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Sample records for putative cis-regulatory elements

  1. 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

  2. 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.

  3. Putative cis-regulatory elements associated with heat shock genes activated during excystation of Cryptosporidium parvum.

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    Benjamin Cohn

    Full Text Available BACKGROUND: Cryptosporidiosis is a ubiquitous infectious disease, caused by the protozoan parasites Cryptosporidium hominis and C. parvum, leading to acute, persistent and chronic diarrhea worldwide. Although the complications of this disease can be serious, even fatal, in immunocompromised patients of any age, they have also been found to lead to long term effects, including growth inhibition and impaired cognitive development, in infected immunocompetent children. The Cryptosporidium life cycle alternates between a dormant stage, the oocyst, and a highly replicative phase that includes both asexual vegetative stages as well as sexual stages, implying fine genetic regulatory mechanisms. The parasite is extremely difficult to study because it cannot be cultured in vitro and animal models are equally challenging. The recent publication of the genome sequence of C. hominis and C. parvum has, however, significantly advanced our understanding of the biology and pathogenesis of this parasite. METHODOLOGY/PRINCIPAL FINDINGS: Herein, our goal was to identify cis-regulatory elements associated with heat shock response in Cryptosporidium using a combination of in silico and real time RT-PCR strategies. Analysis with Gibbs-Sampling algorithms of upstream non-translated regions of twelve genes annotated as heat shock proteins in the Cryptosporidium genome identified a highly conserved over-represented sequence motif in eleven of them. RT-PCR analyses, described herein and also by others, show that these eleven genes bearing the putative element are induced concurrent with excystation of parasite oocysts via heat shock. CONCLUSIONS/SIGNIFICANCE: Our analyses suggest that occurrences of a motif identified in the upstream regions of the Cryptosporidium heat shock genes represent parts of the transcriptional apparatus and function as stress response elements that activate expression of these genes during excystation, and possibly at other stages in the life

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

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    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

    2011-07-01

    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.

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

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    Guo Xiang

    2008-12-01

    Full Text Available Abstract Background Parasites in the genus Theileria cause lymphoproliferative diseases in cattle, resulting in enormous socio-economic losses. The availability of the genome sequences and annotation for T. parva and T. annulata has facilitated the study of parasite biology and their relationship with host cell transformation and tropism. However, the mechanism of transcriptional regulation in this genus, which may be key to understanding fundamental aspects of its parasitology, remains poorly understood. In this study, we analyze the evolution of non-coding sequences in the Theileria genome and identify conserved sequence elements that may be involved in gene regulation of these parasitic species. Results Intergenic regions and introns in Theileria are short, and their length distributions are considerably right-skewed. Intergenic regions flanked by genes in 5'-5' orientation tend to be longer and slightly more AT-rich than those flanked by two stop codons; intergenic regions flanked by genes in 3'-5' orientation have intermediate values of length and AT composition. Intron position is negatively correlated with intron length, and positively correlated with GC content. Using stringent criteria, we identified a set of high-quality orthologous non-coding sequences between T. parva and T. annulata, and determined the distribution of selective constraints across regions, which are shown to be higher close to translation start sites. A positive correlation between constraint and length in both intergenic regions and introns suggests a tight control over length expansion of non-coding regions. Genome-wide searches for functional elements revealed several conserved motifs in intergenic regions of Theileria genomes. Two such motifs are preferentially located within the first 60 base pairs upstream of transcription start sites in T. parva, are preferentially associated with specific protein functional categories, and have significant similarity to know

  6. A universal algorithm for genome-wide in silicio identification of biologically significant gene promoter putative cis-regulatory-elements; identification of new elements for reactive oxygen species and sucrose signaling in Arabidopsis.

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    Geisler, Matt; Kleczkowski, Leszek A; Karpinski, Stanislaw

    2006-02-01

    Short motifs of many cis-regulatory elements (CREs) can be found in the promoters of most Arabidopsis genes, and this raises the question of how their presence can confer specific regulation. We developed a universal algorithm to test the biological significance of CREs by first identifying every Arabidopsis gene with a CRE and then statistically correlating the presence or absence of the element with the gene expression profile on multiple DNA microarrays. This algorithm was successfully verified for previously characterized abscisic acid, ethylene, sucrose and drought responsive CREs in Arabidopsis, showing that the presence of these elements indeed correlates with treatment-specific gene induction. Later, we used standard motif sampling methods to identify 128 putative motifs induced by excess light, reactive oxygen species and sucrose. Our algorithm was able to filter 20 out of 128 novel CREs which significantly correlated with gene induction by either heat, reactive oxygen species and/or sucrose. The position, orientation and sequence specificity of CREs was tested in silicio by analyzing the expression of genes with naturally occurring sequence variations. In three novel CREs the forward orientation correlated with sucrose induction and the reverse orientation with sucrose suppression. The functionality of the predicted novel CREs was experimentally confirmed using Arabidopsis cell-suspension cultures transformed with short promoter fragments or artificial promoters fused with the GUS reporter gene. Our genome-wide analysis opens up new possibilities for in silicio verification of the biological significance of newly discovered CREs, and allows for subsequent selection of such CREs for experimental studies.

  7. Cis-regulatory elements: molecular mechanisms and evolutionary processes underlying divergence.

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    Wittkopp, Patricia J; Kalay, Gizem

    2011-12-06

    Cis-regulatory sequences, such as enhancers and promoters, control development and physiology by regulating gene expression. Mutations that affect the function of these sequences contribute to phenotypic diversity within and between species. With many case studies implicating divergent cis-regulatory activity in phenotypic evolution, researchers have recently begun to elucidate the genetic and molecular mechanisms that are responsible for cis-regulatory divergence. Approaches include detailed functional analysis of individual cis-regulatory elements and comparing mechanisms of gene regulation among species using the latest genomic tools. Despite the limited number of mechanistic studies published to date, this work shows how cis-regulatory activity can diverge and how studies of cis-regulatory divergence can address long-standing questions about the genetic mechanisms of phenotypic evolution.

  8. 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.

  9. A Simple Grammar Defines Activating and Repressing cis-Regulatory Elements in Photoreceptors

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    Michael A. White

    2016-10-01

    Full Text Available Transcription factors often activate and repress different target genes in the same cell. How activation and repression are encoded by different arrangements of transcription factor binding sites in cis-regulatory elements is poorly understood. We investigated how sites for the transcription factor CRX encode both activation and repression in photoreceptors by assaying thousands of genomic and synthetic cis-regulatory elements in wild-type and Crx−/− retinas. We found that sequences with high affinity for CRX repress transcription, whereas sequences with lower affinity activate. This rule is modified by a cooperative interaction between CRX sites and sites for the transcription factor NRL, which overrides the repressive effect of high affinity for CRX. Our results show how simple rearrangements of transcription factor binding sites encode qualitatively different responses to a single transcription factor and explain how CRX plays multiple cis-regulatory roles in the same cell.

  10. [Role of genes and their cis-regulatory elements during animal morphological evolution].

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    Sun, Boyuan; Tu, Jianbo; Li, Ying; Yang, Mingyao

    2014-06-01

    Cis-regulatory hypothesis is one of the most important theories in evolutionary developmental biology (evo-devo), which claims that evolution of cis-regulatory elements (CREs) plays a key role during evolution of morphology. However, an increasing number of experimental results show that cis-regulatory hypothesis alone is not far enough to explain the complexity of evo-devo processes. Other modifications, including mutations of protein coding, gene and genome duplications, and flexibility of homeodomains and CREs, also cause the morphological changes in animals. In this review, we retrospect the recent results of evolution of CREs and genes associated with CREs and discuss new methods and trends for research in evo-devo.

  11. 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

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

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    Mao, Grace; Brody, James P

    2007-11-09

    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 16bp region of Saccharomyces cerevisiae chromosome 5 onto a gold surface. This 16bp 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 0.014s(-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.

  13. Computational identification of new structured cis-regulatory elements in the 3'-untranslated region of human protein coding genes.

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    Chen, Xiaowei Sylvia; Brown, Chris M

    2012-10-01

    Messenger ribonucleic acids (RNAs) contain a large number of cis-regulatory RNA elements that function in many types of post-transcriptional regulation. These cis-regulatory elements are often characterized by conserved structures and/or sequences. Although some classes are well known, given the wide range of RNA-interacting proteins in eukaryotes, it is likely that many new classes of cis-regulatory elements are yet to be discovered. An approach to this is to use computational methods that have the advantage of analysing genomic data, particularly comparative data on a large scale. In this study, a set of structural discovery algorithms was applied followed by support vector machine (SVM) classification. We trained a new classification model (CisRNA-SVM) on a set of known structured cis-regulatory elements from 3'-untranslated regions (UTRs) and successfully distinguished these and groups of cis-regulatory elements not been strained on from control genomic and shuffled sequences. The new method outperformed previous methods in classification of cis-regulatory RNA elements. This model was then used to predict new elements from cross-species conserved regions of human 3'-UTRs. Clustering of these elements identified new classes of potential cis-regulatory elements. The model, training and testing sets and novel human predictions are available at: http://mRNA.otago.ac.nz/CisRNA-SVM.

  14. The identification of cis-regulatory elements: A review from a machine learning perspective.

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    Li, Yifeng; Chen, Chih-Yu; Kaye, Alice M; Wasserman, Wyeth W

    2015-12-01

    The majority of the human genome consists of non-coding regions that have been called junk DNA. However, recent studies have unveiled that these regions contain cis-regulatory elements, such as promoters, enhancers, silencers, insulators, etc. These regulatory elements can play crucial roles in controlling gene expressions in specific cell types, conditions, and developmental stages. Disruption to these regions could contribute to phenotype changes. Precisely identifying regulatory elements is key to deciphering the mechanisms underlying transcriptional regulation. Cis-regulatory events are complex processes that involve chromatin accessibility, transcription factor binding, DNA methylation, histone modifications, and the interactions between them. The development of next-generation sequencing techniques has allowed us to capture these genomic features in depth. Applied analysis of genome sequences for clinical genetics has increased the urgency for detecting these regions. However, the complexity of cis-regulatory events and the deluge of sequencing data require accurate and efficient computational approaches, in particular, machine learning techniques. In this review, we describe machine learning approaches for predicting transcription factor binding sites, enhancers, and promoters, primarily driven by next-generation sequencing data. Data sources are provided in order to facilitate testing of novel methods. The purpose of this review is to attract computational experts and data scientists to advance this field.

  15. 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

  16. Engineering Synthetic cis-Regulatory Elements for Simultaneous Recognition of Three Transcriptional Factors in Bacteria.

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    Amores, Gerardo Ruiz; Guazzaroni, María-Eugenia; Silva-Rocha, Rafael

    2015-12-18

    Recognition of cis-regulatory elements by transcription factors (TF) at target promoters is crucial to gene regulation in bacteria. In this process, binding of TFs to their cognate sequences depends on a set of physical interactions between these proteins and specific nucleotides in the operator region. Previously, we showed that in silico optimization algorithms are able to generate short sequences that are recognized by two different TFs of Escherichia coli, namely, CRP and IHF, thus generating an AND logic gate. Here, we expanded this approach in order to engineer DNA sequences that can be simultaneously recognized by three unrelated TFs (CRP, IHF, and Fis). Using in silico optimization and experimental validation strategies, we were able to obtain a candidate promoter (Plac-CFI1) regulated by only two TFs with an AND logic, thus demonstrating a limitation in the design. Subsequently, we modified the algorithm to allow the optimization of extended sequences, and were able to design two synthetic promoters (PCFI20-1 and PCFI22-5) that were functional in vivo. Expression assays in E. coli mutant strains for each TF revealed that while CRP positively regulates the promoter activities, IHF and Fis are strong repressors of both the promoter variants. Taken together, our results demonstrate the potential of in silico strategies in bacterial synthetic promoter engineering. Furthermore, the study also shows how small modifications in cis-regulatory elements can drastically affect the final logic of the resulting promoter.

  17. Dissection of cis-regulatory elements in the C. elegans Hox gene egl-5 promoter.

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    Teng, Yingqi; Girard, Lisa; Ferreira, Henrique B; Sternberg, Paul W; Emmons, Scott W

    2004-12-15

    Hox genes are highly conserved segmental identity genes well known for their complex expression patterns and divergent targets. Here we present an analysis of cis-regulatory elements in the Caenorhabditis elegans Hox gene egl-5, which is expressed in multiple tissues in the posterior region of the nematode. We have utilized phylogenetic footprinting to efficiently identify cis-regulatory elements and have characterized these with gfp reporters and tissue-specific rescue experiments. We have found that the complex expression pattern of egl-5 is the cumulative result of the activities of multiple tissue or local region-specific activator sequences that are conserved both in sequence and near-perfect order in the related nematode Caenorhabditis briggsae. Two conserved regulatory blocks analyzed in detail contain multiple sites for both positively and negatively acting factors. One of these regions may promote activation of egl-5 in certain cells via the Wnt pathway. Positively acting regions are repressed in inappropriate tissues by additional negative pathways acting at other sites within the promoter. Our analysis has allowed us to implicate several new regulatory factors significant to the control of egl-5 expression.

  18. Lessons from Domestication: Targeting Cis-Regulatory Elements for Crop Improvement.

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    Swinnen, Gwen; Goossens, Alain; Pauwels, Laurens

    2016-06-01

    Domestication of wild plant species has provided us with crops that serve our human nutritional needs. Advanced DNA sequencing has propelled the unveiling of underlying genetic changes associated with domestication. Interestingly, many changes reside in cis-regulatory elements (CREs) that control the expression of an unmodified coding sequence. Sequence variation in CREs can impact gene expression levels, but also developmental timing and tissue specificity of expression. When genes are involved in multiple pathways or active in several organs and developmental stages CRE modifications are favored in contrast to mutations in coding regions, due to the lack of detrimental pleiotropic effects. Therefore, learning from domestication, we propose that CREs are interesting targets for genome editing to create new alleles for plant breeding.

  19. Computational discovery of soybean promoter cis-regulatory elements for the construction of soybean cyst nematode inducible synthetic promoters

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    Computational methods offer great hope but limited accuracy in the prediction of functional cis-regulatory elements; improvements are needed to enable synthetic promoter design. We applied an ensemble strategy for de novo soybean cyst nematode (SCN)-inducible motif discovery among promoters of 18 co...

  20. Analysis of mammalian cis-regulatory DNA elements by homologous recombination.

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    Fiering, S; Bender, M A; Groudine, M

    1999-01-01

    The use of homologous recombination to modify and thereby functionally analyze cis-regulatory DNA elements in mammalian cells has become an important approach in mammalian gene expression research. We have emphasized the necessity of designing a system that allows the removal of selectable markers used in targeting and facilitates the further modification of the region under study. To perform these tasks, we presently favor making an initial HR-mediated replacement of the entire element under study with an active positive selectable marker in combination with either an inactive second positive selectable marker or an active negative selectable marker. The plug and socket system, in which an inactive selectable marker is complemented by HR, is the most dependable and well-characterized option for making secondary modifications. However, the double-replacement system has certain advantages, and the recently developed RMCE approach, which allows replacement of a negative selectable marker by site-specific recombinase-mediated insertion without using a positive selectable marker, will likely prove very valuable in future experiments. Each of the systems, or combinations thereof, should be considered in light of the specifics of any given experiment to select the most appropriate option. Although the emphasis of this article has been the analysis of cis-acting regulatory elements involved in transcription, these same approaches can be used to analyze other regulatory elements (e.g., origins of replication) and to make multiple subtle mutations in polypeptides.

  1. cis-regulatory elements involved in ultraviolet light regulation and plant defense.

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    Wingender, R; Röhrig, H; Höricke, C; Schell, J

    1990-10-01

    An elicitor-regulated transient expression system was established in soybean protoplasts that allowed the identification of cis-regulatory elements involved in plant defense. The 5' region of an ultraviolet (UV) light-inducible and elicitor-inducible chs gene (chs1) of soybean was subjected to deletion analysis with the help of chimeric chs-nptII/gus gene constructs. This analysis delimited the sequences necessary for elicitor inducibility to -175 and -134 of the chs1 promoter. The same soybean sequences were able to direct elicitor inducibility in parsley protoplasts, suggesting a conserved function of cis-acting elements involved in plant defense. In addition, this region of the soybean promoter also promotes UV light inducibility in parsley protoplasts. However, in contrast to the elicitor induction, correct regulation was not observed after UV light induction when sequences downstream of -75 were replaced by a heterologous minimal promoter. This result indicates that at least two cis-acting elements are involved in UV light induction.

  2. Identification of distal cis-regulatory elements at mouse mitoferrin loci using zebrafish transgenesis.

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    Amigo, Julio D; Yu, Ming; Troadec, Marie-Berengere; Gwynn, Babette; Cooney, Jeffrey D; Lambert, Amy J; Chi, Neil C; Weiss, Mitchell J; Peters, Luanne L; Kaplan, Jerry; Cantor, Alan B; Paw, Barry H

    2011-04-01

    Mitoferrin 1 (Mfrn1; Slc25a37) and mitoferrin 2 (Mfrn2; Slc25a28) function as essential mitochondrial iron importers for heme and Fe/S cluster biogenesis. A genetic deficiency of Mfrn1 results in a profound hypochromic anemia in vertebrate species. To map the cis-regulatory modules (CRMs) that control expression of the Mfrn genes, we utilized genome-wide chromatin immunoprecipitation (ChIP) datasets for the major erythroid transcription factor GATA-1. We identified the CRMs that faithfully drive the expression of Mfrn1 during blood and heart development and Mfrn2 ubiquitously. Through in vivo analyses of the Mfrn-CRMs in zebrafish and mouse, we demonstrate their functional and evolutionary conservation. Using knockdowns with morpholinos and cell sorting analysis in transgenic zebrafish embryos, we show that GATA-1 directly regulates the expression of Mfrn1. Mutagenesis of individual GATA-1 binding cis elements (GBE) demonstrated that at least two of the three GBE within this CRM are functionally required for GATA-mediated transcription of Mfrn1. Furthermore, ChIP assays demonstrate switching from GATA-2 to GATA-1 at these elements during erythroid maturation. Our results provide new insights into the genetic regulation of mitochondrial function and iron homeostasis and, more generally, illustrate the utility of genome-wide ChIP analysis combined with zebrafish transgenesis for identifying long-range transcriptional enhancers that regulate tissue development.

  3. Recurrent modification of a conserved cis-regulatory element underlies fruit fly pigmentation diversity.

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    William A Rogers

    2013-08-01

    Full Text Available The development of morphological traits occurs through the collective action of networks of genes connected at the level of gene expression. As any node in a network may be a target of evolutionary change, the recurrent targeting of the same node would indicate that the path of evolution is biased for the relevant trait and network. Although examples of parallel evolution have implicated recurrent modification of the same gene and cis-regulatory element (CRE, little is known about the mutational and molecular paths of parallel CRE evolution. In Drosophila melanogaster fruit flies, the Bric-à-brac (Bab transcription factors control the development of a suite of sexually dimorphic traits on the posterior abdomen. Female-specific Bab expression is regulated by the dimorphic element, a CRE that possesses direct inputs from body plan (ABD-B and sex-determination (DSX transcription factors. Here, we find that the recurrent evolutionary modification of this CRE underlies both intraspecific and interspecific variation in female pigmentation in the melanogaster species group. By reconstructing the sequence and regulatory activity of the ancestral Drosophila melanogaster dimorphic element, we demonstrate that a handful of mutations were sufficient to create independent CRE alleles with differing activities. Moreover, intraspecific and interspecific dimorphic element evolution proceeded with little to no alterations to the known body plan and sex-determination regulatory linkages. Collectively, our findings represent an example where the paths of evolution appear biased to a specific CRE, and drastic changes in function were accompanied by deep conservation of key regulatory linkages.

  4. 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

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

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    Deb, Arindam; Kundu, Sudip

    2015-01-01

    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 sequences and suitable

  6. 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.

  7. 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

  8. Identification of Distal cis-Regulatory Elements at Mouse Mitoferrin Loci Using Zebrafish Transgenesis▿

    OpenAIRE

    Amigo, J. D.; Yu, M.; Troadec, M.-B.; Gwynn, B; Cooney, J. D.; Lambert, A.J.; Chi, N. C.; Weiss, M J; Peters, L. L.; Kaplan, J; Cantor, Alan B.; Paw, Barry Htin

    2011-01-01

    Mitoferrin 1 (Mfrn1; Slc25a37) and mitoferrin 2 (Mfrn2; Slc25a28) function as essential mitochondrial iron importers for heme and Fe/S cluster biogenesis. A genetic deficiency of Mfrn1 results in a profound hypochromic anemia in vertebrate species. To map the cis-regulatory modules (CRMs) that control expression of the Mfrn genes, we utilized genome-wide chromatin immunoprecipitation (ChIP) datasets for the major erythroid transcription factor GATA-1. We identified the CRMs that faithfully dr...

  9. Prioritizing predicted cis-regulatory elements for co-expressed gene sets based on Lasso regression models.

    Science.gov (United States)

    Hu, Hong; Roqueiro, Damian; Dai, Yang

    2011-01-01

    Computational prediction of cis-regulatory elements for a set of co-expressed genes based on sequence analysis provides an overwhelming volume of potential transcription factor binding sites. It presents a challenge to prioritize transcription factors for regulatory functional studies. A novel approach based on the use of Lasso regression models is proposed to address this problem. We examine the ability of the Lasso model using time-course microarray data obtained from a comprehensive study of gene expression profiles in skin and mucosal wounds in mouse over all stages of wound healing.

  10. Creation of cis-regulatory elements during sea urchin evolution by co-option and optimization of a repetitive sequence adjacent to the spec2a gene.

    Science.gov (United States)

    Dayal, Sandeep; Kiyama, Takae; Villinski, Jeffrey T; Zhang, Ning; Liang, Shuguang; Klein, William H

    2004-09-15

    The creation, preservation, and degeneration of cis-regulatory elements controlling developmental gene expression are fundamental genome-level evolutionary processes about which little is known. Here, we identify critical differences in cis-regulatory elements controlling the expression of the sea urchin aboral ectoderm-specific spec genes. We found multiple copies of a repetitive sequence element termed RSR in genomes of species within the Strongylocentrotidae family, but RSRs were not detected in genomes of species outside Strongylocentrotidae. spec genes in Strongylocentrotus purpuratus are invariably associated with RSRs, and the spec2a RSR functioned as a transcriptional enhancer and displayed greater activity than did spec1 or spec2c RSRs. Single-base pair differences at two cis-regulatory elements within the spec2a RSR increased the binding affinities of four transcription factors, SpCCAAT-binding factor at one element and SpOtx, SpGoosecoid, and SpGATA-E at another. The cis-regulatory elements to which these four factors bound were recent evolutionary acquisitions that acted to either activate or repress transcription, depending on the cell type. These elements were found in the spec2a RSR ortholog in Strongylocentrotus pallidus but not in RSR orthologs of Strongylocentrotus droebachiensis or Hemicentrotus pulcherrimus. Our results indicated that a dynamic pattern of cis-regulatory element evolution exists for spec genes despite their conserved aboral ectoderm expression.

  11. Distinct functional constraints partition sequence conservation in a cis-regulatory element.

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    Antoine Barrière

    2011-06-01

    Full Text Available Different functional constraints contribute to different evolutionary rates across genomes. To understand why some sequences evolve faster than others in a single cis-regulatory locus, we investigated function and evolutionary dynamics of the promoter of the Caenorhabditis elegans unc-47 gene. We found that this promoter consists of two distinct domains. The proximal promoter is conserved and is largely sufficient to direct appropriate spatial expression. The distal promoter displays little if any conservation between several closely related nematodes. Despite this divergence, sequences from all species confer robustness of expression, arguing that this function does not require substantial sequence conservation. We showed that even unrelated sequences have the ability to promote robust expression. A prominent feature shared by all of these robustness-promoting sequences is an AT-enriched nucleotide composition consistent with nucleosome depletion. Because general sequence composition can be maintained despite sequence turnover, our results explain how different functional constraints can lead to vastly disparate rates of sequence divergence within a promoter.

  12. Shared Enhancer Activity in the Limbs and Phallus and Functional Divergence of a Limb-Genital cis-Regulatory Element in Snakes.

    Science.gov (United States)

    Infante, Carlos R; Mihala, Alexandra G; Park, Sungdae; Wang, Jialiang S; Johnson, Kenji K; Lauderdale, James D; Menke, Douglas B

    2015-10-12

    The amniote phallus and limbs differ dramatically in their morphologies but share patterns of signaling and gene expression in early development. Thus far, the extent to which genital and limb transcriptional networks also share cis-regulatory elements has remained unexplored. We show that many limb enhancers are retained in snake genomes, suggesting that these elements may function in non-limb tissues. Consistent with this, our analysis of cis-regulatory activity in mice and Anolis lizards reveals that patterns of enhancer activity in embryonic limbs and genitalia overlap heavily. In mice, deletion of HLEB, an enhancer of Tbx4, produces defects in hindlimbs and genitalia, establishing the importance of this limb-genital enhancer for development of these different appendages. Further analyses demonstrate that the HLEB of snakes has lost hindlimb enhancer function while retaining genital activity. Our findings identify roles for Tbx4 in genital development and highlight deep similarities in cis-regulatory activity between limbs and genitalia.

  13. A comparative analysis of the evolution, expression, and cis-regulatory element of polygalacturonase genes in grasses and dicots.

    Science.gov (United States)

    Liang, Ying; Yu, Youjian; Cui, Jinlong; Lyu, Meiling; Xu, Liai; Cao, Jiashu

    2016-11-01

    Cell walls are a distinguishing characteristic of plants essential to their survival. The pectin content of primary cell walls in grasses and dicots is distinctly different. Polygalacturonases (PGs) can degrade pectins and participate in multiple developmental processes of plants. This study comprehensively compared the evolution, expression, and cis-regulatory element of PGs in grasses and dicots. A total of 577 PGs identified from five grasses and five dicots fell into seven clades. Evolutionary analysis demonstrated the distinct differences between grasses and dicots in patterns of gene duplication and loss, and evolutionary rates. Grasses generally contained much fewer clade C and F members than dicots. We found that this disparity was the result of less duplication and more gene losses in grasses. More duplications occurred in clades D and E, and expression analysis showed that most of clade E members were expressed ubiquitously at a high overall level and clade D members were closely related to male reproduction in both grasses and dicots, suggesting their biological functions were highly conserved across species. In addition to the general role in reproductive development, PGs of clades C and F specifically played roles in root development in dicots, shedding light on organ differentiation between the two groups of plants. A regulatory element analysis of clade C and F members implied that possible functions of PGs in specific biological responses contributed to their expansion and preservation. This work can improve the knowledge of PGs in plants generally and in grasses specifically and is beneficial to functional studies.

  14. Characterization of "cis"-regulatory elements ("c"RE) associated with mammary gland function

    Science.gov (United States)

    The Bos taurus genome assembly has propelled dairy science into a new era; still, most of the information encoded in the genome has not yet been decoded. The human Encyclopedia of DNA Elements (ENCODE) project has spearheaded the identification and annotation of functional genomic elements in the hu...

  15. Global reorganisation of cis-regulatory units upon lineage commitment of human embryonic stem cells.

    Science.gov (United States)

    Freire-Pritchett, Paula; Schoenfelder, Stefan; Várnai, Csilla; Wingett, Steven W; Cairns, Jonathan; Collier, Amanda J; García-Vílchez, Raquel; Furlan-Magaril, Mayra; Osborne, Cameron S; Fraser, Peter; Rugg-Gunn, Peter J; Spivakov, Mikhail

    2017-03-23

    Long-range cis-regulatory elements such as enhancers coordinate cell-specific transcriptional programmes by engaging in DNA looping interactions with target promoters. Deciphering the interplay between the promoter connectivity and activity of cis-regulatory elements during lineage commitment is crucial for understanding developmental transcriptional control. Here, we use Promoter Capture Hi-C to generate a high-resolution atlas of chromosomal interactions involving ~22,000 gene promoters in human pluripotent and lineage-committed cells, identifying putative target genes for known and predicted enhancer elements. We reveal extensive dynamics of cis-regulatory contacts upon lineage commitment, including the acquisition and loss of promoter interactions. This spatial rewiring occurs preferentially with predicted changes in the activity of cis-regulatory elements and is associated with changes in target gene expression. Our results provide a global and integrated view of promoter interactome dynamics during lineage commitment of human pluripotent cells.

  16. Evaluation of combinatorial cis-regulatory elements for stable gene expression in chicken cells

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    Seo Hee W

    2010-09-01

    Full Text Available Abstract Background Recent successes in biotechnological application of birds are based on their unique physiological traits such as unlimited manipulability onto developing embryos and simple protein constituents of the eggs. However it is not likely that target protein is produced as kinetically expected because various factors affect target gene expression. Although there have been various attempts to minimize the silencing of transgenes, a generalized study that uses multiple cis-acting elements in chicken has not been made. The aim of the present study was to analyze whether various cis-acting elements can help to sustain transgene expression in chicken fibroblasts. Results We investigated the optimal transcriptional regulatory elements for enhancing stable transgene expression in chicken cells. We generated eight constructs that encode enhanced green fluorescent protein (eGFP driven by either CMV or CAG promoters (including the control, containing three types of key regulatory elements: a chicken lysozyme matrix attachment region (cMAR, 5'-DNase I-hypersensitive sites 4 (cHS4, and the woodchuck hepatitis virus posttranscriptional regulatory element (WPRE. Then we transformed immortalized chicken embryonic fibroblasts with these constructs by electroporation, and after cells were expanded under G418 selection, analyzed mRNA levels and mean fluorescence intensity (MFI by quantitative real-time PCR and flow cytometry, respectively. We found that the copy number of each construct significantly decreased as the size of the construct increased (R2 = 0.701. A significant model effect was found in the expression level among various constructs in both mRNA and protein (P cis-acting elements decreased the level of gene silencing as well as the coefficient of variance of eGFP-expressing cells (P Conclusions Our current data show that an optimal combination of cis-acting elements and promoters/enhancers for sustaining gene expression in chicken cells

  17. Separate elements of the TERMINAL FLOWER 1 cis-regulatory region integrate pathways to control flowering time and shoot meristem identity.

    Science.gov (United States)

    Serrano-Mislata, Antonio; Fernández-Nohales, Pedro; Doménech, María J; Hanzawa, Yoshie; Bradley, Desmond; Madueño, Francisco

    2016-09-15

    TERMINAL FLOWER 1 (TFL1) is a key regulator of Arabidopsis plant architecture that responds to developmental and environmental signals to control flowering time and the fate of shoot meristems. TFL1 expression is dynamic, being found in all shoot meristems, but not in floral meristems, with the level and distribution changing throughout development. Using a variety of experimental approaches we have analysed the TFL1 promoter to elucidate its functional structure. TFL1 expression is based on distinct cis-regulatory regions, the most important being located 3' of the coding sequence. Our results indicate that TFL1 expression in the shoot apical versus lateral inflorescence meristems is controlled through distinct cis-regulatory elements, suggesting that different signals control expression in these meristem types. Moreover, we identified a cis-regulatory region necessary for TFL1 expression in the vegetative shoot and required for a wild-type flowering time, supporting that TFL1 expression in the vegetative meristem controls flowering time. Our study provides a model for the functional organisation of TFL1 cis-regulatory regions, contributing to our understanding of how developmental pathways are integrated at the genomic level of a key regulator to control plant architecture. © 2016. Published by The Company of Biologists Ltd.

  18. An arthropod cis-regulatory element functioning in sensory organ precursor development dates back to the Cambrian

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    Simpson Pat

    2010-09-01

    Full Text Available Abstract Background An increasing number of publications demonstrate conservation of function of cis-regulatory elements without sequence similarity. In invertebrates such functional conservation has only been shown for closely related species. Here we demonstrate the existence of an ancient arthropod regulatory element that functions during the selection of neural precursors. The activity of genes of the achaete-scute (ac-sc family endows cells with neural potential. An essential, conserved characteristic of proneural genes is their ability to restrict their own activity to single or a small number of progenitor cells from their initially broad domains of expression. This is achieved through a process called lateral inhibition. A regulatory element, the sensory organ precursor enhancer (SOPE, is required for this process. First identified in Drosophila, the SOPE contains discrete binding sites for four regulatory factors. The SOPE of the Drosophila asense gene is situated in the 5' UTR. Results Through a manual comparison of consensus binding site sequences we have been able to identify a SOPE in UTR sequences of asense-like genes in species belonging to all four arthropod groups (Crustacea, Myriapoda, Chelicerata and Insecta. The SOPEs of the spider Cupiennius salei and the insect Tribolium castaneum are shown to be functional in transgenic Drosophila. This would place the origin of this regulatory sequence as far back as the last common ancestor of the Arthropoda, that is, in the Cambrian, 550 million years ago. Conclusions The SOPE is not detectable by inter-specific sequence comparison, raising the possibility that other ancient regulatory modules in invertebrates might have escaped detection.

  19. Evaluation of phylogenetic footprint discovery for predicting bacterial cis-regulatory elements and revealing their evolution

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    van Helden Jacques

    2008-01-01

    Full Text Available Abstract Background The detection of conserved motifs in promoters of orthologous genes (phylogenetic footprints has become a common strategy to predict cis-acting regulatory elements. Several software tools are routinely used to raise hypotheses about regulation. However, these tools are generally used as black boxes, with default parameters. A systematic evaluation of optimal parameters for a footprint discovery strategy can bring a sizeable improvement to the predictions. Results We evaluate the performances of a footprint discovery approach based on the detection of over-represented spaced motifs. This method is particularly suitable for (but not restricted to Bacteria, since such motifs are typically bound by factors containing a Helix-Turn-Helix domain. We evaluated footprint discovery in 368 Escherichia coli K12 genes with annotated sites, under 40 different combinations of parameters (taxonomical level, background model, organism-specific filtering, operon inference. Motifs are assessed both at the levels of correctness and significance. We further report a detailed analysis of 181 bacterial orthologs of the LexA repressor. Distinct motifs are detected at various taxonomical levels, including the 7 previously characterized taxon-specific motifs. In addition, we highlight a significantly stronger conservation of half-motifs in Actinobacteria, relative to Firmicutes, suggesting an intermediate state in specificity switching between the two Gram-positive phyla, and thereby revealing the on-going evolution of LexA auto-regulation. Conclusion The footprint discovery method proposed here shows excellent results with E. coli and can readily be extended to predict cis-acting regulatory signals and propose testable hypotheses in bacterial genomes for which nothing is known about regulation.

  20. Prediction of Cis-Regulatory Elements Controlling Genes Differentially Expressed by Retinal and Choroidal Vascular Endothelial Cells.

    Science.gov (United States)

    Choi, Dongseok; Appukuttan, Binoy; Binek, Sierra J; Planck, Stephen R; Stout, J Timothy; Rosenbaum, James T; Smith, Justine R

    2008-01-01

    Cultured endothelial cells of the human retina and choroid demonstrate distinct patterns of gene expression. We hypothesized that differential gene expression reflected differences in the interactions of transcription factors and respective cis-regulatory motifs(s) in these two emdothelial cell subpopulations, recognizing that motifs often exist as modules. We tested this hypothesis in silico by using TRANSFAC Professional and CisModule to identify cis-regulatory motifs and modules in genes that were differentially expressed by human retinal versus choroidal endothelial cells, as identified by analysis of a microarray data set. Motifs corresponding to eight transcription factors were significantly (p < 0.05) differentially abundant in genes that were relatively highly expressed in retinal (i.e., GCCR, HMGIY, HSF1, p53, VDR) or choroidal (i.e., E2F, YY1, ZF5) endothelial cells. Predicted cis-regulatory modules were quite different for these two groups of genes. Our findings raise the possibility of exploiting specific cis-regulatory motifs to target therapy at the ocular endothelial cells subtypes responsible for neovascular age-related macular degeneration or proliferative diabetic retinopathy.

  1. 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

  2. Cis-regulatory element-based genome-wide identification of DREB1/CBF targets in Arabidopsis

    Institute of Scientific and Technical Information of China (English)

    Huan Zhao; Weizhong Liu; Yong Hu; Xianglin Liu; Yikun He

    2008-01-01

    Microarray analysis is used to identify transcriptional targets.However,the direct targets of transcription factors cannot be distin guished from indirect ones;further,genes with low-expression levels cannot be identified by this method.In the present study,we exploi the cis-element dehydration-responsive element(DRE)that is known to be responsible for the transcription of DRE binding factor](DREB1)targets in the promoter region of all Arabidopsis genes.Putative targets whose promoters contain the elements were verified by both microarray and reverse transcription-polymerase chain reaction(RT-PCR)analysis.Five new DREB 1/CBF direct targets were identified.Compared with traditional microarray analysis,our method is convenient and cost-effective for identifying the downstream targets of transcription factors.

  3. Deletion hotspots in AMACR promoter CpG island are cis-regulatory elements controlling the gene expression in the colon.

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    Xiang Zhang

    2009-01-01

    Full Text Available Alpha-methylacyl-coenzyme A racemase (AMACR regulates peroxisomal beta-oxidation of phytol-derived, branched-chain fatty acids from red meat and dairy products -- suspected risk factors for colon carcinoma (CCa. AMACR was first found overexpressed in prostate cancer but not in benign glands and is now an established diagnostic marker for prostate cancer. Aberrant expression of AMACR was recently reported in Cca; however, little is known about how this gene is abnormally activated in cancer. By using a panel of immunostained-laser-capture-microdissected clinical samples comprising the entire colon adenoma-carcinoma sequence, we show that deregulation of AMACR during colon carcinogenesis involves two nonrandom events, resulting in the mutually exclusive existence of double-deletion at CG3 and CG10 and deletion of CG12-16 in a newly identified CpG island within the core promoter of AMACR. The double-deletion at CG3 and CG10 was found to be a somatic lesion. It existed in histologically normal colonic glands and tubular adenomas with low AMACR expression and was absent in villous adenomas and all CCas expressing variable levels of AMACR. In contrast, deletion of CG12-16 was shown to be a constitutional allele with a frequency of 43% in a general population. Its prevalence reached 89% in moderately differentiated CCas strongly expressing AMACR but only existed at 14% in poorly differentiated CCas expressing little or no AMACR. The DNA sequences housing these deletions were found to be putative cis-regulatory elements for Sp1 at CG3 and CG10, and ZNF202 at CG12-16. Chromatin immunoprecipitation, siRNA knockdown, gel shift assay, ectopic expression, and promoter analyses supported the regulation by Sp1 and ZNF202 of AMACR gene expression in an opposite manner. Our findings identified key in vivo events and novel transcription factors responsible for AMACR regulation in CCas and suggested these AMACR deletions may have diagnostic/prognostic value for

  4. Parallel evolution of chordate cis-regulatory code for development.

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    Laura Doglio

    2013-11-01

    Full Text Available Urochordates are the closest relatives of vertebrates and at the larval stage, possess a characteristic bilateral chordate body plan. In vertebrates, the genes that orchestrate embryonic patterning are in part regulated by highly conserved non-coding elements (CNEs, yet these elements have not been identified in urochordate genomes. Consequently the evolution of the cis-regulatory code for urochordate development remains largely uncharacterised. Here, we use genome-wide comparisons between C. intestinalis and C. savignyi to identify putative urochordate cis-regulatory sequences. Ciona conserved non-coding elements (ciCNEs are associated with largely the same key regulatory genes as vertebrate CNEs. Furthermore, some of the tested ciCNEs are able to activate reporter gene expression in both zebrafish and Ciona embryos, in a pattern that at least partially overlaps that of the gene they associate with, despite the absence of sequence identity. We also show that the ability of a ciCNE to up-regulate gene expression in vertebrate embryos can in some cases be localised to short sub-sequences, suggesting that functional cross-talk may be defined by small regions of ancestral regulatory logic, although functional sub-sequences may also be dispersed across the whole element. We conclude that the structure and organisation of cis-regulatory modules is very different between vertebrates and urochordates, reflecting their separate evolutionary histories. However, functional cross-talk still exists because the same repertoire of transcription factors has likely guided their parallel evolution, exploiting similar sets of binding sites but in different combinations.

  5. 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

  6. The contribution of cis-regulatory elements to head-to-head gene pairs’ co-expression pattern

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Transcription regulation is one of the most critical pipelines in biological process,in which cis-elements play the role as gene expression regulators.We attempt to deduce the principles underlying the co-expression of "head-to-head" gene pairs by analyzing activities or behaviors of the shared cis-elements.A network component analysis was performed to estimate the impact of cis-elements on gene promoters and their activities under different conditions.Our discoveries reveal how biological system uses those regulatory elements to control the expression pattern of "head-to-head" gene pairs and the whole transcription regulation system.

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

    Science.gov (United States)

    Hadas, Yoav; Nitzan, Noa; Furley, Andrew J W; Kozlov, Serguei V; Klar, Avihu

    2013-01-01

    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.

  8. 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.

  9. 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.

  10. Identification of Predictive Cis-Regulatory Elements Using a Discriminative Objective Function and a Dynamic Search Space.

    Science.gov (United States)

    Karnik, Rahul; Beer, Michael A

    2015-01-01

    The generation of genomic binding or accessibility data from massively parallel sequencing technologies such as ChIP-seq and DNase-seq continues to accelerate. Yet state-of-the-art computational approaches for the identification of DNA binding motifs often yield motifs of weak predictive power. Here we present a novel computational algorithm called MotifSpec, designed to find predictive motifs, in contrast to over-represented sequence elements. The key distinguishing feature of this algorithm is that it uses a dynamic search space and a learned threshold to find discriminative motifs in combination with the modeling of motifs using a full PWM (position weight matrix) rather than k-mer words or regular expressions. We demonstrate that our approach finds motifs corresponding to known binding specificities in several mammalian ChIP-seq datasets, and that our PWMs classify the ChIP-seq signals with accuracy comparable to, or marginally better than motifs from the best existing algorithms. In other datasets, our algorithm identifies novel motifs where other methods fail. Finally, we apply this algorithm to detect motifs from expression datasets in C. elegans using a dynamic expression similarity metric rather than fixed expression clusters, and find novel predictive motifs.

  11. Kinetics of transcription initiation directed by multiple cis-regulatory elements on the glnAp2 promoter.

    Science.gov (United States)

    Wang, Yaolai; Liu, Feng; Wang, Wei

    2016-12-15

    Transcription initiation is orchestrated by dynamic molecular interactions, with kinetic steps difficult to detect. Utilizing a hybrid method, we aim to unravel essential kinetic steps of transcriptional regulation on the glnAp2 promoter, whose regulatory region includes two enhancers (sites I and II) and three low-affinity sequences (sites III-V), to which the transcriptional activator NtrC binds. By structure reconstruction, we analyze all possible organization architectures of the transcription apparatus (TA). The main regulatory mode involves two NtrC hexamers: one at enhancer II transiently associates with site V such that the other at enhancer I can rapidly approach and catalyze the σ(54)-RNA polymerase holoenzyme. We build a kinetic model characterizing essential steps of the TA operation; with the known kinetics of the holoenzyme interacting with DNA, this model enables the kinetics beyond technical detection to be determined by fitting the input-output function of the wild-type promoter. The model further quantitatively reproduces transcriptional activities of various mutated promoters. These results reveal different roles played by two enhancers and interpret why the low-affinity elements conditionally enhance or repress transcription. This work presents an integrated dynamic picture of regulated transcription initiation and suggests an evolutionarily conserved characteristic guaranteeing reliable transcriptional response to regulatory signals. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

  12. Identification of Predictive Cis-Regulatory Elements Using a Discriminative Objective Function and a Dynamic Search Space.

    Directory of Open Access Journals (Sweden)

    Rahul Karnik

    Full Text Available The generation of genomic binding or accessibility data from massively parallel sequencing technologies such as ChIP-seq and DNase-seq continues to accelerate. Yet state-of-the-art computational approaches for the identification of DNA binding motifs often yield motifs of weak predictive power. Here we present a novel computational algorithm called MotifSpec, designed to find predictive motifs, in contrast to over-represented sequence elements. The key distinguishing feature of this algorithm is that it uses a dynamic search space and a learned threshold to find discriminative motifs in combination with the modeling of motifs using a full PWM (position weight matrix rather than k-mer words or regular expressions. We demonstrate that our approach finds motifs corresponding to known binding specificities in several mammalian ChIP-seq datasets, and that our PWMs classify the ChIP-seq signals with accuracy comparable to, or marginally better than motifs from the best existing algorithms. In other datasets, our algorithm identifies novel motifs where other methods fail. Finally, we apply this algorithm to detect motifs from expression datasets in C. elegans using a dynamic expression similarity metric rather than fixed expression clusters, and find novel predictive motifs.

  13. Cis-regulatory mutations in human disease.

    Science.gov (United States)

    Epstein, Douglas J

    2009-07-01

    Cis-acting regulatory sequences are required for the proper temporal and spatial control of gene expression. Variation in gene expression is highly heritable and a significant determinant of human disease susceptibility. The diversity of human genetic diseases attributed, in whole or in part, to mutations in non-coding regulatory sequences is on the rise. Improvements in genome-wide methods of associating genetic variation with human disease and predicting DNA with cis-regulatory potential are two of the major reasons for these recent advances. This review will highlight select examples from the literature that have successfully integrated genetic and genomic approaches to uncover the molecular basis by which cis-regulatory mutations alter gene expression and contribute to human disease. The fine mapping of disease-causing variants has led to the discovery of novel cis-acting regulatory elements that, in some instances, are located as far away as 1.5 Mb from the target gene. In other cases, the prior knowledge of the regulatory landscape surrounding the gene of interest aided in the selection of enhancers for mutation screening. The success of these studies should provide a framework for following up on the large number of genome-wide association studies that have identified common variants in non-coding regions of the genome that associate with increased risk of human diseases including, diabetes, autism, Crohn's, colorectal cancer, and asthma, to name a few.

  14. The role of cis regulatory evolution in maize domestication.

    Science.gov (United States)

    Lemmon, Zachary H; Bukowski, Robert; Sun, Qi; Doebley, John F

    2014-11-01

    Gene expression differences between divergent lineages caused by modification of cis regulatory elements are thought to be important in evolution. We assayed genome-wide cis and trans regulatory differences between maize and its wild progenitor, teosinte, using deep RNA sequencing in F1 hybrid and parent inbred lines for three tissue types (ear, leaf and stem). Pervasive regulatory variation was observed with approximately 70% of ∼17,000 genes showing evidence of regulatory divergence between maize and teosinte. However, many fewer genes (1,079 genes) show consistent cis differences with all sampled maize and teosinte lines. For ∼70% of these 1,079 genes, the cis differences are specific to a single tissue. The number of genes with cis regulatory differences is greatest for ear tissue, which underwent a drastic transformation in form during domestication. As expected from the domestication bottleneck, maize possesses less cis regulatory variation than teosinte with this deficit greatest for genes showing maize-teosinte cis regulatory divergence, suggesting selection on cis regulatory differences during domestication. Consistent with selection on cis regulatory elements, genes with cis effects correlated strongly with genes under positive selection during maize domestication and improvement, while genes with trans regulatory effects did not. We observed a directional bias such that genes with cis differences showed higher expression of the maize allele more often than the teosinte allele, suggesting domestication favored up-regulation of gene expression. Finally, this work documents the cis and trans regulatory changes between maize and teosinte in over 17,000 genes for three tissues.

  15. The Role of cis Regulatory Evolution in Maize Domestication

    Science.gov (United States)

    Lemmon, Zachary H.; Bukowski, Robert; Sun, Qi; Doebley, John F.

    2014-01-01

    Gene expression differences between divergent lineages caused by modification of cis regulatory elements are thought to be important in evolution. We assayed genome-wide cis and trans regulatory differences between maize and its wild progenitor, teosinte, using deep RNA sequencing in F1 hybrid and parent inbred lines for three tissue types (ear, leaf and stem). Pervasive regulatory variation was observed with approximately 70% of ∼17,000 genes showing evidence of regulatory divergence between maize and teosinte. However, many fewer genes (1,079 genes) show consistent cis differences with all sampled maize and teosinte lines. For ∼70% of these 1,079 genes, the cis differences are specific to a single tissue. The number of genes with cis regulatory differences is greatest for ear tissue, which underwent a drastic transformation in form during domestication. As expected from the domestication bottleneck, maize possesses less cis regulatory variation than teosinte with this deficit greatest for genes showing maize-teosinte cis regulatory divergence, suggesting selection on cis regulatory differences during domestication. Consistent with selection on cis regulatory elements, genes with cis effects correlated strongly with genes under positive selection during maize domestication and improvement, while genes with trans regulatory effects did not. We observed a directional bias such that genes with cis differences showed higher expression of the maize allele more often than the teosinte allele, suggesting domestication favored up-regulation of gene expression. Finally, this work documents the cis and trans regulatory changes between maize and teosinte in over 17,000 genes for three tissues. PMID:25375861

  16. The role of cis regulatory evolution in maize domestication.

    Directory of Open Access Journals (Sweden)

    Zachary H Lemmon

    2014-11-01

    Full Text Available Gene expression differences between divergent lineages caused by modification of cis regulatory elements are thought to be important in evolution. We assayed genome-wide cis and trans regulatory differences between maize and its wild progenitor, teosinte, using deep RNA sequencing in F1 hybrid and parent inbred lines for three tissue types (ear, leaf and stem. Pervasive regulatory variation was observed with approximately 70% of ∼17,000 genes showing evidence of regulatory divergence between maize and teosinte. However, many fewer genes (1,079 genes show consistent cis differences with all sampled maize and teosinte lines. For ∼70% of these 1,079 genes, the cis differences are specific to a single tissue. The number of genes with cis regulatory differences is greatest for ear tissue, which underwent a drastic transformation in form during domestication. As expected from the domestication bottleneck, maize possesses less cis regulatory variation than teosinte with this deficit greatest for genes showing maize-teosinte cis regulatory divergence, suggesting selection on cis regulatory differences during domestication. Consistent with selection on cis regulatory elements, genes with cis effects correlated strongly with genes under positive selection during maize domestication and improvement, while genes with trans regulatory effects did not. We observed a directional bias such that genes with cis differences showed higher expression of the maize allele more often than the teosinte allele, suggesting domestication favored up-regulation of gene expression. Finally, this work documents the cis and trans regulatory changes between maize and teosinte in over 17,000 genes for three tissues.

  17. The role of cis regulatory evolution in maize domestication.

    Directory of Open Access Journals (Sweden)

    Zachary H Lemmon

    2014-11-01

    Full Text Available Gene expression differences between divergent lineages caused by modification of cis regulatory elements are thought to be important in evolution. We assayed genome-wide cis and trans regulatory differences between maize and its wild progenitor, teosinte, using deep RNA sequencing in F1 hybrid and parent inbred lines for three tissue types (ear, leaf and stem. Pervasive regulatory variation was observed with approximately 70% of ∼17,000 genes showing evidence of regulatory divergence between maize and teosinte. However, many fewer genes (1,079 genes show consistent cis differences with all sampled maize and teosinte lines. For ∼70% of these 1,079 genes, the cis differences are specific to a single tissue. The number of genes with cis regulatory differences is greatest for ear tissue, which underwent a drastic transformation in form during domestication. As expected from the domestication bottleneck, maize possesses less cis regulatory variation than teosinte with this deficit greatest for genes showing maize-teosinte cis regulatory divergence, suggesting selection on cis regulatory differences during domestication. Consistent with selection on cis regulatory elements, genes with cis effects correlated strongly with genes under positive selection during maize domestication and improvement, while genes with trans regulatory effects did not. We observed a directional bias such that genes with cis differences showed higher expression of the maize allele more often than the teosinte allele, suggesting domestication favored up-regulation of gene expression. Finally, this work documents the cis and trans regulatory changes between maize and teosinte in over 17,000 genes for three tissues.

  18. Genomic analysis reveals major determinants of cis-regulatory variation in Capsella grandiflora.

    Science.gov (United States)

    Steige, Kim A; Laenen, Benjamin; Reimegård, Johan; Scofield, Douglas G; Slotte, Tanja

    2017-01-31

    Understanding the causes of cis-regulatory variation is a long-standing aim in evolutionary biology. Although cis-regulatory variation has long been considered important for adaptation, we still have a limited understanding of the selective importance and genomic determinants of standing cis-regulatory variation. To address these questions, we studied the prevalence, genomic determinants, and selective forces shaping cis-regulatory variation in the outcrossing plant Capsella grandiflora We first identified a set of 1,010 genes with common cis-regulatory variation using analyses of allele-specific expression (ASE). Population genomic analyses of whole-genome sequences from 32 individuals showed that genes with common cis-regulatory variation (i) are under weaker purifying selection and (ii) undergo less frequent positive selection than other genes. We further identified genomic determinants of cis-regulatory variation. Gene body methylation (gbM) was a major factor constraining cis-regulatory variation, whereas presence of nearby transposable elements (TEs) and tissue specificity of expression increased the odds of ASE. Our results suggest that most common cis-regulatory variation in C. grandiflora is under weak purifying selection, and that gene-specific functional constraints are more important for the maintenance of cis-regulatory variation than genome-scale variation in the intensity of selection. Our results agree with previous findings that suggest TE silencing affects nearby gene expression, and provide evidence for a link between gbM and cis-regulatory constraint, possibly reflecting greater dosage sensitivity of body-methylated genes. Given the extensive conservation of gbM in flowering plants, this suggests that gbM could be an important predictor of cis-regulatory variation in a wide range of plant species.

