish, Fugu rubripes, to identify nearly 1,400 highly conserved non-coding sequences. Given the evolutionary divergence between these species, it is likely that these sequences are found in, and furthermore are essential to, all vertebrates. Most, and possibly all, of these sequences are located in and around genes that act as developmental regulators. Some of these sequences are over 90% identical across more than 500 bases, being more highly conserved than coding sequence between these two species. Despite this, we cannot find any similar sequences in invertebrate genomes. In order to begin to functionally test this set of sequences, we have used a rapid in vivo assay system using zebrafish embryos that allows tissue-specific enhancer activity to be identified. Functional data is presented for highly conserved non-coding sequences associated with four unrelated developmental regulators (SOX21, PAX6, HLXB9, and SHH, in order to demonstrate the suitability of this screen to a wide range of genes and expression patterns. Of 25 sequence elements tested around these four genes, 23 show significant enhancer activity in one or more tissues. We have identified a set of non-coding sequences that are highly conserved throughout vertebrates. They are found in clusters across the human genome, principally around genes that are implicated in the regulation of development

Conserved cell cycle regulatory properties within the amino terminal domain of the Epstein-Barr virus nuclear antigen 3C

International Nuclear Information System (INIS)

Sharma, Nikhil; Knight, Jason S.; Robertson, Erle S.

2006-01-01

The gammaherpesviruses Rhesus lymphocryptovirus (LCV) and Epstein-Barr virus (EBV) are closely related phylogenetically. Rhesus LCV efficiently immortalizes Rhesus B cells in vitro. However, despite a high degree of conservation between the Rhesus LCV and EBV genomes, Rhesus LCV fails to immortalize human B cells in vitro. This species restriction may, at least in part, be linked to the EBV nuclear antigens (EBNAs) and latent membrane proteins (LMPs), known to be essential for B cell transformation. We compared specific properties of EBNA3C, a well-characterized and essential EBV protein, with its Rhesus counterpart to determine whether EBNA3C phenotypes which contribute to cell cycle regulation are conserved in the Rhesus LCV. We show that both EBNA3C and Rhesus EBNA3C bind to a conserved region of mammalian cyclins, regulate pRb stability, and modulate SCF Skp2 -dependent ubiquitination. These results suggest that Rhesus LCV restriction from human B cell immortalization is independent of the conserved cell cycle regulatory functions of the EBNA3C protein

Systematic identification of cis-regulatory sequences active in mouse and human embryonic stem cells.

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Marica Grskovic

2007-08-01

Full Text Available Understanding the transcriptional regulation of pluripotent cells is of fundamental interest and will greatly inform efforts aimed at directing differentiation of embryonic stem (ES cells or reprogramming somatic cells. We first analyzed the transcriptional profiles of mouse ES cells and primordial germ cells and identified genes upregulated in pluripotent cells both in vitro and in vivo. These genes are enriched for roles in transcription, chromatin remodeling, cell cycle, and DNA repair. We developed a novel computational algorithm, CompMoby, which combines analyses of sequences both aligned and non-aligned between different genomes with a probabilistic segmentation model to systematically predict short DNA motifs that regulate gene expression. CompMoby was used to identify conserved overrepresented motifs in genes upregulated in pluripotent cells. We show that the motifs are preferentially active in undifferentiated mouse ES and embryonic germ cells in a sequence-specific manner, and that they can act as enhancers in the context of an endogenous promoter. Importantly, the activity of the motifs is conserved in human ES cells. We further show that the transcription factor NF-Y specifically binds to one of the motifs, is differentially expressed during ES cell differentiation, and is required for ES cell proliferation. This study provides novel insights into the transcriptional regulatory networks of pluripotent cells. Our results suggest that this systematic approach can be broadly applied to understanding transcriptional networks in mammalian species.

Ready access to functionally embellished cis-hydrindanes and cis-decalins: protecting group-free total syntheses of (±)-Nootkatone and (±)-Noreremophilane.

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Handore, Kishor L; Seetharamsingh, B; Reddy, D Srinivasa

2013-08-16

A simple and efficient synthesis of functionalized cis-hydrindanes and cis-decalins was achieved using a sequential Diels-Alder/aldol approach in a highly diastereoselective manner. The scope of this method was tested with a variety of substrates and was successfully applied to the synthesis of two natural products in racemic form. The highlights of the present work provide ready access to 13 new cis-hydrindanes/cis-decalins, a protecting group-free total synthesis of an insect repellent Nootkatone, and the first synthesis of a Noreremophilane using the shortest sequence.

Development and utilization of complementary communication channels for treatment decision making and survivorship issues among cancer patients: The CIS Research Consortium Experience.

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Fleisher, Linda; Wen, Kuang Yi; Miller, Suzanne M; Diefenbach, Michael; Stanton, Annette L; Ropka, Mary; Morra, Marion; Raich, Peter C

2015-11-01

Cancer patients and survivors are assuming active roles in decision-making and digital patient support tools are widely used to facilitate patient engagement. As part of Cancer Information Service Research Consortium's randomized controlled trials focused on the efficacy of eHealth interventions to promote informed treatment decision-making for newly diagnosed prostate and breast cancer patients, and post-treatment breast cancer, we conducted a rigorous process evaluation to examine the actual use of and perceived benefits of two complementary communication channels -- print and eHealth interventions. The three Virtual Cancer Information Service (V-CIS) interventions were developed through a rigorous developmental process, guided by self-regulatory theory, informed decision-making frameworks, and health communications best practices. Control arm participants received NCI print materials; experimental arm participants received the additional V-CIS patient support tool. Actual usage data from the web-based V-CIS was also obtained and reported. Print materials were highly used by all groups. About 60% of the experimental group reported using the V-CIS. Those who did use the V-CIS rated it highly on improvements in knowledge, patient-provider communication and decision-making. The findings show that how patients actually use eHealth interventions either singularly or within the context of other communication channels is complex. Integrating rigorous best practices and theoretical foundations is essential and multiple communication approaches should be considered to support patient preferences.

Regulatory elements of the floral homeotic gene AGAMOUS identified by phylogenetic footprinting and shadowing.

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Hong, R. L., Hamaguchi, L., Busch, M. A., and Weigel, D.

2003-06-01

OAK-B135 In Arabidopsis thaliana, cis-regulatory sequences of the floral homeotic gene AGAMOUS (AG) are located in the second intron. This 3 kb intron contains binding sites for two direct activators of AG, LEAFY (LFY) and WUSCHEL (WUS), along with other putative regulatory elements. We have used phylogenetic footprinting and the related technique of phylogenetic shadowing to identify putative cis-regulatory elements in this intron. Among 29 Brassicaceae, several other motifs, but not the LFY and WUS binding sites previously identified, are largely invariant. Using reporter gene analyses, we tested six of these motifs and found that they are all functionally important for activity of AG regulatory sequences in A. thaliana. Although there is little obvious sequence similarity outside the Brassicaceae, the intron from cucumber AG has at least partial activity in A. thaliana. Our studies underscore the value of the comparative approach as a tool that complements gene-by-gene promoter dissection, but also highlight that sequence-based studies alone are insufficient for a complete identification of cis-regulatory sites.

A minimal murine Msx-1 gene promoter. Organization of its cis-regulatory motifs and their role in transcriptional activation in cells in culture and in transgenic mice.

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Takahashi, T; Guron, C; Shetty, S; Matsui, H; Raghow, R

1997-09-05

To dissect the cis-regulatory elements of the murine Msx-1 promoter, which lacks a conventional TATA element, a putative Msx-1 promoter DNA fragment (from -1282 to +106 base pairs (bp)) or its congeners containing site-specific alterations were fused to luciferase reporter and introduced into NIH3T3 and C2C12 cells, and the expression of luciferase was assessed in transient expression assays. The functional consequences of the sequential 5' deletions of the promotor revealed that multiple positive and negative regulatory elements participate in regulating transcription of the Msx-1 gene. Surprisingly, however, the optimal expression of Msx-1 promoter in either NIH3T3 or C2C12 cells required only 165 bp of the upstream sequence to warrant detailed examination of its structure. Therefore, the functional consequences of site-specific deletions and point mutations of the cis-acting elements of the minimal Msx-1 promoter were systematically examined. Concomitantly, potential transcriptional factor(s) interacting with the cis-acting elements of the minimal promoter were also studied by gel electrophoretic mobility shift assays and DNase I footprinting. Combined analyses of the minimal promoter by DNase I footprinting, electrophoretic mobility shift assays, and super shift assays with specific antibodies revealed that 5'-flanking regions from -161 to -154 and from -26 to -13 of the Msx-1 promoter contains an authentic E box (proximal E box), capable of binding a protein immunologically related to the upstream stimulating factor 1 (USF-1) and a GC-rich sequence motif which can bind to Sp1 (proximal Sp1), respectively. Additionally, we observed that the promoter activation was seriously hampered if the proximal E box was removed or mutated, and the promoter activity was eliminated completely if the proximal Sp1 site was similarly altered. Absolute dependence of the Msx-1 minimal promoter on Sp1 could be demonstrated by transient expression assays in the Sp1-deficient

Small RNAs and the regulation of cis-natural antisense transcripts in Arabidopsis

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Lonardi Stefano

2008-01-01

mitochondrion-targeted proteins are over-represented in the Arabidopsis cis-NATs and that 19% of sense and antisense partner genes of cis-NATs share at least one common Gene Ontology term, which suggests that they encode proteins with possible functional connection. Conclusion The negatively correlated expression patterns of sense and antisense genes as well as the presence of siRNAs in many of the cis-NATs suggest that siRNA regulation of cis-NATs via the RNAi pathway is an important gene regulatory mechanism for at least a subgroup of cis-NATs in Arabidopsis.

A robust approach to identifying tissue-specific gene expression regulatory variants using personalized human induced pluripotent stem cells.

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Je-Hyuk Lee

2009-11-01

Full Text Available Normal variation in gene expression due to regulatory polymorphisms is often masked by biological and experimental noise. In addition, some regulatory polymorphisms may become apparent only in specific tissues. We derived human induced pluripotent stem (iPS cells from adult skin primary fibroblasts and attempted to detect tissue-specific cis-regulatory variants using in vitro cell differentiation. We used padlock probes and high-throughput sequencing for digital RNA allelotyping and measured allele-specific gene expression in primary fibroblasts, lymphoblastoid cells, iPS cells, and their differentiated derivatives. We show that allele-specific expression is both cell type and genotype-dependent, but the majority of detectable allele-specific expression loci remains consistent despite large changes in the cell type or the experimental condition following iPS reprogramming, except on the X-chromosome. We show that our approach to mapping cis-regulatory variants reduces in vitro experimental noise and reveals additional tissue-specific variants using skin-derived human iPS cells.

Ancient and recent positive selection transformed opioid cis-regulation in humans.

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Matthew V Rockman

2005-12-01

Full Text Available Changes in the cis-regulation of neural genes likely contributed to the evolution of our species' unique attributes, but evidence of a role for natural selection has been lacking. We found that positive natural selection altered the cis-regulation of human prodynorphin, the precursor molecule for a suite of endogenous opioids and neuropeptides with critical roles in regulating perception, behavior, and memory. Independent lines of phylogenetic and population genetic evidence support a history of selective sweeps driving the evolution of the human prodynorphin promoter. In experimental assays of chimpanzee-human hybrid promoters, the selected sequence increases transcriptional inducibility. The evidence for a change in the response of the brain's natural opioids to inductive stimuli points to potential human-specific characteristics favored during evolution. In addition, the pattern of linked nucleotide and microsatellite variation among and within modern human populations suggests that recent selection, subsequent to the fixation of the human-specific mutations and the peopling of the globe, has favored different prodynorphin cis-regulatory alleles in different parts of the world.

9-cis-retinoic acid represses estrogen-induced expression of the very low density apolipoprotein II gene.

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Schippers, I J; Kloppenburg, M; Snippe, L; Ab, G

1994-11-01

The chicken very low density apolipoprotein II (apoVLDLII) gene is estrogen-inducible and specifically expressed in liver. We examined the possible involvement of the retinoid X receptor (RXR) and its ligand 9-cis-retinoic acid (9-cis-RA) in the activation of the apoVLDLII promoter. We first concentrated on a potential RXR recognition site, which deviates at only one position from a perfect direct A/GGGTCA repeat spaced by one nucleotide (DR-1) and was earlier identified as a common HNF-4/COUP-TF recognition site. However, band shift analysis revealed that this imperfect DR-1 motif does not interact with RXR alpha-homodimers. In accordance with this observation we found that this regulatory element does not mediate transactivation through RXR alpha in the presence of 9-cis-RA. However, our experiments revealed another, unexpected, effect of 9-cis-RA. Instead of stimulating, 9-cis-RA attenuated estrogen-induced expression of transfected estrogen-responsive VLDL-CAT reporter plasmids. This repression appeared to take place through the main estrogen response element (ERE) of the gene. Importantly, 9-cis-RA also strongly repressed the estrogen-induced expression of the endogenous apoVLDLII gene in cultured chicken hepatoma cells.

High qualitative and quantitative conservation of alternative splicing in Caenorhabditis elegans and Caenorhabditis briggsae

DEFF Research Database (Denmark)

Rukov, Jakob Lewin; Irimia, Manuel; Mørk, Søren

2007-01-01

Alternative splicing (AS) is an important contributor to proteome diversity and is regarded as an explanatory factor for the relatively low number of human genes compared with less complex animals. To assess the evolutionary conservation of AS and its developmental regulation, we have investigated...... the qualitative and quantitative expression of 21 orthologous alternative splice events through the development of 2 nematode species separated by 85-110 Myr of evolutionary time. We demonstrate that most of these alternative splice events present in Caenorhabditis elegans are conserved in Caenorhabditis briggsae....... Moreover, we find that relative isoform expression levels vary significantly during development for 78% of the AS events and that this quantitative variation is highly conserved between the 2 species. Our results suggest that AS is generally tightly regulated through development and that the regulatory...

Dynamic in vivo binding of transcription factors to cis-regulatory modules of cer and gsc in the stepwise formation of the Spemann–Mangold organizer

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Sudou, Norihiro; Yamamoto, Shinji; Ogino, Hajime; Taira, Masanori

2012-01-01

How multiple developmental cues are integrated on cis-regulatory modules (CRMs) for cell fate decisions remains uncertain. The Spemann–Mangold organizer in Xenopus embryos expresses the transcription factors Lim1/Lhx1, Otx2, Mix1, Siamois (Sia) and VegT. Reporter analyses using sperm nuclear transplantation and DNA injection showed that cerberus (cer) and goosecoid (gsc) are activated by the aforementioned transcription factors through CRMs conserved between X. laevis and X. tropicalis. ChIP-qPCR analysis for the five transcription factors revealed that cer and gsc CRMs are initially bound by both Sia and VegT at the late blastula stage, and subsequently bound by all five factors at the gastrula stage. At the neurula stage, only binding of Lim1 and Otx2 to the gsc CRM, among others, persists, which corresponds to their co-expression in the prechordal plate. Based on these data, together with detailed expression pattern analysis, we propose a new model of stepwise formation of the organizer, in which (1) maternal VegT and Wnt-induced Sia first bind to CRMs at the blastula stage; then (2) Nodal-inducible Lim1, Otx2, Mix1 and zygotic VegT are bound to CRMs in the dorsal endodermal and mesodermal regions where all these genes are co-expressed; and (3) these two regions are combined at the gastrula stage to form the organizer. Thus, the in vivo dynamics of multiple transcription factors highlight their roles in the initiation and maintenance of gene expression, and also reveal the stepwise integration of maternal, Nodal and Wnt signaling on CRMs of organizer genes to generate the organizer. PMID:22492356

Dynamic in vivo binding of transcription factors to cis-regulatory modules of cer and gsc in the stepwise formation of the Spemann-Mangold organizer.

Science.gov (United States)

Sudou, Norihiro; Yamamoto, Shinji; Ogino, Hajime; Taira, Masanori

2012-05-01

How multiple developmental cues are integrated on cis-regulatory modules (CRMs) for cell fate decisions remains uncertain. The Spemann-Mangold organizer in Xenopus embryos expresses the transcription factors Lim1/Lhx1, Otx2, Mix1, Siamois (Sia) and VegT. Reporter analyses using sperm nuclear transplantation and DNA injection showed that cerberus (cer) and goosecoid (gsc) are activated by the aforementioned transcription factors through CRMs conserved between X. laevis and X. tropicalis. ChIP-qPCR analysis for the five transcription factors revealed that cer and gsc CRMs are initially bound by both Sia and VegT at the late blastula stage, and subsequently bound by all five factors at the gastrula stage. At the neurula stage, only binding of Lim1 and Otx2 to the gsc CRM, among others, persists, which corresponds to their co-expression in the prechordal plate. Based on these data, together with detailed expression pattern analysis, we propose a new model of stepwise formation of the organizer, in which (1) maternal VegT and Wnt-induced Sia first bind to CRMs at the blastula stage; then (2) Nodal-inducible Lim1, Otx2, Mix1 and zygotic VegT are bound to CRMs in the dorsal endodermal and mesodermal regions where all these genes are co-expressed; and (3) these two regions are combined at the gastrula stage to form the organizer. Thus, the in vivo dynamics of multiple transcription factors highlight their roles in the initiation and maintenance of gene expression, and also reveal the stepwise integration of maternal, Nodal and Wnt signaling on CRMs of organizer genes to generate the organizer.

Novel 9-cis/all-trans β-carotene isomerases from plastidic oil bodies in Dunaliella bardawil catalyze the conversion of all-trans to 9-cis β-carotene.

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Davidi, Lital; Pick, Uri

2017-06-01

We identified and demonstrated the function of 9-cis/all-trans β-carotene isomerases in plastidic globules of Dunaliella bardawil, the species accumulating the highest levels of 9-cis β-carotene that is essential for humans. The halotolerant alga Dunaliella bardawil is unique in that it accumulates under light stress high levels of β-carotene in plastidic lipid globules. The pigment is composed of two major isomers: all-trans β-carotene, the common natural form of this pigment, and 9-cis β-carotene. The biosynthetic pathway of β-carotene is known, but it is not clear how the 9-cis isomer is formed. We identified in plastidic lipid globules that were isolated from D. bardawil two proteins with high sequence homology to the D27 protein-a 9-cis/all-trans β-carotene isomerase from rice (Alder et al. Science 335:1348-1351, 2012). The proteins are enriched in the oil globules by 6- to 17-fold compared to chloroplast proteins. The expression of the corresponding genes, 9-cis-βC-iso1 and 9-cis-βC-iso2, is enhanced under light stress. The synthetic proteins catalyze in vitro conversion of all-trans to 9-cis β-carotene. Expression of the 9-cis-βC-iso1 or of 9-cis-βC-iso2 genes in an E. coli mutant line that harbors β-carotene biosynthesis genes enhanced the conversion of all-trans into 9-cis β-carotene. These results suggest that 9-cis-βC-ISO1 and 9-cis-βC-ISO2 proteins are responsible for the formation of 9-cis β-carotene in D. bardawil under stress conditions.

KIRMES: kernel-based identification of regulatory modules in euchromatic sequences.

Science.gov (United States)

Schultheiss, Sebastian J; Busch, Wolfgang; Lohmann, Jan U; Kohlbacher, Oliver; Rätsch, Gunnar

2009-08-15

Understanding transcriptional regulation is one of the main challenges in computational biology. An important problem is the identification of transcription factor (TF) binding sites in promoter regions of potential TF target genes. It is typically approached by position weight matrix-based motif identification algorithms using Gibbs sampling, or heuristics to extend seed oligos. Such algorithms succeed in identifying single, relatively well-conserved binding sites, but tend to fail when it comes to the identification of combinations of several degenerate binding sites, as those often found in cis-regulatory modules. We propose a new algorithm that combines the benefits of existing motif finding with the ones of support vector machines (SVMs) to find degenerate motifs in order to improve the modeling of regulatory modules. In experiments on microarray data from Arabidopsis thaliana, we were able to show that the newly developed strategy significantly improves the recognition of TF targets. The python source code (open source-licensed under GPL), the data for the experiments and a Galaxy-based web service are available at http://www.fml.mpg.de/raetsch/suppl/kirmes/.

LDsplit: screening for cis-regulatory motifs stimulating meiotic recombination hotspots by analysis of DNA sequence polymorphisms.

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Yang, Peng; Wu, Min; Guo, Jing; Kwoh, Chee Keong; Przytycka, Teresa M; Zheng, Jie

2014-02-17

As a fundamental genomic element, meiotic recombination hotspot plays important roles in life sciences. Thus uncovering its regulatory mechanisms has broad impact on biomedical research. Despite the recent identification of the zinc finger protein PRDM9 and its 13-mer binding motif as major regulators for meiotic recombination hotspots, other regulators remain to be discovered. Existing methods for finding DNA sequence motifs of recombination hotspots often rely on the enrichment of co-localizations between hotspots and short DNA patterns, which ignore the cross-individual variation of recombination rates and sequence polymorphisms in the population. Our objective in this paper is to capture signals encoded in genetic variations for the discovery of recombination-associated DNA motifs. Recently, an algorithm called "LDsplit" has been designed to detect the association between single nucleotide polymorphisms (SNPs) and proximal meiotic recombination hotspots. The association is measured by the difference of population recombination rates at a hotspot between two alleles of a candidate SNP. Here we present an open source software tool of LDsplit, with integrative data visualization for recombination hotspots and their proximal SNPs. Applying LDsplit on SNPs inside an established 7-mer motif bound by PRDM9 we observed that SNP alleles preserving the original motif tend to have higher recombination rates than the opposite alleles that disrupt the motif. Running on SNP windows around hotspots each containing an occurrence of the 7-mer motif, LDsplit is able to guide the established motif finding algorithm of MEME to recover the 7-mer motif. In contrast, without LDsplit the 7-mer motif could not be identified. LDsplit is a software tool for the discovery of cis-regulatory DNA sequence motifs stimulating meiotic recombination hotspots by screening and narrowing down to hotspot associated SNPs. It is the first computational method that utilizes the genetic variation of

Does positive selection drive transcription factor binding site turnover? A test with Drosophila cis-regulatory modules.

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Bin Z He

2011-04-01

Full Text Available Transcription factor binding site(s (TFBS gain and loss (i.e., turnover is a well-documented feature of cis-regulatory module (CRM evolution, yet little attention has been paid to the evolutionary force(s driving this turnover process. The predominant view, motivated by its widespread occurrence, emphasizes the importance of compensatory mutation and genetic drift. Positive selection, in contrast, although it has been invoked in specific instances of adaptive gene expression evolution, has not been considered as a general alternative to neutral compensatory evolution. In this study we evaluate the two hypotheses by analyzing patterns of single nucleotide polymorphism in the TFBS of well-characterized CRM in two closely related Drosophila species, Drosophila melanogaster and Drosophila simulans. An important feature of the analysis is classification of TFBS mutations according to the direction of their predicted effect on binding affinity, which allows gains and losses to be evaluated independently along the two phylogenetic lineages. The observed patterns of polymorphism and divergence are not compatible with neutral evolution for either class of mutations. Instead, multiple lines of evidence are consistent with contributions of positive selection to TFBS gain and loss as well as purifying selection in its maintenance. In discussion, we propose a model to reconcile the finding of selection driving TFBS turnover with constrained CRM function over long evolutionary time.

Mechanistically Distinct Pathways of Divergent Regulatory DNA Creation Contribute to Evolution of Human-Specific Genomic Regulatory Networks Driving Phenotypic Divergence of Homo sapiens.

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Glinsky, Gennadi V

2016-09-19

Thousands of candidate human-specific regulatory sequences (HSRS) have been identified, supporting the hypothesis that unique to human phenotypes result from human-specific alterations of genomic regulatory networks. Collectively, a compendium of multiple diverse families of HSRS that are functionally and structurally divergent from Great Apes could be defined as the backbone of human-specific genomic regulatory networks. Here, the conservation patterns analysis of 18,364 candidate HSRS was carried out requiring that 100% of bases must remap during the alignments of human, chimpanzee, and bonobo sequences. A total of 5,535 candidate HSRS were identified that are: (i) highly conserved in Great Apes; (ii) evolved by the exaptation of highly conserved ancestral DNA; (iii) defined by either the acceleration of mutation rates on the human lineage or the functional divergence from non-human primates. The exaptation of highly conserved ancestral DNA pathway seems mechanistically distinct from the evolution of regulatory DNA segments driven by the species-specific expansion of transposable elements. Genome-wide proximity placement analysis of HSRS revealed that a small fraction of topologically associating domains (TADs) contain more than half of HSRS from four distinct families. TADs that are enriched for HSRS and termed rapidly evolving in humans TADs (revTADs) comprise 0.8-10.3% of 3,127 TADs in the hESC genome. RevTADs manifest distinct correlation patterns between placements of human accelerated regions, human-specific transcription factor-binding sites, and recombination rates. There is a significant enrichment within revTAD boundaries of hESC-enhancers, primate-specific CTCF-binding sites, human-specific RNAPII-binding sites, hCONDELs, and H3K4me3 peaks with human-specific enrichment at TSS in prefrontal cortex neurons (P sapiens is driven by the evolution of human-specific genomic regulatory networks via at least two mechanistically distinct pathways of creation of

A generalized allosteric mechanism for cis-regulated cyclic nucleotide binding domains.

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Alexandr P Kornev

2008-04-01

Full Text Available Cyclic nucleotides (cAMP and cGMP regulate multiple intracellular processes and are thus of a great general interest for molecular and structural biologists. To study the allosteric mechanism of different cyclic nucleotide binding (CNB domains, we compared cAMP-bound and cAMP-free structures (PKA, Epac, and two ionic channels using a new bioinformatics method: local spatial pattern alignment. Our analysis highlights four major conserved structural motifs: 1 the phosphate binding cassette (PBC, which binds the cAMP ribose-phosphate, 2 the "hinge," a flexible helix, which contacts the PBC, 3 the beta(2,3 loop, which provides precise positioning of an invariant arginine from the PBC, and 4 a conserved structural element consisting of an N-terminal helix, an eight residue loop and the A-helix (N3A-motif. The PBC and the hinge were included in the previously reported allosteric model, whereas the definition of the beta(2,3 loop and the N3A-motif as conserved elements is novel. The N3A-motif is found in all cis-regulated CNB domains, and we present a model for an allosteric mechanism in these domains. Catabolite gene activator protein (CAP represents a trans-regulated CNB domain family: it does not contain the N3A-motif, and its long range allosteric interactions are substantially different from the cis-regulated CNB domains.

Exploring the miRNA regulatory network using evolutionary correlations.

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Benedikt Obermayer

2014-10-01

Full Text Available Post-transcriptional regulation by miRNAs is a widespread and highly conserved phenomenon in metazoans, with several hundreds to thousands of conserved binding sites for each miRNA, and up to two thirds of all genes under miRNA regulation. At the same time, the effect of miRNA regulation on mRNA and protein levels is usually quite modest and associated phenotypes are often weak or subtle. This has given rise to the notion that the highly interconnected miRNA regulatory network exerts its function less through any individual link and more via collective effects that lead to a functional interdependence of network links. We present a Bayesian framework to quantify conservation of miRNA target sites using vertebrate whole-genome alignments. The increased statistical power of our phylogenetic model allows detection of evolutionary correlation in the conservation patterns of site pairs. Such correlations could result from collective functions in the regulatory network. For instance, co-conservation of target site pairs supports a selective benefit of combinatorial regulation by multiple miRNAs. We find that some miRNA families are under pronounced co-targeting constraints, indicating a high connectivity in the regulatory network, while others appear to function in a more isolated way. By analyzing coordinated targeting of different curated gene sets, we observe distinct evolutionary signatures for protein complexes and signaling pathways that could reflect differences in control strategies. Our method is easily scalable to analyze upcoming larger data sets, and readily adaptable to detect high-level selective constraints between other genomic loci. We thus provide a proof-of-principle method to understand regulatory networks from an evolutionary perspective.

Interplay of cis- and trans-regulatory mechanisms in the spliceosomal RNA helicase Brr2.

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Absmeier, Eva; Becke, Christian; Wollenhaupt, Jan; Santos, Karine F; Wahl, Markus C

2017-01-02

RNA helicase Brr2 is implicated in multiple phases of pre-mRNA splicing and thus requires tight regulation. Brr2 can be auto-inhibited via a large N-terminal region folding back onto its helicase core and auto-activated by a catalytically inactive C-terminal helicase cassette. Furthermore, it can be regulated in trans by the Jab1 domain of the Prp8 protein, which can inhibit Brr2 by intermittently inserting a C-terminal tail in the enzyme's RNA-binding tunnel or activate the helicase after removal of this tail. Presently it is unclear, whether these regulatory mechanisms functionally interact and to which extent they are evolutionarily conserved. Here, we report crystal structures of Saccharomyces cerevisiae and Chaetomium thermophilum Brr2-Jab1 complexes, demonstrating that Jab1-based inhibition of Brr2 presumably takes effect in all eukaryotes but is implemented via organism-specific molecular contacts. Moreover, the structures show that Brr2 auto-inhibition can act in concert with Jab1-mediated inhibition, and suggest that the N-terminal region influences how the Jab1 C-terminal tail interacts at the RNA-binding tunnel. Systematic RNA binding and unwinding studies revealed that the N-terminal region and the Jab1 C-terminal tail specifically interfere with accommodation of double-stranded and single-stranded regions of an RNA substrate, respectively, mutually reinforcing each other. Additionally, such analyses show that regulation based on the N-terminal region requires the presence of the inactive C-terminal helicase cassette. Together, our results outline an intricate system of regulatory mechanisms, which control Brr2 activities during snRNP assembly and splicing.

Cis-Lunar Base Camp

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Merrill, Raymond G.; Goodliff, Kandyce E.; Mazanek, Daniel D.; Reeves, John D., Jr.

2012-01-01

Historically, when mounting expeditions into uncharted territories, explorers have established strategically positioned base camps to pre-position required equipment and consumables. These base camps are secure, safe positions from which expeditions can depart when conditions are favorable, at which technology and operations can be tested and validated, and facilitate timely access to more robust facilities in the event of an emergency. For human exploration missions into deep space, cis-lunar space is well suited to serve as such a base camp. The outer regions of cis-lunar space, such as the Earth-Moon Lagrange points, lie near the edge of Earth s gravity well, allowing equipment and consumables to be aggregated with easy access to deep space and to the lunar surface, as well as more distant destinations, such as near-Earth Asteroids (NEAs) and Mars and its moons. Several approaches to utilizing a cis-lunar base camp for sustainable human exploration, as well as some possible future applications are identified. The primary objective of the analysis presented in this paper is to identify options, show the macro trends, and provide information that can be used as a basis for more detailed mission development. Compared within are the high-level performance and cost of 15 preliminary cis-lunar exploration campaigns that establish the capability to conduct crewed missions of up to one year in duration, and then aggregate mass in cis-lunar space to facilitate an expedition from Cis-Lunar Base Camp. Launch vehicles, chemical propulsion stages, and electric propulsion stages are discussed and parametric sizing values are used to create architectures of in-space transportation elements that extend the existing in-space supply chain to cis-lunar space. The transportation options to cis-lunar space assessed vary in efficiency by almost 50%; from 0.16 to 0.68 kg of cargo in cis-lunar space for every kilogram of mass in Low Earth Orbit (LEO). For the 15 cases, 5-year campaign

Identification of cis-acting regulatory elements in the human oxytocin gene promoter.

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Richard, S; Zingg, H H

1991-12-01

The expression of hormone-inducible genes is determined by the interaction of trans-acting factors with hormone-inducible elements and elements mediating basal and cell-specific expression. We have shown earlier that the gene encoding the hypothalamic nonapeptide oxytocin (OT) is under the control of an estrogen response element (ERE). The present study was aimed at identifying cis-acting elements mediating basal expression of the OT gene. A construct containing sequences -381 to +36 of the human OT gene was linked to a reporter gene and transiently transfected into a series of neuronal and nonneuronal cell lines. Expression of this construct was cell specific: it was highest in the neuroblastoma-derived cell line, Neuro-2a, and lowest in NIH 3T3 and JEG-3 cells. By 5' deletion analysis, we determined that a segment from -49 to +36 was capable of mediating cells-pecific promoter activity. Within this segment, we identified three proximal promoter elements (PPE-1, PPE-2, and PPE-3) that are each required for promoter activity. Most notably, mutation of a conserved purine-rich element (GAGAGA) contained within PPE-2 leads to a 10-fold decrease in promoter strength. Gel mobility shift analysis with three different double-stranded oligonucleotides demonstrated that each proximal promoter element binds distinct nuclear factors. In each case, only the homologous oligonucleotide, but neither of the oligonucleotides corresponding to adjacent elements, was able to act as a competitor. Thus, a different set of factors appears to bind independently to each element. By reinserting the homologous ERE or a heterologous glucocorticoid response element upstream of intact or altered proximal promoter segments we determined that removal or mutation of proximal promoter elements decreases basal expression, but does not abrogate the hormone responsiveness of the promoter. In conclusion, these results indicate that an important component of the transcriptional activity of the OT

Generation of Chimeric RNAs by cis-splicing of adjacent genes (cis-SAGe) in mammals.

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Zhuo, Jian-Shu; Jing, Xiao-Yan; Du, Xin; Yang, Xiu-Qin

2018-02-20

Chimeric RNA molecules, possessing exons from two or more independent genes, are traditionally believed to be produced by chromosome rearrangement. However, recent studies revealed that cis-splicing of adjacent genes (cis- SAGe) is one of the major mechanisms underlying the formation of chimeric RNAs. cis-SAGe refers to intergenic splicing of directly adjacent genes with the same transcriptional orientation, resulting in read-through transcripts, termed chimeric RNAs, which contain sequences from two or more parental genes. cis-SAGe was first identified in tumor cells, since then its potential in carcinogenesis has attracted extensive attention. More and more scientists are focusing on it. With the development of research, cis-SAGe was found to be ubiquitous in various normal tissues, and might make a crucial contribution to the formation of novel genes in the evolution of genomes. In this review, we summarize the splicing pattern, expression characteristics, possible mechanisms, and significance of cis-SAGe in mammals. This review will be helpful for general understanding of the current status and development tendency of cis-SAGe.

Bioinformatic analysis of cis-regulatory interactions between progesterone and estrogen receptors in breast cancer

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Matloob Khushi

2014-11-01

Full Text Available Chromatin factors interact with each other in a cell and sequence-specific manner in order to regulate transcription and a wealth of publically available datasets exists describing the genomic locations of these interactions. Our recently published BiSA (Binding Sites Analyser database contains transcription factor binding locations and epigenetic modifications collected from published studies and provides tools to analyse stored and imported data. Using BiSA we investigated the overlapping cis-regulatory role of estrogen receptor alpha (ERα and progesterone receptor (PR in the T-47D breast cancer cell line. We found that ERα binding sites overlap with a subset of PR binding sites. To investigate further, we re-analysed raw data to remove any biases introduced by the use of distinct tools in the original publications. We identified 22,152 PR and 18,560 ERα binding sites (<5% false discovery rate with 4,358 overlapping regions among the two datasets. BiSA statistical analysis revealed a non-significant overall overlap correlation between the two factors, suggesting that ERα and PR are not partner factors and do not require each other for binding to occur. However, Monte Carlo simulation by Binary Interval Search (BITS, Relevant Distance, Absolute Distance, Jaccard and Projection tests by Genometricorr revealed a statistically significant spatial correlation of binding regions on chromosome between the two factors. Motif analysis revealed that the shared binding regions were enriched with binding motifs for ERα, PR and a number of other transcription and pioneer factors. Some of these factors are known to co-locate with ERα and PR binding. Therefore spatially close proximity of ERα binding sites with PR binding sites suggests that ERα and PR, in general function independently at the molecular level, but that their activities converge on a specific subset of transcriptional targets.

Regulatory interventions necessitated by non-conservative operator decisions

International Nuclear Information System (INIS)

Ojha, D.; Chande, S.K.; Sharma, S.K.

2005-01-01

Presently, India has 15 nuclear power units in operation and 8 units under construction. Though the safety performance of the Nuclear Power Plants (NPPs) in India has been excellent, a few recent events indicate that conservative decision making process can possibly get affected by perceived production goals. In one of the events, a need for some maintenance work arose while reactor start-up was in progress. After it was realized that the maintenance would require considerable time, the proper course of action would have been to shutdown the reactor and add neutron poison to moderator to ensure sufficient sub criticality. This was not done, as it would have delayed the start up of the reactor on completion of maintenance. In another incident, an unintended slow increase in reactor power occurred due to the adjuster rods becoming inoperable on account of blowing-off of fuses in their power supply. Under this condition, the reactor should have been tripped which was not done. Further, the automatic addition of boron poison to the reactor was inhibited. Regulatory review showed that both the incidents were indicative of degradation in safety culture and reflected operator's overriding concern for keeping the units in operation. Appropriate corrective actions were taken to prevent recurrence of such events in the respective units as also in all other operating units of similar type. In the wake of improved production performance operators may develop a tendency to create new operational records and compete with other units. This aspect points out to a need for careful study of events to check the presence of any element of non-conservative decision-making and to identify leading indicators of degradation in safety performance. (author)

Statistics of high-altitude and high-latitude O+ ion outflows observed by Cluster/CIS

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A. Korth

2005-07-01

Full Text Available The persistent outflows of O+ ions observed by the Cluster CIS/CODIF instrument were studied statistically in the high-altitude (from 3 up to 11 RE and high-latitude (from 70 to ~90 deg invariant latitude, ILAT polar region. The principal results are: (1 Outflowing O+ ions with more than 1keV are observed above 10 RE geocentric distance and above 85deg ILAT location; (2 at 6-8 RE geocentric distance, the latitudinal distribution of O+ ion outflow is consistent with velocity filter dispersion from a source equatorward and below the spacecraft (e.g. the cusp/cleft; (3 however, at 8-12 RE geocentric distance the distribution of O+ outflows cannot be explained by velocity filter only. The results suggest that additional energization or acceleration processes for outflowing O+ ions occur at high altitudes and high latitudes in the dayside polar region. Keywords. Magnetospheric physics (Magnetospheric configuration and dynamics, Solar wind-magnetosphere interactions

Evolutionary growth process of highly conserved sequences in vertebrate genomes.

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Ishibashi, Minaka; Noda, Akiko Ogura; Sakate, Ryuichi; Imanishi, Tadashi

2012-08-01

Genome sequence comparison between evolutionarily distant species revealed ultraconserved elements (UCEs) among mammals under strong purifying selection. Most of them were also conserved among vertebrates. Because they tend to be located in the flanking regions of developmental genes, they would have fundamental roles in creating vertebrate body plans. However, the evolutionary origin and selection mechanism of these UCEs remain unclear. Here we report that UCEs arose in primitive vertebrates, and gradually grew in vertebrate evolution. We searched for UCEs in two teleost fishes, Tetraodon nigroviridis and Oryzias latipes, and found 554 UCEs with 100% identity over 100 bps. Comparison of teleost and mammalian UCEs revealed 43 pairs of common, jawed-vertebrate UCEs (jUCE) with high sequence identities, ranging from 83.1% to 99.2%. Ten of them retain lower similarities to the Petromyzon marinus genome, and the substitution rates of four non-exonic jUCEs were reduced after the teleost-mammal divergence, suggesting that robust conservation had been acquired in the jawed vertebrate lineage. Our results indicate that prototypical UCEs originated before the divergence of jawed and jawless vertebrates and have been frozen as perfect conserved sequences in the jawed vertebrate lineage. In addition, our comparative sequence analyses of UCEs and neighboring regions resulted in a discovery of lineage-specific conserved sequences. They were added progressively to prototypical UCEs, suggesting step-wise acquisition of novel regulatory roles. Our results indicate that conserved non-coding elements (CNEs) consist of blocks with distinct evolutionary history, each having been frozen since different evolutionary era along the vertebrate lineage. Copyright © 2012 Elsevier B.V. All rights reserved.

CisSERS: Customizable In Silico Sequence Evaluation for Restriction Sites.

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Sharpe, Richard M; Koepke, Tyson; Harper, Artemus; Grimes, John; Galli, Marco; Satoh-Cruz, Mio; Kalyanaraman, Ananth; Evans, Katherine; Kramer, David; Dhingra, Amit

2016-01-01

High-throughput sequencing continues to produce an immense volume of information that is processed and assembled into mature sequence data. Data analysis tools are urgently needed that leverage the embedded DNA sequence polymorphisms and consequent changes to restriction sites or sequence motifs in a high-throughput manner to enable biological experimentation. CisSERS was developed as a standalone open source tool to analyze sequence datasets and provide biologists with individual or comparative genome organization information in terms of presence and frequency of patterns or motifs such as restriction enzymes. Predicted agarose gel visualization of the custom analyses results was also integrated to enhance the usefulness of the software. CisSERS offers several novel functionalities, such as handling of large and multiple datasets in parallel, multiple restriction enzyme site detection and custom motif detection features, which are seamlessly integrated with real time agarose gel visualization. Using a simple fasta-formatted file as input, CisSERS utilizes the REBASE enzyme database. Results from CisSERS enable the user to make decisions for designing genotyping by sequencing experiments, reduced representation sequencing, 3'UTR sequencing, and cleaved amplified polymorphic sequence (CAPS) molecular markers for large sample sets. CisSERS is a java based graphical user interface built around a perl backbone. Several of the applications of CisSERS including CAPS molecular marker development were successfully validated using wet-lab experimentation. Here, we present the tool CisSERS and results from in-silico and corresponding wet-lab analyses demonstrating that CisSERS is a technology platform solution that facilitates efficient data utilization in genomics and genetics studies.

CisLunar Habitat Internal Architecture Design Criteria

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Jones, R.; Kennedy, K.; Howard, R.; Whitmore, M.; Martin, C.; Garate, J.

2017-01-01

Lunar Habitat Internal Architecture Study is to become a forcing function to establish a common understanding of CisLunar Phase-1 Habitation Internal Architecture design criteria, processes, and tools. The scope of the CisLunar Habitat Internal Architecture study is to design, develop, demonstrate, and evaluate a Phase-1 CisLunar Habitat common module internal architecture based on design criteria agreed to by NASA, the International Partners, and Commercial Exploration teams. This task is to define the CisLunar Phase-1 Internal Architecture Government Reference Design, assist NASA in becoming a "smart buyer" for Phase-1 Habitat Concepts, and ultimately to derive standards and requirements from the Internal Architecture Design Process. The first step was to define a Habitat Internal Architecture Design Criteria and create a structured philosophy to be used by design teams as a filter by which critical aspects of consideration would be identified for the purpose of organizing and utilizing interior spaces. With design criteria in place, the team will develop a series of iterative internal architecture concept designs which will be assessed by means of an evaluation criteria and process. These assessments will successively drive and refine the design, leading to the combination and down-selection of design concepts. A single refined reference design configuration will be developed into in a medium-to-high fidelity mockup. A multi-day human-in-the-loop mission test will fully evaluate the reference design and validate its configuration. Lessons learned from the design and evaluation will enable the team to identify appropriate standards for Phase-1 CisLunar Habitat Internal Architecture and will enable NASA to develop derived requirements in support of maturing CisLunar Habitation capabilities. This paper will describe the criteria definition process, workshop event, and resulting CisLunar Phase-1 Habitat Internal Architecture Design Criteria.

Human apolipoprotein CIII gene expression is regulated by positive and negative cis-acting elements and tissue-specific protein factors

International Nuclear Information System (INIS)

Reue, K.; Leff, T.; Breslow, J.L.

1988-01-01

Apolipoprotein CIII (apoCIII) is a major protein constituent of triglyceride-rich lipoproteins and is synthesized primarily in the liver. Cis-acting DNA elements required for liver-specific apoCIII gene transcription were identified with transient expression assays in the human hepatoma (HepG2) and epithelial carcinoma (HeLa) cell lines. In liver cells, 821 nucleotides of the human apoCIII gene 5'-flanking sequence were required for maximum levels of gene expression, while the proximal 110 nucleotides alone were sufficient. No expression was observed in similar studies with HeLa cells. The level of expression was modulated by a combination of positive and negative cis-acting sequences, which interact with distinct sets of proteins from liver and HeLa cell nuclear extracts. The proximal positive regulatory region shares homology with similarly located sequences of other genes strongly expressed in the liver, including α 1 -antitrypsin and other apolipoprotein genes. The negative regulatory region is striking homologous to the human β-interferon gene regulatory element. The distal positive region shares homology with some viral enhancers and has properties of a tissue-specific enhancer. The regulation of the apoCIII gene is complex but shares features with other genes, suggesting shuffling of regulatory elements as a common mechanism for cell type-specific gene expression

Conserved regulatory modules in the Sox9 testis-specific enhancer predict roles for SOX, TCF/LEF, Forkhead, DMRT, and GATA proteins in vertebrate sex determination.

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Bagheri-Fam, Stefan; Sinclair, Andrew H; Koopman, Peter; Harley, Vincent R

2010-03-01

While the primary sex determining switch varies between vertebrate species, a key downstream event in testicular development, namely the male-specific up-regulation of Sox9, is conserved. To date, only two sex determining switch genes have been identified, Sry in mammals and the Dmrt1-related gene Dmy (Dmrt1bY) in the medaka fish Oryzias latipes. In mice, Sox9 expression is evidently up-regulated by SRY and maintained by SOX9 both of which directly activate the core 1.3 kb testis-specific enhancer of Sox9 (TESCO). How Sox9 expression is up-regulated and maintained in species without Sry (i.e. non-mammalian species) is not understood. In this study, we have undertaken an in-depth comparative genomics approach and show that TESCO contains an evolutionarily conserved region (ECR) of 180 bp which is present in marsupials, monotremes, birds, reptiles and amphibians. The ECR contains highly conserved modules that predict regulatory roles for SOX, TCF/LEF, Forkhead, DMRT, and GATA proteins in vertebrate sex determination/differentiation. Our data suggest that tetrapods share common aspects of Sox9 regulation in the testis, despite having different sex determining switch mechanisms. They also suggest that Sox9 autoregulation is an ancient mechanism shared by all tetrapods, raising the possibility that in mammals, SRY evolved by mimicking this regulation. The validation of ECR regulatory sequences conserved from human to frogs will provide new insights into vertebrate sex determination. Copyright 2009 Elsevier Ltd. All rights reserved.

A conserved regulatory mechanism in bifunctional biotin protein ligases.

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Wang, Jingheng; Beckett, Dorothy

2017-08-01

Class II bifunctional biotin protein ligases (BirA), which catalyze post-translational biotinylation and repress transcription initiation, are broadly distributed in eubacteria and archaea. However, it is unclear if these proteins all share the same molecular mechanism of transcription regulation. In Escherichia coli the corepressor biotinoyl-5'-AMP (bio-5'-AMP), which is also the intermediate in biotin transfer, promotes operator binding and resulting transcription repression by enhancing BirA dimerization. Like E. coli BirA (EcBirA), Staphylococcus aureus, and Bacillus subtilis BirA (Sa and BsBirA) repress transcription in vivo in a biotin-dependent manner. In this work, sedimentation equilibrium measurements were performed to investigate the molecular basis of this biotin-responsive transcription regulation. The results reveal that, as observed for EcBirA, Sa, and BsBirA dimerization reactions are significantly enhanced by bio-5'-AMP binding. Thus, the molecular mechanism of the Biotin Regulatory System is conserved in the biotin repressors from these three organisms. © 2017 The Protein Society.