  19. Combining computational prediction of cis-regulatory elements with a new enhancer assay to efficiently label neuronal structures in the medaka fish.

    Directory of Open Access Journals (Sweden)

    Emmanuel Mongin

    Full Text Available The developing vertebrate nervous system contains a remarkable array of neural cells organized into complex, evolutionarily conserved structures. The labeling of living cells in these structures is key for the understanding of brain development and function, yet the generation of stable lines expressing reporter genes in specific spatio-temporal patterns remains a limiting step. In this study we present a fast and reliable pipeline to efficiently generate a set of stable lines expressing a reporter gene in multiple neuronal structures in the developing nervous system in medaka. The pipeline combines both the accurate computational genome-wide prediction of neuronal specific cis-regulatory modules (CRMs and a newly developed experimental setup to rapidly obtain transgenic lines in a cost-effective and highly reproducible manner. 95% of the CRMs tested in our experimental setup show enhancer activity in various and numerous neuronal structures belonging to all major brain subdivisions. This pipeline represents a significant step towards the dissection of embryonic neuronal development in vertebrates.

  20. Divergence in cis-regulatory sequences surrounding the opsin gene arrays of African cichlid fishes

    Directory of Open Access Journals (Sweden)

    Streelman J Todd

    2011-05-01

    factor binding site turnover. We also found three SNPs within two opsin promoters and one non-coding element that had weak association with cichlid opsin expression. Conclusions This study is the first to systematically search the opsins of cichlids for putative cis-regulatory sequences. Although many putative regulatory regions are highly conserved across a large number of phenotypically diverse cichlids, we found at least nine divergent sequences that could contribute to opsin expression differences in cis and stand out as candidates for future functional analyses.

  1. Using hexamers to predict cis-regulatory motifs in Drosophila

    Directory of Open Access Journals (Sweden)

    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.

  2. Favorable genomic environments for cis-regulatory evolution: A novel theoretical framework.

    Science.gov (United States)

    Maeso, Ignacio; Tena, Juan J

    2016-09-01

    Cis-regulatory changes are arguably the primary evolutionary source of animal morphological diversity. With the recent explosion of genome-wide comparisons of the cis-regulatory content in different animal species is now possible to infer general principles underlying enhancer evolution. However, these studies have also revealed numerous discrepancies and paradoxes, suggesting that the mechanistic causes and modes of cis-regulatory evolution are still not well understood and are probably much more complex than generally appreciated. Here, we argue that the mutational mechanisms and genomic regions generating new regulatory activities must comply with the constraints imposed by the molecular properties of cis-regulatory elements (CREs) and the organizational features of long-range chromatin interactions. Accordingly, we propose a new integrative evolutionary framework for cis-regulatory evolution based on two major premises for the origin of novel enhancer activity: (i) an accessible chromatin environment and (ii) compatibility with the 3D structure and interactions of pre-existing CREs. Mechanisms and DNA sequences not fulfilling these premises, will be less likely to have a measurable impact on gene expression and as such, will have a minor contribution to the evolution of gene regulation. Finally, we discuss current comparative cis-regulatory data under the light of this new evolutionary model, and propose that the two most prominent mechanisms for the evolution of cis-regulatory changes are the overprinting of ancestral CREs and the exaptation of transposable elements.

  3. Divergence in cis-regulatory sequences surrounding the opsin gene arrays of African cichlid fishes

    National Research Council Canada - National Science Library

    O'Quin, Kelly E; Smith, Daniel; Naseer, Zan; Schulte, Jane; Engel, Samuel D; Loh, Yong-Hwee E; Streelman, J Todd; Boore, Jeffrey L; Carleton, Karen L

    2011-01-01

    .... We use phylogenetic footprinting and shadowing to examine divergence in conserved non-coding elements, promoter sequences, and 3'-UTRs surrounding each opsin in search of candidate cis-regulatory...

  4. Statistical significance of cis-regulatory modules

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    Smith Andrew D

    2007-01-01

    Full Text Available Abstract Background It is becoming increasingly important for researchers to be able to scan through large genomic regions for transcription factor binding sites or clusters of binding sites forming cis-regulatory modules. Correspondingly, there has been a push to develop algorithms for the rapid detection and assessment of cis-regulatory modules. While various algorithms for this purpose have been introduced, most are not well suited for rapid, genome scale scanning. Results We introduce methods designed for the detection and statistical evaluation of cis-regulatory modules, modeled as either clusters of individual binding sites or as combinations of sites with constrained organization. In order to determine the statistical significance of module sites, we first need a method to determine the statistical significance of single transcription factor binding site matches. We introduce a straightforward method of estimating the statistical significance of single site matches using a database of known promoters to produce data structures that can be used to estimate p-values for binding site matches. We next introduce a technique to calculate the statistical significance of the arrangement of binding sites within a module using a max-gap model. If the module scanned for has defined organizational parameters, the probability of the module is corrected to account for organizational constraints. The statistical significance of single site matches and the architecture of sites within the module can be combined to provide an overall estimation of statistical significance of cis-regulatory module sites. Conclusion The methods introduced in this paper allow for the detection and statistical evaluation of single transcription factor binding sites and cis-regulatory modules. The features described are implemented in the Search Tool for Occurrences of Regulatory Motifs (STORM and MODSTORM software.

  5. 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

    have been reported in ZRS, the underlying regulatory mechanism is still unknown. In this study, we analyzed the effect on transcription factor binding of a novel ZRS point mutation (463T>G) in a Pakistani family with preaxial polydactyly and triphalangeal thumb. Electrophoretical mobility shift assay......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...

  6. A Cis-Regulatory Map of the Drosophila Genome

    Science.gov (United States)

    Nègre, Nicolas; Brown, Christopher D.; Ma, Lijia; Bristow, Christopher Aaron; Miller, Steven W.; Wagner, Ulrich; Kheradpour, Pouya; Eaton, Matthew L.; Loriaux, Paul; Sealfon, Rachel; Li, Zirong; Ishii, Haruhiko; Spokony, Rebecca F.; Chen, Jia; Hwang, Lindsay; Cheng, Chao; Auburn, Richard P.; Davis, Melissa B.; Domanus, Marc; Shah, Parantu K.; Morrison, Carolyn A.; Zieba, Jennifer; Suchy, Sarah; Senderowicz, Lionel; Victorsen, Alec; Bild, Nicholas A.; Grundstad, A. Jason; Hanley, David; MacAlpine, David M.; Mannervik, Mattias; Venken, Koen; Bellen, Hugo; White, Robert; Russell, Steven; Grossman, Robert L.; Ren, Bing; Gerstein, Mark; Posakony, James W.; Kellis, Manolis; White, Kevin P.

    2011-01-01

    Systematic annotation of gene regulatory elements is a major challenge in genome science. Direct mapping of chromatin modification marks and transcriptional factor binding sites genome-wide 1,2 has successfully identified specific subtypes of regulatory elements 3. In Drosophila several pioneering studies have provided genome-wide identification of Polycomb-Response Elements 4, chromatin states 5, transcription factor binding sites (TFBS) 6–9, PolII regulation 8, and insulator elements 10; however, comprehensive annotation of the regulatory genome remains a significant challenge. Here we describe results from the modENCODE cis-regulatory annotation project. We produced a map of the Drosophila melanogaster regulatory genome based on more than 300 chromatin immuno-precipitation (ChIP) datasets for eight chromatin features, five histone deacetylases (HDACs) and thirty-eight site-specific transcription factors (TFs) at different stages of development. Using these data we inferred more than 20,000 candidate regulatory elements and we validated a subset of predictions for promoters, enhancers, and insulators in vivo. We also identified nearly 2,000 genomic regions of dense TF binding associated with chromatin activity and accessibility. We discovered hundreds of new TF co-binding relationships and defined a TF network with over 800 potential regulatory relationships. PMID:21430782

  7. Abundant raw material for cis-regulatory evolution in humans

    Science.gov (United States)

    Rockman, Matthew V.; Wray, Gregory A.

    2002-01-01

    Changes in gene expression and regulation--due in particular to the evolution of cis-regulatory DNA sequences--may underlie many evolutionary changes in phenotypes, yet little is known about the distribution of such variation in populations. We present in this study the first survey of experimentally validated functional cis-regulatory polymorphism. These data are derived from more than 140 polymorphisms involved in the regulation of 107 genes in Homo sapiens, the eukaryote species with the most available data. We find that functional cis-regulatory variation is widespread in the human genome and that the consequent variation in gene expression is twofold or greater for 63% of the genes surveyed. Transcription factor-DNA interactions are highly polymorphic, and regulatory interactions have been gained and lost within human populations. On average, humans are heterozygous at more functional cis-regulatory sites (>16,000) than at amino acid positions (human phenotypic variation.

  8. Abundant raw material for cis-regulatory evolution in humans

    Science.gov (United States)

    Rockman, Matthew V.; Wray, Gregory A.

    2002-01-01

    Changes in gene expression and regulation--due in particular to the evolution of cis-regulatory DNA sequences--may underlie many evolutionary changes in phenotypes, yet little is known about the distribution of such variation in populations. We present in this study the first survey of experimentally validated functional cis-regulatory polymorphism. These data are derived from more than 140 polymorphisms involved in the regulation of 107 genes in Homo sapiens, the eukaryote species with the most available data. We find that functional cis-regulatory variation is widespread in the human genome and that the consequent variation in gene expression is twofold or greater for 63% of the genes surveyed. Transcription factor-DNA interactions are highly polymorphic, and regulatory interactions have been gained and lost within human populations. On average, humans are heterozygous at more functional cis-regulatory sites (>16,000) than at amino acid positions (human phenotypic variation.

  9. Conserved cis-regulatory modules in promoters of genes encoding wheat high-molecular-weight glutenin subunits.

    Science.gov (United States)

    Ravel, Catherine; Fiquet, Samuel; Boudet, Julie; Dardevet, Mireille; Vincent, Jonathan; Merlino, Marielle; Michard, Robin; Martre, Pierre

    2014-01-01

    The concentration and composition of the gliadin and glutenin seed storage proteins (SSPs) in wheat flour are the most important determinants of its end-use value. In cereals, the synthesis of SSPs is predominantly regulated at the transcriptional level by a complex network involving at least five cis-elements in gene promoters. The high-molecular-weight glutenin subunits (HMW-GS) are encoded by two tightly linked genes located on the long arms of group 1 chromosomes. Here, we sequenced and annotated the HMW-GS gene promoters of 22 electrophoretic wheat alleles to identify putative cis-regulatory motifs. We focused on 24 motifs known to be involved in SSP gene regulation. Most of them were identified in at least one HMW-GS gene promoter sequence. A common regulatory framework was observed in all the HMW-GS gene promoters, as they shared conserved cis-regulatory modules (CCRMs) including all the five motifs known to regulate the transcription of SSP genes. This common regulatory framework comprises a composite box made of the GATA motifs and GCN4-like Motifs (GLMs) and was shown to be functional as the GLMs are able to bind a bZIP transcriptional factor SPA (Storage Protein Activator). In addition to this regulatory framework, each HMW-GS gene promoter had additional motifs organized differently. The promoters of most highly expressed x-type HMW-GS genes contain an additional box predicted to bind R2R3-MYB transcriptional factors. However, the differences in annotation between promoter alleles could not be related to their level of expression. In summary, we identified a common modular organization of HMW-GS gene promoters but the lack of correlation between the cis-motifs of each HMW-GS gene promoter and their level of expression suggests that other cis-elements or other mechanisms regulate HMW-GS gene expression.

  10. Conservation of shh cis-regulatory architecture of the coelacanth is consistent with its ancestral phylogenetic position

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    Lang Michael

    2010-11-01

    Full Text Available Abstract Background The modern coelacanth (Latimeria is the extant taxon of a basal sarcopterygian lineage and sister group to tetrapods. Apart from certain apomorphic traits, its morphology is characterized by a high degree of retention of ancestral vertebrate structures and little morphological change. An insight into the molecular evolution that may explain the unchanged character of Latimeria morphology requires the analysis of the expression patterns of developmental regulator genes and their cis-regulatory modules (CRMs. Results We describe the comparative and functional analysis of the sonic hedgehog (shh genomic region of Latimeria menadoensis. Several putative enhancers in the Latimeria shh locus have been identified by comparisons to sarcopterygian and actinopterygian extant species. Specific sequence conservation with all known actinopterygian enhancer elements has been detected. However, these elements are selectively missing in more recently diverged actinopterygian and sarcopterygian species. The functionality of the putative Latimeria enhancers was confirmed by reporter gene expression analysis in transient transgenic zebrafish and chick embryos. Conclusions Latimeria shh CRMs represent the ancestral set of enhancers that have emerged before the split of lobe-finned and ray-finned fishes. In contrast to lineage-specific losses and differentiations in more derived lineages, Latimeria shh enhancers reveal low levels of sequence diversification. High overall sequence conservation of shh conserved noncoding elements (CNE is consistent with the general trend of high levels of conservation of noncoding DNA in the slowly evolving Latimeria genome.

  11. Cis-regulatory Mutations in the Caenorhabditis elegans Homeobox Gene Locus cog-1 Affect Neuronal Development

    Science.gov (United States)

    O'Meara, M. Maggie; Bigelow, Henry; Flibotte, Stephane; Etchberger, John F.; Moerman, Donald G.; Hobert, Oliver

    2009-01-01

    We apply here comparative genome hybridization as a novel tool to identify the molecular lesion in two Caenorhabditis elegans mutant strains that affect a neuronal cell fate decision. The phenotype of the mutant strains resembles those of the loss-of-function alleles of the cog-1 homeobox gene, an inducer of the fate of the gustatory neuron ASER. We find that both lesions map to the cis-regulatory control region of cog-1 and affect a phylogenetically conserved binding site for the C2H2 zinc-finger transcription factor CHE-1, a previously known regulator of cog-1 expression in ASER. Identification of this CHE-1-binding site as a critical regulator of cog-1 expression in the ASER in vivo represents one of the rare demonstrations of the in vivo relevance of an experimentally determined or predicted transcription-factor-binding site. Aside from the mutationally defined CHE-1-binding site, cog-1 contains a second, functional CHE-1-binding site, which in isolation is sufficient to drive reporter gene expression in the ASER but in an in vivo context is apparently insufficient for promoting appropriate ASER expression. The cis-regulatory control regions of other ASE-expressed genes also contain ASE motifs that can promote ASE neuron expression when isolated from their genomic context, but appear to depend on multiple ASE motifs in their normal genomic context. The multiplicity of cis-regulatory elements may ensure the robustness of gene expression. PMID:19189954

  12. Are Interactions between cis-Regulatory Variants Evidence for Biological Epistasis or Statistical Artifacts?

    Science.gov (United States)

    Fish, Alexandra E; Capra, John A; Bush, William S

    2016-10-06

    The importance of epistasis-or statistical interactions between genetic variants-to the development of complex disease in humans has been controversial. Genome-wide association studies of statistical interactions influencing human traits have recently become computationally feasible and have identified many putative interactions. However, statistical models used to detect interactions can be confounded, which makes it difficult to be certain that observed statistical interactions are evidence for true molecular epistasis. In this study, we investigate whether there is evidence for epistatic interactions between genetic variants within the cis-regulatory region that influence gene expression after accounting for technical, statistical, and biological confounding factors. We identified 1,119 (FDR = 5%) interactions that appear to regulate gene expression in human lymphoblastoid cell lines, a tightly controlled, largely genetically determined phenotype. Many of these interactions replicated in an independent dataset (90 of 803 tested, Bonferroni threshold). We then performed an exhaustive analysis of both known and novel confounders, including ceiling/floor effects, missing genotype combinations, haplotype effects, single variants tagged through linkage disequilibrium, and population stratification. Every interaction could be explained by at least one of these confounders, and replication in independent datasets did not protect against some confounders. Assuming that the confounding factors provide a more parsimonious explanation for each interaction, we find it unlikely that cis-regulatory interactions contribute strongly to human gene expression, which calls into question the relevance of cis-regulatory interactions for other human phenotypes. We additionally propose several best practices for epistasis testing to protect future studies from confounding. Copyright © 2016 American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

  13. ChIP-Seq-Annotated Heliconius erato Genome Highlights Patterns of cis-Regulatory Evolution in Lepidoptera.

    Science.gov (United States)

    Lewis, James J; van der Burg, Karin R L; Mazo-Vargas, Anyi; Reed, Robert D

    2016-09-13

    Uncovering phylogenetic patterns of cis-regulatory evolution remains a fundamental goal for evolutionary and developmental biology. Here, we characterize the evolution of regulatory loci in butterflies and moths using chromatin immunoprecipitation sequencing (ChIP-seq) annotation of regulatory elements across three stages of head development. In the process we provide a high-quality, functionally annotated genome assembly for the butterfly, Heliconius erato. Comparing cis-regulatory element conservation across six lepidopteran genomes, we find that regulatory sequences evolve at a pace similar to that of protein-coding regions. We also observe that elements active at multiple developmental stages are markedly more conserved than elements with stage-specific activity. Surprisingly, we also find that stage-specific proximal and distal regulatory elements evolve at nearly identical rates. Our study provides a benchmark for genome-wide patterns of regulatory element evolution in insects, and it shows that developmental timing of activity strongly predicts patterns of regulatory sequence evolution.

  14. cis-Regulatory control of the initial neurogenic pattern of onecut gene expression in the sea urchin embryo.

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    Barsi, Julius C; Davidson, Eric H

    2016-01-01

    Specification of the ciliated band (CB) of echinoid embryos executes three spatial functions essential for postgastrular organization. These are establishment of a band about 5 cells wide which delimits and bounds other embryonic territories; definition of a neurogenic domain within this band; and generation within it of arrays of ciliary cells that bear the special long cilia from which the structure derives its name. In Strongylocentrotus purpuratus the spatial coordinates of the future ciliated band are initially and exactly determined by the disposition of a ring of cells that transcriptionally activate the onecut homeodomain regulatory gene, beginning in blastula stage, long before the appearance of the CB per se. Thus the cis-regulatory apparatus that governs onecut expression in the blastula directly reveals the genomic sequence code by which these aspects of the spatial organization of the embryo are initially determined. We screened the entire onecut locus and its flanking region for transcriptionally active cis-regulatory elements, and by means of BAC recombineered deletions identified three separated and required cis-regulatory modules that execute different functions. The operating logic of the crucial spatial control module accounting for the spectacularly precise and beautiful early onecut expression domain depends on spatial repression. Previously predicted oral ectoderm and aboral ectoderm repressors were identified by cis-regulatory mutation as the products of goosecoid and irxa genes respectively, while the pan-ectodermal activator SoxB1 supplies a transcriptional driver function.

  15. Conserved stem-loop structures in the HIV-1 RNA region containing the A3 3' splice site and its cis-regulatory element: possible involvement in RNA splicing.

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    Jacquenet, S; Ropers, D; Bilodeau, P S; Damier, L; Mougin, A; Stoltzfus, C M; Branlant, C

    2001-01-15

    The HIV-1 transcript is alternatively spliced to over 30 different mRNAs. Whether RNA secondary structure can influence HIV-1 RNA alternative splicing has not previously been examined. Here we have determined the secondary structure of the HIV-1/BRU RNA segment, containing the alternative A3, A4a, A4b, A4c and A5 3' splice sites. Site A3, required for tat mRNA production, is contained in the terminal loop of a stem-loop structure (SLS2), which is highly conserved in HIV-1 and related SIVcpz strains. The exon splicing silencer (ESS2) acting on site A3 is located in a long irregular stem-loop structure (SLS3). Two SLS3 domains were protected by nuclear components under splicing condition assays. One contains the A4c branch points and a putative SR protein binding site. The other one is adjacent to ESS2. Unexpectedly, only the 3' A residue of ESS2 was protected. The suboptimal A3 polypyrimidine tract (PPT) is base paired. Using site-directed mutagenesis and transfection of a mini-HIV-1 cDNA into HeLa cells, we found that, in a wild-type PPT context, a mutation of the A3 downstream sequence that reinforced SLS2 stability decreased site A3 utilization. This was not the case with an optimized PPT. Hence, sequence and secondary structure of the PPT may cooperate in limiting site A3 utilization.

  16. The molecular signature and cis-regulatory architecture of a C. elegans gustatory neuron

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    Etchberger, John F.; Lorch, Adam; Sleumer, Monica C.; Zapf, Richard; Jones, Steven J.; Marra, Marco A.; Holt, Robert A.; Moerman, Donald G.; Hobert, Oliver

    2007-01-01

    Taste receptor cells constitute a highly specialized cell type that perceives and conveys specific sensory information to the brain. The detailed molecular composition of these cells and the mechanisms that program their fate are, in general, poorly understood. We have generated serial analysis of gene expression (SAGE) libraries from two distinct populations of single, isolated sensory neuron classes, the gustatory neuron class ASE and the thermosensory neuron class AFD, from the nematode Caenorhabditis elegans. By comparing these two libraries, we have identified >1000 genes that define the ASE gustatory neuron class on a molecular level. This set of genes contains determinants of the differentiated state of the ASE neuron, such as a surprisingly complex repertoire of transcription factors (TFs), ion channels, neurotransmitters, and receptors, as well as seven-transmembrane receptor (7TMR)-type putative gustatory receptor genes. Through the in vivo dissection of the cis-regulatory regions of several ASE-expressed genes, we identified a small cis-regulatory motif, the “ASE motif,” that is required for the expression of many ASE-expressed genes. We demonstrate that the ASE motif is a binding site for the C2H2 zinc finger TF CHE-1, which is essential for the correct differentiation of the ASE gustatory neuron. Taken together, our results provide a unique view of the molecular landscape of a single neuron type and reveal an important aspect of the regulatory logic for gustatory neuron specification in C. elegans. PMID:17606643

  17. 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.

  18. Nucleotide sequence conservation of novel and established cis-regulatory sites within the tyrosine hydroxylase gene promoter.

    Science.gov (United States)

    Wang, Meng; Banerjee, Kasturi; Baker, Harriet; Cave, John W

    2015-02-01

    Tyrosine hydroxylase (TH) is the rate-limiting enzyme in catecholamine biosynthesis and its gene proximal promoter ( electromobility shift assays showed that brain-region specific complexes assemble on these motifs. These studies also identified a non-canonical CRE binding (CREB) protein recognition element in the proximal promoter. Together, these studies provide a detailed analysis of evolutionary conservation within the TH promoter and identify potential cis-regulatory motifs that underlie a core set of regulatory mechanisms in mammals.

  19. cis-Regulatory Circuits Regulating NEK6 Kinase Overexpression in Transformed B Cells Are Super-Enhancer Independent

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    Yue Huang

    2017-03-01

    Full Text Available Alterations in distal regulatory elements that control gene expression underlie many diseases, including cancer. Epigenomic analyses of normal and diseased cells have produced correlative predictions for connections between dysregulated enhancers and target genes involved in pathogenesis. However, with few exceptions, these predicted cis-regulatory circuits remain untested. Here, we dissect cis-regulatory circuits that lead to overexpression of NEK6, a mitosis-associated kinase, in human B cell lymphoma. We find that only a minor subset of predicted enhancers is required for NEK6 expression. Indeed, an annotated super-enhancer is dispensable for NEK6 overexpression and for maintaining the architecture of a B cell-specific regulatory hub. A CTCF cluster serves as a chromatin and architectural boundary to block communication of the NEK6 regulatory hub with neighboring genes. Our findings emphasize that validation of predicted cis-regulatory circuits and super-enhancers is needed to prioritize transcriptional control elements as therapeutic targets.

  20. cis-Regulatory Circuits Regulating NEK6 Kinase Overexpression in Transformed B Cells Are Super-Enhancer Independent.

    Science.gov (United States)

    Huang, Yue; Koues, Olivia I; Zhao, Jiang-Yang; Liu, Regina; Pyfrom, Sarah C; Payton, Jacqueline E; Oltz, Eugene M

    2017-03-21

    Alterations in distal regulatory elements that control gene expression underlie many diseases, including cancer. Epigenomic analyses of normal and diseased cells have produced correlative predictions for connections between dysregulated enhancers and target genes involved in pathogenesis. However, with few exceptions, these predicted cis-regulatory circuits remain untested. Here, we dissect cis-regulatory circuits that lead to overexpression of NEK6, a mitosis-associated kinase, in human B cell lymphoma. We find that only a minor subset of predicted enhancers is required for NEK6 expression. Indeed, an annotated super-enhancer is dispensable for NEK6 overexpression and for maintaining the architecture of a B cell-specific regulatory hub. A CTCF cluster serves as a chromatin and architectural boundary to block communication of the NEK6 regulatory hub with neighboring genes. Our findings emphasize that validation of predicted cis-regulatory circuits and super-enhancers is needed to prioritize transcriptional control elements as therapeutic targets.

  1. A primer on regression methods for decoding cis-regulatory logic

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    Das, Debopriya; Pellegrini, Matteo; Gray, Joe W.

    2009-03-03

    The rapidly emerging field of systems biology is helping us to understand the molecular determinants of phenotype on a genomic scale [1]. Cis-regulatory elements are major sequence-based determinants of biological processes in cells and tissues [2]. For instance, during transcriptional regulation, transcription factors (TFs) bind to very specific regions on the promoter DNA [2,3] and recruit the basal transcriptional machinery, which ultimately initiates mRNA transcription (Figure 1A). Learning cis-Regulatory Elements from Omics Data A vast amount of work over the past decade has shown that omics data can be used to learn cis-regulatory logic on a genome-wide scale [4-6]--in particular, by integrating sequence data with mRNA expression profiles. The most popular approach has been to identify over-represented motifs in promoters of genes that are coexpressed [4,7,8]. Though widely used, such an approach can be limiting for a variety of reasons. First, the combinatorial nature of gene regulation is difficult to explicitly model in this framework. Moreover, in many applications of this approach, expression data from multiple conditions are necessary to obtain reliable predictions. This can potentially limit the use of this method to only large data sets [9]. Although these methods can be adapted to analyze mRNA expression data from a pair of biological conditions, such comparisons are often confounded by the fact that primary and secondary response genes are clustered together--whereas only the primary response genes are expected to contain the functional motifs [10]. A set of approaches based on regression has been developed to overcome the above limitations [11-32]. These approaches have their foundations in certain biophysical aspects of gene regulation [26,33-35]. That is, the models are motivated by the expected transcriptional response of genes due to the binding of TFs to their promoters. While such methods have gathered popularity in the computational domain

  2. Evolution of New cis-Regulatory Motifs Required for Cell-Specific Gene Expression in Caenorhabditis.

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    Michalis Barkoulas

    2016-09-01

    Full Text Available Patterning of C. elegans vulval cell fates relies on inductive signaling. In this induction event, a single cell, the gonadal anchor cell, secretes LIN-3/EGF and induces three out of six competent precursor cells to acquire a vulval fate. We previously showed that this developmental system is robust to a four-fold variation in lin-3/EGF genetic dose. Here using single-molecule FISH, we find that the mean level of expression of lin-3 in the anchor cell is remarkably conserved. No change in lin-3 expression level could be detected among C. elegans wild isolates and only a low level of change-less than 30%-in the Caenorhabditis genus and in Oscheius tipulae. In C. elegans, lin-3 expression in the anchor cell is known to require three transcription factor binding sites, specifically two E-boxes and a nuclear-hormone-receptor (NHR binding site. Mutation of any of these three elements in C. elegans results in a dramatic decrease in lin-3 expression. Yet only a single E-box is found in the Drosophilae supergroup of Caenorhabditis species, including C. angaria, while the NHR-binding site likely only evolved at the base of the Elegans group. We find that a transgene from C. angaria bearing a single E-box is sufficient for normal expression in C. elegans. Even a short 58 bp cis-regulatory fragment from C. angaria with this single E-box is able to replace the three transcription factor binding sites at the endogenous C. elegans lin-3 locus, resulting in the wild-type expression level. Thus, regulatory evolution occurring in cis within a 58 bp lin-3 fragment, results in a strict requirement for the NHR binding site and a second E-box in C. elegans. This single-cell, single-molecule, quantitative and functional evo-devo study demonstrates that conserved expression levels can hide extensive change in cis-regulatory site requirements and highlights the evolution of new cis-regulatory elements required for cell-specific gene expression.

  3. SMCis: An Effective Algorithm for Discovery of Cis-Regulatory Modules

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    Guo, Haitao; Huo, Hongwei; Yu, Qiang

    2016-01-01

    The discovery of cis-regulatory modules (CRMs) is a challenging problem in computational biology. Limited by the difficulty of using an HMM to model dependent features in transcriptional regulatory sequences (TRSs), the probabilistic modeling methods based on HMMs cannot accurately represent the distance between regulatory elements in TRSs and are cumbersome to model the prevailing dependencies between motifs within CRMs. We propose a probabilistic modeling algorithm called SMCis, which builds a more powerful CRM discovery model based on a hidden semi-Markov model. Our model characterizes the regulatory structure of CRMs and effectively models dependencies between motifs at a higher level of abstraction based on segments rather than nucleotides. Experimental results on three benchmark datasets indicate that our method performs better than the compared algorithms. PMID:27637070

  4. Identifying cis-regulatory sequences by word profile similarity.

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    Garmay Leung

    Full Text Available BACKGROUND: Recognizing regulatory sequences in genomes is a continuing challenge, despite a wealth of available genomic data and a growing number of experimentally validated examples. METHODOLOGY/PRINCIPAL FINDINGS: We discuss here a simple approach to search for regulatory sequences based on the compositional similarity of genomic regions and known cis-regulatory sequences. This method, which is not limited to searching for predefined motifs, recovers sequences known to be under similar regulatory control. The words shared by the recovered sequences often correspond to known binding sites. Furthermore, we show that although local word profile clustering is predictive for the regulatory sequences involved in blastoderm segmentation, local dissimilarity is a more universal feature of known regulatory sequences in Drosophila. CONCLUSIONS/SIGNIFICANCE: Our method leverages sequence motifs within a known regulatory sequence to identify co-regulated sequences without explicitly defining binding sites. We also show that regulatory sequences can be distinguished from surrounding sequences by local sequence dissimilarity, a novel feature in identifying regulatory sequences across a genome. Source code for WPH-finder is available for download at http://rana.lbl.gov/downloads/wph.tar.gz.

  5. Computational methods for the detection of cis-regulatory modules.

    Science.gov (United States)

    Van Loo, Peter; Marynen, Peter

    2009-09-01

    Metazoan transcription regulation occurs through the concerted action of multiple transcription factors that bind co-operatively to cis-regulatory modules (CRMs). The annotation of these key regulators of transcription is lagging far behind the annotation of the transcriptome itself. Here, we give an overview of existing computational methods to detect these CRMs in metazoan genomes. We subdivide these methods into three classes: CRM scanners screen sequences for CRMs based on predefined models that often consist of multiple position weight matrices (PWMs). CRM builders construct models of similar CRMs controlling a set of co-regulated or co-expressed genes. CRM genome screeners screen sequences or complete genomes for CRMs as homotypic or heterotypic clusters of binding sites for any combination of transcription factors. We believe that CRM scanners are currently the most advanced methods, although their applicability is limited. Finally, we argue that CRM builders that make use of PWM libraries will benefit greatly from future advances and will prove to be most instrumental for the annotation of regulatory regions in metazoan genomes.

  6. Massively parallel cis-regulatory analysis in the mammalian central nervous system.

    Science.gov (United States)

    Shen, Susan Q; Myers, Connie A; Hughes, Andrew E O; Byrne, Leah C; Flannery, John G; Corbo, Joseph C

    2016-02-01

    Cis-regulatory elements (CREs, e.g., promoters and enhancers) regulate gene expression, and variants within CREs can modulate disease risk. Next-generation sequencing has enabled the rapid generation of genomic data that predict the locations of CREs, but a bottleneck lies in functionally interpreting these data. To address this issue, massively parallel reporter assays (MPRAs) have emerged, in which barcoded reporter libraries are introduced into cells, and the resulting barcoded transcripts are quantified by next-generation sequencing. Thus far, MPRAs have been largely restricted to assaying short CREs in a limited repertoire of cultured cell types. Here, we present two advances that extend the biological relevance and applicability of MPRAs. First, we adapt exome capture technology to instead capture candidate CREs, thereby tiling across the targeted regions and markedly increasing the length of CREs that can be readily assayed. Second, we package the library into adeno-associated virus (AAV), thereby allowing delivery to target organs in vivo. As a proof of concept, we introduce a capture library of about 46,000 constructs, corresponding to roughly 3500 DNase I hypersensitive (DHS) sites, into the mouse retina by ex vivo plasmid electroporation and into the mouse cerebral cortex by in vivo AAV injection. We demonstrate tissue-specific cis-regulatory activity of DHSs and provide examples of high-resolution truncation mutation analysis for multiplex parsing of CREs. Our approach should enable massively parallel functional analysis of a wide range of CREs in any organ or species that can be infected by AAV, such as nonhuman primates and human stem cell-derived organoids.

  7. The cis-regulatory logic of the mammalian photoreceptor transcriptional network.

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    Timothy H-C Hsiau

    Full Text Available The photoreceptor cells of the retina are subject to a greater number of genetic diseases than any other cell type in the human body. The majority of more than 120 cloned human blindness genes are highly expressed in photoreceptors. In order to establish an integrative framework in which to understand these diseases, we have undertaken an experimental and computational analysis of the network controlled by the mammalian photoreceptor transcription factors, Crx, Nrl, and Nr2e3. Using microarray and in situ hybridization datasets we have produced a model of this network which contains over 600 genes, including numerous retinal disease loci as well as previously uncharacterized photoreceptor transcription factors. To elucidate the connectivity of this network, we devised a computational algorithm to identify the photoreceptor-specific cis-regulatory elements (CREs mediating the interactions between these transcription factors and their target genes. In vivo validation of our computational predictions resulted in the discovery of 19 novel photoreceptor-specific CREs near retinal disease genes. Examination of these CREs permitted the definition of a simple cis-regulatory grammar rule associated with high-level expression. To test the generality of this rule, we used an expanded form of it as a selection filter to evolve photoreceptor CREs from random DNA sequences in silico. When fused to fluorescent reporters, these evolved CREs drove strong, photoreceptor-specific expression in vivo. This study represents the first systematic identification and in vivo validation of CREs in a mammalian neuronal cell type and lays the groundwork for a systems biology of photoreceptor transcriptional regulation.

  8. Brachyury, Foxa2 and the cis-Regulatory Origins of the Notochord.

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    Diana S José-Edwards

    2015-12-01

    Full Text Available A main challenge of modern biology is to understand how specific constellations of genes are activated to differentiate cells and give rise to distinct tissues. This study focuses on elucidating how gene expression is initiated in the notochord, an axial structure that provides support and patterning signals to embryos of humans and all other chordates. Although numerous notochord genes have been identified, the regulatory DNAs that orchestrate development and propel evolution of this structure by eliciting notochord gene expression remain mostly uncharted, and the information on their configuration and recurrence is still quite fragmentary. Here we used the simple chordate Ciona for a systematic analysis of notochord cis-regulatory modules (CRMs, and investigated their composition, architectural constraints, predictive ability and evolutionary conservation. We found that most Ciona notochord CRMs relied upon variable combinations of binding sites for the transcription factors Brachyury and/or Foxa2, which can act either synergistically or independently from one another. Notably, one of these CRMs contains a Brachyury binding site juxtaposed to an (AC microsatellite, an unusual arrangement also found in Brachyury-bound regulatory regions in mouse. In contrast, different subsets of CRMs relied upon binding sites for transcription factors of widely diverse families. Surprisingly, we found that neither intra-genomic nor interspecific conservation of binding sites were reliably predictive hallmarks of notochord CRMs. We propose that rather than obeying a rigid sequence-based cis-regulatory code, most notochord CRMs are rather unique. Yet, this study uncovered essential elements recurrently used by divergent chordates as basic building blocks for notochord CRMs.

  9. Overview Article: Identifying transcriptional cis-regulatory modules in animal genomes

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    Suryamohan, Kushal; Halfon, Marc S.

    2014-01-01

    Gene expression is regulated through the activity of transcription factors and chromatin modifying proteins acting on specific DNA sequences, referred to as cis-regulatory elements. These include promoters, located at the transcription initiation sites of genes, and a variety of distal cis-regulatory modules (CRMs), the most common of which are transcriptional enhancers. Because regulated gene expression is fundamental to cell differentiation and acquisition of new cell fates, identifying, characterizing, and understanding the mechanisms of action of CRMs is critical for understanding development. CRM discovery has historically been challenging, as CRMs can be located far from the genes they regulate, have few readily-identifiable sequence characteristics, and for many years were not amenable to high-throughput discovery methods. However, the recent availability of complete genome sequences and the development of next-generation sequencing methods has led to an explosion of both computational and empirical methods for CRM discovery in model and non-model organisms alike. Experimentally, CRMs can be identified through chromatin immunoprecipitation directed against transcription factors or histone post-translational modifications, identification of nucleosome-depleted “open” chromatin regions, or sequencing-based high-throughput functional screening. Computational methods include comparative genomics, clustering of known or predicted transcription factor binding sites, and supervised machine-learning approaches trained on known CRMs. All of these methods have proven effective for CRM discovery, but each has its own considerations and limitations, and each is subject to a greater or lesser number of false-positive identifications. Experimental confirmation of predictions is essential, although shortcomings in current methods suggest that additional means of validation need to be developed. PMID:25704908

  10. ChIP-Seq-Annotated Heliconius erato Genome Highlights Patterns of cis-Regulatory Evolution in Lepidoptera

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    James J. Lewis

    2016-09-01

    Full Text Available Uncovering phylogenetic patterns of cis-regulatory evolution remains a fundamental goal for evolutionary and developmental biology. Here, we characterize the evolution of regulatory loci in butterflies and moths using chromatin immunoprecipitation sequencing (ChIP-seq annotation of regulatory elements across three stages of head development. In the process we provide a high-quality, functionally annotated genome assembly for the butterfly, Heliconius erato. Comparing cis-regulatory element conservation across six lepidopteran genomes, we find that regulatory sequences evolve at a pace similar to that of protein-coding regions. We also observe that elements active at multiple developmental stages are markedly more conserved than elements with stage-specific activity. Surprisingly, we also find that stage-specific proximal and distal regulatory elements evolve at nearly identical rates. Our study provides a benchmark for genome-wide patterns of regulatory element evolution in insects, and it shows that developmental timing of activity strongly predicts patterns of regulatory sequence evolution.

  11. Evidence for the role of transposons in the recruitment of cis-regulatory motifs during the evolution of C4 photosynthesis.

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    Cao, Chensi; Xu, Jiajia; Zheng, Guangyong; Zhu, Xin-Guang

    2016-03-08

    C4 photosynthesis evolved from C3 photosynthesis and has higher light, water, and nitrogen use efficiencies. Several C4 photosynthesis genes show cell-specific expression patterns, which are required for these high resource-use efficiencies. However, the mechanisms underlying the evolution of cis-regulatory elements that control these cell-specific expression patterns remain elusive. In the present study, we tested the hypothesis that the cis-regulatory motifs related to C4 photosynthesis genes were recruited from non-photosynthetic genes and further examined potential mechanisms facilitating this recruitment. We examined 65 predicted bundle sheath cell-specific motifs, 17 experimentally validated cell-specific cis-regulatory elements, and 1,034 motifs derived from gene regulatory networks. Approximately 7, 5, and 1,000 of these three categories of motifs, respectively, were apparently recruited during the evolution of C4 photosynthesis. In addition, we checked 1) the distance between the acceptors and the donors of potentially recruited motifs in a chromosome, and 2) whether the potentially recruited motifs reside within the overlapping region of transposable elements and the promoter of donor genes. The results showed that 7, 4, and 658 of the potentially recruited motifs might have moved via the transposable elements. Furthermore, the potentially recruited motifs showed higher binding affinity to transcription factors compared to randomly generated sequences of the same length as the motifs. This study provides molecular evidence supporting the hypothesis that transposon-driven recruitment of pre-existing cis-regulatory elements from non-photosynthetic genes into photosynthetic genes plays an important role during C4 evolution. The findings of the present study coincide with the observed repetitive emergence of C4 during evolution.

  12. 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

  13. Understanding the Dynamics of Gene Regulatory Systems; Characterisation and Clinical Relevance of cis-Regulatory Polymorphisms

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    Philip Cowie

    2013-01-01

    Full Text Available Modern genetic analysis has shown that most polymorphisms associated with human disease are non-coding. Much of the functional information contained in the non-coding genome consists of cis-regulatory sequences (CRSs that are required to respond to signal transduction cues that direct cell specific gene expression. It has been hypothesised that many diseases may be due to polymorphisms within CRSs that alter their responses to signal transduction cues. However, identification of CRSs, and the effects of allelic variation on their ability to respond to signal transduction cues, is still at an early stage. In the current review we describe the use of comparative genomics and experimental techniques that allow for the identification of CRSs building on recent advances by the ENCODE consortium. In addition we describe techniques that allow for the analysis of the effects of allelic variation and epigenetic modification on CRS responses to signal transduction cues. Using specific examples we show that the interactions driving these elements are highly complex and the effects of disease associated polymorphisms often subtle. It is clear that gaining an understanding of the functions of CRSs, and how they are affected by SNPs and epigenetic modification, is essential to understanding the genetic basis of human disease and stratification whilst providing novel directions for the development of personalised medicine.

  14. A transposon-based chromosomal engineering method to survey a large cis-regulatory landscape in mice.

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    Kokubu, Chikara; Horie, Kyoji; Abe, Koichiro; Ikeda, Ryuji; Mizuno, Sumi; Uno, Yoshihiro; Ogiwara, Sanae; Ohtsuka, Masato; Isotani, Ayako; Okabe, Masaru; Imai, Kenji; Takeda, Junji

    2009-08-01

    A large cis-regulatory landscape is a common feature of vertebrate genomes, particularly at key developmental gene loci with finely tuned expression patterns. Existing genetic tools for surveying large genomic regions of interest spanning over hundreds of kilobases are limited. Here we propose a chromosomal engineering strategy exploiting the local hopping trait of the Sleeping Beauty transposon in the mouse genome. We generated embryonic stem cells with a targeted integration of the transposon vector, carrying an enhancer-detecting lacZ reporter and loxP cassette, into the developmentally critical Pax1 gene locus, followed by efficient local transpositions, nested deletion formation and derivation of embryos by tetraploid complementation. Comparative reporter expression analysis among different insertion/deletion embryos substantially facilitated long-range cis-regulatory element mapping in the genomic neighborhood and demonstrated the potential of the transposon-based approach as a versatile tool for exploration of defined genomic intervals of functional or clinical relevance, such as disease-associated microdeletions.

  15. Studying the functional conservation of cis-regulatory modules and their transcriptional output

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    Bailey Timothy L

    2008-04-01

    Full Text Available Abstract Background Cis-regulatory modules (CRMs are distinct, genomic regions surrounding the target gene that can independently activate the promoter to drive transcription. The activation of a CRM is controlled by the binding of a certain combination of transcription factors (TFs. It would be of great benefit if the transcriptional output mediated by a specific CRM could be predicted. Of equal benefit would be identifying in silico a specific CRM as the driver of the expression in a specific tissue or situation. We extend a recently developed biochemical modeling approach to manage both prediction tasks. Given a set of TFs, their protein concentrations, and the positions and binding strengths of each of the TFs in a putative CRM, the model predicts the transcriptional output of the gene. Our approach predicts the location of the regulating CRM by using predicted TF binding sites in regions near the gene as input to the model and searching for the region that yields a predicted transcription rate most closely matching the known rate. Results Here we show the ability of the model on the example of one of the CRMs regulating the eve gene, MSE2. A model trained on the MSE2 in D. melanogaster was applied to the surrounding sequence of the eve gene in seven other Drosophila species. The model successfully predicts the correct MSE2 location and output in six out of eight Drosophila species we examine. Conclusion The model is able to generalize from D. melanogaster to other Drosophila species and accurately predicts the location and transcriptional output of MSE2 in those species. However, we also show that the current model is not specific enough to function as a genome-wide CRM scanner, because it incorrectly predicts other genomic regions to be MSE2s.

  16. Studying the functional conservation of cis-regulatory modules and their transcriptional output.

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    Bauer, Denis C; Bailey, Timothy L

    2008-04-29

    Cis-regulatory modules (CRMs) are distinct, genomic regions surrounding the target gene that can independently activate the promoter to drive transcription. The activation of a CRM is controlled by the binding of a certain combination of transcription factors (TFs). It would be of great benefit if the transcriptional output mediated by a specific CRM could be predicted. Of equal benefit would be identifying in silico a specific CRM as the driver of the expression in a specific tissue or situation. We extend a recently developed biochemical modeling approach to manage both prediction tasks. Given a set of TFs, their protein concentrations, and the positions and binding strengths of each of the TFs in a putative CRM, the model predicts the transcriptional output of the gene. Our approach predicts the location of the regulating CRM by using predicted TF binding sites in regions near the gene as input to the model and searching for the region that yields a predicted transcription rate most closely matching the known rate. Here we show the ability of the model on the example of one of the CRMs regulating the eve gene, MSE2. A model trained on the MSE2 in D. melanogaster was applied to the surrounding sequence of the eve gene in seven other Drosophila species. The model successfully predicts the correct MSE2 location and output in six out of eight Drosophila species we examine. The model is able to generalize from D. melanogaster to other Drosophila species and accurately predicts the location and transcriptional output of MSE2 in those species. However, we also show that the current model is not specific enough to function as a genome-wide CRM scanner, because it incorrectly predicts other genomic regions to be MSE2s.

  17. Global profiling of rice and poplar transcriptomes highlights key conserved circadian-controlled pathways and cis-regulatory modules.

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    Sergei A Filichkin

    Full Text Available BACKGROUND: Circadian clocks provide an adaptive advantage through anticipation of daily and seasonal environmental changes. In plants, the central clock oscillator is regulated by several interlocking feedback loops. It was shown that a substantial proportion of the Arabidopsis genome cycles with phases of peak expression covering the entire day. Synchronized transcriptome cycling is driven through an extensive network of diurnal and clock-regulated transcription factors and their target cis-regulatory elements. Study of the cycling transcriptome in other plant species could thus help elucidate the similarities and differences and identify hubs of regulation common to monocot and dicot plants. METHODOLOGY/PRINCIPAL FINDINGS: Using a combination of oligonucleotide microarrays and data mining pipelines, we examined daily rhythms in gene expression in one monocotyledonous and one dicotyledonous plant, rice and poplar, respectively. Cycling transcriptomes were interrogated under different diurnal (driven and circadian (free running light and temperature conditions. Collectively, photocycles and thermocycles regulated about 60% of the expressed nuclear genes in rice and poplar. Depending on the condition tested, up to one third of oscillating Arabidopsis-poplar-rice orthologs were phased within three hours of each other suggesting a high degree of conservation in terms of rhythmic gene expression. We identified clusters of rhythmically co-expressed genes and searched their promoter sequences to identify phase-specific cis-elements, including elements that were conserved in the promoters of Arabidopsis, poplar, and rice. CONCLUSIONS/SIGNIFICANCE: Our results show that the cycling patterns of many circadian clock genes are highly conserved across poplar, rice, and Arabidopsis. The expression of many orthologous genes in key metabolic and regulatory pathways is diurnal and/or circadian regulated and phased to similar times of day. Our results confirm

  18. MotifCombinator: a web-based tool to search for combinations of cis-regulatory motifs

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    Tsunoda Tatsuhiko

    2007-03-01

    Full Text Available Abstract Background A combination of multiple types of transcription factors and cis-regulatory elements is often required for gene expression in eukaryotes, and the combinatorial regulation confers specific gene expression to tissues or environments. To reveal the combinatorial regulation, computational methods are developed that efficiently infer combinations of cis-regulatory motifs that are important for gene expression as measured by DNA microarrays. One promising type of computational method is to utilize regression analysis between expression levels and scores of motifs in input sequences. This type takes full advantage of information on expression levels because it does not require that the expression level of each gene be dichotomized according to whether or not it reaches a certain threshold level. However, there is no web-based tool that employs regression methods to systematically search for motif combinations and that practically handles combinations of more than two or three motifs. Results We here introduced MotifCombinator, an online tool with a user-friendly interface, to systematically search for combinations composed of any number of motifs based on regression methods. The tool utilizes well-known regression methods (the multivariate linear regression, the multivariate adaptive regression spline or MARS, and the multivariate logistic regression method for this purpose, and uses the genetic algorithm to search for combinations composed of any desired number of motifs. The visualization systems in this tool help users to intuitively grasp the process of the combination search, and the backup system allows users to easily stop and restart calculations that are expected to require large computational time. This tool also provides preparatory steps needed for systematic combination search – i.e., selecting single motifs to constitute combinations and cutting out redundant similar motifs based on clustering analysis. Conclusion

  19. 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.

  20. Evolved tooth gain in sticklebacks is associated with a cis-regulatory allele of Bmp6.

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    Cleves, Phillip A; Ellis, Nicholas A; Jimenez, Monica T; Nunez, Stephanie M; Schluter, Dolph; Kingsley, David M; Miller, Craig T

    2014-09-23

    Developmental genetic studies of evolved differences in morphology have led to the hypothesis that cis-regulatory changes often underlie morphological evolution. However, because most of these studies focus on evolved loss of traits, the genetic architecture and possible association with cis-regulatory changes of gain traits are less understood. Here we show that a derived benthic freshwater stickleback population has evolved an approximate twofold gain in ventral pharyngeal tooth number compared with their ancestral marine counterparts. Comparing laboratory-reared developmental time courses of a low-toothed marine population and this high-toothed benthic population reveals that increases in tooth number and tooth plate area and decreases in tooth spacing arise at late juvenile stages. Genome-wide linkage mapping identifies largely separate sets of quantitative trait loci affecting different aspects of dental patterning. One large-effect quantitative trait locus controlling tooth number fine-maps to a genomic region containing an excellent candidate gene, Bone morphogenetic protein 6 (Bmp6). Stickleback Bmp6 is expressed in developing teeth, and no coding changes are found between the high- and low-toothed populations. However, quantitative allele-specific expression assays of Bmp6 in developing teeth in F1 hybrids show that cis-regulatory changes have elevated the relative expression level of the freshwater benthic Bmp6 allele at late, but not early, stages of stickleback development. Collectively, our data support a model where a late-acting cis-regulatory up-regulation of Bmp6 expression underlies a significant increase in tooth number in derived benthic sticklebacks.

  1. Expression-guided in silico evaluation of candidate cis regulatory codes for Drosophila muscle founder cells.

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    Anthony A Philippakis

    2006-05-01

    Full Text Available While combinatorial models of transcriptional regulation can be inferred for metazoan systems from a priori biological knowledge, validation requires extensive and time-consuming experimental work. Thus, there is a need for computational methods that can evaluate hypothesized cis regulatory codes before the difficult task of experimental verification is undertaken. We have developed a novel computational framework (termed "CodeFinder" that integrates transcription factor binding site and gene expression information to evaluate whether a hypothesized transcriptional regulatory model (TRM; i.e., a set of co-regulating transcription factors is likely to target a given set of co-expressed genes. Our basic approach is to simultaneously predict cis regulatory modules (CRMs associated with a given gene set and quantify the enrichment for combinatorial subsets of transcription factor binding site motifs comprising the hypothesized TRM within these predicted CRMs. As a model system, we have examined a TRM experimentally demonstrated to drive the expression of two genes in a sub-population of cells in the developing Drosophila mesoderm, the somatic muscle founder cells. This TRM was previously hypothesized to be a general mode of regulation for genes expressed in this cell population. In contrast, the present analyses suggest that a modified form of this cis regulatory code applies to only a subset of founder cell genes, those whose gene expression responds to specific genetic perturbations in a similar manner to the gene on which the original model was based. We have confirmed this hypothesis by experimentally discovering six (out of 12 tested new CRMs driving expression in the embryonic mesoderm, four of which drive expression in founder cells.

  2. Rapid evolution of cis-regulatory sequences via local point mutations

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    Stone, J. R.; Wray, G. A.

    2001-01-01

    Although the evolution of protein-coding sequences within genomes is well understood, the same cannot be said of the cis-regulatory regions that control transcription. Yet, changes in gene expression are likely to constitute an important component of phenotypic evolution. We simulated the evolution of new transcription factor binding sites via local point mutations. The results indicate that new binding sites appear and become fixed within populations on microevolutionary timescales under an assumption of neutral evolution. Even combinations of two new binding sites evolve very quickly. We predict that local point mutations continually generate considerable genetic variation that is capable of altering gene expression.