In silico analysis, mapping of regulatory elements and corresponding dna-protein interaction in polyphenol oxidase gene promoter from different rice varieties

International Nuclear Information System (INIS)

Mahmood, T.; Rehman, M.; Aziz, E.

2015-01-01

Polyphenol oxidase (PPO) is an important enzyme that has positive impact regarding plant resistance against different biotic and abiotic stresses. In the present study PPO promoter from six different rice varieties was amplified and then analyzed for cis- and trans-acting elements. The study revealed a total of 79 different cis-acting regulatory elements including 11 elements restricted to only one or other variety. Among six varieties Pakhal-Basmati had highest number (5) of these elements, whereas C-622 and Rachna-Basmati have no such sequences. Rachna-Basmati, IR-36-Basmati and Kashmir- Basmati had 1, 2 and 3 unique elements, respectively. Different elementsrelated to pathogen, salt and water stresses were found, which may be helpful in controlling PPO activity according to changing environment. Moreover, HADDOCK was used to understand molecular mechanism of PPO regulation and it was found that DNA-protein interactions are stabilized by many potential hydrogen bonds. Adenine and arginine were the most reactive residues in DNA and proteins respectively.Structural comparison of different protein-DNA complexes show that even a highly conserved transcriptional factor can adopt different conformations when they contact a different DNA binding sequence, however their stable interactions depend on the number of hydrogen bonds formed and distance. (author)

Ionizing radiation sources management in the Commonwealth of Independent States - CIS

International Nuclear Information System (INIS)

Iskra, A.; Bufetova, M.

2006-01-01

Ionizing radiation sources cover a broad band of power: from powerful NPP reactors and research reactors to portable radioisotope ionizing radiation sources applied in medicine, agriculture, industry and in the energy supply systems of remote facilities. At present, scales and use field of radionuclide sources in the CIS have the tendency to increase. In this connection, the issues of ionizing radiation sources management safety at all stages of their life cycle, from production to treatment, have been of a great importance. The materials on ionizing radiation sources inventory and treatment in the CIS (Russia, Armenia, Belarus, Georgia, Kazakhstan, Kyrgyzstan, Tajikistan and Ukraine) are presented in the report. It is shown that in some republics, there is difficulty in ionizing radiation sources accounting and control system; the national regulatory and legal framework bases regulating activity on radioactive sources use, localization and treatment require update. Many problems are connected with the sources beyond state accounting. The problem of ionizing radiation sources use safety is complicated by the growing activity of various terrorist groups. The opportunity to use ionizing radiation sources with terrorism goals requires the application of defined systems of security and physical protection at all stages of their management. For this purpose a collective, with all CIS countries, organization of radioactive sources accounting and control as well as countermeasures on their illegal transportation and use are necessary. In this connection, the information collection regarding situation with providing of ionizing radiation sources safety, conditions of equipment and storage facilities, radioactive materials accounting and control system in the CIS countries is vitally needed

CisSERS: Customizable In Silico Sequence Evaluation for Restriction Sites.

Directory of Open Access Journals (Sweden)

Richard M Sharpe

Full Text Available High-throughput sequencing continues to produce an immense volume of information that is processed and assembled into mature sequence data. Data analysis tools are urgently needed that leverage the embedded DNA sequence polymorphisms and consequent changes to restriction sites or sequence motifs in a high-throughput manner to enable biological experimentation. CisSERS was developed as a standalone open source tool to analyze sequence datasets and provide biologists with individual or comparative genome organization information in terms of presence and frequency of patterns or motifs such as restriction enzymes. Predicted agarose gel visualization of the custom analyses results was also integrated to enhance the usefulness of the software. CisSERS offers several novel functionalities, such as handling of large and multiple datasets in parallel, multiple restriction enzyme site detection and custom motif detection features, which are seamlessly integrated with real time agarose gel visualization. Using a simple fasta-formatted file as input, CisSERS utilizes the REBASE enzyme database. Results from CisSERS enable the user to make decisions for designing genotyping by sequencing experiments, reduced representation sequencing, 3'UTR sequencing, and cleaved amplified polymorphic sequence (CAPS molecular markers for large sample sets. CisSERS is a java based graphical user interface built around a perl backbone. Several of the applications of CisSERS including CAPS molecular marker development were successfully validated using wet-lab experimentation. Here, we present the tool CisSERS and results from in-silico and corresponding wet-lab analyses demonstrating that CisSERS is a technology platform solution that facilitates efficient data utilization in genomics and genetics studies.

Readings in Wildlife and Fish Conservation, High School Conservation Curriculum Project.

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Ensminger, Jack

This publication is a tentative edition of readings on Wildlife and Fish Conservation in Louisiana, and as such it forms part of one of the four units of study designed for an experimental high school course, the "High School Conservation Curriculum Project." The other three units are concerned with Forest Conervation, Soil and Water…

Analytical and semipreparative chiral separation of cis-itraconazole on cellulose stationary phases by high-performance liquid chromatography.

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Kurka, Ondřej; Kučera, Lukáš; Bednář, Petr

2016-07-01

cis-Itraconazole is a chiral antifungal drug administered as a racemate. The knowledge of properties of individual cis-itraconazole stereoisomers is vital information for medicine and biosciences as different stereoisomers of cis-itraconazole may possess different affinity to certain biological pathways in the human body. For this purpose, either chiral synthesis of enantiomers or chiral separation of racemate can be used. This paper presents a two-step high-performance liquid chromatography approach for the semipreparative isolation of four stereoisomers (two enantiomeric pairs) of itraconazole using polysaccharide stationary phases and volatile organic mobile phases without additives in isocratic mode. The approach used involves the separation of the racemate into three fractions (i.e. two pure stereoisomers and one mixed fraction containing the remaining two stereoisomers) in the first run and consequent separation of the collected mixed fraction in the second one. For this purpose, combination of cellulose tris-(4-methylbenzoate) and cellulose tris-(3,5-dimehylphenylcarbamate) columns with complementary selectivity for cis-itraconazole provided full separation of all four stereoisomers (with purity of each isomer > 97%). The stereoisomers were collected, their optical rotation determined and their identity confirmed based on the results of a previously published study. Pure separated stereoisomers are subjected to further biological studies. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Prediction of transcriptional regulatory elements for plant hormone responses based on microarray data

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Yamaguchi-Shinozaki Kazuko

2011-02-01

Full Text Available Abstract Background Phytohormones organize plant development and environmental adaptation through cell-to-cell signal transduction, and their action involves transcriptional activation. Recent international efforts to establish and maintain public databases of Arabidopsis microarray data have enabled the utilization of this data in the analysis of various phytohormone responses, providing genome-wide identification of promoters targeted by phytohormones. Results We utilized such microarray data for prediction of cis-regulatory elements with an octamer-based approach. Our test prediction of a drought-responsive RD29A promoter with the aid of microarray data for response to drought, ABA and overexpression of DREB1A, a key regulator of cold and drought response, provided reasonable results that fit with the experimentally identified regulatory elements. With this succession, we expanded the prediction to various phytohormone responses, including those for abscisic acid, auxin, cytokinin, ethylene, brassinosteroid, jasmonic acid, and salicylic acid, as well as for hydrogen peroxide, drought and DREB1A overexpression. Totally 622 promoters that are activated by phytohormones were subjected to the prediction. In addition, we have assigned putative functions to 53 octamers of the Regulatory Element Group (REG that have been extracted as position-dependent cis-regulatory elements with the aid of their feature of preferential appearance in the promoter region. Conclusions Our prediction of Arabidopsis cis-regulatory elements for phytohormone responses provides guidance for experimental analysis of promoters to reveal the basis of the transcriptional network of phytohormone responses.

Exposure of maternal mice to cis-bifenthrin enantioselectively disrupts the transcription of genes related to testosterone synthesis in male offspring.

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Jin, Yuanxiang; Wang, Jiangcong; Sun, Xueqing; Ye, Yang; Xu, Minjie; Wang, Jianai; Chen, Shaoping; Fu, Zhengwei

2013-12-01

The commercial bifenthrin (BF) contains two cis isomers. In the present study, a dose of 15mg/kg of 1R-cis-BF or 1S-cis-BF was orally administered for 3 weeks to female mice before or during pregnancy. Then, the expression of steroidogenesis related genes which were considered as effective biomarkers of endocrine disruption were analyzed in the male offspring. Maternal exposure to 1S-cis-BF during pregnancy significantly reduced the mRNA levels of peripheral benzodiazepine receptor (PBR) and steroidogenic acute regulatory protein (StAR) in the testes of 3- or 6-week old male offspring. In addition, a significant decrease of cytochrome P450 17α-hydroxysteroid dehydrogenase (P450-17α) was also observed in the testes of 6-week old male offspring when dams were treated with 1S-cis-BF during pregnancy but not before pregnancy. Moreover, the scavenger receptor class B type 1 (SRB1) and cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc) decreased significantly in the testes of 6-week old male offspring when dams were treated with 1S-cis-BF during and before pregnancy. Thus, oral administration of the maternal mice to cis-BF for 3 weeks, particularly during pregnancy, resulted in endocrine disruption in the male offspring, with the 1S-cis-BF causing more significant alterations than the 1R-cis-BF form. Copyright © 2013 Elsevier Inc. All rights reserved.

Identification of choriogenin cis-regulatory elements and production of estrogen-inducible, liver-specific transgenic Medaka.

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Ueno, Tetsuro; Yasumasu, Shigeki; Hayashi, Shinji; Iuchi, Ichiro

2004-07-01

Choriogenins (chg-H, chg-L) are precursor proteins of egg envelope of medaka and synthesized in the spawning female liver in response to estrogen. We linked a gene construct chg-L1.5 kb/GFP (a 1.5 kb 5'-upstream region of the chg-L gene fused with a green fluorescence protein (GFP) gene) to another construct emgb/RFP (a cis-regulatory region of embryonic globin gene fused with an RFP gene), injected the double fusion gene construct into 1- or 2-cell-stage embryos, and selected embryos expressing the RFP in erythroid cells. From the embryos, we established two lines of chg-L1.5 kb/GFP-emgb/RFP-transgenic medaka. The 3-month-old spawning females and estradiol-17beta (E2)-exposed males displayed the liver-specific GFP expression. The E2-dependent GFP expression was detected in the differentiating liver of the stage 37-38 embryos. In addition, RT-PCR and whole-mount in situ hybridization showed that the E2-dependent chg expression was found in the liver of the stage 34 embryos of wild medaka, suggesting that such E2-dependency is achieved shortly after differentiation of the liver. Analysis using serial deletion mutants fused with GFP showed that the region -426 to -284 of the chg-L gene or the region -364 to -265 of the chg-H gene had the ability to promote the E2-dependent liver-specific GFP expression of its downstream gene. Further analyses suggested that an estrogen response element (ERE) at -309, an ERE half-site at -330 and a binding site for C/EBP at -363 of the chg-L gene played important roles in its downstream chg-L gene expression. In addition, this transgenic medaka may be useful as one of the test animals for detecting environmental estrogenic steroids.

Outcome of high-risk stage 3 neuroblastoma with myeloablative therapy and 13-cis-retinoic acid: a report from the Children's Oncology Group.

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Park, Julie R; Villablanca, Judith G; London, Wendy B; Gerbing, Robert B; Haas-Kogan, Daphne; Adkins, E Stanton; Attiyeh, Edward F; Maris, John M; Seeger, Robert C; Reynolds, C Patrick; Matthay, Katherine K

2009-01-01

The components of therapy required for patients with INSS Stage 3 neuroblastoma and high-risk features remain controversial. A retrospective cohort design was used to determine if intensive chemoradiotherapy with purged autologous bone marrow rescue (ABMT) and/or 13-cis-retinoic acid (13-cis-RA) improved outcome for patients with high-risk neuroblastoma that was not metastatic to distant sites. We identified 72 patients with INSS Stage 3 neuroblastoma enrolled between 1991 and 1996 on the Phase 3 CCG-3891 randomized trial. Patients were analyzed on an intent-to-treat basis using a log-rank test. The 5-year event-free survival (EFS) and overall survival (OS) rates for patients with Stage 3 neuroblastoma were 55 +/- 6% and 59 +/- 6%, respectively (n = 72). Patients randomized to ABMT (n = 20) had 5-year EFS of 65 +/- 11% and OS of 65 +/- 11% compared to 41 +/- 11 (P = 0.21) and 46 +/- 11% (P = 0.23) for patients randomized to CC (n = 23), respectively. Patients randomized to 13-cis-RA (n = 23) had 5-year EFS of 70 +/- 10% and OS of 78 +/- 9% compared to 63 +/- 12% (P = 0.67) and 67 +/- 12% (P = 0.55) for those receiving no further therapy (n = 16), respectively. Patients randomized to both ABMT and 13-cis-RA (n = 6) had a 5-year EFS of 80 +/- 11% and OS of 100%. Patients with high-risk Stage 3 neuroblastoma have an overall poor prognosis despite aggressive chemoradiotherapy. Further studies are warranted to determine if myeloablative consolidation followed by 13-cis-RA maintenance therapy statistically significantly improves outcome.

Functional Conservation of the Glide/Gcm Regulatory Network Controlling Glia, Hemocyte, and Tendon Cell Differentiation in Drosophila

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Cattenoz, Pierre B.; Popkova, Anna; Southall, Tony D.; Aiello, Giuseppe; Brand, Andrea H.; Giangrande, Angela

2016-01-01

High-throughput screens allow us to understand how transcription factors trigger developmental processes, including cell specification. A major challenge is identification of their binding sites because feedback loops and homeostatic interactions may mask the direct impact of those factors in transcriptome analyses. Moreover, this approach dissects the downstream signaling cascades and facilitates identification of conserved transcriptional programs. Here we show the results and the validation of a DNA adenine methyltransferase identification (DamID) genome-wide screen that identifies the direct targets of Glide/Gcm, a potent transcription factor that controls glia, hemocyte, and tendon cell differentiation in Drosophila. The screen identifies many genes that had not been previously associated with Glide/Gcm and highlights three major signaling pathways interacting with Glide/Gcm: Notch, Hedgehog, and JAK/STAT, which all involve feedback loops. Furthermore, the screen identifies effector molecules that are necessary for cell-cell interactions during late developmental processes and/or in ontogeny. Typically, immunoglobulin (Ig) domain–containing proteins control cell adhesion and axonal navigation. This shows that early and transiently expressed fate determinants not only control other transcription factors that, in turn, implement a specific developmental program but also directly affect late developmental events and cell function. Finally, while the mammalian genome contains two orthologous Gcm genes, their function has been demonstrated in vertebrate-specific tissues, placenta, and parathyroid glands, begging questions on the evolutionary conservation of the Gcm cascade in higher organisms. Here we provide the first evidence for the conservation of Gcm direct targets in humans. In sum, this work uncovers novel aspects of cell specification and sets the basis for further understanding of the role of conserved Gcm gene regulatory cascades. PMID:26567182

A novel method for predicting activity of cis-regulatory modules, based on a diverse training set.

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Yang, Wei; Sinha, Saurabh

2017-01-01

With the rapid emergence of technologies for locating cis-regulatory modules (CRMs) genome-wide, the next pressing challenge is to assign precise functions to each CRM, i.e. to determine the spatiotemporal domains or cell-types where it drives expression. A popular approach to this task is to model the typical k-mer composition of a set of CRMs known to drive a common expression pattern, and assign that pattern to other CRMs exhibiting a similar k-mer composition. This approach does not rely on prior knowledge of transcription factors relevant to the CRM or their binding motifs, and is thus more widely applicable than motif-based methods for predicting CRM activity, but is also prone to false positive predictions. We present a novel strategy to improve the above-mentioned approach: to predict if a CRM drives a specific gene expression pattern, assess not only how similar the CRM is to other CRMs with similar activity but also to CRMs with distinct activities. We use a state-of-the-art statistical method to quantify a CRM's sequence similarity to many different training sets of CRMs, and employ a classification algorithm to integrate these similarity scores into a single prediction of the CRM's activity. This strategy is shown to significantly improve CRM activity prediction over current approaches. Our implementation of the new method, called IMMBoost, is freely available as source code, at https://github.com/weiyangedward/IMMBoost CONTACT: sinhas@illinois.eduSupplementary information: Supplementary data are available at Bioinformatics online. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Identifying cis-mediators for trans-eQTLs across many human tissues using genomic mediation analysis.

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Yang, Fan; Wang, Jiebiao; Pierce, Brandon L; Chen, Lin S

2017-11-01

The impact of inherited genetic variation on gene expression in humans is well-established. The majority of known expression quantitative trait loci (eQTLs) impact expression of local genes ( cis -eQTLs). More research is needed to identify effects of genetic variation on distant genes ( trans -eQTLs) and understand their biological mechanisms. One common trans -eQTLs mechanism is "mediation" by a local ( cis ) transcript. Thus, mediation analysis can be applied to genome-wide SNP and expression data in order to identify transcripts that are " cis -mediators" of trans -eQTLs, including those " cis -hubs" involved in regulation of many trans -genes. Identifying such mediators helps us understand regulatory networks and suggests biological mechanisms underlying trans -eQTLs, both of which are relevant for understanding susceptibility to complex diseases. The multitissue expression data from the Genotype-Tissue Expression (GTEx) program provides a unique opportunity to study cis -mediation across human tissue types. However, the presence of complex hidden confounding effects in biological systems can make mediation analyses challenging and prone to confounding bias, particularly when conducted among diverse samples. To address this problem, we propose a new method: Genomic Mediation analysis with Adaptive Confounding adjustment (GMAC). It enables the search of a very large pool of variables, and adaptively selects potential confounding variables for each mediation test. Analyses of simulated data and GTEx data demonstrate that the adaptive selection of confounders by GMAC improves the power and precision of mediation analysis. Application of GMAC to GTEx data provides new insights into the observed patterns of cis -hubs and trans -eQTL regulation across tissue types. © 2017 Yang et al.; Published by Cold Spring Harbor Laboratory Press.

cis sequence effects on gene expression

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Jacobs Kevin

2007-08-01

Full Text Available Abstract Background Sequence and transcriptional variability within and between individuals are typically studied independently. The joint analysis of sequence and gene expression variation (genetical genomics provides insight into the role of linked sequence variation in the regulation of gene expression. We investigated the role of sequence variation in cis on gene expression (cis sequence effects in a group of genes commonly studied in cancer research in lymphoblastoid cell lines. We estimated the proportion of genes exhibiting cis sequence effects and the proportion of gene expression variation explained by cis sequence effects using three different analytical approaches, and compared our results to the literature. Results We generated gene expression profiling data at N = 697 candidate genes from N = 30 lymphoblastoid cell lines for this study and used available candidate gene resequencing data at N = 552 candidate genes to identify N = 30 candidate genes with sufficient variance in both datasets for the investigation of cis sequence effects. We used two additive models and the haplotype phylogeny scanning approach of Templeton (Tree Scanning to evaluate association between individual SNPs, all SNPs at a gene, and diplotypes, with log-transformed gene expression. SNPs and diplotypes at eight candidate genes exhibited statistically significant (p cis sequence effects in our study, respectively. Conclusion Based on analysis of our results and the extant literature, one in four genes exhibits significant cis sequence effects, and for these genes, about 30% of gene expression variation is accounted for by cis sequence variation. Despite diverse experimental approaches, the presence or absence of significant cis sequence effects is largely supported by previously published studies.

Conserved host response to highly pathogenic avian influenza virus infection in human cell culture, mouse and macaque model systems

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McDermott Jason E

2011-11-01

Full Text Available Abstract Background Understanding host response to influenza virus infection will facilitate development of better diagnoses and therapeutic interventions. Several different experimental models have been used as a proxy for human infection, including cell cultures derived from human cells, mice, and non-human primates. Each of these systems has been studied extensively in isolation, but little effort has been directed toward systematically characterizing the conservation of host response on a global level beyond known immune signaling cascades. Results In the present study, we employed a multivariate modeling approach to characterize and compare the transcriptional regulatory networks between these three model systems after infection with a highly pathogenic avian influenza virus of the H5N1 subtype. Using this approach we identified functions and pathways that display similar behavior and/or regulation including the well-studied impact on the interferon response and the inflammasome. Our results also suggest a primary response role for airway epithelial cells in initiating hypercytokinemia, which is thought to contribute to the pathogenesis of H5N1 viruses. We further demonstrate that we can use a transcriptional regulatory model from the human cell culture data to make highly accurate predictions about the behavior of important components of the innate immune system in tissues from whole organisms. Conclusions This is the first demonstration of a global regulatory network modeling conserved host response between in vitro and in vivo models.

Finding cis-regulatory modules in Drosophila using phylogenetic hidden Markov models

DEFF Research Database (Denmark)

Wong, Wendy S W; Nielsen, Rasmus

2007-01-01

MOTIVATION: Finding the regulatory modules for transcription factors binding is an important step in elucidating the complex molecular mechanisms underlying regulation of gene expression. There are numerous methods available for solving this problem, however, very few of them take advantage of th...

Biodiversity conservation in a changing climate: a review of threats and implications for conservation planning in Myanmar.

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Rao, Madhu; Saw Htun; Platt, Steven G; Tizard, Robert; Poole, Colin; Than Myint; Watson, James E M

2013-11-01

High levels of species richness and endemism make Myanmar a regional priority for conservation. However, decades of economic and political sanctions have resulted in low conservation investment to effectively tackle threats to biodiversity. Recent sweeping political reforms have placed Myanmar on the fast track to economic development-the expectation is increased economic investments focused on the exploitation of the country's rich, and relatively intact, natural resources. Within a context of weak regulatory capacity and inadequate environmental safeguards, rapid economic development is likely to have far-reaching negative implications for already threatened biodiversity and natural-resource-dependent human communities. Climate change will further exacerbate prevailing threats given Myanmar's high exposure and vulnerability. The aim of this review is to examine the implications of increased economic growth and a changing climate within the larger context of biodiversity conservation in Myanmar. We summarize conservation challenges, assess direct climatological impacts on biodiversity and conclude with recommendations for long-term adaptation approaches for biodiversity conservation.

The efficacy of 9-cis retinoic acid in experimental models of cancer.

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Gottardis, M M; Lamph, W W; Shalinsky, D R; Wellstein, A; Heyman, R A

1996-01-01

9-cis retinoic acid (9-cis RA) is a retinoid receptor pan-agonist that binds with high affinity to both retinoic acid receptors (RARs) and retinoid X receptors (RXRs). Using a variety of in vivo and in vitro cancer models, we present experimental data that 9-cis RA has activity as a potential chemotherapeutic agent. Treatment of the human promyelocytic leukemia cell line HL-60 with 9-cis RA decreases cell proliferation, increases cell differentiation, and increases apoptosis. Induction of apoptosis correlates with an increase in tissue transglutaminase (type II) activity. In vivo, 9-cis RA induces complete tumor regression of an early passage human lip squamous cell carcinoma xenograft. Finally, 9-cis RA inhibits the anchorage-independent growth of the human breast cancer cell lines MCF-7 and LY2 (an antiestrogen-resistant MCF-7 variant). Transient co-transfection assays indicate that 9-cis RA inhibits estrogen receptor transcription of an ERE-tk-LUC reporter through RAR or RXR receptors. These data suggest that retinoid receptors can antagonize estrogen-dependent transcription and provides one possible mechanism for the inhibition of cell growth by 9-cis RA in breast cancer cell lines. In summary, these findings present evidence that 9-cis RA has a wide range of activities in human cancer models.

REDfly: a Regulatory Element Database for Drosophila.

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Gallo, Steven M; Li, Long; Hu, Zihua; Halfon, Marc S

2006-02-01

Bioinformatics studies of transcriptional regulation in the metazoa are significantly hindered by the absence of readily available data on large numbers of transcriptional cis-regulatory modules (CRMs). Even the richly annotated Drosophila melanogaster genome lacks extensive CRM information. We therefore present here a database of Drosophila CRMs curated from the literature complete with both DNA sequence and a searchable description of the gene expression pattern regulated by each CRM. This resource should greatly facilitate the development of computational approaches to CRM discovery as well as bioinformatics analyses of regulatory sequence properties and evolution.

Cluster observations of the high-latitude magnetopause and cusp: initial results from the CIS ion instruments

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J. M. Bosqued

2001-09-01

Full Text Available Launched on an elliptical high inclination orbit (apogee: 19.6 RE since January 2001 the Cluster satellites have been conducting the first detailed three-dimensional studies of the high-latitude dayside magnetosphere, including the exterior cusp, neighbouring boundary layers and magnetopause regions. Cluster satellites carry the CIS ion spectrometers that provide high-precision, 3D distributions of low-energy (<35 keV/e ions every 4 s. This paper presents the first two observations of the cusp and/or magnetopause behaviour made under different interplanetary magnetic field (IMF conditions. Flow directions, 3D distribution functions, density profiles and ion composition profiles are analyzed to demonstrate the high variability of high-latitude regions. In the first crossing analyzed (26 January 2001, dusk side, IMF-BZ < 0, multiple, isolated boundary layer, magnetopause and magnetosheath encounters clearly occurred on a quasi-steady basis for ~ 2 hours. CIS ion instruments show systematic accelerated flows in the current layer and adjacent boundary layers on the Earthward side of the magnetopause. Multi-point analysis of the magnetopause, combining magnetic and plasma data from the four Cluster spacecraft, demonstrates that oscillatory outward-inward motions occur with a normal speed of the order of ± 40 km/s; the thickness of the high-latitude current layer is evaluated to be of the order of 900–1000 km. Alfvénic accelerated flows and D-shaped distributions are convincing signatures of a magnetic reconnection occurring equatorward of the Cluster satellites. Moreover, the internal magnetic and plasma structure of a flux transfer event (FTE is analyzed in detail; its size along the magnetopause surface is ~ 12 000 km and it convects with a velocity of ~ 200 km/s. The second event analyzed (2 February 2001 corresponds to the first Cluster pass within the cusp when the IMF-BZ component was northward directed. The analysis of relevant CIS plasma

Characterization of the Promoter Region of an Arabidopsis Gene for 9-cis-Epoxycarotenoid Dioxygenase Involved in Dehydration-Inducible Transcription

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Behnam, Babak; Iuchi, Satoshi; Fujita, Miki; Fujita, Yasunari; Takasaki, Hironori; Osakabe, Yuriko; Yamaguchi-Shinozaki, Kazuko; Kobayashi, Masatomo; Shinozaki, Kazuo

2013-01-01

Plants respond to dehydration stress and tolerate water-deficit status through complex physiological and cellular processes. Many genes are induced by water deficit. Abscisic acid (ABA) plays important roles in tolerance to dehydration stress by inducing many stress genes. ABA is synthesized de novo in response to dehydration. Most of the genes involved in ABA biosynthesis have been identified, and they are expressed mainly in leaf vascular tissues. Of the products of such genes, 9-cis-epoxycarotenoid dioxygenase (NCED) is a key enzyme in ABA biosynthesis. One of the five NCED genes in Arabidopsis, AtNCED3, is significantly induced by dehydration. To understand the regulatory mechanism of the early stages of the dehydration stress response, it is important to analyse the transcriptional regulatory systems of AtNCED3. In the present study, we found that an overlapping G-box recognition sequence (5′-CACGTG-3′) at −2248 bp from the transcriptional start site of AtNCED3 is an important cis-acting element in the induction of the dehydration response. We discuss the possible transcriptional regulatory system of dehydration-responsive AtNCED3 expression, and how this may control the level of ABA under water-deficit conditions. PMID:23604098

Combinatorial binding in human and mouse embryonic stem cells identifies conserved enhancers active in early embryonic development.

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Jonathan Göke

2011-12-01

Full Text Available Transcription factors are proteins that regulate gene expression by binding to cis-regulatory sequences such as promoters and enhancers. In embryonic stem (ES cells, binding of the transcription factors OCT4, SOX2 and NANOG is essential to maintain the capacity of the cells to differentiate into any cell type of the developing embryo. It is known that transcription factors interact to regulate gene expression. In this study we show that combinatorial binding is strongly associated with co-localization of the transcriptional co-activator Mediator, H3K27ac and increased expression of nearby genes in embryonic stem cells. We observe that the same loci bound by Oct4, Nanog and Sox2 in ES cells frequently drive expression in early embryonic development. Comparison of mouse and human ES cells shows that less than 5% of individual binding events for OCT4, SOX2 and NANOG are shared between species. In contrast, about 15% of combinatorial binding events and even between 53% and 63% of combinatorial binding events at enhancers active in early development are conserved. Our analysis suggests that the combination of OCT4, SOX2 and NANOG binding is critical for transcription in ES cells and likely plays an important role for embryogenesis by binding at conserved early developmental enhancers. Our data suggests that the fast evolutionary rewiring of regulatory networks mainly affects individual binding events, whereas "gene regulatory hotspots" which are bound by multiple factors and active in multiple tissues throughout early development are under stronger evolutionary constraints.

Water Well Locations - Conservation Wells

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NSGIC Education | GIS Inventory — The conservation well layer identifies the permitted surface location of oil and gas conservation wells that have not been plugged. These include active, regulatory...

Comprehensive meta-analysis of Signal Transducers and Activators of Transcription (STAT genomic binding patterns discerns cell-specific cis-regulatory modules

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Kang Keunsoo

2013-01-01

Full Text Available Abstract Background Cytokine-activated transcription factors from the STAT (Signal Transducers and Activators of Transcription family control common and context-specific genetic programs. It is not clear to what extent cell-specific features determine the binding capacity of seven STAT members and to what degree they share genetic targets. Molecular insight into the biology of STATs was gained from a meta-analysis of 29 available ChIP-seq data sets covering genome-wide occupancy of STATs 1, 3, 4, 5A, 5B and 6 in several cell types. Results We determined that the genomic binding capacity of STATs is primarily defined by the cell type and to a lesser extent by individual family members. For example, the overlap of shared binding sites between STATs 3 and 5 in T cells is greater than that between STAT5 in T cells and non-T cells. Even for the top 1,000 highly enriched STAT binding sites, ~15% of STAT5 binding sites in mouse female liver are shared by other STATs in different cell types while in T cells ~90% of STAT5 binding sites are co-occupied by STAT3, STAT4 and STAT6. In addition, we identified 116 cis-regulatory modules (CRM, which are recognized by all STAT members across cell types defining a common JAK-STAT signature. Lastly, in liver STAT5 binding significantly coincides with binding of the cell-specific transcription factors HNF4A, FOXA1 and FOXA2 and is associated with cell-type specific gene transcription. Conclusions Our results suggest that genomic binding of STATs is primarily determined by the cell type and further specificity is achieved in part by juxtaposed binding of cell-specific transcription factors.

RNA regulatory elements and polyadenylation in plants

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Arthur G. Hunt

2012-01-01

Full Text Available Alternative poly(A site choice (also known as alternative polyadenylation, or APA has the potential to affect gene expression in qualitative and quantitative ways. Alternative polyadenylation may affect as many as 82% of all expressed genes in a plant. The consequences of APA include the generation of transcripts with differing 3’-UTRs (and thus differing potential regulatory potential and of transcripts with differing protein-coding potential. Genome-wide studies of possible APA suggest a linkage with pre-mRNA splicing, and indicate a coincidence of and perhaps cooperation between RNA regulatory elements that affect splicing efficiency and the recognition of novel intronic poly(A sites. These studies also raise the possibility of the existence of a novel class of polyadenylation-related cis elements that are distinct from the well-characterized plant polyadenylation signal. Many potential APA events, however, have not been associated with identifiable cis elements. The present state of the field reveals a broad scope of APA, and also numerous opportunities for research into mechanisms that govern both choice and regulation of poly(A sites in plants.

The Community Intercomparison Suite (CIS)

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Watson-Parris, Duncan; Schutgens, Nick; Cook, Nick; Kipling, Zak; Kershaw, Phil; Gryspeerdt, Ed; Lawrence, Bryan; Stier, Philip

2017-04-01

Earth observations (both remote and in-situ) create vast amounts of data providing invaluable constraints for the climate science community. Efficient exploitation of these complex and highly heterogeneous datasets has been limited however by the lack of suitable software tools, particularly for comparison of gridded and ungridded data, thus reducing scientific productivity. CIS (http://cistools.net) is an open-source, command line tool and Python library which allows the straight-forward quantitative analysis, intercomparison and visualisation of remote sensing, in-situ and model data. The CIS can read gridded and ungridded remote sensing, in-situ and model data - and many other data sources 'out-of-the-box', such as ESA Aerosol and Cloud CCI product, MODIS, Cloud CCI, Cloudsat, AERONET. Perhaps most importantly however CIS also employs a modular plugin architecture to allow for the reading of limitless different data types. Users are able to write their own plugins for reading the data sources which they are familiar with, and share them within the community, allowing all to benefit from their expertise. To enable the intercomparison of this data the CIS provides a number of operations including: the aggregation of ungridded and gridded datasets to coarser representations using a number of different built in averaging kernels; the subsetting of data to reduce its extent or dimensionality; the co-location of two distinct datasets onto a single set of co-ordinates; the visualisation of the input or output data through a number of different plots and graphs; the evaluation of arbitrary mathematical expressions against any number of datasets; and a number of other supporting functions such as a statistical comparison of two co-located datasets. These operations can be performed efficiently on local machines or large computing clusters - and is already available on the JASMIN computing facility. A case-study using the GASSP collection of in-situ aerosol observations

Piecing together cis-regulatory networks: insights from epigenomics studies in plants.

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Huang, Shao-Shan C; Ecker, Joseph R

2018-05-01

5-Methylcytosine, a chemical modification of DNA, is a covalent modification found in the genomes of both plants and animals. Epigenetic inheritance of phenotypes mediated by DNA methylation is well established in plants. Most of the known mechanisms of establishing, maintaining and modifying DNA methylation have been worked out in the reference plant Arabidopsis thaliana. Major functions of DNA methylation in plants include regulation of gene expression and silencing of transposable elements (TEs) and repetitive sequences, both of which have parallels in mammalian biology, involve interaction with the transcriptional machinery, and may have profound effects on the regulatory networks in the cell. Methylome and transcriptome dynamics have been investigated in development and environmental responses in Arabidopsis and agriculturally and ecologically important plants, revealing the interdependent relationship among genomic context, methylation patterns, and expression of TE and protein coding genes. Analyses of methylome variation among plant natural populations and species have begun to quantify the extent of genetic control of methylome variation vs. true epimutation, and model the evolutionary forces driving methylome evolution in both short and long time scales. The ability of DNA methylation to positively or negatively modulate binding affinity of transcription factors (TFs) provides a natural link from genome sequence and methylation changes to transcription. Technologies that allow systematic determination of methylation sensitivities of TFs, in native genomic and methylation context without confounding factors such as histone modifications, will provide baseline datasets for building cell-type- and individual-specific regulatory networks that underlie the establishment and inheritance of complex traits. This article is categorized under: Laboratory Methods and Technologies > Genetic/Genomic Methods Biological Mechanisms > Regulatory Biology. © 2017 Wiley

Evaluating cis-2,6-Dimethylpiperidide (cis-DMP) as a Base Component in Lithium-Mediated Zincation Chemistry

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Armstrong, David R; Garden, Jennifer A; Kennedy, Alan R; Leenhouts, Sarah M; Mulvey, Robert E; O'Keefe, Philip; O'Hara, Charles T; Steven, Alan

2013-01-01

Most recent advances in metallation chemistry have centred on the bulky secondary amide 2,2,6,6-tetramethylpiperidide (TMP) within mixed metal, often ate, compositions. However, the precursor amine TMP(H) is rather expensive so a cheaper substitute would be welcome. Thus this study was aimed towards developing cheaper non-TMP based mixed-metal bases and, as cis-2,6-dimethylpiperidide (cis-DMP) was chosen as the alternative amide, developing cis-DMP zincate chemistry which has received meagre attention compared to that of its methyl-rich counterpart TMP. A new lithium diethylzincate, [(TMEDA)LiZn(cis-DMP)Et2] (TMEDA=N,N,N′,N′-tetramethylethylenediamine) has been synthesised by co-complexation of Li(cis-DMP), Et2Zn and TMEDA, and characterised by NMR (including DOSY) spectroscopy and X-ray crystallography, which revealed a dinuclear contact ion pair arrangement. By using N,N-diisopropylbenzamide as a test aromatic substrate, the deprotonative reactivity of [(TMEDA)LiZn(cis-DMP)Et2] has been probed and contrasted with that of the known but previously uninvestigated di-tert-butylzincate, [(TMEDA)LiZn(cis-DMP)tBu2]. The former was found to be the superior base (for example, producing the ortho-deuteriated product in respective yields of 78 % and 48 % following D2O quenching of zincated benzamide intermediates). An 88 % yield of 2-iodo-N,N-diisopropylbenzamide was obtained on reaction of two equivalents of the diethylzincate with the benzamide followed by iodination. Comparisons are also drawn using 1,1,1,3,3,3-hexamethyldisilazide (HMDS), diisopropylamide and TMP as the amide component in the lithium amide, Et2Zn and TMEDA system. Under certain conditions, the cis-DMP base system was found to give improved results in comparison to HMDS and diisopropylamide (DA), and comparable results to a TMP system. Two novel complexes isolated from reactions of the di-tert-butylzincate and crystallographically characterised, namely the pre-metallation complex [{(iPr)2N(Ph)C=O}LiZn(cis

Outcome of High-Risk Stage 3 Neuroblastoma with Myeloablative Therapy and 13-cis-Retinoic Acid: A Report from the Children’s Oncology Group

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Park, Julie R.; Villablanca, Judith G.; London, Wendy B.; Gerbing, Robert B.; Haas-Kogan, Daphne; Stanton Adkins, E.; Attiyeh, Edward F.; Maris, John M.; Seeger, Robert C.; Patrick Reynolds, C.; Matthay, Katherine K.

2009-01-01

Background The components of therapy required for patients with INSS Stage 3 neuroblastoma and high risk features remain controversial. Procedure A retrospective cohort design was used to determine if intensive chemoradiotherapy with purged autologous bone marrow rescue (ABMT) and/or 13-cis-retinoic acid (13-cis-RA) improved outcome for patients with high-risk neuroblastoma that was not metastatic to distant sites. We identified 72 patients with INSS Stage 3 neuroblastoma enrolled between 1991 and 1996 on the Phase III CCG 3891 randomized trial. Patients were analyzed on an intent-to-treat basis using a log-rank test. Results The 5-year event-free survival (EFS) and overall survival (OS) rates for patients with Stage 3 neuroblastoma were 55 +/- 6% and 59% +/- 6%, respectively (n=72). Patients randomized to ABMT (n=20) had 5-year EFS of 65% +/- 11% and OS of 65% +/- 11% compared to 41% +/- 11 (p=0.21) and 46% +/- 11% (p=0.23) for patients randomized to CC (n=23), respectively. Patients randomized to 13-cis-RA (n=23) had 5-year EFS of 70% +/- 10% and OS of 78% +/- 9% compared to 63% +/- 12% (p=0.67) and 67% +/- 12% (p=0.55) for those receiving no further therapy (n=16), respectively. Patients randomized to both ABMT and 13-cis-RA (n=6) had a 5-year EFS of 80% ± 11% and OS of 100%. Conclusion Patients with high-risk Stage 3 neuroblastoma have an overall poor prognosis despite aggressive chemoradiotherapy. Further studies are warranted to determine if myeloablative consolidation followed by 13-cis-RA maintenance therapy statistically significantly improves outcome. PMID:18937318

Phylogenetic conservation of the regulatory and functional properties of the Vav oncoprotein family

International Nuclear Information System (INIS)

Couceiro, Jose R.; Martin-Bermudo, Maria D.; Bustelo, Xose R.

2005-01-01

Vav proteins are phosphorylation-dependent GDP/GTP exchange factors for Rho/Rac GTPases. Despite intense characterization of mammalian Vav proteins both biochemically and genetically, there is little information regarding the conservation of their biological properties in lower organisms. To approach this issue, we have performed a characterization of the regulatory, catalytic, and functional properties of the single Vav family member of Drosophila melanogaster. These analyses have shown that the intramolecular mechanisms controlling the enzyme activity of mammalian Vav proteins are already present in Drosophila, suggesting that such properties have been set up before the divergence between protostomes and deuterostomes during evolution. We also show that Drosophila and mammalian Vav proteins have similar catalytic specificities. As a consequence, Drosophila Vav can trigger oncogenic transformation, morphological change, and enhanced cell motility in mammalian cells. Gain-of-function studies using transgenic flies support the implication of this protein in cytoskeletal-dependent processes such as embryonic dorsal closure, myoblast fusion, tracheal development, and the migration/guidance of different cell types. These results highlight the important roles of Vav proteins in the signal transduction pathways regulating cytoskeletal dynamics. Moreover, they indicate that the foundations for the regulatory and enzymatic activities of this protein family have been set up very early during evolution

Pleiotropy constrains the evolution of protein but not regulatory sequences in a transcription regulatory network influencing complex social behaviours

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Daria eMolodtsova

2014-12-01

Full Text Available It is increasingly apparent that genes and networks that influence complex behaviour are evolutionary conserved, which is paradoxical considering that behaviour is labile over evolutionary timescales. How does adaptive change in behaviour arise if behaviour is controlled by conserved, pleiotropic, and likely evolutionary constrained genes? Pleiotropy and connectedness are known to constrain the general rate of protein evolution, prompting some to suggest that the evolution of complex traits, including behaviour, is fuelled by regulatory sequence evolution. However, we seldom have data on the strength of selection on mutations in coding and regulatory sequences, and this hinders our ability to study how pleiotropy influences coding and regulatory sequence evolution. Here we use population genomics to estimate the strength of selection on coding and regulatory mutations for a transcriptional regulatory network that influences complex behaviour of honey bees. We found that replacement mutations in highly connected transcription factors and target genes experience significantly stronger negative selection relative to weakly connected transcription factors and targets. Adaptively evolving proteins were significantly more likely to reside at the periphery of the regulatory network, while proteins with signs of negative selection were near the core of the network. Interestingly, connectedness and network structure had minimal influence on the strength of selection on putative regulatory sequences for both transcription factors and their targets. Our study indicates that adaptive evolution of complex behaviour can arise because of positive selection on protein-coding mutations in peripheral genes, and on regulatory sequence mutations in both transcription factors and their targets throughout the network.

Prioritization of gene regulatory interactions from large-scale modules in yeast

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Bringas Ricardo

2008-01-01

Full Text Available Abstract Background The identification of groups of co-regulated genes and their transcription factors, called transcriptional modules, has been a focus of many studies about biological systems. While methods have been developed to derive numerous modules from genome-wide data, individual links between regulatory proteins and target genes still need experimental verification. In this work, we aim to prioritize regulator-target links within transcriptional modules based on three types of large-scale data sources. Results Starting with putative transcriptional modules from ChIP-chip data, we first derive modules in which target genes show both expression and function coherence. The most reliable regulatory links between transcription factors and target genes are established by identifying intersection of target genes in coherent modules for each enriched functional category. Using a combination of genome-wide yeast data in normal growth conditions and two different reference datasets, we show that our method predicts regulatory interactions with significantly higher predictive power than ChIP-chip binding data alone. A comparison with results from other studies highlights that our approach provides a reliable and complementary set of regulatory interactions. Based on our results, we can also identify functionally interacting target genes, for instance, a group of co-regulated proteins related to cell wall synthesis. Furthermore, we report novel conserved binding sites of a glycoprotein-encoding gene, CIS3, regulated by Swi6-Swi4 and Ndd1-Fkh2-Mcm1 complexes. Conclusion We provide a simple method to prioritize individual TF-gene interactions from large-scale transcriptional modules. In comparison with other published works, we predict a complementary set of regulatory interactions which yields a similar or higher prediction accuracy at the expense of sensitivity. Therefore, our method can serve as an alternative approach to prioritization for

10 points about buying C.I.S

International Nuclear Information System (INIS)

Anon.

1993-01-01

On October 16, 1992, the U.S. Department of Commerce (DOC) settled the antidumping case against the CIS republics by imposing price and volume quotas on CIS uranium imported into the United States. Bound by a suspension agreement, each of the six uranium-producing CIS republics is responsible for restricting the flow of imports to the US-either directly or indirectly. (As the NUKEM Market Report went to press, the Ukraine government notified the DOC of its intent not to terminate the suspension agreement.) This action is to prevent undercutting price levels in the US domestic uranium markets. What follows are ten points about everything you should know about importing uranium from the uranium-producing CIS republics- Kazakhstan, Kyrgyzstan, Russian Federation, Tajikistan, Ukraine and Uzbekistan. Newcomers to the CIS scene should follow this simple roadmap and be aware of the issues they face as importers in terms of Commerce/Customs requirements and documentation and where to get them, when to buy the material and how to transport it, how to deal effectively with CIS exporters, and how to avoid unnecessary complications when buying CIS

Cluster observations of the high-latitude magnetopause and cusp: initial results from the CIS ion instruments

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J. M. Bosqued

Full Text Available Launched on an elliptical high inclination orbit (apogee: 19.6 R_E since January 2001 the Cluster satellites have been conducting the first detailed three-dimensional studies of the high-latitude dayside magnetosphere, including the exterior cusp, neighbouring boundary layers and magnetopause regions. Cluster satellites carry the CIS ion spectrometers that provide high-precision, 3D distributions of low-energy (<35 keV/e ions every 4 s. This paper presents the first two observations of the cusp and/or magnetopause behaviour made under different interplanetary magnetic field (IMF conditions. Flow directions, 3D distribution functions, density profiles and ion composition profiles are analyzed to demonstrate the high variability of high-latitude regions. In the first crossing analyzed (26 January 2001, dusk side, IMF-BZ < 0, multiple, isolated boundary layer, magnetopause and magnetosheath encounters clearly occurred on a quasi-steady basis for ~ 2 hours. CIS ion instruments show systematic accelerated flows in the current layer and adjacent boundary layers on the Earthward side of the magnetopause. Multi-point analysis of the magnetopause, combining magnetic and plasma data from the four Cluster spacecraft, demonstrates that oscillatory outward-inward motions occur with a normal speed of the order of ± 40 km/s; the thickness of the high-latitude current layer is evaluated to be of the order of 900–1000 km. Alfvénic accelerated flows and D-shaped distributions are convincing signatures of a magnetic reconnection occurring equatorward of the Cluster satellites. Moreover, the internal magnetic and plasma structure of a flux transfer event (FTE is analyzed in detail; its size along the magnetopause surface is ~ 12 000 km and it convects with a velocity of ~ 200 km/s. The second event analyzed (2 February 2001 corresponds to the first Cluster pass within the cusp when the IMF-BZ component was northward directed. The analysis of

A Lettuce (Lactuca sativa) Homolog of Human Nogo-B Receptor Interacts with cis-Prenyltransferase and Is Necessary for Natural Rubber Biosynthesis*

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Qu, Yang; Chakrabarty, Romit; Tran, Hue T.; Kwon, Eun-Joo G.; Kwon, Moonhyuk; Nguyen, Trinh-Don; Ro, Dae-Kyun

2015-01-01

Natural rubber (cis-1,4-polyisoprene) is an indispensable biopolymer used to manufacture diverse consumer products. Although a major source of natural rubber is the rubber tree (Hevea brasiliensis), lettuce (Lactuca sativa) is also known to synthesize natural rubber. Here, we report that an unusual cis-prenyltransferase-like 2 (CPTL2) that lacks the conserved motifs of conventional cis-prenyltransferase is required for natural rubber biosynthesis in lettuce. CPTL2, identified from the lettuce rubber particle proteome, displays homology to a human NogoB receptor and is predominantly expressed in latex. Multiple transgenic lettuces expressing CPTL2-RNAi constructs showed that a decrease of CPTL2 transcripts (3–15% CPTL2 expression relative to controls) coincided with the reduction of natural rubber as low as 5%. We also identified a conventional cis-prenyltransferase 3 (CPT3), exclusively expressed in latex. In subcellular localization studies using fluorescent proteins, cytosolic CPT3 was relocalized to endoplasmic reticulum by co-occurrence of CPTL2 in tobacco and yeast at the log phase. Furthermore, yeast two-hybrid data showed that CPTL2 and CPT3 interact. Yeast microsomes containing CPTL2/CPT3 showed enhanced synthesis of short cis-polyisoprenes, but natural rubber could not be synthesized in vitro. Intriguingly, a homologous pair CPTL1/CPT1, which displays ubiquitous expressions in lettuce, showed a potent dolichol biosynthetic activity in vitro. Taken together, our data suggest that CPTL2 is a scaffolding protein that tethers CPT3 on endoplasmic reticulum and is necessary for natural rubber biosynthesis in planta, but yeast-expressed CPTL2 and CPT3 alone could not synthesize high molecular weight natural rubber in vitro. PMID:25477521

cis-Bifenthrin enantioselectively induces hepatic oxidative stress in mice.