  3. Conserved cis-regulatory regions in a large genomic landscape control SHH and BMP-regulated Gremlin1 expression in mouse limb buds

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    Zuniga Aimée

    2012-08-01

    Full Text Available Abstract Background Mouse limb bud is a prime model to study the regulatory interactions that control vertebrate organogenesis. Major aspects of limb bud development are controlled by feedback loops that define a self-regulatory signalling system. The SHH/GREM1/AER-FGF feedback loop forms the core of this signalling system that operates between the posterior mesenchymal organiser and the ectodermal signalling centre. The BMP antagonist Gremlin1 (GREM1 is a critical node in this system, whose dynamic expression is controlled by BMP, SHH, and FGF signalling and key to normal progression of limb bud development. Previous analysis identified a distant cis-regulatory landscape within the neighbouring Formin1 (Fmn1 locus that is required for Grem1 expression, reminiscent of the genomic landscapes controlling HoxD and Shh expression in limb buds. Results Three highly conserved regions (HMCO1-3 were identified within the previously defined critical genomic region and tested for their ability to regulate Grem1 expression in mouse limb buds. Using a combination of BAC and conventional transgenic approaches, a 9 kb region located ~70 kb downstream of the Grem1 transcription unit was identified. This region, termed Grem1 Regulatory Sequence 1 (GRS1, is able to recapitulate major aspects of Grem1 expression, as it drives expression of a LacZ reporter into the posterior and, to a lesser extent, in the distal-anterior mesenchyme. Crossing the GRS1 transgene into embryos with alterations in the SHH and BMP pathways established that GRS1 depends on SHH and is modulated by BMP signalling, i.e. integrates inputs from these pathways. Chromatin immunoprecipitation revealed interaction of endogenous GLI3 proteins with the core cis-regulatory elements in the GRS1 region. As GLI3 is a mediator of SHH signal transduction, these results indicated that SHH directly controls Grem1 expression through the GRS1 region. Finally, all cis-regulatory regions within the Grem1

  4. 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.

  5. 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.

  6. Establishment of the methods for searching eukaryotic gene cis-regulatory modules.

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    Zhong, Dong; Zhang, Zhen-shu; Liu, Yu-hu; Zheng, Guo-qing; Liu, Xiao-yi; Lu, Yang; Zhao, Gui-jun; Xu, An-long

    2004-02-01

    On the basis of the knowledge of eukaryotic gene regulation, we modified the method in three aspects: (1) Searching the cis-regulatory modules (CRM) according Fasta or Blast sequence with multiple sequence and low E value, followed by mutual scoring of these sequence with Smith-Waterman algorithms and finally by clustering analysis; (2) Searching the transcription factor-binding site using International Union of Pure and Applied Chemistry, Position-Weight Matrix(PWM) and Dyed method; (3) Designing and implementation of data analysis based on the software in Windows 2000 and UNIX using object-oriented technology. The results of analysis of the major histocompatibility complex gene family show that this procedure may accurately locate the regions that contain some of the CRMs.

  7. Evolution of metal hyperaccumulation required cis-regulatory changes and triplication of HMA4.

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    Hanikenne, Marc; Talke, Ina N; Haydon, Michael J; Lanz, Christa; Nolte, Andrea; Motte, Patrick; Kroymann, Juergen; Weigel, Detlef; Krämer, Ute

    2008-05-15

    Little is known about the types of mutations underlying the evolution of species-specific traits. The metal hyperaccumulator Arabidopsis halleri has the rare ability to colonize heavy-metal-polluted soils, and, as an extremophile sister species of Arabidopsis thaliana, it is a powerful model for research on adaptation. A. halleri naturally accumulates and tolerates leaf concentrations as high as 2.2% zinc and 0.28% cadmium in dry biomass. On the basis of transcriptomics studies, metal hyperaccumulation in A. halleri has been associated with more than 30 candidate genes that are expressed at higher levels in A. halleri than in A. thaliana. Some of these genes have been genetically mapped to broad chromosomal segments of between 4 and 24 cM co-segregating with Zn and Cd hypertolerance. However, the in planta loss-of-function approaches required to demonstrate the contribution of a given candidate gene to metal hyperaccumulation or hypertolerance have not been pursued to date. Using RNA interference to downregulate HMA4 (HEAVY METAL ATPASE 4) expression, we show here that Zn hyperaccumulation and full hypertolerance to Cd and Zn in A. halleri depend on the metal pump HMA4. Contrary to a postulated global trans regulatory factor governing high expression of numerous metal hyperaccumulation genes, we demonstrate that enhanced expression of HMA4 in A. halleri is attributable to a combination of modified cis-regulatory sequences and copy number expansion, in comparison to A. thaliana. Transfer of an A. halleri HMA4 gene to A. thaliana recapitulates Zn partitioning into xylem vessels and the constitutive transcriptional upregulation of Zn deficiency response genes characteristic of Zn hyperaccumulators. Our results demonstrate the importance of cis-regulatory mutations and gene copy number expansion in the evolution of a complex naturally selected extreme trait. The elucidation of a natural strategy for metal hyperaccumulation enables the rational design of technologies

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

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    He, Xin; Ling, Xu; Sinha, Saurabh

    2009-03-01

    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.

  9. 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.

  10. In silico evolution of the hunchback gene indicates redundancy in cis-regulatory organization and spatial gene expression.

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    Zagrijchuk, Elizaveta A; Sabirov, Marat A; Holloway, David M; Spirov, Alexander V

    2014-04-01

    Biological development depends on the coordinated expression of genes in time and space. Developmental genes have extensive cis-regulatory regions which control their expression. These regions are organized in a modular manner, with different modules controlling expression at different times and locations. Both how modularity evolved and what function it serves are open questions. We present a computational model for the cis-regulation of the hunchback (hb) gene in the fruit fly (Drosophila). We simulate evolution (using an evolutionary computation approach from computer science) to find the optimal cis-regulatory arrangements for fitting experimental hb expression patterns. We find that the cis-regulatory region tends to readily evolve modularity. These cis-regulatory modules (CRMs) do not tend to control single spatial domains, but show a multi-CRM/multi-domain correspondence. We find that the CRM-domain correspondence seen in Drosophila evolves with a high probability in our model, supporting the biological relevance of the approach. The partial redundancy resulting from multi-CRM control may confer some biological robustness against corruption of regulatory sequences. The technique developed on hb could readily be applied to other multi-CRM developmental genes.

  11. Evolutionary processes acting on candidate cis-regulatory regions in humans inferred from patterns of polymorphism and divergence.

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    Dara G Torgerson

    2009-08-01

    Full Text Available Analysis of polymorphism and divergence in the non-coding portion of the human genome yields crucial information about factors driving the evolution of gene regulation. Candidate cis-regulatory regions spanning more than 15,000 genes in 15 African Americans and 20 European Americans were re-sequenced and aligned to the chimpanzee genome in order to identify potentially functional polymorphism and to characterize and quantify departures from neutral evolution. Distortions of the site frequency spectra suggest a general pattern of selective constraint on conserved non-coding sites in the flanking regions of genes (CNCs. Moreover, there is an excess of fixed differences that cannot be explained by a Gamma model of deleterious fitness effects, suggesting the presence of positive selection on CNCs. Extensions of the McDonald-Kreitman test identified candidate cis-regulatory regions with high probabilities of positive and negative selection near many known human genes, the biological characteristics of which exhibit genome-wide trends that differ from patterns observed in protein-coding regions. Notably, there is a higher probability of positive selection in candidate cis-regulatory regions near genes expressed in the fetal brain, suggesting that a larger portion of adaptive regulatory changes has occurred in genes expressed during brain development. Overall we find that natural selection has played an important role in the evolution of candidate cis-regulatory regions throughout hominid evolution.

  12. In silico evolution of the hunchback gene indicates redundancy in cis-regulatory organization and spatial gene expression

    Science.gov (United States)

    Zagrijchuk, Elizaveta A.; Sabirov, Marat A.; Holloway, David M.; Spirov, Alexander V.

    2014-01-01

    Biological development depends on the coordinated expression of genes in time and space. Developmental genes have extensive cis-regulatory regions which control their expression. These regions are organized in a modular manner, with different modules controlling expression at different times and locations. Both how modularity evolved and what function it serves are open questions. We present a computational model for the cis-regulation of the hunchback (hb) gene in the fruit fly (Drosophila). We simulate evolution (using an evolutionary computation approach from computer science) to find the optimal cis-regulatory arrangements for fitting experimental hb expression patterns. We find that the cis-regulatory region tends to readily evolve modularity. These cis-regulatory modules (CRMs) do not tend to control single spatial domains, but show a multi-CRM/multi-domain correspondence. We find that the CRM-domain correspondence seen in Drosophila evolves with a high probability in our model, supporting the biological relevance of the approach. The partial redundancy resulting from multi-CRM control may confer some biological robustness against corruption of regulatory sequences. The technique developed on hb could readily be applied to other multi-CRM developmental genes. PMID:24712536

  13. On the mechanistic nature of epistasis in a canonical cis-regulatory element

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    Lagator, Mato; Paix?o, Tiago; Barton, Nicholas H; Bollback, Jonathan P.; Guet, C?lin C.

    2017-01-01

    eLife digest Mutations are changes to DNA that provide the raw material upon which evolution can act. Therefore, to understand evolution, we need to know the effects of mutations, and how those mutations interact with each other (a phenomenon referred to as epistasis). So far, few mathematical models allow scientists to predict the effects of mutations, and even fewer are able to predict epistasis. Biological systems are complex and consist of many proteins and other molecules. Genes are the ...

  14. New cis-regulatory elements in the Rht-D1b locus region of wheat

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    Fifteen gene-containing BACs with accumulated length of 1.82-Mb from the Rht-D1b locus region weresequenced and compared in detail with the orthologous regions of rice, sorghum, and maize. Our results show that Rht-D1b represents a conserved genomic region as implied by high gene sequence identity...

  15. 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.

  16. 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.

  17. Rearrangements of 2.5 kilobases of noncoding DNA from the Drosophila even-skipped locus define predictive rules of genomic cis-regulatory logic.

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    Ah-Ram Kim

    Full Text Available Rearrangements of about 2.5 kilobases of regulatory DNA located 5' of the transcription start site of the Drosophila even-skipped locus generate large-scale changes in the expression of even-skipped stripes 2, 3, and 7. The most radical effects are generated by juxtaposing the minimal stripe enhancers MSE2 and MSE3 for stripes 2 and 3 with and without small "spacer" segments less than 360 bp in length. We placed these fusion constructs in a targeted transformation site and obtained quantitative expression data for these transformants together with their controlling transcription factors at cellular resolution. These data demonstrated that the rearrangements can alter expression levels in stripe 2 and the 2-3 interstripe by a factor of more than 10. We reasoned that this behavior would place tight constraints on possible rules of genomic cis-regulatory logic. To find these constraints, we confronted our new expression data together with previously obtained data on other constructs with a computational model. The model contained representations of thermodynamic protein-DNA interactions including steric interference and cooperative binding, short-range repression, direct repression, activation, and coactivation. The model was highly constrained by the training data, which it described within the limits of experimental error. The model, so constrained, was able to correctly predict expression patterns driven by enhancers for other Drosophila genes; even-skipped enhancers not included in the training set; stripe 2, 3, and 7 enhancers from various Drosophilid and Sepsid species; and long segments of even-skipped regulatory DNA that contain multiple enhancers. The model further demonstrated that elevated expression driven by a fusion of MSE2 and MSE3 was a consequence of the recruitment of a portion of MSE3 to become a functional component of MSE2, demonstrating that cis-regulatory "elements" are not elementary objects.

  18. Single nucleotide polymorphisms with cis-regulatory effects on long non-coding transcripts in human primary monocytes.

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    Jonas Carlsson Almlöf

    Full Text Available We applied genome-wide allele-specific expression analysis of monocytes from 188 samples. Monocytes were purified from white blood cells of healthy blood donors to detect cis-acting genetic variation that regulates the expression of long non-coding RNAs. We analysed 8929 regions harboring genes for potential long non-coding RNA that were retrieved from data from the ENCODE project. Of these regions, 60% were annotated as intergenic, which implies that they do not overlap with protein-coding genes. Focusing on the intergenic regions, and using stringent analysis of the allele-specific expression data, we detected robust cis-regulatory SNPs in 258 out of 489 informative intergenic regions included in the analysis. The cis-regulatory SNPs that were significantly associated with allele-specific expression of long non-coding RNAs were enriched to enhancer regions marked for active or bivalent, poised chromatin by histone modifications. Out of the lncRNA regions regulated by cis-acting regulatory SNPs, 20% (n = 52 were co-regulated with the closest protein coding gene. We compared the identified cis-regulatory SNPs with those in the catalog of SNPs identified by genome-wide association studies of human diseases and traits. This comparison identified 32 SNPs in loci from genome-wide association studies that displayed a strong association signal with allele-specific expression of non-coding RNAs in monocytes, with p-values ranging from 6.7×10(-7 to 9.5×10(-89. The identified cis-regulatory SNPs are associated with diseases of the immune system, like multiple sclerosis and rheumatoid arthritis.

  19. 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

  20. Conserved Cis-Regulatory Modules Control Robustness in Msx1 Expression at Single-Cell Resolution

    Science.gov (United States)

    Vance, Keith W.; Woodcock, Dan J.; Reid, John E.; Bretschneider, Till; Ott, Sascha; Koentges, Georgy

    2015-01-01

    The process of transcription is highly stochastic leading to cell-to-cell variations and noise in gene expression levels. However, key essential genes have to be precisely expressed at the correct amount and time to ensure proper cellular development and function. Studies in yeast and bacterial systems have shown that gene expression noise decreases as mean expression levels increase, a relationship that is controlled by promoter DNA sequence. However, the function of distal cis-regulatory modules (CRMs), an evolutionary novelty of metazoans, in controlling transcriptional robustness and variability is poorly understood. In this study, we used live cell imaging of transfected reporters combined with a mathematical modelling and statistical inference scheme to quantify the function of conserved Msx1 CRMs and promoters in modulating single-cell real-time transcription rates in C2C12 mouse myoblasts. The results show that the mean expression–noise relationship is solely promoter controlled for this key pluripotency regulator. In addition, we demonstrate that CRMs modulate single-cell basal promoter rate distributions in a graded manner across a population of cells. This extends the rheostatic model of CRM action to provide a more detailed understanding of CRM function at single-cell resolution. We also identify a novel CRM transcriptional filter function that acts to reduce intracellular variability in transcription rates and show that this can be phylogenetically separable from rate modulating CRM activities. These results are important for understanding how the expression of key vertebrate developmental transcription factors is precisely controlled both within and between individual cells. PMID:26342140

  1. Identification of High-Impact cis-Regulatory Mutations Using Transcription Factor Specific Random Forest Models.

    Directory of Open Access Journals (Sweden)

    Dmitry Svetlichnyy

    2015-11-01

    Full Text Available Cancer genomes contain vast amounts of somatic mutations, many of which are passenger mutations not involved in oncogenesis. Whereas driver mutations in protein-coding genes can be distinguished from passenger mutations based on their recurrence, non-coding mutations are usually not recurrent at the same position. Therefore, it is still unclear how to identify cis-regulatory driver mutations, particularly when chromatin data from the same patient is not available, thus relying only on sequence and expression information. Here we use machine-learning methods to predict functional regulatory regions using sequence information alone, and compare the predicted activity of the mutated region with the reference sequence. This way we define the Predicted Regulatory Impact of a Mutation in an Enhancer (PRIME. We find that the recently identified driver mutation in the TAL1 enhancer has a high PRIME score, representing a "gain-of-target" for MYB, whereas the highly recurrent TERT promoter mutation has a surprisingly low PRIME score. We trained Random Forest models for 45 cancer-related transcription factors, and used these to score variations in the HeLa genome and somatic mutations across more than five hundred cancer genomes. Each model predicts only a small fraction of non-coding mutations with a potential impact on the function of the encompassing regulatory region. Nevertheless, as these few candidate driver mutations are often linked to gains in chromatin activity and gene expression, they may contribute to the oncogenic program by altering the expression levels of specific oncogenes and tumor suppressor genes.

  2. cis-Regulatory control of three cell fate-specific genes in vulval organogenesis of Caenorhabditis elegans and C. briggsae.

    Science.gov (United States)

    Kirouac, Martha; Sternberg, Paul W

    2003-05-01

    The great-grandprogeny of the Caenorhabditis elegans vulval precursor cells (VPCs) adopt one of the final vulA, B1, B2, C, D, E, and F cell fates in a precise spatial pattern. This pattern of vulval cell types is likely to depend on the cis-regulatory regions of the transcriptional targets of intercellular signals in vulval development. egl-17, zmp-1, and cdh-3 are expressed differentially in the developing vulva cells, providing a potential readout for different signaling pathways. To understand how such pathways interact to specify unique vulval cell types in a precise pattern, we have identified cis-regulatory regions sufficient to confer vulval cell type-specific regulation when fused in cis to the basal pes-10 promoter. We have identified the C. briggsae homologs of these three genes, with their corresponding control regions, and tested these regions in both C. elegans and C. briggsae. These regions of similarity in C. elegans and C. briggsae upstream of egl-17, zmp-1, and cdh-3 promote expression in vulval cells and the anchor cell (AC). By using the cis-regulatory analysis and phylogenetic footprinting, we have identified overrepresented sequences involved in conferring vulval and AC expression.

  3. 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.

  4. Novel mutations in the GH gene (GH1) uncover putative splicing regulatory elements.

    Science.gov (United States)

    Babu, Deepak; Mellone, Simona; Fusco, Ileana; Petri, Antonella; Walker, Gillian E; Bellone, Simonetta; Prodam, Flavia; Momigliano-Richiardi, Patricia; Bona, Gianni; Giordano, Mara

    2014-05-01

    Mutations affecting exon 3 splicing are the main cause of autosomal dominant Isolated GH Deficiency II (IGHDII) by increasing the level of exon 3-skipped mRNA encoding the functionally inactive dominant-negative 17.5-kDa isoform. The exons and introns of the gene encoding GH (GH1) were screened for the presence of mutations in 103 sporadic isolated GH deficiency cases. Four different variations within exon 3 were identified in 3 patients. One carried c.261C>T (p.Pro87Pro) and c.272A>T (p.Glu91Val), the second c.255G>A (p.Pro85Pro) and c.261 C>T, and the third c.246G>C (p.Glu82Asp). All the variants were likely generated by gene conversion from an homologous gene in the GH1 cluster. In silico analysis predicted that positions c.255 and c.272 were included within 2 putative novel exon splicing enhancers (ESEs). Their effect on splicing was confirmed in vitro. Constructs bearing these 2 variants induced consistently higher levels both of transcript and protein corresponding to the 17.5-kDa isoform. When c.255 and c.272 were combined in cis with the c.261 variant, as in our patients, their effect was weaker. In conclusion, we identified 2 variations, c.255G>A and c.272A>T, located in 2 novel putative exon splicing enhancers and affecting GH1 splicing in vitro by increasing the production of alternatively spliced isoforms. The amount of aberrant isoforms is further regulated by the presence in cis of the c.261 variant. Thus, our results evidenced novel putative splicing regulatory elements within exon 3, confirming the crucial role of this exon in mRNA processing.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  6. 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

    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...

  7. Cis-regulatory underpinnings of human GLI3 expression in embryonic craniofacial structures and internal organs.

    Science.gov (United States)

    Abbasi, Amir A; Minhas, Rashid; Schmidt, Ansgar; Koch, Sabine; Grzeschik, Karl-Heinz

    2013-10-01

    The zinc finger transcription factor Gli3 is an important mediator of Sonic hedgehog (Shh) signaling. During early embryonic development Gli3 participates in patterning and growth of the central nervous system, face, skeleton, limb, tooth and gut. Precise regulation of the temporal and spatial expression of Gli3 is crucial for the proper specification of these structures in mammals and other vertebrates. Previously we reported a set of human intronic cis-regulators controlling almost the entire known repertoire of endogenous Gli3 expression in mouse neural tube and limbs. However, the genetic underpinning of GLI3 expression in other embryonic domains such as craniofacial structures and internal organs remain elusive. Here we demonstrate in a transgenic mice assay the potential of a subset of human/fish conserved non-coding sequences (CNEs) residing within GLI3 intronic intervals to induce reporter gene expression at known regions of endogenous Gli3 transcription in embryonic domains other than central nervous system (CNS) and limbs. Highly specific reporter expression was observed in craniofacial structures, eye, gut, and genitourinary system. Moreover, the comparison of expression patterns directed by these intronic cis-acting regulatory elements in mouse and zebrafish embryos suggests that in accordance with sequence conservation, the target site specificity of a subset of these elements remains preserved among these two lineages. Taken together with our recent investigations, it is proposed here that during vertebrate evolution the Gli3 expression control acquired multiple, independently acting, intronic enhancers for spatiotemporal patterning of CNS, limbs, craniofacial structures and internal organs.

  8. Ancient polymorphism and functional variation in the primate MHC-DQA1 5' cis-regulatory region.

    Science.gov (United States)

    Loisel, Dagan A; Rockman, Matthew V; Wray, Gregory A; Altmann, Jeanne; Alberts, Susan C

    2006-10-31

    Precise regulation of MHC gene expression is critical to vertebrate immune surveillance and response. Polymorphisms in the 5' proximal promoter region of the human class II gene HLA-DQA1 have been shown to influence its transcriptional regulation and may contribute to the pathogenesis of autoimmune diseases. We investigated the evolutionary history of this cis-regulatory region by sequencing the DQA1 5' proximal promoter region in eight nonhuman primate species. We observed unexpectedly high levels of sequence variation and multiple strong signatures of balancing selection in this region. Specifically, the considerable DQA1 promoter region diversity was characterized by abundant shared (or trans-species) polymorphism and a pronounced lack of fixed differences between species. The majority of transcription factor binding sites in the DQA1 promoter region were polymorphic within species, and these binding site polymorphisms were commonly shared among multiple species despite evidence for negative selection eliminating a significant fraction of binding site mutations. We assessed the functional consequences of intraspecific promoter region diversity using a cell line-based reporter assay and detected significant differences among baboon DQA1 promoter haplotypes in their ability to drive transcription in vitro. The functional differentiation of baboon promoter haplotypes, together with the significant deviations from neutral sequence evolution, suggests a role for balancing selection in the evolution of DQA1 transcriptional regulation in primates.

  9. Comparative epigenomics in distantly related teleost species identifies conserved cis-regulatory nodes active during the vertebrate phylotypic period

    Science.gov (United States)

    Tena, Juan J.; González-Aguilera, Cristina; Fernández-Miñán, Ana; Vázquez-Marín, Javier; Parra-Acero, Helena; Cross, Joe W.; Rigby, Peter W.J.; Carvajal, Jaime J.; Wittbrodt, Joachim; Gómez-Skarmeta, José L.; Martínez-Morales, Juan R.

    2014-01-01

    The complex relationship between ontogeny and phylogeny has been the subject of attention and controversy since von Baer’s formulations in the 19th century. The classic concept that embryogenesis progresses from clade general features to species-specific characters has often been revisited. It has become accepted that embryos from a clade show maximum morphological similarity at the so-called phylotypic period (i.e., during mid-embryogenesis). According to the hourglass model, body plan conservation would depend on constrained molecular mechanisms operating at this period. More recently, comparative transcriptomic analyses have provided conclusive evidence that such molecular constraints exist. Examining cis-regulatory architecture during the phylotypic period is essential to understand the evolutionary source of body plan stability. Here we compare transcriptomes and key epigenetic marks (H3K4me3 and H3K27ac) from medaka (Oryzias latipes) and zebrafish (Danio rerio), two distantly related teleosts separated by an evolutionary distance of 115–200 Myr. We show that comparison of transcriptome profiles correlates with anatomical similarities and heterochronies observed at the phylotypic stage. Through comparative epigenomics, we uncover a pool of conserved regulatory regions (≈700), which are active during the vertebrate phylotypic period in both species. Moreover, we show that their neighboring genes encode mainly transcription factors with fundamental roles in tissue specification. We postulate that these regulatory regions, active in both teleost genomes, represent key constrained nodes of the gene networks that sustain the vertebrate body plan. PMID:24709821

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

    Directory of Open Access Journals (Sweden)

    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.

  11. Network discovery pipeline elucidates conserved time-of-day-specific cis-regulatory modules.

    Directory of Open Access Journals (Sweden)

    Todd P Michael

    2008-02-01

    Full Text Available Correct daily phasing of transcription confers an adaptive advantage to almost all organisms, including higher plants. In this study, we describe a hypothesis-driven network discovery pipeline that identifies biologically relevant patterns in genome-scale data. To demonstrate its utility, we analyzed a comprehensive matrix of time courses interrogating the nuclear transcriptome of Arabidopsis thaliana plants grown under different thermocycles, photocycles, and circadian conditions. We show that 89% of Arabidopsis transcripts cycle in at least one condition and that most genes have peak expression at a particular time of day, which shifts depending on the environment. Thermocycles alone can drive at least half of all transcripts critical for synchronizing internal processes such as cell cycle and protein synthesis. We identified at least three distinct transcription modules controlling phase-specific expression, including a new midnight specific module, PBX/TBX/SBX. We validated the network discovery pipeline, as well as the midnight specific module, by demonstrating that the PBX element was sufficient to drive diurnal and circadian condition-dependent expression. Moreover, we show that the three transcription modules are conserved across Arabidopsis, poplar, and rice. These results confirm the complex interplay between thermocycles, photocycles, and the circadian clock on the daily transcription program, and provide a comprehensive view of the conserved genomic targets for a transcriptional network key to successful adaptation.

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

    NARCIS (Netherlands)

    Kusters, E.; Della Pina, S.; Castel, R.; Souer, E.; Koes, R.

    2015-01-01

    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

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

    NARCIS (Netherlands)

    Kusters, E.; Della Pina, S.; Castel, R.; Souer, E.; Koes, R.

    2015-01-01

    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 ch

  14. Expression, subcellular localization, and cis-regulatory structure of duplicated phytoene synthase genes in melon (Cucumis melo L.).

    Science.gov (United States)

    Qin, Xiaoqiong; Coku, Ardian; Inoue, Kentaro; Tian, Li

    2011-10-01

    Carotenoids perform many critical functions in plants, animals, and humans. It is therefore important to understand carotenoid biosynthesis and its regulation in plants. Phytoene synthase (PSY) catalyzes the first committed and rate-limiting step in carotenoid biosynthesis. While PSY is present as a single copy gene in Arabidopsis, duplicated PSY genes have been identified in many economically important monocot and dicot crops. CmPSY1 was previously identified from melon (Cucumis melo L.), but was not functionally characterized. We isolated a second PSY gene, CmPSY2, from melon in this work. CmPSY2 possesses a unique intron/exon structure that has not been observed in other plant PSYs. Both CmPSY1 and CmPSY2 are functional in vitro, but exhibit distinct expression patterns in different melon tissues and during fruit development, suggesting differential regulation of the duplicated melon PSY genes. In vitro chloroplast import assays verified the plastidic localization of CmPSY1 and CmPSY2 despite the lack of an obvious plastid target peptide in CmPSY2. Promoter motif analysis of the duplicated melon and tomato PSY genes and the Arabidopsis PSY revealed distinctive cis-regulatory structures of melon PSYs and identified gibberellin-responsive motifs in all PSYs except for SlPSY1, which has not been reported previously. Overall, these data provide new insights into the evolutionary history of plant PSY genes and the regulation of PSY expression by developmental and environmental signals that may involve different regulatory networks.

  15. Cis-regulatory variants affect CHRNA5 mRNA expression in populations of African and European ancestry.

    Directory of Open Access Journals (Sweden)

    Jen-Chyong Wang

    Full Text Available Variants within the gene cluster encoding α3, α5, and β4 nicotinic receptor subunits are major risk factors for substance dependence. The strongest impact on risk is associated with variation in the CHRNA5 gene, where at least two mechanisms are at work: amino acid variation and altered mRNA expression levels. The risk allele of the non-synonymous variant (rs16969968; D398N primarily occurs on the haplotype containing the low mRNA expression allele. In populations of European ancestry, there are approximately 50 highly correlated variants in the CHRNA5-CHRNA3-CHRNB4 gene cluster and the adjacent PSMA4 gene region that are associated with CHRNA5 mRNA levels. It is not clear which of these variants contribute to the changes in CHRNA5 transcript level. Because populations of African ancestry have reduced linkage disequilibrium among variants spanning this gene cluster, eQTL mapping in subjects of African ancestry could potentially aid in defining the functional variants that affect CHRNA5 mRNA levels. We performed quantitative allele specific gene expression using frontal cortices derived from 49 subjects of African ancestry and 111 subjects of European ancestry. This method measures allele-specific transcript levels in the same individual, which eliminates other biological variation that occurs when comparing expression levels between different samples. This analysis confirmed that substance dependence associated variants have a direct cis-regulatory effect on CHRNA5 transcript levels in human frontal cortices of African and European ancestry and identified 10 highly correlated variants, located in a 9 kb region, that are potential functional variants modifying CHRNA5 mRNA expression levels.

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

    Directory of Open Access Journals (Sweden)

    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.

  17. Identification and Characterization of Putative Integron-Like Elements of the Heavy-Metal-Hypertolerant Strains of Pseudomonas spp.

    Science.gov (United States)

    Ciok, Anna; Adamczuk, Marcin; Bartosik, Dariusz; Dziewit, Lukasz

    2016-11-28

    Pseudomonas strains isolated from the heavily contaminated Lubin copper mine and Zelazny Most post-flotation waste reservoir in Poland were screened for the presence of integrons. This analysis revealed that two strains carried homologous DNA regions composed of a gene encoding a DNA_BRE_C domain-containing tyrosine recombinase (with no significant sequence similarity to other integrases of integrons) plus a three-component array of putative integron gene cassettes. The predicted gene cassettes encode three putative polypeptides with homology to (i) transmembrane proteins, (ii) GCN5 family acetyltransferases, and (iii) hypothetical proteins of unknown function (homologous proteins are encoded by the gene cassettes of several class 1 integrons). Comparative sequence analyses identified three structural variants of these novel integron-like elements within the sequenced bacterial genomes. Analysis of their distribution revealed that they are found exclusively in strains of the genus Pseudomonas.

  18. A Generalized Linear Model for Decomposing Cis-regulatory, Parent-of-Origin, and Maternal Effects on Allele-Specific Gene Expression

    Directory of Open Access Journals (Sweden)

    Yasuaki Takada

    2017-07-01

    Full Text Available Joint quantification of genetic and epigenetic effects on gene expression is important for understanding the establishment of complex gene regulation systems in living organisms. In particular, genomic imprinting and maternal effects play important roles in the developmental process of mammals and flowering plants. However, the influence of these effects on gene expression are difficult to quantify because they act simultaneously with cis-regulatory mutations. Here we propose a simple method to decompose cis-regulatory (i.e., allelic genotype, genomic imprinting [i.e., parent-of-origin (PO], and maternal [i.e., maternal genotype (MG] effects on allele-specific gene expression using RNA-seq data obtained from reciprocal crosses. We evaluated the efficiency of method using a simulated dataset and applied the method to whole-body Drosophila and mouse trophoblast stem cell (TSC and liver RNA-seq data. Consistent with previous studies, we found little evidence of PO and MG effects in adult Drosophila samples. In contrast, we identified dozens and hundreds of mouse genes with significant PO and MG effects, respectively. Interestingly, a similar number of genes with significant PO effect were detect in mouse TSCs and livers, whereas more genes with significant MG effect were observed in livers. Further application of this method will clarify how these three effects influence gene expression levels in different tissues and developmental stages, and provide novel insight into the evolution of gene expression regulation.

  19. Identification of putative PPAR response elements in and around the murine UCP3 gene

    DEFF Research Database (Denmark)

    Siersbæk, Majken

    in thermogenesis in unerring, the physiological functions of UCP2 and UCP3 are at present not fully understood. Synthetic agonists for the peroxisome proliferator-activated receptors (PPARs) as well as fatty acids have been shown to increase murine UCP2 and UCP3 mRNA expression but response elements and mechanisms...... differentiation. Results from sequencing of chromatin immunoprecipitated (ChIP) material from 3T3-L1 adipocytes revealed three PPAR and retinoid X receptor (RXR) binding sites in and around the murine UCP3 gene. The recruitment of PPAR and RXR to the three potential PPREs was determined by ChIP combined...

  20. Identification of putative PPAR response elements in and around the murine UCP3 gene

    DEFF Research Database (Denmark)

    Siersbæk, Majken

    in thermogenesis in unerring, the physiological functions of UCP2 and UCP3 are at present not fully understood. Synthetic agonists for the peroxisome proliferator-activated receptors (PPARs) as well as fatty acids have been shown to increase murine UCP2 and UCP3 mRNA expression but response elements and mechanisms...... are not yet characterized. The aim of this study was to investigate the transcriptional regulation of UCP3 by the PPARs. Results: The PPAR agonists increase UCP2 and UCP3 mRNA expression in skeletal muscle cells (C2C12). In addition, UCP2 and UCP3 mRNA expression is upregulated during 3T3-L1 adipocyte...... differentiation. Results from sequencing of chromatin immunoprecipitated (ChIP) material from 3T3-L1 adipocytes revealed three PPAR and retinoid X receptor (RXR) binding sites in and around the murine UCP3 gene. The recruitment of PPAR and RXR to the three potential PPREs was determined by ChIP combined...

  1. Genome-wide identification of enhancer elements.

    Science.gov (United States)

    Tulin, Sarah; Barsi, Julius C; Bocconcelli, Carlo; Smith, Joel

    2016-01-01

    We present a prospective genome-wide regulatory element database for the sea urchin embryo and the modified chromosome capture-related methodology used to create it. The method we developed is termed GRIP-seq for genome-wide regulatory element immunoprecipitation and combines features of chromosome conformation capture, chromatin immunoprecipitation, and paired-end next-generation sequencing with molecular steps that enrich for active cis-regulatory elements associated with basal transcriptional machinery. The first GRIP-seq database, available to the community, comes from S. purpuratus 24 hpf embryos and takes advantage of the extremely well-characterized cis-regulatory elements in this system for validation. In addition, using the GRIP-seq database, we identify and experimentally validate a novel, intronic cis-regulatory element at the onecut locus. We find GRIP-seq signal sensitively identifies active cis-regulatory elements with a high signal-to-noise ratio for both distal and intronic elements. This promising GRIP-seq protocol has the potential to address a rate-limiting step in resolving comprehensive, predictive network models in all systems.

  2. A cis-regulatory mutation in troponin-I of Drosophila reveals the importance of proper stoichiometry of structural proteins during muscle assembly.

    Science.gov (United States)

    Firdaus, Hena; Mohan, Jayaram; Naz, Sarwat; Arathi, Prabhashankar; Ramesh, Saraf R; Nongthomba, Upendra

    2015-05-01

    Rapid and high wing-beat frequencies achieved during insect flight are powered by the indirect flight muscles, the largest group of muscles present in the thorax. Any anomaly during the assembly and/or structural impairment of the indirect flight muscles gives rise to a flightless phenotype. Multiple mutagenesis screens in Drosophila melanogaster for defective flight behavior have led to the isolation and characterization of mutations that have been instrumental in the identification of many proteins and residues that are important for muscle assembly, function, and disease. In this article, we present a molecular-genetic characterization of a flightless mutation, flightless-H (fliH), originally designated as heldup-a (hdp-a). We show that fliH is a cis-regulatory mutation of the wings up A (wupA) gene, which codes for the troponin-I protein, one of the troponin complex proteins, involved in regulation of muscle contraction. The mutation leads to reduced levels of troponin-I transcript and protein. In addition to this, there is also coordinated reduction in transcript and protein levels of other structural protein isoforms that are part of the troponin complex. The altered transcript and protein stoichiometry ultimately culminates in unregulated acto-myosin interactions and a hypercontraction muscle phenotype. Our results shed new insights into the importance of maintaining the stoichiometry of structural proteins during muscle assembly for proper function with implications for the identification of mutations and disease phenotypes in other species, including humans.

  3. Sex Chromosome-wide Transcriptional Suppression and Compensatory Cis-Regulatory Evolution Mediate Gene Expression in the Drosophila Male Germline.

    Directory of Open Access Journals (Sweden)

    Emily L Landeen

    2016-07-01

    Full Text Available The evolution of heteromorphic sex chromosomes has repeatedly resulted in the evolution of sex chromosome-specific forms of regulation, including sex chromosome dosage compensation in the soma and meiotic sex chromosome inactivation in the germline. In the male germline of Drosophila melanogaster, a novel but poorly understood form of sex chromosome-specific transcriptional regulation occurs that is distinct from canonical sex chromosome dosage compensation or meiotic inactivation. Previous work shows that expression of reporter genes driven by testis-specific promoters is considerably lower-approximately 3-fold or more-for transgenes inserted into X chromosome versus autosome locations. Here we characterize this transcriptional suppression of X-linked genes in the male germline and its evolutionary consequences. Using transgenes and transpositions, we show that most endogenous X-linked genes, not just testis-specific ones, are transcriptionally suppressed several-fold specifically in the Drosophila male germline. In wild-type testes, this sex chromosome-wide transcriptional suppression is generally undetectable, being effectively compensated by the gene-by-gene evolutionary recruitment of strong promoters on the X chromosome. We identify and experimentally validate a promoter element sequence motif that is enriched upstream of the transcription start sites of hundreds of testis-expressed genes; evolutionarily conserved across species; associated with strong gene expression levels in testes; and overrepresented on the X chromosome. These findings show that the expression of X-linked genes in the Drosophila testes reflects a balance between chromosome-wide epigenetic transcriptional suppression and long-term compensatory adaptation by sex-linked genes. Our results have broad implications for the evolution of gene expression in the Drosophila male germline and for genome evolution.

  4. Sex Chromosome-wide Transcriptional Suppression and Compensatory Cis-Regulatory Evolution Mediate Gene Expression in the Drosophila Male Germline.

    Science.gov (United States)

    Landeen, Emily L; Muirhead, Christina A; Wright, Lori; Meiklejohn, Colin D; Presgraves, Daven C

    2016-07-01

    The evolution of heteromorphic sex chromosomes has repeatedly resulted in the evolution of sex chromosome-specific forms of regulation, including sex chromosome dosage compensation in the soma and meiotic sex chromosome inactivation in the germline. In the male germline of Drosophila melanogaster, a novel but poorly understood form of sex chromosome-specific transcriptional regulation occurs that is distinct from canonical sex chromosome dosage compensation or meiotic inactivation. Previous work shows that expression of reporter genes driven by testis-specific promoters is considerably lower-approximately 3-fold or more-for transgenes inserted into X chromosome versus autosome locations. Here we characterize this transcriptional suppression of X-linked genes in the male germline and its evolutionary consequences. Using transgenes and transpositions, we show that most endogenous X-linked genes, not just testis-specific ones, are transcriptionally suppressed several-fold specifically in the Drosophila male germline. In wild-type testes, this sex chromosome-wide transcriptional suppression is generally undetectable, being effectively compensated by the gene-by-gene evolutionary recruitment of strong promoters on the X chromosome. We identify and experimentally validate a promoter element sequence motif that is enriched upstream of the transcription start sites of hundreds of testis-expressed genes; evolutionarily conserved across species; associated with strong gene expression levels in testes; and overrepresented on the X chromosome. These findings show that the expression of X-linked genes in the Drosophila testes reflects a balance between chromosome-wide epigenetic transcriptional suppression and long-term compensatory adaptation by sex-linked genes. Our results have broad implications for the evolution of gene expression in the Drosophila male germline and for genome evolution.

  5. 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.

  6. Exact p-value calculation for heterotypic clusters of regulatory motifs and its application in computational annotation of cis-regulatory modules

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    Roytberg Mikhail A

    2007-10-01

    Full Text Available Abstract Background cis-Regulatory modules (CRMs of eukaryotic genes often contain multiple binding sites for transcription factors. The phenomenon that binding sites form clusters in CRMs is exploited in many algorithms to locate CRMs in a genome. This gives rise to the problem of calculating the statistical significance of the event that multiple sites, recognized by different factors, would be found simultaneously in a text of a fixed length. The main difficulty comes from overlapping occurrences of motifs. So far, no tools have been developed allowing the computation of p-values for simultaneous occurrences of different motifs which can overlap. Results We developed and implemented an algorithm computing the p-value that s different motifs occur respectively k1, ..., ks or more times, possibly overlapping, in a random text. Motifs can be represented with a majority of popular motif models, but in all cases, without indels. Zero or first order Markov chains can be adopted as a model for the random text. The computational tool was tested on the set of cis-regulatory modules involved in D. melanogaster early development, for which there exists an annotation of binding sites for transcription factors. Our test allowed us to correctly identify transcription factors cooperatively/competitively binding to DNA. Method The algorithm that precisely computes the probability of simultaneous motif occurrences is inspired by the Aho-Corasick automaton and employs a prefix tree together with a transition function. The algorithm runs with the O(n|Σ|(m|ℋ MathType@MTEF@5@5@+=feaafiart1ev1aaatCvAUfKttLearuWrP9MDH5MBPbIqV92AaeXatLxBI9gBaebbnrfifHhDYfgasaacH8akY=wiFfYdH8Gipec8Eeeu0xXdbba9frFj0=OqFfea0dXdd9vqai=hGuQ8kuc9pgc9s8qqaq=dirpe0xb9q8qiLsFr0=vr0=vr0dc8meaabaqaciaacaGaaeqabaqabeGadaaakeaat0uy0HwzTfgDPnwy1egaryqtHrhAL1wy0L2yHvdaiqaacqWFlecsaaa@3762@| + K|σ|K ∏i ki time complexity, where n is the length of the text, |Σ| is the alphabet size, m is the

  7. A variant in a Cis-regulatory element enhances claudin-14 expression and is associated with pediatric-onset hypercalciuria and kidney stones

    DEFF Research Database (Denmark)

    Ure, Megan E; Heydari, Emma; Pan, Wanling

    2017-01-01

    The greatest risk factor for kidney stones is hypercalciuria, the etiology of which is largely unknown. A recent genome-wide association study (GWAS) linked hypercalciuria and kidney stones to a claudin-14 (CLDN14) risk haplotype. However, the underlying molecular mechanism was not delineated. Re...

  8. The disruption of a novel limb cis-regulatory element of SHH is associated with autosomal dominant preaxial polydactyly-hypertrichosis.

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    Petit, Florence; Jourdain, Anne-Sophie; Holder-Espinasse, Muriel; Keren, Boris; Andrieux, Joris; Duterque-Coquillaud, Martine; Porchet, Nicole; Manouvrier-Hanu, Sylvie; Escande, Fabienne

    2016-01-01

    The expression gradient of the morphogen Sonic Hedgehog (SHH) is crucial in establishing the number and the identity of the digits during anteroposterior patterning of the limb. Its anterior ectopic expression is responsible for preaxial polydactyly (PPD). Most of these malformations are due to the gain-of-function of the Zone of Polarizing Activity Regulatory Sequence, the only limb-specific enhancer of SHH known to date. We report a family affected with a novel condition associating PPD and hypertrichosis of the upper back, following an autosomal dominant mode of inheritance. This phenotype is consistent with deregulation of SHH expression during limb and follicle development. In affected members, we identified a 2 kb deletion located ~240 kb upstream from the SHH promoter. The deleted sequence is capable of repressing the transcriptional activity of the SHH promoter in vitro, consistent with a silencer activity. We hypothesize that the deletion of this silencer could be responsible for SHH deregulation during development, leading to a PPD-hypertrichosis phenotype.

  9. Arabidopsis Flower and Embryo Developmental Genes are Repressed in Seedlings by Different Combinations of Polycomb Group Proteins in Association with Distinct Sets of Cis-regulatory Elements.

    Science.gov (United States)

    Wang, Hua; Liu, Chunmei; Cheng, Jingfei; Liu, Jian; Zhang, Lei; He, Chongsheng; Shen, Wen-Hui; Jin, Hong; Xu, Lin; Zhang, Yijing

    2016-01-01

    Polycomb repressive complexes (PRCs) play crucial roles in transcriptional repression and developmental regulation in both plants and animals. In plants, depletion of different members of PRCs causes both overlapping and unique phenotypic defects. However, the underlying molecular mechanism determining the target specificity and functional diversity is not sufficiently characterized. Here, we quantitatively compared changes of tri-methylation at H3K27 in Arabidopsis mutants deprived of various key PRC components. We show that CURLY LEAF (CLF), a major catalytic subunit of PRC2, coordinates with different members of PRC1 in suppression of distinct plant developmental programs. We found that expression of flower development genes is repressed in seedlings preferentially via non-redundant role of CLF, which specifically associated with LIKE HETEROCHROMATIN PROTEIN1 (LHP1). In contrast, expression of embryo development genes is repressed by PRC1-catalytic core subunits AtBMI1 and AtRING1 in common with PRC2-catalytic enzymes CLF or SWINGER (SWN). This context-dependent role of CLF corresponds well with the change in H3K27me3 profiles, and is remarkably associated with differential co-occupancy of binding motifs of transcription factors (TFs), including MADS box and ABA-related factors. We propose that different combinations of PRC members distinctively regulate different developmental programs, and their target specificity is modulated by specific TFs.

  10. Arabidopsis Flower and Embryo Developmental Genes are Repressed in Seedlings by Different Combinations of Polycomb Group Proteins in Association with Distinct Sets of Cis-regulatory Elements.

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    Hua Wang

    2016-01-01

    Full Text Available Polycomb repressive complexes (PRCs play crucial roles in transcriptional repression and developmental regulation in both plants and animals. In plants, depletion of different members of PRCs causes both overlapping and unique phenotypic defects. However, the underlying molecular mechanism determining the target specificity and functional diversity is not sufficiently characterized. Here, we quantitatively compared changes of tri-methylation at H3K27 in Arabidopsis mutants deprived of various key PRC components. We show that CURLY LEAF (CLF, a major catalytic subunit of PRC2, coordinates with different members of PRC1 in suppression of distinct plant developmental programs. We found that expression of flower development genes is repressed in seedlings preferentially via non-redundant role of CLF, which specifically associated with LIKE HETEROCHROMATIN PROTEIN1 (LHP1. In contrast, expression of embryo development genes is repressed by PRC1-catalytic core subunits AtBMI1 and AtRING1 in common with PRC2-catalytic enzymes CLF or SWINGER (SWN. This context-dependent role of CLF corresponds well with the change in H3K27me3 profiles, and is remarkably associated with differential co-occupancy of binding motifs of transcription factors (TFs, including MADS box and ABA-related factors. We propose that different combinations of PRC members distinctively regulate different developmental programs, and their target specificity is modulated by specific TFs.

  11. 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

    Identification of cis-regulatory elements in DNA promoters by computational approaches are a great help in genome-wide elucidation of signal transduction, transcriptional regulation and gene expression. However, to accomplish this has proven a difficult problem in computational genomics in part d...

  12. Analyses of fugu hoxa2 genes provide evidence for subfunctionalization of neural crest cell and rhombomere cis-regulatory modules during vertebrate evolution.

    Science.gov (United States)

    McEllin, Jennifer A; Alexander, Tara B; Tümpel, Stefan; Wiedemann, Leanne M; Krumlauf, Robb

    2016-01-15

    Hoxa2 gene is a primary player in regulation of craniofacial programs of head development in vertebrates. Here we investigate the evolution of a Hoxa2 neural crest enhancer identified originally in mouse by comparing and contrasting the fugu hoxa2a and hoxa2b genes with their orthologous teleost and mammalian sequences. Using sequence analyses in combination with transgenic regulatory assays in zebrafish and mouse embryos we demonstrate subfunctionalization of regulatory activity for expression in hindbrain segments and neural crest cells between these two fugu co-orthologs. hoxa2a regulatory sequences have retained the ability to mediate expression in neural crest cells while those of hoxa2b include cis-elements that direct expression in rhombomeres. Functional dissection of the neural crest regulatory potential of the fugu hoxa2a and hoxa2b genes identify the previously unknown cis-element NC5, which is implicated in generating the differential activity of the enhancers from these genes. The NC5 region plays a similar role in the ability of this enhancer to mediate reporter expression in mice, suggesting it is a conserved component involved in control of neural crest expression of Hoxa2 in vertebrate craniofacial development.

  13. 3'-coterminal subgenomic RNAs and putative cis-acting elements of Grapevine leafroll-associated virus 3 reveals 'unique' features of gene expression strategy in the genus Ampelovirus

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    Dawson William O

    2010-08-01

    Full Text Available Abstract Background The family Closteroviridae comprises genera with monopartite genomes, Closterovirus and Ampelovirus, and with bipartite and tripartite genomes, Crinivirus. By contrast to closteroviruses in the genera Closterovirus and Crinivirus, much less is known about the molecular biology of viruses in the genus Ampelovirus, although they cause serious diseases in agriculturally important perennial crops like grapevines, pineapple, cherries and plums. Results The gene expression and cis-acting elements of Grapevine leafroll-associated virus 3 (GLRaV-3; genus Ampelovirus was examined and compared to that of other members of the family Closteroviridae. Six putative 3'-coterminal subgenomic (sg RNAs were abundantly present in grapevine (Vitis vinifera infected with GLRaV-3. The sgRNAs for coat protein (CP, p21, p20A and p20B were confirmed using gene-specific riboprobes in Northern blot analysis. The 5'-termini of sgRNAs specific to CP, p21, p20A and p20B were mapped in the 18,498 nucleotide (nt virus genome and their leader sequences determined to be 48, 23, 95 and 125 nt, respectively. No conserved motifs were found around the transcription start site or in the leader sequence of these sgRNAs. The predicted secondary structure analysis of sequences around the start site failed to reveal any conserved motifs among the four sgRNAs. The GLRaV-3 isolate from Washington had a 737 nt long 5' nontranslated region (NTR with a tandem repeat of 65 nt sequence and differed in sequence and predicted secondary structure with a South Africa isolate. Comparison of the dissimilar sequences of the 5'NTRs did not reveal any common predicted structures. The 3'NTR was shorter and more conserved. The lack of similarity among the cis-acting elements of the diverse viruses in the family Closteroviridae is another measure of the complexity of their evolution. Conclusions The results indicate that transcription regulation of GLRaV-3 sgRNAs appears to be different

  14. The nuclear OXPHOS genes in insecta: a common evolutionary origin, a common cis-regulatory motif, a common destiny for gene duplicates

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    Pesole Graziano

    2007-11-01

    Full Text Available Abstract Background When orthologous sequences from species distributed throughout an optimal range of divergence times are available, comparative genomics is a powerful tool to address problems such as the identification of the forces that shape gene structure during evolution, although the functional constraints involved may vary in different genes and lineages. Results We identified and annotated in the MitoComp2 dataset the orthologs of 68 nuclear genes controlling oxidative phosphorylation in 11 Drosophilidae species and in five non-Drosophilidae insects, and compared them with each other and with their counterparts in three vertebrates (Fugu rubripes, Danio rerio and Homo sapiens and in the cnidarian Nematostella vectensis, taking into account conservation of gene structure and regulatory motifs, and preservation of gene paralogs in the genome. Comparative analysis indicates that the ancestral insect OXPHOS genes were intron rich and that extensive intron loss and lineage-specific intron gain occurred during evolution. Comparison with vertebrates and cnidarians also shows that many OXPHOS gene introns predate the cnidarian/Bilateria evolutionary split. The nuclear respiratory gene element (NRG has played a key role in the evolution of the insect OXPHOS genes; it is constantly conserved in the OXPHOS orthologs of all the insect species examined, while their duplicates either completely lack the element or possess only relics of the motif. Conclusion Our observations reinforce the notion that the common ancestor of most animal phyla had intron-rich gene, and suggest that changes in the pattern of expression of the gene facilitate the fixation of duplications in the genome and the development of novel genetic functions.

  15. Integration of bioinformatics and synthetic promoters leads to the discovery of novel elicitor-responsive cis-regulatory sequences in Arabidopsis.

    Science.gov (United States)

    Koschmann, Jeannette; Machens, Fabian; Becker, Marlies; Niemeyer, Julia; Schulze, Jutta; Bülow, Lorenz; Stahl, Dietmar J; Hehl, Reinhard

    2012-09-01

    A combination of bioinformatic tools, high-throughput gene expression profiles, and the use of synthetic promoters is a powerful approach to discover and evaluate novel cis-sequences in response to specific stimuli. With Arabidopsis (Arabidopsis thaliana) microarray data annotated to the PathoPlant database, 732 different queries with a focus on fungal and oomycete pathogens were performed, leading to 510 up-regulated gene groups. Using the binding site estimation suite of tools, BEST, 407 conserved sequence motifs were identified in promoter regions of these coregulated gene sets. Motif similarities were determined with STAMP, classifying the 407 sequence motifs into 37 families. A comparative analysis of these 37 families with the AthaMap, PLACE, and AGRIS databases revealed similarities to known cis-elements but also led to the discovery of cis-sequences not yet implicated in pathogen response. Using a parsley (Petroselinum crispum) protoplast system and a modified reporter gene vector with an internal transformation control, 25 elicitor-responsive cis-sequences from 10 different motif families were identified. Many of the elicitor-responsive cis-sequences also drive reporter gene expression in an Agrobacterium tumefaciens infection assay in Nicotiana benthamiana. This work significantly increases the number of known elicitor-responsive cis-sequences and demonstrates the successful integration of a diverse set of bioinformatic resources combined with synthetic promoter analysis for data mining and functional screening in plant-pathogen interaction.

  16. Gene expression profiling of cultured human NF1 heterozygous (NF1+/-) melanocytes reveals downregulation of a transcriptional cis-regulatory network mediating activation of the melanocyte-specific dopachrome tautomerase (DCT) gene.

    Science.gov (United States)

    Boucneau, Joachim; De Schepper, Sofie; Vuylsteke, Marnik; Van Hummelen, Paul; Naeyaert, Jean-Marie; Lambert, Jo

    2005-08-01

    One of the major primary features of the neurocutaneous genetic disorder Neurofibromatosis type 1 are the hyperpigmentary café-au-lait macules where disregulation of melanocyte biology is supposed to play a key etiopathogenic role. To gain better insight into the possible role of the tumor suppressor gene NF1, a transcriptomic microarray analysis was performed on human NF1 heterozygous (NF1+/-) melanocytes of a Neurofibromatosis type 1 patient and NF1 wild type (NF1+/+) melanocytes of a healthy control patient, both cultured from normally pigmented skin and hyperpigmented lesional café-au-lait skin. From the magnitude of gene effects, we found that gene expression was affected most strongly by genotype and less so by lesional type. A total of 137 genes had a significant twofold or more up- (72) or downregulated (65) expression in NF1+/- melanocytes compared with NF1+/+ melanocytes. Melanocytes cultured from hyperpigmented café-au-lait skin showed 37 upregulated genes whereas only 14 were downregulated compared with normal skin melanocytes. In addition, significant genotype xlesional type interactions were observed for 465 genes. Differentially expressed genes were mainly involved in regulating cell proliferation and cell adhesion. A high number of transcription factor genes, among which a specific subset important in melanocyte lineage development, were downregulated in the cis-regulatory network governing the activation of the melanocyte-specific dopachrome tautomerase (DCT) gene. Although the results presented have been obtained with a restricted number of patients (one NF1 patient and one control) and using cDNA microarrays that may limit their interpretation, the data nevertheless addresses for the first time the effect of a heterozygous NF1 gene on the expression of the human melanocyte transcriptome and has generated several interesting candidate genes helpful in elucidating the etiopathology of café-au-lait macules in NF1 patients.

  17. 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. PMID:28005945

  18. High-resolution analysis of four efficient yeast replication origins reveals new insights into the ORC and putative MCM binding elements.

    Science.gov (United States)

    Chang, Fujung; May, Caitlin D; Hoggard, Timothy; Miller, Jeremy; Fox, Catherine A; Weinreich, Michael

    2011-08-01

    In budding yeast, the eukaryotic initiator protein ORC (origin recognition complex) binds to a bipartite sequence consisting of an 11 bp ACS element and an adjacent B1 element. However, the genome contains many more matches to this consensus than actually bind ORC or function as origins in vivo. Although ORC-dependent loading of the replicative MCM helicase at origins is enhanced by a distal B2 element, less is known about this element. Here, we analyzed four highly active origins (ARS309, ARS319, ARS606 and ARS607) by linker scanning mutagenesis and found that sequences adjacent to the ACS contributed substantially to origin activity and ORC binding. Using the sequences of four additional B2 elements we generated a B2 multiple sequence alignment and identified a shared, degenerate 8 bp sequence that was enriched within 228 known origins. In addition, our high-resolution analysis revealed that not all origins exist within nucleosome free regions: a class of Sir2-regulated origins has a stably positioned nucleosome overlapping or near B2. This study illustrates the conserved yet flexible nature of yeast origin architecture to promote ORC binding and origin activity, and helps explain why a strong match to the ORC binding site is insufficient to identify origins within the genome.

  19. A Novel Peroxisome Proliferator Response Element Modulates Hepatic Low Density Lipoprotein Receptor Gene Transcription in Response to PPARδ Activation

    Science.gov (United States)

    Shende, Vikram R.; Singh, Amar Bahadur; Liu, Jingwen

    2016-01-01

    The hepatic expression of LDLR gene is regulated primarily at the transcriptional level by a sterol-regulatory element (SRE) in its proximal promoter region which is the site of action of SRE-binding protein 2 (SREBP2). However whether additional cis-regulatory elements contribute to LDLR transcription has not been fully explored. We investigated the function of a putative PPAR-response element (PPRE) sequence motif located at −768 to −752 bases upstream of the transcription start site of human LDLR gene in response to PPARδ activation. Promoter luciferase reporter analyses showed that treating HepG2 cells with PPARδ agonist L165041 markedly increased the activity of a full-length LDLR promoter construct (pLDLR-1192) without any effects on the shorter promoter reporter pLDLR-234 that contains only the core regulatory elements SRE-1 and SP1 sites. Importantly, mutation of the PPRE sequence greatly attenuated the induction of the full-length LDLR promoter activity by L165041 without affecting rosuvastatin mediated transactivation. Electrophoretic mobility shift and chromatin immunoprecipitation assays further confirmed the binding of PPARδ to the LDLR-PPRE site. Treating HepG2 cells with L165041 elevated the mRNA and protein expressions of LDLR without affecting the LDLR mRNA decay rate. The induction of LDLR expression by PPARδ agonist was further observed in liver tissue of mice and hamsters treated with L165041. Altogether, our studies identify a novel PPRE-mediated regulatory mechanism for LDLR transcription and suggest that combined treatment of statin with PPARδ agonists may have advantageous effects on LDLR expression. PMID:26443862

  20. A novel peroxisome proliferator response element modulates hepatic low-density lipoprotein receptor gene transcription in response to PPARδ activation.

    Science.gov (United States)

    Shende, Vikram R; Singh, Amar Bahadur; Liu, Jingwen

    2015-12-15

    The hepatic expression of low-density lipoprotein (LDL) receptor (LDLR) gene is regulated primarily at the transcriptional level by a sterol-regulatory element (SRE) in its proximal promoter region which is the site of action of SRE-binding protein 2 (SREBP2). However whether additional cis-regulatory elements contribute to LDLR transcription has not been fully explored. We investigated the function of a putative peroxisome proliferator-activated receptor (PPAR)-response element (PPRE) sequence motif located at -768 to -752 bases upstream of the transcription start site of human LDLR gene in response to PPARδ activation. Promoter luciferase reporter analyses showed that treating HepG2 cells with PPARδ agonist L165041 markedly increased the activity of a full-length LDLR promoter construct (pLDLR-1192) without any effects on the shorter promoter reporter pLDLR-234 that contains only the core regulatory elements SRE-1 and SP1 sites. Importantly, mutation of the PPRE sequence greatly attenuated the induction of the full-length LDLR promoter activity by L165041 without affecting rosuvastatin (RSV)-mediated transactivation. EMSA and ChIP assay further confirmed the binding of PPARδ to the LDLR-PPRE site. Treating HepG2 cells with L165041 elevated the mRNA and protein expressions of LDLR without affecting the LDLR mRNA decay rate. The induction of LDLR expression by PPARδ agonist was further observed in liver tissue of mice and hamsters treated with L165041. Altogether, our studies identify a novel PPRE-mediated regulatory mechanism for LDLR transcription and suggest that combined treatment of statin with PPARδ agonists may have advantageous effects on LDLR expression.

  1. Genome-wide prediction of transcriptional regulatory elements of human promoters using gene expression and promoter analysis data

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    Kim Seon-Young

    2006-07-01

    Full Text Available Abstract Background A complete understanding of the regulatory mechanisms of gene expression is the next important issue of genomics. Many bioinformaticians have developed methods and algorithms for predicting transcriptional regulatory mechanisms from sequence, gene expression, and binding data. However, most of these studies involved the use of yeast which has much simpler regulatory networks than human and has many genome wide binding data and gene expression data under diverse conditions. Studies of genome wide transcriptional networks of human genomes currently lag behind those of yeast. Results We report herein a new method that combines gene expression data analysis with promoter analysis to infer transcriptional regulatory elements of human genes. The Z scores from the application of gene set analysis with gene sets of transcription factor binding sites (TFBSs were successfully used to represent the activity of TFBSs in a given microarray data set. A significant correlation between the Z scores of gene sets of TFBSs and individual genes across multiple conditions permitted successful identification of many known human transcriptional regulatory elements of genes as well as the prediction of numerous putative TFBSs of many genes which will constitute a good starting point for further experiments. Using Z scores of gene sets of TFBSs produced better predictions than the use of mRNA levels of a transcription factor itself, suggesting that the Z scores of gene sets of TFBSs better represent diverse mechanisms for changing the activity of transcription factors in the cell. In addition, cis-regulatory modules, combinations of co-acting TFBSs, were readily identified by our analysis. Conclusion By a strategic combination of gene set level analysis of gene expression data sets and promoter analysis, we were able to identify and predict many transcriptional regulatory elements of human genes. We conclude that this approach will aid in decoding

  2. Mutations in CG8878, a Novel Putative Protein Kinase, Enhance P Element Dependent Silencing (PDS) and Position Effect Variegation (PEV) in Drosophila melanogaster

    Science.gov (United States)

    McCracken, Allen; Locke, John

    2014-01-01

    Genes in multicellular organisms are expressed as part of a developmental program that is largely dependent on self-perpetuating higher-order chromatin states. The mechanism of establishing and maintaining these epigenetic events is well studied in Drosophila. The first known example of an epigenetic effect was that of (PEV) in Drosophila, which has been shown to be due to gene silencing via heterochromatin formation. We are investigating a process similar to Position Effect Variegation (PEV) using a mini-w transgene, called Pci, inserted in the upstream regulatory region of ci. The mini-white+ transgene in Pci is expressed throughout the adult eye; however, when other P or KP elements are present, a variegated eye phenotype results indicating random w+ silencing during development. This P element dependent silencing (PDS) can be modified by the haplo-suppressors/triplo-enhancers, Su(var)205 and Su(var)3–7, indicating that these heterochromatic modifiers also act dose dependently in PDS. Here we use a spontaneous derivative mutation of Pci called PciE1 (E1) that variegates like PDS in the absence of P elements, presumably due to an adjacent gypsy element insertion, to screen for second-site modifier mutations that enhance variable silencing of white+ in E1. We isolated 7 mutations in CG8878, an essential gene, that enhance the E1 variegated phenotype. CG8878, a previously uncharacterized gene, potentially encodes a serine/threonine kinase whose closest Drosophila paralogue, ballchen (nhk-1), phosphorylates histones. These mutant alleles enhance both PDS at E1 and Position Effect Variegation (PEV) at wm4, indicating a previously unknown common silencing mechanism between the two. PMID:24614804

  3. Mutations in CG8878, a novel putative protein kinase, enhance P element dependent silencing (PDS and position effect variegation (PEV in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Allen McCracken

    Full Text Available Genes in multicellular organisms are expressed as part of a developmental program that is largely dependent on self-perpetuating higher-order chromatin states. The mechanism of establishing and maintaining these epigenetic events is well studied in Drosophila. The first known example of an epigenetic effect was that of (PEV in Drosophila, which has been shown to be due to gene silencing via heterochromatin formation. We are investigating a process similar to Position Effect Variegation (PEV using a mini-w transgene, called Pci, inserted in the upstream regulatory region of ci. The mini-white+ transgene in Pci is expressed throughout the adult eye; however, when other P or KP elements are present, a variegated eye phenotype results indicating random w+ silencing during development. This P element dependent silencing (PDS can be modified by the haplo-suppressors/triplo-enhancers, Su(var205 and Su(var3-7, indicating that these heterochromatic modifiers also act dose dependently in PDS. Here we use a spontaneous derivative mutation of Pci called PciE1 (E1 that variegates like PDS in the absence of P elements, presumably due to an adjacent gypsy element insertion, to screen for second-site modifier mutations that enhance variable silencing of white+ in E1. We isolated 7 mutations in CG8878, an essential gene, that enhance the E1 variegated phenotype. CG8878, a previously uncharacterized gene, potentially encodes a serine/threonine kinase whose closest Drosophila paralogue, ballchen (nhk-1, phosphorylates histones. These mutant alleles enhance both PDS at E1 and Position Effect Variegation (PEV at w(m4, indicating a previously unknown common silencing mechanism between the two.

  4. A biophysical model for identifying splicing regulatory elements and their interactions.

    Directory of Open Access Journals (Sweden)

    Ji Wen

    Full Text Available Alternative splicing (AS of precursor mRNA (pre-mRNA is a crucial step in the expression of most eukaryotic genes. Splicing factors (SFs play an important role in AS regulation by binding to the cis-regulatory elements on the pre-mRNA. Although many splicing factors (SFs and their binding sites have been identified, their combinatorial regulatory effects remain to be elucidated. In this paper, we derive a biophysical model for AS regulation that integrates combinatorial signals of cis-acting splicing regulatory elements (SREs and their interactions. We also develop a systematic framework for model inference. Applying the biophysical model to a human RNA-Seq data set, we demonstrate that our model can explain 49.1%-66.5% variance of the data, which is comparable to the best result achieved by biophysical models for transcription. In total, we identified 119 SRE pairs between different regions of cassette exons that may regulate exon or intron definition in splicing, and 77 SRE pairs from the same region that may arise from a long motif or two different SREs bound by different SFs. Particularly, putative binding sites of polypyrimidine tract-binding protein (PTB, heterogeneous nuclear ribonucleoprotein (hnRNP F/H and E/K are identified as interacting SRE pairs, and have been shown to be consistent with the interaction models proposed in previous experimental results. These results show that our biophysical model and inference method provide a means of quantitative modeling of splicing regulation and is a useful tool for identifying SREs and their interactions. The software package for model inference is available under an open source license.

  5. Highly conserved elements discovered in vertebrates are present in non-syntenic loci of tunicates, act as enhancers and can be transcribed during development

    Science.gov (United States)

    Sanges, Remo; Hadzhiev, Yavor; Gueroult-Bellone, Marion; Roure, Agnes; Ferg, Marco; Meola, Nicola; Amore, Gabriele; Basu, Swaraj; Brown, Euan R.; De Simone, Marco; Petrera, Francesca; Licastro, Danilo; Strähle, Uwe; Banfi, Sandro; Lemaire, Patrick; Birney, Ewan; Müller, Ferenc; Stupka, Elia

    2013-01-01

    Co-option of cis-regulatory modules has been suggested as a mechanism for the evolution of expression sites during development. However, the extent and mechanisms involved in mobilization of cis-regulatory modules remains elusive. To trace the history of non-coding elements, which may represent candidate ancestral cis-regulatory modules affirmed during chordate evolution, we have searched for conserved elements in tunicate and vertebrate (Olfactores) genomes. We identified, for the first time, 183 non-coding sequences that are highly conserved between the two groups. Our results show that all but one element are conserved in non-syntenic regions between vertebrate and tunicate genomes, while being syntenic among vertebrates. Nevertheless, in all the groups, they are significantly associated with transcription factors showing specific functions fundamental to animal development, such as multicellular organism development and sequence-specific DNA binding. The majority of these regions map onto ultraconserved elements and we demonstrate that they can act as functional enhancers within the organism of origin, as well as in cross-transgenesis experiments, and that they are transcribed in extant species of Olfactores. We refer to the elements as ‘Olfactores conserved non-coding elements’. PMID:23393190

  6. V(D)J recombination frequency is affected by the sequence interposed between a pair of recombination signals: sequence comparison reveals a putative recombinational enhancer element.

    Science.gov (United States)

    Roch, F A; Hobi, R; Berchtold, M W; Kuenzle, C C

    1997-06-15

    The immunoglobulin heavy chain intron enhancer (Emu) not only stimulates transcription but also V(D)J recombination of chromosomally integrated recombination substrates. We aimed at reproducing this effect in recombination competent cells by transient transfection of extrachromosomal substrates. These we prepared by interposing between the recombination signal sequences (RSS) of the plasmid pBlueRec various fragments, including Emu, possibly affecting V(D)J recombination. Our work shows that sequences inserted between RSS 23 and RSS 12, with distances from their proximal ends of 26 and 284 bp respectively, can markedly affect the frequency of V(D)J recombination. We report that the entire Emu, the Emu core as well as its flanking 5' and 3' matrix associated regions (5' and 3' MARs) upregulate V(D)J recombination while the downstream section of the 3' MAR of Emu does not. Also, prokaryotic sequences markedly suppress V(D)J recombination. This confirms previous results obtained with chromosomally integrated substrates, except for the finding that the full length 3' MAR of Emu stimulates V(D)J recombination in an episomal but not in a chromosomal context. The fact that other MARs do not share this activity suggests that the effect is no mediated through attachment of the recombination substrate to a nuclear matrix-associated recombination complex but through cis-activation. The presence of a 26 bp A-T-rich sequence motif in the 5' and 3' MARs of Emu and in all of the other upregulating fragments investigated, leads us to propose that the motif represents a novel recombinational enhancer element distinct from those constituting the Emu core.

  7. An adaptive transposable element insertion in the regulatory region of the EO gene in the domesticated silkworm, Bombyx mori.

    Science.gov (United States)

    Sun, Wei; Shen, Yi-Hong; Han, Min-Jin; Cao, Yun-Feng; Zhang, Ze

    2014-12-01

    Although there are many studies to show a key role of transposable elements (TEs) in adaptive evolution of higher organisms, little is known about the molecular mechanisms. In this study, we found that a partial TE (Taguchi) inserted in the cis-regulatory region of the silkworm ecdysone oxidase (EO) gene, which encodes a crucial enzyme to reduce the titer of molting hormone (20-hydroxyecdysone, 20E). The TE insertion occurred during domestication of silkworm and the frequency of the TE insertion in the domesticated silkworm (Bombyx mori) is high, 54.24%. The linkage disequilibrium in the TE inserted strains of the domesticated silkworm was elevated. Molecular population genetics analyses suggest that this TE insertion is adaptive for the domesticated silkworm. Luminescent reporter assay shows that the TE inserted in the cis-regulatory region of the EO gene functions as a 20E-induced enhancer of the gene expression. Further, phenotypic bioassay indicates that the silkworm with the TE insertion exhibited more stable developmental phenotype than the silkworm without the TE insertion when suffering from food shortage. Thus, the inserted TE in the cis-regulatory region of the EO gene increased developmental uniformity of silkworm individuals through regulating 20E metabolism, partially explaining transformation of a domestication developmental trait in the domesticated silkworm. Our results emphasize the exceptional role of gene expression regulation in developmental transition of domesticated animals.

  8. Close Sequence Comparisons are Sufficient to Identify Humancis-Regulatory Elements

    Energy Technology Data Exchange (ETDEWEB)

    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.

  9. Conserved Noncoding Elements in the Most Distant Genera of Cephalochordates: The Goldilocks Principle

    Science.gov (United States)

    Yue, Jia-Xing; Kozmikova, Iryna; Ono, Hiroki; Nossa, Carlos W.; Kozmik, Zbynek; Putnam, Nicholas H.; Yu, Jr-Kai; Holland, Linda Z.

    2016-01-01

    Cephalochordates, the sister group of vertebrates + tunicates, are evolving particularly slowly. Therefore, genome comparisons between two congeners of Branchiostoma revealed so many conserved noncoding elements (CNEs), that it was not clear how many are functional regulatory elements. To more effectively identify CNEs with potential regulatory functions, we compared noncoding sequences of genomes of the most phylogenetically distant cephalochordate genera, Asymmetron and Branchiostoma, which diverged approximately 120–160 million years ago. We found 113,070 noncoding elements conserved between the two species, amounting to 3.3% of the genome. The genomic distribution, target gene ontology, and enriched motifs of these CNEs all suggest that many of them are probably cis-regulatory elements. More than 90% of previously verified amphioxus regulatory elements were re-captured in this study. A search of the cephalochordate CNEs around 50 developmental genes in several vertebrate genomes revealed eight CNEs conserved between cephalochordates and vertebrates, indicating sequence conservation over >500 million years of divergence. The function of five CNEs was tested in reporter assays in zebrafish, and one was also tested in amphioxus. All five CNEs proved to be tissue-specific enhancers. Taken together, these findings indicate that even though Branchiostoma and Asymmetron are distantly related, as they are evolving slowly, comparisons between them are likely optimal for identifying most of their tissue-specific cis-regulatory elements laying the foundation for functional characterizations and a better understanding of the evolution of developmental regulation in cephalochordates. PMID:27412606

  10. Patterns of cis regulatory variation in diverse human populations.

    Directory of Open Access Journals (Sweden)

    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 the transferability of complex trait variants across populations.

  11. Computational methods to dissect cis-regulatory transcriptional networks

    Indian Academy of Sciences (India)

    Vibha Rani

    2007-12-01

    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 deciphering the regulation of metazoan genes as well as interpreting the results of high-throughput screening. The integration of computer science with biology has expedited molecular modelling and processing of large-scale data inputs such as microarrays, analysis of genomes, transcriptomes and proteomes. Many bioinformaticians have developed various algorithms for predicting transcriptional regulatory mechanisms from the sequence, gene expression and interaction data. This review contains compiled information of various computational methods adopted to dissect gene expression pathways.

  12. Patterns of cis regulatory variation in diverse human populations.

    Directory of Open Access Journals (Sweden)

    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 the transferability of complex trait variants across populations.

  13. Short 5'-flanking regions of the Amy gene of Drosophila kikkawai affect amylase gene expression and respond to food environments.

    Science.gov (United States)

    Inomata, Nobuyuki; Nakashima, Shuichi

    2008-04-15

    Evolution of the duplicated genes and regulation in gene expression is of great interest, especially in terms of adaptation. Molecular population genetic and evolutionary studies on the duplicated amylase genes of Drosophila species have suggested that their 5'-flanking (cis-regulatory) regions play an important role in evolution of these genes. For better understanding of evolution of the duplicated amylase genes and gene expression, we studied functional significance of the Amy1 gene of Drosophila kikkawai using in vitro deletion mutagenesis followed by P-element-mediated germline transformation. We found that a 1.6-kb of the 5'-flanking region can produce strikingly higher level of larval amylase activity on starch food compared with that on glucose food. We found two cis-regulatory elements, which increase larval amylase activity on starch food. We also found a larval cis-regulatory element, which responds to the food difference. This food-response element is necessary for the function of the element increasing larval activity on starch food. A 5-bp deletion in a putative GRE caused high amylase activity, indicating a cis-regulatory element decreasing amylase activity. These cis-regulatory elements identified in the 5'-flanking region could be the targets of natural selection.

  14. Transcriptional Regulation of Hhex in Hematopoiesis and Hematopoietic Stem Cell Ontogeny

    DEFF Research Database (Denmark)

    Portero Migueles, Rosa; Shaw, Louise; Rodrigues, Neil P

    2017-01-01

    in endothelium of the dorsal aorta (DA) and in clusters of putative HSCs as they are specified during murine development. We exploited this observation, using the Hhex locus to define cis regulatory elements, enhancers and interacting transcription factors that are both necessary and sufficient to support gene...

  15. Phylogeny based discovery of regulatory elements

    Directory of Open Access Journals (Sweden)

    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.

  16. Identification of gene co-regulatory modules and associated cis-elements involved in degenerative heart disease

    Directory of Open Access Journals (Sweden)

    Danko Charles G

    2009-05-01

    odds ratios of 2.7 (contractile, 1.9 (energy generation, and 5.5 (protein translation. Conclusion We identified patterns comprised of putative cis-regulatory motifs enriched in the upstream promoter sequence of genes that undergo similar changes in expression secondary to cardiomyopathies of various etiologies. Our analysis is a first step towards understanding transcription factor networks that are active in regulating gene expression during degenerative heart disease.

  17. Recombinant Trichomonas vaginalis eIF-5A protein expressed from a eukaryotic system binds specifically to mammalian and putative trichomonal eIF-5A response elements (EREs).

    Science.gov (United States)

    Carvajal-Gamez, Bertha Isabel; Carrillo, Laura Vázquez; Torres-Romero, Julio César; Camacho-Nuez, Minerva; Ponce-Regalado, María Dolores; Camarillo, César López; Alvarez-Sánchez, María Elizbeth

    2016-12-01

    Trichomonas vaginalis eIF-5A-like protein (TveIF-5A) belongs to the highly conserved eIF-5A family of proteins that contains a unique polyamine-derived amino acid, hypusine. Recently, we determined that the polyamine putrescine is required for tveif-5a mRNA stability, and it is necessary for stability and maturation of the TveIF-5A protein. Eukaryotic eIF-5A is known to be involved in mRNA turnover and is capable of sequence-specific RNA binding to eIF-5A response elements (EREs). These ERE sequences are present in diverse mammalian mRNAs, including human cyclooxygenase-2 (cox-2). Here, we cloned the complete coding sequence of TveIF-5A and overexpressed it in a eukaryotic system. The recombinant protein (rTveIF-5A) was purified in soluble form using size-exclusion chromatography. Because of the polyamine-dependent regulation of TvCP39 (a protease of T. vaginalis) at the protein and RNA messenger (mRNA) levels, we looked for an ERE-like structure in the 3' region of tvcp39 mRNA. In RNA gel-shift assays, rTveIF-5A bound to transcripts at the EREs of cox-2 or tvcp39 mRNAs. This work shows the eIF-5A/ERE-like interaction in T. vaginalis.

  18. Amino acid transport in taxonomically diverse cyanobacteria and identification of two genes encoding elements of a neutral amino acid permease putatively involved in recapture of leaked hydrophobic amino acids.

    Science.gov (United States)

    Montesinos, M L; Herrero, A; Flores, E

    1997-02-01

    The activities of uptake of thirteen 14C-labeled amino acids were determined in nine cyanobacteria, including the unicellular strains Synechococcus sp. strain PCC 7942 and Synechocystis sp. strain PCC 6803; the filamentous strain Pseudanabaena sp. strain PCC 6903, and the filamentous, heterocyst-forming strains Anabaena sp. strains PCC 7120 and PCC 7937; Nostoc sp. strains PCC 7413 and PCC 7107; Calothrix sp. strain PCC 7601 (which is a mutant unable to develop heterocysts); and Fischerella muscicola UTEX 1829. Amino acid transport mutants, selected as mutants resistant to some amino acid analogs, were isolated from the Anabaena, Nostoc, Calothrix, and Pseudanabaena strains. All of the tested cyanobacteria bear at least a neutral amino acid transport system, and some strains also bear transport systems specific for basic or acidic amino acids. Two genes, natA and natB, encoding elements (conserved component, NatA, and periplasmic binding protein, NatB) of an ABC-type permease for neutral amino acids were identified by insertional mutagenesis of strain PCC 6803 open reading frames from the recently published genomic DNA sequence of this cyanobacterium. DNA sequences homologous to natA and natB from strain PCC 6803 were detected by hybridization in eight cyanobacterial strains tested. Mutants unable to transport neutral amino acids, including natA and natB insertional mutants, accumulated in the extracellular medium a set of amino acids that always included Ala, Val, Phe, Ile, and Leu. A general role for a cyanobacterial neutral amino acid permease in recapture of hydrophobic amino acids leaked from the cells is suggested.

  19. Multiple Regulatory Elements in the Arabidopsis NIA1 Promoter Act Synergistically to Form a Nitrate Enhancer1[W][OA

    Science.gov (United States)

    Wang, Rongchen; Guan, Peizhu; Chen, Mingsheng; Xing, Xiujuan; Zhang, Yali; Crawford, Nigel M.

    2010-01-01

    To accommodate fluctuating nutrient levels in the soil, plants modulate their metabolism and root development via signaling mechanisms that rapidly reprogram the plant transcriptome. In the case of nitrate, over 1,000 genes are induced or repressed within minutes of nitrate exposure. To identify cis-regulatory elements that mediate these responses, an enhancer screen was performed in transgenic Arabidopsis (Arabidopsis thaliana) plants. A 1.8-kb promoter fragment from the nitrate reductase gene NIA1 was identified that acts as a nitrate enhancer when fused to a 35S minimal promoter. Enhancer activity was localized to a 180-bp fragment, and this activity could be enhanced by the addition of a 131-bp fragment from the nitrite reductase promoter. A promoter construct containing the 180- and 131-bp fragments was also induced by nitrite and repressed by ammonium, indicating that it was responsive to multiple nitrogen signals. To identify specific regulatory elements within the 180-bp NIA1 fragment, a transient expression system using agroinfiltration of Nicotiana benthamiana was developed. Deletion analysis identified three elements corresponding to predicted binding motifs for homeodomain/E-box, Myb, and Alfin1 transcription factors. A fully active promoter showing nitrate and nitrite enhancer activity equivalent to that of the wild-type 180-bp fragment could be built from these three elements if the spacing between the homeodomain/E-box and Myb-Alfin1 sites was equivalent to that of the native promoter. These findings were validated in transgenic Arabidopsis plants and identify a cis-regulatory module containing three elements that comprise a nitrate enhancer in the NIA1 promoter. PMID:20668061

  20. Trypanosoma brucei: a putative RNA polymerase II promoter.

    Science.gov (United States)

    Bayele, Henry K

    2009-12-01

    RNA polymerase II (pol II) promoters are rare in the African trypanosome Trypanosoma brucei because gene regulation in the parasite is complex and polycistronic. Here, we describe a putative pol II promoter and its structure-function relationship. The promoter has features of an archetypal eukaryotic pol II promoter including putative canonical CCAAT and TATA boxes, and an initiator element. However, the spatial arrangement of these elements is only similar to yeast pol II promoters. Deletion mapping and transcription assays enabled delineation of a minimal promoter that could drive orientation-independent reporter gene expression suggesting that it may be a bidirectional promoter. In vitro transcription in a heterologous nuclear extract revealed that the promoter can be recognized by the basal eukaryotic transcription complex. This suggests that the transcription machinery in the parasite may be very similar to those of other eukaryotes.

  1. Transposable element recruitments in the mammalian placenta: impacts and mechanisms.

    Science.gov (United States)

    Emera, Deena; Wagner, Günter P

    2012-07-01

    Transposable elements (TEs) are mobile DNA elements found at high frequency in mammalian genomes. Although these elements are generally perceived as genomic parasites, they have the potential to influence host genome function in many beneficial ways. This article discusses the role TEs have played in the evolution of the placenta and pregnancy in viviparous mammals. Using examples from our own research and the literature, we argue that frequent recruitment of TEs, in particular of retroelements, has facilitated the extreme diversification of tissues at the maternal-fetal interface. We also discuss the mechanisms by which TEs have been recruited for functions during pregnancy. We argue that retroelements are pre-adapted to becoming cis-regulatory elements for host genomes because they need to utilize host regulatory signals for their own life cycle. However, although TEs contain some of the signals necessary for host functions upon insertion, they often require modification before acquiring a biological role in a host tissue. We discuss the process by which one TE was transformed into a promoter for prolactin expression in the endometrium, describing a model for TE domestication called 'epistatic capture'.

  2. Cloning of partial putative gonadotropin hormone receptor sequence from fish

    Indian Academy of Sciences (India)

    G Kumaresan; T Venugopal; A Vikas; T J Pandian; S M Athavan

    2000-03-01

    A search for the presence of mariner-like elements in the Labeo rohita genome by polymerase chain reaction led to the amplification of a partial DNA sequence coding for a putative transmembrane domain of gonadotropin hormone receptor. The amplified DNA sequence shows a high degree of homology to the available turkey and human luteinizing and follicle stimulating hormone receptor coding sequences. This is the first report on cloning such sequences of piscine origin.

  3. Wnt3a signal pathways activate MyoD expression by targeting cis-elements inside and outside its distal enhancer.

    Science.gov (United States)

    Pan, Yu Chih; Wang, Xiao Wen; Teng, Han Feng; Wu, Yi Ju; Chang, Hsuan Chia; Chen, Shen Liang

    2015-03-18

    Wnt proteins are secreted cytokines and several Wnts are expressed in the developing somites and surrounding tissues. Without proper Wnt stimulation, the organization of the dermomyotome and myotome can become defective. These Wnt signals received by somitic cells can lead to activation of Pax3/Pax7 and myogenic regulatory factors (MRFs), especially Myf5 and MyoD. However, it is currently unknown whether Wnts activate Myf5 and MyoD through direct targeting of their cis-regulatory elements or via indirect pathways. To clarify this issue, in the present study, we tested the regulation of MyoD cis-regulatory elements by Wnt3a secreted from human embryonic kidney (HEK)-293T cells. We found that Wnt3a activated the MyoD proximal 6.0k promoter (P6P) only marginally, but highly enhanced the activity of the composite P6P plus distal enhancer (DE) reporter through canonical and non-canonical pathways. Further screening of the intervening fragments between the DE and the P6P identified a strong Wnt-response element (WRE) in the upstream -8 to -9k region (L fragment) that acted independently of the DE, but was dependent on the P6P. Deletion of a Pax3/Pax7-targeted site in the L fragment significantly reduced its response to Wnt3a, implying that Wnt3a activates the L fragment partially through Pax3/Pax7 action. Binding of β-catenin and Pax7 to their target sites in the DE and the L fragment respectively was also demonstrated by ChIP. These observations demonstrated the first time that Wnt3a can directly activate MyoD expression through targeting cis-elements in the DE and the L fragment.

  4. 脂肪细胞分化过程中影响PAI-1基因转录表达的Dex和C/EBPs顺式调控元件的分析%Analysis of a Novel Dexamethasone Response Element and a Putative C/EBPs cis-Motif: Controlling PAI-1 Gene Expression During Adipocyte Differentiation

    Institute of Scientific and Technical Information of China (English)

    陈可洋; 马春姑; 汤其群; 宋后燕

    2002-01-01

    在研究胰岛素(Ins)、地塞米松(Dex)和甲基异丁基黄嘌呤(Mix)对脂肪细胞分化过程中PAI-1基因表达的影响基础上,为进一步探讨Ins、Dex调控PAI-1基因转录表达的调控机制,应用DNA重组技术,构建含萤光素酶(luciferase)报告基因和PAI-1启动子不同长度片段的嵌合质粒,转染3T3-L1前脂肪细胞并测定报告基因荧光素酶的活性.结果表明,小鼠PAI-1基因起动子-690至-850碱基序列之间有一个Dex的正调控元件.用计算机软件进行分析发现:Dex顺式元件位于PAI-1启动子的-750至-770碱基序列.其组成为:5′ GGTAACCTCTGTTCTCAT 3′.同时还发现在PAI-1启动子的-720至-740碱基序列中,存在一个C/EBPs的结合元件5′CCAAT3′并用凝胶电泳迁移实验对这些元件进行了鉴定.表明Dex正是通过激活转录因子(糖皮质激素受体,GR)和C/EBPα一起与各自的顺式元件结合来促进PAI-1基因的表达.%It has been reported that there is a significant increase in PAI-1 expression level in obese subjects. To explore the linkage between PAI-1 gene expression and obesity, the restriction enzymes and DNA recombination technologies were used to construct the chimeric plasmids with luciferase and different lengths of PAI-1 promoter. After transfection of the chimeric plasmids into 3T3-L1 preadipocyte and detection of luciferase activity, the results indicated that a positive dexamethasone cis-acting element (bases -690 to -850) may be present in mouse PAI-1 promoter. In addition, computer analysis using Match-Search Software found that a new motif of DexRE (dexamethasone response element) 5′ GGTAACCTCTGTTCTCAT 3′ and a putative C/EBPs binding site (cis-motif) exist respectively in the fragment (nucleotides -751 to -770) of, and a sequence (bases -720 to -740) of, mouse PAI-1 promoter,and GMSA and competition assays identified that the trans-acting factors induced by dexamethasone can specifically bind to those cis-motifs. Meanwhile

  5. 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.

  6. Sequences upstream of the homologous cis-elements of the Adh adult enhancer of Drosophila are required for maximal levels of Adh gene transcription in adults of Scaptodrosophila lebanonensis.

    Science.gov (United States)

    Papaceit, Montserrat; Orengo, Dorcas; Juan, Elvira

    2004-01-01

    The evolution of cis-regulatory elements is of particular interest for our understanding of the evolution of gene regulation. The Adh gene of Drosophilidae shows interspecific differences in tissue-specific expression and transcript levels during development. In Scaptodrosophila lebanonensis adults, the level of distal transcripts is maximal between the fourth and eighth day after eclosion and is around five times higher than that in D. melanogaster Adh(S). To examine whether these quantitative differences are regulated by sequences lying upstream of the distal promoter, we performed in vitro deletion mutagenesis of the Adh gene of S. lebanonensis, followed by P-element-mediated germ-line transformation. All constructs included, as a cotransgene, a modified Adh gene of D. melanogaster (dAdh) in a fixed position and orientation that acted as a chromosomal position control. Using this approach, we have identified a fragment of 1.5 kb in the 5' region, 830 bp upstream of the distal start site, which is required to achieve maximal levels of distal transcript in S. lebanonensis. The presence of this fragment produces a 3.5-fold higher level of distal mRNA (as determined by real time quantitative PCR) compared with the D. melanogaster dAdh cotransgene. This region contains the degenerated end of a minisatellite sequence expanding farther upstream and does not correspond to the Adh adult enhancer (AAE) of D. melanogaster. Indeed, the cis-regulatory elements of the AAE have been identified by phylogenetic footprinting within the region 830 bp upstream of the distal start site of S. lebanonensis. Furthermore, the deletions Delta-830 and Delta-2358 yield the same pattern of tissue-specific expression, indicating that all tissue-specific elements are contained within the region 830 bp upstream of the distal start site. PMID:15166155

  7. Histone H3 K27 acetylation marks a potent enhancer element on the adipogenic master regulator gene Pparg2

    Science.gov (United States)

    Ramlee, Muhammad Khairul; Zhang, Qiongyi; Idris, Muhammad; Peng, Xu; Sim, Choon Kiat; Han, Weiping; Xu, Feng

    2014-01-01

    PPARγ2 is expressed almost exclusively in adipose tissue and plays a central role in adipogenesis. Despite intensive studies over the last 2 decades, the mechanism regulating the expression of the Pparg2 gene, especially the role of cis-regulatory elements, is still not completely understood. Here, we report a comprehensive investigation of the enhancer elements within the murine Pparg2 gene. Utilizing the combined techniques of sequence conservation analysis and chromatin marker examination, we identified a potent enhancer element that augmented the expression of a reporter gene under the control of the Pparg2 promoter by 20-fold. This enhancer element was first identified as highly conserved non-coding sequence 10 (CNS10) and was later shown to be enriched with the enhancer marker H3 K27 acetylation. Further studies identified a binding site for p300 as the essential enhancer element in CNS10. Moreover, p300 physically binds to CNS10 and is required for the enhancer activity of CNS10. The depletion of p300 by siRNA resulted in significantly impaired activation of Pparg2 at the early stages of 3T3-L1 adipogenesis. In summary, our study identified a novel enhancer element on the murine Pparg2 gene and suggested a novel mechanism for the regulation of Pparg2 expression by p300 in 3T3-L1 adipogenesis. PMID:25485585

  8. Evolution of anterior Hox regulatory elements among chordates

    Directory of Open Access Journals (Sweden)

    Natale Alfonso

    2011-11-01

    Full Text Available Abstract Background The Hox family of transcription factors has a fundamental role in segmentation pathways and axial patterning of embryonic development and their clustered organization is linked with the regulatory mechanisms governing their coordinated expression along embryonic axes. Among chordates, of particular interest are the Hox paralogous genes in groups 1-4 since their expression is coupled to the control of regional identity in the anterior nervous system, where the highest structural diversity is observed. Results To investigate the degree of conservation in cis-regulatory components that form the basis of Hox expression in the anterior nervous system, we have used assays for transcriptional activity in ascidians and vertebrates to compare and contrast regulatory potential. We identified four regulatory sequences located near the CiHox1, CiHox2 and CiHox4 genes of the ascidian Ciona intestinalis which direct neural specific domains of expression. Using functional assays in Ciona and vertebrate embryos in combination with sequence analyses of enhancer fragments located in similar positions adjacent to Hox paralogy group genes, we compared the activity of these four Ciona cis-elements with a series of neural specific enhancers from the amphioxus Hox1-3 genes and from mouse Hox paralogous groups 1-4. Conclusions This analysis revealed that Kreisler and Krox20 dependent enhancers critical in segmental regulation of the hindbrain appear to be specific for the vertebrate lineage. In contrast, neural enhancers that function as Hox response elements through the action of Hox/Pbx binding motifs have been conserved during chordate evolution. The functional assays reveal that these Hox response cis-elements are recognized by the regulatory components of different and extant species. Together, our results indicate that during chordate evolution, cis-elements dependent upon Hox/Pbx regulatory complexes, are responsible for key aspects of

  9. Detection and characterization of regulatory elements using probabilistic conditional random field and hidden Markov models

    Institute of Scientific and Technical Information of China (English)

    Hongyan Wang; Xiaobo Zhou

    2013-01-01

    By altering the electrostatic charge of histones or providing binding sites to protein recognition molecules,Chromatin marks have been proposed to regulate gene expression,a property that has motivated researchers to link these marks to cis-regulatory elements.With the help of next generation sequencing technologies,we can now correlate one specific chromatin mark with regulatory elements (e.g.enhancers or promoters) and also build tools,such as hidden Markov models,to gain insight into mark combinations.However,hidden Markov models have limitation for their character of generative models and assume that a current observation depends only on a current hidden state in the chain.Here,we employed two graphical probabilistic models,namely the linear conditional random field model and multivariate hidden Markov model,to mark gene regions with different states based on recurrent and spatially coherent character of these eight marks.Both models revealed chromatin states that may correspond to enhancers and promoters,transcribed regions,transcriptional elongation,and low-signal regions.We also found that the linear conditional random field model was more effective than the hidden Markov model in recognizing regulatory elements,such as promoter-,enhancer-,and transcriptional elongation-associated regions,which gives us a better choice.

  10. Ancient Transposable Elements Transformed the Uterine Regulatory Landscape and Transcriptome during the Evolution of Mammalian Pregnancy

    Directory of Open Access Journals (Sweden)

    Vincent J. Lynch

    2015-02-01

    Full Text Available A major challenge in biology is determining how evolutionarily novel characters originate; however, mechanistic explanations for the origin of new characters are almost completely unknown. The evolution of pregnancy is an excellent system in which to study the origin of novelties because mammals preserve stages in the transition from egg laying to live birth. To determine the molecular bases of this transition, we characterized the pregnant/gravid uterine transcriptome from tetrapods to trace the evolutionary history of uterine gene expression. We show that thousands of genes evolved endometrial expression during the origins of mammalian pregnancy, including genes that mediate maternal-fetal communication and immunotolerance. Furthermore, thousands of cis-regulatory elements that mediate decidualization and cell-type identity in decidualized stromal cells are derived from ancient mammalian transposable elements (TEs. Our results indicate that one of the defining mammalian novelties evolved from DNA sequences derived from ancient mammalian TEs co-opted into hormone-responsive regulatory elements distributed throughout the genome.

  11. Discovery and analysis of evolutionarily conserved intronic splicing regulatory elements.

    Directory of Open Access Journals (Sweden)

    Gene W Yeo

    2007-05-01

    Full Text Available Knowledge of the functional cis-regulatory elements that regulate constitutive and alternative pre-mRNA splicing is fundamental for biology and medicine. Here we undertook a genome-wide comparative genomics approach using available mammalian genomes to identify conserved intronic splicing regulatory elements (ISREs. Our approach yielded 314 ISREs, and insertions of ~70 ISREs between competing splice sites demonstrated that 84% of ISREs altered 5' and 94% altered 3' splice site choice in human cells. Consistent with our experiments, comparisons of ISREs to known splicing regulatory elements revealed that 40%-45% of ISREs might have dual roles as exonic splicing silencers. Supporting a role for ISREs in alternative splicing, we found that 30%-50% of ISREs were enriched near alternatively spliced (AS exons, and included almost all known binding sites of tissue-specific alternative splicing factors. Further, we observed that genes harboring ISRE-proximal exons have biases for tissue expression and molecular functions that are ISRE-specific. Finally, we discovered that for Nova1, neuronal PTB, hnRNP C, and FOX1, the most frequently occurring ISRE proximal to an alternative conserved exon in the splicing factor strongly resembled its own known RNA binding site, suggesting a novel application of ISRE density and the propensity for splicing factors to auto-regulate to associate RNA binding sites to splicing factors. Our results demonstrate that ISREs are crucial building blocks in understanding general and tissue-specific AS regulation and the biological pathways and functions regulated by these AS events.

  12. Putative archaeal viruses from the mesopelagic ocean.

    Science.gov (United States)

    Vik, Dean R; Roux, Simon; Brum, Jennifer R; Bolduc, Ben; Emerson, Joanne B; Padilla, Cory C; Stewart, Frank J; Sullivan, Matthew B

    2017-01-01

    Oceanic viruses that infect bacteria, or phages, are known to modulate host diversity, metabolisms, and biogeochemical cycling, while the viruses that infect marine Archaea remain understudied despite the critical ecosystem roles played by their hosts. Here we introduce "MArVD", for Metagenomic Archaeal Virus Detector, an annotation tool designed to identify putative archaeal virus contigs in metagenomic datasets. MArVD is made publicly available through the online iVirus analytical platform. Benchmarking analysis of MArVD showed it to be >99% accurate and 100% sensitive in identifying the 127 known archaeal viruses among the 12,499 viruses in the VirSorter curated dataset. Application of MArVD to 10 viral metagenomes from two depth profiles in the Eastern Tropical North Pacific (ETNP) oxygen minimum zone revealed 43 new putative archaeal virus genomes and large genome fragments ranging in size from 10 to 31 kb. Network-based classifications, which were consistent with marker gene phylogenies where available, suggested that these putative archaeal virus contigs represented six novel candidate genera. Ecological analyses, via fragment recruitment and ordination, revealed that the diversity and relative abundances of these putative archaeal viruses were correlated with oxygen concentration and temperature along two OMZ-spanning depth profiles, presumably due to structuring of the host Archaea community. Peak viral diversity and abundances were found in surface waters, where Thermoplasmata 16S rRNA genes are prevalent, suggesting these archaea as hosts in the surface habitats. Together these findings provide a baseline for identifying archaeal viruses in sequence datasets, and an initial picture of the ecology of such viruses in non-extreme environments.

  13. Escherichia coli genome-wide promoter analysis: Identification of additional AtoC binding target elements

    Directory of Open Access Journals (Sweden)

    Kolisis Fragiskos N

    2011-05-01

    Full Text Available Abstract Background Studies on bacterial signal transduction systems have revealed complex networks of functional interactions, where the response regulators play a pivotal role. The AtoSC system of E. coli activates the expression of atoDAEB operon genes, and the subsequent catabolism of short-chain fatty acids, upon acetoacetate induction. Transcriptome and phenotypic analyses suggested that atoSC is also involved in several other cellular activities, although we have recently reported a palindromic repeat within the atoDAEB promoter as the single, cis-regulatory binding site of the AtoC response regulator. In this work, we used a computational approach to explore the presence of yet unidentified AtoC binding sites within other parts of the E. coli genome. Results Through the implementation of a computational de novo motif detection workflow, a set of candidate motifs was generated, representing putative AtoC binding targets within the E. coli genome. In order to assess the biological relevance of the motifs and to select for experimental validation of those sequences related robustly with distinct cellular functions, we implemented a novel approach that applies Gene Ontology Term Analysis to the motif hits and selected those that were qualified through this procedure. The computational results were validated using Chromatin Immunoprecipitation assays to assess the in vivo binding of AtoC to the predicted sites. This process verified twenty-two additional AtoC binding sites, located not only within intergenic regions, but also within gene-encoding sequences. Conclusions This study, by tracing a number of putative AtoC binding sites, has indicated an AtoC-related cross-regulatory function. This highlights the significance of computational genome-wide approaches in elucidating complex patterns of bacterial cell regulation.

  14. A SINE-derived element constitutes a unique modular enhancer for mammalian diencephalic Fgf8.

    Directory of Open Access Journals (Sweden)

    Akiko Nakanishi

    Full Text Available Transposable elements, including short interspersed repetitive elements (SINEs, comprise nearly half the mammalian genome. Moreover, they are a major source of conserved non-coding elements (CNEs, which play important functional roles in regulating development-related genes, such as enhancing and silencing, serving for the diversification of morphological and physiological features among species. We previously reported a novel SINE family, AmnSINE1, as part of mammalian-specific CNEs. One AmnSINE1 locus, named AS071, showed an enhancer property in the developing mouse diencephalon. Indeed, AS071 appears to recapitulate the expression of diencephalic fibroblast growth factor 8 (Fgf8. Here we established three independent lines of AS071-transgenic mice and performed detailed expression profiling of AS071-enhanced lacZ in comparison with that of Fgf8 across embryonic stages. We demonstrate that AS071 is a distal enhancer that directs Fgf8 expression in the developing diencephalon. Furthermore, enhancer assays with constructs encoding partially deleted AS071 sequence revealed a unique modular organization in which AS071 contains at least three functionally distinct sub-elements that cooperatively direct the enhancer activity in three diencephalic domains, namely the dorsal midline and the lateral wall of the diencephalon, and the ventral midline of the hypothalamus. Interestingly, the AmnSINE1-derived sub-element was found to specify the enhancer activity to the ventral midline of the hypothalamus. To our knowledge, this is the first discovery of an enhancer element that could be separated into respective sub-elements that determine regional specificity and/or the core enhancing activity. These results potentiate our understanding of the evolution of retroposon-derived cis-regulatory elements as well as the basis for future studies of the molecular mechanism underlying the determination of domain-specificity of an enhancer.

  15. Whorl-specific expression of the SUPERMAN gene of Arabidopsis is mediated by cis elements in the transcribed region.

    Science.gov (United States)

    Ito, Toshiro; Sakai, Hajime; Meyerowitz, Elliot M

    2003-09-02

    The SUPERMAN (SUP) gene of Arabidopsis is involved in controlling cell proliferation in stamen and carpel primordia and in ovules during flower development. The SUP gene encodes a transcription factor with a C2H2-type zinc finger motif, a serine/proline-rich domain, a basic domain, and a leucine-zipper-like domain and is expressed in a very limited region in stamen primordia and in the developing ovary during flower development. The SUP gene is susceptible to methylation, resulting in epigenetic gene silencing. To understand how the SUP gene is expressed spatially and temporally in its restricted domain, and why methylation of the transcribed region affects early-stage SUP expression, we have identified the SUP cis regulatory elements by characterizing SUP gene fusions. These studies show that the SUP gene has discrete upstream promoter elements required for expression in stamen primordia in early stages and in the ovary in later stages. The promoter activity for stamen primordia is modulated by several positive and negative elements located in the transcribed and translated regions. Several regulatory elements in the transcribed region correlate with the areas of the gene that are heavily methylated in epigenetic alleles; these data provide a possible explanation of how methylation of the transcribed region represses transcription.

  16. Copy number variation and transposable elements feature in recent, ongoing adaptation at the Cyp6g1 locus

    National Research Council Canada - National Science Library

    Schmidt, Joshua M; Good, Robert T; Appleton, Belinda; Sherrard, Jayne; Raymant, Greta C; Bogwitz, Michael R; Martin, Jon; Daborn, Phillip J; Goddard, Mike E; Batterham, Philip; Robin, Charles

    2010-01-01

    The increased transcription of the Cyp6g1 gene of Drosophila melanogaster, and consequent resistance to insecticides such as DDT, is a widely cited example of adaptation mediated by cis-regulatory change...

  17. Evolutionary origin of Rosaceae-specific active non-autonomous hAT elements and their contribution to gene regulation and genomic structural variation.

    Science.gov (United States)

    Wang, Lu; Peng, Qian; Zhao, Jianbo; Ren, Fei; Zhou, Hui; Wang, Wei; Liao, Liao; Owiti, Albert; Jiang, Quan; Han, Yuepeng

    2016-05-01

    Transposable elements account for approximately 30 % of the Prunus genome; however, their evolutionary origin and functionality remain largely unclear. In this study, we identified a hAT transposon family, termed Moshan, in Prunus. The Moshan elements consist of three types, aMoshan, tMoshan, and mMoshan. The aMoshan and tMoshan types contain intact or truncated transposase genes, respectively, while the mMoshan type is miniature inverted-repeat transposable element (MITE). The Moshan transposons are unique to Rosaceae, and the copy numbers of different Moshan types are significantly correlated. Sequence homology analysis reveals that the mMoshan MITEs are direct deletion derivatives of the tMoshan progenitors, and one kind of mMoshan containing a MuDR-derived fragment were amplified predominately in the peach genome. The mMoshan sequences contain cis-regulatory elements that can enhance gene expression up to 100-fold. The mMoshan MITEs can serve as potential sources of micro and long noncoding RNAs. Whole-genome re-sequencing analysis indicates that mMoshan elements are highly active, and an insertion into S-haplotype-specific F-box gene was reported to cause the breakdown of self-incompatibility in sour cherry. Taken together, all these results suggest that the mMoshan elements play important roles in regulating gene expression and driving genomic structural variation in Prunus.

  18. Extensive cis-regulatory variation robust to environmental perturbation in Arabidopsis.

    Science.gov (United States)

    Cubillos, Francisco A; Stegle, Oliver; Grondin, Cécile; Canut, Matthieu; Tisné, Sébastien; Gy, Isabelle; Loudet, Olivier

    2014-11-01

    cis- and trans-acting factors affect gene expression and responses to environmental conditions. However, for most plant systems, we lack a comprehensive map of these factors and their interaction with environmental variation. Here, we examined allele-specific expression (ASE) in an F1 hybrid to study how alleles from two Arabidopsis thaliana accessions affect gene expression. To investigate the effect of the environment, we used drought stress and developed a variance component model to estimate the combined genetic contributions of cis- and trans-regulatory polymorphisms, environmental factors, and their interactions. We quantified ASE for 11,003 genes, identifying 3318 genes with consistent ASE in control and stress conditions, demonstrating that cis-acting genetic effects are essentially robust to changes in the environment. Moreover, we found 1618 genes with genotype x environment (GxE) interactions, mostly cis x E interactions with magnitude changes in ASE. We found fewer trans x E interactions, but these effects were relatively less robust across conditions, showing more changes in the direction of the effect between environments; this confirms that trans-regulation plays an important role in the response to environmental conditions. Our data provide a detailed map of cis- and trans-regulation and GxE interactions in A. thaliana, laying the ground for mechanistic investigations and studies in other plants and environments.