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Jin, Yuanxiang; Wang, Jiangcong; Pan, Xiuhong; Wang, Linggang; Fu, Zhengwei

2013-09-01

Bifenthrin (BF), as a chiral synthetic pyrethroid, is widely used to control field and household pests. In China, the commercial cis-BF contained two enantiomers including 1R-cis-BF and 1S-cis-BF. However, the difference in oxidative stress induced by the two enantiomers in mice still remains unclear. In the present study, 4 week-old adolescent male ICR mice were orally administered cis-BF, 1R-cis-BF or 1S-cis-BF daily for 2, 4 and 6 weeks at doses of 5 mg/kg/day, respectively. We found that the hepatic reactive oxygen species (ROS) levels, as well as the malondialdehyde (MDA) and glutathione (GSH) content both in the serum and liver increased significantly in the 4 or 6 weeks 1S-cis-BF treated groups. The activities of superoxide dismutase (SOD) and catalase (CAT) also changed significantly in the serum and liver of 1S-cis-BF treated mice. More importantly, the significant differences in MDA content and CAT activity both in the serum and liver, and the activities of total antioxidant capacity (T-AOC) and SOD in serum were also observed between the 1S-cis-BF and 1R-cis-BF treated groups. Moreover, the transcription of oxidative stress response related genes including Sod1, Cat and heme oxygenase-1(Ho-1) in the liver of 1S-cis-BF treated groups were also significant higher than those in 1R-cis-BF treated group. Thus, it was concluded that cis-BF induced hepatic oxidative stress in an enantiomer specific manner in mice when exposed during the puberty, and that 1S-cis-BF showed much more toxic in hepatic oxidative stress than 1R-cis-BF. Copyright © 2013 Elsevier Inc. All rights reserved.

cisTEM, user-friendly software for single-particle image processing

Science.gov (United States)

2018-01-01

We have developed new open-source software called cisTEM (computational imaging system for transmission electron microscopy) for the processing of data for high-resolution electron cryo-microscopy and single-particle averaging. cisTEM features a graphical user interface that is used to submit jobs, monitor their progress, and display results. It implements a full processing pipeline including movie processing, image defocus determination, automatic particle picking, 2D classification, ab-initio 3D map generation from random parameters, 3D classification, and high-resolution refinement and reconstruction. Some of these steps implement newly-developed algorithms; others were adapted from previously published algorithms. The software is optimized to enable processing of typical datasets (2000 micrographs, 200 k – 300 k particles) on a high-end, CPU-based workstation in half a day or less, comparable to GPU-accelerated processing. Jobs can also be scheduled on large computer clusters using flexible run profiles that can be adapted for most computing environments. cisTEM is available for download from cistem.org. PMID:29513216

cisTEM, user-friendly software for single-particle image processing.

Science.gov (United States)

Grant, Timothy; Rohou, Alexis; Grigorieff, Nikolaus

2018-03-07

We have developed new open-source software called cis TEM (computational imaging system for transmission electron microscopy) for the processing of data for high-resolution electron cryo-microscopy and single-particle averaging. cis TEM features a graphical user interface that is used to submit jobs, monitor their progress, and display results. It implements a full processing pipeline including movie processing, image defocus determination, automatic particle picking, 2D classification, ab-initio 3D map generation from random parameters, 3D classification, and high-resolution refinement and reconstruction. Some of these steps implement newly-developed algorithms; others were adapted from previously published algorithms. The software is optimized to enable processing of typical datasets (2000 micrographs, 200 k - 300 k particles) on a high-end, CPU-based workstation in half a day or less, comparable to GPU-accelerated processing. Jobs can also be scheduled on large computer clusters using flexible run profiles that can be adapted for most computing environments. cis TEM is available for download from cistem.org. © 2018, Grant et al.

Identification and Characterization of 5′ Untranslated Regions (5′UTRs in Zymomonas mobilis as Regulatory Biological Parts

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Seung Hee Cho

2017-12-01

Full Text Available Regulatory RNA regions within a transcript, particularly in the 5′ untranslated region (5′UTR, have been shown in a variety of organisms to control the expression levels of these mRNAs in response to various metabolites or environmental conditions. Considering the unique tolerance of Zymomonas mobilis to ethanol and the growing interest in engineering microbial strains with enhanced tolerance to industrial inhibitors, we searched natural cis-regulatory regions in this microorganism using transcriptomic data and bioinformatics analysis. Potential regulatory 5′UTRs were identified and filtered based on length, gene function, relative gene counts, and conservation in other organisms. An in vivo fluorescence-based screening system was developed to confirm the responsiveness of 36 5′UTR candidates to ethanol, acetate, and xylose stresses. UTR_ZMO0347 (5′UTR of gene ZMO0347 encoding the RNA binding protein Hfq was found to down-regulate downstream gene expression under ethanol stress. Genomic deletion of UTR_ZMO0347 led to a general decrease of hfq expression at the transcript level and increased sensitivity for observed changes in Hfq expression at the protein level. The role of UTR_ZMO0347 and other 5′UTRs gives us insight into the regulatory network of Z. mobilis in response to stress and unlocks new strategies for engineering robust industrial strains as well as for harvesting novel responsive regulatory biological parts for controllable gene expression platforms in this organism.

Synthesis of racemic [methyl-d3]-labeled cis- and trans-3'-hydroxycotinine

International Nuclear Information System (INIS)

Ravard, A.; Crooks, P.A.

1994-01-01

A method is described for the synthesis of the racemic [methyl-d 3 ] forms of the nicotine metabolites cis-3'-hydroxycotinine and trans-3'-hydroxycotinine. The key intermediate was [methyl-d 3 ]-N-methylhydroxylamine, obtained from a selective hydrogenation of d 3 -nitro-methane. This intermediate was converted to [methyl-d 3 ]-α-3-pyridyl-N-methylnitrone, which was condensed with methyl acrylate to give a mixture of isomeric isoxazolidines. The hydrogenolysis of this mixture afforded a 70:30 mixture of [methyl-d 3 ] cis- and trans-3'-hydroxycotinine, from which the pure cis-isomer could be isolated by recrystallization from acetone. [Methyl-d 3 ]-trans-3'-hydroxycotinine could be prepared in high yield from the cis-isomer via chiral inversion utilizing a Mitsunobu reaction, or by chromatographic separation from a mixture of the cis- and trans-3'-benzoyloxycotinine, followed by O-debenzoylation in methanolic NaOH. (author)

The Role of Cis-Lunar Space in Future Global Space Exploration

Science.gov (United States)

Bobskill, Marianne R.; Lupisella, Mark L.

2012-01-01

Cis-lunar space offers affordable near-term opportunities to help pave the way for future global human exploration of deep space, acting as a bridge between present missions and future deep space missions. While missions in cis-lunar space have value unto themselves, they can also play an important role in enabling and reducing risk for future human missions to the Moon, Near-Earth Asteroids (NEAs), Mars, and other deep space destinations. The Cis-Lunar Destination Team of NASA's Human Spaceflight Architecture Team (HAT) has been analyzing cis-lunar destination activities and developing notional missions (or "destination Design Reference Missions" [DRMs]) for cis-lunar locations to inform roadmap and architecture development, transportation and destination elements definition, operations, and strategic knowledge gaps. The cis-lunar domain is defined as that area of deep space under the gravitational influence of the earth-moon system. This includes a set of earth-centered orbital locations in low earth orbit (LEO), geosynchronous earth orbit (GEO), highly elliptical and high earth orbits (HEO), earth-moon libration or "Lagrange" points (E-ML1 through E-ML5, and in particular, E-ML1 and E-ML2), and low lunar orbit (LLO). To help explore this large possibility space, we developed a set of high level cis-lunar mission concepts in the form of a large mission tree, defined primarily by mission duration, pre-deployment, type of mission, and location. The mission tree has provided an overall analytical context and has helped in developing more detailed design reference missions that are then intended to inform capabilities, operations, and architectures. With the mission tree as context, we will describe two destination DRMs to LEO and GEO, based on present human space exploration architectural considerations, as well as our recent work on defining mission activities that could be conducted with an EML1 or EML2 facility, the latter of which will be an emphasis of this

Cytoplasmic protein binding to highly conserved sequences in the 3' untranslated region of mouse protamine 2 mRNA, a translationally regulated transcript of male germ cells

International Nuclear Information System (INIS)

Kwon, Y.K.; Hecht, N.B.

1991-01-01

The expression of the protamines, the predominant nuclear proteins of mammalian spermatozoa, is regulated translationally during male germ-cell development. The 3' untranslated region (UTR) of protamine 1 mRNA has been reported to control its time of translation. To understand the mechanisms controlling translation of the protamine mRNAs, we have sought to identify cis elements of the 3' UTR of protamine 2 mRNA that are recognized by cytoplasmic factors. From gel retardation assays, two sequence elements are shown to form specific RNA-protein complexes. Protein binding sites of the two complexes were determined by RNase T1 mapping, by blocking the putative binding sites with antisense oligonucleotides, and by competition assays. The sequences of these elements, located between nucleotides + 537 and + 572 in protamine 2 mRNA, are highly conserved among postmeiotic translationally regulated nuclear proteins of the mammalian testis. Two closely linked protein binding sites were detected. UV-crosslinking studies revealed that a protein of about 18 kDa binds to one of the conserved sequences. These data demonstrate specific protein binding to a highly conserved 3' UTR of translationally regulated testicular mRNA

A highly conserved basidiomycete peptide synthetase produces a trimeric hydroxamate siderophore.

Science.gov (United States)

Brandenburger, Eileen; Gressler, Markus; Leonhardt, Robin; Lackner, Gerald; Habel, Andreas; Hertweck, Christian; Brock, Matthias; Hoffmeister, Dirk

2017-08-25

The model white-rot basidiomycete Ceriporiopsis ( Gelatoporia ) subvermispora B encodes putative natural product biosynthesis genes. Among them is the gene for the seven-domain nonribosomal peptide synthetase CsNPS2. It is a member of the as-yet uncharacterized fungal type VI siderophore synthetase family which is highly conserved and widely distributed among the basidiomycetes. These enzymes include only one adenylation (A) domain, i.e., one complete peptide synthetase module and two thiolation/condensation (T-C) di-domain partial modules which, together, constitute an AT 1 C 1 T 2 C 2 T 3 C 3 domain setup. The full-length CsNPS2 enzyme (274.5 kDa) was heterologously produced as polyhistidine fusion in Aspergillus niger as soluble and active protein. N 5 -acetyl- N 5 -hydroxy-l-ornithine (l-AHO) and N 5 - cis -anhydromevalonyl- N 5 -hydroxy-l-ornithine (l-AMHO) were accepted as substrates, as assessed in vitro using the substrate-dependent [ 32 P]ATP-pyrophosphate radioisotope exchange assay. Full-length holo -CsNPS2 catalyzed amide bond formation between three l-AHO molecules to release the linear l-AHO trimer, called basidioferrin, as product in vitro , which was verified by LC-HRESIMS. Phylogenetic analyses suggest that type VI family siderophore synthetases are widespread in mushrooms and have evolved in a common ancestor of basidiomycetes. Importance : The basidiomycete nonribosomal peptide synthetase CsNPS2 represents a member of a widely distributed but previously uninvestigated class (type VI) of fungal siderophore synthetases. Genes orthologous to CsNPS2 are highly conserved across various phylogenetic clades of the basidiomycetes. Hence, our work serves as a broadly applicable model for siderophore biosynthesis and iron metabolism in higher fungi. Also, our results on the amino acid substrate preference of CsNPS2 supports further understanding of the substrate selectivity of fungal adenylation domains. Methodologically, this report highlights the

Identification of distal regulatory regions in the human alpha IIb gene locus necessary for consistent, high-level megakaryocyte expression.

Science.gov (United States)

Thornton, Michael A; Zhang, Chunyan; Kowalska, Maria A; Poncz, Mortimer

2002-11-15

The alphaIIb/beta3-integrin receptor is present at high levels only in megakaryocytes and platelets. Its presence on platelets is critical for hemostasis. The tissue-specific nature of this receptor's expression is secondary to the restricted expression of alphaIIb, and studies of the alphaIIb proximal promoter have served as a model of a megakaryocyte-specific promoter. We have examined the alphaIIb gene locus for distal regulatory elements. Sequence comparison between the human (h) and murine (m) alphaIIb loci revealed high levels of conservation at intergenic regions both 5' and 3' to the alphaIIb gene. Additionally, deoxyribonuclease (DNase) I sensitivity mapping defined tissue-specific hypersensitive (HS) sites that coincide, in part, with these conserved regions. Transgenic mice containing various lengths of the h(alpha)IIb gene locus, which included or excluded the various conserved/HS regions, demonstrated that the proximal promoter was sufficient for tissue specificity, but that a region 2.5 to 7.1 kb upstream of the h(alpha)IIb gene was necessary for consistent expression. Another region 2.2 to 7.4 kb downstream of the gene enhanced expression 1000-fold and led to levels of h(alpha)IIb mRNA that were about 30% of the native m(alpha)IIb mRNA level. These constructs also resulted in detectable h(alpha)IIb/m(beta)3 on the platelet surface. This work not only confirms the importance of the proximal promoter of the alphaIIb gene for tissue specificity, but also characterizes the distal organization of the alphaIIb gene locus and provides an initial localization of 2 important regulatory regions needed for the expression of the alphaIIb gene at high levels during megakaryopoiesis.

Cis-acting regulatory sequences promote high-frequency gene conversion between repeated sequences in mammalian cells.

Science.gov (United States)

Raynard, Steven J; Baker, Mark D

2004-01-01

In mammalian cells, little is known about the nature of recombination-prone regions of the genome. Previously, we reported that the immunoglobulin heavy chain (IgH) mu locus behaved as a hotspot for mitotic, intrachromosomal gene conversion (GC) between repeated mu constant (Cmu) regions in mouse hybridoma cells. To investigate whether elements within the mu gene regulatory region were required for hotspot activity, gene targeting was used to delete a 9.1 kb segment encompassing the mu gene promoter (Pmu), enhancer (Emu) and switch region (Smu) from the locus. In these cell lines, GC between the Cmu repeats was significantly reduced, indicating that this 'recombination-enhancing sequence' (RES) is necessary for GC hotspot activity at the IgH locus. Importantly, the RES fragment stimulated GC when appended to the same Cmu repeats integrated at ectopic genomic sites. We also show that deletion of Emu and flanking matrix attachment regions (MARs) from the RES abolishes GC hotspot activity at the IgH locus. However, no stimulation of ectopic GC was observed with the Emu/MARs fragment alone. Finally, we provide evidence that no correlation exists between the level of transcription and GC promoted by the RES. We suggest a model whereby Emu/MARS enhances mitotic GC at the endogenous IgH mu locus by effecting chromatin modifications in adjacent DNA.

Dilemmas of compliance in the CIS

International Nuclear Information System (INIS)

Vorobyev, A.

1996-01-01

The objective of this paper is to examine some of the difficulties faced by Russia and other Common Independent States (CIS) in the field of compliance with disarmament treaties and non-proliferation regimes, as well as ways and means, particularly with regard to the legal framework, designed to overcome these difficulties. Naturally, the fate and pace of overcoming the existing problems will depend only partially on development of CIS States. A large variety of international factors and the general security will be essential for progress in resolving disarmament and arms control issues in the CIS

Whole-body kinetics and dosimetry of cis-4-[18F]fluoro-l-proline

International Nuclear Information System (INIS)

Boerner, Anne R.; Langen, Karl-J.; Herzog, Hans; Hamacher, Kurt; Mueller-Mattheis, Volker; Schmitz, Thomas; Ackermann, Rolf; Coenen, Heinz H.

2001-01-01

The whole-body distribution of 4-cis[ 18 F]fluoro-L-proline (cis-FPro) was studied in six patients with urological tumors by PET. Based on the IMEDOSE and MIRDOSE procedures radiation absorbed doses were estimated from whole-body PET scans acquired at 1 and 3-5 h after i.v. injection of 400 MBq cis-FPro. Cis-FPro showed high retention in the renal cortex and a slight uptake in liver and pancreas. Urinary excretion ranged from 12 to 19% at 5 h p.i. Highest absorbed doses were found for the urinary bladder wall and the kidneys (44.1/44.0 μGy/MBq). The effective dose according to ICRP 60 was 15.1 μSv/MBq for adults. This leads to an effective dose of 6.0 mSv in a PET study using 400 MBq cis-FPro

Whole-body kinetics and dosimetry of cis-4-[(18)F]fluoro-L-proline.

Science.gov (United States)

Börner, A R; Langen, K J; Herzog, H; Hamacher, K; Müller-Mattheis, V; Schmitz, T; Ackermann, R; Coenen, H H

2001-04-01

The whole-body distribution of 4-cis[(18)F]fluoro-L-proline (cis-FPro) was studied in six patients with urological tumors by PET. Based on the IMEDOSE and MIRDOSE procedures radiation absorbed doses were estimated from whole-body PET scans acquired at 1 and 3-5 h after i.v. injection of 400 MBq cis-FPro. Cis-FPro showed high retention in the renal cortex and a slight uptake in liver and pancreas. Urinary excretion ranged from 12 to 19% at 5 h p.i. Highest absorbed doses were found for the urinary bladder wall and the kidneys (44.1/44.0 microGy/mbq). The effective dose according to ICRP 60 was 15.1 microSv/mbq for adults. This leads to an effective dose of 6.0 mSv in a PET study using 400 MBq cis-FPro.

Up front in the CIS

International Nuclear Information System (INIS)

Grey, C.A.

1994-01-01

A picture is drawn of the current supply side of the front-end fuel cycle production capacities in the CIS. Uranium production has been steadily declining, as in the West. Market realities have been reflected in local costs of production since the break-up of the former Soviet Union and some uneconomic mines have been closed. In terms of actual production, Kazakhstan, Russia and Uzbekistan, remain among the top five uranium producers in the world. Western government action has been taken to restrict the market access for natural uranium from the CIS. Reactors in the CIS continue to be supplied with fabricated fuel solely by Russian, though Western fuel fabricators have reduced Russian supplies to Eastern Europe. Russia's current dominance in conversion and enrichment services in both the CIS and Eastern Europe is likely to continue as long as the present surplus low enriched uranium stocks last and surplus production capacity exists. Market penetration in the West has been limited by government action but Russia in 1993 still held about 20% of the world's conversion market and nearly 19% of the enrichment market. (6 figures, 2 tables, 4 references) (UK)

A lettuce (Lactuca sativa) homolog of human Nogo-B receptor interacts with cis-prenyltransferase and is necessary for natural rubber biosynthesis.

Science.gov (United States)

Qu, Yang; Chakrabarty, Romit; Tran, Hue T; Kwon, Eun-Joo G; Kwon, Moonhyuk; Nguyen, Trinh-Don; Ro, Dae-Kyun

2015-01-23

Natural rubber (cis-1,4-polyisoprene) is an indispensable biopolymer used to manufacture diverse consumer products. Although a major source of natural rubber is the rubber tree (Hevea brasiliensis), lettuce (Lactuca sativa) is also known to synthesize natural rubber. Here, we report that an unusual cis-prenyltransferase-like 2 (CPTL2) that lacks the conserved motifs of conventional cis-prenyltransferase is required for natural rubber biosynthesis in lettuce. CPTL2, identified from the lettuce rubber particle proteome, displays homology to a human NogoB receptor and is predominantly expressed in latex. Multiple transgenic lettuces expressing CPTL2-RNAi constructs showed that a decrease of CPTL2 transcripts (3-15% CPTL2 expression relative to controls) coincided with the reduction of natural rubber as low as 5%. We also identified a conventional cis-prenyltransferase 3 (CPT3), exclusively expressed in latex. In subcellular localization studies using fluorescent proteins, cytosolic CPT3 was relocalized to endoplasmic reticulum by co-occurrence of CPTL2 in tobacco and yeast at the log phase. Furthermore, yeast two-hybrid data showed that CPTL2 and CPT3 interact. Yeast microsomes containing CPTL2/CPT3 showed enhanced synthesis of short cis-polyisoprenes, but natural rubber could not be synthesized in vitro. Intriguingly, a homologous pair CPTL1/CPT1, which displays ubiquitous expressions in lettuce, showed a potent dolichol biosynthetic activity in vitro. Taken together, our data suggest that CPTL2 is a scaffolding protein that tethers CPT3 on endoplasmic reticulum and is necessary for natural rubber biosynthesis in planta, but yeast-expressed CPTL2 and CPT3 alone could not synthesize high molecular weight natural rubber in vitro. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Re-resection rates and risk characteristics following breast conserving surgery for breast cancer and carcinoma in situ

DEFF Research Database (Denmark)

Kryh, C G; Pietersen, C A; Rahr, Hans

2014-01-01

OBJECTIVES: To examine the frequency of re-resections and describe risk characteristics: invasive carcinoma or carcinoma in situ (CIS), palpability of the lesion, and neoadjuvant chemotherapy. RESULTS: 1703 breast conserving surgeries were performed: 1575 primary breast conserving surgeries (BCS...

A conserved RNA structural element within the hepatitis B virus post-transcriptional regulatory element enhance nuclear export of intronless transcripts and repress the splicing mechanism.

Science.gov (United States)

Visootsat, Akasit; Payungporn, Sunchai; T-Thienprasert, Nattanan P

2015-12-01

Hepatitis B virus (HBV) infection is a primary cause of hepatocellular carcinoma and liver cirrhosis worldwide. To develop novel antiviral drugs, a better understanding of HBV gene expression regulation is vital. One important aspect is to understand how HBV hijacks the cellular machinery to export unspliced RNA from the nucleus. The HBV post-transcriptional regulatory element (HBV PRE) has been proposed to be the HBV RNA nuclear export element. However, the function remains controversial, and the core element is unclear. This study, therefore, aimed to identify functional regulatory elements within the HBV PRE and investigate their functions. Using bioinformatics programs based on sequence conservation and conserved RNA secondary structures, three regulatory elements were predicted, namely PRE 1151-1410, PRE 1520-1620 and PRE 1650-1684. PRE 1151-1410 significantly increased intronless and unspliced luciferase activity in both HepG2 and COS-7 cells. Likewise, PRE 1151-1410 significantly elevated intronless and unspliced HBV surface transcripts in liver cancer cells. Moreover, motif analysis predicted that PRE 1151-1410 contains several regulatory motifs. This study reported the roles of PRE 1151-1410 in intronless transcript nuclear export and the splicing mechanism. Additionally, these results provide knowledge in the field of HBV RNA regulation. Moreover, PRE 1151-1410 may be used to enhance the expression of other mRNAs in intronless reporter plasmids.

14 CFR 313.4 - Major regulatory actions.

Science.gov (United States)

2010-01-01

...) PROCEDURAL REGULATIONS IMPLEMENTATION OF THE ENERGY POLICY AND CONSERVATION ACT § 313.4 Major regulatory... of actions shall not be deemed as major regulatory actions requiring an energy statement: (1) Tariff...

[Separation and identification of beta-carotene and its cis isomers by high pressure liquid chromatography (HPLC)].

Science.gov (United States)

Carrillo de Padilla, F

1996-06-01

The separation and identification by HPLC of the cis isomers of beta-carotene was studied. A 1.26 mg/ml beta-carotene solution previously isomerized with iodine as a catalyst, was eluted with 2% acetone in hexane, from a Ca(OH)2 chromatographic column in three bands. The fractions were identified by spectrophotometry and the retention times of 2.05, 2.4 and 2.8 min for the 13 cis, all-trans, and 9 cis beta-carotene isomers, determined by HPLC, with 1% acetone in hexane as movil phase. 22.13 mg % of all-trans beta-carotene were found in a sample of canned carrots. It is recommended the analyses of a greater number of samples, the determination of the method's sensitivity, reproductibility, and the use of a standard of reference of a response factor for calculations.

Separation and identification of beta-carotene and its cis isomers by high pressure liquid chromatography (HPLC)

International Nuclear Information System (INIS)

Carrillo de Padilla, F.

1996-01-01

The separation and identification by HPLC of the cis isomers of beta-carotene was studied. A 1.26 mg/ml beta-carotene solution previously isomerized with iodine as a catalyst, was eluted with 2% acetone in hexane, from a Ca(OH)2 chromatographic column in three bands. The fractions were identified by spectrophotometry and the retention times of 2.05, 2.4 and 2.8 min for the 13 cis, all-trans, and 9 cis beta-carotene isomers, determined by HPLC, with 1% acetone in hexane as Mobil phase. 22.13 mg % of all-trans beta-carotene were found in a sample of canned carrots. It is recommended the analyses of a greater number of samples, the determination of the method's sensitivity, reproducibility, and the use of a standard of reference of a response factor for calculations

A randomized phase II chemoprevention trial of 13-CIS retinoic acid with or without alpha tocopherol or observation in subjects at high risk for lung cancer.

Science.gov (United States)

Kelly, Karen; Kittelson, John; Franklin, Wilbur A; Kennedy, Timothy C; Klein, Catherine E; Keith, Robert L; Dempsey, Edward C; Lewis, Marina; Jackson, Mary K; Hirsch, Fred R; Bunn, Paul A; Miller, York E

2009-05-01

No chemoprevention strategies have been proven effective for lung cancer. We evaluated the effect of 13-cis retinoic acid (13-cis RA), with or without alpha tocopherol, as a lung cancer chemoprevention agent in a phase II randomized controlled clinical trial of adult subjects at high risk for lung cancer as defined by the presence of sputum atypia, history of smoking, and airflow obstruction, or a prior surgically cured nonsmall cell lung cancer (disease free, >3 years). Subjects were randomly assigned to receive either 13-cis RA, 13-cis RA plus alpha tocopherol (13-cis RA/alpha toco) or observation for 12 months. Outcome measures are derived from histologic evaluation of bronchial biopsy specimens obtained by bronchoscopy at baseline and follow-up. The primary outcome measure is treatment "failure" defined as histologic progression (any increase in the maximum histologic score) or failure to return for follow-up bronchoscopy. Seventy-five subjects were randomized (27/22/26 to observations/13-cis RA/13-cis RA/alpha toco); 59 completed the trial; 55 had both baseline and follow-up bronchoscopy. The risk of treatment failure was 55.6% (15 of 27) and 50% (24 of 48) in the observation and combined (13 cis RA plus 13 cis RA/alpha toco) treatment arms, respectively (odds ratio adjusted for baseline histology, 0.97; 95% confidence interval, 0.36-2.66; P = 0.95). Among subjects with complete histology data, maximum histology score in the observation arm increased by 0.37 units and by 0.03 units in the treated arms (difference adjusted for baseline, -0.18; 95% confidence interval, -1.16 to 0.81; P = 0.72). Similar (nonsignificant) results were observed for treatment effects on endobronchial proliferation as assessed by Ki-67 immunolabeling. Twelve-month treatment with 13-cis RA produced nonsignificant changes in bronchial histology, consistent with results in other trials. Agents advancing to phase III randomized trials should produce greater histologic changes. The

A Randomized Phase II Chemoprevention Trial of 13-CIS Retinoic Acid with Or without α Tocopherol or Observation in Subjects at High Risk for Lung Cancer

Science.gov (United States)

Kelly, Karen; Kittelson, John; Franklin, Wilbur A.; Kennedy, Timothy C.; Klein, Catherine E.; Keith, Robert L.; Dempsey, Edward C.; Lewis, Marina; Jackson, Mary K.; Hirsch, Fred R.; Bunn, Paul A.; Miller, York E.

2011-01-01

No chemoprevention strategies have been proven effective for lung cancer. We evaluated the effect of 13-cis retinoic acid (13-cis RA), with or without α tocopherol, as a lung cancer chemoprevention agent in a phase II randomized controlled clinical trial of adult subjects at high risk for lung cancer as defined by the presence of sputum atypia, history of smoking, and airflow obstruction, or a prior surgically cured nonsmall cell lung cancer (disease free, >3 years). Subjects were randomly assigned to receive either 13-cis RA, 13-cis RA plus α tocopherol (13-cis RA/α toco) or observation for 12 months. Outcome measures are derived from histologic evaluation of bronchial biopsy specimens obtained by bronchoscopy at baseline and follow-up. The primary outcome measure is treatment “failure” defined as histologic progression (any increase in the maximum histologic score) or failure to return for follow-up bronchoscopy. Seventy-five subjects were randomized (27/22/26 to obervations/13-cis RA/13-cis RA/α toco); 59 completed the trial; 55 had both baseline and follow-up bronchoscopy. The risk of treatment failure was 55.6% (15 of 27) and 50% (24 of 48) in the observation and combined (13 cis RA plus 13 cis RA/α toco) treatment arms, respectively (odds ratio adjusted for baseline histology, 0.97; 95% confidence interval, 0.36–2.66; P = 0.95). Among subjects with complete histology data, maximum histology score in the observation arm increased by 0.37 units and by 0.03 units in the treated arms (difference adjusted for baseline, −0.18; 95% confidence interval, −1.16 to 0.81; P = 0.72). Similar (nonsignificant) results were observed for treatment effects on endobronchial proliferation as assessed by Ki-67 immunolabeling. Twelve-month treatment with 13-cis RA produced nonsignificant changes in bronchial histology, consistent with results in other trials. Agents advancing to phase III randomized trials should produce greater histologic changes. The addition of �

The Evolution of Lineage-Specific Regulatory Activities in the Human Embryonic Limb

OpenAIRE

Cotney, Justin; Leng, Jing; Yin, Jun; Reilly, Steven K.; DeMare, Laura E.; Emera, Deena; Ayoub, Albert E.; Rakic, Pasko; Noonan, James P.

2013-01-01

The evolution of human anatomical features likely involved changes in gene regulation during development. However, the nature and extent of human-specific developmental regulatory functions remain unknown. We obtained a genome-wide view of cis-regulatory evolution in human embryonic tissues by comparing the histone modification H3K27ac, which provides a quantitative readout of promoter and enhancer activity, during human, rhesus, and mouse limb development. Based on increased H3K27ac, we find...

Sheep skeletal muscle transcriptome analysis reveals muscle growth regulatory lncRNAs.

Science.gov (United States)

Chao, Tianle; Ji, Zhibin; Hou, Lei; Wang, Jin; Zhang, Chunlan; Wang, Guizhi; Wang, Jianmin

2018-01-01

As widely distributed domestic animals, sheep are an important species and the source of mutton. In this study, we aimed to evaluate the regulatory lncRNAs associated with muscle growth and development between high production mutton sheep (Dorper sheep and Qianhua Mutton Merino sheep) and low production mutton sheep (Small-tailed Han sheep). In total, 39 lncRNAs were found to be differentially expressed. Using co-expression analysis and functional annotation, 1,206 co-expression interactions were found between 32 lncRNAs and 369 genes, and 29 of these lncRNAs were found to be associated with muscle development, metabolism, cell proliferation and apoptosis. lncRNA-mRNA interactions revealed 6 lncRNAs as hub lncRNAs. Moreover, three lncRNAs and their associated co-expressed genes were demonstrated by cis-regulatory gene analyses, and we also found a potential regulatory relationship between the pseudogene lncRNA LOC101121401 and its parent gene FTH1. This study provides a genome-wide resolution of lncRNA and mRNA regulation in muscles from mutton sheep.

Conservation of lipid metabolic gene transcriptional regulatory networks in fish and mammals.

Science.gov (United States)

Carmona-Antoñanzas, Greta; Tocher, Douglas R; Martinez-Rubio, Laura; Leaver, Michael J

2014-01-15

Lipid content and composition in aquafeeds have changed rapidly as a result of the recent drive to replace ecologically limited marine ingredients, fishmeal and fish oil (FO). Terrestrial plant products are the most economic and sustainable alternative; however, plant meals and oils are devoid of physiologically important cholesterol and long-chain polyunsaturated fatty acids (LC-PUFA), eicosapentaenoic (EPA), docosahexaenoic (DHA) and arachidonic (ARA) acids. Although replacement of dietary FO with vegetable oil (VO) has little effect on growth in Atlantic salmon (Salmo salar), several studies have shown major effects on the activity and expression of genes involved in lipid homeostasis. In vertebrates, sterols and LC-PUFA play crucial roles in lipid metabolism by direct interaction with lipid-sensing transcription factors (TFs) and consequent regulation of target genes. The primary aim of the present study was to elucidate the role of key TFs in the transcriptional regulation of lipid metabolism in fish by transfection and overexpression of TFs. The results show that the expression of genes of LC-PUFA biosynthesis (elovl and fads2) and cholesterol metabolism (abca1) are regulated by Lxr and Srebp TFs in salmon, indicating highly conserved regulatory mechanism across vertebrates. In addition, srebp1 and srebp2 mRNA respond to replacement of dietary FO with VO. Thus, Atlantic salmon adjust lipid metabolism in response to dietary lipid composition through the transcriptional regulation of gene expression. It may be possible to further increase efficient and effective use of sustainable alternatives to marine products in aquaculture by considering these important molecular interactions when formulating diets. © 2013.

Whole-body kinetics and dosimetry of cis-4-[{sup 18}F]fluoro-l-proline

Energy Technology Data Exchange (ETDEWEB)

Boerner, Anne R.; Langen, Karl-J. E-mail: k.j.langen@fz-juelich.de; Herzog, Hans; Hamacher, Kurt; Mueller-Mattheis, Volker; Schmitz, Thomas; Ackermann, Rolf; Coenen, Heinz H

2001-04-01

The whole-body distribution of 4-cis[{sup 18}F]fluoro-L-proline (cis-FPro) was studied in six patients with urological tumors by PET. Based on the IMEDOSE and MIRDOSE procedures radiation absorbed doses were estimated from whole-body PET scans acquired at 1 and 3-5 h after i.v. injection of 400 MBq cis-FPro. Cis-FPro showed high retention in the renal cortex and a slight uptake in liver and pancreas. Urinary excretion ranged from 12 to 19% at 5 h p.i. Highest absorbed doses were found for the urinary bladder wall and the kidneys (44.1/44.0 {mu}Gy/MBq). The effective dose according to ICRP 60 was 15.1 {mu}Sv/MBq for adults. This leads to an effective dose of 6.0 mSv in a PET study using 400 MBq cis-FPro.

First update to the US Nuclear Regulatory Commission's regulatory strategy for the high-level waste repository program

International Nuclear Information System (INIS)

Johnson, R.L.; Linehan, J.J.

1991-01-01

The US Nuclear Regulatory Commission (NRC) staff has updated its initial regulatory strategy for the High-Level Waste Repository Licensing Program. The update describes changes to the initial strategy and summarizes progress and future activities. This paper summarizes the first update of the regulatory strategy. In general the overall strategy of identifying and reducing uncertainties is unchanged. Identifying regulatory and institutional uncertainties is essentially complete, and therefore, the current and future emphasis is on reducing those regulatory and institutional uncertainties identified to date. The NRC staff has improved the methods of reducing regulatory uncertainties by (1) enhancing the technical basis preparation process for potential rulemakings and guidance and (2) designing a new guidance document, called a staff position, for clarifying regulatory uncertainties. For guiding the US DOE's reduction of technical uncertainties, the NRC staff will give more emphasis to prelicense application reviews and less emphasis on preparing staff technical positions

Synthesis of mixed-ligand cobalt complexes and their applications in high cis-1,4-selective butadiene polymerization

KAUST Repository

Liu, Wen; Pan, Weijing; Wang, Peng; Li, Wei; Mu, Jingshan; Weng, Gengsheng; Jia, Xiaoyu; Gong, Dirong; Huang, Kuo-Wei

2015-01-01

Incomplete oxidation of (N-di-tert-butylphosphino)-6-(2-methyl-2’H-benzoimidazole)-2-aminepyridine dichlorocobalt (PN3CoCl2) in DMF results in a unique co-crystal I formed with three parts including DMF, unit A and unit B complex with Co1 and Co2, respectively, (PN3 ligand in unit A: (N-di-tert-butylphosphino)-6-(2’-methyl-2’H-benzoimidazole)-2-aminepyridine, and O=PN3 ligand in unit B: (N-di-tert-butylphosphinoxide)-6-(2’-methyl-2’H-benzoimidazole)-2-aminepyridine) with 1:1:1 molar ratio. Co1 and Co2 complexes both display a five-coordinated distorted-square-pyramidal geometry around the metal center. The Co1 center is coordinated with PN3 ligand via two N atoms from pyridine, benzoimidazole moiety as well as one P atom, and the Co2 center is coordinated with the oxidized ligandO=PN3 via two N atoms from pyridine, benzoimidazole moiety as well as one O atom from DMF molecule, while the oxidized phosphine moiety (O=P) being excluded from the coordination sphere. Activated with AlEt2Cl, the co-crystallized complexes I are able to actively convert butadiene to polybutadiene, affording cis-1,4 polybutadiene with cis-1,4 unit up to 95.5-97.8% and number average molecular weight of cal. 105g/mol. The high cis-1,4 selectivity and monomodal GPC curve of resultant polymer imply that the identical active species generated from two distinctive cobalt centers.

Synthesis of mixed-ligand cobalt complexes and their applications in high cis-1,4-selective butadiene polymerization

KAUST Repository

Liu, Wen

2015-08-03

Incomplete oxidation of (N-di-tert-butylphosphino)-6-(2-methyl-2’H-benzoimidazole)-2-aminepyridine dichlorocobalt (PN3CoCl2) in DMF results in a unique co-crystal I formed with three parts including DMF, unit A and unit B complex with Co1 and Co2, respectively, (PN3 ligand in unit A: (N-di-tert-butylphosphino)-6-(2’-methyl-2’H-benzoimidazole)-2-aminepyridine, and O=PN3 ligand in unit B: (N-di-tert-butylphosphinoxide)-6-(2’-methyl-2’H-benzoimidazole)-2-aminepyridine) with 1:1:1 molar ratio. Co1 and Co2 complexes both display a five-coordinated distorted-square-pyramidal geometry around the metal center. The Co1 center is coordinated with PN3 ligand via two N atoms from pyridine, benzoimidazole moiety as well as one P atom, and the Co2 center is coordinated with the oxidized ligandO=PN3 via two N atoms from pyridine, benzoimidazole moiety as well as one O atom from DMF molecule, while the oxidized phosphine moiety (O=P) being excluded from the coordination sphere. Activated with AlEt2Cl, the co-crystallized complexes I are able to actively convert butadiene to polybutadiene, affording cis-1,4 polybutadiene with cis-1,4 unit up to 95.5-97.8% and number average molecular weight of cal. 105g/mol. The high cis-1,4 selectivity and monomodal GPC curve of resultant polymer imply that the identical active species generated from two distinctive cobalt centers.

Optimization of Pseudomonas putida KT2440 as host for the production of cis, cis-muconate from benzoate

NARCIS (Netherlands)

Duuren, van J.B.J.H.

2011-01-01

Optimization of Pseudomonas putida KT2440 as host for the production of cis, cis-muconate

from benzoate P. putida KT2440 was used as biocatalyst given its versatile and energetically robust metabolism.

Therefore, a mutant was generated and a process developed based on which a

Nominal Anchors in the CIS

OpenAIRE

Peter M Keller; Thomas J Richardson

2003-01-01

Monetary policy has become increasingly important in the countries of the Commonwealth of Independent States (CIS) as fiscal adjustment and structural reforms have taken root. Inflation has been brought down to relatively low levels in almost all of these countries, raising the question of what should be the appropriate nominal anchor at this stage. Formally, almost all CIS countries have floating exchange rate regimes, yet in practice they manage their exchange rates very heavily, perhaps be...

Identification of sparsely distributed clusters of cis-regulatory elements in sets of co-expressed genes

OpenAIRE

Kreiman, Gabriel

2004-01-01

Sequence information and high‐throughput methods to measure gene expression levels open the door to explore transcriptional regulation using computational tools. Combinatorial regulation and sparseness of regulatory elements throughout the genome allow organisms to control the spatial and temporal patterns of gene expression. Here we study the organization of cis‐regulatory elements in sets of co‐regulated genes. We build an algorithm to search for combinations of transcription factor binding...

The lytic origin of herpesvirus papio is highly homologous to Epstein-Barr virus ori-Lyt: evolutionary conservation of transcriptional activation and replication signals.

Science.gov (United States)

Ryon, J J; Fixman, E D; Houchens, C; Zong, J; Lieberman, P M; Chang, Y N; Hayward, G S; Hayward, S D

1993-01-01

Herpesvirus papio (HVP) is a B-lymphotropic baboon virus with an estimated 40% homology to Epstein-Barr virus (EBV). We have cloned and sequenced ori-Lyt of herpesvirus papio and found a striking degree of nucleotide homology (89%) with ori-Lyt of EBV. Transcriptional elements form an integral part of EBV ori-Lyt. The promoter and enhancer domains of EBV ori-Lyt are conserved in herpesvirus papio. The EBV ori-Lyt promoter contains four binding sites for the EBV lytic cycle transactivator Zta, and the enhancer includes one Zta and two Rta response elements. All five of the Zta response elements and one of the Rta motifs are conserved in HVP ori-Lyt, and the HVP DS-L leftward promoter and the enhancer were activated in transient transfection assays by the EBV Zta and Rta transactivators. The EBV ori-Lyt enhancer contains a palindromic sequence, GGTCAGCTGACC, centered on a PvuII restriction site. This sequence, with a single base change, is also present in the HVP ori-Lyt enhancer. DNase I footprinting demonstrated that the PvuII sequence was bound by a protein present in a Raji nuclear extract. Mobility shift and competition assays using oligonucleotide probes identified this sequence as a binding site for the cellular transcription factor MLTF. Mutagenesis of the binding site indicated that MLTF contributes significantly to the constitutive activity of the ori-Lyt enhancer. The high degree of conservation of cis-acting signal sequences in HVP ori-Lyt was further emphasized by the finding that an HVP ori-Lyt-containing plasmid was replicated in Vero cells by a set of cotransfected EBV replication genes. The central domain of EBV ori-Lyt contains two related AT-rich palindromes, one of which is partially duplicated in the HVP sequence. The AT-rich palindromes are functionally important cis-acting motifs. Deletion of these palindromes severely diminished replication of an ori-Lyt target plasmid. Images PMID:8389916

The pineapple genome and the evolution of CAM photosynthesis.

Science.gov (United States)

Ming, Ray; VanBuren, Robert; Wai, Ching Man; Tang, Haibao; Schatz, Michael C; Bowers, John E; Lyons, Eric; Wang, Ming-Li; Chen, Jung; Biggers, Eric; Zhang, Jisen; Huang, Lixian; Zhang, Lingmao; Miao, Wenjing; Zhang, Jian; Ye, Zhangyao; Miao, Chenyong; Lin, Zhicong; Wang, Hao; Zhou, Hongye; Yim, Won C; Priest, Henry D; Zheng, Chunfang; Woodhouse, Margaret; Edger, Patrick P; Guyot, Romain; Guo, Hao-Bo; Guo, Hong; Zheng, Guangyong; Singh, Ratnesh; Sharma, Anupma; Min, Xiangjia; Zheng, Yun; Lee, Hayan; Gurtowski, James; Sedlazeck, Fritz J; Harkess, Alex; McKain, Michael R; Liao, Zhenyang; Fang, Jingping; Liu, Juan; Zhang, Xiaodan; Zhang, Qing; Hu, Weichang; Qin, Yuan; Wang, Kai; Chen, Li-Yu; Shirley, Neil; Lin, Yann-Rong; Liu, Li-Yu; Hernandez, Alvaro G; Wright, Chris L; Bulone, Vincent; Tuskan, Gerald A; Heath, Katy; Zee, Francis; Moore, Paul H; Sunkar, Ramanjulu; Leebens-Mack, James H; Mockler, Todd; Bennetzen, Jeffrey L; Freeling, Michael; Sankoff, David; Paterson, Andrew H; Zhu, Xinguang; Yang, Xiaohan; Smith, J Andrew C; Cushman, John C; Paull, Robert E; Yu, Qingyi

2015-12-01

Pineapple (Ananas comosus (L.) Merr.) is the most economically valuable crop possessing crassulacean acid metabolism (CAM), a photosynthetic carbon assimilation pathway with high water-use efficiency, and the second most important tropical fruit. We sequenced the genomes of pineapple varieties F153 and MD2 and a wild pineapple relative, Ananas bracteatus accession CB5. The pineapple genome has one fewer ancient whole-genome duplication event than sequenced grass genomes and a conserved karyotype with seven chromosomes from before the ρ duplication event. The pineapple lineage has transitioned from C3 photosynthesis to CAM, with CAM-related genes exhibiting a diel expression pattern in photosynthetic tissues. CAM pathway genes were enriched with cis-regulatory elements associated with the regulation of circadian clock genes, providing the first cis-regulatory link between CAM and circadian clock regulation. Pineapple CAM photosynthesis evolved by the reconfiguration of pathways in C3 plants, through the regulatory neofunctionalization of preexisting genes and not through the acquisition of neofunctionalized genes via whole-genome or tandem gene duplication.