  19. Extensive cis-Regulatory Variation Robust to Environmental Perturbation in Arabidopsis[W

    Science.gov (United States)

    Cubillos, Francisco A.; Stegle, Oliver; Grondin, Cécile; Canut, Matthieu; Tisné, Sébastien; Gy, Isabelle

    2014-01-01

    cis- and trans-acting factors affect gene expression and responses to environmental conditions. However, for most plant systems, we lack a comprehensive map of these factors and their interaction with environmental variation. Here, we examined allele-specific expression (ASE) in an F1 hybrid to study how alleles from two Arabidopsis thaliana accessions affect gene expression. To investigate the effect of the environment, we used drought stress and developed a variance component model to estimate the combined genetic contributions of cis- and trans-regulatory polymorphisms, environmental factors, and their interactions. We quantified ASE for 11,003 genes, identifying 3318 genes with consistent ASE in control and stress conditions, demonstrating that cis-acting genetic effects are essentially robust to changes in the environment. Moreover, we found 1618 genes with genotype x environment (GxE) interactions, mostly cis x E interactions with magnitude changes in ASE. We found fewer trans x E interactions, but these effects were relatively less robust across conditions, showing more changes in the direction of the effect between environments; this confirms that trans-regulation plays an important role in the response to environmental conditions. Our data provide a detailed map of cis- and trans-regulation and GxE interactions in A. thaliana, laying the ground for mechanistic investigations and studies in other plants and environments. PMID:25428981

  20. Evolution of metal hyperaccumulation required cis-regulatory changes and triplication of HMA4

    OpenAIRE

    Hanikenne, Marc; Talke, Ina N.; Haydon, Michael J.; Lanz, Christa; Nolte, Andrea; Motte, Patrick; Kroymann, Juergen; Weigel, Detlef; Krämer, Ute

    2008-01-01

    Little is known about the types of mutations underlying the evolution of species-specific traits. The metal hyperaccumulator Arabidopsis halleri has the rare ability to colonize heavy-metal-polluted soils, and, as an extremophile sister species of Arabidopsis thaliana, it is a powerful model for research on adaptation. A. halleri naturally accumulates and tolerates leaf concentrations as high as 2.2% zinc and 0.28% cadmium in dry biomass. On the basis of transcriptomics studies, metal hyperac...

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

    DEFF Research Database (Denmark)

    Wong, Wendy S W; Nielsen, Rasmus

    2007-01-01

    of the increasing availability of comparative genomic data. RESULTS: We develop a method for finding regulatory modules in Eukaryotic species using phylogenetic data. Using computer simulations and analysis of real data, we show that the use of phylogenetic hidden Markov model can lead to an increase in accuracy...

  2. Subfunctionalization of duplicated zebrafish pax6 genes by cis-regulatory divergence

    National Research Council Canada - National Science Library

    Kleinjan, Dirk A; Bancewicz, Ruth M; Gautier, Philippe; Dahm, Ralf; Schonthaler, Helia B; Damante, Giuseppe; Seawright, Anne; Hever, Ann M; Yeyati, Patricia L; van Heyningen, Veronica; Coutinho, Pedro

    2008-01-01

    Gene duplication is a major driver of evolutionary divergence. In most vertebrates a single PAX6 gene encodes a transcription factor required for eye, brain, olfactory system, and pancreas development...

  3. Interactions between glucocorticoid treatment and cis-regulatory polymorphisms contribute to cellular response phenotypes.

    Directory of Open Access Journals (Sweden)

    Joseph C Maranville

    2011-07-01

    Full Text Available Glucocorticoids (GCs mediate physiological responses to environmental stress and are commonly used as pharmaceuticals. GCs act primarily through the GC receptor (GR, a transcription factor. Despite their clear biomedical importance, little is known about the genetic architecture of variation in GC response. Here we provide an initial assessment of variability in the cellular response to GC treatment by profiling gene expression and protein secretion in 114 EBV-transformed B lymphocytes of African and European ancestry. We found that genetic variation affects the response of nearby genes and exhibits distinctive patterns of genotype-treatment interactions, with genotypic effects evident in either only GC-treated or only control-treated conditions. Using a novel statistical framework, we identified interactions that influence the expression of 26 genes known to play central roles in GC-related pathways (e.g. NQO1, AIRE, and SGK1 and that influence the secretion of IL6.

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

    Directory of Open Access Journals (Sweden)

    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.

  5. Landscape of histone modifications in a sponge reveals the origin of animal cis-regulatory complexity

    Science.gov (United States)

    Gaiti, Federico; Jindrich, Katia; Fernandez-Valverde, Selene L; Roper, Kathrein E; Degnan, Bernard M; Tanurdžić, Miloš

    2017-01-01

    Combinatorial patterns of histone modifications regulate developmental and cell type-specific gene expression and underpin animal complexity, but it is unclear when this regulatory system evolved. By analysing histone modifications in a morphologically-simple, early branching animal, the sponge Amphimedonqueenslandica, we show that the regulatory landscape used by complex bilaterians was already in place at the dawn of animal multicellularity. This includes distal enhancers, repressive chromatin and transcriptional units marked by H3K4me3 that vary with levels of developmental regulation. Strikingly, Amphimedon enhancers are enriched in metazoan-specific microsyntenic units, suggesting that their genomic location is extremely ancient and likely to place constraints on the evolution of surrounding genes. These results suggest that the regulatory foundation for spatiotemporal gene expression evolved prior to the divergence of sponges and eumetazoans, and was necessary for the evolution of animal multicellularity. DOI: http://dx.doi.org/10.7554/eLife.22194.001 PMID:28395144

  6. Ten Putative Contributors to the Obesity Epidemic

    Science.gov (United States)

    McAllister, Emily J.; Dhurandhar, Nikhil V.; Keith, Scott W.; Aronne, Louis J.; Barger, Jamie; Baskin, Monica; Benca, Ruth M.; Biggio, Joseph; Boggiano, Mary M.; Eisenmann, Joe C.; Elobeid, Mai; Fontaine, Kevin R.; Gluckman, Peter; Hanlon, Erin C.; Katzmarzyk, Peter; Pietrobelli, Angelo; Redden, David T.; Ruden, Douglas M.; Wang, Chenxi; Waterland, Robert A.; Wright, Suzanne M.; Allison, David B.

    2010-01-01

    The obesity epidemic is a global issue and shows no signs of abating, while the cause of this epidemic remains unclear. Marketing practices of energy-dense foods and institutionally-driven declines in physical activity are the alleged perpetrators for the epidemic, despite a lack of solid evidence to demonstrate their causal role. While both may contribute to obesity, we call attention to their unquestioned dominance in program funding and public efforts to reduce obesity, and propose several alternative putative contributors that would benefit from equal consideration and attention. Evidence for microorganisms, epigenetics, increasing maternal age, greater fecundity among people with higher adiposity, assortative mating, sleep debt, endocrine disruptors, pharmaceutical iatrogenesis, reduction in variability of ambient temperatures, and intrauterine and intergenerational effects, as contributing factors to the obesity epidemic are reviewed herein. While the evidence is strong for some contributors such as pharmaceutical-induced weight gain, it is still emerging for other reviewed factors. Considering the role of such putative etiological factors of obesity may lead to comprehensive, cause specific, and effective strategies for prevention and treatment of this global epidemic. PMID:19960394

  7. Evolutionary Novelty in a Butterfly Wing Pattern through Enhancer Shuffling

    National Research Council Canada - National Science Library

    Wallbank, Richard W R; Baxter, Simon W; Pardo-Diaz, Carolina; Hanly, Joseph J; Martin, Simon H; Mallet, James; Dasmahapatra, Kanchon K; Salazar, Camilo; Joron, Mathieu; Nadeau, Nicola; McMillan, W Owen; Jiggins, Chris D

    2016-01-01

    .... Phylogenetic analysis of the two elements demonstrated that they have distinct evolutionary histories and that novel adaptive morphological variation was created by shuffling these cis-regulatory...

  8. Putative Nitrogen Sensing Systems in Higher Plants

    Institute of Scientific and Technical Information of China (English)

    Hon-Ming Lam; Ying Ann Chiao; Man-Wah Li; Yuk-Kwong Yung; Sang Ji

    2006-01-01

    Nitrogen (N) metabolism is essential for the biosynthesis of vital biomolecules. N status thus exerts profound effects on plant growth and development, and must be closely monitored. In bacteria and fungi, a few sophisticated N sensing systems have been extensively studied. In animals, the ability to receive amino acid signals has evolved to become an integral part of the nervous coordination system. In this review, we will summarize recent developments in the search for putative N sensing systems in higher plants based on homologous systems in bacteria, fungi, and animals. Apparently, although plants have separated and diversified from other organisms during the evolution process, striking similarities can be found in their N sensing systems compared with those of their counterparts; however, our understanding of these systems is still incomplete. Significant modifications of the N sensing systems (including cross-talk with other signal transduction pathways) in higher plants may be a strategy of adaptation to their unique mode of life.

  9. Putative respiratory chain of Porphyromonas gingivalis.

    Science.gov (United States)

    Meuric, Vincent; Rouillon, Astrid; Chandad, Fatiha; Bonnaure-Mallet, Martine

    2010-05-01

    The electron transfer chain in Porphyromonas gingivalis, or periodontopathogens, has not yet been characterized. P. gingivalis, a strict anaerobic bacteria and the second colonizer of the oral cavity, is considered to be a major causal agent involved in periodontal diseases. Primary colonizers create a favorable environment for P. gingivalis growth by decreasing oxygen pressure. Oxygen does not appear to be the final electron acceptor of the respiratory chain. Fumarate and cytochrome b have been implicated as major components of the respiratory activity. However, the P. gingivalis genome shows many other enzymes that could be implicated in aerobic or nitrite respiration. Using bioinformatic tools and literature studies of respiratory pathways, the ATP synthesis mechanism from the sodium cycle and nutrients metabolism, the putative respirasome of P. gingivalis has been proposed.

  10. 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.

  11. Genome-wide profiling of p63 DNA-binding sites identifies an element that regulates gene expression during limb development in the 7q21 SHFM1 locus.

    Directory of Open Access Journals (Sweden)

    Evelyn N Kouwenhoven

    2010-08-01

    Full Text Available Heterozygous mutations in p63 are associated with split hand/foot malformations (SHFM, orofacial clefting, and ectodermal abnormalities. Elucidation of the p63 gene network that includes target genes and regulatory elements may reveal new genes for other malformation disorders. We performed genome-wide DNA-binding profiling by chromatin immunoprecipitation (ChIP, followed by deep sequencing (ChIP-seq in primary human keratinocytes, and identified potential target genes and regulatory elements controlled by p63. We show that p63 binds to an enhancer element in the SHFM1 locus on chromosome 7q and that this element controls expression of DLX6 and possibly DLX5, both of which are important for limb development. A unique micro-deletion including this enhancer element, but not the DLX5/DLX6 genes, was identified in a patient with SHFM. Our study strongly indicates disruption of a non-coding cis-regulatory element located more than 250 kb from the DLX5/DLX6 genes as a novel disease mechanism in SHFM1. These data provide a proof-of-concept that the catalogue of p63 binding sites identified in this study may be of relevance to the studies of SHFM and other congenital malformations that resemble the p63-associated phenotypes.

  12. Putative bronchopulmonary flagellated protozoa in immunosuppressed patients.

    Science.gov (United States)

    Kilimcioglu, Ali Ahmet; Havlucu, Yavuz; Girginkardesler, Nogay; Celik, Pınar; Yereli, Kor; Özbilgin, Ahmet

    2014-01-01

    Flagellated protozoa that cause bronchopulmonary symptoms in humans are commonly neglected. These protozoal forms which were presumed to be "flagellated protozoa" have been previously identified in immunosuppressed patients in a number of studies, but have not been certainly classified so far. Since no human cases of bronchopulmonary flagellated protozoa were reported from Turkey, we aimed to investigate these putative protozoa in immunosuppressed patients who are particularly at risk of infectious diseases. Bronchoalveolar lavage fluid samples of 110 immunosuppressed adult patients who were admitted to the Department of Chest Diseases, Hafsa Sultan Hospital of Celal Bayar University, Manisa, Turkey, were examined in terms of parasites by light microscopy. Flagellated protozoal forms were detected in nine (8.2%) of 110 cases. Metronidazole (500 mg b.i.d. for 30 days) was given to all positive cases and a second bronchoscopy was performed at the end of the treatment, which revealed no parasites. In conclusion, immunosuppressed patients with bronchopulmonary symptoms should attentively be examined with regard to flagellated protozoa which can easily be misidentified as epithelial cells.

  13. The Biogeography of Putative Microbial Antibiotic Production.

    Directory of Open Access Journals (Sweden)

    Hélène Morlon

    Full Text Available Understanding patterns in the distribution and abundance of functional traits across a landscape is of fundamental importance to ecology. Mapping these distributions is particularly challenging for species-rich groups with sparse trait measurement coverage, such as flowering plants, insects, and microorganisms. Here, we use likelihood-based character reconstruction to infer and analyze the spatial distribution of unmeasured traits. We apply this framework to a microbial dataset comprised of 11,732 ketosynthase alpha gene sequences extracted from 144 soil samples from three continents to document the spatial distribution of putative microbial polyketide antibiotic production. Antibiotic production is a key competitive strategy for soil microbial survival and performance. Additionally, novel antibiotic discovery is highly relevant to human health, making natural antibiotic production by soil microorganisms a major target for bioprospecting. Our comparison of trait-based biogeographical patterns to patterns based on taxonomy and phylogeny is relevant to our basic understanding of microbial biogeography as well as the pressing need for new antibiotics.

  14. Mechanosensory neurons, cutaneous mechanoreceptors, and putative mechanoproteins.

    Science.gov (United States)

    Del Valle, M E; Cobo, T; Cobo, J L; Vega, J A

    2012-08-01

    The mammalian skin has developed sensory structures (mechanoreceptors) that are responsible for different modalities of mechanosensitivity like touch, vibration, and pressure sensation. These specialized sensory organs are anatomically and functionally connected to a special subset of sensory neurons called mechanosensory neurons, which electrophysiologically correspond with Aβ fibers. Although mechanosensory neurons and cutaneous mechanoreceptors are rather well known, the biology of the sense of touch still remains poorly understood. Basically, the process of mechanosensitivity requires the conversion of a mechanical stimulus into an electrical signal through the activation of ion channels that gate in response to mechanical stimuli. These ion channels belong primarily to the family of the degenerin/epithelium sodium channels, especially the subfamily acid-sensing ion channels, and to the family of transient receptor potential channels. This review compiles the current knowledge on the occurrence of putative mechanoproteins in mechanosensory neurons and mechanoreceptors, as well as the involvement of these proteins on the biology of touch. Furthermore, we include a section about what the knock-out mice for mechanoproteins are teaching us. Finally, the possibilities for mechanotransduction in mechanoreceptors, and the common involvement of the ion channels, extracellular membrane, and cytoskeleton, are revisited.

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

    Directory of Open Access Journals (Sweden)

    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.

  16. Comprehensive Phenotyping in Multiple Sclerosis: Discovery Based Proteomics and the Current Understanding of Putative Biomarkers

    Directory of Open Access Journals (Sweden)

    Kevin C. O’Connor

    2006-01-01

    Full Text Available Currently, there is no single test for multiple sclerosis (MS. Diagnosis is confirmed through clinical evaluation, abnormalities revealed by magnetic resonance imaging (MRI, and analysis of cerebrospinal fluid (CSF chemistry. The early and accurate diagnosis of the disease, monitoring of progression, and gauging of therapeutic intervention are important but elusive elements of patient care. Moreover, a deeper understanding of the disease pathology is needed, including discovery of accurate biomarkers for MS. Herein we review putative biomarkers of MS relating to neurodegeneration and contributions to neuropathology, with particular focus on autoimmunity. In addition, novel assessments of biomarkers not driven by hypotheses are discussed, featuring our application of advanced proteomics and metabolomics for comprehensive phenotyping of CSF and blood. This strategy allows comparison of component expression levels in CSF and serum between MS and control groups. Examination of these preliminary data suggests that several CSF proteins in MS are differentially expressed, and thus, represent putative biomarkers deserving of further evaluation.

  17. A distal intergenic region controls pancreatic endocrine differentiation by acting as a transcriptional enhancer and as a polycomb response element

    Science.gov (United States)

    Pardanaud-Glavieux, Corinne; García-Hurtado, Javier; Sauty, Claire; Guerci, Aline; Ferrer, Jorge

    2017-01-01

    Lineage-selective expression of developmental genes is dependent on the interplay between activating and repressive mechanisms. Gene activation is dependent on cell-specific transcription factors that recognize transcriptional enhancer sequences. Gene repression often depends on the recruitment of Polycomb group (PcG) proteins, although the sequences that underlie the recruitment of PcG proteins, also known as Polycomb response elements (PREs), remain poorly understood in vertebrates. While distal PREs have been identified in mammals, a role for positive-acting enhancers in PcG-mediated repression has not been described. Here we have used a highly efficient procedure based on lentiviral-mediated transgenesis to carry out in vivo fine-mapping of, cis-regulatory sequences that control lineage-specific activation of Neurog3, a master regulator of pancreatic endocrine differentiation. Our findings reveal an enhancer region that is sufficient to drive correct spacio-temporal expression of Neurog3 and demonstrate that this same region serves as a PRE in alternative lineages where Neurog3 is inactive. PMID:28225770

  18. A method of predicting changes in human gene splicing induced by genetic variants in context of cis-acting elements

    Directory of Open Access Journals (Sweden)

    Hicks Chindo

    2010-01-01

    Full Text Available Abstract Background Polymorphic variants and mutations disrupting canonical splicing isoforms are among the leading causes of human hereditary disorders. While there is a substantial evidence of aberrant splicing causing Mendelian diseases, the implication of such events in multi-genic disorders is yet to be well understood. We have developed a new tool (SpliceScan II for predicting the effects of genetic variants on splicing and cis-regulatory elements. The novel Bayesian non-canonical 5'GC splice site (SS sensor used in our tool allows inference on non-canonical exons. Results Our tool performed favorably when compared with the existing methods in the context of genes linked to the Autism Spectrum Disorder (ASD. SpliceScan II was able to predict more aberrant splicing isoforms triggered by the mutations, as documented in DBASS5 and DBASS3 aberrant splicing databases, than other existing methods. Detrimental effects behind some of the polymorphic variations previously associated with Alzheimer's and breast cancer could be explained by changes in predicted splicing patterns. Conclusions We have developed SpliceScan II, an effective and sensitive tool for predicting the detrimental effects of genomic variants on splicing leading to Mendelian and complex hereditary disorders. The method could potentially be used to screen resequenced patient DNA to identify de novo mutations and polymorphic variants that could contribute to a genetic disorder.

  19. Quantitative analysis of polycomb response elements (PREs at identical genomic locations distinguishes contributions of PRE sequence and genomic environment

    Directory of Open Access Journals (Sweden)

    Okulski Helena

    2011-03-01

    Full Text Available Abstract Background Polycomb/Trithorax response elements (PREs are cis-regulatory elements essential for the regulation of several hundred developmentally important genes. However, the precise sequence requirements for PRE function are not fully understood, and it is also unclear whether these elements all function in a similar manner. Drosophila PRE reporter assays typically rely on random integration by P-element insertion, but PREs are extremely sensitive to genomic position. Results We adapted the ΦC31 site-specific integration tool to enable systematic quantitative comparison of PREs and sequence variants at identical genomic locations. In this adaptation, a miniwhite (mw reporter in combination with eye-pigment analysis gives a quantitative readout of PRE function. We compared the Hox PRE Frontabdominal-7 (Fab-7 with a PRE from the vestigial (vg gene at four landing sites. The analysis revealed that the Fab-7 and vg PREs have fundamentally different properties, both in terms of their interaction with the genomic environment at each site and their inherent silencing abilities. Furthermore, we used the ΦC31 tool to examine the effect of deletions and mutations in the vg PRE, identifying a 106 bp region containing a previously predicted motif (GTGT that is essential for silencing. Conclusions This analysis showed that different PREs have quantifiably different properties, and that changes in as few as four base pairs have profound effects on PRE function, thus illustrating the power and sensitivity of ΦC31 site-specific integration as a tool for the rapid and quantitative dissection of elements of PRE design.

  20. Toxic Elements

    DEFF Research Database (Denmark)

    Hajeb, Parvaneh; Shakibazadeh, Shahram; Sloth, Jens Jørgen

    2016-01-01

    Food is considered the main source of toxic element (arsenic, cadmium, lead, and mercury) exposure to humans, and they can cause major public health effects. In this chapter, we discuss the most important sources for toxic element in food and the foodstuffs which are significant contributors...... to human exposure. The occurrence of each element in food classes from different regions is presented. Some of the current toxicological risk assessments on toxic elements, the human health effect of each toxic element, and their contents in the food legislations are presented. An overview of analytical...... techniques and challenges for determination of toxic elements in food is also given....

  1. A putative viral defence mechanism in archaeal cells

    Directory of Open Access Journals (Sweden)

    Reidun Lillestøl

    2006-01-01

    Full Text Available Clusters of regularly spaced direct repeats, separated by unconserved spacer sequences, are ubiquitous in archaeal chromosomes and occur in some plasmids. Some clusters constitute around 1% of chromosomal DNA. Similarly structured clusters, generally smaller, also occur in some bacterial chromosomes. Although early studies implicated these clusters in segregation/partition functions, recent evidence suggests that the spacer sequences derive from extrachromosomal elements, and, primarily, viruses. This has led to the proposal that the clusters provide a defence against viral propagation in cells, and that both the mode of inhibition of viral propagation and the mechanism of adding spacer-repeat units to clusters, are dependent on RNAs transcribed from the clusters. Moreover, the putative inhibitory apparatus (piRNA-based may be evolutionarily related to the interference RNA systems (siRNA and miRNA, which are common in eukarya. Here, we analyze all the current data on archaeal repeat clusters and provide some new insights into their diverse structures, transcriptional properties and mode of structural development. The results are consistent with larger cluster transcripts being processed at the centers of the repeat sequences and being further trimmed by exonucleases to yield a dominant, intracellular RNA species, which corresponds approximately to the size of a spacer. Furthermore, analysis of the extensive clusters of Sulfolobus solfataricus strains P1 and P2B provides support for the presence of a flanking sequence adjoining a cluster being a prerequisite for the incorporation of new spacer-repeat units, which occurs between the flanking sequence and the cluster. An archaeal database summarizing the data will be maintained at http://dac.molbio.ku.dk/dbs/SRSR/.

  2. Transcriptome-Based Examination of Putative Pollen Allergens of Rice(Oryza sativa ssp.japonica)

    Institute of Scientific and Technical Information of China (English)

    Scott D.Russell; Prem L Bhalla; Mohan B.Singh

    2008-01-01

    Pollen allergens are among the most abundantly transcribed and translated products in the Iife history of plants,and particularly grasses.To identify different pollen allergens in rice,putative allergens were identified in the rice genome and their expression characterized using the Affymetrix 57K rice GeneChip microarray.Among the most abundant pollen-specific candidate transcripts were Ory s 1 beta-expansin.Ory s 2,Ory s 7 EFhand,Ory s 11,Ory s 12 profilin A,Ory s 23,glycosyl hydrolase family 28(polygalacturonase).and FAD binding proteins.Highly expressed pollen proteins are frequently present in multiple copy numbers,sometimes with mirror images Iocated on nearby regions of the opposite DNA strand.Many of these are intronless and inserted as copies that retain nearly exact copies of their regulatory elements.Ory s 23 reflects low variability and high copy number,suggesting recent gene amplification.Some copies contain pseudogenes,which may reflect their origin through activity of retrotransposition;some putative allergenic sequences bear fusion products with repeat sequences of transposable elements(LTRs).The abundance of nearby repetitive sequences,activation of transposable elements.and high production of mRNA transcripts appear to coincide in pollen and may contribute to a syndrome in which highly transcribed proteins may be copied and inserted with streamlined features for translation,including grouping and removaI of introns.

  3. An element

    Energy Technology Data Exchange (ETDEWEB)

    Nakamura, K.; Iidzima, K.

    1983-03-30

    An anode of a light metal is used in the element, along with an electrolyte which consists of an ether solvent and an ionogenic additive in the form of a salt of dithiocarbamic acid. The element has good discharge characteristics.

  4. Putative Corneal Neuralgia Responding to Vitamin D Supplementation

    Directory of Open Access Journals (Sweden)

    Eric L. Singman

    2013-09-01

    Full Text Available A patient with putative corneal neuralgia was incidentally discovered to have hypovitaminosis D. Supplementation of vitamin D appears to have led to a resolution of the patient's pain, whereas other efforts to treat the patient were unsuccessful.

  5. Structure, regulation, and putative function of the arginine deiminase system of Streptococcus suis.

    Science.gov (United States)

    Gruening, Petra; Fulde, Marcus; Valentin-Weigand, Peter; Goethe, Ralph

    2006-01-01

    Streptococcus suis is an important cause of infectious diseases in young pigs. Little is known about the virulence factors or protective antigens of S. suis. Recently, we have identified two proteins of the arginine deiminase system (ADS) of S. suis, which were temperature induced and expressed on the streptococcal surface (N. Winterhoff, R. Goethe, P. Gruening, M. Rohde, H. Kalisz, H. E. Smith, and P. Valentin-Weigand, J. Bacteriol. 184:6768-6776, 2002). In the present study, we analyzed the complete ADS of S. suis. Due to their homologies to the recently published S. gordonii ADS genes, the genes for arginine deiminase, ornithine carbamoyl-transferase, and carbamate kinase, which were previously designated adiS, octS, and ckS, respectively, were renamed arcA, arcB, and arcC, respectively. Our data revealed that arcA, arcB, and arcC of the S. suis ADS are transcribed from an operon (arcABC operon). Additionally, putative ADS-associated genes were cloned and sequenced which, however, did not belong to the arcABC operon. These were the flpS gene upstream of the arcABC operon with homology to the flp transcription regulator of S. gordonii and the arcD, arcT, arcH, and argR genes downstream of the arcABC operon with high homologies to a putative arginine-ornithine antiporter, a putative dipeptidase of S. gordonii, a putative beta-N-acetylhexosaminidase of S. pneumoniae, and a putative arginine repressor of S. gordonii, respectively. The transcriptional start point of the arcABC operon was determined, and promoter analysis provided evidence that multiple factors contribute to the regulation of the ADS. Thus, a putative binding site for a transcription regulator of the Crp/Fnr family, an ArgR-binding site, and two cis-acting catabolite response elements were identified in the promoter-operator region of the operon. Consistent with this, we could demonstrate that the ADS of S. suis is inducible by arginine and reduced O2 tension and subject to carbon catabolite

  6. Human Papillomavirus Type 18 cis-Elements Crucial for Segregation and Latency.

    Directory of Open Access Journals (Sweden)

    Mart Ustav

    Full Text Available Stable maintenance replication is characteristic of the latency phase of HPV infection, during which the viral genomes are actively maintained as extrachromosomal genetic elements in infected proliferating basal keratinocytes. Active replication in the S-phase and segregation of the genome into daughter cells in mitosis are required for stable maintenance replication. Most of our knowledge about papillomavirus genome segregation has come from studies of bovine papillomavirus type 1 (BPV-1, which have demonstrated that the E2 protein cooperates with cellular trans-factors and that E2 binding sites act as cis-regulatory elements in the viral genome that are essential for the segregation process. However, the genomic organization of the regulatory region in HPVs, and the properties of the viral proteins are different from those of their BPV-1 counterparts. We have designed a segregation assay for HPV-18 and used it to demonstrate that the E2 protein performs segregation in combination with at least two E2 binding sites. The cooperative binding of the E2 protein to two E2 binding sites is a major determinant of HPV-18 genome segregation, as demonstrated by the change in spacing between adjacent binding sites #1 and #2 in the HPV-18 Upstream Regulatory Region (URR. Duplication or triplication of the natural 4 bp 5'-CGGG-3' spacer between the E2 binding sites increased the cooperative binding of the E2 molecules as well as E2-dependent segregation. Removal of any spacing between these sites eliminated cooperative binding of the E2 protein and disabled segregation of the URR and HPV-18 genome. Transfer of these configurations of the E2 binding sites into viral genomes confirmed the role of the E2 protein and binding sites #1 and #2 in the segregation process. Additional analysis demonstrated that these sites also play an important role in the transcriptional regulation of viral gene expression from different HPV-18 promoters.

  7. Two estrogen response element sequences near the PCNA gene are not responsible for its estrogen-enhanced expression in MCF7 cells.

    Directory of Open Access Journals (Sweden)

    Cheng Wang

    Full Text Available BACKGROUND: The proliferating cell nuclear antigen (PCNA is an essential component of DNA replication, cell cycle regulation, and epigenetic inheritance. High expression of PCNA is associated with poor prognosis in patients with breast cancer. The 5'-region of the PCNA gene contains two computationally-detected estrogen response element (ERE sequences, one of which is evolutionarily conserved. Both of these sequences are of undocumented cis-regulatory function. We recently demonstrated that estradiol (E2 enhances PCNA mRNA expression in MCF7 breast cancer cells. MCF7 cells proliferate in response to E2. METHODOLOGY/PRINCIPAL FINDINGS: Here, we demonstrate that E2 rapidly enhanced PCNA mRNA and protein expression in a process that requires ERalpha as well as de novo protein synthesis. One of the two upstream ERE sequences was specifically bound by ERalpha-containing protein complexes, in vitro, in gel shift analysis. Yet, each ERE sequence, when cloned as a single copy, or when engineered as two tandem copies of the ERE-containing sequence, was not capable of activating a luciferase reporter construct in response to E2. In MCF7 cells, neither ERE-containing genomic region demonstrated E2-dependent recruitment of ERalpha by sensitive ChIP-PCR assays. CONCLUSION/SIGNIFICANCE: We conclude that E2 enhances PCNA gene expression by an indirect process and that computational detection of EREs, even when evolutionarily conserved and when near E2-responsive genes, requires biochemical validation.

  8. The alr-groEL1 operon in Mycobacterium tuberculosis: an interplay of multiple regulatory elements

    Science.gov (United States)

    Bhat, Aadil H.; Pathak, Deepika; Rao, Alka

    2017-01-01

    Threonylcarbamoyladenosine is a universally conserved essential modification of tRNA that ensures translational fidelity in cellular milieu. TsaD, TsaB and TsaE are identified as tRNA-A37-threonylcarbamoyl (t6A)-transferase enzymes that have been reconstituted in vitro, in few bacteria recently. However, transcriptional organization and regulation of these genes are not known in any of these organisms. This study describes the intricate architecture of a complex multicistronic alr-groEL1 operon, harboring essential genes, namely tsaD, tsaB, tsaE, groES, groEL1, and alr (required for cell wall synthesis), and rimI encoding an N-α- acetyltransferase in Mycobacterium tuberculosis. Using northern blotting, RT-PCR and in vivo fluorescence assays, genes alr to groEL1 were found to constitute an ~6.3 kb heptacistronic operon with multiple internal promoters and an I-shaped intrinsic hairpin-like cis-regulatory element. A strong promoter PtsaD within the coding sequence of rimI gene is identified in M. tuberculosis, in addition. The study further proposes an amendment in the known bicistronic groESL1 operon annotation by providing evidence that groESL1 is co-transcribed as sub-operon of alr-groEL1 operon. The architecture of alr-groEL1 operon, conservation of the genetic context and a mosaic transcriptional profile displayed under various stress conditions convincingly suggest the involvement of this operon in stress adaptation in M. tuberculosis. PMID:28256563

  9. A putative viral defence mechanism in archaeal cells

    DEFF Research Database (Denmark)

    Lillestøl, Reidun K; Redder, Peter; Garrett, Roger Antony

    2006-01-01

    in cells, and that both the mode of inhibition of viral propagation and the mechanism of adding spacer-repeat units to clusters, are dependent on RNAs transcribed from the clusters. Moreover, the putative inhibitory apparatus (piRNA-based) may be evolutionarily related to the interference RNA systems (si...

  10. Putative Lineage of Novel African Usutu Virus, Central Europe

    Centers for Disease Control (CDC) Podcasts

    2015-10-15

    Sarah Gregory reads an abridged version of "Putative Lineage of Novel African Usutu Virus, Central Europe.".  Created: 10/15/2015 by National Center for Emerging and Zoonotic Infectious Diseases (NCEZID).   Date Released: 10/15/2015.

  11. Putative golden proportions as predictors of facial esthetics in adolescents.

    NARCIS (Netherlands)

    Kiekens, R.M.A.; Kuijpers-Jagtman, A.M.; Hof, M.A. van 't; Hof, BE van 't; Maltha, J.C.

    2008-01-01

    INTRODUCTION: In orthodontics, facial esthetics is assumed to be related to golden proportions apparent in the ideal human face. The aim of the study was to analyze the putative relationship between facial esthetics and golden proportions in white adolescents. METHODS: Seventy-six adult laypeople

  12. Geographic polymorphism of P element in populations of Drosophila sturtevanti

    Directory of Open Access Journals (Sweden)

    Luciane M. de Almeida

    2003-01-01

    Full Text Available The aim of this report was to detect full-sized P element sequences in eight strains of Drosophila sturtevanti populations from distant geographic regions and to assess the structural geographic variation among P element sequences. PCR analysis confirmed the presence of a putative complete P element in all strains. Southern blot analysis indicated bands shared by all strains, and bands restricted to geographically related strains. Parsimony analysis corroborated the hybridization pattern that reflected the geographic relationships.

  13. A novel putative enterococcal pathogenicity island linked to the esp virulence gene of Enterococcus faecium and associated with epidemicity.

    Science.gov (United States)

    Leavis, Helen; Top, Janetta; Shankar, Nathan; Borgen, Katrine; Bonten, Marc; van Embden, Jan; Willems, Rob J L

    2004-02-01

    Enterococcus faecalis harbors a virulence-associated surface protein encoded by the esp gene. This gene has been shown to be part of a 150-kb putative pathogenicity island. A gene similar to esp has recently been found in Enterococcus faecium isolates recovered from hospitalized patients. In the present study we analyzed the polymorphism in the esp gene of E. faecium, and we investigated the association of esp with neighboring chromosomal genes. The esp gene showed considerable sequence heterogeneity in the regions encoding the nonrepeat N- and C-terminal domains of the Esp protein as well as differences in the number of repeats. DNA sequencing of chromosomal regions flanking the esp gene of E. faecium revealed seven open reading frames, representing putative genes implicated in virulence, regulation of transcription, and antibiotic resistance. These flanking regions were invariably associated with the presence or absence of the esp gene in E. faecium, indicating that esp in E. faecium is part of a distinct genetic element. Because of the presence of virulence genes in this gene cluster, the lower G+C content relative to that of the genome, and the presence of esp in E. faecium isolates associated with nosocomial outbreaks and clinically documented infections, we conclude that this genetic element constitutes a putative pathogenicity island, the first one described in E. faecium. Except for the presence of esp and araC, this pathogenicity island is completely different from the esp-containing pathogenicity island previously disclosed in E. faecalis.

  14. Powerful identification of cis-regulatory SNPs in human primary monocytes using allele-specific gene expression.

    Directory of Open Access Journals (Sweden)

    Jonas Carlsson Almlöf

    Full Text Available A large number of genome-wide association studies have been performed during the past five years to identify associations between SNPs and human complex diseases and traits. The assignment of a functional role for the identified disease-associated SNP is not straight-forward. Genome-wide expression quantitative trait locus (eQTL analysis is frequently used as the initial step to define a function while allele-specific gene expression (ASE analysis has not yet gained a wide-spread use in disease mapping studies. We compared the power to identify cis-acting regulatory SNPs (cis-rSNPs by genome-wide allele-specific gene expression (ASE analysis with that of traditional expression quantitative trait locus (eQTL mapping. Our study included 395 healthy blood donors for whom global gene expression profiles in circulating monocytes were determined by Illumina BeadArrays. ASE was assessed in a subset of these monocytes from 188 donors by quantitative genotyping of mRNA using a genome-wide panel of SNP markers. The performance of the two methods for detecting cis-rSNPs was evaluated by comparing associations between SNP genotypes and gene expression levels in sample sets of varying size. We found that up to 8-fold more samples are required for eQTL mapping to reach the same statistical power as that obtained by ASE analysis for the same rSNPs. The performance of ASE is insensitive to SNPs with low minor allele frequencies and detects a larger number of significantly associated rSNPs using the same sample size as eQTL mapping. An unequivocal conclusion from our comparison is that ASE analysis is more sensitive for detecting cis-rSNPs than standard eQTL mapping. Our study shows the potential of ASE mapping in tissue samples and primary cells which are difficult to obtain in large numbers.

  15. A cis-regulatory antisense RNA represses translation in Vibrio cholerae through extensive complementarity and proximity to the target locus.

    Science.gov (United States)

    Chang, Howard; Replogle, John Michael; Vather, Naomi; Tsao-Wu, Maya; Mistry, Ronak; Liu, Jane M

    2015-01-01

    As with all facultative pathogens, Vibrio cholerae must optimize its cellular processes to adapt to different environments with varying carbon sources and to environmental stresses. More specifically, in order to metabolize mannitol, V. cholerae must regulate the synthesis of MtlA, a mannitol transporter protein produced exclusively in the presence of mannitol. We previously showed that a cis-acting small RNA (sRNA) expressed by V. cholerae, MtlS, appears to post-transcriptionally downregulate the expression of mtlA and is produced in the absence of mannitol. We hypothesized that since it is complementary to the 5' untranslated region (UTR) of mtlA mRNA, MtlS may affect synthesis of MtlA by forming an mtlA-MtlS complex that blocks translation of the mRNA through occlusion of its ribosome binding site. To test this hypothesis, we used in vitro translation assays in order to examine the role MtlS plays in mtlA regulation and found that MtlS is sufficient to suppress translation of transcripts harboring the 5' UTR of mtlA. However, in a cellular context, the 5' UTR of mtlA is not sufficient for targeted repression by endogenous MtlS; additional segments from the coding region of mtlA play a role in the ability of the sRNA to regulate translation of mtlA mRNA. Additionally, proximity of transcription sites between the sRNA and mRNA significantly affects the efficacy of MtlS.

  16. The meta-epigenomic structure of purified human stem cell populations is defined at cis-regulatory sequences

    Science.gov (United States)

    Zhao, Yong Mei; Golden, Aaron; Mar, Jessica C.; Einstein, Francine H.; Greally, John M.

    2014-01-01

    The mechanism and significance of epigenetic variability in the same cell type between healthy individuals are not clear. Here, we purify human CD34+ hematopoietic stem and progenitor cells (HSPCs) from different individuals and find that there is increased variability of DNA methylation at loci with properties of promoters and enhancers. The variability is especially enriched at candidate enhancers near genes transitioning between silent and expressed states, and encoding proteins with leukocyte differentiation properties. Our findings of increased variability at loci with intermediate DNA methylation values, at candidate “poised” enhancers, and at genes involved in HSPC lineage commitment suggest that CD34+ cell subtype heterogeneity between individuals is a major mechanism for the variability observed. Epigenomic studies performed on cell populations, even when purified, are testing collections of epigenomes, or meta-epigenomes. Our findings show that meta-epigenomic approaches to data analysis can provide insights into cell subpopulation structure. PMID:25327398

  17. The Role of Cis-Regulatory Motifs and Genetical Control of Expression in the Divergence of Yeast Duplicate Genes

    National Research Council Canada - National Science Library

    Leach, Lindsey J; Zhang, Ze; Lu, Chenqi; Kearsey, Michael J; Luo, Zewei

    2007-01-01

    Expression divergence of duplicate genes is widely believed to be important for their retention and evolution of new function, although the mechanism that determines their expression divergence remains unclear...

  18. A Survey of 6,300 Genomic Fragments for cis-Regulatory Activity in the Imaginal Discs of Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Aurélie Jory

    2012-10-01

    Full Text Available Over 6,000 fragments from the genome of Drosophila melanogaster were analyzed for their ability to drive expression of GAL4 reporter genes in the third-instar larval imaginal discs. About 1,200 reporter genes drove expression in the eye, antenna, leg, wing, haltere, or genital imaginal discs. The patterns ranged from large regions to individual cells. About 75% of the active fragments drove expression in multiple discs; 20% were expressed in ventral, but not dorsal, discs (legs, genital, and antenna, whereas ∼23% were expressed in dorsal but not ventral discs (wing, haltere, and eye. Several patterns, for example, within the leg chordotonal organ, appeared a surprisingly large number of times. Unbiased searches for DNA sequence motifs suggest candidate transcription factors that may regulate enhancers with shared activities. Together, these expression patterns provide a valuable resource to the community and offer a broad overview of how transcriptional regulatory information is distributed in the Drosophila genome.

  19. PELTIER ELEMENTS

    CERN Document Server

    Tani, Laurits

    2015-01-01

    To control Peltier elements, temperature controller was used. I used TEC-1091 that was manufactured my Meerstetter Engineering. To gain control with the temperature controller, software had to be intalled on a controlling PC. There were different modes to control the Peltier: Tempererature controller to control temperature, Static current/voltage to control voltage and current and LIVE ON/OFF to auto-tune the controller respectively to the system. Also, since near the collision pipe there is much radiation, radiation-proof Peltier elements have to be used. To gain the best results, I had to find the most efficient Peltier elements and try to get their cold side to -40 degrees Celsius.

  20. Two-Component Signal Transduction Systems of Desulfovibrio Vulgaris: Structural and Phylogenetic Analysis and Deduction of Putative Cognate Pairs

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Weiwen; Culley, David E.; Wu, Gang; Brockman, Fred J.

    2006-01-20

    ABSTRACT-Two-component signal transduction systems (TCSTS) composed of sensory histidine kinases (HK) and response regulators (RR), constitute a key element of the mechanism by which bacteria sense and respond to changes in environments. A large number of TCSTSs including 59 putative HKs and 55 RRs were identified from the Desulfovibrio vulgaris genome, indicating their important roles in regulation of cellular metabolism. In this study, the structural and phylogenetic analysis of all putative TCSTSs in D. vulgaris was performed. The results showed D. vulgaris contained an unexpectedly large number of hybrid-type HKs, implying that multiple-step phosphorelay may be a common signal transduction mechanism in D. vulgaris. Most TCSTS components of D. vulgaris were found clustered into several subfamilies previously recognized in other bacteria and extensive co-evolution between D. vulgaris HKs and RRs was observed, suggesting that the concordance of HKs and RRs in cognate phylogenetic groups could be indicative of cognate TCSTSs...

  1. Element Research.

    Science.gov (United States)

    Herald, Christine

    2001-01-01

    Describes a research assignment for 8th grade students on the elements of the periodic table. Students use web-based resources and a chemistry handbook to gather information, construct concept maps, and present the findings to the full class using the mode of their choice: a humorous story, a slideshow or gameboard, a brochure, a song, or skit.…

  2. Antiviral Activities and Putative Identification of Compounds in Microbial Extracts from the Hawaiian Coastal Waters

    Directory of Open Access Journals (Sweden)

    Yuanan Lu

    2012-02-01

    Full Text Available Marine environments are a rich source of significant bioactive compounds. The Hawaiian archipelago, located in the middle of the Pacific Ocean, hosts diverse microorganisms, including many endemic species. Thirty-eight microbial extracts from Hawaiian coastal waters were evaluated for their antiviral activity against four mammalian viruses including herpes simplex virus type one (HSV-1, vesicular stomatitis virus (VSV, vaccinia virus and poliovirus type one (poliovirus-1 using in vitro cell culture assay. Nine of the 38 microbial crude extracts showed antiviral potencies and three of these nine microbial extracts exhibited significant activity against the enveloped viruses. A secosteroid, 5α(H,17α(H,(20R-beta-acetoxyergost-8(14-ene was putatively identified and confirmed to be the active compound in these marine microbial extracts. These results warrant future in-depth tests on the isolation of these active elements in order to explore and validate their antiviral potential as important therapeutic remedies.

  3. Putative melatonin receptors in a human biological clock

    Energy Technology Data Exchange (ETDEWEB)

    Reppert, S.M.; Weaver, D.R.; Rivkees, S.A.; Stopa, E.G.

    1988-10-07

    In vitro autoradiography with /sup 125/I-labeled melatonin was used to examine melatonin binding sites in human hypothalamus. Specific /sup 125/I-labeled melatonin binding was localized to the suprachiasmatic nuclei, the site of a putative biological clock, and was not apparent in other hypothalamic regions. Specific /sup 125/I-labeled melatonin binding was consistently found in the suprachiasmatic nuclei of hypothalami from adults and fetuses. Densitometric analysis of competition experiments with varying concentrations of melatonin showed monophasic competition curves, with comparable half-maximal inhibition values for the suprachiasmatic nuclei of adults (150 picomolar) and fetuses (110 picomolar). Micromolar concentrations of the melatonin agonist 6-chloromelatonin completely inhibited specific /sup 125/I-labeled melatonin binding, whereas the same concentrations of serotonin and norepinephrine caused only a partial reduction in specific binding. The results suggest that putative melatonin receptors are located in a human biological clock.

  4. Chromosomal Abnormalities and Putative Susceptibility Genes in Autism Spectrum Disorders

    DEFF Research Database (Denmark)

    Nielsen, Mette Gilling

    Autism spectrum disorders (ASDs) is a heterogeneous group of neurodevelopmental disorders with a significant genetic component as shown by family and twin studies. However, only a few genes have repeatedly been shown to be involved in the development of ASDs. The aim of this study has been...... to identify possible ASD susceptibility genes. Genome screens in ASD patients suggest possible susceptibility gene regions on almost every chromosome. We identified four ASD patients with chromosomal rearrangements, two of which were familial rearrangements involving one of these putative susceptibility gene......) was performed for all four patients. By combination of these methods we identified several putative susceptibility genes for ASDs. Expression patterns were established for several of these genes by Quantitative PCR (Q-PCR) or in situ hybridization and one gene was sequenced in 157 ASD patients. Our results...

  5. A putative role for apelin in the etiology of obesity.

    Science.gov (United States)

    Rayalam, Srujana; Della-Fera, Mary Anne; Krieg, Paul A; Cox, Christopher M; Robins, Allan; Baile, Clifton A

    2008-04-11

    Apelin, the endogenous ligand of the G protein-coupled APJ receptor has been shown to promote tumor angiogenesis. However, the effect of apelin on inducing angiogenesis in adipose tissue has not been investigated. In this review, we propose a putative role for apelin in promoting angiogenesis in adipose tissue. We further propose that targeting adipose tissue vasculature by blocking apelin signaling with anti-apelin antibodies will lead not only to inhibition of angiogenesis in adipose tissue but also to decreased adiposity.

  6. Isolation and Identification of Putative Oral Cancer Stem Cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Min; ZHAO Yan-Hua; TANG Xiao-Fei

    2011-01-01

    Objective: To isolate and characterize putative cancer stem cells in Tea8113 oral squmous cell carcinoma cell line. Methods: Putative cancer stem cells were isolated by limited dilution assay in Tea8113 cell line. Biological features of putative cancer stem cells were detected by MTT assay, flow cytometry, immunofluorescence, Colony Forming Efficiency assays, cell motility assay and in vivo tumor formation experiment. Results: Compared with untreated Tea8113 cells, the putative cancer stem cells proliferated more quickly and showed heteroploid cell cycle,higher G0/G1-arrested cells, higher CFE and higher expression levels of ABCG2 belonged to tumor stem cell phenotypes. The putative cancer stem cells had stronger capacity to generate tumors in vivo. Conclusion: The holoclone cells have higher proliferation and self-renewal abilities, which may be cancer stem cells existed in Tea8113 oral squmous cell carcinoma cell line.%目的:分离鉴定口腔鳞癌细胞系Tca8113中的肿瘤干细胞.方法:利用有限稀释的方法分离Tca8113细胞系中的肿瘤干细胞.通过MTT法、流式细胞技术、细胞免疫荧光、克隆形成率分析、细胞迁移能力检测和裸鼠皮下成瘤实验确定分离得到的肿瘤干细胞的生物学特点.结果:分离得到的紧密型克隆肿瘤细胞表现为异倍体样细胞周期,大部分细胞处于G0/G1期,增殖能力、克隆形成率和体外迁移能力都明显高于未分离的肿瘤细胞.紧密型克隆肿瘤细胞肿瘤干细胞标记物ABCG2表达也高于未分离的肿瘤细胞,并且具有更强的裸鼠皮下成瘤能力.结论:我们分离得到的紧密型克隆细胞具有较强的细胞增殖和自我更新能力,可能就是口腔鳞癌细胞系Tca8113中的肿瘤干细胞.

  7. The Schizosaccharomyces pombe inv1+ regulatory region is unusually large and contains redundant cis-acting elements that function in a SAGA- and Swi/Snf-dependent fashion.

    Science.gov (United States)

    Ahn, Sejin; Spatt, Dan; Winston, Fred

    2012-08-01

    The Schizosaccharomyces pombe inv1(+) gene encodes invertase, the enzyme required for hydrolysis of sucrose and raffinose. Transcription of inv1(+) is regulated by glucose levels, with transcription tightly repressed in high glucose and strongly induced in low glucose. To understand this regulation, we have analyzed the inv1(+) cis-regulatory region and the requirement for the trans-acting coactivators SAGA and Swi/Snf. Surprisingly, deletion of the entire 1-kilobase intergenic region between the inv1(+) TATA element and the upstream open reading frame SPCC191.10 does not significantly alter regulation of inv1(+) transcription. However, a longer deletion that extends through SPCC191.10 abolishes inv1(+) induction in low glucose. Additional analysis demonstrates that there are multiple, redundant regulatory regions spread over 1.5 kb 5' of inv1(+), including within SPCC191.10, that can confer glucose-mediated transcriptional regulation to inv1(+). Furthermore, SPCC191.10 can regulate inv1(+) transcription in an orientation-independent fashion and from a distance as great as 3 kb. With respect to trans-acting factors, both SAGA and Swi/Snf are recruited to SPCC191.10 and to other locations in the large inv1(+) regulatory region in a glucose-dependent fashion, and both are required for inv1(+) derepression. Taken together, these results demonstrate that inv1(+) regulation in S. pombe occurs via the use of multiple regulatory elements and that activation can occur over a great distance, even from elements within other open reading frames.

  8. Improved recombinant antibody production by CHO cells using a production enhancer DNA element with repeated transgene integration at a predetermined chromosomal site.

    Science.gov (United States)

    Kawabe, Yoshinori; Inao, Takanori; Komatsu, Shodai; Huang, Guan; Ito, Akira; Omasa, Takeshi; Kamihira, Masamichi

    2017-03-01

    Chinese hamster ovary (CHO) cells are one of the most useful host cell lines for the production of biopharmaceutical proteins. Although a series of production processes have been refined to improve protein productivity and cost performance, establishing producer cells is still time-consuming and labor-intensive. Recombinase-mediated site-specific gene integration into a predetermined chromosomal locus may enable predictable protein expression, reducing the laborious process of cell screening. We previously developed an accumulative site-specific gene integration system (AGIS) using Cre recombinase and mutated loxP sites for transgene integration and amplification in the CHO cell genome. Epigenetic modifier elements such as insulators are effective DNA cis-regulatory elements for stabilizing transgene expression. Here, we attempted to enhance transgene expression in recombinant CHO cells generated by AGIS using a production enhancer DNA element (PE) derived from the CHO genome. The PE was introduced into an expression unit for a recombinant scFv-Fc antibody. The effect on scFv-Fc productivity of PE position and orientation within the transgene was evaluated, while keeping the background chromosomal structure constant. For the optimal PE arrangement, scFv-Fc productivity was enhanced 2.6-fold compared with an expression unit without a PE. The enhancing effect of the PE on transgene expression was also observed when two or three PE-flanked expression units were inserted as tandem repeats. These results indicate that AGIS using the PE-flanked expression unit is a promising approach for establishing producer cell lines for biopharmaceutical protein production.

  9. DNA element downstream of the κB site in the Lcn2 promoter is required for transcriptional activation by IκBζ and NF-κB p50.

    Science.gov (United States)

    Kohda, Akira; Yamazaki, Soh; Sumimoto, Hideki

    2014-08-01

    The nuclear protein IκBζ activates transcription of a subset of NF-κB-dependent innate immune genes such as Lcn2 encoding the antibacterial protein lipocalin-2. IκBζ functions as a coactivator via its interaction with NF-κB p50, which contains a DNA-binding Rel-homology domain but lacks a transcriptional activation domain. However cis-regulatory elements involved in IκBζ function have remained unknown. Here, we show that, although IκBζ by itself is unable to associate with the Lcn2 promoter, IκBζ interacts with the promoter via p50 binding to the NF-κB-binding site (κB site) and the interaction also requires the pyrimidine-rich site (CCCCTC) that localizes seven bases downstream of the κB site. The pyrimidine-rich site is also essential for IκBζ-mediated activation of the Lcn2 gene. Introduction of both sites into an IκBζ-independent gene culminates in IκBζ-p50-DNA complex formation and transcriptional activation. Furthermore, spacing between the two sites is crucial for both IκBζ-DNA interaction and IκBζ-mediated gene activation. Thus, the pyrimidine-rich IκBζ-responsive site plays an essential role in productive interaction of IκBζ with the p50-DNA complex.