Studies on radiosensitization of Escherichia coli cells by cis-platinum complexes

International Nuclear Information System (INIS)

Zimbrick, J.D.; Sukrochana, A.; Richmond, R.C.

1979-01-01

We recently reported that the antitumor drug cis-Pt(NH 3 ) 2 Cl 2 (cis-DDP) produces significant radiosensitization of anoxic E coli C cells. We have extended these studies to three other platinum drugs, all of which have been shown to be more effective antitumor drugs than cis-DDP. The drugs are: cis-dichloro bis(ethylene imine) Pt(II) (cis-DEP); cis-dichlorobicyclopentylamine Pt(II) (cis-PAD); and Pt-thymine blue (cis-PTB). Survival curve studies indicate that these drugs all produce greater anoxic radiosensitization of E coli C than cis-DDP at concentrations which are less toxic to the cells than similar concentrations of cis-DDP. If the cells are treated with any one of these drugs for two hours and then washed to remove the drug before irradiation, no detectable radiosensitization is found. We conclude that these drugs have the potential for being useful agents in combined modality therapy and that they warrant further study in mammalian systems

Cross-cohort analysis identifies a TEAD4 ↔ MYCN positive-feedback loop as the core regulatory element of high-risk neuroblastoma. | Office of Cancer Genomics

Science.gov (United States)

High-risk neuroblastomas show a paucity of recurrent somatic mutations at diagnosis. As a result, the molecular basis for this aggressive phenotype remains elusive. Recent progress in regulatory network analysis helped us elucidate disease-driving mechanisms downstream of genomic alterations, including recurrent chromosomal alterations. Our analysis identified three molecular subtypes of high-risk neuroblastomas, consistent with chromosomal alterations, and identified subtype-specific master regulator (MR) proteins that were conserved across independent cohorts.

Hepatitis B virus nuclear export elements: RNA stem-loop α and β, key parts of the HBV post-transcriptional regulatory element.

Science.gov (United States)

Lim, Chun Shen; Brown, Chris M

2016-09-01

Many viruses contain RNA elements that modulate splicing and/or promote nuclear export of their RNAs. The RNAs of the major human pathogen, hepatitis B virus (HBV) contain a large (~600 bases) composite cis-acting 'post-transcriptional regulatory element' (PRE). This element promotes expression from these naturally intronless transcripts. Indeed, the related woodchuck hepadnavirus PRE (WPRE) is used to enhance expression in gene therapy and other expression vectors. These PRE are likely to act through a combination of mechanisms, including promotion of RNA nuclear export. Functional components of both the HBV PRE and WPRE are 2 conserved RNA cis-acting stem-loop (SL) structures, SLα and SLβ. They are within the coding regions of polymerase (P) gene, and both P and X genes, respectively. Based on previous studies using mutagenesis and/or nuclear magnetic resonance (NMR), here we propose 2 covariance models for SLα and SLβ. The model for the 30-nucleotide SLα contains a G-bulge and a CNGG(U) apical loop of which the first and the fourth loop residues form a CG pair and the fifth loop residue is bulged out, as observed in the NMR structure. The model for the 23-nucleotide SLβ contains a 7-base-pair stem and a 9-nucleotide loop. Comparison of the models with other RNA structural elements, as well as similarity searches of human transcriptome and viral genomes demonstrate that SLα and SLβ are specific to HBV transcripts. However, they are well conserved among the hepadnaviruses of non-human primates, the woodchuck and ground squirrel.

Generation of monoclonal antibodies against highly conserved antigens.

Directory of Open Access Journals (Sweden)

Hongzhe Zhou

Full Text Available BACKGROUND: Therapeutic antibody development is one of the fastest growing areas of the pharmaceutical industry. Generating high-quality monoclonal antibodies against a given therapeutic target is very crucial for the success of the drug development. However, due to immune tolerance, some proteins that are highly conserved between mice and humans are not very immunogenic in mice, making it difficult to generate antibodies using a conventional approach. METHODOLOGY/PRINCIPAL FINDINGS: In this report, the impaired immune tolerance of NZB/W mice was exploited to generate monoclonal antibodies against highly conserved or self-antigens. Using two highly conserved human antigens (MIF and HMGB1 and one mouse self-antigen (TNF-alpha as examples, we demonstrate here that multiple clones of high affinity, highly specific antibodies with desired biological activities can be generated, using the NZB/W mouse as the immunization host and a T cell-specific tag fused to a recombinant antigen to stimulate the immune system. CONCLUSIONS/SIGNIFICANCE: We developed an efficient and universal method for generating surrogate or therapeutic antibodies against "difficult antigens" to facilitate the development of therapeutic antibodies.

Single d(ApG)/cis-diamminedichloroplatinum(II) adduct-induced mutagenesis in Escherichia coli

International Nuclear Information System (INIS)

Burnouf, D.; Fuchs, R.P.P.; Gauthier, C.; Chottard, J.C.

1990-01-01

The mutation spectrum induced by the widely used antitumor drug cis-diamminedichloroplatinum(II) (cis-DDP) showed that cisDDP[d(ApG)] adducts, although they account for only 25% of the lesions formed are ∼5 times more mutagenic than the major GG adduct. The authors report the construction of vectors bearing a single cisDDP[d(ApG)] lesion and their use in mutagenesis experiments in Escherichia coli. The mutagenic processing of the lesion is found to depend strictly on induction of the SOS system of the bacterial host cells. In SOS-induced cells, mutation frequencies of 1-2% were detected. All these mutations are targeted to the 5' base of the adduct. Single A → T transversions are mainly observed (80%), whereas A → G transitions account for 10% of the total mutations. Tandem base-pair substitutions involving the adenine residue and the thymine residue immediately 5' to the adduct occur at a comparable frequency (10%). No selective loss of the strand bearing the platinum adduct was seen, suggesting that, in vivo, cisDDP[d(ApG)] adducts are not blocking lesions. The high mutation specificity of cisDDP-[d(ApG)]-induced mutagenesis is discussed in relation to structural data

BIP induces mice CD19(hi) regulatory B cells producing IL-10 and highly expressing PD-L1, FasL.

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Tang, Youfa; Jiang, Qing; Ou, Yanghui; Zhang, Fan; Qing, Kai; Sun, Yuanli; Lu, Wenjie; Zhu, Huifen; Gong, Feili; Lei, Ping; Shen, Guanxin

2016-01-01

Many studies have shown that B cells possess a regulatory function in mouse models of autoimmune diseases. Regulatory B cells can modulate immune response through many types of molecular mechanisms, including the production of IL-10 and the expression of PD-1 Ligand and Fas Ligand, but the microenvironmental factors and mechanisms that induce regulatory B cells have not been fully identified. BIP (binding immunoglobulin protein), a member of the heat shock protein 70 family, is a type of evolutionarily highly conserved protein. In this article, we have found that IL-10(+), PD-L1(hi) and FasL(hi) B cells are discrete cell populations, but enriched in CD19(hi) cells. BIP can induce IL-10-producing splenic B cells, IL-10 secretion and B cells highly expressing PD-L1 and FasL. CD40 signaling acts in synergy with BIP to induce regulatory B cells. BIP increased surface CD19 molecule expression intensity and IL-10(+), PD-L1(hi) and FasL(hi) B cells induced by BIP share the CD19(hi) phenotype. Furthermore, B cells treated with BIP and anti-CD40 can lead to suppression of T cell proliferation and the effect is partially IL-10-dependent and mainly BIP-induced. Taken together, our findings identify a novel function of BIP in the induction of regulatory B cells and add a new reason for the therapy of autoimmune disorders or other inflammatory conditions. Copyright © 2015. Published by Elsevier Ltd.

African wildlife conservation and the evolution of hunting institutions

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't Sas-Rolfes, Michael

2017-11-01

Hunting regulation presents a significant challenge for contemporary global conservation governance. Motivated by various incentives, hunters may act legally or illegally, for or against the interests of conservation. Hunter incentives are shaped by the interactions between unevenly evolving formal and informal institutions, embedded in socio-ecological systems. To work effectively for conservation, regulatory interventions must take these evolving institutional interactions into account. Drawing on analytical tools from evolutionary institutional economics, this article examines the trajectory of African hunting regulation and its consequences. Concepts of institutional dynamics, fit, scale, and interplay are applied to case studies of rhinoceros and lion hunting to highlight issues of significance to conservation outcomes. These include important links between different forms of hunting and dynamic interplay with institutions of trade. The case studies reveal that inappropriate formal regulatory approaches may be undermined by adaptive informal market responses. Poorly regulated hunting may lead to calls for stricter regulations or bans, but such legal restrictions may in turn perversely lead to more intensified and organised illegal hunting activity, further undermining conservation objectives. I conclude by offering insights and recommendations to guide more effective future regulatory interventions and priorities for further research. Specifically, I advocate approaches that move beyond simplistic regulatory interventions toward more complex, but supportive, institutional arrangements that align formal and informal institutions through inclusive stakeholder engagement.

Protection against the clastogenic effect of CIS-PLATINUM by WR-2721, ATP and Vitamin C

International Nuclear Information System (INIS)

Rupova, I.; Yagova, A.

1993-01-01

The mutagenic effect of antineoplastic drug Cis-Platinum has been assessed by the frequency of micronuclei in polychromatic erythrocytes (PE) in the mouse bone marrow at time intervals of 24, 48 and 73 h. The maximal clastogenic effect of 2 mg/kg of Cis-Platinum has been found at 24 h interval after treatment (3.6%). The protection against mutagenic activity of Cis-Platinum has been studied by pretreatment of the mice with the thiophosphate compound WR-2721, adenosine triphosphate (ATP) and Vitamin C alone or in combinations at the time of maximal response. The radioprotector WR-2721 exerts a high anti-mutagenic activity when applied prior to Cis-Platinum (2.2%; p 0.05). (author)

Hydrogenation of fluoroarenes: Direct access to all-cis-(multi)fluorinated cycloalkanes.

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Wiesenfeldt, Mario P; Nairoukh, Zackaria; Li, Wei; Glorius, Frank

2017-09-01

All-c is -multifluorinated cycloalkanes exhibit intriguing electronic properties. In particular, they display extremely high dipole moments perpendicular to the aliphatic ring, making them highly desired motifs in material science. Very few such motifs have been prepared, as their syntheses require multistep sequences from diastereoselectively prefunctionalized precursors. Herein we report a synthetic strategy to access these valuable materials via the rhodium-cyclic (alkyl)(amino)carbene (CAAC)-catalyzed hydrogenation of readily available fluorinated arenes in hexane. This route enables the scalable single-step preparation of an abundance of multisubstituted and multifluorinated cycloalkanes, including all- cis -1,2,3,4,5,6-hexafluorocyclohexane as well as cis-configured fluorinated aliphatic heterocycles. Copyright © 2017, American Association for the Advancement of Science.

Cis-eQTL analysis and functional validation of candidate susceptibility genes for high-grade serous ovarian cancer.

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Lawrenson, Kate; Li, Qiyuan; Kar, Siddhartha; Seo, Ji-Heui; Tyrer, Jonathan; Spindler, Tassja J; Lee, Janet; Chen, Yibu; Karst, Alison; Drapkin, Ronny; Aben, Katja K H; Anton-Culver, Hoda; Antonenkova, Natalia; Baker, Helen; Bandera, Elisa V; Bean, Yukie; Beckmann, Matthias W; Berchuck, Andrew; Bisogna, Maria; Bjorge, Line; Bogdanova, Natalia; Brinton, Louise A; Brooks-Wilson, Angela; Bruinsma, Fiona; Butzow, Ralf; Campbell, Ian G; Carty, Karen; Chang-Claude, Jenny; Chenevix-Trench, Georgia; Chen, Anne; Chen, Zhihua; Cook, Linda S; Cramer, Daniel W; Cunningham, Julie M; Cybulski, Cezary; Dansonka-Mieszkowska, Agnieszka; Dennis, Joe; Dicks, Ed; Doherty, Jennifer A; Dörk, Thilo; du Bois, Andreas; Dürst, Matthias; Eccles, Diana; Easton, Douglas T; Edwards, Robert P; Eilber, Ursula; Ekici, Arif B; Fasching, Peter A; Fridley, Brooke L; Gao, Yu-Tang; Gentry-Maharaj, Aleksandra; Giles, Graham G; Glasspool, Rosalind; Goode, Ellen L; Goodman, Marc T; Grownwald, Jacek; Harrington, Patricia; Harter, Philipp; Hasmad, Hanis Nazihah; Hein, Alexander; Heitz, Florian; Hildebrandt, Michelle A T; Hillemanns, Peter; Hogdall, Estrid; Hogdall, Claus; Hosono, Satoyo; Iversen, Edwin S; Jakubowska, Anna; James, Paul; Jensen, Allan; Ji, Bu-Tian; Karlan, Beth Y; Kruger Kjaer, Susanne; Kelemen, Linda E; Kellar, Melissa; Kelley, Joseph L; Kiemeney, Lambertus A; Krakstad, Camilla; Kupryjanczyk, Jolanta; Lambrechts, Diether; Lambrechts, Sandrina; Le, Nhu D; Lee, Alice W; Lele, Shashi; Leminen, Arto; Lester, Jenny; Levine, Douglas A; Liang, Dong; Lissowska, Jolanta; Lu, Karen; Lubinski, Jan; Lundvall, Lene; Massuger, Leon F A G; Matsuo, Keitaro; McGuire, Valerie; McLaughlin, John R; Nevanlinna, Heli; McNeish, Ian; Menon, Usha; Modugno, Francesmary; Moysich, Kirsten B; Narod, Steven A; Nedergaard, Lotte; Ness, Roberta B; Azmi, Mat Adenan Noor; Odunsi, Kunle; Olson, Sara H; Orlow, Irene; Orsulic, Sandra; Weber, Rachel Palmieri; Pearce, Celeste L; Pejovic, Tanja; Pelttari, Liisa M; Permuth-Wey, Jennifer; Phelan, Catherine M; Pike, Malcolm C; Poole, Elizabeth M; Ramus, Susan J; Risch, Harvey A; Rosen, Barry; Rossing, Mary Anne; Rothstein, Joseph H; Rudolph, Anja; Runnebaum, Ingo B; Rzepecka, Iwona K; Salvesen, Helga B; Schildkraut, Joellen M; Schwaab, Ira; Sellers, Thomas A; Shu, Xiao-Ou; Shvetsov, Yurii B; Siddiqui, Nadeem; Sieh, Weiva; Song, Honglin; Southey, Melissa C; Sucheston, Lara; Tangen, Ingvild L; Teo, Soo-Hwang; Terry, Kathryn L; Thompson, Pamela J; Timorek, Agnieszka; Tsai, Ya-Yu; Tworoger, Shelley S; van Altena, Anne M; Van Nieuwenhuysen, Els; Vergote, Ignace; Vierkant, Robert A; Wang-Gohrke, Shan; Walsh, Christine; Wentzensen, Nicolas; Whittemore, Alice S; Wicklund, Kristine G; Wilkens, Lynne R; Woo, Yin-Ling; Wu, Xifeng; Wu, Anna H; Yang, Hannah; Zheng, Wei; Ziogas, Argyrios; Monteiro, Alvaro; Pharoah, Paul D; Gayther, Simon A; Freedman, Matthew L

2015-09-22

Genome-wide association studies have reported 11 regions conferring risk of high-grade serous epithelial ovarian cancer (HGSOC). Expression quantitative trait locus (eQTL) analyses can identify candidate susceptibility genes at risk loci. Here we evaluate cis-eQTL associations at 47 regions associated with HGSOC risk (P≤10(-5)). For three cis-eQTL associations (P<1.4 × 10(-3), FDR<0.05) at 1p36 (CDC42), 1p34 (CDCA8) and 2q31 (HOXD9), we evaluate the functional role of each candidate by perturbing expression of each gene in HGSOC precursor cells. Overexpression of HOXD9 increases anchorage-independent growth, shortens population-doubling time and reduces contact inhibition. Chromosome conformation capture identifies an interaction between rs2857532 and the HOXD9 promoter, suggesting this SNP is a leading causal variant. Transcriptomic profiling after HOXD9 overexpression reveals enrichment of HGSOC risk variants within HOXD9 target genes (P=6 × 10(-10) for risk variants (P<10(-4)) within 10 kb of a HOXD9 target gene in ovarian cells), suggesting a broader role for this network in genetic susceptibility to HGSOC.

A Slow Conformational Switch in the BMAL1 Transactivation Domain Modulates Circadian Rhythms.

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Gustafson, Chelsea L; Parsley, Nicole C; Asimgil, Hande; Lee, Hsiau-Wei; Ahlbach, Christopher; Michael, Alicia K; Xu, Haiyan; Williams, Owen L; Davis, Tara L; Liu, Andrew C; Partch, Carrie L

2017-05-18

The C-terminal transactivation domain (TAD) of BMAL1 (brain and muscle ARNT-like 1) is a regulatory hub for transcriptional coactivators and repressors that compete for binding and, consequently, contributes to period determination of the mammalian circadian clock. Here, we report the discovery of two distinct conformational states that slowly exchange within the dynamic TAD to control timing. This binary switch results from cis/trans isomerization about a highly conserved Trp-Pro imide bond in a region of the TAD that is required for normal circadian timekeeping. Both cis and trans isomers interact with transcriptional regulators, suggesting that isomerization could serve a role in assembling regulatory complexes in vivo. Toward this end, we show that locking the switch into the trans isomer leads to shortened circadian periods. Furthermore, isomerization is regulated by the cyclophilin family of peptidyl-prolyl isomerases, highlighting the potential for regulation of BMAL1 protein dynamics in period determination. Copyright © 2017 Elsevier Inc. All rights reserved.

Genome-wide analysis of the regulatory function mediated by the small regulatory psm-mec RNA of methicillin-resistant Staphylococcus aureus.

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Cheung, Gordon Y C; Villaruz, Amer E; Joo, Hwang-Soo; Duong, Anthony C; Yeh, Anthony J; Nguyen, Thuan H; Sturdevant, Daniel E; Queck, S Y; Otto, M

2014-07-01

Several methicillin resistance (SCCmec) clusters characteristic of hospital-associated methicillin-resistant Staphylococcus aureus (MRSA) strains harbor the psm-mec locus. In addition to encoding the cytolysin, phenol-soluble modulin (PSM)-mec, this locus has been attributed gene regulatory functions. Here we employed genome-wide transcriptional profiling to define the regulatory function of the psm-mec locus. The immune evasion factor protein A emerged as the primary conserved and strongly regulated target of psm-mec, an effect we show is mediated by the psm-mec RNA. Furthermore, the psm-mec locus exerted regulatory effects that were more moderate in extent. For example, expression of PSM-mec limited expression of mecA, thereby decreasing methicillin resistance. Our study shows that the psm-mec locus has a rare dual regulatory RNA and encoded cytolysin function. Furthermore, our findings reveal a specific mechanism underscoring the recently emerging concept that S. aureus strains balance pronounced virulence and high expression of antibiotic resistance. Published by Elsevier GmbH.

Inferring the conservative causal core of gene regulatory networks

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Emmert-Streib Frank

2010-09-01

Full Text Available Abstract Background Inferring gene regulatory networks from large-scale expression data is an important problem that received much attention in recent years. These networks have the potential to gain insights into causal molecular interactions of biological processes. Hence, from a methodological point of view, reliable estimation methods based on observational data are needed to approach this problem practically. Results In this paper, we introduce a novel gene regulatory network inference (GRNI algorithm, called C3NET. We compare C3NET with four well known methods, ARACNE, CLR, MRNET and RN, conducting in-depth numerical ensemble simulations and demonstrate also for biological expression data from E. coli that C3NET performs consistently better than the best known GRNI methods in the literature. In addition, it has also a low computational complexity. Since C3NET is based on estimates of mutual information values in conjunction with a maximization step, our numerical investigations demonstrate that our inference algorithm exploits causal structural information in the data efficiently. Conclusions For systems biology to succeed in the long run, it is of crucial importance to establish methods that extract large-scale gene networks from high-throughput data that reflect the underlying causal interactions among genes or gene products. Our method can contribute to this endeavor by demonstrating that an inference algorithm with a neat design permits not only a more intuitive and possibly biological interpretation of its working mechanism but can also result in superior results.

Inferring the conservative causal core of gene regulatory networks.

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Altay, Gökmen; Emmert-Streib, Frank

2010-09-28

Inferring gene regulatory networks from large-scale expression data is an important problem that received much attention in recent years. These networks have the potential to gain insights into causal molecular interactions of biological processes. Hence, from a methodological point of view, reliable estimation methods based on observational data are needed to approach this problem practically. In this paper, we introduce a novel gene regulatory network inference (GRNI) algorithm, called C3NET. We compare C3NET with four well known methods, ARACNE, CLR, MRNET and RN, conducting in-depth numerical ensemble simulations and demonstrate also for biological expression data from E. coli that C3NET performs consistently better than the best known GRNI methods in the literature. In addition, it has also a low computational complexity. Since C3NET is based on estimates of mutual information values in conjunction with a maximization step, our numerical investigations demonstrate that our inference algorithm exploits causal structural information in the data efficiently. For systems biology to succeed in the long run, it is of crucial importance to establish methods that extract large-scale gene networks from high-throughput data that reflect the underlying causal interactions among genes or gene products. Our method can contribute to this endeavor by demonstrating that an inference algorithm with a neat design permits not only a more intuitive and possibly biological interpretation of its working mechanism but can also result in superior results.

Cross-species conservation of endocrine pathways provides a basis for reevaluation of EDSP tiered testing paradigm

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Many structural and functional aspects of the vertebrate hypothalamic-pituitary-gonadal (HPG) axis are known to be highly conserved, but the relative significance of this from a regulatory toxicology perspective has received comparatively little attention. High-quality data gene...

CIS-based registration of quality of life in a single source approach.

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Fritz, Fleur; Ständer, Sonja; Breil, Bernhard; Riek, Markus; Dugas, Martin

2011-04-21

Documenting quality of life (QoL) in routine medical care and using it both for treatment and for clinical research is not common, although such information is absolutely valuable for physicians and patients alike. We therefore aimed at developing an efficient method to integrate quality of life information into the clinical information system (CIS) and thus make it available for clinical care and secondary use. We piloted our method in three different medical departments, using five different QoL questionnaires. In this setting we used structured interviews and onsite observations to perform workflow and form analyses. The forms and pertinent data reports were implemented using the integrated tools of the local CIS. A web-based application for mobile devices was developed based on XML schemata to facilitate data import into the CIS. Data exports of the CIS were analysed with statistical software to perform an analysis of data quality. The quality of life questionnaires are now regularly documented by patients and physicians. The resulting data is available in the Electronic Health Record (EHR) and can be used for treatment purposes and communication as well as research functionalities. The completion of questionnaires by the patients themselves using a mobile device (iPad) and the import of the respective data into the CIS forms were successfully tested in a pilot installation. The quality of data is rendered high by the use of automatic score calculations as well as the automatic creation of forms for follow-up documentation. The QoL data was exported to research databases for use in scientific analysis. The CIS-based QoL is technically feasible, clinically accepted and provides an excellent quality of data for medical treatment and clinical research. Our approach with a commercial CIS and the web-based application is transferable to other sites.

Genome-wide conserved non-coding microsatellite (CNMS) marker-based integrative genetical genomics for quantitative dissection of seed weight in chickpea.

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Bajaj, Deepak; Saxena, Maneesha S; Kujur, Alice; Das, Shouvik; Badoni, Saurabh; Tripathi, Shailesh; Upadhyaya, Hari D; Gowda, C L L; Sharma, Shivali; Singh, Sube; Tyagi, Akhilesh K; Parida, Swarup K

2015-03-01

Phylogenetic footprinting identified 666 genome-wide paralogous and orthologous CNMS (conserved non-coding microsatellite) markers from 5'-untranslated and regulatory regions (URRs) of 603 protein-coding chickpea genes. The (CT)n and (GA)n CNMS carrying CTRMCAMV35S and GAGA8BKN3 regulatory elements, respectively, are abundant in the chickpea genome. The mapped genic CNMS markers with robust amplification efficiencies (94.7%) detected higher intraspecific polymorphic potential (37.6%) among genotypes, implying their immense utility in chickpea breeding and genetic analyses. Seventeen differentially expressed CNMS marker-associated genes showing strong preferential and seed tissue/developmental stage-specific expression in contrasting genotypes were selected to narrow down the gene targets underlying seed weight quantitative trait loci (QTLs)/eQTLs (expression QTLs) through integrative genetical genomics. The integration of transcript profiling with seed weight QTL/eQTL mapping, molecular haplotyping, and association analyses identified potential molecular tags (GAGA8BKN3 and RAV1AAT regulatory elements and alleles/haplotypes) in the LOB-domain-containing protein- and KANADI protein-encoding transcription factor genes controlling the cis-regulated expression for seed weight in the chickpea. This emphasizes the potential of CNMS marker-based integrative genetical genomics for the quantitative genetic dissection of complex seed weight in chickpea. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

MYC cis-Elements in PsMPT Promoter Is Involved in Chilling Response of Paeonia suffruticosa.

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Yuxi Zhang

Full Text Available The MPT transports Pi to synthesize ATP. PsMPT, a chilling-induced gene, was previously reported to promote energy metabolism during bud dormancy release in tree peony. In this study, the regulatory elements of PsMPT promoter involved in chilling response were further analyzed. The PsMPT transcript was detected in different tree peony tissues and was highly expressed in the flower organs, including petal, stigma and stamen. An 1174 bp of the PsMPT promoter was isolated by TAIL-PCR, and the PsMPT promoter::GUS transgenic Arabidopsis was generated and analyzed. GUS staining and qPCR showed that the promoter was active in mainly the flower stigma and stamen. Moreover, it was found that the promoter activity was enhanced by chilling, NaCl, GA, ACC and NAA, but inhibited by ABA, mannitol and PEG. In transgenic plants harboring 421 bp of the PsMPT promoter, the GUS gene expression and the activity were significantly increased by chilling treatment. When the fragment from -421 to -408 containing a MYC cis-element was deleted, the chilling response could not be observed. Further mutation analysis confirmed that the MYC element was one of the key motifs responding to chilling in the PsMPT promoter. The present study provides useful information for further investigation of the regulatory mechanism of PsMPT during the endo-dormancy release.

The value of bladder mapping and prostatic urethra biopsies for detection of carcinoma in situ (CIS).

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Gudjónsson, Sigurdur; Bläckberg, Mats; Chebil, Gunilla; Jahnson, Staffan; Olsson, Hans; Bendahl, Pär-Ola; Månsson, Wiking; Liedberg, Fredrik

2012-07-01

It is well known that CIS is a major risk factor for muscle-invasive bladder cancer and that this entity can be difficult to diagnose. Taking cold-cup mapping biopsies from different areas of the bladder (BMAP) is commonly used in patients at risk of harbouring CIS. The diagnostic accuracy of this approach has not been assessed until now. By using the CIS found in the cystoprostatectomy specimen as an indicator of the true occurrence of CIS and comparing that with the findings of BMAP, it is clear that the sensitivity of BMAP to detect CIS when present is low and that negative findings should be considered unreliable. To assess the value of bladder mapping and prostatic urethra biopsies for detection of urothelial carcinoma in situ (CIS). CIS of the urinary bladder is a flat high-grade lesion of the mucosa associated with a significant risk of progression to muscle-invasive disease. CIS is difficult to identify on cystoscopy, and definite diagnosis requires histopathology. Traditionally, if CIS is suspected, multiple cold-cup biopsies are taken from the bladder mucosa, and resection biopsies are obtained from the prostatic urethra in males. This approach is often called bladder mapping (BMAP). The accuracy of BMAP as a diagnostic tool is not known. Male patients with bladder cancer scheduled for cystectomy underwent cold-cup bladder biopsies (sidewalls, posterior wall, dome, trigone), and resection biopsies were taken from the prostatic urethra. After cystectomy, the surgical specimen was investigated in a standardised manner and subsequently compared with the BMAP biopsies for the presence of CIS. The histopathology reports of 162 patients were analysed. CIS was detected in 46% of the cystoprostatectomy specimens, and multiple (≥2) CIS lesions were found in 30%. BMAP (cold-cup bladder biopsies + resection biopsies from the prostatic urethra) provided sensitivity of 51% for any CIS, and 55% for multiple CIS lesions. The cold-cup biopsies for CIS in the bladder

Two potential hookworm DAF-16 target genes, SNR-3 and LPP-1: gene structure, expression profile, and implications of a cis-regulatory element in the regulation of gene expression.

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Gao, Xin; Goggin, Kevin; Dowling, Camille; Qian, Jason; Hawdon, John M

2015-01-08

Hookworms infect nearly 700 million people, causing anemia and developmental stunting in heavy infections. Little is known about the genomic structure or gene regulation in hookworms, although recent publication of draft genome assemblies has allowed the first investigations of these topics to be undertaken. The transcription factor DAF-16 mediates multiple developmental pathways in the free living nematode Caenorhabditis elegans, and is involved in the recovery from the developmentally arrested L3 in hookworms. Identification of downstream targets of DAF-16 will provide a better understanding of the molecular mechanism of hookworm infection. Genomic Fragment 2.23 containing a DAF-16 binding element (DBE) was used to identify overlapping complementary expressed sequence tags (ESTs). These sequences were used to search a draft assembly of the Ancylostoma caninum genome, and identified two neighboring genes, snr-3 and lpp-1, in a tail-to-tail orientation. Expression patterns of both genes during parasitic development were determined by qRT-PCR. DAF-16 dependent cis-regulatory activity of fragment 2.23 was investigated using an in vitro reporter system. The snr-3 gene spans approximately 5.6 kb in the genome and contains 3 exons and 2 introns, and contains the DBE in its 3' untranslated region. Downstream from snr-3 in a tail-to-tail arrangement is the gene lpp-1. The lpp-1 gene spans more than 6 kb and contains 10 exons and 9 introns. The A. caninum genome contains 2 apparent splice variants, but there are 7 splice variants in the A. ceylanicum genome. While the gene order is similar, the gene structures of the hookworm genes differ from their C. elegans orthologs. Both genes show peak expression in the late L4 stage. Using a cell culture based expression system, fragment 2.23 was found to have both DAF-16-dependent promoter and enhancer activity that required an intact DBE. Two putative DAF-16 targets were identified by genome wide screening for DAF-16 binding

A novel cis-acting element required for DNA damage-inducible expression of yeast DIN7

International Nuclear Information System (INIS)

Yoshitani, Ayako; Yoshida, Minoru; Ling Feng

2008-01-01

Din7 is a DNA damage-inducible mitochondrial nuclease that modulates the stability of mitochondrial DNA (mtDNA) in Saccharomyces cerevisiae. How DIN7 gene expression is regulated, however, has remained largely unclear. Using promoter sequence alignment, we found a highly conserved 19-bp sequence in the promoter regions of DIN7 and NTG1, which encodes an oxidative stress-inducible base-excision-repair enzyme. Deletion of the 19-bp sequence markedly reduced the hydroxyurea (HU)-enhanced DIN7 promoter activity. In addition, nuclear fractions prepared from HU-treated cells were used in in vitro band shift assays to reveal the presence of currently unidentified trans-acting factor(s) that preferentially bound to the 19-bp region. These results suggest that the 19-bp sequence is a novel cis-acting element that is required for the regulation of DIN7 expression in response to HU-induced DNA damage

Manganese catalyzed cis-dihydroxylation of electron deficient alkenes with H(2)O(2).

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Saisaha, Pattama; Pijper, Dirk; van Summeren, Ruben P; Hoen, Rob; Smit, Christian; de Boer, Johannes W; Hage, Ronald; Alsters, Paul L; Feringa, Ben L; Browne, Wesley R

2010-10-07

A practical method for the multigram scale selective cis-dihydroxylation of electron deficient alkenes such as diethyl fumarate and N-alkyl and N-aryl-maleimides using H(2)O(2) is described. High turnovers (>1000) can be achieved with this efficient manganese based catalyst system, prepared in situ from a manganese salt, pyridine-2-carboxylic acid, a ketone and a base, under ambient conditions. Under optimized conditions, for diethyl fumarate at least 1000 turnovers could be achieved with only 1.5 equiv. of H(2)O(2) with d/l-diethyl tartrate (cis-diol product) as the sole product. For electron rich alkenes, such as cis-cyclooctene, this catalyst provides for efficient epoxidation.

Stepwise encapsulation and controlled two-stage release system for cis-Diamminediiodoplatinum.

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Chen, Yun; Li, Qian; Wu, Qingsheng

2014-01-01

cis-Diamminediiodoplatinum (cis-DIDP) is a cisplatin-like anticancer drug with higher anticancer activity, but lower stability and price than cisplatin. In this study, a cis-DIDP carrier system based on micro-sized stearic acid was prepared by an emulsion solvent evaporation method. The maximum drug loading capacity of cis-DIDP-loaded solid lipid nanoparticles was 22.03%, and their encapsulation efficiency was 97.24%. In vitro drug release in phosphate-buffered saline (pH =7.4) at 37.5°C exhibited a unique two-stage process, which could prove beneficial for patients with tumors and malignancies. MTT (3-[4,5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide) assay results showed that cis-DIDP released from cis-DIDP-loaded solid lipid nanoparticles had better inhibition activity than cis-DIDP that had not been loaded.

Mutational robustness of gene regulatory networks.

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Aalt D J van Dijk

Full Text Available Mutational robustness of gene regulatory networks refers to their ability to generate constant biological output upon mutations that change network structure. Such networks contain regulatory interactions (transcription factor-target gene interactions but often also protein-protein interactions between transcription factors. Using computational modeling, we study factors that influence robustness and we infer several network properties governing it. These include the type of mutation, i.e. whether a regulatory interaction or a protein-protein interaction is mutated, and in the case of mutation of a regulatory interaction, the sign of the interaction (activating vs. repressive. In addition, we analyze the effect of combinations of mutations and we compare networks containing monomeric with those containing dimeric transcription factors. Our results are consistent with available data on biological networks, for example based on evolutionary conservation of network features. As a novel and remarkable property, we predict that networks are more robust against mutations in monomer than in dimer transcription factors, a prediction for which analysis of conservation of DNA binding residues in monomeric vs. dimeric transcription factors provides indirect evidence.

Comparative Evolution of Morphological Regulatory Functions in Candida Species

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Lackey, Erika; Vipulanandan, Geethanjali; Childers, Delma S.

2013-01-01

Morphological transitions play an important role in virulence and virulence-related processes in a wide variety of pathogenic fungi, including the most commonly isolated human fungal pathogen Candida albicans. While environmental signals, transcriptional regulators, and target genes associated with C. albicans morphogenesis are well-characterized, considerably little is known about morphological regulatory mechanisms and the extent to which they are evolutionarily conserved in less pathogenic and less filamentous non-albicans Candida species (NACS). We have identified specific optimal filament-inducing conditions for three NACS (C. tropicalis, C. parapsilosis, and C. guilliermondii), which are very limited, suggesting that these species may be adapted for niche-specific filamentation in the host. Only a subset of evolutionarily conserved C. albicans filament-specific target genes were induced upon filamentation in C. tropicalis, C. parapsilosis, and C. guilliermondii. One of the genes showing conserved expression was UME6, a key filament-specific regulator of C. albicans hyphal development. Constitutive high-level expression of UME6 was sufficient to drive increased filamentation as well as biofilm formation and partly restore conserved filament-specific gene expression in both C. tropicalis and C. parapsilosis, suggesting that evolutionary differences in filamentation ability among pathogenic Candida species may be partially attributed to alterations in the expression level of a conserved filamentous growth machinery. In contrast to UME6, NRG1, an important repressor of C. albicans filamentation, showed only a partly conserved role in controlling NACS filamentation. Overall, our results suggest that C. albicans morphological regulatory functions are partially conserved in NACS and have evolved to respond to more specific sets of host environmental cues. PMID:23913541

Systematic discovery of regulatory motifs in Fusarium graminearum by comparing four Fusarium genomes

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Kistler Corby

2010-03-01

Full Text Available Abstract Background Fusarium graminearum (Fg, a major fungal pathogen of cultivated cereals, is responsible for billions of dollars in agriculture losses. There is a growing interest in understanding the transcriptional regulation of this organism, especially the regulation of genes underlying its pathogenicity. The generation of whole genome sequence assemblies for Fg and three closely related Fusarium species provides a unique opportunity for such a study. Results Applying comparative genomics approaches, we developed a computational pipeline to systematically discover evolutionarily conserved regulatory motifs in the promoter, downstream and the intronic regions of Fg genes, based on the multiple alignments of sequenced Fusarium genomes. Using this method, we discovered 73 candidate regulatory motifs in the promoter regions. Nearly 30% of these motifs are highly enriched in promoter regions of Fg genes that are associated with a specific functional category. Through comparison to Saccharomyces cerevisiae (Sc and Schizosaccharomyces pombe (Sp, we observed conservation of transcription factors (TFs, their binding sites and the target genes regulated by these TFs related to pathways known to respond to stress conditions or phosphate metabolism. In addition, this study revealed 69 and 39 conserved motifs in the downstream regions and the intronic regions, respectively, of Fg genes. The top intronic motif is the splice donor site. For the downstream regions, we noticed an intriguing absence of the mammalian and Sc poly-adenylation signals among the list of conserved motifs. Conclusion This study provides the first comprehensive list of candidate regulatory motifs in Fg, and underscores the power of comparative genomics in revealing functional elements among related genomes. The conservation of regulatory pathways among the Fusarium genomes and the two yeast species reveals their functional significance, and provides new insights in their

Comparative Bioinformatics Analysis of Transcription Factor Genes Indicates Conservation of Key Regulatory Domains among Babesia bovis, Babesia microti, and Theileria equi.

Science.gov (United States)

Alzan, Heba F; Knowles, Donald P; Suarez, Carlos E

2016-11-01

Apicomplexa tick-borne hemoparasites, including Babesia bovis, Babesia microti, and Theileria equi are responsible for bovine and human babesiosis and equine theileriosis, respectively. These parasites of vast medical, epidemiological, and economic impact have complex life cycles in their vertebrate and tick hosts. Large gaps in knowledge concerning the mechanisms used by these parasites for gene regulation remain. Regulatory genes coding for DNA binding proteins such as members of the Api-AP2, HMG, and Myb families are known to play crucial roles as transcription factors. Although the repertoire of Api-AP2 has been defined and a HMG gene was previously identified in the B. bovis genome, these regulatory genes have not been described in detail in B. microti and T. equi. In this study, comparative bioinformatics was used to: (i) identify and map genes encoding for these transcription factors among three parasites' genomes; (ii) identify a previously unreported HMG gene in B. microti; (iii) define a repertoire of eight conserved Myb genes; and (iv) identify AP2 correlates among B. bovis and the better-studied Plasmodium parasites. Searching the available transcriptome of B. bovis defined patterns of transcription of these three gene families in B. bovis erythrocyte stage parasites. Sequence comparisons show conservation of functional domains and general architecture in the AP2, Myb, and HMG proteins, which may be significant for the regulation of common critical parasite life cycle transitions in B. bovis, B. microti, and T. equi. A detailed understanding of the role of gene families encoding DNA binding proteins will provide new tools for unraveling regulatory mechanisms involved in B. bovis, B. microti, and T. equi life cycles and environmental adaptive responses and potentially contributes to the development of novel convergent strategies for improved control of babesiosis and equine piroplasmosis.

Comparative Bioinformatics Analysis of Transcription Factor Genes Indicates Conservation of Key Regulatory Domains among Babesia bovis, Babesia microti, and Theileria equi.

Directory of Open Access Journals (Sweden)

Heba F Alzan

2016-11-01

Full Text Available Apicomplexa tick-borne hemoparasites, including Babesia bovis, Babesia microti, and Theileria equi are responsible for bovine and human babesiosis and equine theileriosis, respectively. These parasites of vast medical, epidemiological, and economic impact have complex life cycles in their vertebrate and tick hosts. Large gaps in knowledge concerning the mechanisms used by these parasites for gene regulation remain. Regulatory genes coding for DNA binding proteins such as members of the Api-AP2, HMG, and Myb families are known to play crucial roles as transcription factors. Although the repertoire of Api-AP2 has been defined and a HMG gene was previously identified in the B. bovis genome, these regulatory genes have not been described in detail in B. microti and T. equi. In this study, comparative bioinformatics was used to: (i identify and map genes encoding for these transcription factors among three parasites' genomes; (ii identify a previously unreported HMG gene in B. microti; (iii define a repertoire of eight conserved Myb genes; and (iv identify AP2 correlates among B. bovis and the better-studied Plasmodium parasites. Searching the available transcriptome of B. bovis defined patterns of transcription of these three gene families in B. bovis erythrocyte stage parasites. Sequence comparisons show conservation of functional domains and general architecture in the AP2, Myb, and HMG proteins, which may be significant for the regulation of common critical parasite life cycle transitions in B. bovis, B. microti, and T. equi. A detailed understanding of the role of gene families encoding DNA binding proteins will provide new tools for unraveling regulatory mechanisms involved in B. bovis, B. microti, and T. equi life cycles and environmental adaptive responses and potentially contributes to the development of novel convergent strategies for improved control of babesiosis and equine piroplasmosis.

Relative Stability of cis- and trans-Hydrindanones

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Motoo Tori

2015-01-01

Full Text Available The relative stabilities of several cis- and trans-hydrindanones were compared using both isomerization experiments and MM2 calculations. The generally believed rule that cis-hydrindanones are more stable than trans-isomers is applicable, but is not always true. This review introduces examples, mainly from studies in our laboratory, to explain these facts.

Mutations in the newly identified RAX regulatory sequence are not a frequent cause of micro/anophthalmia.

Science.gov (United States)

Chassaing, Nicolas; Vigouroux, Adeline; Calvas, Patrick

2009-06-01

Microphthalmia and anophthalmia are at the severe end of the spectrum of abnormalities in ocular development. A few genes (SOX2, OTX2, RAX, and CHX10) have been implicated in isolated micro/anophthalmia, but causative mutations of these genes explain less than a quarter of these developmental defects. A specifically conserved SOX2/OTX2-mediated RAX expression regulatory sequence has recently been identified. We postulated that mutations in this sequence could lead to micro/anophthalmia, and thus we performed molecular screening of this regulatory element in patients suffering from micro/anophthalmia. Fifty-one patients suffering from nonsyndromic microphthalmia (n = 40) or anophthalmia (n = 11) were included in this study after negative molecular screening for SOX2, OTX2, RAX, and CHX10 mutations. Mutation screening of the RAX regulatory sequence was performed by direct sequencing for these patients. No mutations were identified in the highly conserved RAX regulatory sequence in any of the 51 patients. Mutations in the newly identified RAX regulatory sequence do not represent a frequent cause of nonsyndromic micro/anophthalmia.

Optimization of Pseudomonas putida KT2440 as host for the production of cis, cis-muconate from benzoate

OpenAIRE

Duuren, van, J.B.J.H.

2011-01-01

Optimization of Pseudomonas putida KT2440 as host for the production of cis, cis-muconate from benzoate P. putida KT2440 was used as biocatalyst given its versatile and energetically robust metabolism. Therefore, a mutant was generated and a process developed based on which a life cycle assessment (LCA) was performed. Additionally, the growth related parameters were experimentally obtained to constrain the metabolic model iJP815 further. The mutant Pseudomonas putida KT2440-JD1 was deri...

Separation and identification of beta-carotene and its cis isomers by high pressure liquid chromatography (HPLC); Separacion e identificacion del beta-caroteno y sus isomeros cis por cromatografia liquida de alta resolucion (HPLC)

Energy Technology Data Exchange (ETDEWEB)

Carrillo de Padilla, F [Universidad Central de Venezuela (UCV), Facultad de Farmacia, Catedra de Analisis de Alimentos, Caracas (Venezuela)

1996-07-01

The separation and identification by HPLC of the cis isomers of beta-carotene was studied. A 1.26 mg/ml beta-carotene solution previously isomerized with iodine as a catalyst, was eluted with 2% acetone in hexane, from a Ca(OH)2 chromatographic column in three bands. The fractions were identified by spectrophotometry and the retention times of 2.05, 2.4 and 2.8 min for the 13 cis, all-trans, and 9 cis beta-carotene isomers, determined by HPLC, with 1% acetone in hexane as Mobil phase. 22.13 mg % of all-trans beta-carotene were found in a sample of canned carrots. It is recommended the analyses of a greater number of samples, the determination of the method's sensitivity, reproducibility, and the use of a standard of reference of a response factor for calculations.

Polibutadieno alto-cis: estudo viscosimétrico em tolueno e ciclo-hexano High-cis polybutadiene: viscometric study in toluene and cyclohexane

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Ivana L. Mello

2006-03-01

toluene and cyclohexane solutions, at 30 °C, were carried out on high-cis polybutadiene synthesized with neodymium catalysts. Six different equations were employed to calculate intrinsic viscosity values and viscometric constants: Huggins; Kraemer; Martin and Schulz-Blaschke, by graphic extrapolation; and Solomon-Ciuta; Deb-Chanterjee; and again Schulz-Blaschke, by single point determination. The values obtained for intrinsic viscosity in both solvents and also the molecular weight (in toluene, calculated by applying Mark-Houwink-Sakurada equation, were compared in order to verify the validity of single point determination, which was confirmed. Solomon-Ciuta equation seemed to be suitable for toluene solutions whereas the Deb-Chanterjee equation was adequate for cyclohexane solutions. End-to-end distances of polymeric chains were also determined by applying Flory equation. Based on the higher values of intrinsic viscosity and end-to-end distance of polymeric chains for all samples in toluene, the latter was found to be better solvent than cyclohexane for polybutadiene.

Chiral synthesis of (Z)-3-cis-6,7-cis-9,10-diepoxyhenicosenes, sex pheromone components of the satin moth, Leucoma salicis.

Science.gov (United States)

Wimalaratne, Priyantha D C; Slessor, Keith N

2004-06-01

All four isomers of (Z)-3-cis-6,7-cis-9,10-diepoxyhenicosenes, 1-4, have been synthesized using D-xylose as the chirally pure starting material. D-Xylose was first converted to 2-deoxy-4,5-O-isopropylidene-3-t-butyldimethylsilyl-D-threopentose 11, via several steps of selective protection, dehydroxylation, and deprotection. Wittig coupling of 11 with nonyltriphenylphosphonium bromide followed by hydrogenation and acid catalyzed deprotection of hydroxyl groups yielded the chiral (2R,3R)-1,2,3-triol, 14, which was used as the precursor for the C-8 to C-21 unit of the (Z)-3-cis-6,7-cis-9,10-diepoxyhenicosenes. Selective tosylation of 14 followed by stereospecific cyclization yielded (2R,3R)-1,2-epoxytetradecan-3-ol, 16, which was then divergently converted to the t-butyldimethylsilyl ether 17 and tosylate 22, respectively. Establishment of the C-5 through C-7 unit of the target molecules was accomplished via regiospecific coupling of 17 with 1-t-butyldimethylsiloxy-2-propyne to form 18. Stepwise transformation of 18 via the formation of tosylate 19, desilylation, and stereospecific cyclization to form epoxy alcohol 20, followed by P2-Ni reduction yielded a key intermediate, allylic epoxy alcohol (Z)-2-(5S,6R)-cis-5,6-epoxyheptadecen-1-ol, 21. Similarly, the coupling of 22 with 1-t-butyldimethylsiloxy-2-propyne yielded 23, which was stereospecifically cyclized to form 24. Desilylation and P2-Ni reduction of 24 gave the antipodal intermediate, (Z)-2-(5R,6S)-cis-5,6-epoxyheptadecen-1-ol, 26. Asymmetric epoxidation of antipodes 21 and 26 with (L)- or (D)-diethyl tartrates resulted in the formation of diepoxy alcohols 27 and 29 from 21, and 33 and 31 from 26, respectively. Tosylation of these diepoxy alcohols followed by coupling with lithium dibutenyl cuprate yielded the four stereoisomers of (Z)-3-cis-6,7-cis-9,10-diepoxyhenicosenes, 1-4. Analysis of the retention characteristics of these materials revealed that one or both of the S*,R*,S*,R* stereoisomers comprise the

Highly accessible AU-rich regions in 3’ untranslated regions are hotspots for binding of regulatory factors

Science.gov (United States)

2017-01-01

Post-transcriptional regulation is regarded as one of the major processes involved in the regulation of gene expression. It is mainly performed by RNA binding proteins and microRNAs, which target RNAs and typically affect their stability. Recent efforts from the scientific community have aimed at understanding post-transcriptional regulation at a global scale by using high-throughput sequencing techniques such as cross-linking and immunoprecipitation (CLIP), which facilitates identification of binding sites of these regulatory factors. However, the diversity in the experimental procedures and bioinformatics analyses has hindered the integration of multiple datasets and thus limited the development of an integrated view of post-transcriptional regulation. In this work, we have performed a comprehensive analysis of 107 CLIP datasets from 49 different RBPs in HEK293 cells to shed light on the complex interactions that govern post-transcriptional regulation. By developing a more stringent CLIP analysis pipeline we have discovered the existence of conserved regulatory AU-rich regions in the 3’UTRs where miRNAs and RBPs that regulate several processes such as polyadenylation or mRNA stability bind. Analogous to promoters, many factors have binding sites overlapping or in close proximity in these hotspots and hence the regulation of the mRNA may depend on their relative concentrations. This hypothesis is supported by RBP knockdown experiments that alter the relative concentration of RBPs in the cell. Upon AGO2 knockdown (KD), transcripts containing “free” target sites show increased expression levels compared to those containing target sites in hotspots, which suggests that target sites within hotspots are less available for miRNAs to bind. Interestingly, these hotspots appear enriched in genes with regulatory functions such as DNA binding and RNA binding. Taken together, our results suggest that hotspots are functional regulatory elements that define an extra layer

Identification and characterization of promoters and cis-regulatory elements of genes involved in secondary metabolites production in hop (Humulus lupulus. L)

Czech Academy of Sciences Publication Activity Database

Duraisamy, Ganesh Selvaraj; Mishra, Ajay Kumar; Kocábek, Tomáš; Matoušek, Jaroslav

2016-01-01

Roč. 84, October (2016), s. 346-352 ISSN 1476-9271 R&D Projects: GA ČR GA13-03037S Institutional support: RVO:60077344 Keywords : Cis-acting elements * Gene regulation * Humulus lupulus Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.331, year: 2016

cis-Tetrachlorido-bis(indazole)osmium(iv) and its osmium(iii) analogues: paving the way towards the cis-isomer of the ruthenium anticancer drugs KP1019 and/or NKP1339

KAUST Repository

Bü chel, Gabriel E.; Kossatz, Susanne; Sadique, Ahmad; Rapta, Peter; Zalibera, Michal; Bucinsky, Lukas; Komorovsky, Stanislav; Telser, Joshua; Eppinger, Jö rg; Reiner, Thomas; Arion, Vladimir B.

2017-01-01

The relationship between cis-trans isomerism and anticancer activity has been mainly addressed for square-planar metal complexes, in particular, for platinum(II), e.g., cis- and trans-[PtCl2(NH3)(2)], and a number of related compounds, of which, however, only cis-counterparts are in clinical use today. For octahedral metal complexes, this effect of geometrical isomerism on anticancer activity has not been investigated systematically, mainly because the relevant isomers are still unavailable. An example of such an octahedral complex is trans-[RuCl4(Hind)(2)](-), which is in clinical trials now as its indazolium (KP1019) or sodium salt (NKP1339), but the corresponding cis-isomers remain inaccessible. We report the synthesis of Na[cis-(OsCl4)-Cl-III(kappa N2-1H-ind)(2)] . (Na[1]) suggesting a route to the cis-isomer of NKP1339. The procedure involves heating (H(2)ind)[(OsCl5)-Cl-IV(kappa N1-2H-ind)] in a high boiling point organic solvent resulting in an Anderson rearrangement with the formation of cis-[(OsCl4)-Cl-IV(kappa N2-1H-ind)(2)] ([1]) in high yield. The transformation is accompanied by an indazole coordination mode switch from kappa N1 to kappa N2 and stabilization of the 1H-indazole tautomer. Fully reversible spectroelectrochemical reduction of [1] in acetonitrile at 0.46 V vs. NHE is accompanied by a change in electronic absorption bands indicating the formation of cis-[(OsCl4)-Cl-III(kappa N2-1H-ind)(2)](-) ([1](-)). Chemical reduction of [1] in methanol with NaBH4 followed by addition of nBu(4)NCl afforded the osmium(III) complex nBu(4)N[cis-(OsCl4)-Cl-III(kappa N2-1H-ind)(2)] (nBu(4)N [1]). A metathesis reaction of nBu(4)N[1] with an ion exchange resin led to the isolation of the water-soluble salt Na[1]. The X-ray diffraction crystal structure of [1] . Me2CO was determined and compared with that of trans-[(OsCl4)-Cl-IV(kappa N2-1H-ind)(2)] . 2Me(2)SO (2 . 2Me(2)SO), also prepared in this work. EPR spectroscopy was performed on the Os-III complexes and

cis-Tetrachlorido-bis(indazole)osmium(iv) and its osmium(iii) analogues: paving the way towards the cis-isomer of the ruthenium anticancer drugs KP1019 and/or NKP1339

KAUST Repository

Büchel, Gabriel E.

2017-08-15

The relationship between cis-trans isomerism and anticancer activity has been mainly addressed for square-planar metal complexes, in particular, for platinum(II), e.g., cis- and trans-[PtCl2(NH3)(2)], and a number of related compounds, of which, however, only cis-counterparts are in clinical use today. For octahedral metal complexes, this effect of geometrical isomerism on anticancer activity has not been investigated systematically, mainly because the relevant isomers are still unavailable. An example of such an octahedral complex is trans-[RuCl4(Hind)(2)](-), which is in clinical trials now as its indazolium (KP1019) or sodium salt (NKP1339), but the corresponding cis-isomers remain inaccessible. We report the synthesis of Na[cis-(OsCl4)-Cl-III(kappa N2-1H-ind)(2)] . (Na[1]) suggesting a route to the cis-isomer of NKP1339. The procedure involves heating (H(2)ind)[(OsCl5)-Cl-IV(kappa N1-2H-ind)] in a high boiling point organic solvent resulting in an Anderson rearrangement with the formation of cis-[(OsCl4)-Cl-IV(kappa N2-1H-ind)(2)] ([1]) in high yield. The transformation is accompanied by an indazole coordination mode switch from kappa N1 to kappa N2 and stabilization of the 1H-indazole tautomer. Fully reversible spectroelectrochemical reduction of [1] in acetonitrile at 0.46 V vs. NHE is accompanied by a change in electronic absorption bands indicating the formation of cis-[(OsCl4)-Cl-III(kappa N2-1H-ind)(2)](-) ([1](-)). Chemical reduction of [1] in methanol with NaBH4 followed by addition of nBu(4)NCl afforded the osmium(III) complex nBu(4)N[cis-(OsCl4)-Cl-III(kappa N2-1H-ind)(2)] (nBu(4)N [1]). A metathesis reaction of nBu(4)N[1] with an ion exchange resin led to the isolation of the water-soluble salt Na[1]. The X-ray diffraction crystal structure of [1] . Me2CO was determined and compared with that of trans-[(OsCl4)-Cl-IV(kappa N2-1H-ind)(2)] . 2Me(2)SO (2 . 2Me(2)SO), also prepared in this work. EPR spectroscopy was performed on the Os-III complexes and

Conservation and reclamation at Alberta's mineable oil sands

Energy Technology Data Exchange (ETDEWEB)

Purdy, B.; Richens, T. [Alberta Environment, Edmonton, AB (Canada)

2010-07-01

The regulatory foundation for oil sands in this region is established by the Energy Resources Conservation Board, Environmental Protection and Enhancement Act (EPEA), as well as the Water Act. This presentation discussed the regulatory foundation for conservation and reclamation in the mineable oil sands region. EPEA requirements and conservation objectives were identified. EPEA conservation and reclamation requirements stipulate that an operator must conserve and reclaim and obtain a reclamation certificate. EPEA approvals that were presented compared prescriptive standards versus meeting outcomes at certification. Operational and management challenges as well as the role of research networks and multi-stakeholder organizations were also addressed. Challenge facing the industry include progressive reclamation; tailings management and process-affected water; reclamation certification; integrated landscapes; soil handling and revegetation and monitoring and research. The presentation demonstrated that reclamation begins with mine planning and ends with certification. figs.

Monoclonal antibodies to DNA modified with cis- or trans-diamminedichloroplatinum(II)

International Nuclear Information System (INIS)

Sundquist, W.I.; Lippard, S.J.; Stollar, B.D.

1987-01-01

Murine monoclonal antibodies that bind selectively to adducts formed on DNA by the antitumor drug cis-diamminedichloroplatinum(II), cis-DDP, or to the chemothrapeutically inactive trans isomer trans-DDP were elicited by immunization with calf thymus DNA modified with either cis- or trans-DDP at ratios of bound platinum per nucleotide, (D/N)/sub b/, of 0.06-0.08. The binding of two monoclonal antibodies to cis-DDP-modified DNA was competitively inhibited in an enzyme-linked immunosorbent assay (ELISA) by 4-6 nM concentrations of cis-DDP bound to DNA. Adducts formed by cis-DDP on other synthetic DNA polymers did not inhibit antibody binding to cis-DDP-DNA. The biologically active compounds [Pt(en)Cl 2 ], [Pt(dach)Cl 2 ], and [Pt(NH 3 ) 2 (cbdca)] (carboplatin) all formed antibody-detectable adducts on DNA, whereas the inactive platinum complexes trans-DDP and [Pt(dien)Cl]Cl (dien, diethylenetriamine) did not. The monoclonal antibodies therefore recognize a bifunctional Pt-DNA adduct with cis stereochemistry in which platinum is coordinated by two adjacent guanines or, to a lesser degree, by adjacent adenine and guanine. A monoclonal antibody raised against trans-DDP-DNA was competitively inhibited in an ELISA by 40 nM trans-DDP bound to DNA. This antibody crossreacted with unmodified, denatured DNA. The recognition of cis- or trans-DDP-modified DNAs by monoclonal antibodies thus parallels the known modes of DNA binding of these compounds and may correlate with their biological activities

Sequence-based model of gap gene regulatory network.

Science.gov (United States)

Kozlov, Konstantin; Gursky, Vitaly; Kulakovskiy, Ivan; Samsonova, Maria

2014-01-01

) functional important sites are not exclusively located in cis-regulatory elements, but are rather dispersed through regulatory region. It is of importance that some of the sites with high functional impact in hb, Kr and kni regulatory regions coincide with strong sites annotated and verified in Dnase I footprint assays.

Stepwise encapsulation and controlled two-stage release system for cis-Diamminediiodoplatinum

Directory of Open Access Journals (Sweden)

Chen Y

2014-06-01

Full Text Available Yun Chen,1,* Qian Li,1,2,* Qingsheng Wu1 1Department of Chemistry, Key Laboratory of Yangtze River Water Environment, Ministry of Education, Tongji University, Shanghai; 2Shanghai Institute of Quality Inspection and Technical Research, Shanghai, People’s Republic of China *These authors contributed equally to this work Abstract: cis-Diamminediiodoplatinum (cis-DIDP is a cisplatin-like anticancer drug with higher anticancer activity, but lower stability and price than cisplatin. In this study, a cis-DIDP carrier system based on micro-sized stearic acid was prepared by an emulsion solvent evaporation method. The maximum drug loading capacity of cis-DIDP-loaded solid lipid nanoparticles was 22.03%, and their encapsulation efficiency was 97.24%. In vitro drug release in phosphate-buffered saline (pH =7.4 at 37.5°C exhibited a unique two-stage process, which could prove beneficial for patients with tumors and malignancies. MTT (3-[4,5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide assay results showed that cis-DIDP released from cis-DIDP-loaded solid lipid nanoparticles had better inhibition activity than cis-DIDP that had not been loaded. Keywords: stearic acid, emulsion solvent evaporation method, drug delivery, cis-DIDP, in vitro

Connections between Transcription Downstream of Genes and cis-SAGe Chimeric RNA.

Science.gov (United States)

Chwalenia, Katarzyna; Qin, Fujun; Singh, Sandeep; Tangtrongstittikul, Panjapon; Li, Hui

2017-11-22

cis-Splicing between adjacent genes (cis-SAGe) is being recognized as one way to produce chimeric fusion RNAs. However, its detail mechanism is not clear. Recent study revealed induction of transcriptions downstream of genes (DoGs) under osmotic stress. Here, we investigated the influence of osmotic stress on cis-SAGe chimeric RNAs and their connection to DoGs. We found,the absence of induction of at least some cis-SAGe fusions and/or their corresponding DoGs at early time point(s). In fact, these DoGs and their cis-SAGe fusions are inversely correlated. This negative correlation was changed to positive at a later time point. These results suggest a direct competition between the two categories of transcripts when total pool of readthrough transcripts is limited at an early time point. At a later time point, DoGs and corresponding cis-SAGe fusions are both induced, indicating that total readthrough transcripts become more abundant. Finally, we observed overall enhancement of cis-SAGe chimeric RNAs in KCl-treated samples by RNA-Seq analysis.

Enantioselective disruption of the endocrine system by Cis-Bifenthrin in the male mice.

Science.gov (United States)

Jin, Yuanxiang; Wang, Jiangcong; Pan, Xiuhong; Miao, Wenyu; Lin, Xiaojian; Wang, Linggang; Fu, Zhengwei

2015-07-01

Bifenthrin (BF), as a chiral pyrethroid, is widely used to control field and household pests in China. At present, the commercial BF is a mixed compound containing cis isomers (cis-BF) including two enantiomers of 1R-cis-BF and 1S-cis-BF. In the present study, the two individual cis-BF enantiomers were separated by a preparative supercritical fluid chromatography. Then, four week-old adolescent male ICR mice were orally administered 1R-cis-BF and 1S-cis-BF separately daily for 3 weeks at doses of 0, 7.5 and 15 mg/kg/day, respectively. Results showed that the transcription status of some genes involved in cholesterol synthesis and transport as well as testosterone (T) synthesis in the testes were influenced by cis-BF enantiomers. Especially, we observed that the transcription status of key genes on the pathway of T synthesis including cytochrome P450 cholesterol side-chain cleavage enzyme (P450scc) and cytochrome P450 17α-hydroxysteroid dehydrogenase (P45017α)) were selectively altered in the testis of mice when treated with 1S-cis-BF, suggesting that it is the possible reason to explain why the lower serum T concentration in 1S-cis-BF treated group. Taken together, it concluded that both of the cis-BF enantiomers have the endocrine disruption activities, while 1S-cis-BF was higher than 1R-cis-BF in mice when exposed during the puberty. The data was helpful to understand the toxicity of cis-BF in mammals under enantiomeric level. © 2014 Wiley Periodicals, Inc.

Synthesis of High cis-Polybutadiene in Styrene Solution with Neodymium-Based Catalysts: Towards the Preparation of HIPS and ABS via In Situ Bulk Polymerization

Directory of Open Access Journals (Sweden)

Ramón Díaz de León

2016-01-01

Full Text Available In a first step, 1,3-butadiene was selectively polymerized at 60°C in styrene as solvent using NdV3/DIBAH/EASC as the catalyst system. The catalyst system activation process, the addition order of monomers and catalyst components, and the molar ratios [Al]/[Nd] and [Cl]/[Nd] were studied. The catalyst system allowed the selective 1,3-butadiene polymerization, reaching conversions between 57.5 and 88.1% with low polystyrene contents in the order of 6.3 to 15.4%. Molecular weights ranging from 39,000 to 150,000 g/mol were obtained, while cis-1,4 content was found in the interval of 94.4 to 96.4%. On the other hand, the glass transition temperatures of synthesized materials were established in the range of −101.9 to −107.4°C, explained by the presence of polystyrene segments in the polybutadiene chains; in the same sense, the polybutadienes did not show the typical melting endotherm of high cis-polybutadienes. In a second step, the resulting styrene/high cis-1,4 polybutadiene solutions were used to synthesize ABS (adding a fraction of acrylonitrile monomer and HIPS via in situ bulk polymerizations and the results were discussed in terms of morphological development, molecular parameters, dynamical mechanical behavior, and mechanical properties.

The role of 11-cis-retinyl esters in vertebrate cone vision.

Science.gov (United States)

Babino, Darwin; Perkins, Brian D; Kindermann, Aljoscha; Oberhauser, Vitus; von Lintig, Johannes

2015-01-01

A cycle of cis-to-trans isomerization of the chromophore is intrinsic to vertebrate vision where rod and cone photoreceptors mediate dim- and bright-light vision, respectively. Daylight illumination can greatly exceed the rate at which the photoproduct can be recycled back to the chromophore by the canonical visual cycle. Thus, an additional supply pathway(s) must exist to sustain cone-dependent vision. Two-photon microscopy revealed that the eyes of the zebrafish (Danio rerio) contain high levels of 11-cis-retinyl esters (11-REs) within the retinal pigment epithelium. HPLC analyses demonstrate that 11-REs are bleached by bright light and regenerated in the dark. Pharmacologic treatment with all-trans-retinylamine (Ret-NH2), a potent and specific inhibitor of the trans-to-cis reisomerization reaction of the canonical visual cycle, impeded the regeneration of 11-REs. Intervention with 11-cis-retinol restored the regeneration of 11-REs in the presence of all-trans-Ret-NH2. We used the XOPS:mCFP transgenic zebrafish line with a functional cone-only retina to directly demonstrate that this 11-RE cycle is critical to maintain vision under bright-light conditions. Thus, our analyses reveal that a dark-generated pool of 11-REs helps to supply photoreceptors with the chromophore under the varying light conditions present in natural environments. © FASEB.

A High-Level Petri Net Framework for Genetic Regulatory Networks

Directory of Open Access Journals (Sweden)

Banks Richard

2007-12-01

Full Text Available To understand the function of genetic regulatory networks in the development of cellular systems, we must not only realise the individual network entities, but also the manner by which they interact. Multi-valued networks are a promising qualitative approach for modelling such genetic regulatory networks, however, at present they have limited formal analysis techniques and tools. We present a flexible formal framework for modelling and analysing multi-valued genetic regulatory networks using high-level Petri nets and logic minimization techniques. We demonstrate our approach with a detailed case study in which part of the genetic regulatory network responsible for the carbon starvation stress response in Escherichia coli is modelled and analysed. We then compare and contrast this multivalued model to a corresponding Boolean model and consider their formal relationship.

Single nucleotide polymorphism in transcriptional regulatory regions and expression of environmentally responsive genes

International Nuclear Information System (INIS)

Wang, Xuting; Tomso, Daniel J.; Liu Xuemei; Bell, Douglas A.

2005-01-01

Single nucleotide polymorphisms (SNPs) in the human genome are DNA sequence variations that can alter an individual's response to environmental exposure. SNPs in gene coding regions can lead to changes in the biological properties of the encoded protein. In contrast, SNPs in non-coding gene regulatory regions may affect gene expression levels in an allele-specific manner, and these functional polymorphisms represent an important but relatively unexplored class of genetic variation. The main challenge in analyzing these SNPs is a lack of robust computational and experimental methods. Here, we first outline mechanisms by which genetic variation can impact gene regulation, and review recent findings in this area; then, we describe a methodology for bioinformatic discovery and functional analysis of regulatory SNPs in cis-regulatory regions using the assembled human genome sequence and databases on sequence polymorphism and gene expression. Our method integrates SNP and gene databases and uses a set of computer programs that allow us to: (1) select SNPs, from among the >9 million human SNPs in the NCBI dbSNP database, that are similar to cis-regulatory element (RE) consensus sequences; (2) map the selected dbSNP entries to the human genome assembly in order to identify polymorphic REs near gene start sites; (3) prioritize the candidate polymorphic RE containing genes by searching the existing genotype and gene expression data sets. The applicability of this system has been demonstrated through studies on p53 responsive elements and is being extended to additional pathways and environmentally responsive genes

Targeting cytokine signaling checkpoint CIS activates NK cells to protect from tumor initiation and metastasis

Science.gov (United States)

Putz, Eva M.; Guillerey, Camille; Kos, Kevin; Stannard, Kimberley; Miles, Kim; Delconte, Rebecca B.; Nicholson, Sandra E.; Huntington, Nicholas D.; Smyth, Mark J.

2017-01-01

ABSTRACT The cytokine-induced SH2-containing protein CIS belongs to the suppressor of cytokine signaling (SOCS) protein family. Here, we show the critical role of CIS in suppressing natural killer (NK) cell control of tumor initiation and metastasis. Cish-deficient mice were highly resistant to methylcholanthrene-induced sarcoma formation and protected from lung metastasis of B16F10 melanoma and RM-1 prostate carcinoma cells. In contrast, the growth of primary subcutaneous tumors, including those expressing the foreign antigen OVA, was unchanged in Cish-deficient mice. The combination of Cish deficiency and relevant targeted and immuno-therapies such as combined BRAF and MEK inhibitors, immune checkpoint blockade antibodies, IL-2 and type I interferon revealed further improved control of metastasis. The data clearly indicate that targeting CIS promotes NK cell antitumor functions and CIS holds great promise as a novel target in NK cell immunotherapy. PMID:28344878

RNA-ID, a Powerful Tool for Identifying and Characterizing Regulatory Sequences.

Science.gov (United States)

Brule, C E; Dean, K M; Grayhack, E J

2016-01-01

The identification and analysis of sequences that regulate gene expression is critical because regulated gene expression underlies biology. RNA-ID is an efficient and sensitive method to discover and investigate regulatory sequences in the yeast Saccharomyces cerevisiae, using fluorescence-based assays to detect green fluorescent protein (GFP) relative to a red fluorescent protein (RFP) control in individual cells. Putative regulatory sequences can be inserted either in-frame or upstream of a superfolder GFP fusion protein whose expression, like that of RFP, is driven by the bidirectional GAL1,10 promoter. In this chapter, we describe the methodology to identify and study cis-regulatory sequences in the RNA-ID system, explaining features and variations of the RNA-ID reporter, as well as some applications of this system. We describe in detail the methods to analyze a single regulatory sequence, from construction of a single GFP variant to assay of variants by flow cytometry, as well as modifications required to screen libraries of different strains simultaneously. We also describe subsequent analyses of regulatory sequences. © 2016 Elsevier Inc. All rights reserved.

Adaptive evolution of conserved noncoding elements in mammals.

Directory of Open Access Journals (Sweden)

Su Yeon Kim

2007-09-01

Full Text Available Conserved noncoding elements (CNCs are an abundant feature of vertebrate genomes. Some CNCs have been shown to act as cis-regulatory modules, but the function of most CNCs remains unclear. To study the evolution of CNCs, we have developed a statistical method called the "shared rates test" to identify CNCs that show significant variation in substitution rates across branches of a phylogenetic tree. We report an application of this method to alignments of 98,910 CNCs from the human, chimpanzee, dog, mouse, and rat genomes. We find that approximately 68% of CNCs evolve according to a null model where, for each CNC, a single parameter models the level of constraint acting throughout the phylogeny linking these five species. The remaining approximately 32% of CNCs show departures from the basic model including speed-ups and slow-downs on particular branches and occasionally multiple rate changes on different branches. We find that a subset of the significant CNCs have evolved significantly faster than the local neutral rate on a particular branch, providing strong evidence for adaptive evolution in these CNCs. The distribution of these signals on the phylogeny suggests that adaptive evolution of CNCs occurs in occasional short bursts of evolution. Our analyses suggest a large set of promising targets for future functional studies of adaptation.

Identification of unique cis-element pattern on simulated microgravity treated Arabidopsis by in silico and gene expression

Science.gov (United States)

Soh, Hyuncheol; Choi, Yongsang; Lee, Taek-Kyun; Yeo, Up-Dong; Han, Kyeongsik; Auh, Chungkyun; Lee, Sukchan

2012-08-01

Arabidopsis gene expression microarray (44 K) was used to detect genes highly induced under simulated microgravity stress (SMS). Ten SMS-inducible genes were selected from the microarray data and these 10 genes were found to be abundantly expressed in 3-week-old plants. Nine out of the 10 SMS-inducible genes were also expressed in response to the three abiotic stresses of drought, touch, and wounding in 3-week-old Arabidopsis plants respectively. However, WRKY46 was elevated only in response to SMS. Six other WRKY genes did not respond to SMS. To clarify the characteristics of the genes expressed at high levels in response to SMS, 20 cis-elements in the promoters of the 40 selected genes including the 10 SMS-inducible genes, the 6 WRKY genes, and abiotic stress-inducible genes were analyzed and their spatial positions on each promoter were determined. Four cis-elements (M/T-G-T-P from MYB1AT or TATABOX5, GT1CONSENSUS, TATABOX5, and POLASIG1) showed a unique spatial arrangement in most SMS-inducible genes including WRKY46. Therefore the M/T-G-T-P cis-element patterns identified in the promoter of WRKY46 may play important roles in regulating gene expression in response to SMS. The presences of the cis-element patterns suggest that the order or spatial positioning of certain groups of cis-elements is more important than the existence or numbers of specific cis-elements. Taken together, our data indicate that WRKY46 is a novel SMS inducible transcription factor and the unique spatial arrangement of cis-elements shown in WRKY46 promoter may play an important role for its response to SMS.

The Complexity of Posttranscriptional Small RNA Regulatory Networks Revealed by In Silico Analysis of Gossypium arboreum L. Leaf, Flower and Boll Small Regulatory RNAs.

Directory of Open Access Journals (Sweden)

Hongtao Hu

Full Text Available MicroRNAs (miRNAs and secondary small interfering RNAs (principally phased siRNAs or trans-acting siRNAs are two distinct subfamilies of small RNAs (sRNAs that are emerging as key regulators of posttranscriptional gene expression in plants. Both miRNAs and secondary-siRNAs (sec-siRNAs are processed from longer RNA precursors by DICER-LIKE proteins (DCLs. Gossypium arboreum L., also known as tree cotton or Asian cotton, is a diploid, possibly ancestral relative of tetraploid Gossypium hirsutum L., the predominant type of commercially grown cotton worldwide known as upland cotton. To understand the biological significance of these gene regulators in G. arboreum, a bioinformatics analysis was performed on G. arboreum small RNAs produced from G. arboreum leaf, flower, and boll tissues. Consequently, 263 miRNAs derived from 353 precursors, including 155 conserved miRNAs (cs-miRNAs and 108 novel lineage-specific miRNAs (ls-miRNAs. Along with miRNAs, 2,033 miRNA variants (isomiRNAs were identified as well. Those isomiRNAs with variation at the 3'-miRNA end were expressed at the highest levels, compared to other types of variants. In addition, 755 pha-siRNAs derived 319 pha-siRNA gene transcripts (PGTs were identified, and the potential pha-siRNA initiators were predicted. Also, 2,251 non-phased siRNAs were found as well, of which 1,088 appeared to be produced by so-called cis- or trans-cleavage of the PGTs observed at positions differing from pha-siRNAs. Of those sRNAs, 148 miRNAs/isomiRNAs and 274 phased/non-phased siRNAs were differentially expressed in one or more pairs of tissues examined. Target analysis revealed that target genes for both miRNAs and pha-siRNAs are involved a broad range of metabolic and enzymatic activities. We demonstrate that secondary siRNA production could result from initial cleavage of precursors by both miRNAs or isomiRNAs, and that subsequently produced phased and unphased siRNAs could result that also serve as triggers

The Complexity of Posttranscriptional Small RNA Regulatory Networks Revealed by In Silico Analysis of Gossypium arboreum L. Leaf, Flower and Boll Small Regulatory RNAs.

Science.gov (United States)

Hu, Hongtao; Rashotte, Aaron M; Singh, Narendra K; Weaver, David B; Goertzen, Leslie R; Singh, Shree R; Locy, Robert D

2015-01-01

MicroRNAs (miRNAs) and secondary small interfering RNAs (principally phased siRNAs or trans-acting siRNAs) are two distinct subfamilies of small RNAs (sRNAs) that are emerging as key regulators of posttranscriptional gene expression in plants. Both miRNAs and secondary-siRNAs (sec-siRNAs) are processed from longer RNA precursors by DICER-LIKE proteins (DCLs). Gossypium arboreum L., also known as tree cotton or Asian cotton, is a diploid, possibly ancestral relative of tetraploid Gossypium hirsutum L., the predominant type of commercially grown cotton worldwide known as upland cotton. To understand the biological significance of these gene regulators in G. arboreum, a bioinformatics analysis was performed on G. arboreum small RNAs produced from G. arboreum leaf, flower, and boll tissues. Consequently, 263 miRNAs derived from 353 precursors, including 155 conserved miRNAs (cs-miRNAs) and 108 novel lineage-specific miRNAs (ls-miRNAs). Along with miRNAs, 2,033 miRNA variants (isomiRNAs) were identified as well. Those isomiRNAs with variation at the 3'-miRNA end were expressed at the highest levels, compared to other types of variants. In addition, 755 pha-siRNAs derived 319 pha-siRNA gene transcripts (PGTs) were identified, and the potential pha-siRNA initiators were predicted. Also, 2,251 non-phased siRNAs were found as well, of which 1,088 appeared to be produced by so-called cis- or trans-cleavage of the PGTs observed at positions differing from pha-siRNAs. Of those sRNAs, 148 miRNAs/isomiRNAs and 274 phased/non-phased siRNAs were differentially expressed in one or more pairs of tissues examined. Target analysis revealed that target genes for both miRNAs and pha-siRNAs are involved a broad range of metabolic and enzymatic activities. We demonstrate that secondary siRNA production could result from initial cleavage of precursors by both miRNAs or isomiRNAs, and that subsequently produced phased and unphased siRNAs could result that also serve as triggers of a second

Cis-urocanic acid, a sunlight-induced immunosuppressive factor, activates immune suppression via the 5-HT2A receptor

Science.gov (United States)

Walterscheid, Jeffrey P.; Nghiem, Dat X.; Kazimi, Nasser; Nutt, Leta K.; McConkey, David J.; Norval, Mary; Ullrich, Stephen E.

2006-01-01

Exposure to UV radiation induces skin cancer and suppresses the immune response. To induce immune suppression, the electromagnetic energy of UV radiation must be absorbed by an epidermal photoreceptor and converted into a biologically recognizable signal. Two photoreceptors have been recognized: DNA and trans-urocanic acid (UCA). Trans-UCA is normally found in the outermost layer of skin and isomerizes to the cis isomer upon exposure to UV radiation. Although UCA was identified as a UV photoreceptor years ago, and many have documented its ability to induce immune suppression, its exact mode of action remains elusive. Particularly vexing has been the identity of the molecular pathway by which cis-UCA mediates immune suppression. Here we provide evidence that cis-UCA binds to the serotonin [5-hydroxytryptamine (5-HT)] receptor with relatively high affinity (Kd = 4.6 nM). Anti-cis-UCA antibody precipitates radiolabeled 5-HT, and the binding is inhibited by excess 5-HT and/or excess cis-UCA. Similarly, anti-5-HT antibody precipitates radiolabeled cis-UCA, and the binding is inhibited by excess 5-HT or excess cis-UCA. Calcium mobilization was activated when a mouse fibroblast line, stably transfected with the human 5-HT2A receptor, was treated with cis-UCA. Cis-UCA-induced calcium mobilization was blocked with a selective 5-HT2A receptor antagonist. UV- and cis-UCA-induced immune suppression was blocked by antiserotonin antibodies or by treating the mice with 5-HT2A receptor antagonists. Our findings identify cis-UCA as a serotonin receptor ligand and indicate that the immunosuppressive effects of cis-UCA and UV radiation are mediated by activation of the 5-HT2A receptor. PMID:17085585

Diversity of antisense and other non-coding RNAs in Archaea revealed by comparative small RNA sequencing in four Pyrobaculum species

Directory of Open Access Journals (Sweden)

David L Bernick

2012-07-01

Full Text Available A great diversity of small, non-coding RNA molecules with roles in gene regulation and RNA processing have been intensely studied in eukaryotic and bacterial model organisms, yet our knowledge of possible parallel roles for small RNAs in archaea is limited. We employed RNA-seq to identify novel small RNA across multiple species of the hyperthermophilic genus Pyrobaculum, known for unusual RNA gene characteristics. By comparing transcriptional data collected in parallel among four species, we were able to identify conserved RNA genes fitting into known and novel families. Among our findings, we highlight three novel cis-antisense small RNAs encoded opposite to key regulatory (ferric uptake regulator, metabolic (triose-phosphate isomerase, and core transcriptional apparatus genes (transcription factor B. We also found a large increase in the number of conserved C/D box small RNA genes over what had been previously recognized; many of these genes are encoded antisense to protein coding genes. The conserved opposition to orthologous genes across the Pyrobaculum genus suggests similarities to other cis-antisense regulatory systems. Furthermore, the genus-specific nature of these small RNAs indicates they are relatively recent, stable adaptations.

Effects of cis-9, trans-11 and trans-10, cis-12 conjugated linoleic acid (CLA) isomers on immune function in healthy men

NARCIS (Netherlands)

Albers, R.; Wielen, R.P.J. van der; Brink, E.J.; Hendriks, H.F.J.; Dorovska-Taran, V.N.; Mohede, I.C.M.

2003-01-01

Objectives: To study the effects of two different mixtures of the main conjugated linoleic acid (CLA) isomers cis-9, trans-11 CLA and trans-10, cis-12 CLA on human immune function. Design: Double-blind, randomized, parallel, reference-controlled intervention study. Subjects and intervention:

Identification of a cis-regulatory region of a gene in Arabidopsis thaliana whose induction by dehydration is mediated by abscisic acid and requires protein synthesis.

Science.gov (United States)

Iwasaki, T; Yamaguchi-Shinozaki, K; Shinozaki, K

1995-05-20

In Arabidopsis thaliana, the induction of a dehydration-responsive gene, rd22, is mediated by abscisic acid (ABA) but the gene does not include any sequence corresponding to the consensus ABA-responsive element (ABRE), RYACGTGGYR, in its promoter region. The cis-regulatory region of the rd22 promoter was identified by monitoring the expression of beta-glucuronidase (GUS) activity in leaves of transgenic tobacco plants transformed with chimeric gene fusions constructed between 5'-deleted promoters of rd22 and the coding region of the GUS reporter gene. A 67-bp nucleotide fragment corresponding to positions -207 to -141 of the rd22 promoter conferred responsiveness to dehydration and ABA on a non-responsive promoter. The 67-bp fragment contains the sequences of the recognition sites for some transcription factors, such as MYC, MYB, and GT-1. The fact that accumulation of rd22 mRNA requires protein synthesis raises the possibility that the expression of rd22 might be regulated by one of these trans-acting protein factors whose de novo synthesis is induced by dehydration or ABA. Although the structure of the RD22 protein is very similar to that of a non-storage seed protein, USP, of Vicia faba, the expression of the GUS gene driven by the rd22 promoter in non-stressed transgenic Arabidopsis plants was found mainly in flowers and bolted stems rather than in seeds.

Single nucleotide resolution RNA-seq uncovers new regulatory mechanisms in the opportunistic pathogen Streptococcus agalactiae.

Science.gov (United States)

Rosinski-Chupin, Isabelle; Sauvage, Elisabeth; Sismeiro, Odile; Villain, Adrien; Da Cunha, Violette; Caliot, Marie-Elise; Dillies, Marie-Agnès; Trieu-Cuot, Patrick; Bouloc, Philippe; Lartigue, Marie-Frédérique; Glaser, Philippe

2015-05-30

Streptococcus agalactiae, or Group B Streptococcus, is a leading cause of neonatal infections and an increasing cause of infections in adults with underlying diseases. In an effort to reconstruct the transcriptional networks involved in S. agalactiae physiology and pathogenesis, we performed an extensive and robust characterization of its transcriptome through a combination of differential RNA-sequencing in eight different growth conditions or genetic backgrounds and strand-specific RNA-sequencing. Our study identified 1,210 transcription start sites (TSSs) and 655 transcript ends as well as 39 riboswitches and cis-regulatory regions, 39 cis-antisense non-coding RNAs and 47 small RNAs potentially acting in trans. Among these putative regulatory RNAs, ten were differentially expressed in response to an acid stress and two riboswitches sensed directly or indirectly the pH modification. Strikingly, 15% of the TSSs identified were associated with the incorporation of pseudo-templated nucleotides, showing that reiterative transcription is a pervasive process in S. agalactiae. In particular, 40% of the TSSs upstream genes involved in nucleotide metabolism show reiterative transcription potentially regulating gene expression, as exemplified for pyrG and thyA encoding the CTP synthase and the thymidylate synthase respectively. This comprehensive map of the transcriptome at the single nucleotide resolution led to the discovery of new regulatory mechanisms in S. agalactiae. It also provides the basis for in depth analyses of transcriptional networks in S. agalactiae and of the regulatory role of reiterative transcription following variations of intra-cellular nucleotide pools.

Effects of pyrethroid pesticide cis-bifenthrin on lipogenesis in hepatic cell line.

Science.gov (United States)

Xiang, Dandan; Chu, Tianyi; Li, Meng; Wang, Qiangwei; Zhu, Guonian

2018-06-01

Mounting evidence suggests there is a link between exposure to synthetic pyrethroids (SPs) and the development of obesity. The information presented in this study suggests that cis-bifenthrin (cis-BF) could activate pregnane X receptor (PXR) mediated pathway and lead to the lipid accumulation of human hepatoma (HepG2) cells. Cells were incubated in the control or different concentrations of cis-BF for 24 h. The 1 × 10 -7  M and 1 × 10 -6  M cis-BF exposure were found to induce cellular triglyceride (TG) accumulation significantly. This phenomenon was further supported by Oil Red O Staining assay. The cis-BF exposure caused upregulation of PXR gene and protein. Correspondingly, we also observed the increased expression of downstream genes involved in lipid formation and the inhibition of the expression of β-oxidation. As chiral pesticide，cis-BF was further conformed to behave enantioselectivity in the lipid metabolism. Rather than 1R-cis-BF, HepG2 cells incubated with 1S-cis-BF exhibited a significant TG accumulation. 1S-cis-BF also showed a higher binding level, of which the KD value was 9.184 × 10 -8  M in the SPR assay, compared with 1R-cis-BF (3.463 × 10 -6  M). In addition, the molecular docking simulation analyses correlated well with the KD values measured by the SPR, indicating that 1S-cis-BF showed a better binding affinity with PXR. The results in this study also elucidates the differences between the two enantiomers of pyrethroid-induced toxicity in lipid metabolism of non-target organism. Copyright © 2018 Elsevier Ltd. All rights reserved.

Network University of the CIS as a tool for development of academic mobility within a single (unified educational space of the CIS member states

Directory of Open Access Journals (Sweden)

Гульнара Амангельдиновна Краснова

2010-03-01

Full Text Available The article presents the results of the project «Establishment of a network University of the CIS (2008-2010 years» realization, aimed to establish a joint master's programs in the commonwealth, to strengthen international cooperation in the training of highly qualified specialists, specified tasks to further its implementation in 2010.

Enzymatic study on AtCCD4 and AtCCD7 and their potential to form acyclic regulatory metabolites

KAUST Repository

Bruno, Mark

2016-09-29

The Arabidopsis carotenoid cleavage dioxygenase 4 (AtCCD4) is a negative regulator of the carotenoid content of seeds and has recently been suggested as a candidate for the generation of retrograde signals that are thought to derive from the cleavage of poly-cis-configured carotene desaturation intermediates. In this work, we investigated the activity of AtCCD4 in vitro and used dynamic modeling to determine its substrate preference. Our results document strict regional specificity for cleavage at the C9–C10 double bond in carotenoids and apocarotenoids, with preference for carotenoid substrates and an obstructing effect on hydroxyl functions, and demonstrate the specificity for all-trans-configured carotenes and xanthophylls. AtCCD4 cleaved substrates with at least one ionone ring and did not convert acyclic carotene desaturation intermediates, independent of their isomeric states. These results do not support a direct involvement of AtCCD4 in generating the supposed regulatory metabolites. In contrast, the strigolactone biosynthetic enzyme AtCCD7 converted 9-cis-configured acyclic carotenes, such as 9-cis-ζ-carotene, 9\\'-cis-neurosporene, and 9-cis-lycopene, yielding 9-cis-configured products and indicating that AtCCD7, rather than AtCCD4, is the candidate for forming acyclic retrograde signals.

Enzymatic study on AtCCD4 and AtCCD7 and their potential to form acyclic regulatory metabolites

KAUST Repository

Bruno, Mark; Koschmieder, Julian; Wuest, Florian; Schaub, Patrick; Fehling-Kaschek, Mirjam; Timmer, Jens; Beyer, Peter; Al-Babili, Salim

2016-01-01

The Arabidopsis carotenoid cleavage dioxygenase 4 (AtCCD4) is a negative regulator of the carotenoid content of seeds and has recently been suggested as a candidate for the generation of retrograde signals that are thought to derive from the cleavage of poly-cis-configured carotene desaturation intermediates. In this work, we investigated the activity of AtCCD4 in vitro and used dynamic modeling to determine its substrate preference. Our results document strict regional specificity for cleavage at the C9–C10 double bond in carotenoids and apocarotenoids, with preference for carotenoid substrates and an obstructing effect on hydroxyl functions, and demonstrate the specificity for all-trans-configured carotenes and xanthophylls. AtCCD4 cleaved substrates with at least one ionone ring and did not convert acyclic carotene desaturation intermediates, independent of their isomeric states. These results do not support a direct involvement of AtCCD4 in generating the supposed regulatory metabolites. In contrast, the strigolactone biosynthetic enzyme AtCCD7 converted 9-cis-configured acyclic carotenes, such as 9-cis-ζ-carotene, 9'-cis-neurosporene, and 9-cis-lycopene, yielding 9-cis-configured products and indicating that AtCCD7, rather than AtCCD4, is the candidate for forming acyclic retrograde signals.

Enzymatic study on AtCCD4 and AtCCD7 and their potential to form acyclic regulatory metabolites

Science.gov (United States)

Bruno, Mark; Koschmieder, Julian; Wuest, Florian; Schaub, Patrick; Fehling-Kaschek, Mirjam; Timmer, Jens; Beyer, Peter; Al-Babili, Salim

2016-01-01

The Arabidopsis carotenoid cleavage dioxygenase 4 (AtCCD4) is a negative regulator of the carotenoid content of seeds and has recently been suggested as a candidate for the generation of retrograde signals that are thought to derive from the cleavage of poly-cis-configured carotene desaturation intermediates. In this work, we investigated the activity of AtCCD4 in vitro and used dynamic modeling to determine its substrate preference. Our results document strict regional specificity for cleavage at the C9–C10 double bond in carotenoids and apocarotenoids, with preference for carotenoid substrates and an obstructing effect on hydroxyl functions, and demonstrate the specificity for all-trans-configured carotenes and xanthophylls. AtCCD4 cleaved substrates with at least one ionone ring and did not convert acyclic carotene desaturation intermediates, independent of their isomeric states. These results do not support a direct involvement of AtCCD4 in generating the supposed regulatory metabolites. In contrast, the strigolactone biosynthetic enzyme AtCCD7 converted 9-cis-configured acyclic carotenes, such as 9-cis-ζ-carotene, 9'-cis-neurosporene, and 9-cis-lycopene, yielding 9-cis-configured products and indicating that AtCCD7, rather than AtCCD4, is the candidate for forming acyclic retrograde signals. PMID:27811075

Genomic regulatory landscapes and chromosomal rearrangements

DEFF Research Database (Denmark)

Ladegaard, Elisabete L Engenheiro

2008-01-01

The main objectives of the PhD study are to identify and characterise chromosomal rearrangements within evolutionarily conserved regulatory landscapes around genes involved in the regulation of transcription and/or development (trans-dev genes). A frequent feature of trans-dev genes is that they ......The main objectives of the PhD study are to identify and characterise chromosomal rearrangements within evolutionarily conserved regulatory landscapes around genes involved in the regulation of transcription and/or development (trans-dev genes). A frequent feature of trans-dev genes...... the complex spatio-temporal expression of the associated trans-dev gene. Rare chromosomal breakpoints that disrupt the integrity of these regulatory landscapes may be used as a tool, not only to make genotype-phenotype associations, but also to link the associated phenotype with the position and tissue...... specificity of the individual CNEs. In this PhD study I have studied several chromosomal rearrangements with breakpoints in the vicinity of trans-dev genes. This included chromosomal rearrangements compatible with known phenotype-genotype associations (Rieger syndrome-PITX2, Mowat-Wilson syndrome-ZEB2...

U.S./CIS eye joint nuclear rocket venture

Science.gov (United States)

Clark, John S.; Mcilwain, Melvin C.; Smetanikov, Vladimir; D'Yakov, Evgenij K.; Pavshuk, Vladimir A.

1993-01-01

An account is given of the significance for U.S. spacecraft development of a nuclear thermal rocket (NTR) reactor concept that has been developed in the (formerly Soviet) Commonwealth of Independent States (CIS). The CIS NTR reactor employs a hydrogen-cooled zirconium hydride moderator and ternary carbide fuels; the comparatively cool operating temperatures associated with this design promise overall robustness.

Evidence for multiple major histocompatibility class II X-box binding proteins.

OpenAIRE

Celada, A; Maki, R

1989-01-01

The X box is a loosely conserved DNA sequence that is located upstream of all major histocompatibility class II genes and is one of the cis-acting regulatory elements. Despite the similarity between all X-box sequences, each promoter-proximal X box in the mouse appears to bind a separate nuclear factor.

Detecting the limits of regulatory element conservation anddivergence estimation using pairwise and multiple alignments

Energy Technology Data Exchange (ETDEWEB)

Pollard, Daniel A.; Moses, Alan M.; Iyer, Venky N.; Eisen,Michael B.