  10. Molecular genetics: DNA analysis of a putative dog clone.

    Science.gov (United States)

    Parker, Heidi G; Kruglyak, Leonid; Ostrander, Elaine A

    2006-03-09

    In August 2005, Lee et al. reported the first cloning of a domestic dog from adult somatic cells. This putative dog clone was the result of somatic-cell nuclear transfer from a fibroblast cell of a three-year-old male Afghan hound into a donor oocyte provided by a dog of mixed breed. In light of recent concerns regarding the creation of cloned human cell lines from the same institution, we have undertaken an independent test to determine the validity of the claims made by Lee et al..

  11. Comparative and functional analysis of cardiovascular-related genes

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, Jan-Fang; Pennacchio, Len A.

    2003-09-01

    The ability to detect putative cis-regulatory elements in cardiovascular-related genes has been accelerated by the availability of genomic sequence data from numerous vertebrate species and the recent development of comparative genomic tools. This improvement is anticipated to lead to a better understanding of the complex regulatory architecture of cardiovascular (CV) genes and how genetic variants in these non-coding regions can potentially play a role in cardiovascular disease. This manuscript reviews a recently established database dedicated to the comparative sequence analysis of 250 human CV genes of known importance, 37 of which currently contain sequence comparison data for organisms beyond those of human, mouse and rat. These data have provided a glimpse into the variety of possible insights from deep vertebrate sequence comparisons and the identification of putative gene regulatory elements.

  12. Putative cryptoendolithic life in Devonian pillow basalt, Rheinisches Schiefergebirge, Germany.

    Science.gov (United States)

    Peckmann, J; Bach, W; Behrens, K; Reitner, J

    2008-03-01

    Middle Devonian (Givetian) pillow basalt and inter-pillow breccia from the Rheinisches Schiefergebirge in Germany were found to contain putative biogenic filaments that indicate that life once proliferated within these volcanic rocks. Mineralized filaments are found in carbonate amygdules (vesicles filled by carbonate cement) in the volcanic rock, where they started to form on the internal surface of the once water-filled vesicles. Biogenicity of the filaments is indicated by (1) their size and shape resembling modern microorganisms including a constant diameter along the length of curved filaments, (2) their independence of crystal faces or cleavage planes, (3) branching patterns reminiscent of modern microorganisms, and (4) their spatial clustering and preferential occurrence close to the margin of pillows and in the inter-pillow breccias. A time lag between the deposition of pillow basalt and the activity of endoliths is revealed by the sequence of carbonate cements filling the amygdules. The putative filamentous microorganisms thrived after the formation of early fibrous rim cement, but before later equant calcite spar filled most of the remaining porosity. Microbial clay authigenesis analogous to the encrustation of prokaryotes in modern iron-rich environments led to the preservation of filaments. The filaments predominantly consist of the clay minerals chamosite and illite. Having dwelled in water-filled vesicles, the Devonian basalt-hosted filaments apparently represent cryptoendoliths. This finding suggests that a previously unrecognized niche for life exists within volcanic rock.

  13. Putative golden proportions as predictors of facial esthetics in adolescents.

    Science.gov (United States)

    Kiekens, Rosemie M A; Kuijpers-Jagtman, Anne Marie; van 't Hof, Martin A; van 't Hof, Bep E; Maltha, Jaap C

    2008-10-01

    In orthodontics, facial esthetics is assumed to be related to golden proportions apparent in the ideal human face. The aim of the study was to analyze the putative relationship between facial esthetics and golden proportions in white adolescents. Seventy-six adult laypeople evaluated sets of photographs of 64 adolescents on a visual analog scale (VAS) from 0 to 100. The facial esthetic value of each subject was calculated as a mean VAS score. Three observers recorded the position of 13 facial landmarks included in 19 putative golden proportions, based on the golden proportions as defined by Ricketts. The proportions and each proportion's deviation from the golden target (1.618) were calculated. This deviation was then related to the VAS scores. Only 4 of the 19 proportions had a significant negative correlation with the VAS scores, indicating that beautiful faces showed less deviation from the golden standard than less beautiful faces. Together, these variables explained only 16% of the variance. Few golden proportions have a significant relationship with facial esthetics in adolescents. The explained variance of these variables is too small to be of clinical importance.

  14. CRYSTAL STRUCTURE ANALYSIS OF A PUTATIVE OXIDOREDUCTASE FROM KLEBSIELLA PNEUMONIAE

    Energy Technology Data Exchange (ETDEWEB)

    Baig, M.; Brown, A.; Eswaramoorthy, S.; Swaminathan, S.

    2009-01-01

    Klebsiella pneumoniae, a gram-negative enteric bacterium, is found in nosocomial infections which are acquired during hospital stays for about 10% of hospital patients in the United States. The crystal structure of a putative oxidoreductase from K. pneumoniae has been determined. The structural information of this K. pneumoniae protein was used to understand its function. Crystals of the putative oxidoreductase enzyme were obtained by the sitting drop vapor diffusion method using Polyethylene glycol (PEG) 3350, Bis-Tris buffer, pH 5.5 as precipitant. These crystals were used to collect X-ray data at beam line X12C of the National Synchrotron Light Source (NSLS) at Brookhaven National Laboratory (BNL). The crystal structure was determined using the SHELX program and refi ned with CNS 1.1. This protein, which is involved in the catalysis of an oxidation-reduction (redox) reaction, has an alpha/beta structure. It utilizes nicotinamide adenine dinucleotide phosphate (NADP) or nicotine adenine dinucleotide (NAD) to perform its function. This structure could be used to determine the active and co-factor binding sites of the protein, information that could help pharmaceutical companies in drug design and in determining the protein’s relationship to disease treatment such as that for pneumonia and other related pathologies.

  15. A de novo 1.58 Mb deletion, including MAP2K6 and mapping 1.28 Mb upstream to SOX9, identified in a patient with Pierre Robin sequence and osteopenia with multiple fractures.

    Science.gov (United States)

    Smyk, Marta; Roeder, Elizabeth; Cheung, Sau Wai; Szafranski, Przemyslaw; Stankiewicz, Paweł

    2015-08-01

    Defects of long-range regulatory elements of dosage-sensitive genes represent an under-recognized mechanism underlying genetic diseases. Haploinsufficiency of SOX9, the gene essential for development of testes and differentiation of chondrocytes, results in campomelic dysplasia, a skeletal malformation syndrome often associated with sex reversal. Chromosomal rearrangements with breakpoints mapping up to 1.6 Mb up- and downstream to SOX9, and disrupting its distant cis-regulatory elements, have been described in patients with milder forms of campomelic dysplasia, Pierre Robin sequence, and sex reversal. We present an ∼1.58 Mb deletion mapping ∼1.28 Mb upstream to SOX9 that encompasses its putative long-range cis-regulatory element(s) and MAP2K6 in a patient with Pierre Robin sequence and osteopenia with multiple fractures. Low bone mass panel testing using massively parallel sequencing of 23 nuclear genes, including COL1A1 and COL1A2 was negative. Based on the previous mouse model of Map2k6, suggesting that Sox9 is likely a downstream target of the p38 MAPK pathway, and our previous chromosome conformation capture-on-chip (4C) data showing potential interactions between SOX9 promoter and MAP2K6, we hypothesize that deletion of MAP2K6 might have affected SOX9 expression and contributed to our patient's phenotype.

  16. Identification and Characterization of a Putative Manganese Export Protein in Vibrio cholerae.

    Science.gov (United States)

    Fisher, Carolyn R; Wyckoff, Elizabeth E; Peng, Eric D; Payne, Shelley M

    2016-10-15

    Manganese plays an important role in the cellular physiology and metabolism of bacterial species, including the human pathogen Vibrio cholerae The intracellular level of manganese ions is controlled through coordinated regulation of the import and export of this element. We have identified a putative manganese exporter (VC0022), named mneA (manganese exporter A), which is highly conserved among Vibrio spp. An mneA mutant exhibited sensitivity to manganese but not to other cations. Under high-manganese conditions, the mneA mutant showed an almost 50-fold increase in intracellular manganese levels and reduced intracellular iron relative to those of its wild-type parent, suggesting that the mutant's manganese sensitivity is due to the accumulation of toxic levels of manganese and reduced iron. Expression of mneA suppressed the manganese-sensitive phenotype of an Escherichia coli strain carrying a mutation in the nonhomologous manganese export gene, mntP, further supporting a manganese export function for V. cholerae MneA. The level of mneA mRNA was induced approximately 2.5-fold after addition of manganese to the medium, indicating regulation of this gene by manganese. This study offers the first insights into understanding manganese homeostasis in this important pathogen. Bacterial cells control intracellular metal concentrations by coordinating acquisition in metal-limited environments with export in metal-excess environments. We identified a putative manganese export protein, MneA, in Vibrio cholerae An mneA mutant was sensitive to manganese, and this effect was specific to manganese. The mneA mutant accumulated high levels of intracellular manganese with a concomitant decrease in intracellular iron levels when grown in manganese-supplemented medium. Expression of mneA in trans suppressed the manganese sensitivity of an E. coli mntP mutant. This study is the first to investigate manganese export in V. cholerae. Copyright © 2016, American Society for Microbiology

  17. Identification of a novel variant of staphylococcal cassette chromosome mec, type II.5, and Its truncated form by insertion of putative conjugative transposon Tn6012.

    Science.gov (United States)

    Han, Xiao; Ito, Teruyo; Takeuchi, Fumihiko; Ma, Xiao Xue; Takasu, Michihiko; Uehara, Yoshio; Oliveira, Duarte C; de Lencastre, Hermínia; Hiramatsu, Keiichi

    2009-06-01

    We identified two novel staphylococcal cassette chromosome mec (SCCmec) elements in sequence type 8 methicillin-resistant Staphylococcus aureus strains isolated in Japan: type II.5 SCCmec, whose J1 region was highly homologous to that of type I.2 SCCmec of strain PL72 (previously isolated in Poland), and its J1 region variant caused by the deletion/insertion of putative conjugative transposon Tn6012, identified in four S. aureus genomes.

  18. Exceptional error minimization in putative primordial genetic codes

    Directory of Open Access Journals (Sweden)

    Koonin Eugene V

    2009-11-01

    Full Text Available Abstract Background The standard genetic code is redundant and has a highly non-random structure. Codons for the same amino acids typically differ only by the nucleotide in the third position, whereas similar amino acids are encoded, mostly, by codon series that differ by a single base substitution in the third or the first position. As a result, the code is highly albeit not optimally robust to errors of translation, a property that has been interpreted either as a product of selection directed at the minimization of errors or as a non-adaptive by-product of evolution of the code driven by other forces. Results We investigated the error-minimization properties of putative primordial codes that consisted of 16 supercodons, with the third base being completely redundant, using a previously derived cost function and the error minimization percentage as the measure of a code's robustness to mistranslation. It is shown that, when the 16-supercodon table is populated with 10 putative primordial amino acids, inferred from the results of abiotic synthesis experiments and other evidence independent of the code's evolution, and with minimal assumptions used to assign the remaining supercodons, the resulting 2-letter codes are nearly optimal in terms of the error minimization level. Conclusion The results of the computational experiments with putative primordial genetic codes that contained only two meaningful letters in all codons and encoded 10 to 16 amino acids indicate that such codes are likely to have been nearly optimal with respect to the minimization of translation errors. This near-optimality could be the outcome of extensive early selection during the co-evolution of the code with the primordial, error-prone translation system, or a result of a unique, accidental event. Under this hypothesis, the subsequent expansion of the code resulted in a decrease of the error minimization level that became sustainable owing to the evolution of a high

  19. Basal ganglia calcification as a putative cause for cognitive decline

    Directory of Open Access Journals (Sweden)

    João Ricardo Mendes de Oliveira

    Full Text Available ABSTRACT Basal ganglia calcifications (BGC may be present in various medical conditions, such as infections, metabolic, psychiatric and neurological diseases, associated with different etiologies and clinical outcomes, including parkinsonism, psychosis, mood swings and dementia. A literature review was performed highlighting the main neuropsychological findings of BGC, with particular attention to clinical reports of cognitive decline. Neuroimaging studies combined with neuropsychological analysis show that some patients have shown progressive disturbances of selective attention, declarative memory and verbal perseveration. Therefore, the calcification process might represent a putative cause for dementia syndromes, suggesting a probable link among calcinosis, the aging process and eventually with neuronal death. The increasing number of reports available will foster a necessary discussion about cerebral calcinosis and its role in determining symptomatology in dementia patients

  20. Probing the putative active site of YjdL

    DEFF Research Database (Denmark)

    Jensen, Johanne Mørch; Ismat, Fouzia; Szakonyi, Gerda;

    2012-01-01

    YjdL from E. coli is an unusual proton-coupled oligopeptide transporter (POT). Unlike prototypical POTs, dipeptides are preferred over tripeptides, in particular dipeptides with a positively charged C-terminal residue. To further understand this difference in peptide specificity, the sequences...... of YjdL and YdgR, a prototypical E. coli POT, were compared in light of the crystal structure of a POT from Shewanella oneidensis. Several residues found in the putative active site were mutated and the activities of the mutated variants were assessed in terms of substrate uptake assays, and changes...... pocket that opens towards the extracellular space. The C-terminal side chain faces in the opposite direction into a sub pocket that faces the cytoplasm. These data indicated a stabilizing effect on a bulky N-terminal residue by an Ala281Phe variant and on the dipeptide backbone by Trp278...

  1. Novel putative mechanisms to link circadian clocks to healthy aging.

    Science.gov (United States)

    Popa-Wagner, Aurel; Catalin, Bogdan; Buga, Ana-Maria

    2015-08-01

    The circadian clock coordinates the internal physiology to increase the homeostatic capacity thereby providing both a survival advantage to the system and an optimization of energy budgeting. Multiple-oscillator circadian mechanisms are likely to play a role in regulating human health and may contribute to the aging process. Our aim is to give an overview of how the central clock in the hypothalamus and peripheral clocks relate to aging and metabolic disorders, including hyperlipidemia and hyperglycemia. In particular, we unravel novel putative mechanisms to link circadian clocks to healthy aging. This review may lead to the design of large-scale interventions to help people stay healthy as they age by adjusting daily activities, such as feeding behavior, and or adaptation to age-related changes in individual circadian rhythms.

  2. Ballistic gelatin as a putative substrate for EEG phantom devices

    CERN Document Server

    Hairston, W David; Yu, Alfred B

    2016-01-01

    Phantom devices allow the human variable to be controlled for in order to allow clear comparison and validation of biomedical imaging hardware and software. There is currently no standard phantom for electroencephalography (EEG). To be useful, such a device would need to: (a) accurately recreate the real and imaginary components of scalp electrical impedance, (b) contain internal emitters to create electrical dipoles, and (c) be easily replicable across various labs and research groups. Cost-effective materials, which are conductive, repeatable, and easily formed are a missing key enabler for EEG phantoms. Here, we explore the use of ballistics gelatin, an inexpensive, easily-formable and repeatable material, as a putative substrate by examining its electrical properties and physical stability over time. We show that varied concentrations of NaCl salt relative to gelatin powder shifts the phase/frequency response profile, allowing for selective tuning of the material electrical properties.

  3. Putative benefits of microalgal astaxanthin on exercise and human health

    Directory of Open Access Journals (Sweden)

    Marcelo P. Barros

    2011-04-01

    Full Text Available Astaxanthin (ASTA is a pinkish-orange carotenoid produced by microalgae, but also commonly found in shrimp, lobster and salmon, which accumulate ASTA from the aquatic food chain. Numerous studies have addressed the benefits of ASTA for human health, including the inhibition of LDL oxidation, UV-photoprotection and prophylaxis of bacterial stomach ulcers. ASTA is recognized as a powerful scavenger of reactive oxygen species (ROS, especially those involved in lipid peroxidation. Both aerobic and anaerobic exercise are closely related to overproduction of ROS in muscle tissue. Post-exercise inflammatory processes can even exacerbate the oxidative stress imposed by exercise. Thus, ASTA is suggested here as a putative nutritional alternative/coadjutant for antioxidant therapy to afford additional protection to muscle tissues against oxidative damage induced by exercise, as well as for an (overall integrative redox re-balance and general human health.

  4. Cryptic species in putative ancient asexual darwinulids (Crustacea, Ostracoda.

    Directory of Open Access Journals (Sweden)

    Isa Schön

    Full Text Available BACKGROUND: Fully asexually reproducing taxa lack outcrossing. Hence, the classic Biological Species Concept cannot be applied. METHODOLOGY/PRINCIPAL FINDINGS: We used DNA sequences from the mitochondrial COI gene and the nuclear ITS2 region to check species boundaries according to the evolutionary genetic (EG species concept in five morphospecies in the putative ancient asexual ostracod genera, Penthesilenula and Darwinula, from different continents. We applied two methods for detecting cryptic species, namely the K/θ method and the General Mixed Yule Coalescent model (GMYC. We could confirm the existence of species in all five darwinulid morphospecies and additional cryptic diversity in three morphospecies, namely in Penthesilenula brasiliensis, Darwinula stevensoni and in P. aotearoa. The number of cryptic species within one morphospecies varied between seven (P. brasiliensis, five to six (D. stevensoni and two (P. aotearoa, respectively, depending on the method used. Cryptic species mainly followed continental distributions. We also found evidence for coexistence at the local scale for Brazilian cryptic species of P. brasiliensis and P. aotearoa. Our ITS2 data confirmed that species exist in darwinulids but detected far less EG species, namely two to three cryptic species in P. brasiliensis and no cryptic species at all in the other darwinulid morphospecies. CONCLUSIONS/SIGNIFICANCE: Our results clearly demonstrate that both species and cryptic diversity can be recognized in putative ancient asexual ostracods using the EG species concept, and that COI data are more suitable than ITS2 for this purpose. The discovery of up to eight cryptic species within a single morphospecies will significantly increase estimates of biodiversity in this asexual ostracod group. Which factors, other than long-term geographic isolation, are important for speciation processes in these ancient asexuals remains to be investigated.

  5. Putative regulatory factors associated with intramuscular fat content.

    Directory of Open Access Journals (Sweden)

    Aline S M Cesar

    Full Text Available Intramuscular fat (IMF content is related to insulin resistance, which is an important prediction factor for disorders, such as cardiovascular disease, obesity and type 2 diabetes in human. At the same time, it is an economically important trait, which influences the sensorial and nutritional value of meat. The deposition of IMF is influenced by many factors such as sex, age, nutrition, and genetics. In this study Nellore steers (Bos taurus indicus subspecies were used to better understand the molecular mechanisms involved in IMF content. This was accomplished by identifying differentially expressed genes (DEG, biological pathways and putative regulatory factors. Animals included in this study had extreme genomic estimated breeding value (GEBV for IMF. RNA-seq analysis, gene set enrichment analysis (GSEA and co-expression network methods, such as partial correlation coefficient with information theory (PCIT, regulatory impact factor (RIF and phenotypic impact factor (PIF were utilized to better understand intramuscular adipogenesis. A total of 16,101 genes were analyzed in both groups (high (H and low (L GEBV and 77 DEG (FDR 10% were identified between the two groups. Pathway Studio software identified 13 significantly over-represented pathways, functional classes and small molecule signaling pathways within the DEG list. PCIT analyses identified genes with a difference in the number of gene-gene correlations between H and L group and detected putative regulatory factors involved in IMF content. Candidate genes identified by PCIT include: ANKRD26, HOXC5 and PPAPDC2. RIF and PIF analyses identified several candidate genes: GLI2 and IGF2 (RIF1, MPC1 and UBL5 (RIF2 and a host of small RNAs, including miR-1281 (PIF. These findings contribute to a better understanding of the molecular mechanisms that underlie fat content and energy balance in muscle and provide important information for the production of healthier beef for human consumption.

  6. Phylum-Level Conservation of Regulatory Information in Nematodes despite Extensive Non-coding Sequence Divergence

    Science.gov (United States)

    Gordon, Kacy L.; Arthur, Robert K.; Ruvinsky, Ilya

    2015-01-01

    Gene regulatory information guides development and shapes the course of evolution. To test conservation of gene regulation within the phylum Nematoda, we compared the functions of putative cis-regulatory sequences of four sets of orthologs (unc-47, unc-25, mec-3 and elt-2) from distantly-related nematode species. These species, Caenorhabditis elegans, its congeneric C. briggsae, and three parasitic species Meloidogyne hapla, Brugia malayi, and Trichinella spiralis, represent four of the five major clades in the phylum Nematoda. Despite the great phylogenetic distances sampled and the extensive sequence divergence of nematode genomes, all but one of the regulatory elements we tested are able to drive at least a subset of the expected gene expression patterns. We show that functionally conserved cis-regulatory elements have no more extended sequence similarity to their C. elegans orthologs than would be expected by chance, but they do harbor motifs that are important for proper expression of the C. elegans genes. These motifs are too short to be distinguished from the background level of sequence similarity, and while identical in sequence they are not conserved in orientation or position. Functional tests reveal that some of these motifs contribute to proper expression. Our results suggest that conserved regulatory circuitry can persist despite considerable turnover within cis elements. PMID:26020930

  7. Phylum-Level Conservation of Regulatory Information in Nematodes despite Extensive Non-coding Sequence Divergence.

    Directory of Open Access Journals (Sweden)

    Kacy L Gordon

    2015-05-01

    Full Text Available Gene regulatory information guides development and shapes the course of evolution. To test conservation of gene regulation within the phylum Nematoda, we compared the functions of putative cis-regulatory sequences of four sets of orthologs (unc-47, unc-25, mec-3 and elt-2 from distantly-related nematode species. These species, Caenorhabditis elegans, its congeneric C. briggsae, and three parasitic species Meloidogyne hapla, Brugia malayi, and Trichinella spiralis, represent four of the five major clades in the phylum Nematoda. Despite the great phylogenetic distances sampled and the extensive sequence divergence of nematode genomes, all but one of the regulatory elements we tested are able to drive at least a subset of the expected gene expression patterns. We show that functionally conserved cis-regulatory elements have no more extended sequence similarity to their C. elegans orthologs than would be expected by chance, but they do harbor motifs that are important for proper expression of the C. elegans genes. These motifs are too short to be distinguished from the background level of sequence similarity, and while identical in sequence they are not conserved in orientation or position. Functional tests reveal that some of these motifs contribute to proper expression. Our results suggest that conserved regulatory circuitry can persist despite considerable turnover within cis elements.

  8. Reference: OCTAMERMOTIFTAH3H4 [PLACE

    Lifescience Database Archive (English)

    Full Text Available OCTAMERMOTIFTAH3H4 Nakayama T, Sakamoto A, Yang P, Minami M, Fujimoto Y, Ito T, Iwa...buchi M Highly conserved hexamer, octamer and nonamer motifs are positive cis-regulatory elements of the whe

  9. Reference: SRENTTTO1 [PLACE

    Lifescience Database Archive (English)

    Full Text Available SRENTTTO1 Takeda S, Sugimoto K, Otsuki H, Hirochika H A 13-bp cis-regulatory elemen...t in the LTR promoter of the tobacco retrotransposon Tto1 is involved in responsiveness to tissue culture, w

  10. Dynamic regulation of the transcription initiation landscape at single nucleotide resolution during vertebrate embryogenesis

    NARCIS (Netherlands)

    C. Nepal (Chirag); Y. Hadzhiev (Yavor); C. Previti (Christopher); V. Haberle (Vanja); N. Li (Nan); H. Takahashi (Hiroyuki); A.M. Suzuki (Ana Maria); Y. Sheng (Ying); R.F. Abdelhamid (Rehab); S. Anand (Santosh); P.A. Gehrig (Paola A.); A. Akalin (Altuna); C. Kockx (Christel); A. Van Der Sloot (Antoine); W.F.J. van IJcken (Wilfred); O. Armant (Olivier); S. Rastegar (Sepand); C. Watson (Craig); U. Strähle (Uwe); E. Stupka (Elia); P. Carninci (Piero); B. Lenhard (Boris); F. Müller (Ferenc)

    2013-01-01

    textabstractSpatiotemporal control of gene expression is central to animal development. Core promoters represent a previously unanticipated regulatory level by interacting with cis-regulatory elements and transcription initiation in different physiological and developmental contexts. Here, we provid

  11. Functional Characterization of a Putative Nitrate Transporter Gene Promoter from Rice

    Institute of Scientific and Technical Information of China (English)

    Ting-Zhang HU; Kai-Ming CAO; Mian XIA; Xi-Ping WANG

    2006-01-01

    Drought is one of the most significant abiotic stresses that influence plant growth and development. Expression analysis revealed that OsNRT1.3, a putative nitrate transporter gene in rice, was induced by drought. To confirm if the OsNRT1.3 promoter can respond to drought stress, a 2019 bp upstream sequence of OsNRT1.3 was cloned. Three OsNRT1.3 promoter fragments were generated by 5'-deletion, and fused to the β-glucuronidase (GUS) gene. The chimeric genes were introduced into rice plants. NRT2019::GUS, NRT1196: :GUS and NRT719::GUS showed similar expression patterns in seeds,roots, leaves and flowers in all transgenic rice, and GUS activity conferred by different OsNRT1.3 promoter fragments was significantly upregulated by drought stress, indicating that OsNRT1.3 promoter responds to drought stress and the 719 bp upstream sequence of OsNRT1.3 contains the drought response elements.

  12. Putative uremic encephalopathy in horses: five cases (1978-1998).

    Science.gov (United States)

    Frye, M A; Johnson, J S; Traub-Dargatz, J L; Savage, C J; Fettman, M J; Gould, D H

    2001-02-15

    To determine historical, physical examination, clinicopathologic, and postmortem findings in horses with putative uremic encephalopathy. Design-Retrospective study. Animals-5 horses with renal failure and neurologic disease not attributable to abnormalities in any other organ system. Medical records from 1978 to 1998 were examined for horses with renal disease and neurologic signs not attributable to primary neurologic, hepatic, or other diseases. Signalment, history, physical examination findings, clinicopathologic data, renal ultrasonographic findings, and postmortem data were reviewed. Of 332 horses with renal disease, 5 met selection criteria. Historical findings, physical examination findings, clinicopathologic data, ultrasonographic data, and postmortem findings were consistent with chronic renal failure. Swollen astrocytes were detected in all 4 horses examined at necropsy. A single criterion was not determined to be pathognomonic for uremic encephalopathy in horses. Uremic encephalopathy should be considered as a differential diagnosis in horses with evidence of chronic renal failure and encephalopathic neurologic sign not attributable to other causes. Astrocyte swelling, which was common to all 4 horses examined at necropsy, may serve as a microscopic indicator of uremic encephalopathy in horses.

  13. Phytophthora infestans specific phosphorylation patterns and new putative control targets.

    Science.gov (United States)

    Frades, Itziar; Andreasson, Erik

    2016-04-01

    In this study we applied biomathematical searches of gene regulatory mechanisms to learn more about oomycete biology and to identify new putative targets for pesticides or biological control against Phytophthora infestans. First, oomycete phylum-specific phosphorylation motifs were found by discriminative n-gram analysis. We found 11.600 P. infestans specific n-grams, mapping 642 phosphoproteins. The most abundant group among these related to phosphatidylinositol metabolism. Due to the large number of possible targets found and our hypothesis that multi-level control is a sign of usefulness as targets for intervention, we identified overlapping targets with a second screen. This was performed to identify proteins dually regulated by small RNA and phosphorylation. We found 164 proteins to be regulated by both sRNA and phosphorylation and the dominating functions where phosphatidylinositol signalling/metabolism, endocytosis, and autophagy. Furthermore we performed a similar regulatory study and discriminative n-gram analysis of proteins with no clear orthologs in other species and proteins that are known to be unique to P. infestans such as the RxLR effectors, Crinkler (CRN) proteins and elicitins. We identified CRN proteins with specific phospho-motifs present in all life stages. PITG_12626, PITG_14042 and PITG_23175 are CRN proteins that have species-specific phosphorylation motifs and are subject to dual regulation.

  14. Rapid Discrimination Among Putative Mechanistic Models of Biochemical Systems.

    Science.gov (United States)

    Lomnitz, Jason G; Savageau, Michael A

    2016-08-31

    An overarching goal in molecular biology is to gain an understanding of the mechanistic basis underlying biochemical systems. Success is critical if we are to predict effectively the outcome of drug treatments and the development of abnormal phenotypes. However, data from most experimental studies is typically noisy and sparse. This allows multiple potential mechanisms to account for experimental observations, and often devising experiments to test each is not feasible. Here, we introduce a novel strategy that discriminates among putative models based on their repertoire of qualitatively distinct phenotypes, without relying on knowledge of specific values for rate constants and binding constants. As an illustration, we apply this strategy to two synthetic gene circuits exhibiting anomalous behaviors. Our results show that the conventional models, based on their well-characterized components, cannot account for the experimental observations. We examine a total of 40 alternative hypotheses and show that only 5 have the potential to reproduce the experimental data, and one can do so with biologically relevant parameter values.

  15. Putative role of Tat-Env interaction in HIV infection.

    Science.gov (United States)

    Poon, Selina; Moscoso, Carlos G; Xing, Li; Kan, Elaine; Sun, Yide; Kolatkar, Prasanna R; Vahlne, Anders G; Srivastava, Indresh K; Barnett, Susan W; Cheng, R Holland

    2013-09-24

    To study the complex formed between Tat protein and Env soluble trimeric immunogen, and compare with previously determined structures of Env native trimers and Env-CD4m complexes. The soluble Env trimer was used to mimic the spike glycoprotein on the virus surface for the study. To overcome limitations of other structural determination methods, cryoelectron microscopy was employed to image the complex, and single particle reconstruction was utilized to reconstruct the structure of the complex from collected micrographs. Molecular modeling of gp120-Tat was performed to provide atomic coordinates for docking. Images were preprocessed by multivariate statistical analysis to identify principal components of variation then submitted for reconstruction. Reconstructed structures were docked with modeled gp120-Tat atomic coordinates to study the positions of crucial epitopes. Analysis of the Env-Tat complex demonstrated an intermediate structure between Env native trimers and Env-CD4m structures. Docking results indicate that the CD4-binding site and the V3 loop are exposed in the Env-Tat complex. The integrin-binding sequence in Tat was also exposed in Env-Tat docking. The intermediate structure induced by Tat-interaction with Env could potentially provide an explanation for increased virus infection in the presence of Tat protein. Consequently, exposure of CD4-binding sites and a putative integrin-binding sequence on Tat in the complex may provide a new avenue for rational design of an effective HIV vaccine. © 2013 Wolters Kluwer Health | Lippincott Williams & Wilkins

  16. Phosphoglycerate Dehydrogenase: Potential Therapeutic Target and Putative Metabolic Oncogene

    Directory of Open Access Journals (Sweden)

    Cheryl K. Zogg

    2014-01-01

    Full Text Available Exemplified by cancer cells’ preference for glycolysis, for example, the Warburg effect, altered metabolism in tumorigenesis has emerged as an important aspect of cancer in the past 10–20 years. Whether due to changes in regulatory tumor suppressors/oncogenes or by acting as metabolic oncogenes themselves, enzymes involved in the complex network of metabolic pathways are being studied to understand their role and assess their utility as therapeutic targets. Conversion of glycolytic intermediate 3-phosphoglycerate into phosphohydroxypyruvate by the enzyme phosphoglycerate dehydrogenase (PHGDH—a rate-limiting step in the conversion of 3-phosphoglycerate to serine—represents one such mechanism. Forgotten since classic animal studies in the 1980s, the role of PHGDH as a potential therapeutic target and putative metabolic oncogene has recently reemerged following publication of two prominent papers near-simultaneously in 2011. Since that time, numerous studies and a host of metabolic explanations have been put forward in an attempt to understand the results observed. In this paper, I review the historic progression of our understanding of the role of PHGDH in cancer from the early work by Snell through its reemergence and rise to prominence, culminating in an assessment of subsequent work and what it means for the future of PHGDH.

  17. Small intestinal mucosa expression of putative chaperone fls485

    Directory of Open Access Journals (Sweden)

    Raupach Kerstin

    2010-03-01

    Full Text Available Abstract Background Maturation of enterocytes along the small intestinal crypt-villus axis is associated with significant changes in gene expression profiles. fls485 coding a putative chaperone protein has been recently suggested as a gene involved in this process. The aim of the present study was to analyze fls485 expression in human small intestinal mucosa. Methods fls485 expression in purified normal or intestinal mucosa affected with celiac disease was investigated with a molecular approach including qRT-PCR, Western blotting, and expression strategies. Molecular data were corroborated with several in situ techniques and usage of newly synthesized mouse monoclonal antibodies. Results fls485 mRNA expression was preferentially found in enterocytes and chromaffine cells of human intestinal mucosa as well as in several cell lines including Rko, Lovo, and CaCo2 cells. Western blot analysis with our new anti-fls485 antibodies revealed at least two fls485 proteins. In a functional CaCo2 model, an increase in fls485 expression was paralleled by cellular maturation stage. Immunohistochemistry demonstrated fls485 as a cytosolic protein with a slightly increasing expression gradient along the crypt-villus axis which was impaired in celiac disease Marsh IIIa-c. Conclusions Expression and synthesis of fls485 are found in surface lining epithelia of normal human intestinal mucosa and deriving epithelial cell lines. An interdependence of enterocyte differentiation along the crypt-villus axis and fls485 chaperone activity might be possible.

  18. Small intestinal mucosa expression of putative chaperone fls485.

    Science.gov (United States)

    Reinartz, Andrea; Ehling, Josef; Franz, Susanne; Simon, Verena; Bravo, Ignacio G; Tessmer, Claudia; Zentgraf, Hanswalter; Lyer, Stefan; Schneider, Ursula; Köster, Jan; Raupach, Kerstin; Kämmerer, Elke; Klaus, Christina; Tischendorf, Jens J W; Kopitz, Jürgen; Alonso, Angel; Gassler, Nikolaus

    2010-03-07

    Maturation of enterocytes along the small intestinal crypt-villus axis is associated with significant changes in gene expression profiles. fls485 coding a putative chaperone protein has been recently suggested as a gene involved in this process. The aim of the present study was to analyze fls485 expression in human small intestinal mucosa. fls485 expression in purified normal or intestinal mucosa affected with celiac disease was investigated with a molecular approach including qRT-PCR, Western blotting, and expression strategies. Molecular data were corroborated with several in situ techniques and usage of newly synthesized mouse monoclonal antibodies. fls485 mRNA expression was preferentially found in enterocytes and chromaffine cells of human intestinal mucosa as well as in several cell lines including Rko, Lovo, and CaCo2 cells. Western blot analysis with our new anti-fls485 antibodies revealed at least two fls485 proteins. In a functional CaCo2 model, an increase in fls485 expression was paralleled by cellular maturation stage. Immunohistochemistry demonstrated fls485 as a cytosolic protein with a slightly increasing expression gradient along the crypt-villus axis which was impaired in celiac disease Marsh IIIa-c. Expression and synthesis of fls485 are found in surface lining epithelia of normal human intestinal mucosa and deriving epithelial cell lines. An interdependence of enterocyte differentiation along the crypt-villus axis and fls485 chaperone activity might be possible.

  19. Putative impact of RNA editing on drug discovery.

    Science.gov (United States)

    Decher, Niels; Netter, Michael F; Streit, Anne K

    2013-01-01

    Virtually all organisms use RNA editing as a powerful post-transcriptional mechanism to recode genomic information and to increase functional protein diversity. The enzymatic editing of pre-mRNA by ADARs and CDARs is known to change the functional properties of neuronal receptors and ion channels regulating cellular excitability. However, RNA editing is also an important mechanism for genes expressed outside the brain. The fact that RNA editing breaks the 'one gene encodes one protein' hypothesis is daunting for scientists and a probable drawback for drug development, as scientists might search for drugs targeting the 'wrong' protein. This possible difficulty for drug discovery and development became more evident from recent publications, describing that RNA editing events have profound impact on the pharmacology of some common drug targets. These recent studies highlight that RNA editing can cause massive discrepancies between the in vitro and in vivo pharmacology. Here, we review the putative impact of RNA editing on drug discovery, as RNA editing has to be considered before using high-throughput screens, rational drug design or choosing the right model organism for target validation.

  20. Epigenetic regulation of putative tumor suppressor TGFBI in human leukemias

    Institute of Scientific and Technical Information of China (English)

    Fang Hongbo; Liu Jing; Guo Dan; Liu Peixiang; Zhao Yongliang

    2014-01-01

    Background Both in vitro and in vivo data have demonstrated the TGFBI gene functions as a putative tumor suppressor and is frequently downregulated in human tumors of different histological types.The hypermethylation of the TGFBI promoter,as one of the main regulatory mechanisms,is associated with TGFBI silencing.In this study,we used a methylation-specific PCR (MSP) method to evaluate the methylation status of the TGFBI promoter in human leukemias.Methods Real-time RT-PCR and methylation-specific PCR approaches were performed to define the TGFBI expression and promoter methylation in human leukemia call lines and clinical samples.Genomic DNA was isolated from peripheral blood mononuclear cells from leukemia patients,bisulfite-converted,and analyzed by the MSP method.Results Hypermethylation of the TGFBI promoter occurred in leukemia cell lines and demethylation treatment reexpressed TGFBI at a substantially increased level in most of leukemia cell lines tested.Furthermore,a much higher level of CpG island methylation and a significantly lower TGFBI expression were also identified in clinical leukemia samples.Conclusion The results suggest an important role of promoter methylation in regulating TGFBI expression in leukemia,which provides a useful diagnostic marker for clinical management of human leukemias.

  1. Expression and characterization of rice putative PAUSED gene

    Institute of Scientific and Technical Information of China (English)

    Chengguo Yao; Liangfa Ge; Wei Li; Botao Zhao; Chaoqun Li; Kangcheng Ruan; Hongxuan Lin; Youxin Jin

    2008-01-01

    In Arab idopsis, PA USED ( PSD ) encodes the ortholog of loslp/ exportin-t, which mediates the nuclear export of transfer RNA (tRNA) in yeast and mammals. However, in monocot plants such as rice, knowledge of the corresponding ortholog is limited, and its effects on growth development and productivity remain unknown. In this study, we verified a rice transfer-DNA insertional mutantpsd line and analyzed its phenotypes;the mutant displayed severe morphological defects including retarded development and low fertility compared with wild-type rice. Examining intronless tRNA-Tyr and intron-containing pre-tRNA-Ala expression levels in cytoplasmic and nuclear fraction with Northern blot analysis between wild -type and mutant leaf tissue suggested that rice PSD might be involved in tRNA export from the nucleus to the cytoplasm.Additionally, reverse transcription-polymerase chain reaction analysis revealed that PSD transcript was expressed throughout normal rice plant development, and subcellular localization assays showed that rice PSD protein was present in both the nucleus and cytoplasm. In summary, our data implied that the putative PSD gene might be indispensable for normal rice development and its function might be the same as that ofArabidopsis PSD.

  2. Conformational study of a putative HLTV-1 retroviral protease inhibitor.

    Science.gov (United States)

    Llido, S; d'Estaintot, B L; Dautant, A; Geoffre, S; Picard, P; Precigoux, G

    1993-05-01

    The crystal structure of prolyl-glutaminyl-valyl-statyl-alanyl-leucine (Pro-Gln-Val-Sta-Ala-Leu, C(32)H(57)N(7)0(9).5H(2)0, M(r) = 683.9 + 90.1), a putative HTLV-1 protease inhibitor based on one of the consensus retroviral protease cleavage sequences, and containing the statine residue [(4S,3S)-4-amino-3-hydroxy-6-methylheptanoic acid], has been determined by X-ray diffraction. The same molecule has been modelled in the active site of the HTLV-1 protease and both conformations have been compared. The peptide crystallizes as a pentahydrate in space group P2(1) with a = 10.874(2), b = 9.501(2), c = 21.062(5) A, beta = 103.68 (1) degrees, Z = 2, V= 2114.3 A(3), D(x) = 1.21 g cm(-3), micro = 8.02 cm(-1), T= 293 K, lambda(Cu Kalpha) = 1.5418 A. The structure has been refined to an R value of 0.070 for 2152 observed reflections. The peptide main chain can be described as extended and adopts the usual zigzag conformation from the prolyl to the statyl residue. The main difference in conformation between the individual observed and modelled molecules is located on the Sta, Ala and Leu residues with the main chain of the modelled molecule rotated by about 180 degrees as compared to the observed conformation in the crystal state.

  3. A new putative sigma factor of Myxococcus xanthus.

    Science.gov (United States)

    Apelian, D; Inouye, S

    1993-06-01

    A third putative sigma factor gene, sigC, has been isolated from Myxococcus xanthus by using the sigA gene (formerly rpoD of M. xanthus) as a probe. The nucleotide sequence of sigC has been determined, and an open reading frame of 295 residues (M(r) = 33,430) has been identified. The deduced amino acid sequence of sigC exhibits the features which are characteristic of other bacterial sigma factors. The characterization of a sigC-lacZ strain has demonstrated that sigC expression is induced immediately after cells enter into the developmental cycle and is dramatically reduced at the onset of sporulation. A deletion mutant of sigC grows normally in vegetative culture and is able to develop normally. However, in contrast to the wild-type cells, the sigC deletion mutant cells became capable of forming fruiting bodies and myxospores on semirich agar plates. This suggests that sigC may play a role in expression of genes involved in negatively regulating the initiation of fruiting body formation.

  4. Isolation and characterization of a new chemokine receptor gene, the putative chicken CXCR1.

    Science.gov (United States)

    Li, Q J; Lu, S; Ye, R D; Martins-Green, M

    2000-10-31

    This study delineates the isolation and characterization of a novel chemokine receptor gene, the putative chicken CXC receptor 1 (cCXCR1). Using a human CXCR1 probe, we isolated several positive clones from a chicken genomic library. One of the clones contained a fragment of approximately 5000bp that hybridized strongly with the hCXCR1 probe. This fragment was sequenced and subjected to a variety of computer analyses. The open reading frame for this gene predicts a seven transmembrane domain protein with all the characteristics of a chemokine receptor and with 67% sequence homology to hCXCR1, 65% to hCXCR2 and also with considerable sequence homology to other human chemokine receptors such as hCXCR4 (50%), hCCR2 (49%) and hCCR1 (49%). However, the homology to a previously isolated potential G-protein-coupled receptor for chickens (AvCRL1) is only 47%. Using 5' RACE, two transcription initiation sites were identified suggesting the potential for the expression of two protein isoforms (I and II) in vivo. The promoter for the putative cCXCR1 contains a variety of consensus transcription factor binding elements that can potentially be involved in the expression of this chicken receptor upon stimulation by stress-inducing agents. RT-PCR analysis was used to determine the pattern of expression of the larger isoform (I) of this receptor in a variety of tissues. This form of the receptor is expressed primarily in the organs of the gastrointestinal tract, tissues that are frequently exposed to stress-inducing agents, but not in the central nervous system, tissues that are protected from insult by the blood barrier. Using the same RT-PCR approach we show that stress-inducing agents, such as 'first-hand' and 'second-hand' cigarette smoke components, tumor promoters and thrombin, differentially stimulate the expression of the isoform I in primary fibroblasts. Thrombin is an enzyme that plays many important roles in thrombosis, angiogenesis and wound healing and exposure to

  5. Putative molecular mechanism underlying sperm chromatin remodelling is regulated by reproductive hormones

    Directory of Open Access Journals (Sweden)

    Gill-Sharma Manjeet Kaur

    2012-12-01

    Full Text Available Abstract Background The putative regulatory role of the male reproductive hormones in the molecular mechanism underlying chromatin condensation remains poorly understood. In the past decade, we developed two adult male rat models wherein functional deficits of testosterone or FSH, produced after treatments with 20 mg/Kg/d of cyproterone acetate (CPA per os, for a period of 15 days or 3 mg/Kg/d of fluphenazine decanoate (FD subcutaneously, for a period of 60 days, respectively, affected the rate of sperm chromatin decondensation in vitro. These rat models have been used in the current study in order to delineate the putative roles of testosterone and FSH in the molecular mechanism underlying remodelling of sperm chromatin. Results We report that deficits of both testosterone and FSH affected the turnover of polyubiquitylated histones and led to their accumulation in the testis. Functional deficits of testosterone reduced expression of MIWI, the 5-methyl cap binding RNA-binding protein (PIWIlike murine homologue of the Drosophila protein PIWI/P-element induced wimpy testis containing a PAZ/Piwi-Argonaut-Zwille domain and levels of histone deacetylase1 (HDAC1, ubiquitin ligating enzyme (URE-B1/E3, 20S proteasome α1 concomitant with reduced expression of ubiquitin activating enzyme (ube1, conjugating enzyme (ube2d2, chromodomain Y like protein (cdyl, bromodomain testis specific protein (brdt, hdac6 (histone deacetylase6, androgen-dependent homeobox placentae embryonic protein (pem/RhoX5, histones h2b and th3 (testis-specific h3. Functional deficits of FSH reduced the expression of cdyl and brdt genes in the testis, affected turnover of ubiquitylated histones, stalled the physiological DNA repair mechanism and culminated in spermiation of DNA damaged sperm. Conclusions We aver that deficits of both testosterone and FSH differentially affected the process of sperm chromatin remodelling through subtle changes in the ‘chromatin condensation

  6. The aerosols' fate in a putative ammonia ocean on Titan

    Science.gov (United States)

    Ramírez, S. I.; Coll, P.; Buch, A.; Brassé, C.; Poch, O.; Raulin, F.

    2010-04-01

    A laboratory study on the chemical transformation of Titan's aerosol analogues placed under putative surface conditions of the satellite was performed. The surface of Titan was one of the targets of the Cassini-Huygens mission and of several of the Cassini orbiter instruments, especially ISS, VIMS and Radar. The first images revealed an interesting solid surface with features that suggest aeolian, tectonic, fluvial processes and even an impact structure[1]. Since then, more detailed descriptions of dunes, channels, lakes, impact craters and cryovolcanic structures have been documented[2]. The existence of an internal liquid water ocean, containing a few percent ammonia has been proposed[2, 3]. It has also been proposed that ammonia-water mixtures can erupt from the putative subsurface ocean leading to cryovolcanism[4]. The Cassini Titan Radar Mapper obtained Synthetic Aperture Radar (SAR) images during 2004 and 2005 that revealed a highly complex geology occurring at Titan's surface[5], among which cryovolcanic features play a central role. The composition of the cryomagma is mainly proposed to be a mixture of water ice and ammonia[6, 7, 8], although ammonia has not been directly detected on Titan, but suggested by recent Cassini-VIMS observations[9]. In order to understand the role that ammonia may play on the chemical transformation of atmospheric aerosols once they reach the surface, we designed the following protocol: laboratory analogues of Titan's aerosols were synthesized from a N2:CH4 (98:2) mixture irradiated under a continuous flow regime of 845 sccm inside which, a cold plasma of 180 W was established. The synthesized analogues were recovered and partitioned in several 10.0 mg samples that were placed in 4.0 mL-volume of aqueous ammonia solutions (25.00, 12.50, 6.25 and 3.125%) at different temperatures (298, 277, 253 and 93 K) for 10 weeks. After a derivatization process performed to the aerosols' refractory phase with N

  7. Molecular diagnosis of putative Stargardt disease probands by exome sequencing

    Directory of Open Access Journals (Sweden)

    Strom Samuel P

    2012-08-01

    Full Text Available Abstract Background The commonest genetic form of juvenile or early adult onset macular degeneration is Stargardt Disease (STGD caused by recessive mutations in the gene ABCA4. However, high phenotypic and allelic heterogeneity and a small but non-trivial amount of locus heterogeneity currently impede conclusive molecular diagnosis in a significant proportion of cases. Methods We performed whole exome sequencing (WES of nine putative Stargardt Disease probands and searched for potentially disease-causing genetic variants in previously identified retinal or macular dystrophy genes. Follow-up dideoxy sequencing was performed for confirmation and to screen for mutations in an additional set of affected individuals lacking a definitive molecular diagnosis. Results Whole exome sequencing revealed seven likely disease-causing variants across four genes, providing a confident genetic diagnosis in six previously uncharacterized participants. We identified four previously missed mutations in ABCA4 across three individuals. Likely disease-causing mutations in RDS/PRPH2, ELOVL, and CRB1 were also identified. Conclusions Our findings highlight the enormous potential of whole exome sequencing in Stargardt Disease molecular diagnosis and research. WES adequately assayed all coding sequences and canonical splice sites of ABCA4 in this study. Additionally, WES enables the identification of disease-related alleles in other genes. This work highlights the importance of collecting parental genetic material for WES testing as the current knowledge of human genome variation limits the determination of causality between identified variants and disease. While larger sample sizes are required to establish the precision and accuracy of this type of testing, this study supports WES for inherited early onset macular degeneration disorders as an alternative to standard mutation screening techniques.

  8. A putatively novel form of spontaneous coordination in neural activity.

    Science.gov (United States)

    Hermer-Vazquez, Raymond; Hermer-Vazquez, Linda; Srinivasan, Sridhar

    2009-04-06

    We simultaneously recorded local field potentials from three sites along the olfactory-entorhinal axis in rats lightly anesthetized with isoflurane, as part of another experiment. While analyzing the initial data from that experiment with spectrograms, we discovered a potentially novel form of correlated neural activity, with near-simultaneous occurrence across the three widely separated brain sites. After validating their existence further, we named these events Synchronous Frequency Bursts (SFBs). Here we report our initial investigations into their properties and their potential functional significance. In Experiment 1, we found that SFBs have highly regular properties, consisting of brief (approximately 250 ms), high amplitude bursts of LFP energy spanning frequency ranges from the delta band (1-4 Hz) to at least the low gamma band (30-50 Hz). SFBs occurred almost simultaneously across recording sites, usually with onsets sites. While the SFBs had fairly typical, exponentially decaying power spectral density plots, their coherence structure was unusual, with high peaks in several narrow frequency ranges and little coherence in other bands. In Experiment 2, we found that SFBs occurred far more often under light anesthesia than deeper anesthetic states, and were especially prevalent as the animals regained consciousness. Finally, in Experiment 3 we showed that SFBs occur simultaneously at a significant rate across brain sites from putatively different functional subsystems--olfactory versus motor pathways. We suggest that SFBs do not carry information per se, but rather, play a role in coordinating activity in different frequency bands, potentially brain-wide, as animals progress from sleep or anesthesia toward full consciousness.

  9. Tissue factor residues that putatively interact with membrane phospholipids.

    Directory of Open Access Journals (Sweden)

    Ke Ke

    Full Text Available Blood clotting is initiated by the two-subunit enzyme consisting of the plasma protease, factor VIIa (the catalytic subunit, bound to the integral membrane protein, tissue factor (the regulatory subunit. Molecular dynamics simulations have predicted that certain residues in the tissue factor ectodomain interact with phosphatidylserine headgroups to ensure optimal positioning of the tissue factor/factor VIIa complex relative to its membrane-bound protein substrates, factors IX and X. In this study, we individually mutated to alanine all the putative phosphatidylserine-interactive residues in the tissue factor ectodomain and measured their effects on tissue factor cofactor function (activation of factors IX and X by tissue factor/factor VIIa, and clotting of plasma. Some tissue factor mutants exhibited decreased activity in all three assays, with the most profound defects observed from mutations in or near the flexible loop from Lys159 to Gly164. The decreased activity of all of these tissue factor mutants could be partially or completely overcome by increasing the phosphatidylserine content of tissue factor-liposomes. Additionally, yeast surface display was used to screen a random library of tissue factor mutants for enhanced factor VIIa binding. Surprisingly, mutations at a single amino acid (Lys165 predominated, with the Lys165→Glu mutant exhibiting a 3-fold enhancement in factor VIIa binding affinity. Our studies reveal the functional contributions of residues in the C-terminal half of the tissue factor ectodomain that are implicated in interacting with phosphatidylserine headgroups to enhance tissue factor cofactor activity, possibly by allosterically modulating the conformation of the adjacent substrate-binding exosite region of tissue factor.

  10. Characterization of a spontaneous nonmagnetic mutant of Magnetospirillum gryphiswaldense reveals a large deletion comprising a putative magnetosome island.

    Science.gov (United States)

    Schübbe, Sabrina; Kube, Michael; Scheffel, André; Wawer, Cathrin; Heyen, Udo; Meyerdierks, Anke; Madkour, Mohamed H; Mayer, Frank; Reinhardt, Richard; Schüler, Dirk

    2003-10-01

    Frequent spontaneous loss of the magnetic phenotype was observed in stationary-phase cultures of the magnetotactic bacterium Magnetospirillum gryphiswaldense MSR-1. A nonmagnetic mutant, designated strain MSR-1B, was isolated and characterized. The mutant lacked any structures resembling magnetosome crystals as well as internal membrane vesicles. The growth of strain MSR-1B was impaired under all growth conditions tested, and the uptake and accumulation of iron were drastically reduced under iron-replete conditions. A large chromosomal deletion of approximately 80 kb was identified in strain MSR-1B, which comprised both the entire mamAB and mamDC clusters as well as further putative operons encoding a number of magnetosome-associated proteins. A bacterial artificial chromosome clone partially covering the deleted region was isolated from the genomic library of wild-type M. gryphiswaldense. Sequence analysis of this fragment revealed that all previously identified mam genes were closely linked with genes encoding other magnetosome-associated proteins within less than 35 kb. In addition, this region was remarkably rich in insertion elements and harbored a considerable number of unknown gene families which appeared to be specific for magnetotactic bacteria. Overall, these findings suggest the existence of a putative large magnetosome island in M. gryphiswaldense and other magnetotactic bacteria.

  11. The putative Leishmania telomerase RNA (LeishTER undergoes trans-splicing and contains a conserved template sequence.

    Directory of Open Access Journals (Sweden)

    Elton J R Vasconcelos

    Full Text Available Telomerase RNAs (TERs are highly divergent between species, varying in size and sequence composition. Here, we identify a candidate for the telomerase RNA component of Leishmania genus, which includes species that cause leishmaniasis, a neglected tropical disease. Merging a thorough computational screening combined with RNA-seq evidence, we mapped a non-coding RNA gene localized in a syntenic locus on chromosome 25 of five Leishmania species that shares partial synteny with both Trypanosoma brucei TER locus and a putative TER candidate-containing locus of Crithidia fasciculata. Using target-driven molecular biology approaches, we detected a ∼2,100 nt transcript (LeishTER that contains a 5' spliced leader (SL cap, a putative 3' polyA tail and a predicted C/D box snoRNA domain. LeishTER is expressed at similar levels in the logarithmic and stationary growth phases of promastigote forms. A 5'SL capped LeishTER co-immunoprecipitated and co-localized with the telomerase protein component (TERT in a cell cycle-dependent manner. Prediction of its secondary structure strongly suggests the existence of a bona fide single-stranded template sequence and a conserved C[U/C]GUCA motif-containing helix II, representing the template boundary element. This study paves the way for further investigations on the biogenesis of parasite TERT ribonucleoproteins (RNPs and its role in parasite telomere biology.

  12. Putative Risk Factors in Developmental Dyslexia: A Case-Control Study of Italian Children

    Science.gov (United States)

    Mascheretti, Sara; Marino, Cecilia; Simone, Daniela; Quadrelli, Ermanno; Riva, Valentina; Cellino, Maria Rosaria; Maziade, Michel; Brombin, Chiara; Battaglia, Marco

    2015-01-01

    Although dyslexia runs in families, several putative risk factors that cannot be immediately identified as genetic predict reading disability. Published studies analyzed one or a few risk factors at a time, with relatively inconsistent results. To assess the contribution of several putative risk factors to the development of dyslexia, we conducted…

  13. Putative Risk Factors in Developmental Dyslexia: A Case-Control Study of Italian Children

    Science.gov (United States)

    Mascheretti, Sara; Marino, Cecilia; Simone, Daniela; Quadrelli, Ermanno; Riva, Valentina; Cellino, Maria Rosaria; Maziade, Michel; Brombin, Chiara; Battaglia, Marco

    2015-01-01

    Although dyslexia runs in families, several putative risk factors that cannot be immediately identified as genetic predict reading disability. Published studies analyzed one or a few risk factors at a time, with relatively inconsistent results. To assess the contribution of several putative risk factors to the development of dyslexia, we conducted…

  14. An "in-out" strategy using gene targeting and FLP recombinase for the functional dissection of complex DNA regulatory elements: analysis of the beta-globin locus control region.

    Science.gov (United States)

    Fiering, S; Kim, C G; Epner, E M; Groudine, M

    1993-01-01

    The human beta-globin locus control region (LCR) is a complex DNA regulatory element that controls the expression of the cis-linked beta-like globin genes located in the 55 kilobases 3' of the LCR. We have initiated the functional analysis of the LCR by homologous recombination in murine erythroleukemia cell somatic hybrids that carry a single copy of human chromosome 11 on which the beta-globin locus is situated. High-level expression of the human beta-globin gene normally occurs when these hybrid cells are induced to differentiate. We have reported that the insertion of an expressed selectable marker gene (driven by the Friend virus enhancer/promoter) into the LCR disrupts the LCR-mediated regulation of globin transcription. In these cells, beta-globin is no longer expressed when the cells differentiate; instead, expression of the selectable marker gene increases significantly after differentiation. Since present techniques for homologous recombination require the insertion of a selectable marker, further progress in using homologous recombination to analyze the LCR depends on deletion of the selectable marker and demonstration that the locus functions normally after the insertion, expression, and deletion of the selectable marker. Here we show that after precise deletion of the selectable marker by using the FLP recombinase/FRT (FLP recombinase target) system, the locus functions as it did before the homologous recombination event. These studies demonstrate the feasibility of using homologous recombination to analyze the LCR in particular, and other complex cis-regulatory DNA elements in general, in their normal chromosomal context. Images Fig. 2 Fig. 3 Fig. 4 PMID:8378321

  15. Universal Elements in Human Nature: The Putative Connection between Moral Development and Formal-Pragmatic Presupposition Awareness.

    Science.gov (United States)

    Stroud, Scott R.

    This study attempts to demonstrate a correlation between college students' moral stage level and their recognition of formal-pragmatic presuppositions of communicative action. Jurgen Habermas has posited the existence of formal-pragmatic presuppositions that are assumed in every act of communicative behavior. Lawrence Kohlberg has posited…

  16. Transposable elements in the Anopheles funestus transcriptome.

    Science.gov (United States)

    Fernández-Medina, Rita D; Carareto, Claudia M A; Struchiner, Cláudio J; Ribeiro, José M C

    2017-06-01

    Transposable elements (TEs) are present in most of the eukaryotic genomes and their impact on genome evolution is increasingly recognized. Although there is extensive information on the TEs present in several eukaryotic genomes, less is known about the expression of these elements at the transcriptome level. Here we present a detailed analysis regarding the expression of TEs in Anopheles funestus, the second most important vector of human malaria in Africa. Several transcriptionally active TE families belonging both to Class I and II were identified and characterized. Interestingly, we have identified a full-length putative active element (including the presence of full length TIRs in the genomic sequence) belonging to the hAT superfamily, which presents active members in other insect genomes. This work contributes to a comprehensive understanding of the landscape of transposable elements in A. funestus transcriptome. Our results reveal that TEs are abundant and diverse in the mosquito and that most of the TE families found in the genome are represented in the mosquito transcriptome, a fact that could indicate activity of these elements.The vast diversity of TEs expressed in A. funestus suggests that there is ongoing amplification of several families in this organism.

  17. Putative functions of extracellular matrix glycoproteins in secondary palate morphogenesis

    Science.gov (United States)

    d'Amaro, Rocca; Scheidegger, Rolf; Blumer, Susan; Pazera, Pawel; Katsaros, Christos; Graf, Daniel; Chiquet, Matthias

    2012-01-01

    Cleft palate is a common birth defect in humans. Elevation and fusion of paired palatal shelves are coordinated by growth and transcription factors, and mutations in these can cause malformations. Among the effector genes for growth factor signaling are extracellular matrix (ECM) glycoproteins. These provide substrates for cell adhesion (e.g., fibronectin, tenascins), but also regulate growth factor availability (e.g., fibrillins). Cleft palate in Bmp7 null mouse embryos is caused by a delay in palatal shelf elevation. In contrast, palatal shelves of Tgf-β3 knockout mice elevate normally, but a cleft develops due to their failure to fuse. However, nothing is known about a possible functional interaction between specific ECM proteins and Tgf-β/Bmp family members in palatogenesis. To start addressing this question, we studied the mRNA and protein distribution of relevant ECM components during secondary palate development, and compared it to growth factor expression in wildtypewild type and mutant mice. We found that fibrillin-2 (but not fibrillin-1) mRNA appeared in the mesenchyme of elevated palatal shelves adjacent to the midline epithelial cells, which were positive for Tgf-β3 mRNA. Moreover, midline epithelial cells started expressing fibronectin upon contact of the two palatal shelves. These findings support the hypothesis that fibrillin-2 and fibronectin are involved in regulating the activity of Tgf-β3 at the fusing midline. In addition, we observed that tenascin-W (but not tenascin-C) was misexpressed in palatal shelves of Bmp7-deficient mouse embryos. In contrast to tenascin-C, tenascin-W secretion was strongly induced by Bmp7 in embryonic cranial fibroblasts in vitro. These results are consistent with a putative function for tenascin-W as a target of Bmp7 signaling during palate elevation. Our results indicate that distinct ECM proteins are important for morphogenesis of the secondary palate, both as downstream effectors and as regulators of Tgf

  18. Discrete Element Modeling

    Energy Technology Data Exchange (ETDEWEB)

    Morris, J; Johnson, S

    2007-12-03

    The Distinct Element Method (also frequently referred to as the Discrete Element Method) (DEM) is a Lagrangian numerical technique where the computational domain consists of discrete solid elements which interact via compliant contacts. This can be contrasted with Finite Element Methods where the computational domain is assumed to represent a continuum (although many modern implementations of the FEM can accommodate some Distinct Element capabilities). Often the terms Discrete Element Method and Distinct Element Method are used interchangeably in the literature, although Cundall and Hart (1992) suggested that Discrete Element Methods should be a more inclusive term covering Distinct Element Methods, Displacement Discontinuity Analysis and Modal Methods. In this work, DEM specifically refers to the Distinct Element Method, where the discrete elements interact via compliant contacts, in contrast with Displacement Discontinuity Analysis where the contacts are rigid and all compliance is taken up by the adjacent intact material.

  19. Functional validation of a constitutive autonomous silencer element.

    Directory of Open Access Journals (Sweden)

    Heyuan Qi

    Full Text Available Sequences of the genome that are capable of silencing gene expression are thought to play a key role in gene regulation. However, very few silencer elements capable of functioning in mammalian cells have been described, and only a fraction of these have been tested for the ability to function in an autonomous fashion. We report here the characterization and functional validation of a constitutive autonomous silencer element from the human genome called T39, and the comparison of T39 to three other putative silencer elements previously described by others. Functional analysis included one assay for enhancer-blocking insulator activity and two independent assays for silencer activity, all based on stable transfection and comparison to a neutral spacer control. In erythroid K562 cells, T39 exhibited potent silencer activity, the previously described element PRE2-S5 exhibited modest silencer activity, and the two other previously described elements exhibited no silencer activity. T39 was further found to be capable of silencing three disparate promoters, of silencing gene expression in three disparate cell lines, and of functioning as a single copy in a topology-independent manner. Of the four elements analyzed, only T39 exhibits a constitutive pattern of DNase hypersensitivity and binding by CTCF. In its native location the T39 element also exhibits a unique interaction profile with a subset of distal putative regulatory elements. Taken together, these studies validate T39 as a constitutive autonomous silencer, identify T39 as a defined control for future studies of other regulatory elements such as insulators, and provide a basic chromatin profile for one highly potent silencer element.

  20. Prioritization of putative metabolite identifications in LC-MS/MS experiments using a computational pipeline.

    Science.gov (United States)

    Zhou, Bin; Xiao, Jun Feng; Ressom, Habtom W

    2013-01-01

    One of the major bottle-necks in current LC-MS-based metabolomic investigations is metabolite identification. An often-used approach is to first look up metabolites from databases through peak mass, followed by verification of the obtained putative identifications using MS/MS data. However, the mass-based search may provide inappropriate putative identifications when the observed peak is from isotopes, fragments, or adducts. In addition, a large fraction of peaks is often left with multiple putative identifications. To differentiate these putative identifications, manual verification of metabolites through comparison between biological samples and authentic compounds is necessary. However, such experiments are laborious, especially when multiple putative identifications are encountered. It is desirable to use computational approaches to obtain more reliable putative identifications and prioritize them before performing experimental verification of the metabolites. In this article, a computational pipeline is proposed to assist metabolite identification with improved metabolome coverage and prioritization capability. Multiple publicly available software tools and databases, along with in-house developed algorithms, are utilized to fully exploit the information acquired from LC-MS/MS experiments. The pipeline is successfully applied to identify metabolites on the basis of LC-MS as well as MS/MS data. Using accurate masses, retention time values, MS/MS spectra, and metabolic pathways/networks, more appropriate putative identifications are retrieved and prioritized to guide subsequent metabolite verification experiments.

  1. Structure determination and biochemical characterization of a putative HNH endonuclease from Geobacter metallireducens GS-15.

    Directory of Open Access Journals (Sweden)

    Shuang-yong Xu

    Full Text Available The crystal structure of a putative HNH endonuclease, Gmet_0936 protein from Geobacter metallireducens GS-15, has been determined at 2.6 Å resolution using single-wavelength anomalous dispersion method. The structure contains a two-stranded anti-parallel β-sheet that are surrounded by two helices on each face, and reveals a Zn ion bound in each monomer, coordinated by residues Cys38, Cys41, Cys73, and Cys76, which likely plays an important structural role in stabilizing the overall conformation. Structural homologs of Gmet_0936 include Hpy99I endonuclease, phage T4 endonuclease VII, and other HNH endonucleases, with these enzymes sharing 15-20% amino acid sequence identity. An overlay of Gmet_0936 and Hpy99I structures shows that most of the secondary structure elements, catalytic residues as well as the zinc binding site (zinc ribbon are conserved. However, Gmet_0936 lacks the N-terminal domain of Hpy99I, which mediates DNA binding as well as dimerization. Purified Gmet_0936 forms dimers in solution and a dimer of the protein is observed in the crystal, but with a different mode of dimerization as compared to Hpy99I. Gmet_0936 and its N77H variant show a weak DNA binding activity in a DNA mobility shift assay and a weak Mn²⁺-dependent nicking activity on supercoiled plasmids in low pH buffers. The preferred substrate appears to be acid and heat-treated DNA with AP sites, suggesting Gmet_0936 may be a DNA repair enzyme.

  2. REVISITING THE PUTATIVE TCR Cα DIMERIZATION MODEL THROUGH STRUCTURAL ANALYSIS

    Directory of Open Access Journals (Sweden)

    Jia-huai eWang

    2013-01-01

    Full Text Available Despite major advances in T cell receptor (TCR biology and structure, how peptide-MHC complex (pMHC ligands trigger αβ TCR activation remains unresolved. Two views exist. One model postulates that monomeric TCR-pMHC ligation events are sufficient while a second proposes that TCR-TCR dimerization in cis via Cα domain interaction plus pMHC binding is critical. We scrutinized 22 known TCR/pMHC complex crystal structures, and did not find any predicted molecular Cα-Cα contacts in these crystals that would allow for physiological TCR dimerization. Moreover, the presence of conserved glycan adducts on the outer face of the Cα domain preclude the hypothesized TCR dimerization through the Cα domain. Observed functional consequences of Cα mutations are likely indirect, with TCR microclusters at the immunological synapse driven by TCR transmembrane/cytoplasmic interactions via signaling molecules, scaffold proteins and/or cytoskeletal elements.

  3. Tropomyosin-1, A Putative Tumor-Suppressor and a Biomarker of Human Breast Cancer

    Science.gov (United States)

    2004-10-01

    cDNA. Lobular carcinoma - 2 A polyclonal pan-TM antibody that recognizes multiple TM Phyllodes tumor - 1 Not determined from the initial pathology...AD Award Number: DAMD17-98-1-8162 TITLE: Tropomyosin-1, A Putative Tumor -Suppressor and a Biomarker of Human Breast Cancer PRINCIPAL INVESTIGATOR...4. TITLE AND SUBTITLE 5. FUNDING NUMBERS Tropomyosin-l, A Putative Tumor -Suppressor and a Biomarker DAMD17-98-1-8162 of Human Breast Cancer 6. A UTHOR

  4. Fluoroquinolone-Resistant Haemophilus parasuis Isolates Exhibit More Putative Virulence Factors than Their Susceptible Counterparts

    OpenAIRE

    Zhang, Qiang; Liu, Jiantao; Yan, Shuxian; Yang, Yujie; Zhang, Anding; Jin, Meilin

    2013-01-01

    The prevalence of 23 putative virulence factors among fluoroquinolone-susceptible and -resistant Haemophilus parasuis isolates was analyzed. Putative hemolysin precursor, fimbrial assembly chaperone, and type I site-specific restriction modification system R subunit genes were more prevalent among fluoroquinolone-resistant H. parasuis isolates than among fluoroquinolone-susceptible H. parasuis isolates. Fluoroquinolone resistance may be associated with an increase in the presence of some viru...

  5. A novel putative tyrosine kinase receptor with oncogenic potential.

    Science.gov (United States)

    Janssen, J W; Schulz, A S; Steenvoorden, A C; Schmidberger, M; Strehl, S; Ambros, P F; Bartram, C R

    1991-11-01

    We have detected transforming activity by a tumorigenicity assay using NIH3T3 cells transfected with DNA from a chronic myeloproliferative disorder patient. Here, we report the cDNA cloning of the corresponding oncogene, designated UFO, in allusion to the as yet unidentified function of its protein. Nucleotide sequence analysis of a 3116bp cDNA clone revealed a 2682-bp-long open reading frame capable of directing the synthesis of a 894 amino acid polypeptide. The predicted UFO protein exhibits characteristic features of a transmembrane receptor with associated tyrosine kinase activity. The UFO proto-oncogene maps to human chromosome 19q13.1 and is transcribed into two 5.0 kb and 3.2 kb mRNAs in human bone marrow and human tumor cell lines. The UFO locus is evolutionarily conserved between vertebrate species. A 4.0 kb mRNA of the murine UFO homolog is expressed in a variety of different mouse tissues. We thus have identified a novel element of the complex signaling network involved in the control of cell proliferation and differentiation.

  6. Prevalence and characteristics of Streptococcus pneumoniae "putative serotype 6E" isolates from Asian countries.

    Science.gov (United States)

    Baek, Jin Yang; Park, In Ho; So, Thomas Man-kit; Lalitha, M K; Shimono, Nobuyuki; Yasin, Rohani Md; Carlos, Celia C; Perera, Jennifer; Thamlikitkul, Visanu; Hsueh, Po-Ren; Van, Pham Hung; Shibl, Atef M; Song, Jae-Hoon; Ko, Kwan Soo

    2014-12-01

    The prevalence, antimicrobial susceptibility, and genotypes of Streptococcus pneumoniae “putative serotype 6E” isolates from Asian countries were investigated. A total of 244 S. pneumoniae serogroup 6 isolates obtained from 11 Asian countries were included in this study. Of the 244 serogroup 6 isolates, 101 (41.4%) were typed as "putative serotype 6E," followed by serotypes 6A, 6B, 6C, and 6D (27.0, 20.1, 5.7, and 5.7%, respectively). Multilocus sequence typing revealed that clonal complex (CC) 90, including ST90 and its variants, was the most prevalent clonal group of "putative serotype 6E" isolates (n = 63; 62.4%). CC146 and CC315 were also found frequently in some of the countries. Most of the "putative serotype 6E" isolates showed very high resistance rates against cefuroxime, erythromycin, azithromycin, clarithromycin, clindamycin, and trimethoprim/sulfamethoxazole, probably due to their highly resistant to antimicrobials clone, CC90. Our results indicate that “putative serotype 6E” is prevalent in Asian countries. The clonal dissemination of "putative serotype 6E" isolates was also identified.

  7. Characterization of putative effectors from the cereal cyst nematode Heterodera avenae.

    Science.gov (United States)

    Cui, Jiangkuan; Peng, Huan; Qiao, Fen; Wang, Gaofeng; Huang, Wenkun; Wu, Duqign; Peng, Deliang

    2017-09-20

    Few molecular details of effectors of Heterodera avenae parasitism are known. We performed a high-throughput sequencing analysis of the H. avenae transcriptome at five developmental stages. A total of 82,549 unigenes were ultimately obtained, and 747 transcripts showed best hits to genes putatively encoding carbohydrate-active enzymes in plant parasitic nematodes that play an important role in the invasion process. A total of 1480 unigenes were homologous to known phytonematode effectors, and 63 putative novel effectors were identified in the H. avenae transcriptomes. Twenty-three unigenes were analyzed by qRT-PCR and confirmed to be highly expressed during at least one developmental stage. For in situ hybridization, 17 of the 22 tested putative effectors were specifically expressed and located in the subventral gland cells, and five putative novel effectors were specifically expressed in the dorsal gland. Furthermore, 115 transcripts were found to have putative lethal RNA interference (RNAi) phenotypes. Three target genes with lethal RNAi phenotypes and two of the four tested putative effectors were associated with a decrease in the number of cysts through in vitro RNAi technology. These transcriptomic data lay a foundation for further studies of interactions of H. avenae with cereal and H. avenae parasitic control.

  8. Molecular cloning and primary sequence analysis of a gene encoding a putative shitinase gene in Brassica oleracea var.capitata

    Institute of Scientific and Technical Information of China (English)

    TANGGUOQING; YONGYANBAI; 等

    1996-01-01

    Chitinase,which catalyzes the hydrolysis of the β-1,4-acetyl-D-glucosamine linkages of the fungal cell wall polymer chitin,is involved in inducible plants defense system.By construction of cabbage(Brassica oleracea var. capitata) genomic library and screening the library with pRCH8,a probe of rice chitinase gene fragment,a chitinase genomic sequence was isolated.The complete uncleotide sequence of the putative cabbage chitinase gene (cabch29) was determined,with its longest open reading frame (ORF) encoding a polypeptide of 413 aa.This polypeptide consists of a 21 aa N-terminal signal peptide,two chitin-binding domains different from those of other classes of plant chitinases,and a catalytic domain.Homology analysis illustrated that this cabch29 gene has 58.8% identity at the nucleotide level with the pRCH8 ORF probe and has 50% identity at the amino acid level tiwh the catalytic domains of chitinase from bean,maize and sugar beet.Meanwhile,several kinds of cis-elements,such as TATA box,CAAT box,GATA motif,ASF-1 binding site,wound-response elements and AATAAA,have also been discovered in the flanking region of cabch29 gene.

  9. Data Element Registry Services

    Data.gov (United States)

    U.S. Environmental Protection Agency — Data Element Registry Services (DERS) is a resource for information about value lists (aka code sets / pick lists), data dictionaries, data elements, and EPA data...

  10. Element-ary Development.

    Science.gov (United States)

    Schamp, Homer W., Jr.

    1989-01-01

    Describes the historic development of the periodic table from the four-element theory to the Lavoisier's table. Presents a table listing the old and new names of chemicals and the Lavoisier's table of elements. Lists two references. (YP)

  11. On Element SDD Approximability

    CERN Document Server

    Avron, Haim; Toledo, Sivan

    2009-01-01

    This short communication shows that in some cases scalar elliptic finite element matrices cannot be approximated well by an SDD matrix. We also give a theoretical analysis of a simple heuristic method for approximating an element by an SDD matrix.

  12. Identification of multiple putative S-layer genes partly expressed by Lysinibacillus sphaericus JG-B53.

    Science.gov (United States)

    Lederer, Franziska L; Weinert, Ulrike; Günther, Tobias J; Raff, Johannes; Weiß, Stephan; Pollmann, Katrin

    2013-06-01

    Lysinibacillus sphaericus JG-B53 was isolated from the uranium mining waste pile Haberland near Johanngeorgenstadt, Germany. Previous studies have shown that many bacteria that have been isolated from these heavy metal contaminated environments possess surface layer (S-layer) proteins that enable the bacteria to survive by binding metals with high affinity. Conversely, essential trace elements are able to cross the filter layer and reach the interior of the cell. This is especially true of the S-layer of L. sphaericus JG-B53, which possesses outstanding recrystallization and metal-binding properties. In this study, S-layer protein gene sequences encoded in the genome of L. sphaericus JG-B53 were identified using next-generation sequencing technology followed by bioinformatic analyses. The genome of L. sphaericus JG-B53 encodes at least eight putative S-layer protein genes with distinct differences. Using mRNA analysis the expression of the putative S-layer protein genes was studied. The functional S-layer protein B53 Slp1 was identified as the dominantly expressed S-layer protein in L. sphaericus JG-B53 by mRNA studies, SDS-PAGE and N-terminal sequencing. B53 Slp1 is characterized by square lattice symmetry and a molecular mass of 116 kDa. The S-layer protein B53 Slp1 shows a high similarity to the functional S-layer protein of L. sphaericus JG-A12, which was isolated from the same uranium mining waste pile Haberland and has been described by previous research. These similarities indicate horizontal gene transfer and DNA rearrangements between these bacteria. The presence of multiple S-layer gene copies may enable the bacterial strains to quickly adapt to changing environments.

  13. Identification of putative regulatory motifs in the upstream regions of co-expressed functional groups of genes in Plasmodium falciparum

    Directory of Open Access Journals (Sweden)

    Joshi NV

    2009-01-01

    Full Text Available Abstract Background Regulation of gene expression in Plasmodium falciparum (Pf remains poorly understood. While over half the genes are estimated to be regulated at the transcriptional level, few regulatory motifs and transcription regulators have been found. Results The study seeks to identify putative regulatory motifs in the upstream regions of 13 functional groups of genes expressed in the intraerythrocytic developmental cycle of Pf. Three motif-discovery programs were used for the purpose, and motifs were searched for only on the gene coding strand. Four motifs – the 'G-rich', the 'C-rich', the 'TGTG' and the 'CACA' motifs – were identified, and zero to all four of these occur in the 13 sets of upstream regions. The 'CACA motif' was absent in functional groups expressed during the ring to early trophozoite transition. For functional groups expressed in each transition, the motifs tended to be similar. Upstream motifs in some functional groups showed 'positional conservation' by occurring at similar positions relative to the translational start site (TLS; this increases their significance as regulatory motifs. In the ribonucleotide synthesis, mitochondrial, proteasome and organellar translation machinery genes, G-rich, C-rich, CACA and TGTG motifs, respectively, occur with striking positional conservation. In the organellar translation machinery group, G-rich motifs occur close to the TLS. The same motifs were sometimes identified for multiple functional groups; differences in location and abundance of the motifs appear to ensure different modes of action. Conclusion The identification of positionally conserved over-represented upstream motifs throws light on putative regulatory elements for transcription in Pf.

  14. Elemental Chemical Puzzlers

    Science.gov (United States)

    Thomas, Nicholas C.

    2009-01-01

    This paper provides nine short chemically based puzzles or problems extensible for use with students from middle school to college. Some of these will strengthen students' recognition of individual elements and element names. Others require students to focus on the salient properties of given chemical elements.

  15. Rare (Earth Elements [score

    Directory of Open Access Journals (Sweden)

    Camilo Méndez

    2014-12-01

    Full Text Available Rare (Earth Elements is a cycle of works for solo piano. The cycle was inspired by James Dillon’s Book of Elements (Vol. I-V. The complete cycle will consist of 14 pieces; one for each selected rare (earth element. The chosen elements are Neodymium, Erbium, Tellurium, Hafnium, Tantalum, Technetium, Indium, Dysprosium, Lanthanium, Cerium, Europium, Terbium, Yttrium and Darmstadtium. These elements were selected due to their special atomic properties that in many cases make them extremely valuable for the development of new technologies, and also because of their scarcity. To date, only 4 works have been completed Yttrium, Technetium, Indium and Tellurium.

  16. Musikalske elementer i musikaler

    OpenAIRE

    Jensen, Linnea Reitan

    2014-01-01

    Denne masteroppgaven undersøker hvordan musikalske elementer kan bli brukt i musikaler for å påvirke den dramaturgiske helheten. Gjennom både musikalsk og dramaturgisk analyse viser jeg hvordan elementer i musikk kan ha innvirkning på forestillingens dramaturgi. In this master thesis I am aiming to examine the musical elements used in the songs in musicals. Music is an important element in the musical, and therefore I want to look at how some musical elements can contribute to the musical'...

  17. Evolutionary conservation of regulatory elements in vertebrate HOX gene clusters

    Energy Technology Data Exchange (ETDEWEB)

    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.

  18. Ancient Pbx-Hox signatures define hundreds of vertebrate developmental enhancers

    Directory of Open Access Journals (Sweden)

    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.

  19. Low level of polymorphism in two putative NPR1 homologs in the Vitaceae family

    Directory of Open Access Journals (Sweden)

    Walter Bernard

    2010-02-01

    Full Text Available Abstract Background Grapevine is subjected to numerous pests and diseases resulting in the use of phytochemicals in large quantities. The will to decrease the use of phytochemicals leads to attempts to find alternative strategies, implying knowledge of defence mechanisms. Numerous studies have led to the identification of signalling pathways and regulatory elements involved in defence in various plant species. Nonexpressor of Pathogenesis Related 1 (NPR1 is an important regulatory component of systemic acquired resistance (SAR in Arabidopsis thaliana. Results Two putative homologs of NPR1 gene were found in the two sequenced grapevine genomes available in the Genoscope database for line 40024 and in the IASMA database for Pinot noir ENTAV 115. We named these two NPR1 genes of Vitis vinifera : VvNPR1.1 and VvNPR1.2. A PCR-based strategy with primers designed on exons was used to successfully amplify NPR1 gene fragments from different Vitaceae accessions. Sequence analyses show that NPR1.1 and NPR1.2 are highly conserved among the different accessions not only V. vinifera cultivars but also other species. We report nucleotide polymorphisms in NPR1.1 and NPR1.2 from fifteen accessions belonging to the Vitaceae family. The ratio of nonsynonymous to synonymous nucleotide substitutions determines the evolutionary pressures acting on the Vitaceae NPR1 genes. These genes appear to be experiencing purifying selection. In some of the species we have analysed one of the two alleles of NPR1.1 contains a premature stop codon. The deduced amino acid sequences share structural features with known NPR1-like proteins: ankyrin repeats, BTB/POZ domains, nuclear localization signature and cysteines. Phylogenetic analyses of deduced amino acid sequences show that VvNPR1.1 belongs to a first group of NPR1 proteins known as positive regulators of SAR and VvNPR1.2 belongs to a second group of NPR1 proteins whose principal members are AtNPR3 and AtNPR4 defined as

  20. Stringent comparative sequence analysis reveals SOX10 as a putative inhibitor of glial cell differentiation.

    Science.gov (United States)

    Gopinath, Chetna; Law, William D; Rodríguez-Molina, José F; Prasad, Arjun B; Song, Lingyun; Crawford, Gregory E; Mullikin, James C; Svaren, John; Antonellis, Anthony

    2016-11-07

    The transcription factor SOX10 is essential for all stages of Schwann cell development including myelination. SOX10 cooperates with other transcription factors to activate the expression of key myelin genes in Schwann cells and is therefore a context-dependent, pro-myelination transcription factor. As such, the identification of genes regulated by SOX10 will provide insight into Schwann cell biology and related diseases. While genome-wide studies have successfully revealed SOX10 target genes, these efforts mainly focused on myelinating stages of Schwann cell development. We propose that less-biased approaches will reveal novel functions of SOX10 outside of myelination. We developed a stringent, computational-based screen for genome-wide identification of SOX10 response elements. Experimental validation of a pilot set of predicted binding sites in multiple systems revealed that SOX10 directly regulates a previously unreported alternative promoter at SOX6, which encodes a transcription factor that inhibits glial cell differentiation. We further explored the utility of our computational approach by combining it with DNase-seq analysis in cultured Schwann cells and previously published SOX10 ChIP-seq data from rat sciatic nerve. Remarkably, this analysis enriched for genomic segments that map to loci involved in the negative regulation of gliogenesis including SOX5, SOX6, NOTCH1, HMGA2, HES1, MYCN, ID4, and ID2. Functional studies in Schwann cells revealed that: (1) all eight loci are expressed prior to myelination and down-regulated subsequent to myelination; (2) seven of the eight loci harbor validated SOX10 binding sites; and (3) seven of the eight loci are down-regulated upon repressing SOX10 function. Our computational strategy revealed a putative novel function for SOX10 in Schwann cells, which suggests a model where SOX10 activates the expression of genes that inhibit myelination during non-myelinating stages of Schwann cell development. Importantly, the

  1. Diversity of secondary endosymbionts among different putative species of the whitefly Bemisia tabaci.

    Science.gov (United States)

    Bing, Xiao-Li; Ruan, Yong-Ming; Rao, Qiong; Wang, Xiao-Wei; Liu, Shu-Sheng

    2013-04-01

    Endosymbionts are important components of arthropod biology. The whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae) is a cryptic species complex composed of ≥ 28 putative species. In addition to the primary endosymbiont Portiera aleyrodidarum, six secondary endosymbionts (S-endosymbionts), Hamiltonella, Rickettsia, Wolbachia, Cardinium, Arsenophonus and Fritschea, have been identified in B. tabaci thus far. Here, we tested five of the six S-endosymbiont lineages (excluding Fritschea) from 340 whitely individuals representing six putative species from China. Hamiltonella was detected only in the two exotic invaders, Middle East-Asia Minor 1 (MEAM1) and Mediterranean (MED). Rickettsia was absent in Asia II 1 and MED, scarce in Asia II 3 (13%), but abundant in Asia II 7 (63.2%), China 1 (84.7%) and MEAM1 (100%). Wolbachia, Cardinium and Arsenophonus were absent in the invasive MEAM1 and MED but mostly abundant in the native putative species. Furthermore, phylogenetic analyses revealed that some S-endosymbionts have several clades and different B. tabaci putative species can harbor different clades of a given S-endosymbiont, demonstrating further the complexity of S-endosymbionts in B. tabaci. All together, our results demonstrate the variation and diversity of S-endosymbionts in different putative species of B. tabaci, especially between invasive and native whiteflies.

  2. Diversity of secondary endosymbionts among different putative species of the whitefly Bemisia tabaci

    Institute of Scientific and Technical Information of China (English)

    Xiao-Li Bing; Yong-Ming Ruan; Qiong Rao; Xiao-Wei Wang; Shu-Sheng Liu

    2013-01-01

    Endosymbionts are important components of arthropod biology.The whitefly Bemisia tabaci (Gennadius) (Hemiptera:Aleyrodidae) is a cryptic species complex composed of≥28 putative species.In addition to the primary endosymbiont Portiera aleyrodidarum,six secondary endosymbionts (S-endosymbionts),Hamiltonella,Rickettsia,Wolbachia,Cardinium,Arsenophonus and Fritschea,have been identified in B.tabaci thus far.Here,we tested five of the six S-endosymbiont lineages (excluding Fritschea) from 340 whitely individuals representing six putative species from China.Hamiltonella was detected only in the two exotic invaders,Middle East-Asia Minor 1 (MEAM 1) and Mediterranean (MED).Rickettsia was absent in Asia Ⅱ 1 and MED,scarce in Asia Ⅱ 3 (13%),but abundant in Asia Ⅱ 7 (63.2%),China 1 (84.7%) and MEAM1 (100%).Wolbachia,Cardinium and Arsenophonus were absent in the invasive MEAM 1 and MED but mostly abundant in the native putative species.Furthermore,phylogenetic analyses revealed that some S-endosymbionts have several clades and different B.tabaci putative species can harbor different clades of a given S-endosymbiont,demonstrating further the complexity of S-endosymbionts in B.tabaci.All together,our results demonstrate the variation and diversity of S-endosymbionts in different putative species ofB.tabaci,especially between invasive and native whiteflies.

  3. Conjugative Transfer and cis-Mobilization of a Genomic Island by an Integrative and Conjugative Element of Streptococcus agalactiae

    OpenAIRE

    Puymège, Aurore; Bertin, Stéphane; Chuzeville, Sarah; Guédon, Gérard; Payot, Sophie

    2013-01-01

    Putative integrative and conjugative elements (ICEs), i.e., genomic islands which could excise, self-transfer by conjugation, and integrate into the chromosome of the bacterial host strain, were previously identified by in silico analysis in the sequenced genomes of Streptococcus agalactiae (M. Brochet et al., J. Bacteriol. 190:6913–6917, 2008). We investigated here the mobility of the elements integrated into the 3′ end of a tRNALys gene. Three of the four putative ICEs tested were found to ...

  4. Eliciting maltreated and nonmaltreated children's transgression disclosures: narrative practice rapport building and a putative confession.

    Science.gov (United States)

    Lyon, Thomas D; Wandrey, Lindsay; Ahern, Elizabeth; Licht, Robyn; Sim, Megan P Y; Quas, Jodi A

    2014-01-01

    This study tested the effects of narrative practice rapport building (asking open-ended questions about a neutral event) and a putative confession (telling the child an adult "told me everything that happened and he wants you to tell the truth") on 4- to 9-year-old maltreated and nonmaltreated children's reports of an interaction with a stranger who asked them to keep toy breakage a secret (n = 264). Only one third of children who received no interview manipulations disclosed breakage; in response to a putative confession, one half disclosed. Narrative practice rapport building did not affect the likelihood of disclosure. Maltreated children and nonmaltreated children responded similarly to the manipulations. Neither narrative practice rapport building nor a putative confession increased false reports.

  5. Molecular diagnosis of putative Stargardt disease by capture next generation sequencing.

    Science.gov (United States)

    Zhang, Xiao; Ge, Xianglian; Shi, Wei; Huang, Ping; Min, Qingjie; Li, Minghan; Yu, Xinping; Wu, Yaming; Zhao, Guangyu; Tong, Yi; Jin, Zi-Bing; Qu, Jia; Gu, Feng

    2014-01-01

    Stargardt Disease (STGD) is the commonest genetic form of juvenile or early adult onset macular degeneration, which is a genetically heterogeneous disease. Molecular diagnosis of STGD remains a challenge in a significant proportion of cases. To address this, seven patients from five putative STGD families were recruited. We performed capture next generation sequencing (CNGS) of the probands and searched for potentially disease-causing genetic variants in previously identified retinal or macular dystrophy genes. Seven disease-causing mutations in ABCA4 and two in PROM1 were identified by CNGS, which provides a confident genetic diagnosis in these five families. We also provided a genetic basis to explain the differences among putative STGD due to various mutations in different genes. Meanwhile, we show for the first time that compound heterozygous mutations in PROM1 gene could cause cone-rod dystrophy. Our findings support the enormous potential of CNGS in putative STGD molecular diagnosis.

  6. Putative epidermal stem cell convert into corneal epithelium-like cell under corneal tissue in vitro

    Institute of Scientific and Technical Information of China (English)

    GAO Nan; CUI GuangHui; WANG ZhiChong; HUANG Bing; GE Jian; LU Rong; ZHANG KeFei; FAN ZhiGang; LU Li; PENG Zhan

    2007-01-01

    Rhesus putative epidermal stem cells are being investigated for their potential use in regenerative corneal epithelium-like cells, which may provide a practical source of autologous seed cells for the construction of bioengineered corneas. The goal of this study was to investigate the potential of epidermal stem cells for trans-differentiation into corneal epithelium-like cells. Rhesus putative epidermal stem cells were isolated by type IV collagen attachment method. Flow cytometry analysis, immunohistology and RT-PCR were conducted to identify the expression of specific markers (β1, α6 integrin, K15, K1/K10, K3/K12 and CD71) on the isolated rapid attaching cells. The isolated cells were cocultured with human corneal limbal stroma and corneal epithelial cells. After coculture, the expression of the same specific markers was evaluated in order to identify expression difference caused by the coculture conditions. K3/K12 expression was analyzed in coculture cells on day 2, 4, 6, 8 and 10. Putative epidermal stem cells in conditioned culture media were used as control. Putative epidermal stem cells were predominant in rapid attaching cells by type IV collagen attachment isolation. Before being cocultured, the rhesus putative epidermal stem cells expressed K15, α6 and β1 integrin, but no CD71, K1/K10 and K3/K12. After coculture, these cells expressed K3/K12 (a marker of corneal epithelial cells), K15 and β 1 integrin, but no K1/K10. Cells being not coculture converted into terminally differentiated cells expressing K1/K10. These results indicate that rhesus putative epidermal stem cells can trans-differentiate into corneal epithelium-like cells and, therefore, may have potential therapeutic application as autologous seed cells for the construction of bioengineered corneas.

  7. CLONING, SEQUENCE ANALYSIS, AND CHARACTERIZATION OF PUTATIVE BETA-LACTAMASE OF STENOTROPHOMONAS MALTOPHILIA

    Directory of Open Access Journals (Sweden)

    Chong Seng Shueh

    2012-10-01

    Full Text Available The main objective of current study was to explore the function of chromosomal putative beta-lactamase gene (smlt 0115 in clinical Stenotrophomonas maltophilia. Antibiotic susceptibility test (AST screening for current antimicrobial drugs was done and Minimum Inhibitory Concentration (MIC level towards beta-lactams was determined by E-test. Putative beta-lactamase gene of S. maltophilia was amplified via PCR, with specific primers, then cloned into pET-15 expression plasmid and transformed into Escherichia coli BL21. The gene was sequenced and analyzed. The expressed protein was purified by affinity chromatography and the kinetic assay was performed. S. maltophilia ATCC 13637 was included in this experiment. Besides, a hospital strain which exhibited resistant to a series of beta-lactams including cefepime was identified via AST and MIC, hence it was named as S2 strain and was considered in this study. Sequencing result showed that putative beta-lactamase gene obtained from ATCC 13637 and S2 strains were predicted to have cephalosporinase activity by National Center for Biotechnology Information (NCBI blast program. Differences in the sequences of both ATCC 13637 and S2 strains were found via ClustalW alignment software. Kinetic assay proved a cephalosporinase characteristic produced by E. coli BL21 clone that overexpressed the putative beta-lactamase gene cloned under the control of an external promoter. Yet, expressed protein purified from S2 strain had high catalytic activity against beta-lactam antibiotics which was 14-fold higher than expressed protein purified from ATCC 13637 strain. This study represents the characterization analysis of putative beta-lactamase gene (smlt 0115 of S. maltophilia. The presence of the respective gene in the chromosome of S. maltophilia suggested that putative beta-lactamase gene (smlt 0115 of S. maltophilia plays a role in beta-lactamase resistance.

  8. Putative epidermal stem cell convert into corneal epithelium-like cell under corneal tissue in vitro

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Rhesus putative epidermal stem cells are being investigated for their potential use in regenerative corneal epithelium-like cells, which may provide a practical source of autologous seed cells for the construction of bioengineered corneas. The goal of this study was to investigate the potential of epi-dermal stem cells for trans-differentiation into corneal epithelium-like cells. Rhesus putative epidermal stem cells were isolated by type IV collagen attachment method. Flow cytometry analysis, immuno-histology and RT-PCR were conducted to identify the expression of specific markers (β1, α6 integrin, K15, K1/K10, K3/K12 and CD71) on the isolated rapid attaching cells. The isolated cells were cocultured with human corneal limbal stroma and corneal epithelial cells. After coculture, the expression of the same specific markers was evaluated in order to identify expression difference caused by the coculture conditions. K3/K12 expression was analyzed in coculture cells on day 2, 4, 6, 8 and 10. Putative epi-dermal stem cells in conditioned culture media were used as control. Putative epidermal stem cells were predominant in rapid attaching cells by type IV collagen attachment isolation. Before being co-cultured, the rhesus putative epidermal stem cells expressed K15, α6 and β1 integrin, but no CD71, K1/K10 and K3/K12. After coculture, these cells expressed K3/K12 (a marker of corneal epithelial cells), K15 and β 1 integrin, but no K1/K10. Cells being not coculture converted into terminally differentiated cells expressing K1/K10. These results indicate that rhesus putative epidermal stem cells can trans-differentiate into corneal epithelium-like cells and, therefore, may have potential therapeutic application as autologous seed cells for the construction of bioengineered corneas.

  9. Detection of putative new mutacins by bioinformatic analysis using available web tools

    Directory of Open Access Journals (Sweden)

    Nicolas Guillaume G

    2011-07-01

    Full Text Available Abstract In order to characterise new bacteriocins produced by Streptococcus mutans we perform a complete bioinformatic analyses by scanning the genome sequence of strains UA159 and NN2025. By searching in the adjacent genomic context of the two-component signal transduction system we predicted the existence of many putative new bacteriocins' maturation pathways and some of them were only exclusive to a group of Streptococcus. Computational genomic and proteomic analysis combined to predictive functionnal analysis represent an alternative way for rapid identification of new putative bacteriocins as well as new potential antimicrobial drugs compared to the more traditional methods of drugs discovery using antagonism tests.

  10. Finite element procedures

    CERN Document Server

    Bathe, Klaus-Jürgen

    2015-01-01

    Finite element procedures are now an important and frequently indispensable part of engineering analyses and scientific investigations. This book focuses on finite element procedures that are very useful and are widely employed. Formulations for the linear and nonlinear analyses of solids and structures, fluids, and multiphysics problems are presented, appropriate finite elements are discussed, and solution techniques for the governing finite element equations are given. The book presents general, reliable, and effective procedures that are fundamental and can be expected to be in use for a long time. The given procedures form also the foundations of recent developments in the field.

  11. Main: -300ELEMENT [PLACE

    Lifescience Database Archive (English)

    Full Text Available -300ELEMENT S000122 11-May-2006 (last modified) kehi Present upstream of the promot...er from the B-hordein gene of barley and the alpha-gliadin, gamma-gliadin, and low molecular weight glutenin... genes of wheat; See S000001 -300CORE; See S000002 -300MOTIF; -300 element; hordein; gliadin; glutenin; seed; wheat (Triticum aestivum) TGHAAARK ...

  12. A dry element

    Energy Technology Data Exchange (ETDEWEB)

    Kimura, T.; Ota, A.

    1983-08-11

    The agglomerate for the element is made from activated charcoal powder, an electrically conducting additive and a neutral electrolyte. The activated charcoal makes up 30 to 50 percent of the weight of the agglomerate. It is a mixture of hydrophobized and unhydrophobized powder in a ratio of 85 to 70 to 15 to 30. The element has high discharge characteristics.

  13. A battery element

    Energy Technology Data Exchange (ETDEWEB)

    Nakai, M.; Koboyasi, S.; Oisi, K.; Okadzaki, R.; Ota, A.

    1983-07-29

    An anode made of an alkaline or an alkaline earth metal and an electrolyte based on an organic solvent are used in the element. A mixture of Mn203 and Mn304 in a 9 to 1 to 3 to 7 ratio serves as the cathode. The element has a stable discharge curve at a nominal voltage of 1.5 volts.

  14. The solar element

    DEFF Research Database (Denmark)

    Kragh, Helge

    2009-01-01

    Apart from hydrogen, helium is the most abundant chemical element in the universe, and yet it was only discovered on the Earth in 1895. Its early history is unique because it encompasses astronomy as well as chemistry, two sciences which the spectroscope brought into contact during the second half...... of the nineteenth century. In the modest form of a yellow spectral line known as D3, 'helium' was sometimes supposed to exist in the Sun's atmosphere, an idea which is traditionally ascribed to J. Norman Lockyer. Did Lockyer discover helium as a solar element? How was the suggestion received by chemists, physicists...... elements might be different. The complex story of how helium became established as both a solar and terrestrial element involves precise observations as well as airy speculations. It is a story that is unique among the discovery histories of the chemical elements....

  15. Evidence for regulation of columnar habit in apple by a putative 2OG-Fe(II) oxygenase.

    Science.gov (United States)

    Wolters, Pieter J; Schouten, Henk J; Velasco, Riccardo; Si-Ammour, Azeddine; Baldi, Paolo

    2013-12-01

    Understanding the genetic mechanisms controlling columnar-type growth in the apple mutant 'Wijcik' will provide insights on how tree architecture and growth are regulated in fruit trees. In apple, columnar-type growth is controlled by a single major gene at the Columnar (Co) locus. By comparing the genomic sequence of the Co region of 'Wijcik' with its wild-type 'McIntosh', a novel non-coding DNA element of 1956 bp specific to Pyreae was found to be inserted in an intergenic region of 'Wijcik'. Expression analysis of selected genes located in the vicinity of the insertion revealed the upregulation of the MdCo31 gene encoding a putative 2OG-Fe(II) oxygenase in axillary buds of 'Wijcik'. Constitutive expression of MdCo31 in Arabidopsis thaliana resulted in compact plants with shortened floral internodes, a phenotype reminiscent of the one observed in columnar apple trees. We conclude that MdCo31 is a strong candidate gene for the control of columnar growth in 'Wijcik'.

  16. Genome-wide transcriptome analysis of the plant pathogen Xanthomonas identifies sRNAs with putative virulence functions

    Science.gov (United States)

    Schmidtke, Cornelius; Findeiß, Sven; Sharma, Cynthia M.; Kuhfuß, Juliane; Hoffmann, Steve; Vogel, Jörg; Stadler, Peter F.; Bonas, Ulla

    2012-01-01

    The Gram-negative plant-pathogenic bacterium Xanthomonas campestris pv. vesicatoria (Xcv) is an important model to elucidate the mechanisms involved in the interaction with the host. To gain insight into the transcriptome of the Xcv strain 85–10, we took a differential RNA sequencing (dRNA-seq) approach. Using a novel method to automatically generate comprehensive transcription start site (TSS) maps we report 1421 putative TSSs in the Xcv genome. Genes in Xcv exhibit a poorly conserved −10 promoter element and no consensus Shine-Dalgarno sequence. Moreover, 14% of all mRNAs are leaderless and 13% of them have unusually long 5′-UTRs. Northern blot analyses confirmed 16 intergenic small RNAs and seven cis-encoded antisense RNAs in Xcv. Expression of eight intergenic transcripts was controlled by HrpG and HrpX, key regulators of the Xcv type III secretion system. More detailed characterization identified sX12 as a small RNA that controls virulence of Xcv by affecting the interaction of the pathogen and its host plants. The transcriptional landscape of Xcv is unexpectedly complex, featuring abundant antisense transcripts, alternative TSSs and clade-specific small RNAs. PMID:22080557

  17. Occurrence of Putative Pathogenicity Islands in Enterococci from Distinct Species and of Differing Origins▿ †

    OpenAIRE

    Semedo-Lemsaddek, Teresa; Barreto-Crespo, Maria Teresa; Tenreiro, Rogério

    2009-01-01

    Enterococci isolated from ewe's milk and cheese, clinical isolates of human and veterinary origins, and reference strains obtained from culture collections were screened for the occurrence of putative pathogenicity island (PAIs). Results obtained after PCR amplification and hybridization point toward PAI dissemination among enterococci of diverse origins (food/clinical) and species (Enterococcus faecalis/non-E. faecalis).

  18. Cloning and characterization of prunus serotina AGAMOUS, a putative flower homeotic gene

    Science.gov (United States)

    Xiaomei Liu; Joseph Anderson; Paula Pijut

    2010-01-01

    Members of the AGAMOUS subfamily of MADS-box transcription factors play an important role in regulating the development of reproductive organs in flowering plants. To help understand the mechanism of floral development in black cherry (Prunus serotina), PsAG (a putative flower homeotic identity gene) was isolated...

  19. A new putative alphapartitivirus recovered from the powdery mildew fungus Erysiphe palczewskii.

    Science.gov (United States)

    Xiong, Guihong; Qiu, Ping; Li, Cong; Chen, Zhuo; Islam, Saif Ul; Fang, Shouguo; Wu, Zujian; Zhang, Songbai; Du, Zhenguo

    2017-02-27

    Two double-stranded RNAs (dsRNA) likely representing the genome of a novel alphapartitivirus which we provisionally named Erysiphe palczewskii alphapartitivirus 1 (EpV1) were recovered from the powdery mildew fungus E. palczewskii infecting Sophora japonica in Jingzhou, Hubei province of China. The two dsRNAs, 1955 (dsRNA1) and 1917 (dsRNA2) bp in size, respectively, each contains a single open reading frame (ORF) encoding a 585- and 528-aa protein, respectively. The 585-aa protein contains a conserved RNA-dependent RNA polymerase (RdRp) domain and shows significant homology to RdRps of approved or putative partitiviruses, particularly those belonging to the genus Alphapartitivirus. However, it shares an aa sequence identity lower than 80% with its closest relative, the RdRp of the putative alphapartitivirus Grapevine partitivirus, and lower than 60% with the RdRps of other partitiviruses. In a phylogenetic tree constructed with RdRp aa sequences of selected partitiviruses, the putative virus EpV1 clustered with Grapevine partitivirus and formed a well-supported monophyletic clade with known or putative alphapartitiviruses.

  20. A rapid approach to evaluate putative nursery sites for penaeid prawns

    Science.gov (United States)

    Taylor, Matthew D.; Smith, James A.; Boys, Craig A.; Whitney, Hannah

    2016-08-01

    Identifying nursery habitats for an aquatic species generally requires tracing adult individuals back through time and space to the area or habitat in which they developed as juveniles. We develop and trial a study design and analytical approach to evaluate the suitability of using stable isotopes to trace emigrating prawns to putative nursery sites, and evaluate assumptions inherent in the application of the approach using two penaeid species with Type-II life cycles: Penaeus (Melicertus) plebejus and Metapenaeus macleayi. Prawns were collected in putative nursery sites within the Hunter River, Australia, and analysed as composite samples of 6 individuals to provide habitat-specific isotopic signatures. Prawns emigrating from the mouth of the river were used as a proxy for individuals recruiting to the adult population, and assigned to putative nursery sites using a probabilistic mixing model and a simple, distance-based approach. Bivariate (δ15N and δ13C) isotopic data was sufficient to distinguish prawns from different putative nursery sites, and isotopic composition correlated closely with salinity. Approximately 90% of emigrating prawns collected could be assigned to these sites using bivariate isotopic data, and both analytical approaches gave similar results. The design developed here is broadly applicable to a suite of penaeid species, but its application will be most powerful when sampling is also aimed at understanding nursery function by simultaneous monitoring of size structure/growth, density, and trophic relationships within nursery habitats.