2006-08-14

Background: Molecular evolutionary studies of noncodingsequences rely on multiple alignments. Yet how multiple alignmentaccuracy varies across sequence types, tree topologies, divergences andtools, and further how this variation impacts specific inferences,remains unclear. Results: Here we develop a molecular evolutionsimulation platform, CisEvolver, with models of background noncoding andtranscription factor binding site evolution, and use simulated alignmentsto systematically examine multiple alignment accuracy and its impact ontwo key molecular evolutionary inferences: transcription factor bindingsite conservation and divergence estimation. We find that the accuracy ofmultiple alignments is determined almost exclusively by the pairwisedivergence distance of the two most diverged species and that additionalspecies have a negligible influence on alignment accuracy. Conservedtranscription factor binding sites align better than surroundingnoncoding DNA yet are often found to be misaligned at relatively shortdivergence distances, such that studies of binding site gain and losscould easily be confounded by alignment error. Divergence estimates frommultiple alignments tend to be overestimated at short divergencedistances but reach a tool specific divergence at which they cease toincrease, leading to underestimation at long divergences. Our moststriking finding was that overall alignment accuracy, binding sitealignment accuracy and divergence estimation accuracy vary greatly acrossbranches in a tree and are most accurate for terminal branches connectingsister taxa and least accurate for internal branches connectingsub-alignments. Conclusions: Our results suggest that variation inalignment accuracy can lead to errors in molecular evolutionaryinferences that could be construed as biological variation. Thesefindings have implications for which species to choose for analyses, whatkind of errors would be expected for a given set of species and howmultiple alignment tools and

Cis By Trans

OpenAIRE

Rodovalho,Amara Moira

2017-01-01

Cis, trans: above all, metaphors. Cisjordan, region skirting the Jordan River. Cisplatin, Uruguay’s ancient name, region occupying one of the banks of the Prata River. Trans- Amazonian, that which crosses the Amazon; transatlantic, that which crosses the Atlantic. Cisalpine, transalpine. The geometric isomerism of Organic Chemistry, where “cis” are atoms that, when molecules are divided in half, remain on the same side, and “trans” those remaining on opposite sides. Ev...

Proteome-wide analysis of lysine acetylation suggests its broad regulatory scope in Saccharomyces cerevisiae

DEFF Research Database (Denmark)

Henriksen, Peter; Wagner, Sebastian Alexander; Weinert, Brian Tate

2012-01-01

Post-translational modification of proteins by lysine acetylation plays important regulatory roles in living cells. The budding yeast Saccharomyces cerevisiae is a widely used unicellular eukaryotic model organism in biomedical research. S. cerevisiae contains several evolutionary conserved lysine...... acetyltransferases and deacetylases. However, only a few dozen acetylation sites in S. cerevisiae are known, presenting a major obstacle for further understanding the regulatory roles of acetylation in this organism. Here we use high resolution mass spectrometry to identify about 4000 lysine acetylation sites in S....... cerevisiae. Acetylated proteins are implicated in the regulation of diverse cytoplasmic and nuclear processes including chromatin organization, mitochondrial metabolism, and protein synthesis. Bioinformatic analysis of yeast acetylation sites shows that acetylated lysines are significantly more conserved...

Regulatory heterochronies and loose temporal scaling between sea star and sea urchin regulatory circuits.

Science.gov (United States)

Gildor, Tsvia; Hinman, Veronica; Ben-Tabou-De-Leon, Smadar

2017-01-01

It has long been argued that heterochrony, a change in relative timing of a developmental process, is a major source of evolutionary innovation. Heterochronic changes of regulatory gene activation could be the underlying molecular mechanism driving heterochronic changes through evolution. Here, we compare the temporal expression profiles of key regulatory circuits between sea urchin and sea star, representative of two classes of Echinoderms that shared a common ancestor about 500 million years ago. The morphologies of the sea urchin and sea star embryos are largely comparable, yet, differences in certain mesodermal cell types and ectodermal patterning result in distinct larval body plans. We generated high resolution temporal profiles of 17 mesodermally-, endodermally- and ectodermally-expressed regulatory genes in the sea star, Patiria miniata, and compared these to their orthologs in the Mediterranean sea urchin, Paracentrotus lividus. We found that the maternal to zygotic transition is delayed in the sea star compared to the sea urchin, in agreement with the longer cleavage stage in the sea star. Interestingly, the order of gene activation shows the highest variation in the relatively diverged mesodermal circuit, while the correlations of expression dynamics are the highest in the strongly conserved endodermal circuit. We detected loose scaling of the developmental rates of these species and observed interspecies heterochronies within all studied regulatory circuits. Thus, after 500 million years of parallel evolution, mild heterochronies between the species are frequently observed and the tight temporal scaling observed for closely related species no longer holds.

Tilapia and human CLIC2 structures are highly conserved.

Science.gov (United States)

Zeng, Jiao; Li, Zhengjun; Lui, Eei Yin; Lam, Siew Hong; Swaminathan, Kunchithapadam

2018-01-08

Chloride intracellular channels (CLICs) exist in soluble and membrane bound forms. We have determined the crystal structure of soluble Clic2 from the euryhaline teleost fish Oreochromis mossambicus. Structural comparison of tilapia and human CLIC2 with other CLICs shows that these proteins are highly conserved. We have also compared the expression levels of clic2 in selected osmoregulatory organs of tilapia, acclimated to freshwater, seawater and hypersaline water. Structural conservation of vertebrate CLICs implies that they might play conserved roles. Also, tissue-specific responsiveness of clic2 suggests that it might be involved in iono-osmoregulation under extreme conditions in tilapia. Copyright © 2017 Elsevier Inc. All rights reserved.

Communications and Information Sharing (CIS) Laboratory

Data.gov (United States)

Federal Laboratory Consortium — TheCommunications and Information Sharing (CIS) Laboratory is a Public Safety interoperable communications technology laboratory with analog and digital radios, and...

EIA for mining projects in the CIS

Energy Technology Data Exchange (ETDEWEB)

Coppin, N.J.; Wheeler, P. [Wardell Armstrong, Newcastle under Lyme (United Kingdom)

1996-12-31

This paper examines the Environmental Impact Assessment (EIA) requirements and procedures encountered during work on gold and coal mining projects in Kazakhstan, Uzbekistan and Mongolia. Observations on the implementation of former-Soviet inspired EIA in the Commonwealth of Independent States (CIS), and the differences with North American and European requirements and procedures are highlighted, particularly where these indicate lessons for the West. The main implications for mining companies considering or developing projects in the CIS are discussed, particularly the procedures that have to be followed for environmental permitting. 2 figs.

cis,cis-Muconic acid: separation and catalysis to bio-adipic acid for nylon-6,6 polymerization

Energy Technology Data Exchange (ETDEWEB)

Vardon, Derek R.; Rorrer, Nicholas A.; Salvachúa, Davinia; Settle, Amy E.; Johnson, Christopher W.; Menart, Martin J.; Cleveland, Nicholas S.; Ciesielski, Peter N.; Steirer, K. Xerxes; Dorgan, John R.; Beckham, Gregg T.

2016-01-01

cis,cis-Muconic acid is a polyunsaturated dicarboxylic acid that can be produced renewably via the biological conversion of sugars and lignin-derived aromatic compounds. Subsequently, muconic acid can be catalytically converted to adipic acid -- the most commercially significant dicarboxylic acid manufactured from petroleum. Nylon-6,6 is the major industrial application for adipic acid, consuming 85% of market demand; however, high purity adipic acid (99.8%) is required for polymer synthesis. As such, process technologies are needed to effectively separate and catalytically transform biologically derived muconic acid to adipic acid in high purity over stable catalytic materials. To that end, this study: (1) demonstrates bioreactor production of muconate at 34.5 g L-1 in an engineered strain of Pseudomonas putida KT2440, (2) examines the staged recovery of muconic acid from culture media, (3) screens platinum group metals (e.g., Pd, Pt, Rh, Ru) for activity and leaching stability on activated carbon (AC) and silica supports, (4) evaluates the time-on-stream performance of Rh/AC in a trickle bed reactor, and (5) demonstrates the polymerization of bio-adipic acid to nylon-6,6. Separation experiments confirmed AC effectively removed broth color compounds, but subsequent pH/temperature shift crystallization resulted in significant levels of Na, P, K, S and N in the crystallized product. Ethanol dissolution of muconic acid precipitated bulk salts, achieving a purity of 99.8%. Batch catalysis screening reactions determined that Rh and Pd were both highly active compared to Pt and Ru, but Pd leached significantly (1-9%) from both AC and silica supports. Testing of Rh/AC in a continuous trickle bed reactor for 100 h confirmed stable performance after 24 h, although organic adsorption resulted in reduced steady-state activity. Lastly, polymerization of bio-adipic acid with hexamethyldiamine produced nylon-6,6 with comparable properties to its petrochemical counterpart

ECONOMIC AND LEGAL ASPECTS OF CREATION OF AN INTERNATIONAL FINANCIAL CENTER AS IMPORTANT CIS AND EVRASES COUNTRIES INTERGATION FACTOR

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V. S. Balabanov

2011-01-01

Full Text Available The process of forming in Moscow a regional (to be in future transformed into a global international Single Economic Space (SES financial center should become for the Commonwealth of Independent States (CIS and Euroasian Economic Community (EvrAsES countries an universal integration instrument to be used to create their common economic and commercial space. The international center along with SES national financial centers will form an internationally competitive polycentric financial network with single institutional (regulatory, law, customs, etc.agreements. A mechanism should be formed to attract countries outside Customs Union to participate in creation of the international financial center.

Mechanically induced cis to trans reisomerization of azobenzene

Science.gov (United States)

Turansky, Robert; Konopka, Martin; Stich, Ivan; Marx, Dominik

2007-03-01

Using density functional techniques we study mechanochemistry of the azobenzene molecule. Azobenzene is an optically switchable molecule. Laser light is normally used to achieve molecular switching between the cis and trans isomers. We use mechanochemistry to achieve the switching. Thiolate-gold bond can used to exert mechanical energy on the molecule bonded between two gold electrodes in static AFM apparatus. Our model consists of two realistic gold electrodes bridged by dithioazobenzene. We find that pulling the transisomer leads just to formation of gold nanowires and mechanical breakage of the electrodes. However, mechanochemistry with modest applied forces leads to cis trans reisomerization via rotation mechanism. Contrary, use of simple constraints instead of realistic gold electrodes, leads to cis trans reisomerization, albeit with significantly larger applied forces and via inversion mechanism. Important experimental and theoretical ramifications of these simulations will be discussed.

The effect of cis-diammine dichloro platinum(II) on radiation injury in the rat bowel

International Nuclear Information System (INIS)

Lee, Kyung Ja; Rhee, Chung Sik

1995-01-01

This experimental study was performed for evaluate the effects of cis-diamminedichloroplatinum(II) (cis-DDP) on the radiation injury of rat bowel by histopathologic changes. Rats were exposed to entire abdomen by a single doses of X-ray(6-10 Gy) without or with cis-DDP(2.5mg/kg). Rats were divided into 3 groups such as radiation alone, cis-DDP alone and combined group. In combined group, cis-DDP was given 30 minutes before or immediately after irradiation. Cis-DDP induced the inflammatory cell infiltrations with focal necrosis of the mucosa in the small bowel and no abnormal change in the large bowel. In radiation alone group, mucosal necrosis, submucosal fibrosis and muscular necrosis were prominent changes in small bowel and submucosal fibrosis in the large bowel. The submucosal fibrosis in the small bowel was appeared in 10 Gy of radiation alone group and 8 Gy of cis-DDP infusion after radiation and 6 Gy of cis-DDP infusion before radiation of combined group. In the large bowel, submucosal fibrosis was noted in 8 Gy of radiation alone group 8 Gy of cis-DDP infusion after radiation and 6 Gy of cis-DDP infusion before radiation of combined group. In the small bowel, the enhancement ratio was 1.67 in a group of cis-DDP infusion before radiation and 1.25 in group of cis-DDP infusion after radiation as the end point was the submucosal fibrosis. In the large bowel, the enhancement ratio was 1.33 in a group of cis-DDP infusion before radiation and 1.0 in a group of cis-DDP infusion after radiation as the end point was the submucosal fibrosis. This study suggested that cis-DDP enhance the radiation effect in the small and large bowel especially when cis-DDP was infused before radiation

Oxidation reactions of derivatives of cis-octalins promoted by thallium trinitrate (TTN); Reacoes de oxidacao de cis-octalinas promovidas por trinitrato de talio (TTN)

Energy Technology Data Exchange (ETDEWEB)

Ferraz, Helena M.C.; Carneiro, Vania M.T.; Vieira, Tiago O.; Silva Junior, Luiz F. [Universidade de Sao Paulo (USP), SP (Brazil). Inst. de Quimica]. E-mail: luizfsjr@iq.usp.br

2008-07-01

The reaction of ten cis-octalins and cis-octalones with thallium trinitrate (TTN) leads to different products, depending mainly on the substitution pattern of the substrate. Functionalized cis-hydrindanes were obtained from the reaction of 1,2,3,4,4a,5,8,8a-octahydro- 4a-methylnaphthalene and of 1,2,3,4,4a,5,8,8a-octahydro-4a,7-dimethylnaphthalene with TTN in acetonitrile, whereas a cyclic ether was formed treating 1,2,3,4,4a,5,8,8a-octahydro-6,8a-dimethylnaphthalene-1-ol with TTN in trimethylorthoformate (TMOF). (author)

Biodiversity conservation and climate mitigation: What role can economic instruments play?

NARCIS (Netherlands)

Ring, I.; Drechsler, M.; Teeffelen, van A.J.A.; Irawan, S.; Venter, O.

2010-01-01

Tradable permits and intergovernmental fiscal transfers play an increasing role in both biodiversity conservation and climate mitigation. In comparison to regulatory and planning approaches these economic instruments offer a more flexible and cost-effective approach to biodiversity conservation.

Delay-independent stability of genetic regulatory networks.

Science.gov (United States)

Wu, Fang-Xiang

2011-11-01

Genetic regulatory networks can be described by nonlinear differential equations with time delays. In this paper, we study both locally and globally delay-independent stability of genetic regulatory networks, taking messenger ribonucleic acid alternative splicing into consideration. Based on nonnegative matrix theory, we first develop necessary and sufficient conditions for locally delay-independent stability of genetic regulatory networks with multiple time delays. Compared to the previous results, these conditions are easy to verify. Then we develop sufficient conditions for global delay-independent stability for genetic regulatory networks. Compared to the previous results, this sufficient condition is less conservative. To illustrate theorems developed in this paper, we analyze delay-independent stability of two genetic regulatory networks: a real-life repressilatory network with three genes and three proteins, and a synthetic gene regulatory network with five genes and seven proteins. The simulation results show that the theorems developed in this paper can effectively determine the delay-independent stability of genetic regulatory networks.

Differential trypanosome surface coat regulation by a CCCH protein that co-associates with procyclin mRNA cis-elements.

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Pegine Walrad

2009-02-01

Full Text Available The genome of Trypanosoma brucei is unusual in being regulated almost entirely at the post-transcriptional level. In terms of regulation, the best-studied genes are procyclins, which encode a family of major surface GPI-anchored glycoproteins (EP1, EP2, EP3, GPEET that show differential expression in the parasite's tsetse-fly vector. Although procyclin mRNA cis-regulatory sequences have provided the paradigm for post-transcriptional control in kinetoplastid parasites, trans-acting regulators of procyclin mRNAs are unidentified, despite intensive effort over 15 years. Here we identify the developmental regulator, TbZFP3, a CCCH-class predicted RNA binding protein, as an isoform-specific regulator of Procyclin surface coat expression in trypanosomes. We demonstrate (i that endogenous TbZFP3 shows sequence-specific co-precipitation of EP1 and GPEET, but not EP2 and EP3, procyclin mRNA isoforms, (ii that ectopic overexpression of TbZFP3 does not perturb the mRNA abundance of procyclin transcripts, but rather that (iii their protein expression is regulated in an isoform-specific manner, as evidenced by mass spectrometric analysis of the Procyclin expression signature in the transgenic cell lines. The TbZFP3 mRNA-protein complex (TbZFP3mRNP is identified as a trans-regulator of differential surface protein expression in trypanosomes. Moreover, its sequence-specific interactions with procyclin mRNAs are compatible with long-established predictions for Procyclin regulation. Combined with the known association of TbZFP3 with the translational apparatus, this study provides a long-sought missing link between surface protein cis-regulatory signals and the gene expression machinery in trypanosomes.

Ancient Pbx-Hox signatures define hundreds of vertebrate developmental enhancers

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Parker Hugo J

2011-12-01

Full Text Available Abstract Background Gene regulation through cis-regulatory elements plays a crucial role in development and disease. A major aim of the post-genomic era is to be able to read the function of cis-regulatory elements through scrutiny of their DNA sequence. Whilst comparative genomics approaches have identified thousands of putative regulatory elements, our knowledge of their mechanism of action is poor and very little progress has been made in systematically de-coding them. Results Here, we identify ancient functional signatures within vertebrate conserved non-coding elements (CNEs through a combination of phylogenetic footprinting and functional assay, using genomic sequence from the sea lamprey as a reference. We uncover a striking enrichment within vertebrate CNEs for conserved binding-site motifs of the Pbx-Hox hetero-dimer. We further show that these predict reporter gene expression in a segment specific manner in the hindbrain and pharyngeal arches during zebrafish development. Conclusions These findings evoke an evolutionary scenario in which many CNEs evolved early in the vertebrate lineage to co-ordinate Hox-dependent gene-regulatory interactions that pattern the vertebrate head. In a broader context, our evolutionary analyses reveal that CNEs are composed of tightly linked transcription-factor binding-sites (TFBSs, which can be systematically identified through phylogenetic footprinting approaches. By placing a large number of ancient vertebrate CNEs into a developmental context, our findings promise to have a significant impact on efforts toward de-coding gene-regulatory elements that underlie vertebrate development, and will facilitate building general models of regulatory element evolution.

Repetitive Elements in Mycoplasma hyopneumoniae Transcriptional Regulation.

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Amanda Malvessi Cattani

Full Text Available Transcriptional regulation, a multiple-step process, is still poorly understood in the important pig pathogen Mycoplasma hyopneumoniae. Basic motifs like promoters and terminators have already been described, but no other cis-regulatory elements have been found. DNA repeat sequences have been shown to be an interesting potential source of cis-regulatory elements. In this work, a genome-wide search for tandem and palindromic repetitive elements was performed in the intergenic regions of all coding sequences from M. hyopneumoniae strain 7448. Computational analysis demonstrated the presence of 144 tandem repeats and 1,171 palindromic elements. The DNA repeat sequences were distributed within the 5' upstream regions of 86% of transcriptional units of M. hyopneumoniae strain 7448. Comparative analysis between distinct repetitive sequences found in related mycoplasma genomes demonstrated different percentages of conservation among pathogenic and nonpathogenic strains. qPCR assays revealed differential expression among genes showing variable numbers of repetitive elements. In addition, repeats found in 206 genes already described to be differentially regulated under different culture conditions of M. hyopneumoniae strain 232 showed almost 80% conservation in relation to M. hyopneumoniae strain 7448 repeats. Altogether, these findings suggest a potential regulatory role of tandem and palindromic DNA repeats in the M. hyopneumoniae transcriptional profile.

Repetitive Elements in Mycoplasma hyopneumoniae Transcriptional Regulation.

Science.gov (United States)

Cattani, Amanda Malvessi; Siqueira, Franciele Maboni; Guedes, Rafael Lucas Muniz; Schrank, Irene Silveira

2016-01-01

Transcriptional regulation, a multiple-step process, is still poorly understood in the important pig pathogen Mycoplasma hyopneumoniae. Basic motifs like promoters and terminators have already been described, but no other cis-regulatory elements have been found. DNA repeat sequences have been shown to be an interesting potential source of cis-regulatory elements. In this work, a genome-wide search for tandem and palindromic repetitive elements was performed in the intergenic regions of all coding sequences from M. hyopneumoniae strain 7448. Computational analysis demonstrated the presence of 144 tandem repeats and 1,171 palindromic elements. The DNA repeat sequences were distributed within the 5' upstream regions of 86% of transcriptional units of M. hyopneumoniae strain 7448. Comparative analysis between distinct repetitive sequences found in related mycoplasma genomes demonstrated different percentages of conservation among pathogenic and nonpathogenic strains. qPCR assays revealed differential expression among genes showing variable numbers of repetitive elements. In addition, repeats found in 206 genes already described to be differentially regulated under different culture conditions of M. hyopneumoniae strain 232 showed almost 80% conservation in relation to M. hyopneumoniae strain 7448 repeats. Altogether, these findings suggest a potential regulatory role of tandem and palindromic DNA repeats in the M. hyopneumoniae transcriptional profile.

Cis-bifenthrin induces immunotoxicity in adolescent male C57BL/6 mice.

Science.gov (United States)

Wang, Xia; Gao, Xingli; He, Bingnan; Zhu, Jiawei; Lou, Huihui; Hu, Qinglian; Jin, Yuanxiang; Fu, Zhengwei

2017-07-01

Bifenthrin (BF) is an important synthetic pyrethroid. Previous studies have demonstrated that cis-BF exhibits toxic effects on development, the neurological, reproductive and endocrine system. In this study, we evaluated the immunotoxicity caused by cis-BF in adolescent male C57BL/6 mice. Mice were exposed orally to 0, 5, 10, and 20 mg/kg/d for 3 weeks. The results showed that body weight, spleen weight, and splenic cellularity decreased in mice exposed to 20 mg/kg/d cis-BF. Additionally, we found that the mRNA levels of the pro-inflammatory factors IL-1β, IL-6, CXCL-1, and TNF-α, in peritoneal macrophages, the spleen, and the thymus were inhibited in the cis-BF-treated groups. Moreover, MTT assays demonstrated that cis-BF inhibited splenocyte proliferation stimulated by LPS or Con A, as well as the secretion of IFN-γ on Con A stimulation. Collectively, the results of this study suggest that exposure to cis-BF has the potential to induce immunotoxicity in adolescent male C57BL/6 mice. © 2017 Wiley Periodicals, Inc.

Innate immune activity conditions the effect of regulatory variants upon monocyte gene expression.

Science.gov (United States)

Fairfax, Benjamin P; Humburg, Peter; Makino, Seiko; Naranbhai, Vivek; Wong, Daniel; Lau, Evelyn; Jostins, Luke; Plant, Katharine; Andrews, Robert; McGee, Chris; Knight, Julian C

2014-03-07

To systematically investigate the impact of immune stimulation upon regulatory variant activity, we exposed primary monocytes from 432 healthy Europeans to interferon-γ (IFN-γ) or differing durations of lipopolysaccharide and mapped expression quantitative trait loci (eQTLs). More than half of cis-eQTLs identified, involving hundreds of genes and associated pathways, are detected specifically in stimulated monocytes. Induced innate immune activity reveals multiple master regulatory trans-eQTLs including the major histocompatibility complex (MHC), coding variants altering enzyme and receptor function, an IFN-β cytokine network showing temporal specificity, and an interferon regulatory factor 2 (IRF2) transcription factor-modulated network. Induced eQTL are significantly enriched for genome-wide association study loci, identifying context-specific associations to putative causal genes including CARD9, ATM, and IRF8. Thus, applying pathophysiologically relevant immune stimuli assists resolution of functional genetic variants.

A transcription factor collective defines the HSN serotonergic neuron regulatory landscape.

Science.gov (United States)

Lloret-Fernández, Carla; Maicas, Miren; Mora-Martínez, Carlos; Artacho, Alejandro; Jimeno-Martín, Ángela; Chirivella, Laura; Weinberg, Peter; Flames, Nuria

2018-03-22

Cell differentiation is controlled by individual transcription factors (TFs) that together activate a selection of enhancers in specific cell types. How these combinations of TFs identify and activate their target sequences remains poorly understood. Here, we identify the cis -regulatory transcriptional code that controls the differentiation of serotonergic HSN neurons in Caenorhabditis elegans . Activation of the HSN transcriptome is directly orchestrated by a collective of six TFs. Binding site clusters for this TF collective form a regulatory signature that is sufficient for de novo identification of HSN neuron functional enhancers. Among C. elegans neurons, the HSN transcriptome most closely resembles that of mouse serotonergic neurons. Mouse orthologs of the HSN TF collective also regulate serotonergic differentiation and can functionally substitute for their worm counterparts which suggests deep homology. Our results identify rules governing the regulatory landscape of a critically important neuronal type in two species separated by over 700 million years. © 2018, Lloret-Fernández et al.

77 FR 58022 - Montana Regulatory Program

Science.gov (United States)

2012-09-19

... precludes in situ gasification projects from including carbon capture and sequestration (CCS) under the... Conservation as the regulatory authority for CCS activities within the State. SB498 generally established that..., the Board would regulate any proposed CCS activities appropriately. CCS operations have potential...

Inhibition of Hepatitis C Virus in Mice by a Small Interfering RNA Targeting a Highly Conserved Sequence in Viral IRES Pseudoknot.

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Jae-Su Moon

Full Text Available The hepatitis C virus (HCV internal ribosome entry site (IRES that directs cap-independent viral translation is a primary target for small interfering RNA (siRNA-based HCV antiviral therapy. However, identification of potent siRNAs against HCV IRES by bioinformatics-based siRNA design is a challenging task given the complexity of HCV IRES secondary and tertiary structures and association with multiple proteins, which can also dynamically change the structure of this cis-acting RNA element. In this work, we utilized siRNA tiling approach whereby siRNAs were tiled with overlapping sequences that were shifted by one or two nucleotides over the HCV IRES stem-loop structures III and IV spanning nucleotides (nts 277-343. Based on their antiviral activity, we mapped a druggable region (nts 313-343 where the targets of potent siRNAs were enriched. siIE22, which showed the greatest anti-HCV potency, targeted a highly conserved sequence across diverse HCV genotypes, locating within the IRES subdomain IIIf involved in pseudoknot formation. Stepwise target shifting toward the 5' or 3' direction by 1 or 2 nucleotides reduced the antiviral potency of siIE22, demonstrating the importance of siRNA accessibility to this highly structured and sequence-conserved region of HCV IRES for RNA interference. Nanoparticle-mediated systemic delivery of the stability-improved siIE22 derivative gs_PS1 siIE22, which contains a single phosphorothioate linkage on the guide strand, reduced the serum HCV genome titer by more than 4 log10 in a xenograft mouse model for HCV replication without generation of resistant variants. Our results provide a strategy for identifying potent siRNA species against a highly structured RNA target and offer a potential pan-HCV genotypic siRNA therapy that might be beneficial for patients resistant to current treatment regimens.

Fused Regression for Multi-source Gene Regulatory Network Inference.

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Kari Y Lam

2016-12-01

Full Text Available Understanding gene regulatory networks is critical to understanding cellular differentiation and response to external stimuli. Methods for global network inference have been developed and applied to a variety of species. Most approaches consider the problem of network inference independently in each species, despite evidence that gene regulation can be conserved even in distantly related species. Further, network inference is often confined to single data-types (single platforms and single cell types. We introduce a method for multi-source network inference that allows simultaneous estimation of gene regulatory networks in multiple species or biological processes through the introduction of priors based on known gene relationships such as orthology incorporated using fused regression. This approach improves network inference performance even when orthology mapping and conservation are incomplete. We refine this method by presenting an algorithm that extracts the true conserved subnetwork from a larger set of potentially conserved interactions and demonstrate the utility of our method in cross species network inference. Last, we demonstrate our method's utility in learning from data collected on different experimental platforms.

Inference of gene regulatory networks with sparse structural equation models exploiting genetic perturbations.

Directory of Open Access Journals (Sweden)

Xiaodong Cai

Full Text Available Integrating genetic perturbations with gene expression data not only improves accuracy of regulatory network topology inference, but also enables learning of causal regulatory relations between genes. Although a number of methods have been developed to integrate both types of data, the desiderata of efficient and powerful algorithms still remains. In this paper, sparse structural equation models (SEMs are employed to integrate both gene expression data and cis-expression quantitative trait loci (cis-eQTL, for modeling gene regulatory networks in accordance with biological evidence about genes regulating or being regulated by a small number of genes. A systematic inference method named sparsity-aware maximum likelihood (SML is developed for SEM estimation. Using simulated directed acyclic or cyclic networks, the SML performance is compared with that of two state-of-the-art algorithms: the adaptive Lasso (AL based scheme, and the QTL-directed dependency graph (QDG method. Computer simulations demonstrate that the novel SML algorithm offers significantly better performance than the AL-based and QDG algorithms across all sample sizes from 100 to 1,000, in terms of detection power and false discovery rate, in all the cases tested that include acyclic or cyclic networks of 10, 30 and 300 genes. The SML method is further applied to infer a network of 39 human genes that are related to the immune function and are chosen to have a reliable eQTL per gene. The resulting network consists of 9 genes and 13 edges. Most of the edges represent interactions reasonably expected from experimental evidence, while the remaining may just indicate the emergence of new interactions. The sparse SEM and efficient SML algorithm provide an effective means of exploiting both gene expression and perturbation data to infer gene regulatory networks. An open-source computer program implementing the SML algorithm is freely available upon request.

Zirconium-doped magnetic microspheres for the selective enrichment of cis-diol-containing ribonucleosides.

Science.gov (United States)

Fan, Hua; Chen, Peihong; Wang, Chaozhan; Wei, Yinmao

2016-05-27

Zirconium-doped magnetic microspheres (Zr-Fe3O4) for the selective enrichment of cis-diol-containing biomolecules were easily synthesized via a one-step hydrothermal method. Characterization of the microspheres revealed that zirconium was successfully doped into the lattice of Fe3O4 at a doping level of 4.0 at%. Zr-Fe3O4 possessed good magnetic properties and high specificity towards cis-diol molecules, as shown using 28 compounds. For ribonucleosides, the adsorbent not only has favorable anti-interferential abilities but also has a high adsorption capacity up to 159.4μmol/g. As an example of a real application, four ribonucleosides in urine were efficiently enriched and detected via magnetic solid-phase extraction coupled with high-performance liquid chromatography. Under the optimized extraction conditions, the detection limits were determined to be between 0.005 and 0.017μg/mL, and the linearities ranged from 0.02 to 5.00μg/mL (R≥0.996) for these analytes. The accuracy of the analytical method was examined by studying the relative recoveries of the analytes in real urine samples, with recoveries varying from 77.8% to 119.6% (RSDs<10.6%, n=6). The results indicate that Zr-Fe3O4 is a suitable adsorbent for the analysis of cis-diol-containing biomolecules in practical applications. Copyright © 2016 Elsevier B.V. All rights reserved.

Enhanced binding capacity of boronate affinity adsorbent via surface modification of silica by combination of atom transfer radical polymerization and chain-end functionalization for high-efficiency enrichment of cis-diol molecules

Energy Technology Data Exchange (ETDEWEB)

Wang, Wei; He, Maofang; Wang, Chaozhan; Wei, Yinmao, E-mail: ymwei@nwu.edu.cn

2015-07-30

Boronate affinity materials have been widely used for specific separation and preconcentration of cis-diol molecules, but most do not have sufficient capacity due to limited binding sites on the material surface. In this work, we prepared a phenylboronic acid-functionalized adsorbent with a high binding capacity via the combination of surface-initiated atom transfer radical polymerization (SI-ATRP) and chain-end functionalization. With this method, the terminal chlorides of the polymer chains were used fully, and the proposed adsorbent contains dense boronic acid polymers chain with boronic acid on the chain end. Consequently, the proposed adsorbent possesses excellent selectivity and a high binding capacity of 513.6 μmol g{sup −1} for catechol and 736.8 μmol g{sup −1} for fructose, which are much higher than those of other reported adsorbents. The dispersed solid-phase extraction (dSPE) based on the prepared adsorbent was used for extraction of three cis-diol drugs (i.e., epinephrine, isoprenaline and caffeic acid isopropyl ester) from plasma; the eluates were analyzed by HPLC-UV. The reduced amount of adsorbent (i.e., 2.0 mg) could still eliminate interferences efficiently and yielded a recovery range of 85.6–101.1% with relative standard deviations ranging from 2.5 to 9.7% (n = 5). The results indicated that the proposed strategy could serve as a promising alternative to increase the density of surface functional groups on the adsorbent; thus, the prepared adsorbent has the potential to effectively enrich cis-diol substances in real samples. - Highlights: • Boronate adsorbent is prepared via ATRP and chain-end functionalization. • The adsorbent has quite high binding capacity for cis-diols. • Binding capacity is easily manipulated by ATRP condition. • Chain-end functionalization can improve binding capacity significantly. • Reduced adsorbent is consumed in dispersed solid-phase extraction of cis-diols.

An in vivo comparison of cis- and trans-[{sup 18}F]mefway in the nonhuman primate

Energy Technology Data Exchange (ETDEWEB)

Wooten, Dustin, E-mail: dwooten@wisc.edu [Department of Medical Physics, Waisman Laboratory for Brain Imaging and Behavior, University of Wisconsin-Madison, Madison, WI 53705 (United States); Hillmer, Ansel; Murali, Dhanabalan; Barnhart, Todd [Department of Medical Physics, Waisman Laboratory for Brain Imaging and Behavior, University of Wisconsin-Madison, Madison, WI 53705 (United States); Schneider, Mary L. [Department of Kinesiology, University of Wisconsin-Madison, Madison, WI 53706 (United States); Mukherjee, Jogeshwar [Preclinical Imaging Center, Department of Psychiatry and Human Behavior, University of California-Irvine, Irvine 92697 (United States); Christian, Bradley T. [Department of Medical Physics, Waisman Laboratory for Brain Imaging and Behavior, University of Wisconsin-Madison, Madison, WI 53705 (United States)

2011-10-15

Introduction: [{sup 18}F]Mefway is a serotonin 5-HT{sub 1A} PET radiotracer with high specificity and favorable in vivo imaging properties. The chemical structure of [{sup 18}F]mefway permits {sup 18}F labeling in either the cis or trans positions at the 4-cyclohexyl site. We have previously reported on the in vivo kinetics of trans-[{sup 18}F]mefway in the nonhuman primate. In this work, we compare the in vivo binding of cis-[{sup 18}F]mefway and trans-[{sup 18}F]mefway to evaluate the properties of cis-[{sup 18}F]mefway for 5-HT{sub 1A} PET imaging. Methods: The cis- and trans-[{sup 18}F]mefway tracers were synthesized via nucleophilic substitution with their respective tosyl precursors. Two monkeys (one male, one female) were given bolus injections of both cis- and trans-labeled [{sup 18}F]mefway in separate experiments. Dynamic scans were acquired for 90 min with a microPET P4 scanner. Time-activity curves were extracted in the areas of the mesial temporal cortex (MTC), anterior cingulate gyrus (aCG), insular cortex (IC), raphe nuclei (RN) and cerebellum (CB). The in vivo behavior of the radiotracers was compared based upon the nondisplaceable binding potential (BP{sub ND}) using the CB as a reference region. Results: Averaged over the two subjects, BP{sub ND} values were as follows: MTC: 7.7, 0.58; aCG: 4.95, 0.32; IC: 3.27, 0.2; and RN: 3.05, 0.13, for trans-[{sup 18}F]mefway and cis-[{sup 18}F]mefway, respectively. Conclusion: The cis-labeled [{sup 18}F]mefway tracer has low specific binding throughout the 5-HT{sub 1A} regions of the brain compared to trans-[{sup 18}F]mefway, suggesting that the target-to-background binding of cis-[{sup 18}F]mefway may limit its use for in vivo assessment of 5-HT{sub 1A} binding.

C.I.S. natural gas-1

International Nuclear Information System (INIS)

Carson, M.; Stram, B.

1993-01-01

This paper reports that in the countries that make up the Commonwealth of Independent States (C.I.S.), with their vast resources and a considerable existing production base, prospects are good for further growth of the region's exportable gas surplus. Investment fundamentals are stronger for gas than for any other energy resources in the area. But the pipeline infrastructure to move large amounts of gas will need extensive refurbishment to ensure export reliability and growth. Given the potential in terms of production and markets, significant amounts of outside investment in oil, natural gas, and NGL infrastructure will likely increase dramatically in these countries in the near future. These are some of the major conclusions of Enron Corp.'s recent investigations in the C.I.S. and other former Soviet republics

Hijacking Complement Regulatory Proteins for Bacterial Immune Evasion.

Science.gov (United States)

Hovingh, Elise S; van den Broek, Bryan; Jongerius, Ilse

2016-01-01

The human complement system plays an important role in the defense against invading pathogens, inflammation and homeostasis. Invading microbes, such as bacteria, directly activate the complement system resulting in the formation of chemoattractants and in effective labeling of the bacteria for phagocytosis. In addition, formation of the membrane attack complex is responsible for direct killing of Gram-negative bacteria. In turn, bacteria have evolved several ways to evade complement activation on their surface in order to be able to colonize and invade the human host. One important mechanism of bacterial escape is attraction of complement regulatory proteins to the microbial surface. These molecules are present in the human body for tight regulation of the complement system to prevent damage to host self-surfaces. Therefore, recruitment of complement regulatory proteins to the bacterial surface results in decreased complement activation on the microbial surface which favors bacterial survival. This review will discuss recent advances in understanding the binding of complement regulatory proteins to the bacterial surface at the molecular level. This includes, new insights that have become available concerning specific conserved motives on complement regulatory proteins that are favorable for microbial binding. Finally, complement evasion molecules are of high importance for vaccine development due to their dominant role in bacterial survival, high immunogenicity and homology as well as their presence on the bacterial surface. Here, the use of complement evasion molecules for vaccine development will be discussed.

Preparation and characterization of fluorophenylboronic acid-functionalized affinity monolithic columns for the selective enrichment of cis-diol-containing biomolecules.

Science.gov (United States)

Li, Qianjin; Liu, Zhen

2015-01-01

Boronate affinity monolithic columns have been developed into an important means for the selective recognition and capture of cis-diol-containing biomolecules, such as glycoproteins, nucleosides and saccharides. The ligands of boronic acids are playing an important role in boronate affinity monolithic columns. Although several boronate affinity monoliths with high affinity toward cis-diol-containing biomolecules have been reported, only few publications are focused on their detailed procedures for preparation and characterization. This chapter describes in detail the preparation and characterization of a boronate affinity monolithic column applying 2,4-difluoro-3-formyl-phenylboronic acid (DFFPBA) as a ligand. The DFFPBA-functionalized monolithic column not only exhibited an ultrahigh boronate affinity toward cis-diol-containing biomolecules, but also showed great potential for the selective enrichment of cis-diol-containing biomolecules in real samples.

Evolutionary analysis reveals regulatory and functional landscape of coding and non-coding RNA editing.

Science.gov (United States)

Zhang, Rui; Deng, Patricia; Jacobson, Dionna; Li, Jin Billy

2017-02-01

Adenosine-to-inosine RNA editing diversifies the transcriptome and promotes functional diversity, particularly in the brain. A plethora of editing sites has been recently identified; however, how they are selected and regulated and which are functionally important are largely unknown. Here we show the cis-regulation and stepwise selection of RNA editing during Drosophila evolution and pinpoint a large number of functional editing sites. We found that the establishment of editing and variation in editing levels across Drosophila species are largely explained and predicted by cis-regulatory elements. Furthermore, editing events that arose early in the species tree tend to be more highly edited in clusters and enriched in slowly-evolved neuronal genes, thus suggesting that the main role of RNA editing is for fine-tuning neurological functions. While nonsynonymous editing events have been long recognized as playing a functional role, in addition to nonsynonymous editing sites, a large fraction of 3'UTR editing sites is evolutionarily constrained, highly edited, and thus likely functional. We find that these 3'UTR editing events can alter mRNA stability and affect miRNA binding and thus highlight the functional roles of noncoding RNA editing. Our work, through evolutionary analyses of RNA editing in Drosophila, uncovers novel insights of RNA editing regulation as well as its functions in both coding and non-coding regions.

Multi-arrangement quantum dynamics in 6D: cis-trans isomerization and 1,3-hydrogen transfer in HONO

International Nuclear Information System (INIS)

Luckhaus, David

2004-01-01

The overtone spectrum and wave packet dynamics of nitrous acid (HONO) are studied with a global six-dimensional potential energy function interpolated directly from density functional calculations together with the corresponding dipole hypersurfaces. The quantum dynamics for the cis-trans isomerization and the symmetric 1,3-hydrogen transfer are treated in full dimensionality in terms of the generalized Z-matrix discrete variable representation. For the quantum mechanical description of complicated rearrangements a new approach to multi-arrangement quantum dynamics is introduced and applied to the symmetric hydrogen exchange tunneling in cis-HONO. The cis-trans isomerization is found to be dominated by adiabatic barrier crossing with only minor tunneling contributions, but with pronounced mode selectivity. The OH-stretching overtones of trans-HONO are adiabatically almost completely separated from the OH torsional dynamics with extremely slow intramolecular energy redistribution. The 1,3-hydrogen transfer, by contrast, proceeds largely via coherent tunneling even significantly below the barrier. The process is clearly non-adiabatic (at least in terms of valence coordinates) but remains highly state specific. While the absorption spectrum of trans-HONO remains largely unaffected, OH-stretching overtones of cis-HONO (above the barrier between 2ν OH and 3ν OH ) decompose into highly fragmented absorption patterns with corresponding tunneling periods on the picosecond time scale

High-resolution satellite imagery is an important yet underutilized resource in conservation biology.

Science.gov (United States)

Boyle, Sarah A; Kennedy, Christina M; Torres, Julio; Colman, Karen; Pérez-Estigarribia, Pastor E; de la Sancha, Noé U

2014-01-01

Technological advances and increasing availability of high-resolution satellite imagery offer the potential for more accurate land cover classifications and pattern analyses, which could greatly improve the detection and quantification of land cover change for conservation. Such remotely-sensed products, however, are often expensive and difficult to acquire, which prohibits or reduces their use. We tested whether imagery of high spatial resolution (≤5 m) differs from lower-resolution imagery (≥30 m) in performance and extent of use for conservation applications. To assess performance, we classified land cover in a heterogeneous region of Interior Atlantic Forest in Paraguay, which has undergone recent and dramatic human-induced habitat loss and fragmentation. We used 4 m multispectral IKONOS and 30 m multispectral Landsat imagery and determined the extent to which resolution influenced the delineation of land cover classes and patch-level metrics. Higher-resolution imagery more accurately delineated cover classes, identified smaller patches, retained patch shape, and detected narrower, linear patches. To assess extent of use, we surveyed three conservation journals (Biological Conservation, Biotropica, Conservation Biology) and found limited application of high-resolution imagery in research, with only 26.8% of land cover studies analyzing satellite imagery, and of these studies only 10.4% used imagery ≤5 m resolution. Our results suggest that high-resolution imagery is warranted yet under-utilized in conservation research, but is needed to adequately monitor and evaluate forest loss and conversion, and to delineate potentially important stepping-stone fragments that may serve as corridors in a human-modified landscape. Greater access to low-cost, multiband, high-resolution satellite imagery would therefore greatly facilitate conservation management and decision-making.

78 FR 14840 - U.S.-EU High Level Regulatory Cooperation Forum-Stakeholder Session

Science.gov (United States)

2013-03-07

... OFFICE OF MANAGEMENT AND BUDGET U.S.-EU High Level Regulatory Cooperation Forum--Stakeholder... the future. Time constraints may prevent some stakeholders from making presentations on April 10th or... stakeholder session agenda will in no way prejudge the specific elements of a future transatlantic regulatory...

Challenges posed to the European pharmaceutical regulatory system by highly personalized medicines.

Science.gov (United States)

Johnston, John D; Feldschreiber, Peter

2014-03-01

The European pharmaceutical regulatory system has not yet been challenged by issues related to highly personalized medicines such as those to be found with active substances that affect RNA biochemistry. We review the current status of RNA-based pharmacology and present three possible case histories. The implications for the European pharmaceutical regulatory system are discussed. © 2013 The British Pharmacological Society.

Unique ATPase site architecture triggers cis-mediated synchronized ATP binding in heptameric AAA+-ATPase domain of flagellar regulatory protein FlrC.

Science.gov (United States)

Dey, Sanjay; Biswas, Maitree; Sen, Udayaditya; Dasgupta, Jhimli

2015-04-03

Bacterial enhancer-binding proteins (bEBPs) oligomerize through AAA(+) domains and use ATP hydrolysis-driven energy to isomerize the RNA polymerase-σ(54) complex during transcriptional initiation. Here, we describe the first structure of the central AAA(+) domain of the flagellar regulatory protein FlrC (FlrC(C)), a bEBP that controls flagellar synthesis in Vibrio cholerae. Our results showed that FlrC(C) forms heptamer both in nucleotide (Nt)-free and -bound states without ATP-dependent subunit remodeling. Unlike the bEBPs such as NtrC1 or PspF, a novel cis-mediated "all or none" ATP binding occurs in the heptameric FlrC(C), because constriction at the ATPase site, caused by loop L3 and helix α7, restricts the proximity of the trans-protomer required for Nt binding. A unique "closed to open" movement of Walker A, assisted by trans-acting "Glu switch" Glu-286, facilitates ATP binding and hydrolysis. Fluorescence quenching and ATPase assays on FlrC(C) and mutants revealed that although Arg-349 of sensor II, positioned by trans-acting Glu-286 and Tyr-290, acts as a key residue to bind and hydrolyze ATP, Arg-319 of α7 anchors ribose and controls the rate of ATP hydrolysis by retarding the expulsion of ADP. Heptameric state of FlrC(C) is restored in solution even with the transition state mimicking ADP·AlF3. Structural results and pulldown assays indicated that L3 renders an in-built geometry to L1 and L2 causing σ(54)-FlrC(C) interaction independent of Nt binding. Collectively, our results underscore a novel mechanism of ATP binding and σ(54) interaction that strives to understand the transcriptional mechanism of the bEBPs, which probably interact directly with the RNA polymerase-σ(54) complex without DNA looping. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Questions on maintenance of ecological safety in space of the CIS

International Nuclear Information System (INIS)

Maharramov, A.A.

2009-01-01

Disintegration of the USSR has led to disintegration uniform economic and legal space and it has led to occurrence of some problems in ecological sphere. In given article it is shown some directions of maintenance of ecological safety, agreements on this sphere and organizational mechanisms, forms of mutual relations of the CIS countries in this sphere. At the end it is shownthe basic directions on maintenance of ecological safety in framework of CIS and elements of development of mutual relations of the CIS countries

Chromatin Heterogeneity and Distribution of Regulatory Elements in the Late-Replicating Intercalary Heterochromatin Domains of Drosophila melanogaster Chromosomes.

Directory of Open Access Journals (Sweden)

Varvara A Khoroshko

Full Text Available Late-replicating domains (intercalary heterochromatin in the Drosophila genome display a number of features suggesting their organization is quite unique. Typically, they are quite large and encompass clusters of functionally unrelated tissue-specific genes. They correspond to the topologically associating domains and conserved microsynteny blocks. Our study aims at exploring further details of molecular organization of intercalary heterochromatin and has uncovered surprising heterogeneity of chromatin composition in these regions. Using the 4HMM model developed in our group earlier, intercalary heterochromatin regions were found to host chromatin fragments with a particular epigenetic profile. Aquamarine chromatin fragments (spanning 0.67% of late-replicating regions are characterized as a class of sequences that appear heterogeneous in terms of their decompactization. These fragments are enriched with enhancer sequences and binding sites for insulator proteins. They likely mark the chromatin state that is related to the binding of cis-regulatory proteins. Malachite chromatin fragments (11% of late-replicating regions appear to function as universal transitional regions between two contrasting chromatin states. Namely, they invariably delimit intercalary heterochromatin regions from the adjacent active chromatin of interbands. Malachite fragments also flank aquamarine fragments embedded in the repressed chromatin of late-replicating regions. Significant enrichment of insulator proteins CP190, SU(HW, and MOD2.2 was observed in malachite chromatin. Neither aquamarine nor malachite chromatin types appear to correlate with the positions of highly conserved non-coding elements (HCNE that are typically replete in intercalary heterochromatin. Malachite chromatin found on the flanks of intercalary heterochromatin regions tends to replicate earlier than the malachite chromatin embedded in intercalary heterochromatin. In other words, there exists a

Initial conservative treatment for grade 3 Ta-1 superficial bladder cancer

International Nuclear Information System (INIS)

Fujimoto, Kiyohide; Chihara, Yoshitomo; Kondo, Hideaki; Hirao, Yoshihiko

2006-01-01

We retrospectively investigated the therapeutic outcomes of our series of 7 Ta and 62 T1 bladder cancers with grade 3 (G3) malignancy in 61 men and 8 women having a mean age of 66.2 years. Following transurethral resection of bladder tumor (TURBT), 35 and 6 patients received intravesical instillations of bacillus Calmette-Guerin (BCG) and anthracycline-derivants, respectively, whereas 15 received no adjuvant therapy. Five and 2 patients received systemic and local chemotherapy with irradiation, respectively, and six underwent radical cystectomy for invasive potential. The 5-year non-recurrence, progression-free, and overall (cancer-specific) survival rates were 66, 82%, and 76 (88)%, respectively, after a median follow-up of 52 months. The 5-year non-recurrence rates were 24% in non-adjuvant, 85% in BCG, 0% in anthracycline-derivants, 65% in systemic and local chemoradiation therapy, and 68% in cystectomy. The 5-year progression-free and overall (cancer-specific) survival rates of the patients treated with BCG instillation were 91% and 94 (100)%. There were no significant differences in the 5-year non-recurrence and progression-free rates between 12 patients with carcinoma in situ (CIS) and 23 patients without CIS. Complete TUR of all visible tumors and a reliable histopathological diagnosis of appropriate specimens bearing the muscle layer are mandatory for assessment of recurrence. G3 Ta-1 bladder cancers and CIS showed a high risk of recurrence, and required aggressive treatment. Since BCG therapy following TURBT significantly reduced the risk of recurrence and progression, adjuvant BCG therapy is considered to be the most promising initial conservative treatment for G3 Ta-1 bladder cancers. (author)

Reconstruction of the yeast Snf1 kinase regulatory network reveals its role as a global energy regulator

DEFF Research Database (Denmark)

Usaite, Renata; Jewett, Michael Christopher; Soberano de Oliveira, Ana Paula

2009-01-01

Highly conserved among eukaryotic cells, the AMP-activated kinase (AMPK) is a central regulator of carbon metabolism. To map the complete network of interactions around AMPK in yeast (Snf1) and to evaluate the role of its regulatory subunit Snf4, we measured global mRNA, protein and metabolite...

In Silico Expression Analysis.

Science.gov (United States)

Bolívar, Julio; Hehl, Reinhard; Bülow, Lorenz

2016-01-01

Information on the specificity of cis-sequences enables the design of functional synthetic plant promoters that are responsive to specific stresses. Potential cis-sequences may be experimentally tested, however, correlation of genomic sequence with gene expression data enables an in silico expression analysis approach to bioinformatically assess the stress specificity of candidate cis-sequences prior to experimental verification. The present chapter demonstrates an example for the in silico validation of a potential cis-regulatory sequence responsive to cold stress. The described online tool can be applied for the bioinformatic assessment of cis-sequences responsive to most abiotic and biotic stresses of plants. Furthermore, a method is presented based on a reverted in silico expression analysis approach that predicts highly specific potentially functional cis-regulatory elements for a given stress.

Mutation of the regulatory phosphorylation site of tobacco nitrate reductase results in high nitrite excretion and NO emission from leaf and root tissue

NARCIS (Netherlands)

Lea, US; ten Hoopen, F; Provan, F; Kaiser, WM; Meyer, C; Lillo, C

In wild-type Nicotiana plumbaginifolia Viv. and other higher plants, nitrate reductase (NR) is regulated at the post-translational level and is rapidly inactivated in response to, for example, a light-to-dark transition. This inactivation is caused by phosphorylation of a conserved regulatory serine

Action of cis-dichlorobis (cyclopentylamine) platinum (2) (cis-PAD) on L5178Y cells of two strains inversely cross-sensitive to X-rays and UV-light. Part 3. Differential effects of caffeine

International Nuclear Information System (INIS)

Szumiel, I.

1977-01-01

Two strains of L5178Y murine lymphoma, inversely cross-sensitive to X-rays and UV-light, were shown previously to respond to treatment with an antitumour platinum complex, cis-PAD, in a similar manner as to UV. The difference in sensitivity to cis-PAD, found in L5178Y-R and L5178Y-S cells is not caused by a difference in ability to bind platinum complex. Enhancement of chromosomal damage and potentiation of lethal effect of cis-PAD by 0.75 mM caffeine were found in cis-PAD and UV-light-resistant L5178Y-S strain but not in cis-PAD and UV-light-sensitive L5178Y-R strain. These results suggest that the extreme sensitivity of L5178Y-R strain to cis-PAD and UV-light is caused to some extent by deficiency in a caffeine-sensitive post-replication repair system. (author)

Action of cis-dichlorobis (cyclopentylamine) platinum (2) (cis-PAD) on L5178Y cells of two strains inversely cross-sensitive to X-rays and UV-light. Part 1. Cytotoxicity

International Nuclear Information System (INIS)

Szumiel, I.

1977-01-01

The response to cis-PAD, an antitumour platinum complex, was studied in two strains of murine lymphoma L5178Y cross-sensitive to X-rays and UV light. Dose-survival relationship, DNA synthesis, formation of chromatid aberrations, progression through the cell cycle, and growth and viability changes after 1 h cis-PAD treatment at 37 0 C were examined and compared with respective effects of X-rays and UV-light. In both strains studied, cis-PAD causes immediate inhibition of progression through the cell cycle, reduced rate of DNA synthesis, delayed appearance of chromatid aberrations and delayed death. However, there is a marked difference in sensitivity to cis-PAD between L5178Y-S strain (D 0 ca.5.8 μg/ml) and L5178Y-R strain (D 0 ca. 2.5μg/ml). In both strains a close resemblance was found between dose-survival relationship after cis-PAD and UV-light treatment, respectively. (author)

Adenovirus delivered short hairpin RNA targeting a conserved site in the 5' non-translated region inhibits all four serotypes of dengue viruses.

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Anil Babu Korrapati

Full Text Available BACKGROUND: Dengue is a mosquito-borne viral disease caused by four closely related serotypes of Dengue viruses (DENVs. This disease whose symptoms range from mild fever to potentially fatal haemorrhagic fever and hypovolemic shock, threatens nearly half the global population. There is neither a preventive vaccine nor an effective antiviral therapy against dengue disease. The difference between severe and mild disease appears to be dependent on the viral load. Early diagnosis may enable timely therapeutic intervention to blunt disease severity by reducing the viral load. Harnessing the therapeutic potential of RNA interference (RNAi to attenuate DENV replication may offer one approach to dengue therapy. METHODOLOGY/PRINCIPAL FINDINGS: We screened the non-translated regions (NTRs of the RNA genomes of representative members of the four DENV serotypes for putative siRNA targets mapping to known transcription/translation regulatory elements. We identified a target site in the 5' NTR that maps to the 5' upstream AUG region, a highly conserved cis-acting element essential for viral replication. We used a replication-defective human adenovirus type 5 (AdV5 vector to deliver a short-hairpin RNA (shRNA targeting this site into cells. We show that this shRNA matures to the cognate siRNA and is able to inhibit effectively antigen secretion, viral RNA replication and infectious virus production by all four DENV serotypes. CONCLUSION/SIGNIFICANCE: The data demonstrate the feasibility of using AdV5-mediated delivery of shRNAs targeting conserved sites in the viral genome to achieve inhibition of all four DENV serotypes. This paves the way towards exploration of RNAi as a possible therapeutic strategy to curtail DENV infection.

Both positive and negative regulatory elements mediate expression of a photoregulated CAB gene from Nicotiana plumbaginifolia.

Science.gov (United States)

Castresana, C; Garcia-Luque, I; Alonso, E; Malik, V S; Cashmore, A R

1988-01-01

We have analyzed promoter regulatory elements from a photoregulated CAB gene (Cab-E) isolated from Nicotiana plumbaginifolia. These studies have been performed by introducing chimeric gene constructs into tobacco cells via Agrobacterium tumefaciens-mediated transformation. Expression studies on the regenerated transgenic plants have allowed us to characterize three positive and one negative cis-acting elements that influence photoregulated expression of the Cab-E gene. Within the upstream sequences we have identified two positive regulatory elements (PRE1 and PRE2) which confer maximum levels of photoregulated expression. These sequences contain multiple repeated elements related to the sequence-ACCGGCCCACTT-. We have also identified within the upstream region a negative regulatory element (NRE) extremely rich in AT sequences, which reduces the level of gene expression in the light. We have defined a light regulatory element (LRE) within the promoter region extending from -396 to -186 bp which confers photoregulated expression when fused to a constitutive nopaline synthase ('nos') promoter. Within this region there is a 132-bp element, extending from -368 to -234 bp, which on deletion from the Cab-E promoter reduces gene expression from high levels to undetectable levels. Finally, we have demonstrated for a full length Cab-E promoter conferring high levels of photoregulated expression, that sequences proximal to the Cab-E TATA box are not replaceable by corresponding sequences from a 'nos' promoter. This contrasts with the apparent equivalence of these Cab-E and 'nos' TATA box-proximal sequences in truncated promoters conferring low levels of photoregulated expression. Images PMID:2901343

Characterization of a novel and potentially lethal designer drug (±)-cis-para-methyl-4-methylaminorex (4,4'-DMAR, or 'Serotoni').

Science.gov (United States)

Brandt, Simon D; Baumann, Michael H; Partilla, John S; Kavanagh, Pierce V; Power, John D; Talbot, Brian; Twamley, Brendan; Mahony, Olivia; O'Brien, John; Elliott, Simon P; Archer, Roland P; Patrick, Julian; Singh, Kuldip; Dempster, Nicola M; Cosbey, Simon H

2014-01-01

During the second half of 2013, a total of 26 deaths involving para-methyl-4-methylaminorex (4,4'-DMAR) were reported to the European Monitoring Centre for Drugs and Drug Addiction. While aminorex and 4-methylaminorex (4-MAR) are known psychostimulants, nothing is known about the comparatively new para-methyl analog. Analytical characterization of two independent samples obtained from online vendors confirmed the presence of the (±)-cis isomer that also appeared to be associated with at least 18 of the 26 deaths. Extensive characterizations included crystal structure analysis, single, tandem, and high-resolution mass spectrometry, liquid and gas chromatography, and nuclear magnetic resonance spectroscopy. For the work described here, both the (±)-cis and (±)-trans racemates were also synthesized, confirming that the differentiation between these two forms was straight-forward. Monoamine transporter activity was studied using rat brain synaptosomes which included the comparison with d-amphetamine, aminorex and (±)-cis-4-MAR. (±)-cis-4,4'-DMAR was a potent, efficacious substrate-type releaser at transporters for dopamine, norepinephrine and serotonin with EC50 values of 8.6 ± 1.1 nM (DAT), 26.9 ± 5.9 nM (NET) and 18.5 ± 2.8 nM (SERT), respectively. The potency of (±)-cis-4,4'-DMAR at DAT and NET rivalled that of other psychomotor stimulant drugs like d-amphetamine and aminorex. However, (±)-cis-4,4'-DMAR had much more potent actions at SERT and activity at SERT varied more than 100-fold across the four drugs. The potent releasing activity of (±)-cis-4,4'-DMAR at all three monoamine transporters predicts a potential for serious side-effects such as psychotic symptoms, agitation, hyperthermia and cardiovascular stimulation, especially after high-dose exposure or following combination with other psychostimulants. Copyright © 2014 John Wiley & Sons, Ltd.

Narrowband NIR-Induced In Situ Generation of the High-Energy Trans Conformer of Trichloroacetic Acid Isolated in Solid Nitrogen and its Spontaneous Decay by Tunneling to the Low-Energy Cis Conformer

Directory of Open Access Journals (Sweden)

R. F. G. Apóstolo

2015-12-01

Full Text Available The monomeric form of trichloroacetic acid (CCl3COOH; TCA was isolated in a cryogenic nitrogen matrix (15 K and the higher energy trans conformer (O=C–O–H dihedral: 180° was generated in situ by narrowband near-infrared selective excitation the 1st OH stretching overtone of the low-energy cis conformer (O=C–O–H dihedral: 0°. The spontaneous decay, by tunneling, of the generated high-energy conformer into the cis form was then evaluated and compared with those observed previously for the trans conformers of acetic and formic acids in identical experimental conditions. The much faster decay of the high-energy conformer of TCA compared to both formic and acetic acids (by ~35 and ca. 25 times, respectively was found to correlate well with the lower energy barrier for the trans→cis isomerization in the studied compound. The experimental studies received support from quantum chemistry calculations undertaken at the DFT(B3LYP/cc-pVDZ level of approximation, which allowed a detailed characterization of the potential energy surface of the molecule and the detailed assignment of the infrared spectra of the two conformers.

Biodiversity maintenance in food webs with regulatory environmental feedbacks.

Science.gov (United States)

Bagdassarian, Carey K; Dunham, Amy E; Brown, Christopher G; Rauscher, Daniel

2007-04-21

Although the food web is one of the most fundamental and oldest concepts in ecology, elucidating the strategies and structures by which natural communities of species persist remains a challenge to empirical and theoretical ecologists. We show that simple regulatory feedbacks between autotrophs and their environment when embedded within complex and realistic food-web models enhance biodiversity. The food webs are generated through the niche-model algorithm and coupled with predator-prey dynamics, with and without environmental feedbacks at the autotroph level. With high probability and especially at lower, more realistic connectance levels, regulatory environmental feedbacks result in fewer species extinctions, that is, in increased species persistence. These same feedback couplings, however, also sensitize food webs to environmental stresses leading to abrupt collapses in biodiversity with increased forcing. Feedback interactions between species and their material environments anchor food-web persistence, adding another dimension to biodiversity conservation. We suggest that the regulatory features of two natural systems, deep-sea tubeworms with their microbial consortia and a soil ecosystem manifesting adaptive homeostatic changes, can be embedded within niche-model food-web dynamics.

Development of cooperation of the CIS member states in the peaceful use of atomic energy

International Nuclear Information System (INIS)

Sobolev, A.Ye.

2012-01-01

Full text: Cooperation platform: Attraction of potential investors; Promotion of national goods and services; Pursuit of national and commercial interests. The Commission of the CIS Member States for the Peaceful Use of Atomic Energy is a nuclear cooperation body and the CIS intergovernmental coordinating and advisory authority. The Commission of the CIS Member States for the Peaceful Use of Atomic Energy coordinates and expands the spheres of cooperation. Members of the Commission- state-appointed heads of the authorized CIS member state bodies in the peaceful use of atomic energy; Secretariat is the working body of the Commission. Expert work groups formed within the CIS members States Commission: On the status of the draft Agreement on Coordination of Interstate Relations in the Peaceful Use of Atomic Energy in the CIS Territory; On the establishment of the CIS regional center for advanced training of medical physicists; Formation of an integrated system for the maintenance of safety of the nuclear research facilities. Issues of establishing the Coalition of the CIS Nuclear Research reactors; Formation of mechanisms for the convergence of the CIS member states legal and technical regulations in the peaceful use of atomic energy; Adaptation and introduction in the CIS members states of international standards in the field of using industrial radiation technologies and ensuring radiation safety; Basic forms of the CIS cooperation in ensuring economic security of projects for the peaceful use of atomic energy; Establishment of a system for the management of intellectual assets of the CIS members states; On the use of tele medical technologies of Ros atom State Cooperation- FMBA-MEPHI in diagnosis of oncologic diseases; Development of the major components of the Concept of Ensuring Nuclear, radiation and Radio ecological; Policy of the CIS Member States in the Peaceful Use of Atomic Energy; Joint implementation of the project to establish and implement a program of

Ultraviolet irradiation produces cytotoxic synergy and increased DNA interstrand crosslinking with cis- and trans-diamminedichloroplatinum(II)

International Nuclear Information System (INIS)

Swinnen, L.J.; Erickson, L.C.

1989-01-01

The excision-repair mechanism responsible for the removal of UV-induced thymine dimers may also play a role in the repair of cis-diamminedichloroplatinum(II) (cis-DDP)-induced DNA adducts in both bacteria and mammalian cells. It was hypothesized that UV dimers and cis-DDP adducts, when present simultaneously, might compete for a common repair system. Colony survival assays were performed in HT-29 human colon carcinoma cells exposed either to cis-DDP alone or to cis-DDP immediately followed by UV exposure. Progressively greater cytotoxic synergy with both increasing UV dose and cis-DDP dose was observed, to a point of saturation beyond which further toxicity was purely additive. An approximate doubling in DNA crosslink frequency, relative to cis-DDP alone, was found in cells exposed to cis-DDP plus UV. Since cis-DDP produces both inter- and intrastrand DNA crosslinks similar studies were performed with trans-DDP, which is incapable of producing intrastrand crosslinks, but does produce interstrand crosslinks. Cytotoxic synergy and increased interstrand crosslinking again resulted from the addition of UV exposure, but not to the same extent as seen with cis-DDP. (author)

Conjugated linoleic acid (CLA. Cis 9, trans 11 and trans 10, cis 12 isomer detection in crude and refined corn oils by capillary GC

Directory of Open Access Journals (Sweden)

Özlem Tokuşoğlu

2008-06-01

Full Text Available Conjugated linoleic acids (CLAs exhibit protective effects against various types of cancer and heart diseases. With the newly developed capillary gas chromatographic method (GC, cis9, trans11 and trans10, cis12 octadecadienoic acid isomers of CLA (C18:2 were determined in crude and refined corn oils as qualitative and quantitative measurements. Cis 9, trans11 C18:2 (c9, t11 CLA was the major CLA isomer in both oils. It was found that c9, t11 CLA was 0.62% of the total lipid in crude oil and 1.24% of the total lipid in refined oil. Using the refining process, the total CLA was 1.38% whereas that of crude corn oil was 0.62%. An approximate 2.2 fold increase in the total CLA was found in refined oil (n = 9 (p y = 2.782x + 0.046 (R2 = 0.9999] were performed (p El ácido linoleico conjugado (CLA parece exhibir efecto protector frente a enfermedades cardiovasculares y varios tipos de cáncer. En este trabajo, se establece un mátodo analítico mediante cromatografía de gases con columna capilar para la determinación cualitativa y cuantitativa de los isómeros cis 9,trans 11 y trans 10, cis 12 en aceites de maiz crudo y refinado. El isómero cis 9, trans11 C18:2 fue el mayoritario encontrándose en concentraciones de 0.62% en el aceite cru,do y de 1.24 % en el aceite refinado. La cantidad total de CLA encontrada en el aceite refinado (n = 9 (p 2 = 0.9999 y de recuperación [y = 2.782x+0.046 (R2 = 0.9999]. El método cromatográfico propuesto podría ser usado para el control de calidad de los aceites vegetales.

A system-level model for the microbial regulatory genome.

Science.gov (United States)

Brooks, Aaron N; Reiss, David J; Allard, Antoine; Wu, Wei-Ju; Salvanha, Diego M; Plaisier, Christopher L; Chandrasekaran, Sriram; Pan, Min; Kaur, Amardeep; Baliga, Nitin S

2014-07-15

Microbes can tailor transcriptional responses to diverse environmental challenges despite having streamlined genomes and a limited number of regulators. Here, we present data-driven models that capture the dynamic interplay of the environment and genome-encoded regulatory programs of two types of prokaryotes: Escherichia coli (a bacterium) and Halobacterium salinarum (an archaeon). The models reveal how the genome-wide distributions of cis-acting gene regulatory elements and the conditional influences of transcription factors at each of those elements encode programs for eliciting a wide array of environment-specific responses. We demonstrate how these programs partition transcriptional regulation of genes within regulons and operons to re-organize gene-gene functional associations in each environment. The models capture fitness-relevant co-regulation by different transcriptional control mechanisms acting across the entire genome, to define a generalized, system-level organizing principle for prokaryotic gene regulatory networks that goes well beyond existing paradigms of gene regulation. An online resource (http://egrin2.systemsbiology.net) has been developed to facilitate multiscale exploration of conditional gene regulation in the two prokaryotes. © 2014 The Authors. Published under the terms of the CC BY 4.0 license.

Immobilization of defense high-level waste: an assessment of technological strategies and potential regulatory goals. Volume I

International Nuclear Information System (INIS)

1979-06-01

An investigation was made of the high-level radioactive waste immobilization technology programs in the U.S. and Europe, and of the associated regulatory programs and waste management perspectives in the countries studied. Purpose was to assess the ability of those programs to satisfy DOE waste management needs and U.S. regulatory requirements. This volume includes: introduction, immobilization strategies in the context of waste isolation program needs, high-level waste management as an integrated system, regulatory goals, engineered-barrier characteristics, barrier technology, high-level waste disposal programs, analysis of HLW immobilization technology in the context of policy and regulatory requirements, and waste immobilization program option

Columbia River Coordinated Information System (CIS), 1992-1993 Annual Report.

Energy Technology Data Exchange (ETDEWEB)

Rowe, Mike; Roger, Phillip B.; O' Connor, Dick

1993-11-01

The purposes of this report are to: (1) describe the project to date; (2) to document the work and accomplishments of the (CIS) project for Fiscal Year 1993; and (3) to provide a glimpse of future project direction. The concept of a Coordinated Information System (CIS) as an approach to meeting the growing needs for regionally standardized anadromous fish information.

Hybrid macrocycle formation and spiro annulation on cis-syn-cis-tricyclo[6.3.0.02,6]undeca-3,11-dione and its congeners via ring-closing metathesis

Directory of Open Access Journals (Sweden)

Sambasivarao Kotha

2015-07-01

Full Text Available We have developed a simple methodology to transform cis-syn-cis-triquinane derivative 2 into the diindole based macrocycle 6 involving Fischer indolization and ring-closing metathesis (RCM. Various spiro-polyquinane derivatives have been assembled via RCM as a key step.

Challenges for modeling global gene regulatory networks during development: insights from Drosophila.

Science.gov (United States)

Wilczynski, Bartek; Furlong, Eileen E M

2010-04-15

Development is regulated by dynamic patterns of gene expression, which are orchestrated through the action of complex gene regulatory networks (GRNs). Substantial progress has been made in modeling transcriptional regulation in recent years, including qualitative "coarse-grain" models operating at the gene level to very "fine-grain" quantitative models operating at the biophysical "transcription factor-DNA level". Recent advances in genome-wide studies have revealed an enormous increase in the size and complexity or GRNs. Even relatively simple developmental processes can involve hundreds of regulatory molecules, with extensive interconnectivity and cooperative regulation. This leads to an explosion in the number of regulatory functions, effectively impeding Boolean-based qualitative modeling approaches. At the same time, the lack of information on the biophysical properties for the majority of transcription factors within a global network restricts quantitative approaches. In this review, we explore the current challenges in moving from modeling medium scale well-characterized networks to more poorly characterized global networks. We suggest to integrate coarse- and find-grain approaches to model gene regulatory networks in cis. We focus on two very well-studied examples from Drosophila, which likely represent typical developmental regulatory modules across metazoans. Copyright (c) 2009 Elsevier Inc. All rights reserved.

Divergent evolutionary rates in vertebrate and mammalian specific conserved non-coding elements (CNEs) in echolocating mammals.

Science.gov (United States)

Davies, Kalina T J; Tsagkogeorga, Georgia; Rossiter, Stephen J

2014-12-19

The majority of DNA contained within vertebrate genomes is non-coding, with a certain proportion of this thought to play regulatory roles during development. Conserved Non-coding Elements (CNEs) are an abundant group of putative regulatory sequences that are highly conserved across divergent groups and thus assumed to be under strong selective constraint. Many CNEs may contain regulatory factor binding sites, and their frequent spatial association with key developmental genes - such as those regulating sensory system development - suggests crucial roles in regulating gene expression and cellular patterning. Yet surprisingly little is known about the molecular evolution of CNEs across diverse mammalian taxa or their role in specific phenotypic adaptations. We examined 3,110 vertebrate-specific and ~82,000 mammalian-specific CNEs across 19 and 9 mammalian orders respectively, and tested for changes in the rate of evolution of CNEs located in the proximity of genes underlying the development or functioning of auditory systems. As we focused on CNEs putatively associated with genes underlying the development/functioning of auditory systems, we incorporated echolocating taxa in our dataset because of their highly specialised and derived auditory systems. Phylogenetic reconstructions of concatenated CNEs broadly recovered accepted mammal relationships despite high levels of sequence conservation. We found that CNE substitution rates were highest in rodents and lowest in primates, consistent with previous findings. Comparisons of CNE substitution rates from several genomic regions containing genes linked to auditory system development and hearing revealed differences between echolocating and non-echolocating taxa. Wider taxonomic sampling of four CNEs associated with the homeobox genes Hmx2 and Hmx3 - which are required for inner ear development - revealed family-wise variation across diverse bat species. Specifically within one family of echolocating bats that utilise

The implications of RCRA [Resource Conservation and Recovery Act] regulation for the disposal of transuranic and high-level waste

International Nuclear Information System (INIS)

Sigmon, C.F.; Sharples, F.E.; Smith, E.D.

1988-01-01

In May of 1987 the Department of Energy (DOE) published a rule interpreting the definition of ''byproduct'' under the Atomic Energy Act. This byproduct rule clarified the role of the Resource Conservation and Recovery Act (RCRA) in the regulation of DOE's radioactive waste management activities. According to the rule, only the radioactive portion of DOE's mixed radioactive and hazardous waste (mixed waste), including mixed transuranic (TRU) and high-level waste (HLW), is exempt from RCRA under the byproduct exemption. The portion of a waste that is hazardous as defined by RCRA is subject to full regulation under RCRA. Because the radioactive and hazardous portions of m any, if not most, DOE wastes are likely to be inseparable, the rule in effect makes most mixed wastes subject to dual regulation. The potential application of RCRA to facilities such as the Waste Isolation Pilot Plant (WIPP) and the HLW repository creates unique challenges for both the DOE and regulatory authorities. Strategies must be developed to assure compliance with RCRA without either causing excessive administrative burdens or abandoning the goal of minimizing radiation exposure. This paper will explore some of the potential regulatory options for and recent trends in the regulation of TRU and HLW under RCRA

PBPK modeling of the cis- and trans-permethrin isomers and their major urinary metabolites in rats

Energy Technology Data Exchange (ETDEWEB)

Willemin, Marie-Emilie [Institut National de l' Environnement Industriel et des Risques (INERIS), Unité Modèles pour l' Ecotoxicologie et la Toxicologie (METO), Parc ALATA BP2, 60550, Verneuil en Halatte (France); Sorbonne University, Université de Technologie de Compiègne, CNRS, UMR 7338 Biomechanics and Bioengineering, Centre de recherche Royallieu CS 60319,60203 Compiègnee Cedex (France); Desmots, Sophie [Institut National de l' Environnement Industriel et des Risques (INERIS), Unité Toxicologie Expérimentale (TOXI), Parc ALATA BP2, 60550, Verneuil en Halatte (France); Le Grand, Rozenn [Centre Hospitalo-Universitaire de Limoges, Service de Pharmacologie et de Toxicologie — Pharmacovigilance, 2, avenue Martin Luther King, 87042 Limoges (France); Lestremau, François [Institut National de l' Environnement Industriel et des Risques (INERIS), Unité Innovation pour la Mesure (NOVA), Parc ALATA BP2, 60550, Verneuil en Halatte (France); Zeman, Florence A. [Institut National de l' Environnement Industriel et des Risques (INERIS), Unité Modèles pour l' Ecotoxicologie et la Toxicologie (METO), Parc ALATA BP2, 60550, Verneuil en Halatte (France); Leclerc, Eric [Sorbonne University, Université de Technologie de Compiègne, CNRS, UMR 7338 Biomechanics and Bioengineering, Centre de recherche Royallieu CS 60319,60203 Compiègnee Cedex (France); Moesch, Christian [Centre Hospitalo-Universitaire de Limoges, Service de Pharmacologie et de Toxicologie — Pharmacovigilance, 2, avenue Martin Luther King, 87042 Limoges (France); and others

2016-03-01

Permethrin, a pyrethroid insecticide, is suspected to induce neuronal and hormonal disturbances in humans. The widespread exposure of the populations has been confirmed by the detection of the urinary metabolites of permethrin in biomonitoring studies. Permethrin is a chiral molecule presenting two forms, the cis and the trans isomers. Because in vitro studies indicated a metabolic interaction between the trans and cis isomers of permethrin, we adapted and calibrated a PBPK model for trans- and cis-permethrin separately in rats. The model also describes the toxicokinetics of three urinary metabolites, cis- and trans-3-(2,2 dichlorovinyl)-2,2-dimethyl-(1-cyclopropane) carboxylic acid (cis- and trans-DCCA), 3-phenoxybenzoic acid (3-PBA) and 4′OH-phenoxybenzoic acid (4′-OH-PBA). In vivo experiments performed in Sprague–Dawley rats were used to calibrate the PBPK model in a Bayesian framework. The model captured well the toxicokinetics of permethrin isomers and their metabolites including the rapid absorption, the accumulation in fat, the extensive metabolism of the parent compounds, and the rapid elimination of metabolites in urine. Average hepatic clearances in rats were estimated to be 2.4 and 5.7 L/h/kg for cis- and trans-permethrin, respectively. High concentrations of the metabolite 4′-OH-PBA were measured in urine compared to cis- and trans-DCCA and 3-PBA. The confidence in the extended PBPK model was then confirmed by good predictions of published experimental data obtained using the isomers mixture. The extended PBPK model could be extrapolated to humans to predict the internal dose of exposure to permethrin from biomonitoring data in urine. - Highlights: • A PBPK model of isomers of permethrin and its urinary metabolites was developed. • A quantitative link was established for permethrin and its biomarkers of exposure. • The bayesian framework allows getting confidence interval on the estimated parameters. • The PBPK model can be extrapolated

PBPK modeling of the cis- and trans-permethrin isomers and their major urinary metabolites in rats

International Nuclear Information System (INIS)

Willemin, Marie-Emilie; Desmots, Sophie; Le Grand, Rozenn; Lestremau, François; Zeman, Florence A.; Leclerc, Eric; Moesch, Christian

2016-01-01

Permethrin, a pyrethroid insecticide, is suspected to induce neuronal and hormonal disturbances in humans. The widespread exposure of the populations has been confirmed by the detection of the urinary metabolites of permethrin in biomonitoring studies. Permethrin is a chiral molecule presenting two forms, the cis and the trans isomers. Because in vitro studies indicated a metabolic interaction between the trans and cis isomers of permethrin, we adapted and calibrated a PBPK model for trans- and cis-permethrin separately in rats. The model also describes the toxicokinetics of three urinary metabolites, cis- and trans-3-(2,2 dichlorovinyl)-2,2-dimethyl-(1-cyclopropane) carboxylic acid (cis- and trans-DCCA), 3-phenoxybenzoic acid (3-PBA) and 4′OH-phenoxybenzoic acid (4′-OH-PBA). In vivo experiments performed in Sprague–Dawley rats were used to calibrate the PBPK model in a Bayesian framework. The model captured well the toxicokinetics of permethrin isomers and their metabolites including the rapid absorption, the accumulation in fat, the extensive metabolism of the parent compounds, and the rapid elimination of metabolites in urine. Average hepatic clearances in rats were estimated to be 2.4 and 5.7 L/h/kg for cis- and trans-permethrin, respectively. High concentrations of the metabolite 4′-OH-PBA were measured in urine compared to cis- and trans-DCCA and 3-PBA. The confidence in the extended PBPK model was then confirmed by good predictions of published experimental data obtained using the isomers mixture. The extended PBPK model could be extrapolated to humans to predict the internal dose of exposure to permethrin from biomonitoring data in urine. - Highlights: • A PBPK model of isomers of permethrin and its urinary metabolites was developed. • A quantitative link was established for permethrin and its biomarkers of exposure. • The bayesian framework allows getting confidence interval on the estimated parameters. • The PBPK model can be extrapolated

Separation properties of the MIL-125(Ti) Metal-Organic Framework in high-performance liquid chromatography revealing cis/trans selectivity.

Science.gov (United States)

Van der Perre, Stijn; Liekens, Anuschka; Bueken, Bart; De Vos, Dirk E; Baron, Gino V; Denayer, Joeri F M

2016-10-21

Monodisperse MIL-125(Ti) Metal-Organic Framework crystals were synthesized and studied as stationary phase in high performance liquid chromatography (HPLC). Different pure compounds and model mixtures (including stereoisomer mixtures) were injected from which chromatographic parameters, including selectivities and resolution factors, were determined to evaluate the adsorption properties and separation performance of MIL-125(Ti) in liquid phase. The MIL-125(Ti) framework displayed a trans selectivity for cis/trans difunctionalized cyclohexane molecules with high selectivity and resolution for 1,3-dimethylcyclohexane and 4-ethylcyclohexanol. The slurry-packed column was further characterized via van Deemter analysis. Fitting of the van Deemter equation through the experimental points allowed to define the contributions of the different processes to plate height with a significant proportion of the A-term, reflecting the importance of a good crystal packing. Although high in comparison to commercial HPLC stationary phases, a very good plate height of around 50μm was found. Copyright © 2016 Elsevier B.V. All rights reserved.

Crystal structure of cis-anti-cis-dicyclohexane-18-crown-6 acetonitrile disolvate

Directory of Open Access Journals (Sweden)

Alexander Nazarenko

2015-07-01

Full Text Available The title compound (systematic name: cis-anti-cis-2,5,8,15,18,21-hexaoxatricyclo[20.4.0.09,14]hexacosane acetonitrile disolvate, C20H36O6·2CH3CN, crystallizes from an acetonitrile solution of dicyclohexane-18-crown-6 on evaporation. The molecule is arranged around a center of symmetry with half the crown ether molecule and one molecule of acetonitrile symmetry independent. All O—C—C—O torsion angles are gauche while all C—O—C—C angles are trans. The sequence of torsion angles is [(tg+t(tg−t]3; the geometry of oxygen atoms is close to pseudo-D3d with three atoms below and three atoms above the mean plane, with an average deviation of ±0.16 (1 Å from the mean plane. This geometry is identical to that observed in metal ion complexes of dicyclohexane-18-crown-6 but differs significantly from the conformation of a free unsolvated molecule. Each acetonitrile molecule connects to a crown ether molecule via two of its methyl group H atoms (C—H...O. Weaker interactions exist between the third H atom of the acetonitrile methyl group and an O atom of a neighbouring crown ether molecule (C—H...O; and between the N atom of the acetonitrile molecule and a H atom of another neighbouring crown ether molecule. All these intermolecular interactions create a three-dimensional network stabilizing the disolvate.

RNA SURVEILLANCE– AN EMERGING ROLE FOR RNA REGULATORY NETWORKS IN AGING

OpenAIRE

Montano, Monty; Long, Kimberly

2010-01-01

In this review, we describe recent advances in the field of RNA regulatory biology and relate these advances to aging science. We introduce a new term, RNA surveillance, an RNA regulatory process that is conserved in metazoans, and describe how RNA surveillance represents molecular cross-talk between two emerging RNA regulatory systems – RNA interference and RNA editing. We discuss how RNA surveillance mechanisms influence mRNA and microRNA expression and activity during lifespan. Additionall...

New families of human regulatory RNA structures identified by comparative analysis of vertebrate genomes.

Science.gov (United States)

Parker, Brian J; Moltke, Ida; Roth, Adam; Washietl, Stefan; Wen, Jiayu; Kellis, Manolis; Breaker, Ronald; Pedersen, Jakob Skou

2011-11-01

Regulatory RNA structures are often members of families with multiple paralogous instances across the genome. Family members share functional and structural properties, which allow them to be studied as a whole, facilitating both bioinformatic and experimental characterization. We have developed a comparative method, EvoFam, for genome-wide identification of families of regulatory RNA structures, based on primary sequence and secondary structure similarity. We apply EvoFam to a 41-way genomic vertebrate alignment. Genome-wide, we identify 220 human, high-confidence families outside protein-coding regions comprising 725 individual structures, including 48 families with known structural RNA elements. Known families identified include both noncoding RNAs, e.g., miRNAs and the recently identified MALAT1/MEN β lincRNA family; and cis-regulatory structures, e.g., iron-responsive elements. We also identify tens of new families supported by strong evolutionary evidence and other statistical evidence, such as GO term enrichments. For some of these, detailed analysis has led to the formulation of specific functional hypotheses. Examples include two hypothesized auto-regulatory feedback mechanisms: one involving six long hairpins in the 3'-UTR of MAT2A, a key metabolic gene that produces the primary human methyl donor S-adenosylmethionine; the other involving a tRNA-like structure in the intron of the tRNA maturation gene POP1. We experimentally validate the predicted MAT2A structures. Finally, we identify potential new regulatory networks, including large families of short hairpins enriched in immunity-related genes, e.g., TNF, FOS, and CTLA4, which include known transcript destabilizing elements. Our findings exemplify the diversity of post-transcriptional regulation and provide a resource for further characterization of new regulatory mechanisms and families of noncoding RNAs.

Paradigms for parasite conservation.

Science.gov (United States)

Dougherty, Eric R; Carlson, Colin J; Bueno, Veronica M; Burgio, Kevin R; Cizauskas, Carrie A; Clements, Christopher F; Seidel, Dana P; Harris, Nyeema C

2016-08-01

Parasitic species, which depend directly on host species for their survival, represent a major regulatory force in ecosystems and a significant component of Earth's biodiversity. Yet the negative impacts of parasites observed at the host level have motivated a conservation paradigm of eradication, moving us farther from attainment of taxonomically unbiased conservation goals. Despite a growing body of literature highlighting the importance of parasite-inclusive conservation, most parasite species remain understudied, underfunded, and underappreciated. We argue the protection of parasitic biodiversity requires a paradigm shift in the perception and valuation of their role as consumer species, similar to that of apex predators in the mid-20th century. Beyond recognizing parasites as vital trophic regulators, existing tools available to conservation practitioners should explicitly account for the unique threats facing dependent species. We built upon concepts from epidemiology and economics (e.g., host-density threshold and cost-benefit analysis) to devise novel metrics of margin of error and minimum investment for parasite conservation. We define margin of error as the risk of accidental host extinction from misestimating equilibrium population sizes and predicted oscillations, while minimum investment represents the cost associated with conserving the additional hosts required to maintain viable parasite populations. This framework will aid in the identification of readily conserved parasites that present minimal health risks. To establish parasite conservation, we propose an extension of population viability analysis for host-parasite assemblages to assess extinction risk. In the direst cases, ex situ breeding programs for parasites should be evaluated to maximize success without undermining host protection. Though parasitic species pose a considerable conservation challenge, adaptations to conservation tools will help protect parasite biodiversity in the face of

The evolutionary capacitor HSP90 buffers the regulatory effects of mammalian endogenous retroviruses.

Science.gov (United States)

Hummel, Barbara; Hansen, Erik C; Yoveva, Aneliya; Aprile-Garcia, Fernando; Hussong, Rebecca; Sawarkar, Ritwick

2017-03-01

Understanding how genotypes are linked to phenotypes is important in biomedical and evolutionary studies. The chaperone heat-shock protein 90 (HSP90) buffers genetic variation by stabilizing proteins with variant sequences, thereby uncoupling phenotypes from genotypes. Here we report an unexpected role of HSP90 in buffering cis-regulatory variation affecting gene expression. By using the tripartite-motif-containing 28 (TRIM28; also known as KAP1)-mediated epigenetic pathway, HSP90 represses the regulatory influence of endogenous retroviruses (ERVs) on neighboring genes that are critical for mouse development. Our data based on natural variations in the mouse genome show that genes respond to HSP90 inhibition in a manner dependent on their genomic location with regard to strain-specific ERV-insertion sites. The evolutionary-capacitor function of HSP90 may thus have facilitated the exaptation of ERVs as key modifiers of gene expression and morphological diversification. Our findings add a new regulatory layer through which HSP90 uncouples phenotypic outcomes from individual genotypes.

Manganese catalyzed cis-dihydroxylation of electron deficient alkenes with H2O2

NARCIS (Netherlands)

Saisaha, Pattama; Pijper, Dirk; van Summeren, Ruben P.; Hoen, Robert; Smit, Christian; de Boer, Johannes W.; Hage, Ronald; Alsters, Paul L.; Feringa, Bernard; Browne, Wesley R.

2010-01-01

A practical method for the multigram scale selective cis-dihydroxylation of electron deficient alkenes such as diethyl fumarate and N-alkyl and N-aryl-maleimides using H2O2 is described. High turnovers (>1000) can be achieved with this efficient manganese based catalyst system, prepared in situ from

Effects of metals on enantioselective toxicity and biotransformation of cis-bifenthrin in zebrafish.

Science.gov (United States)

Yang, Ye; Ji, Dapeng; Huang, Xin; Zhang, Jianyun; Liu, Jing

2017-08-01

Co-occurrence of pyrethroids and metals in watersheds previously has been reported to pose great risk to aquatic species. Pyrethroids are a class of chiral insecticides that have been shown to have enantioselective toxicity and biotransformation. However, the influence of metals on enantioselectivity of pyrethroids has not yet been evaluated. In the present study, the effects of cadmium (Cd), copper (Cu), and lead (Pb) on the enantioselective toxicity and metabolism of cis-bifenthrin (cis-BF) were investigated in zebrafish at environmentally relevant concentrations. The addition of Cd, Cu, or Pb significantly increased the mortality of zebrafish in racemate and R-enantiomer of cis-BF-treated groups. In rac-cis-BF- or 1R-cis-BF-treated groups, the addition of Cd, Cu, or Pb caused a decrease in enantiomeric fraction (EF) and an increased ratio of R-enantiomer residues in zebrafish. In 1S-cis-BF-treated groups, coexposure to Cd led to a lower EF and decreased residue levels of S-enantiomer. In addition, coexposure to the 3 metals resulted in different biodegradation characteristics of each enantiomer accompanied with differential changes in the expression of cytochrome P450 (CYP)1, CYP2, and CYP3 genes, which might be responsible for the enantioselective biodegradation of cis-BF in zebrafish. These results suggest that the influence of coexistent metals should be considered in the ecological risk assessment of chiral pyrethroids in aquatic environments. Environ Toxicol Chem 2017;36:2139-2146. © 2017 SETAC. © 2017 SETAC.

Multi-species comparative analysis of the equine ACE gene identifies a highly conserved potential transcription factor binding site in intron 16.

Directory of Open Access Journals (Sweden)

Natasha A Hamilton

Full Text Available Angiotensin converting enzyme (ACE is essential for control of blood pressure. The human ACE gene contains an intronic Alu indel (I/D polymorphism that has been associated with variation in serum enzyme levels, although the functional mechanism has not been identified. The polymorphism has also been associated with cardiovascular disease, type II diabetes, renal disease and elite athleticism. We have characterized the ACE gene in horses of breeds selected for differing physical abilities. The equine gene has a similar structure to that of all known mammalian ACE genes. Nine common single nucleotide polymorphisms (SNPs discovered in pooled DNA were found to be inherited in nine haplotypes. Three of these SNPs were located in intron 16, homologous to that containing the Alu polymorphism in the human. A highly conserved 18 bp sequence, also within that intron, was identified as being a potential binding site for the transcription factors Oct-1, HFH-1 and HNF-3β, and lies within a larger area of higher than normal homology. This putative regulatory element may contribute to regulation of the documented inter-individual variation in human circulating enzyme levels, for which a functional mechanism is yet to be defined. Two equine SNPs occurred within the conserved area in intron 16, although neither of them disrupted the putative binding site. We propose a possible regulatory mechanism of the ACE gene in mammalian species which was previously unknown. This advance will allow further analysis leading to a better understanding of the mechanisms underpinning the associations seen between the human Alu polymorphism and enzyme levels, cardiovascular disease states and elite athleticism.

Multi-species comparative analysis of the equine ACE gene identifies a highly conserved potential transcription factor binding site in intron 16.

Science.gov (United States)

Hamilton, Natasha A; Tammen, Imke; Raadsma, Herman W

2013-01-01

Angiotensin converting enzyme (ACE) is essential for control of blood pressure. The human ACE gene contains an intronic Alu indel (I/D) polymorphism that has been associated with variation in serum enzyme levels, although the functional mechanism has not been identified. The polymorphism has also been associated with cardiovascular disease, type II diabetes, renal disease and elite athleticism. We have characterized the ACE gene in horses of breeds selected for differing physical abilities. The equine gene has a similar structure to that of all known mammalian ACE genes. Nine common single nucleotide polymorphisms (SNPs) discovered in pooled DNA were found to be inherited in nine haplotypes. Three of these SNPs were located in intron 16, homologous to that containing the Alu polymorphism in the human. A highly conserved 18 bp sequence, also within that intron, was identified as being a potential binding site for the transcription factors Oct-1, HFH-1 and HNF-3β, and lies within a larger area of higher than normal homology. This putative regulatory element may contribute to regulation of the documented inter-individual variation in human circulating enzyme levels, for which a functional mechanism is yet to be defined. Two equine SNPs occurred within the conserved area in intron 16, although neither of them disrupted the putative binding site. We propose a possible regulatory mechanism of the ACE gene in mammalian species which was previously unknown. This advance will allow further analysis leading to a better understanding of the mechanisms underpinning the associations seen between the human Alu polymorphism and enzyme levels, cardiovascular disease states and elite athleticism.

Investing in CIS: obstacles and strategies

International Nuclear Information System (INIS)

Anon.

1996-01-01

The CIS (Community of Independent States) needs the help of foreign investments to develop its huge hydrocarbon potentialities. Because of the uncertainties concerning the legal and fiscal context of their activities, the role of the international oil companies has remained limited so far. These problems were discussed in May 1996 during an important ''General Session'' of the OTC 96 congress. Proven and explored onshore oil reserves in Russia are enormous, in particular in Western and Eastern Siberia. However, the economically most interesting resources are located in the offshore zones of the Barents, Pechora and Kara seas and of Eastern Siberia and Okhotsk. This paper describes the last years decay of oil production in CIS and its causes, in particular the obstacles to foreign investments (exportation and transport taxes), the recent contracts and joint-ventures with American major companies (Exxon, Amoco..), and the situation of the Russian industry in petroleum engineering and field exploitation materials. (J.S.)

Temporary wetlands: Challenges and solutions to conserving a ‘disappearing’ ecosystem

Science.gov (United States)

Calhoun, Aram J.K.; Mushet, David M.; Bell, Kathleen P.; Boix, Dani; Fitzsimons, James A.; Isselin-Nondedeu, Francis

2017-01-01

Frequent drying of ponded water, and support of unique, highly specialized assemblages of often rare species, characterize temporary wetlands, such as vernal pools, gilgais, and prairie potholes. As small aquatic features embedded in a terrestrial landscape, temporary wetlands enhance biodiversity and provide aesthetic, biogeochemical, and hydrologic functions. Challenges to conserving temporary wetlands include the need to: (1) integrate freshwater and terrestrial biodiversity priorities; (2) conserve entire ‘pondscapes’ defined by connections to other aquatic and terrestrial systems; (3) maintain natural heterogeneity in environmental gradients across and within wetlands, especially gradients in hydroperiod; (4) address economic impact on landowners and developers; (5) act without complete inventories of these wetlands; and (6) work within limited or non-existent regulatory protections. Because temporary wetlands function as integral landscape components, not singly as isolated entities, their cumulative loss is ecologically detrimental yet not currently part of the conservation calculus. We highlight approaches that use strategies for conserving temporary wetlands in increasingly human-dominated landscapes that integrate top-down management and bottom-up collaborative approaches. Diverse conservation activities (including education, inventory, protection, sustainable management, and restoration) that reduce landowner and manager costs while achieving desired ecological objectives will have the greatest probability of success in meeting conservation goals.

The Non-Coding Regulatory RNA Revolution in Archaea

Directory of Open Access Journals (Sweden)

Diego Rivera Gelsinger

2018-03-01

Full Text Available Small non-coding RNAs (sRNAs are ubiquitously found in the three domains of life playing large-scale roles in gene regulation, transposable element silencing and defense against foreign elements. While a substantial body of experimental work has been done to uncover function of sRNAs in Bacteria and Eukarya, the functional roles of sRNAs in Archaea are still poorly understood. Recently, high throughput studies using RNA-sequencing revealed that sRNAs are broadly expressed in the Archaea, comprising thousands of transcripts within the transcriptome during non-challenged and stressed conditions. Antisense sRNAs, which overlap a portion of a gene on the opposite strand (cis-acting, are the most abundantly expressed non-coding RNAs and they can be classified based on their binding patterns to mRNAs (3′ untranslated region (UTR, 5′ UTR, CDS-binding. These antisense sRNAs target many genes and pathways, suggesting extensive roles in gene regulation. Intergenic sRNAs are less abundantly expressed and their targets are difficult to find because of a lack of complete overlap between sRNAs and target mRNAs (trans-acting. While many sRNAs have been validated experimentally, a regulatory role has only been reported for very few of them. Further work is needed to elucidate sRNA-RNA binding mechanisms, the molecular determinants of sRNA-mediated regulation, whether protein components are involved and how sRNAs integrate with complex regulatory networks.

Electronic structure description of the cis-MoOS unit in models for molybdenum hydroxylases.

Science.gov (United States)

Doonan, Christian J; Rubie, Nick D; Peariso, Katrina; Harris, Hugh H; Knottenbelt, Sushilla Z; George, Graham N; Young, Charles G; Kirk, Martin L

2008-01-09

The molybdenum hydroxylases catalyze the oxidation of numerous aromatic heterocycles and simple organics and, unlike other hydroxylases, utilize water as the source of oxygen incorporated into the product. The electronic structures of the cis-MoOS units in CoCp2[TpiPrMoVOS(OPh)] and TpiPrMoVIOS(OPh) (TpiPr = hydrotris(3-isopropylpyrazol-1-yl)borate), new models for molybdenum hydroxylases, have been studied in detail using S K-edge X-ray absorption spectroscopy, vibrational spectroscopy, and detailed bonding calculations. The results show a highly delocalized Mo=S pi* LUMO redox orbital that is formally Mo(dxy) with approximately 35% sulfido ligand character. Vibrational spectroscopy has been used to quantitate Mo-Ssulfido bond order changes in the cis-MoOS units as a function of redox state. Results support a redox active molecular orbital that has a profound influence on MoOS bonding through changes to the relative electro/nucleophilicity of the terminal sulfido ligand accompanying oxidation state changes. The bonding description for these model cis-MoOS systems supports enzyme mechanisms that are under orbital control and dominantly influenced by the unique electronic structure of the cis-MoOS site. The electronic structure of the oxidized enzyme site is postulated to play a role in polarizing a substrate carbon center for nucleophilic attack by metal activated water and acting as an electron sink in the two-electron oxidation of substrates.

Studies of Binary Complexes of Tripodal Ligand cis,cis-1,3,5-tris(methylaminocyclohexane with Cr(III and Fe(III

Directory of Open Access Journals (Sweden)

S. Esakki Muthu

2005-01-01

Full Text Available The formation of binary complexes of Cr(III and Fe(III with a tripodal ligand cis,cis-1,3,5-tris(methylaminocyclohexane (tmach (L has been investigated in solution. The overall stability constants of tmach with Cr(III and Fe(III were determined by potentiometric method at an ionic strength of 0.1 M NaClO4 at 25±1°C in aqueous medium. The formation of species like MLH25+, MLH4+, ML3+, ML(OH2+ and ML(OH3 were observed. Fe(III was found to form more stable complexes than Cr(III. Molecular mechanics calculations were performed to explain the mode of coordination in solution.

Identification of conserved drought-adaptive genes using a cross-species meta-analysis approach.

Science.gov (United States)

Shaar-Moshe, Lidor; Hübner, Sariel; Peleg, Zvi

2015-05-03

Drought is the major environmental stress threatening crop-plant productivity worldwide. Identification of new genes and metabolic pathways involved in plant adaptation to progressive drought stress at the reproductive stage is of great interest for agricultural research. We developed a novel Cross-Species meta-Analysis of progressive Drought stress at the reproductive stage (CSA:Drought) to identify key drought adaptive genes and mechanisms and to test their evolutionary conservation. Empirically defined filtering criteria were used to facilitate a robust integration of 17 deposited microarray experiments (148 arrays) of Arabidopsis, rice, wheat and barley. By prioritizing consistency over intensity, our approach was able to identify 225 differentially expressed genes shared across studies and taxa. Gene ontology enrichment and pathway analyses classified the shared genes into functional categories involved predominantly in metabolic processes (e.g. amino acid and carbohydrate metabolism), regulatory function (e.g. protein degradation and transcription) and response to stimulus. We further investigated drought related cis-acting elements in the shared gene promoters, and the evolutionary conservation of shared genes. The universal nature of the identified drought-adaptive genes was further validated in a fifth species, Brachypodium distachyon that was not included in the meta-analysis. qPCR analysis of 27, randomly selected, shared orthologs showed similar expression pattern as was found by the CSA:Drought.In accordance, morpho-physiological characterization of progressive drought stress, in B. distachyon, highlighted the key role of osmotic adjustment as evolutionary conserved drought-adaptive mechanism. Our CSA:Drought strategy highlights major drought-adaptive genes and metabolic pathways that were only partially, if at all, reported in the original studies included in the meta-analysis. These genes include a group of unclassified genes that could be involved

The evolutionarily conserved leprecan gene: its regulation by Brachyury and its role in the developing Ciona notochord.

Science.gov (United States)

Dunn, Matthew P; Di Gregorio, Anna

2009-04-15

In Ciona intestinalis, leprecan was identified as a target of the notochord-specific transcription factor Ciona Brachyury (Ci-Bra) (Takahashi, H., Hotta, K., Erives, A., Di Gregorio, A., Zeller, R.W., Levine, M., Satoh, N., 1999. Brachyury downstream notochord differentiation in the ascidian embryo. Genes Dev. 13, 1519-1523). By screening approximately 14 kb of the Ci-leprecan locus for cis-regulatory activity, we have identified a 581-bp minimal notochord-specific cis-regulatory module (CRM) whose activity depends upon T-box binding sites located at the 3'-end of its sequence. These sites are specifically bound in vitro by a GST-Ci-Bra fusion protein, and mutations that abolish binding in vitro result in loss or decrease of regulatory activity in vivo. Serial deletions of the 581-bp notochord CRM revealed that this sequence is also able to direct expression in muscle cells through the same T-box sites that are utilized by Ci-Bra in the notochord, which are also bound in vitro by the muscle-specific T-box activators Ci-Tbx6b and Ci-Tbx6c. Additionally, we created plasmids aimed to interfere with the function of Ci-leprecan and categorized the resulting phenotypes, which consist of variable dislocations of notochord cells along the anterior-posterior axis. Together, these observations provide mechanistic insights generally applicable to T-box transcription factors and their target sequences, as well as a first set of clues on the function of Leprecan in early chordate development.

The Regulatory Independence of FANR

International Nuclear Information System (INIS)

ALNuaimi, Fatema; Choi, Kwang Shik

2012-01-01

Regulatory independence is meant to provide a conservative system of policy making in order to comply with the problems that are forecasted upon the basis of assumptions. The Federal Authorization of Nuclear Regulation (FANR) is a regulatory commission that was formed to be regulatory body that governs the generation of nuclear power in United Arab Emirates. It was established under the UAE nuclear law (9/2009) as an independent regulatory body that was tasked with the regulation of all nuclear activities in the United Arab Emirates. As an independent body, FANR was tasked with ensuring that the regulation of the nuclear sector is done in effective and transparent manner to ensure its accountability to the people. Being independent, the regulatory body develops national nuclear regulations based on laid down safety standards by the International Atomic Energy Agency, ensuring that they are based on scientific and proven technologies The role of FANR is to ensure that the all corporations that undertake nuclear activities follow the laid down procedures and objectives and ensure safety measures are taken keenly to ensure the safety of the workers and the general public while at the same time ensuring the environment is free from nuclear radiations

The Regulatory Independence of FANR

Energy Technology Data Exchange (ETDEWEB)

ALNuaimi, Fatema; Choi, Kwang Shik [Korea Advanced Institute of Science and Technology, Daejeon (Korea, Republic of)

2012-05-15

Regulatory independence is meant to provide a conservative system of policy making in order to comply with the problems that are forecasted upon the basis of assumptions. The Federal Authorization of Nuclear Regulation (FANR) is a regulatory commission that was formed to be regulatory body that governs the generation of nuclear power in United Arab Emirates. It was established under the UAE nuclear law (9/2009) as an independent regulatory body that was tasked with the regulation of all nuclear activities in the United Arab Emirates. As an independent body, FANR was tasked with ensuring that the regulation of the nuclear sector is done in effective and transparent manner to ensure its accountability to the people. Being independent, the regulatory body develops national nuclear regulations based on laid down safety standards by the International Atomic Energy Agency, ensuring that they are based on scientific and proven technologies The role of FANR is to ensure that the all corporations that undertake nuclear activities follow the laid down procedures and objectives and ensure safety measures are taken keenly to ensure the safety of the workers and the general public while at the same time ensuring the environment is free from nuclear radiations

Linkage disequilibrium of evolutionarily conserved regions in the human genome

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Johnson Todd A

2006-12-01

Full Text Available Abstract Background The strong linkage disequilibrium (LD recently found in genic or exonic regions of the human genome demonstrated that LD can be increased by evolutionary mechanisms that select for functionally important loci. This suggests that LD might be stronger in regions conserved among species than in non-conserved regions, since regions exposed to natural selection tend to be conserved. To assess this hypothesis, we used genome-wide polymorphism data from the HapMap project and investigated LD within DNA sequences conserved between the human and mouse genomes. Results Unexpectedly, we observed that LD was significantly weaker in conserved regions than in non-conserved regions. To investigate why, we examined sequence features that may distort the relationship between LD and conserved regions. We found that interspersed repeats, and not other sequence features, were associated with the weak LD tendency in conserved regions. To appropriately understand the relationship between LD and conserved regions, we removed the effect of repetitive elements and found that the high degree of sequence conservation was strongly associated with strong LD in coding regions but not with that in non-coding regions. Conclusion Our work demonstrates that the degree of sequence conservation does not simply increase LD as predicted by the hypothesis. Rather, it implies that purifying selection changes the polymorphic patterns of coding sequences but has little influence on the patterns of functional units such as regulatory elements present in non-coding regions, since the former are generally restricted by the constraint of maintaining a functional protein product across multiple exons while the latter may exist more as individually isolated units.

Sunscreens with broad-spectrum absorption decrease the trans TO cis photoisomerization of urocanic acid in the human stratum corneum after multiple UV light exposures

International Nuclear Information System (INIS)

Krien, P.M.; Moyal, D.

1994-01-01

The trans to cis photoisomerization of urocanic acid (UCA) in skin is considered to play an important role in the mechanism of immunosuppression. We have investigated the effects of skin type and various sunscreens with low sun protection factor (SPF) on the UV-induced cis-UCA formation in human skin after exposure to artificial UV light. The rate of cis-UCA formation depends little on the skin type and is reduced by topical application of sunscreens. The rate of cis-UCA formation decreases with increasing SPF and only broad-spectrum, highly protective sunscreens offer protection against the UV-induced formation of cis-UCA, which accumulates in the stratum corneum after multiple UV exposures. A theoretical approach to estimate the distribution of cis-UCA after irradiation indicates that this compound may diffuse into the deeper layers of the epidermis with D ∼ 10 -17 m 2 /s, and that its elimination from the stratum corneum is mainly due to desquamation. (author)

CIS-Type PV Device Fabrication by Novel Techniques; Phase I Annual Technical Report, 1 July 1998 - 30 June 1999

International Nuclear Information System (INIS)

Basol, B.M.; Halani, A.; Kapur, V.K.; Leidholm, C.R.; Norsworthy, G.; Roe, R.

1999-01-01

This report describes work performed by International Solar Electric Technology, Inc. (ISET) during phase I of the R and D partnership subcontract titled ''CIS-Type PV Device Fabrication by Novel Techniques.'' The objective of this program is to bring ISET's novel non-vacuum CIS technology closer to commercialization by concentrating on issues such as device-efficiency improvement, larger-bandgap absorber growth, and module fabrication. Advances made in CIS and related compound solar cell fabrication processes have clearly shown that these materials and device structures can yield power conversion efficiencies in the 15%-20% range. However, many of the laboratory results on CIS-type devices have been obtained using relatively high-cost vacuum-based deposition techniques. The present project was specifically geared toward developing a low-cost, non-vacuum ''particle deposition'' method for CIS-type absorber growth. There are four major processing steps in this technique: i) preparation of a starting powder containing all or some of the chemical species constituting CIS, ii) preparation of an ink using the starting powder, iii) deposition of the ink on a substrate in the form of a thin precursor layer, and iv) conversion of the precursor layer into a fused photovoltaic absorber through annealing steps. During this Phase I program, ISET worked on tasks that were geared toward the following goals: i) elimination of back-contact problems, ii) growth of large-bandgap absorbers, and iii) fabrication of mini-modules. As a result of the Phase I research, a Mo back-contact structure was developed that eliminated problems that resulted in poor mechanical integrity of the absorber layers. Sulfur inclusion into CIS films through high-temperature sulfurization in H2S gas was also studied. It was determined that S diffusion was a strong function of the stoichiometry of the CIS layer. Sulfur was found to diffuse rapidly through the Cu-rich films, whereas the diffusion constant was

Regulatory Mechanisms of a Highly Pectinolytic Mutant of Penicillium occitanis and Functional Analysis of a Candidate Gene in the Plant Pathogen Fusarium oxysporum

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Gustavo Bravo-Ruiz

2017-09-01

Full Text Available Penicillium occitanis is a model system for enzymatic regulation. A mutant strain exhibiting constitutive overproduction of different pectinolytic enzymes both under inducing (pectin or repressing conditions (glucose was previously isolated after chemical mutagenesis. In order to identify the molecular basis of this regulatory mechanism, the genomes of the wild type and the derived mutant strain were sequenced and compared, providing the first reference genome for this species. We used a phylogenomic approach to compare P. occitanis with other pectinolytic fungi and to trace expansions of gene families involved in carbohydrate degradation. Genome comparison between wild type and mutant identified seven mutations associated with predicted proteins. The most likely candidate was a mutation in a highly conserved serine residue of a conserved fungal protein containing a GAL4-like Zn2Cys6 binuclear cluster DNA-binding domain and a fungus-specific transcription factor regulatory middle homology region. To functionally characterize the role of this candidate gene, the mutation was recapitulated in the predicted orthologue Fusarium oxysporum, a vascular wilt pathogen which secretes a wide array of plant cell wall degrading enzymes, including polygalacturonases, pectate lyases, xylanases and proteases, all of which contribute to infection. However, neither the null mutant nor a mutant carrying the analogous point mutation exhibited a deregulation of pectinolytic enzymes. The availability, annotation and phylogenomic analysis of the P. occitanis genome sequence represents an important resource for understanding the evolution and biology of this species, and sets the basis for the discovery of new genes of biotechnological interest for the degradation of complex polysaccharides.

The conservation pattern of short linear motifs is highly correlated with the function of interacting protein domains

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Wang Yiguo

2008-10-01

Full Text Available Abstract Background Many well-represented domains recognize primary sequences usually less than 10 amino acids in length, called Short Linear Motifs (SLiMs. Accurate prediction of SLiMs has been difficult because they are short (often Results Our combined approach revealed that SLiMs are highly conserved in proteins from functional classes that are known to interact with a specific domain, but that they are not conserved in most other protein groups. We found that SLiMs recognized by SH2 domains were highly conserved in receptor kinases/phosphatases, adaptor molecules, and tyrosine kinases/phosphatases, that SLiMs recognized by SH3 domains were highly conserved in cytoskeletal and cytoskeletal-associated proteins, that SLiMs recognized by PDZ domains were highly conserved in membrane proteins such as channels and receptors, and that SLiMs recognized by S/T kinase domains were highly conserved in adaptor molecules, S/T kinases/phosphatases, and proteins involved in transcription or cell cycle control. We studied Tyr-SLiMs recognized by SH2 domains in more detail, and found that SH2-recognized Tyr-SLiMs on the cytoplasmic side of membrane proteins are more highly conserved than those on the extra-cellular side. Also, we found that SH2-recognized Tyr-SLiMs that are associated with SH3 motifs and a tyrosine kinase phosphorylation motif are more highly conserved. Conclusion The interactome of protein domains is reflected by the evolutionary conservation of SLiMs recognized by these domains. Combining scoring matrixes derived from peptide libraries and conservation analysis, we would be able to find those protein groups that are more likely to interact with specific domains.

Water conservation and allocation guideline for oilfield injection

International Nuclear Information System (INIS)

2006-01-01

This paper was prepared as a guide for regulatory agencies and developers using non-saline water sources in enhanced oil recovery (EOR) schemes. A systems approach was used to achieve specific environmental outcomes that adhered to the Water Conservation and Allocation Policy for Oilfield Injection. The guide was applicable to licence renewal applications for projects operating and licensed to use non-saline water resources, as well as new licence applications for oilfield injection use. The guide provided recommended water conservation practices and application requirements, and outlined regulatory procedures and steps for obtaining a Water Act licence. The guideline was prepared to eliminate the use of non-saline water in EOR projects where feasible alternatives existed, as well as to identify areas with water shortages and reduce the use of non-saline water. The guide included monitoring and reporting requirements to improve the evaluation of water use practices and outlined current initiatives to address water conservation and research. It was concluded that outcomes from the program will include reliable quality water supplies for a sustainable economy, healthy aquatic ecosystems, and safe, secure drinking water supplies for Albertans. 3 tabs., 5 figs

Transcriptional regulation by competing transcription factor modules.

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Rutger Hermsen

2006-12-01

Full Text Available Gene regulatory networks lie at the heart of cellular computation. In these networks, intracellular and extracellular signals are integrated by transcription factors, which control the expression of transcription units by binding to cis-regulatory regions on the DNA. The designs of both eukaryotic and prokaryotic cis-regulatory regions are usually highly complex. They frequently consist of both repetitive and overlapping transcription factor binding sites. To unravel the design principles of these promoter architectures, we have designed in silico prokaryotic transcriptional logic gates with predefined input-output relations using an evolutionary algorithm. The resulting cis-regulatory designs are often composed of modules that consist of tandem arrays of binding sites to which the transcription factors bind cooperatively. Moreover, these modules often overlap with each other, leading to competition between them. Our analysis thus identifies a new signal integration motif that is based upon the interplay between intramodular cooperativity and intermodular competition. We show that this signal integration mechanism drastically enhances the capacity of cis-regulatory domains to integrate signals. Our results provide a possible explanation for the complexity of promoter architectures and could be used for the rational design of synthetic gene circuits.

Ethical Challenges of Educational Leadership in the Countries of the CIS

Science.gov (United States)

Zagoumennov, Iouri

2010-01-01

There is no school administrators' pre-service training in the CIS (Former USSR) countries. As for the in-service training, it has been traditionally focused on educational management. As a result, schools in the CIS countries are run as well-oiled machines but there is a big question mark over whether these machines are running in the right…

Genome-wide haplotype analysis of cis expression quantitative trait loci in monocytes.

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Sophie Garnier

Full Text Available In order to assess whether gene expression variability could be influenced by several SNPs acting in cis, either through additive or more complex haplotype effects, a systematic genome-wide search for cis haplotype expression quantitative trait loci (eQTL was conducted in a sample of 758 individuals, part of the Cardiogenics Transcriptomic Study, for which genome-wide monocyte expression and GWAS data were available. 19,805 RNA probes were assessed for cis haplotypic regulation through investigation of ~2,1 × 10(9 haplotypic combinations. 2,650 probes demonstrated haplotypic p-values >10(4-fold smaller than the best single SNP p-value. Replication of significant haplotype effects were tested for 412 probes for which SNPs (or proxies that defined the detected haplotypes were available in the Gutenberg Health Study composed of 1,374 individuals. At the Bonferroni correction level of 1.2 × 10(-4 (~0.05/412, 193 haplotypic signals replicated. 1000 G imputation was then conducted, and 105 haplotypic signals still remained more informative than imputed SNPs. In-depth analysis of these 105 cis eQTL revealed that at 76 loci genetic associations were compatible with additive effects of several SNPs, while for the 29 remaining regions data could be compatible with a more complex haplotypic pattern. As 24 of the 105 cis eQTL have previously been reported to be disease-associated loci, this work highlights the need for conducting haplotype-based and 1000 G imputed cis eQTL analysis before commencing functional studies at disease-associated loci.

Oxidation reactions of derivatives of cis-octalins promoted by thallium trinitrate (TTN)

International Nuclear Information System (INIS)

Ferraz, Helena M.C.; Carneiro, Vania M.T.; Vieira, Tiago O.; Silva Junior, Luiz F.

2008-01-01

The reaction of ten cis-octalins and cis-octalones with thallium trinitrate (TTN) leads to different products, depending mainly on the substitution pattern of the substrate. Functionalized cis-hydrindanes were obtained from the reaction of 1,2,3,4,4a,5,8,8a-octahydro- 4a-methylnaphthalene and of 1,2,3,4,4a,5,8,8a-octahydro-4a,7-dimethylnaphthalene with TTN in acetonitrile, whereas a cyclic ether was formed treating 1,2,3,4,4a,5,8,8a-octahydro-6,8a-dimethylnaphthalene-1-ol with TTN in trimethylorthoformate (TMOF). (author)

Enantiodiscrimination of flexible cyclic solutes using NMR spectroscopy in polypeptide chiral mesophases: investigation of cis-decalin and THF.

Science.gov (United States)

Aroulanda, Christie; Lafon, Olivier; Lesot, Philippe

2009-08-06

The conformational dynamics and orientational behavior of two model cyclic molecules, cis-decalin (cis-dec) and tetrahydrofurane (THF), dissolved in weakly ordering, polypeptidic chiral liquid crystals (CLCs) are theoretically discussed and experimentally investigated using deuterium and carbon-13 NMR spectroscopies. The analysis of enantiomeric and enantiotopic discriminations in these compounds is shown to depend on the rate of conformational exchange regime, slow or fast. The slow exchange regime is illustrated through the case of cis-dec at low temperature (243 K). We show that the deuterium NMR spectra in this regime can be qualitatively and quantitatively interpreted by restricting the conformational pathway of cis-dec to two enantiomeric conformers of C(2)-symmetry. The orientational order parameters of these interconverting enantiomers are calculated by matching the (2)H quadrupolar splittings with calculated conformer structures. The fast exchange regime is investigated through the examples of cis-dec at high temperature (356 K) and THF at room temperature (300 K). The (2)H NMR spectra above the coalescence temperature are analyzed by introducing the concept of "average molecular structure". This fictitious structure allows easily identifying NMR equivalences of solutes dissolved in CLC. However, it cannot be applied to determine consistent orientational order parameters. This study emphasizes that enantiotopic discriminations observed for flexible molecules in the fast exchange regime can be quantitatively interpreted only by considering the orientational order of each conformer.

ABCE1 is a highly conserved RNA silencing suppressor.

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Kairi Kärblane

Full Text Available ATP-binding cassette sub-family E member 1 (ABCE1 is a highly conserved protein among eukaryotes and archaea. Recent studies have identified ABCE1 as a ribosome-recycling factor important for translation termination in mammalian cells, yeast and also archaea. Here we report another conserved function of ABCE1. We have previously described AtRLI2, the homolog of ABCE1 in the plant Arabidopsis thaliana, as an endogenous suppressor of RNA silencing. In this study we show that this function is conserved: human ABCE1 is able to suppress RNA silencing in Nicotiana benthamiana plants, in mammalian HEK293 cells and in the worm Caenorhabditis elegans. Using co-immunoprecipitation and mass spectrometry, we found a number of potential ABCE1-interacting proteins that might support its function as an endogenous suppressor of RNA interference. The interactor candidates are associated with epigenetic regulation, transcription, RNA processing and mRNA surveillance. In addition, one of the identified proteins is translin, which together with its binding partner TRAX supports RNA interference.

Structure-aided prediction of mammalian transcription factor complexes in conserved non-coding elements

KAUST Repository

Guturu, H.

2013-11-11

Mapping the DNA-binding preferences of transcription factor (TF) complexes is critical for deciphering the functions of cis-regulatory elements. Here, we developed a computational method that compares co-occurring motif spacings in conserved versus unconserved regions of the human genome to detect evolutionarily constrained binding sites of rigid TF complexes. Structural data were used to estimate TF complex physical plausibility, explore overlapping motif arrangements seldom tackled by non-structure-aware methods, and generate and analyse three-dimensional models of the predicted complexes bound to DNA. Using this approach, we predicted 422 physically realistic TF complex motifs at 18% false discovery rate, the majority of which (326, 77%) contain some sequence overlap between binding sites. The set of mostly novel complexes is enriched in known composite motifs, predictive of binding site configurations in TF-TF-DNA crystal structures, and supported by ChIP-seq datasets. Structural modelling revealed three cooperativity mechanisms: direct protein-protein interactions, potentially indirect interactions and \\'through-DNA\\' interactions. Indeed, 38% of the predicted complexes were found to contain four or more bases in which TF pairs appear to synergize through overlapping binding to the same DNA base pairs in opposite grooves or strands. Our TF complex and associated binding site predictions are available as a web resource at http://bejerano.stanford.edu/complex.

Structure-aided prediction of mammalian transcription factor complexes in conserved non-coding elements

KAUST Repository

Guturu, H.; Doxey, A. C.; Wenger, A. M.; Bejerano, G.

2013-01-01

Mapping the DNA-binding preferences of transcription factor (TF) complexes is critical for deciphering the functions of cis-regulatory elements. Here, we developed a computational method that compares co-occurring motif spacings in conserved versus unconserved regions of the human genome to detect evolutionarily constrained binding sites of rigid TF complexes. Structural data were used to estimate TF complex physical plausibility, explore overlapping motif arrangements seldom tackled by non-structure-aware methods, and generate and analyse three-dimensional models of the predicted complexes bound to DNA. Using this approach, we predicted 422 physically realistic TF complex motifs at 18% false discovery rate, the majority of which (326, 77%) contain some sequence overlap between binding sites. The set of mostly novel complexes is enriched in known composite motifs, predictive of binding site configurations in TF-TF-DNA crystal structures, and supported by ChIP-seq datasets. Structural modelling revealed three cooperativity mechanisms: direct protein-protein interactions, potentially indirect interactions and 'through-DNA' interactions. Indeed, 38% of the predicted complexes were found to contain four or more bases in which TF pairs appear to synergize through overlapping binding to the same DNA base pairs in opposite grooves or strands. Our TF complex and associated binding site predictions are available as a web resource at http://bejerano.stanford.edu/complex.

In-silico analysis of cis-acting regulatory elements of pathogenesis-related proteins of Arabidopsis thaliana and Oryza sativa.

Science.gov (United States)

Kaur, Amritpreet; Pati, Pratap Kumar; Pati, Aparna Maitra; Nagpal, Avinash Kaur

2017-01-01

Pathogenesis related (PR) proteins are low molecular weight family of proteins induced in plants under various biotic and abiotic stresses. They play an important role in plant-defense mechanism. PRs have wide range of functions, acting as hydrolases, peroxidases, chitinases, anti-fungal, protease inhibitors etc. In the present study, an attempt has been made to analyze promoter regions of PR1, PR2, PR5, PR9, PR10 and PR12 of Arabidopsis thaliana and Oryza sativa. Analysis of cis-element distribution revealed the functional multiplicity of PRs and provides insight into the gene regulation. CpG islands are observed only in rice PRs, which indicates that monocot genome contains more GC rich motifs than dicots. Tandem repeats were also observed in 5' UTR of PR genes. Thus, the present study provides an understanding of regulation of PR genes and their versatile roles in plants.

Identifying TF-MiRNA Regulatory Relationships Using Multiple Features.

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Mingyu Shao

Full Text Available MicroRNAs are known to play important roles in the transcriptional and post-transcriptional regulation of gene expression. While intensive research has been conducted to identify miRNAs and their target genes in various genomes, there is only limited knowledge about how microRNAs are regulated. In this study, we construct a pipeline that can infer the regulatory relationships between transcription factors and microRNAs from ChIP-Seq data with high confidence. In particular, after identifying candidate peaks from ChIP-Seq data, we formulate the inference as a PU learning (learning from only positive and unlabeled examples problem. Multiple features including the statistical significance of the peaks, the location of the peaks, the transcription factor binding site motifs, and the evolutionary conservation are derived from peaks for training and prediction. To further improve the accuracy of our inference, we also apply a mean reciprocal rank (MRR-based method to the candidate peaks. We apply our pipeline to infer TF-miRNA regulatory relationships in mouse embryonic stem cells. The experimental results show that our approach provides very specific findings of TF-miRNA regulatory relationships.

Molecular mechanism of 9-cis-retinoic acid inhibition of adipogenesis in 3T3-L1 cells

International Nuclear Information System (INIS)

Sagara, Chiaki; Takahashi, Katsuhiko; Kagechika, Hiroyuki; Takahashi, Noriko

2013-01-01

Highlights: ► We examined the effects of 9-cis-RA on adipogenesis in mouse preadipocyte 3T3-L1. ► 9-cis-RA inhibited lipid accumulation in adipogenetically-induced 3T3-L1 cells. ► A RXR pan-antagonist suppressed the inhibitory effects of 9-cis-RA on adipogenesis. ► This antagonist had no effects on RXRα and PPARγ levels in 9-cis-RA-treated cells. ► 9-cis-RA-induced decrease in both RXRα and PPARγ was independent of RXR activation. -- Abstract: Retinoic acid (RA) signaling is mediated by specific nuclear hormone receptors. Here we examined the effects of 9-cis-RA on adipogenesis in mouse preadipocyte 3T3-L1 cells. 9-cis-RA inhibits the lipid accumulation of adipogenetically induced 3T3-L1 cells. The complex of retinoid X receptor α (RXRα) with peroxisome proliferator-activated receptor γ (PPARγ) is a major transcription factor in the process of adipogenesis, and the levels of these molecules were decreased by 9-cis-RA treatment. A RXR pan-antagonist suppressed 9-cis-RA’s inhibitory effects on adipogenesis, but not on the intracellular levels of both RXRα and PPARγ. These results suggest that 9-cis-RA could inhibit adipogenesis by activating RXR, and decrease both RXR and PPARγs levels in a RXR activation-independent manner

Current status of herbal product: Regulatory overview

Science.gov (United States)

Sharma, Sanjay

2015-01-01

A review of the regulatory status of herbal drugs/products was done for few countries forming part of Asia, Africa, America, Europe, and Australia, to understand various categories under which the trade of herbal products is permitted and their premarketing requirements. A critical assessment was done, to know the hindrances in the process of harmonization of herbal products. It has been found that there is a lack of harmonization in the regulatory requirements of herbal products internationally, besides the issues of availability of herbs and their conservation. These are hindering the international trade and growth of the herbal products segment. PMID:26681886

Information Spreadsheet for Engines and Vehicles Compliance Information System (EV-CIS) User Registration

Science.gov (United States)

In this spreadsheet, user(s) provide their company’s manufacturer code, user contact information for EV-CIS, and user roles. This spreadsheet is used for the Company Authorizing Official (CAO), CROMERR Signer, and EV-CIS Submitters.

Decoupling mechanisms-paying for conservation

Energy Technology Data Exchange (ETDEWEB)

Cross, P.S.

1993-07-15

In 1988, the National Association of Regulatory Utility Commissioners issued a policy statement that said [open quotes]ratemaking practices should align utilities' pursuit of profit with least-cost planning.[close quotes] This policy coincided with then-current thinkingg at a number of state commissions about the much-touted goal of encouraging utilities to invest in conservation, or demand-side management (DSM) programs, rather than in generating resources to meet system load requirements. Besides utility concerns about recovering conservation program investments, regulators also notices a built-in [open quotes]disincentive[close quotes] to investment in the traditional ratemaking format: If profit is tied to sales, then utilities will always shy away from aggressively promoting conservation. Or so the thinkin went. [open quotes]Decoupling mechanisms[close quotes] were born to remove this disincentive. A number of states have implemented these mechanisms, while several others are investigating the issue. One chief drawback of the mechanisms is that if sales go down, rates go up to cover the shortfall. (Of course, rates go down if sales exceed forecasted levels.) A major problem has been that rate increases have occurred at exactly the wrong time, during economic slowdowns when utilities are struggling to retain price-sensitive customers and residential ratepayers are least likely to bear with quiet stoicism the burden placed on family budgets. Decoupling is seen by some as a step backwards in the move to competitive regulatory reforms that seek to encourage utilities to behave like free-market companies. Indeed, the newest decoupling mechanisms face serious challenge.

A systemic identification approach for primary transcription start site of Arabidopsis miRNAs from multidimensional omics data.

Science.gov (United States)

You, Qi; Yan, Hengyu; Liu, Yue; Yi, Xin; Zhang, Kang; Xu, Wenying; Su, Zhen

2017-05-01

The 22-nucleotide non-coding microRNAs (miRNAs) are mostly transcribed by RNA polymerase II and are similar to protein-coding genes. Unlike the clear process from stem-loop precursors to mature miRNAs, the primary transcriptional regulation of miRNA, especially in plants, still needs to be further clarified, including the original transcription start site, functional cis-elements and primary transcript structures. Due to several well-characterized transcription signals in the promoter region, we proposed a systemic approach integrating multidimensional "omics" (including genomics, transcriptomics, and epigenomics) data to improve the genome-wide identification of primary miRNA transcripts. Here, we used the model plant Arabidopsis thaliana to improve the ability to identify candidate promoter locations in intergenic miRNAs and to determine rules for identifying primary transcription start sites of miRNAs by integrating high-throughput omics data, such as the DNase I hypersensitive sites, chromatin immunoprecipitation-sequencing of polymerase II and H3K4me3, as well as high throughput transcriptomic data. As a result, 93% of refined primary transcripts could be confirmed by the primer pairs from a previous study. Cis-element and secondary structure analyses also supported the feasibility of our results. This work will contribute to the primary transcriptional regulatory analysis of miRNAs, and the conserved regulatory pattern may be a suitable miRNA characteristic in other plant species.

The synthesis and preliminary pharmacological evaluation of the racemic cis and trans 3-alkylfentanyl analogues

Directory of Open Access Journals (Sweden)

LJ. DOSEN-MICOVIC

2004-07-01

Full Text Available A general, five step method for the synthesis of 3-alkylfentanyl analogues (i.e., cis and trans 3-alkyl-4-anilidopiperidines 6.1–6.6 has been developed. The starting N-phenethyl-4-piperidone 1 was first converted into the cyclohexylimine derivative 2, a-deprotonated with butyllithium and the resulting imine anion efficiently monoalkylated with primary and secondary alkyl halides. After mild acid hydrolysis, the obtained 3-alkyl-4-piperidones 3.1–3.6 were isolated in good yields (79–85 %, then condensed with aniline to form imines 4.1–4.6. Subsequent reduction of the imines (LiAlH4/THF yielded cis/trans mixtures of 3-alkyl-4-anilinopiperidines 5.1–5.6. Quantitative separation of the diastereoisomers by column chromatography of Al2O3 gave pure cis 5.1–5.6 (29–51 % yield and trans 5.1–5.6 (19–27 % yield, with the cis/trans ratio in the range 7/3–6/4. The synthesis was concluded by N-acylation of the purified 5.1–5.6, with propionyl chloride, to afford cis and trans 3-alkyl-4-anilidopiperidines 6.1–6.6 (~95 % yield, as monooxalate salts. No enatioseparation was attempted at any stage. The relative cis/trans stereochemistry was provisionally assigned from the 1H-NMR spectra. Of the twelve synthesized 3-alkylfentanyls, ten compounds (two known and eight novel derivatives, all as the monooxalate salts were preliminarily tested as analgesics in rats, comparing the potency to fentanyl. Except for the known (±-cis-3-Me fentanyl 6.1cis, (8 × fentanyl, and the novel (±-cis-3-Et fentanyl 6.2cis, (1.5 × fentanyl, all of the others were less active than fentanyl or inactive. Some tentative conclusions on the structure-activity relationship (SAR in this series of derivatives have been made.

A highly conserved tyrosine of Tim-3 is phosphorylated upon stimulation by its ligand galectin-9

International Nuclear Information System (INIS)

Weyer, Philipp S. van de; Muehlfeit, Michael; Klose, Christoph; Bonventre, Joseph V.; Walz, Gerd; Kuehn, E. Wolfgang

2006-01-01

Tim-3 is a member of the TIM family of proteins (T-cell immunoglobulin mucin) involved in the regulation of CD4+ T-cells. Tim-3 is a T H 1-specific type 1 membrane protein and regulates T H 1 proliferation and the development of tolerance. Binding of galectin-9 to the extracellular domain of Tim-3 results in apoptosis of T H 1 cells, but the intracellular pathways involved in the regulatory function of Tim-3 are unknown. Unlike Tim-1, which is expressed in renal epithelia and cancer, Tim-3 has not been described in cells other than neuronal or T-cells. Using RT-PCR we demonstrate that Tim-3 is expressed in malignant and non-malignant epithelial tissues. We have cloned Tim-3 from an immortalized liver cell carcinoma line and identified a highly conserved tyrosine in the intracellular tail of Tim-3 (Y265). We demonstrate that Y265 is specifically phosphorylated in vivo by the interleukin inducible T cell kinase (ITK), a kinase which is located in close proximity of the TIM genes on the allergy susceptibility locus 5q33.3. Stimulation of Tim-3 by its ligand galectin-9 results in increased phosphorylation of Y265, suggesting that this tyrosine residue plays an important role in downstream signalling events regulating T-cell fate. Given the role of TIM proteins in autoimmunity and cancer, the conserved SH2 binding domain surrounding Y265 could represent a possible target site for pharmacological intervention

Homologous high-throughput expression and purification of highly conserved E coli proteins

Directory of Open Access Journals (Sweden)

Duchmann Rainer

2007-06-01

Full Text Available Abstract Background Genetic factors and a dysregulated immune response towards commensal bacteria contribute to the pathogenesis of Inflammatory Bowel Disease (IBD. Animal models demonstrated that the normal intestinal flora is crucial for the development of intestinal inflammation. However, due to the complexity of the intestinal flora, it has been difficult to design experiments for detection of proinflammatory bacterial antigen(s involved in the pathogenesis of the disease. Several studies indicated a potential association of E. coli with IBD. In addition, T cell clones of IBD patients were shown to cross react towards antigens from different enteric bacterial species and thus likely responded to conserved bacterial antigens. We therefore chose highly conserved E. coli proteins as candidate antigens for abnormal T cell responses in IBD and used high-throughput techniques for cloning, expression and purification under native conditions of a set of 271 conserved E. coli proteins for downstream immunologic studies. Results As a standardized procedure, genes were PCR amplified and cloned into the expression vector pQTEV2 in order to express proteins N-terminally fused to a seven-histidine-tag. Initial small-scale expression and purification under native conditions by metal chelate affinity chromatography indicated that the vast majority of target proteins were purified in high yields. Targets that revealed low yields after purification probably due to weak solubility were shuttled into Gateway (Invitrogen destination vectors in order to enhance solubility by N-terminal fusion of maltose binding protein (MBP, N-utilizing substance A (NusA, or glutathione S-transferase (GST to the target protein. In addition, recombinant proteins were treated with polymyxin B coated magnetic beads in order to remove lipopolysaccharide (LPS. Thus, 73% of the targeted proteins could be expressed and purified in large-scale to give soluble proteins in the range of 500

Identification of the 11-cis-specific retinyl-ester synthase in retinal Müller cells as multifunctional O-acyltransferase (MFAT)

Science.gov (United States)

Kaylor, Joanna J.; Cook, Jeremy D.; Makshanoff, Jacob; Bischoff, Nicholas; Yong, Jennifer; Travis, Gabriel H.

2014-01-01

Absorption of a photon by a rhodopsin or cone-opsin pigment isomerizes its 11-cis-retinaldehyde (11-cis-RAL) chromophore to all-trans-retinaldehyde (all-trans-RAL), which dissociates after a brief period of activation. Light sensitivity is restored to the resulting apo-opsin when it recombines with another 11-cis-RAL. Conversion of all-trans-RAL to 11-cis-RAL is carried out by an enzyme pathway called the visual cycle in cells of the retinal pigment epithelium. A second visual cycle is present in Müller cells of the retina. The retinol isomerase for this noncanonical pathway is dihydroceramide desaturase (DES1), which catalyzes equilibrium isomerization of retinol. Because 11-cis-retinol (11-cis-ROL) constitutes only a small fraction of total retinols in an equilibrium mixture, a subsequent step involving selective removal of 11-cis-ROL is required to drive synthesis of 11-cis-retinoids for production of visual chromophore. Selective esterification of 11-cis-ROL is one possibility. Crude homogenates of chicken retinas rapidly convert all-trans-ROL to 11-cis-retinyl esters (11-cis-REs) with minimal formation of other retinyl-ester isomers. This enzymatic activity implies the existence of an 11-cis-specific retinyl-ester synthase in Müller cells. Here, we evaluated multifunctional O-acyltransferase (MFAT) as a candidate for this 11-cis-RE-synthase. MFAT exhibited much higher catalytic efficiency as a synthase of 11-cis-REs versus other retinyl-ester isomers. Further, we show that MFAT is expressed in Müller cells. Finally, homogenates of cells coexpressing DES1 and MFAT catalyzed the conversion of all-trans-ROL to 11-cis-RP, similar to what we observed with chicken-retina homogenates. MFAT is therefore an excellent candidate for the retinyl-ester synthase that cooperates with DES1 to drive synthesis of 11-cis-retinoids by mass action. PMID:24799687

Detection of trans-cis flips and peptide-plane flips in protein structures

NARCIS (Netherlands)

Touw, W.G.; Joosten, R.P.; Vriend, G.

2015-01-01

A coordinate-based method is presented to detect peptide bonds that need correction either by a peptide-plane flip or by a trans-cis inversion of the peptide bond. When applied to the whole Protein Data Bank, the method predicts 4617 trans-cis flips and many thousands of hitherto unknown

CRISPR-Cas9 epigenome editing enables high-throughput screening for functional regulatory elements in the human genome.

Science.gov (United States)

Klann, Tyler S; Black, Joshua B; Chellappan, Malathi; Safi, Alexias; Song, Lingyun; Hilton, Isaac B; Crawford, Gregory E; Reddy, Timothy E; Gersbach, Charles A

2017-06-01

Large genome-mapping consortia and thousands of genome-wide association studies have identified non-protein-coding elements in the genome as having a central role in various biological processes. However, decoding the functions of the millions of putative regulatory elements discovered in these studies remains challenging. CRISPR-Cas9-based epigenome editing technologies have enabled precise perturbation of the activity of specific regulatory elements. Here we describe CRISPR-Cas9-based epigenomic regulatory element screening (CERES) for improved high-throughput screening of regulatory element activity in the native genomic context. Using dCas9 KRAB repressor and dCas9 p300 activator constructs and lentiviral single guide RNA libraries to target DNase I hypersensitive sites surrounding a gene of interest, we carried out both loss- and gain-of-function screens to identify regulatory elements for the β-globin and HER2 loci in human cells. CERES readily identified known and previously unidentified regulatory elements, some of which were dependent on cell type or direction of perturbation. This technology allows the high-throughput functional annotation of putative regulatory elements in their native chromosomal context.

Enhancing Conservation with High Resolution Productivity Datasets for the Conterminous United States

Science.gov (United States)

Robinson, Nathaniel Paul

Human driven alteration of the earth's terrestrial surface is accelerating through land use changes, intensification of human activity, climate change, and other anthropogenic pressures. These changes occur at broad spatio-temporal scales, challenging our ability to effectively monitor and assess the impacts and subsequent conservation strategies. While satellite remote sensing (SRS) products enable monitoring of the earth's terrestrial surface continuously across space and time, the practical applications for conservation and management of these products are limited. Often the processes driving ecological change occur at fine spatial resolutions and are undetectable given the resolution of available datasets. Additionally, the links between SRS data and ecologically meaningful metrics are weak. Recent advances in cloud computing technology along with the growing record of high resolution SRS data enable the development of SRS products that quantify ecologically meaningful variables at relevant scales applicable for conservation and management. The focus of my dissertation is to improve the applicability of terrestrial gross and net primary productivity (GPP/NPP) datasets for the conterminous United States (CONUS). In chapter one, I develop a framework for creating high resolution datasets of vegetation dynamics. I use the entire archive of Landsat 5, 7, and 8 surface reflectance data and a novel gap filling approach to create spatially continuous 30 m, 16-day composites of the normalized difference vegetation index (NDVI) from 1986 to 2016. In chapter two, I integrate this with other high resolution datasets and the MOD17 algorithm to create the first high resolution GPP and NPP datasets for CONUS. I demonstrate the applicability of these products for conservation and management, showing the improvements beyond currently available products. In chapter three, I utilize this dataset to evaluate the relationships between land ownership and terrestrial production