  1. Mapping the flow of information within the putative mirror neuron system during gesture observation

    NARCIS (Netherlands)

    Schippers, Marleen B.; Keysers, Christian

    2011-01-01

    The putative mirror neuron system may either function as a strict feed-forward system or as a dynamic control system. A strict feed-forward system would predict that action observation leads to a predominantly temporal -> parietal -> premotor flow of information in which a visual representation is t

  2. Complete Genome Sequence of a Putative Densovirus of the Asian Citrus Psyllid, Diaphorina citri.

    Science.gov (United States)

    Nigg, Jared C; Nouri, Shahideh; Falk, Bryce W

    2016-07-28

    Here, we report the complete genome sequence of a putative densovirus of the Asian citrus psyllid, Diaphorina citri Diaphorina citri densovirus (DcDNV) was originally identified through metagenomics, and here, we obtained the complete nucleotide sequence using PCR-based approaches. Phylogenetic analysis places DcDNV between viruses of the Ambidensovirus and Iteradensovirus genera.

  3. Expression of putative expansin genes in phylloxera (Daktulosphaira vitifoliae Fitch) induced root galls of Vitis spp.

    Science.gov (United States)

    Lawo, N C; Griesser, M; Forneck, A

    Grape phylloxera (Daktulosphaira vitifoliae Fitch) is a serious global pest in viticulture. The insects are sedentary feeders and require a gall to feed and reproduce. The insects induce their feeding site within the meristematic zone of the root tip, where they stay attached, feeding both intra- and intercellularly, and causing damage by reducing plant vigour. Several changes in cell structure and composition, including increased cell division and tissue swelling close to the feeding site, cause an organoid gall called a nodosity to develop. Because alpha expansin genes are involved in cell enlargement and cell wall loosening in many plant tissues it may be anticipated that they are also involved in nodosity formation. To identify expansin genes in Vitis vinifera cv. Pinot noir, we mined for orthologues genes in a comparative analysis. Eleven putative expansin genes were identified and shown to be present in the rootstock Teleki 5C (V. berlandieri Planch. x V. riparia Michx.) using specific PCR followed by DNA sequencing. Expression analysis of young and mature nodosities and uninfested root tips were conducted via quantitative real time PCR (qRT-PCR). Up-regulation was measured for three putative expansin genes (VvEXPA15, -A17 and partly -A20) or down-regulation for three other putative genes (VvEXPA7, -A12, -A20) in nodosities. The present study clearly shows the involvement of putative expansin genes in the phylloxera-root interaction.

  4. A review on the putative association between beta-blockers and depression

    NARCIS (Netherlands)

    Verbeek, D.E.; van Riezen, J.; de Boer, R.A.; van Melle, J.P.; de Jonge, P.

    Several kinds of systematic studies have been conducted verifying the putative association between beta-blockers and depressive symptoms. However, many of these studies had important limitations in their design. In most of the studies, no effect of beta-blockers on depressive symptoms was seen.

  5. A review on the putative association between beta-blockers and depression

    NARCIS (Netherlands)

    Verbeek, D.E.; van Riezen, J.; de Boer, R.A.; van Melle, J.P.; de Jonge, P.

    2011-01-01

    Several kinds of systematic studies have been conducted verifying the putative association between beta-blockers and depressive symptoms. However, many of these studies had important limitations in their design. In most of the studies, no effect of beta-blockers on depressive symptoms was seen. Beca

  6. DETERMINATION OF ROCURONIUM AND ITS PUTATIVE METABOLITES IN BODY-FLUIDS AND TISSUE-HOMOGENATES

    NARCIS (Netherlands)

    KLEEF, UW; PROOST, JH; ROGGEVELD, J

    1993-01-01

    A sensitive and selective HPLC method was developed for the quantification of the neuromuscular blocking agent rocuronium and its putative metabolites (the 17-desacetyl derivative and the N-desallyl derivative of rocuronium) in plasma, urine, bile, tissue homogenates and stoma fluid. Samples were pr

  7. Search strings for the study of putative occupational determinants of disease

    NARCIS (Netherlands)

    S. Mattioli; F. Zanardi; A. Baldasseroni; F. Schaafsma; R.M.T. Cooke; G. Mancini; M. Fierro; C. Santangelo; A. Farioli; S. Fucksia; S. Curti; F.S. Violante; J. Verbeek

    2010-01-01

    Objective To identify efficient PubMed search strategies to retrieve articles regarding putative occupational determinants of conditions not generally considered to be work related. Methods Based on MeSH definitions and expert knowledge, we selected as candidate search terms the four MeSH terms desc

  8. Search strings for the study of putative occupational determinants of disease

    NARCIS (Netherlands)

    Mattioli, S.; Zanardi, F.; Baldasseroni, A.; Schaafsma, F.; Cooke, R.M.T.; Mancini, G.; Fierro, M.; Santangelo, C.; Farioli, A.; Fucksia, S.; Curti, S.; Violante, F.S.; Verbeek, J.

    2010-01-01

    Objective To identify efficient PubMed search strategies to retrieve articles regarding putative occupational determinants of conditions not generally considered to be work related. Methods Based on MeSH definitions and expert knowledge, we selected as candidate search terms the four MeSH terms

  9. Isolation of Bartonella quintana from a Woman and a Cat following Putative Bite Transmission▿

    OpenAIRE

    Breitschwerdt, Edward B; Maggi, Ricardo G; Sigmon, Betsy; Nicholson, William L.

    2006-01-01

    We report here the detection of Bartonella quintana, after putative bite transmission, in pre-enrichment blood cultures from a woman and from two feral barn cats. Prospective molecular epidemiological studies are necessary to characterize the risk of human Bartonella quintana infection following cat bites.

  10. Isolation and characterization of 17 different genes encoding putative endopolygalacturonase genes from Rhizopus oryzae

    Science.gov (United States)

    Polygalacturonase enzymes are a valuable aid in the retting of flax for production of linens and, more recently, production of biofuels from citrus wastes. In a search of the recently sequenced Rhizopus oryzae strain 99-880 genome database, 18 putative endopolygalacturonase genes were identified, w...

  11. A putative morphological substrate of the catecholamine-influenced neuropeptide Y (NPY) release in the human hypothalamus.

    Science.gov (United States)

    Ko, Laam; Rotoli, Giorgio; Grignol, George; Hu, Walter; Merchenthaler, Istvan; Dudas, Bertalan

    2011-06-01

    Neuropeptide Y (NPY) is a 36 amino acid peptide, which among others, plays a pivotal role in stress response. Although previous studies confirmed that NPY release is increased by stress in several species, the exact mechanism of the stress-induced NPY release has not been elucidated yet. In the present study, we examined, with morphological means, the possibility that catecholamines directly influence NPY release in the human hypothalamus. Since the use of electron microscopic techniques is virtually impossible in immunostained human samples due to the long post mortem time, double-label immunohistochemistry was utilised in order to reveal the putative catecholaminergic-NPY associations. The present study is the first to demonstrate juxtapositions between the catecholaminergic, tyrosine hydroxylase (TH)/dopamine-beta hydroxylase (DBH)-immunoreactive (IR) and NPY-IR neural elements in the human hypothalamus. These en passant type associations are most numerous in the infundibular and periventricular areas of the human diencephalon. Here, NPY-IR neurons often form several contacts with catecholaminergic fibre varicosities, without any observable gaps between the contacting elements, suggesting that these juxtapositions may represent functional synapses. The lack of phenylethanolamine N-methyltransferase (PNMT)-NPY juxtapositions and the relatively few observed DBH-NPY associations suggest that the vast majority of the observed TH-NPY juxtapositions represent dopaminergic synapses. Since catecholamines are known to be the crucial components of the stress response, the presence of direct, catecholaminergic (primarily dopaminergic)-NPY-IR synapses may explain the increased NPY release during stress. The released NPY in turn is believed to play an active role in the responses that are directed to maintain the homeostasis during stressful conditions.

  12. A novel human polycomb binding site acts as a functional polycomb response element in Drosophila.

    Directory of Open Access Journals (Sweden)

    Suresh Cuddapah

    Full Text Available Polycomb group (PcG proteins are key chromatin regulators implicated in multiple processes including embryonic development, tissue homeostasis, genomic imprinting, X-chromosome inactivation, and germ cell differentiation. The PcG proteins recognize target genomic loci through cis DNA sequences known as Polycomb Response Elements (PREs, which are well characterized in Drosophila. However, mammalian PREs have been elusive until two groups reported putative mammalian PREs recently. Consistent with the existence of mammalian PREs, here we report the identification and characterization of a potential PRE from human T cells. The putative human PRE has enriched binding of PcG proteins, and such binding is dependent on a key PcG component SUZ12. We demonstrate that the putative human PRE carries both genetic and molecular features of Drosophila PRE in transgenic flies, implying that not only the trans PcG proteins but also certain features of the cis PREs are conserved between mammals and Drosophila.

  13. Divergence of conserved non-coding sequences: rate estimates and relative rate tests.

    Science.gov (United States)

    Wagner, Günter P; Fried, Claudia; Prohaska, Sonja J; Stadler, Peter F

    2004-11-01

    In many eukaryotic genomes only a small fraction of the DNA codes for proteins, but the non-protein coding DNA harbors important genetic elements directing the development and the physiology of the organisms, like promoters, enhancers, insulators, and micro-RNA genes. The molecular evolution of these genetic elements is difficult to study because their functional significance is hard to deduce from sequence information alone. Here we propose an approach to the study of the rate of evolution of functional non-coding sequences at a macro-evolutionary scale. We identify functionally important non-coding sequences as Conserved Non-Coding Nucleotide (CNCN) sequences from the comparison of two outgroup species. The CNCN sequences so identified are then compared to their homologous sequences in a pair of ingroup species, and we monitor the degree of modification these sequences suffered in the two ingroup lineages. We propose a method to test for rate differences in the modification of CNCN sequences among the two ingroup lineages, as well as a method to estimate their rate of modification. We apply this method to the full sequences of the HoxA clusters from six gnathostome species: a shark, Heterodontus francisci; a basal ray finned fish, Polypterus senegalus; the amphibian, Xenopus tropicalis; as well as three mammalian species, human, rat and mouse. The results show that the evolutionary rate of CNCN sequences is not distinguishable among the three mammalian lineages, while the Xenopus lineage has a significantly increased rate of evolution. Furthermore the estimates of the rate parameters suggest that in the stem lineage of mammals the rate of CNCN sequence evolution was more than twice the rate observed within the placental amniotes clade, suggesting a high rate of evolution of cis-regulatory elements during the origin of amniotes and mammals. We conclude that the proposed methods can be used for testing hypotheses about the rate and pattern of evolution of putative

  14. Flow Element Models

    DEFF Research Database (Denmark)

    Heiselberg, Per; Nielsen, Peter V.

    Air distribution in ventilated rooms is a flow process that can be divided into different elements such as supply air jets, exhaust flows, thermal plumes, boundary layer flows, infiltration and gravity currents. These flow elements are isolated volumes where the air movement is controlled...... by a restricted number of parameters, and the air movement is fairly independent of the general flow in the enclosure. In many practical situations, the most convenient· method is to design the air distribution system using flow element theory....

  15. Discovery of element 112

    Energy Technology Data Exchange (ETDEWEB)

    Hofmann, S. [GSI, Darmstadt (Germany)

    1996-12-31

    The new elements 110, 111, and 112 were synthesized and unambiguously identified in experiments at SHIP. Due to strong shell effects the dominant decay mode is not fission, but emission of alpha particles. Theoretical investigations predict that maximum shell effects should exist in nuclei near proton number 114 and neutron number 184. Measurements give hope that isotopes of element 114 close to the island of spherical Superheavy Elements could be produced by fusion reactions using {sup 118}Pb as target. systematic studies of the reaction cross-sections indicate that transfer of nucleons is the important process to initiate the fusion.

  16. Finite element mesh generation

    CERN Document Server

    Lo, Daniel SH

    2014-01-01

    Highlights the Progression of Meshing Technologies and Their ApplicationsFinite Element Mesh Generation provides a concise and comprehensive guide to the application of finite element mesh generation over 2D domains, curved surfaces, and 3D space. Organised according to the geometry and dimension of the problem domains, it develops from the basic meshing algorithms to the most advanced schemes to deal with problems with specific requirements such as boundary conformity, adaptive and anisotropic elements, shape qualities, and mesh optimization. It sets out the fundamentals of popular techniques

  17. Exon B of human surfactant protein A2 mRNA, alone or within its surrounding sequences, interacts with 14-3-3; role of cis-elements and secondary structure.

    Science.gov (United States)

    Noutsios, Georgios T; Silveyra, Patricia; Bhatti, Faizah; Floros, Joanna

    2013-06-01

    Human surfactant protein A, an innate immunity molecule, is encoded by two genes: SFTPA1 (SP-A1) and SFTPA2 (SP-A2). The 5' untranslated (5'UTR) splice variant of SP-A2 (ABD), but not of SP-A1 (AD), contains exon B (eB), which is an enhancer for transcription and translation. We investigated whether eB contains cis-regulatory elements that bind trans-acting factors in a sequence-specific manner as well as the role of the eB mRNA secondary structure. Binding of cytoplasmic NCI-H441 proteins to wild-type eB, eB mutant, AD, and ABD 5'UTR mRNAs were studied by RNA electromobility shift assays (REMSAs). The bound proteins were identified by mass spectroscopy and specific antibodies (Abs). We found that 1) proteins bind eB mRNA in a sequence-specific manner, with two cis-elements identified within eB to be important; 2) eB secondary structure is necessary for binding; 3) mass spectroscopy and specific Abs in REMSAs identified 14-3-3 proteins to bind (directly or indirectly) eB and the natural SP-A2 (ABD) splice variant but not the SP-A1 (AD) splice variant; 4) other ribosomal and cytoskeletal proteins, and translation factors, are also present in the eB mRNA-protein complex; 5) knockdown of 14-3-3 β/α isoform resulted in a downregulation of SP-A2 expression. In conclusion, proteins including the 14-3-3 family bind two cis-elements within eB of hSP-A2 mRNA in a sequence- and secondary structure-specific manner. Differential regulation of SP-A1 and SP-A2 is mediated by the 14-3-3 protein family as well as by a number of other proteins that bind UTRs with or without eB mRNA.

  18. Divergent picornavirus IRES elements

    DEFF Research Database (Denmark)

    Belsham, Graham

    2009-01-01

    Internal ribosome entry site (IRES) elements were first identified about 20 years ago within the 5' untranslated region of picornavirus RNAs. They direct a cap-independent mechanism of translation initiation on the viral RNA. Within the picornavirus family it is now known that there are four...... classes of IRES element which vary in size (450-270nt), they also have different, complex, secondary structures and distinct requirements for cellular proteins to allow them to function. This review describes the features of each class of picornavirus IRES element but focuses on the characteristics...... of the most recently described group, initially identified within the porcine teschovirus-1 RNA, which has strong similarities to the IRES elements from within the genomes of hepatitis C virus and the pestiviruses which are members of the flavivirus family. The selection of the initiation codon...

  19. New functionalities in abundant element oxides: ubiquitous element strategy.

    Science.gov (United States)

    Hosono, Hideo; Hayashi, Katsuro; Kamiya, Toshio; Atou, Toshiyuki; Susaki, Tomofumi

    2011-06-01

    While most ceramics are composed of ubiquitous elements (the ten most abundant elements within the Earth's crust), many advanced materials are based on rare elements. A 'rare-element crisis' is approaching owing to the imbalance between the limited supply of rare elements and the increasing demand. Therefore, we propose a 'ubiquitous element strategy' for materials research, which aims to apply abundant elements in a variety of innovative applications. Creation of innovative oxide materials and devices based on conventional ceramics is one specific challenge. This review describes the concept of ubiquitous element strategy and gives some highlights of our recent research on the synthesis of electronic, thermionic and structural materials using ubiquitous elements.

  20. New roof element system

    DEFF Research Database (Denmark)

    Ditlev, Jesper; Rudbeck, Claus Christian

    1997-01-01

    The aim of the project has been to develop an element system for warm deck roofs which, from a thermal and economical point of view, can deal with the future demands for heat loss coefficients for low slope roofs.......The aim of the project has been to develop an element system for warm deck roofs which, from a thermal and economical point of view, can deal with the future demands for heat loss coefficients for low slope roofs....

  1. Novel porcine repetitive elements

    Directory of Open Access Journals (Sweden)

    Nonneman Dan J

    2006-12-01

    Full Text Available Abstract Background Repetitive elements comprise ~45% of mammalian genomes and are increasingly known to impact genomic function by contributing to the genomic architecture, by direct regulation of gene expression and by affecting genomic size, diversity and evolution. The ubiquity and increasingly understood importance of repetitive elements contribute to the need to identify and annotate them. We set out to identify previously uncharacterized repetitive DNA in the porcine genome. Once found, we characterized the prevalence of these repeats in other mammals. Results We discovered 27 repetitive elements in 220 BACs covering 1% of the porcine genome (Comparative Vertebrate Sequencing Initiative; CVSI. These repeats varied in length from 55 to 1059 nucleotides. To estimate copy numbers, we went to an independent source of data, the BAC-end sequences (Wellcome Trust Sanger Institute, covering approximately 15% of the porcine genome. Copy numbers in BAC-ends were less than one hundred for 6 repeat elements, between 100 and 1000 for 16 and between 1,000 and 10,000 for 5. Several of the repeat elements were found in the bovine genome and we have identified two with orthologous sites, indicating that these elements were present in their common ancestor. None of the repeat elements were found in primate, rodent or dog genomes. We were unable to identify any of the replication machinery common to active transposable elements in these newly identified repeats. Conclusion The presence of both orthologous and non-orthologous sites indicates that some sites existed prior to speciation and some were generated later. The identification of low to moderate copy number repetitive DNA that is specific to artiodactyls will be critical in the assembly of livestock genomes and studies of comparative genomics.

  2. New roof element system

    DEFF Research Database (Denmark)

    Ditlev, Jesper; Rudbeck, Claus Christian

    1997-01-01

    The aim of the project has been to develop an element system for warm deck roofs which, from a thermal and economical point of view, can deal with the future demands for heat loss coefficients for low slope roofs.......The aim of the project has been to develop an element system for warm deck roofs which, from a thermal and economical point of view, can deal with the future demands for heat loss coefficients for low slope roofs....

  3. Structural elements design manual

    CERN Document Server

    Draycott, Trevor

    2012-01-01

    Gives clear explanations of the logical design sequence for structural elements. The Structural Engineer says: `The book explains, in simple terms, and with many examples, Code of Practice methods for sizing structural sections in timber, concrete,masonry and steel. It is the combination into one book of section sizing methods in each of these materials that makes this text so useful....Students will find this an essential support text to the Codes of Practice in their study of element sizing'.

  4. A battery element

    Energy Technology Data Exchange (ETDEWEB)

    Vatanabe, U.; Aoki, K.; Ito, K.; Ogava, K.; Okadzaki, R.

    1983-07-29

    An anode made of a light metal is used in the element, along with an anhydrous liquid electrolyte and a cathode made of CuC12(CFn)x or another material. The current tap of the anode is made from aluminum, gold, silver or another metal of the platinum group and the current tap may be coated with this metal. The thickness of the coating is 0.1 to 1 micrometer. The element has a long storage life.

  5. Atoms, molecules & elements

    CERN Document Server

    Graybill, George

    2007-01-01

    Young scientists will be thrilled to explore the invisible world of atoms, molecules and elements. Our resource provides ready-to-use information and activities for remedial students using simplified language and vocabulary. Students will label each part of the atom, learn what compounds are, and explore the patterns in the periodic table of elements to find calcium (Ca), chlorine (Cl), and helium (He) through hands-on activities.

  6. Mobilization of horizontally acquired island 2 is induced in planta in the phytopathogen Pectobacterium atrosepticum SCRI1043 and involves the putative relaxase ECA0613 and quorum sensing.

    Science.gov (United States)

    Vanga, Bhanupratap R; Ramakrishnan, Pavithra; Butler, Ruth C; Toth, Ian K; Ronson, Clive W; Jacobs, Jeanne M E; Pitman, Andrew R

    2015-11-01

    Integrative and conjugative elements (ICEs) contribute to the rapid evolution of bacterial pathogens via horizontal gene transfer of virulence determinants. ICEs have common mechanisms for transmission, yet the cues triggering this process under natural environmental or physiological conditions are largely unknown. In this study, mobilization of the putative ICE horizontally acquired island 2 (HAI2), present in the chromosome of the phytopathogen Pectobacterium atrosepticum SCRI1043, was examined during infection of the host plant potato. Under these conditions, mobilization of HAI2 increased markedly compared with in vitro cultures. In planta-induced mobilization of HAI2 was regulated by quorum sensing and involved the putative ICE-encoded relaxase ECA0613. Disruption of ECA0613 also reduced transcription of genes involved in production of coronafacic acid (Cfa), the major virulence factor harboured on HAI2, whereas their expression was unaffected in the quorum-sensing (expI) mutant. Thus, suppression of cfa gene expression was not regulated by the mobilization of the ICE per se, but was due directly to inactivation of the relaxase. The identification of genetic factors associated solely with in planta mobilization of an ICE demonstrates that this process is highly adapted to the natural environment of the bacterial host and can influence the expression of virulence determinants.

  7. Putative and unique gene sequence utilization for the design of species specific probes as modeled by Lactobacillus plantarum

    Science.gov (United States)

    The concept of utilizing putative and unique gene sequences for the design of species specific probes was tested. The abundance profile of assigned functions within the Lactobacillus plantarum genome was used for the identification of the putative and unique gene sequence, csh. The targeted gene (cs...

  8. Trace elements in dialysis.

    Science.gov (United States)

    Filler, Guido; Felder, Sarah

    2014-08-01

    In end-stage chronic kidney disease (CKD), pediatric nephrologists must consider the homeostasis of the multiple water-soluble ions that are influenced by renal replacement therapy (RRT). While certain ions such as potassium and calcium are closely monitored, little is known about the handling of trace elements in pediatric dialysis. RRT may lead to accumulation of toxic trace elements, either due to insufficient elimination or due to contamination, or to excessive removal of essential trace elements. However, trace elements are not routinely monitored in dialysis patients and no mechanism for these deficits or toxicities has been established. This review summarizes the handling of trace elements, with particular attention to pediatric data. The best data describe lead and indicate that there is a higher prevalence of elevated lead (Pb, atomic number 82) levels in children on RRT when compared to adults. Lead is particularly toxic in neurodevelopment and lead levels should therefore be monitored. Monitoring of zinc (Zn, atomic number 30) and selenium (Se, atomic number 34) may be indicated in the monitoring of all pediatric dialysis patients to reduce morbidity from deficiency. Prospective studies evaluating the impact of abnormal trace elements and the possible therapeutic value of intervention are required.

  9. Negative regulation of P element excision by the somatic product and terminal sequences of P in drosophila melanogaster

    Science.gov (United States)

    A transient in vivo P element excision assay was used to test the regulatory properties of putative repressor-encoding plasmids in Drosophila melanogaster embryos. The somatic expression of an unmodified transposase transcription unit under the control of a heat shock gene promoter (phsn) effectivel...

  10. Hippocampal and thalamic neuronal metabolism in a putative rat model of schizophrenia○

    Institute of Scientific and Technical Information of China (English)

    Guolin Ma; Tianbin Song; Min Chen; Yuan Fu; Yong Xu; Ensen Ma; Wu Wang; Jiang Du; Mingxiong Huang

    2013-01-01

    The transcription factor early growth response protein 3 (EGR3) is involved in schizophrenia. We developed a putative rat model of schizophrenia by transfecting lentiviral particles carrying the Egr3 gene into bilateral hippocampal dentate gyrus. We assessed spatial working memory using the Morris water maze test, and neuronal metabolite levels in bilateral hippocampus and thalamus were determined by 3.0 T proton magnetic resonance spectroscopy. Choline content was significantly greater in the hippocampus after transfection, while N-acetylaspartate and the ratio of N-acetylaspartate to creatine/phosphocreatine in the thalamus were lower than in controls. This study is the first to report evaluation of brain metabolites using 3.0 T proton magnetic resonance spectroscopy in rats transfected with Egr3, and reveals metabolic abnormalities in the hippocampus and thalamus in this putative model of schizophrenia.

  11. Functional characterization of a putative β-lactamase gene in the genome of Zymomonas mobilis.

    Science.gov (United States)

    Rajnish, K Narayanan; Asraf, Sheik Abdul Kader Sheik; Manju, Nagarajan; Gunasekaran, Paramasamy

    2011-12-01

    Zymomonas mobilis ZM4 is resistant to β-lactam antibiotics but there are no reports of a β-lactam resistance gene and its regulation. A putative β-lactamase gene sequence (ZMO0103) in the genome of Z. mobilis showed a 55% amino acid sequence identity with class C β-lactamase genes. qPCR analysis of the β-lactamase transcript indicated a higher level expression of the β-lactamase compared to the relative transcript quantities in antibiotic-susceptible bacteria. The putative β-lactamase gene was cloned, expressed in Escherichia coli BL21 and the product, AmpC, was purified to homogeneity. Its optimal activity was at pH 6 and 30 °C. Further, the β-lactamase had a higher affinity towards penicillins than cephalosporin antibiotics. © Springer Science+Business Media B.V. 2011

  12. Putative Biomarkers and Targets of Estrogen Receptor Negative Human Breast Cancer

    Directory of Open Access Journals (Sweden)

    Stephen W. Byers

    2011-07-01

    Full Text Available Breast cancer is a progressive and potentially fatal disease that affects women of all ages. Like all progressive diseases, early and reliable diagnosis is the key for successful treatment and annihilation. Biomarkers serve as indicators of pathological, physiological, or pharmacological processes. Her2/neu, CA15.3, estrogen receptor (ER, progesterone receptor (PR, and cytokeratins are biomarkers that have been approved by the Food and Drug Administration for disease diagnosis, prognosis, and therapy selection. The structural and functional complexity of protein biomarkers and the heterogeneity of the breast cancer pathology present challenges to the scientific community. Here we review estrogen receptor-related putative breast cancer biomarkers, including those of putative breast cancer stem cells, a minor population of estrogen receptor negative tumor cells that retain the stem cell property of self renewal. We also review a few promising cytoskeleton targets for ER alpha negative breast cancer.

  13. A putative ABC transporter is involved in negative regulation of biofilm formation by Listeria monocytogenes

    DEFF Research Database (Denmark)

    Zhu, Xinna; Long, Fei; Chen, Yonghui

    2008-01-01

    Listeria monocytogenes may persist for long periods in food processing environments. In some instances, this may be due to aggregation or biofilm formation. To investigate the mechanism controlling biofilm formation in the food-borne pathogen L. monocytogenes, we characterized LM-49, a mutant...... with enhanced ability of biofilm-formation generated via transposon Tn917 mutagenesis of L. monocytogenes 4b G. In this mutant, a Tn917 insertion has disrupted the coding region of the gene encoding a putative ATP binding cassette (ABC) transporter permease identical to Lmof2365_1771 (a putative ABC......-transporter permease) presented in the sequenced strain L. monocytogenes str. 4b F2365. This disrupted gene, denoted lm.G_1771, encoded a protein with 10 transmembrane helixes. The revertant, LM-49RE, was obtained by replacing lm.G_1771::Tn917 with lm.G_1771 via homologous recombination. We found that LM-49RE formed...

  14. The solution structure of ChaB, a putative membrane ion antiporter regulator from Escherichia coli

    Directory of Open Access Journals (Sweden)

    Iannuzzi Pietro

    2004-08-01

    Full Text Available Abstract Background ChaB is a putative regulator of ChaA, a Na+/H+ antiporter that also has Ca+/H+ activity in E. coli. ChaB contains a conserved 60-residue region of unknown function found in other bacteria, archaeabacteria and a series of baculoviral proteins. As part of a structural genomics project, the structure of ChaB was elucidated by NMR spectroscopy. Results The structure of ChaB is composed of 3 α-helices and a small sheet that pack tightly to form a fold that is found in the cyclin-box family of proteins. Conclusion ChaB is distinguished from its putative DNA binding sequence homologues by a highly charged flexible loop region that has weak affinity to Mg2+ and Ca2+ divalent metal ions.

  15. Exploring Universal Partnerships and Putative Marriages as Tools for Awarding Partnership Property in Contemporary Family Law

    Directory of Open Access Journals (Sweden)

    Elsje Bonthuys

    2016-12-01

    Full Text Available Following upon the Supreme Court of Appeal's judgment in Butters v Mncora 2012 4 SA 1 (SCA, which broadened the criteria and consequences of universal partnerships in cohabitation relationships, this article investigates the potential of universal partnerships and putative marriages to allocate rights to share in partnership property in other intimate relationships. It traverses several instances in which marriages are not recognised - bigamous marriages, Muslim and Hindu religious marriages and invalid customary marriages – examining whether the wives in these marriages could use universal partnerships and putative marriages to claim a share in property. It then considers the use of universal partnerships to obtain a share of property in civil marriages out of community of property. It concludes by pointing out several issues which are in need of clarification and where the common law should be developed to give effect to fundamental constitutional rights.

  16. Enrichment of putative stem cells from adipose tissue using dielectrophoretic field-flow fractionation

    Science.gov (United States)

    Vykoukal, Jody; Vykoukal, Daynene M.; Freyberg, Susanne; Alt, Eckhard U.; Gascoyne, Peter R. C.

    2009-01-01

    We have applied the microfluidic cell separation method of dielectrophoretic field-flow fractionation (DEP-FFF) to the enrichment of a putative stem cell population from an enzyme-digested adipose tissue derived cell suspension. A DEP-FFF separator device was constructed using a novel microfluidic-microelectronic hybrid flex-circuit fabrication approach that is scaleable and anticipates future low-cost volume manufacturing. We report the separation of a nucleated cell fraction from cell debris and the bulk of the erythrocyte population, with the relatively rare (<2% starting concentration) NG2-positive cell population (pericytes and/or putative progenitor cells) being enriched up to 14-fold. This work demonstrates a potential clinical application for DEP-FFF and further establishes the utility of the method for achieving label-free fractionation of cell subpopulations. PMID:18651083

  17. Composition of the Putative Prepore Complex of Bacillus thuringiensis Cry1Ab Toxin

    Science.gov (United States)

    Nair, Manoj S.; Dean, Donald H.

    2015-01-01

    Prepore formation is hypothesized to be an obligate step in the insertion of Cry1Ab toxin into insect brush border membrane vesicles. We examined the architecture of the putative prepore when isolated using the published protocols [1] [2]. Our results demonstrate that the putative prepore form of Cry1Ab is a combination of receptor proteins attached to the toxin, when purified. The results also suggest that this prepore form as prepared by the methods published is different from other membrane-extracted oligomeric forms of Cry toxins and prepore of other toxins in general. While most other known prepores are composed of multimers of a single protein, the Cry1Ab prepore, as generated, is a protein-receptor complex oligomer and monomers of Cry toxins. PMID:26702367

  18. A putative autonomous 20.5 kb-CACTA transposon insertion in an F3'H allele identifies a new CACTA transposon subfamily in Glycine max

    Directory of Open Access Journals (Sweden)

    Vodkin Lila

    2008-12-01

    Full Text Available Abstract Background The molecular organization of very few genetically defined CACTA transposon systems have been characterized thoroughly as those of Spm/En in maize, Tam1 of Antirrhinum majus Candystripe1 (Cs1 from Sorghum bicolor and CAC1 from Arabidopsis thaliana, for example. To date, only defective deletion derivatives of CACTA elements have been described for soybean, an economically important plant species whose genome sequence will be completed in 2008. Results We identified a 20.5 kb insertion in a soybean flavonoid 3'-hydroxylase (F3'H gene representing the t* allele (stable gray trichome color whose origin traces to a single mutable chimeric plant displaying both tawny and gray trichomes. This 20.5 kb insertion has the molecular structure of a putative autonomous transposon of the CACTA family, designated Tgmt*. It encodes a large gene that was expressed in two sister isolines (T* and tm of the stable gray line (t* from which Tgmt* was isolated. RT-PCR derived cDNAs uncovered the structure of a large precursor mRNA as well as alternatively spliced transcripts reminiscent of the TNPA-mRNA generated by the En-1 element of maize but without sequence similarity to the maize TNPA. The larger mRNA encodes a transposase with a tnp2 and TNP1-transposase family domains. Because the two soybean lines expressing Tgmt* were derived from the same mutable chimeric plant that created the stable gray trichome t* allele line from which the element was isolated, Tgmt* has the potential to be an autonomous element that was rapidly inactivated in the stable gray trichome t* line. Comparison of Tgmt* to previously described Tgm elements demonstrated that two subtypes of CACTA transposon families exist in soybean based on divergence of their characteristic subterminal repeated motifs and their transposases. In addition, we report the sequence and annotation of a BAC clone containing the F3'H gene (T locus which was interrupted by the novel Tgmt* element

  19. Identification of putative effector genes and their transcripts in three strains related to 'Candidatus Phytoplasma aurantifolia'.

    Science.gov (United States)

    Anabestani, Ameneh; Izadpanah, Keramat; Abbà, Simona; Galetto, Luciana; Ghorbani, Abozar; Palmano, Sabrina; Siampour, Majid; Veratti, Flavio; Marzachì, Cristina

    2017-06-01

    Molecular mechanisms underlying phytoplasma interactions with host plants are largely unknown. In this study attempts were made to identify effectors of three phytoplasma strains related to 'Ca. P. aurantifolia', crotalaria phyllody (CrP), faba bean phyllody (FBP), and witches' broom disease of lime (WBDL), using information from draft genome of peanut witches' broom phytoplasma. Seven putative effectors were identified in WBDL genome (SAP11, SAP21, Eff64, Eff115, Eff197, Eff211 and EffSAP67), five (SAP11, SAP21, Eff64, Eff99 and Eff197) in CrP and two (SAP11, Eff64) in FBP. No homologs to Eff64, Eff197 and Eff211 in phytoplasmas of other phylogenetic groups were found. SAP11 and Eff64 homologs of 'Ca. P. aurantifolia' strains shared at least 95.9% identity and were detected in the three phytoplasmas, supporting their role within the group. Five of the putative effectors (SAP11, SAP21, Eff64, Eff115, and Eff99) were transcribed from total RNA extracts of periwinkle plants infected with these phytoplasmas. Transcription profiles of selected putative effectors of CrP, FBP and WBDL indicated that SAP11 transcripts were the most abundant in the three phytoplasmas. SAP21 transcript levels were comparable to those of SAP11 for CrP and not measurable for the other phytoplasmas. Eff64 had the lowest transcription level irrespective of sampling date and phytoplasma isolate. Eff115 transcript levels were the highest in WBDL infected plants. This work reports the first sequence information for 14 putative effectors in three strains related to 'Ca. P. aurantifolia', and offers novel insight into the transcription profile of five of them during infection of periwinkle. Copyright © 2017 Elsevier GmbH. All rights reserved.

  20. A novel putative auxin carrier family regulates intracellular auxin homeostasis in plants

    OpenAIRE

    2012-01-01

    The phytohormone auxin acts as a prominent signal, providing, by its local accumulation or depletion in selected cells, a spatial and temporal reference for changes in the developmental program. The distribution of auxin depends on both auxin metabolism (biosynthesis, conjugation and degradation) and cellular auxin transport. We identified in silico a novel putative auxin transport facilitator family, called PIN-LIKES (PILS). Here we illustrate that PILS proteins are required for auxin-depend...

  1. ANTIBIOTICS RESISTANCE AND PUTATIVE VIRULENCE FACTORS OF AEROMONAS HYDROPHILA ISOLATED FROM ESTUARY

    OpenAIRE

    Olumide Adedokun Odeyemi; Ahmad Asmat; Gires Usup

    2012-01-01

    This study aim to investigate antibiotics resistance profile and putative virulence factors of Aeromonas hydrophila isolated from estuary. Bacteria used for this study were isolated from water and sediment samples obtained from Sungai Melayu, Johor, Malaysia. Serially diluted 100 µL water and 1g sediment were inoculated on modified Rimler - Shott (mRS) agar. Colonies with distinct cultural characteristics were picked for further studies. Isolates were tested for biofilm productions, protease ...

  2. Bioinformatic Analysis of Putative Gene Products Encoded in SARS-HCoV Genome

    Institute of Scientific and Technical Information of China (English)

    赵心刚; 韩敬东; 宁元亨; 孟安明; 陈晔光

    2003-01-01

    The cause of severe acute respiratory syndrome (SARS) has been identified as a new coronavirus named as SARS-HCoV.Using bioinformatic methods, we have performed a detailed domain search.In addition to the viral structure proteins, we have found that several putative polypeptides share sequence similarity to known domains or proteins.This study may provide a basis for future studies on the infection and replication process of this notorious virus.

  3. Detection of putative periodontopathic bacteria in type 1 diabetic and healthy children: A comparative study

    OpenAIRE

    2013-01-01

    Aim: The aim of this study was to compare and assess the risk of periodontitis due to the presence of four putative periodontopathic bacteria (Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, and Aggregatibacter actinomycetemcomitans) in type 1 diabetic and healthy children. Materials and Methods: Fifty type 1 diabetic and 50 healthy children in the age group of 7-14 years were recruited for the study. Subgingival plaque samples collected from permanent first molars w...

  4. Transposable elements belonging to the Tc1-Mariner superfamily are heavily mutated in Colletotrichum graminicola.

    Science.gov (United States)

    Braga, Raíssa Mesquita; Santana, Mateus Ferreira; Veras da Costa, Rodrigo; Brommonschenkel, Sergio Herminio; de Araújo, Elza Fernandes; de Queiroz, Marisa Vieira

    2014-01-01

    Transposable elements are ubiquitous and constitute an important source of genetic variation in addition to generating deleterious mutations. Several filamentous fungi are able to defend against transposable elements using RIP(repeat-induced point mutation)-like mechanisms, which induce mutations in duplicated sequences. The sequenced Colletotrichum graminicola genome and the availability of transposable element databases provide an efficient approach for identifying and characterizing transposable elements in this fungus, which was the subject of this study. We identified 132 full-sized Tc1-Mariner transposable elements in the sequenced C. graminicola genome, which were divided into six families. Several putative transposases that have been found in these elements have conserved DDE motifs, but all are interrupted by stop codons. An in silico analysis showed evidence for RIP-generated mutations. The TCg1 element, which was cloned from the Brazilian 2908 m isolate, has a putative transposase sequence with three characteristic conserved motifs. However, this sequence is interrupted by five stop codons. Genomic DNA from various isolates was analyzed by hybridization with an internal region of TCg1. All of the isolates featured transposable elements that were similar to TCg1, and several hybridization profiles were identified. C. graminicola has many Tc1-Mariner transposable elements that have been degenerated by characteristic RIP mutations. It is unlikely that any of the characterized elements are autonomous in the sequenced isolate. The possible existence of active copies in field isolates from Brazil was shown. The TCg1 element is present in several C. graminicola isolates and is a potentially useful molecular marker for population studies of this phytopathogen. © 2014 by The Mycological Society of America.

  5. Immunodiagnosis of episomal Banana streak MY virus using polyclonal antibodies to an expressed putative coat protein.

    Science.gov (United States)

    Sharma, Susheel Kumar; Kumar, P Vignesh; Baranwal, Virendra Kumar

    2014-10-01

    A cryptic Badnavirus species complex, known as banana streak viruses (BSV) poses a serious threat to banana production and genetic improvement worldwide. Due to the presence of integrated BSV sequences in the banana genome, routine detection is largely based on serological and nucleo-serological diagnostic methods which require high titre specific polyclonal antiserum. Viral structural proteins like coat protein (CP) are the best target for in vitro expression, to be used as antigen for antiserum production. However, in badnaviruses precise CP sequences are not known. In this study, two putative CP coding regions (p48 and p37) of Banana streak MY virus (BSMYV) were identified in silico by comparison with caulimoviruses, retroviruses and Rice tungro bacilliform virus. The putative CP coding region (p37) was in vitro expressed in pMAL system and affinity purified. The purified fusion protein was used as antigen for raising polyclonal antiserum in rabbit. The specificity of antiserum was confirmed in Western blots, immunosorbent electron microscopy (ISEM) and antigen coated plate-enzyme linked immunosorbent assay (ACP-ELISA). The antiserum (1:2000) was successfully used in ACP-ELISA for specific detection of BSMYV infection in field and tissue culture raised banana plants. The antiserum was also utilized in immuno-capture PCR (IC-PCR) based indexing of episomal BSMYV infection. This is the first report of in silico identification of putative CP region of BSMYV, production of polyclonal antiserum against recombinant p37 and its successful use in immunodetection.

  6. Identification of Putative Coffee Rust Mycoparasites via Single-Molecule DNA Sequencing of Infected Pustules.

    Science.gov (United States)

    James, Timothy Y; Marino, John A; Perfecto, Ivette; Vandermeer, John

    2015-11-13

    The interaction of crop pests with their natural enemies is a fundament to their control. Natural enemies of fungal pathogens of crops are poorly known relative to those of insect pests, despite the diversity of fungal pathogens and their economic importance. Currently, many regions across Latin America are experiencing unprecedented epidemics of coffee rust (Hemileia vastatrix). Identification of natural enemies of coffee rust could aid in developing management strategies or in pinpointing species that could be used for biocontrol. In the present study, we characterized fungal communities associated with coffee rust lesions by single-molecule DNA sequencing of fungal rRNA gene bar codes from leaf discs (≈28 mm(2)) containing rust lesions and control discs with no rust lesions. The leaf disc communities were hyperdiverse in terms of fungi, with up to 69 operational taxonomic units (putative species) per control disc, and the diversity was only slightly reduced in rust-infected discs, with up to 63 putative species. However, geography had a greater influence on the fungal community than whether the disc was infected by coffee rust. Through comparisons between control and rust-infected leaf discs, as well as taxonomic criteria, we identified 15 putative mycoparasitic fungi. These fungi are concentrated in the fungal family Cordycipitaceae and the order Tremellales. These data emphasize the complexity of diverse fungi of unknown ecological function within a leaf that might influence plant disease epidemics or lead to the development of species for biocontrol of fungal disease.

  7. Emissions of putative isoprene oxidation products from mango branches under abiotic stress

    Science.gov (United States)

    Jardine, Kolby J.; Meyers, Kimberly; Abrell, Leif; Alves, Eliane G.; Yanez Serrano, Ana Maria; Kesselmeier, Jürgen; Karl, Thomas; Guenther, Alex; Vickers, Claudia; Chambers, Jeffrey Q.

    2013-01-01

    Although several per cent of net carbon assimilation can be re-released as isoprene emissions to the atmosphere by many tropical plants, much uncertainty remains regarding its biological significance. In a previous study, we detected emissions of isoprene and its oxidation products methyl vinyl ketone (MVK) and methacrolein (MACR) from tropical plants under high temperature/light stress, suggesting that isoprene is oxidized not only in the atmosphere but also within plants. However, a comprehensive analysis of the suite of isoprene oxidation products in plants has not been performed and production relationships with environmental stress have not been described. In this study, putative isoprene oxidation products from mango (Mangifera indica) branches under abiotic stress were first identified. High temperature/light and freeze–thaw treatments verified direct emissions of the isoprene oxidation products MVK and MACR together with the first observations of 3-methyl furan (3-MF) and 2-methyl-3-buten-2-ol (MBO) as putative novel isoprene oxidation products. Mechanical wounding also stimulated emissions of MVK and MACR. Photosynthesis under 13CO2 resulted in rapid (<30min) labelling of up to five carbon atoms of isoprene, with a similar labelling pattern observed in the putative oxidation products. These observations highlight the need to investigate further the mechanisms of isoprene oxidation within plants under stress and its biological and atmospheric significance. PMID:23881400

  8. Cloning and molecular characterization of a putative voltage-gated sodium channel gene in the crayfish.

    Science.gov (United States)

    Coskun, Cagil; Purali, Nuhan

    2016-06-01

    Voltage-gated sodium channel genes and associated proteins have been cloned and studied in many mammalian and invertebrate species. However, there is no data available about the sodium channel gene(s) in the crayfish, although the animal has frequently been used as a model to investigate various aspects of neural cellular and circuit function. In the present work, by using RNA extracts from crayfish abdominal ganglia samples, the complete open reading frame of a putative sodium channel gene has firstly been cloned and molecular properties of the associated peptide have been analyzed. The open reading frame of the gene has a length of 5793 bp that encodes for the synthesis of a peptide, with 1930 amino acids, that is 82% similar to the α-peptide of a sodium channel in a neighboring species, Cancer borealis. The transmembrane topology analysis of the crayfish peptide indicated a pattern of four folding domains with several transmembrane segments, as observed in other known voltage-gated sodium channels. Upon analysis of the obtained sequence, functional regions of the putative sodium channel responsible for the selectivity filter, inactivation gate, voltage sensor, and phosphorylation have been predicted. The expression level of the putative sodium channel gene, as defined by a qPCR method, was measured and found to be the highest in nervous tissue.

  9. Putative periodontopathic bacteria and herpesviruses in pregnant women: a case-control study

    Science.gov (United States)

    Lu, Haixia; Zhu, Ce; Li, Fei; Xu, Wei; Tao, Danying; Feng, Xiping

    2016-01-01

    Little is known about herpesvirus and putative periodontopathic bacteria in maternal chronic periodontitis. The present case-control study aimed to explore the potential relationship between putative periodontopathic bacteria and herpesviruses in maternal chronic periodontitis.Saliva samples were collected from 36 pregnant women with chronic periodontitis (cases) and 36 pregnant women with healthy periodontal status (controls). Six putative periodontopathic bacteria (Porphyromonas gingivalis [Pg], Aggregatibacer actinomycetemcomitans [Aa], Fusobacterium nucleatum [Fn], Prevotella intermedia [Pi], Tannerella forsythia [Tf], and Treponema denticola [Td]) and three herpesviruses (Epstein-Barr virus [EBV], human cytomegalovirus [HCMV], and herpes simplex virus [HSV]) were detected. Socio-demographic data and oral health related behaviors, and salivary estradiol and progesterone levels were also collected. The results showed no significant differences in socio-demographic background, oral health related behaviors, and salivary estradiol and progesterone levels between the two groups (all P > 0.05). The detection rates of included periodontopathic microorganisms were not significantly different between the two groups (all P > 0.05), but the coinfection rate of EBV and Pg was significantly higher in the case group than in the control group (P = 0.028). EBV and Pg coinfection may promote the development of chronic periodontitis among pregnant women. PMID:27301874

  10. Genomic identification of a putative circadian system in the cladoceran crustacean Daphnia pulex

    Science.gov (United States)

    Tilden, Andrea R.; McCoole, Matthew D.; Harmon, Sarah M.; Baer, Kevin N.; Christie, Andrew E.

    2011-01-01

    Essentially nothing is known about the molecular underpinnings of crustacean circadian clocks. The genome of Daphnia pulex, the only crustacean genome available for public use, provides a unique resource for identifying putative circadian proteins in this species. Here, the Daphnia genome was mined for putative circadian protein genes using Drosophila melanogaster queries. The sequences of core clock (e.g. CLOCK, CYCLE, PERIOD, TIMELESS and CRYPTOCHROME 2), clock input (CRYPTOCHROME 1) and clock output (PIGMENT DISPERSING HORMONE RECEPTOR) proteins were deduced. Structural analyses and alignment of the Daphnia proteins with their Drosophila counterparts revealed extensive sequence conservation, particularly in functional domains. Comparisons of the Daphnia proteins with other sequences showed that they are, in most cases, more similar to homologs from other species, including vertebrates, than they are to those of Drosophila. The presence of both CRYPTOCHROME 1 and 2 in Daphnia suggests the organization of its clock may be more similar to that of the butterfly Danaus plexippus than to that of Drosophila (which possesses CRYPTOCHROME 1 but not CRYPTOCHROME 2). These data represent the first description of a putative circadian system from any crustacean, and provide a foundation for future molecular, anatomical and physiological investigations of circadian signaling in Daphnia. PMID:21798832

  11. Phylogeny of algal sequences encoding carbohydrate sulfotransferases, formylglycine-dependent sulfatases and putative sulfatase modifying factors

    Directory of Open Access Journals (Sweden)

    Chai-Ling eHo

    2015-11-01

    Full Text Available Many algae are rich sources of sulfated polysaccharides with biological activities. The physicochemical/rheological properties and biological activities of sulfated polysaccharides are affected by the pattern and number of sulfate moieties. Sulfation of carbohydrates is catalyzed by carbohydrate sulfotransferases (CHSTs while modification of sulfate moieties on sulfated polysaccharides was presumably catalyzed by sulfatases including formylglycine-dependent sulfatases (FGly-SULFs. Post-translationally modification of Cys to FGly in FGly-SULFs by sulfatase modifiying factors (SUMFs is necessary for the activity of this enzyme. The aims of this study are to mine for sequences encoding algal CHSTs, FGly-SULFs and putative SUMFs from the fully sequenced algal genomes and to infer their phylogenetic relationships to their well characterized counterparts from other organisms. Algal sequences encoding CHSTs, FGly-SULFs, SUMFs and SUMF-like proteins were successfully identified from green and brown algae. However, red algal FGly-SULFs and SUMFs were not identified. In addition, a group of SUMF-like sequences with different gene structure and possibly different functions were identified for green, brown and red algae. The phylogeny of these putative genes contributes to the corpus of knowledge of an unexplored area. The analyses of these putative genes contribute towards future production of existing and new sulfated carbohydrate polymers through enzymatic synthesis and metabolic engineering.

  12. Fuel Element Technical Manual

    Energy Technology Data Exchange (ETDEWEB)

    Burley, H.H. [ed.

    1956-08-01

    It is the purpose of the Fuel Element Technical Manual to Provide a single document describing the fabrication processes used in the manufacture of the fuel element as well as the technical bases for these processes. The manual will be instrumental in the indoctrination of personnel new to the field and will provide a single data reference for all personnel involved in the design or manufacture of the fuel element. The material contained in this manual was assembled by members of the Engineering Department and the Manufacturing Department at the Hanford Atomic Products Operation between the dates October, 1955 and June, 1956. Arrangement of the manual. The manual is divided into six parts: Part I--introduction; Part II--technical bases; Part III--process; Part IV--plant and equipment; Part V--process control and improvement; and VI--safety.

  13. Creativity Management Key Elements

    Directory of Open Access Journals (Sweden)

    Rosa María Fuchs Ángeles

    2015-09-01

    Full Text Available Organizations are constantly looking towards innovation. In order to reach it they must foment creativity. This paper analyzes a series of elements considered in the organizational creativity management and proposes a model with the indispensable factors that organizations should consider to reach it. These elements are: culture and organizational environment, strategy, structure, communication, relation with customers, human resources (recruiting, training, job design, compensation, promotion, and performance evaluation, long term orientation and the organizational life cycle. Having the analysis of those elements as a basis, the indispensable pillars on management creativity are identified. The proposed model is based on 5 pillars: the alignment between strategic, culture and organizational structure, called by the authors 'Holy Trinity'; intern publicity; customer’s voice; recognition and a look towards future. Finally, the case of an innovative Peruvian enterprise is presented from the model’s perspective and the study conclusions.

  14. Elemental analysis in biotechnology.

    Science.gov (United States)

    Hann, Stephan; Dernovics, Mihaly; Koellensperger, Gunda

    2015-02-01

    This article focuses on analytical strategies integrating atomic spectroscopy in biotechnology. The rationale behind developing such methods is inherently linked to unique features of the key technique in elemental analysis, which is inductively coupled plasma mass spectrometry: (1) the high sensitivity and selectivity of state of the art instrumentation, (2) the possibility of accurate absolute quantification even in complex matrices, (3) the capability of combining elemental detectors with chromatographic separation methods and the versatility of the latter approach, (4) the complementarity of inorganic and organic mass spectrometry, (5) the multi-element capability and finally (6) the capability of isotopic analysis. The article highlights the most recent bio-analytical developments exploiting these methodological advantages and shows the potential in biotechnological applications.

  15. Advanced finite element technologies

    CERN Document Server

    Wriggers, Peter

    2016-01-01

    The book presents an overview of the state of research of advanced finite element technologies. Besides the mathematical analysis, the finite element development and their engineering applications are shown to the reader. The authors give a survey of the methods and technologies concerning efficiency, robustness and performance aspects. The book covers the topics of mathematical foundations for variational approaches and the mathematical understanding of the analytical requirements of modern finite element methods. Special attention is paid to finite deformations, adaptive strategies, incompressible, isotropic or anisotropic material behavior and the mathematical and numerical treatment of the well-known locking phenomenon. Beyond that new results for the introduced approaches are presented especially for challenging nonlinear problems.

  16. An alkaline element

    Energy Technology Data Exchange (ETDEWEB)

    Arita, T.; Murakami, K.; Okha, K.

    1983-04-28

    A cathode with a dual layer active mass is installed in the disk shaped alkaline silver and zinc element. The first layer, which is turned towards the anode, contains 85 parts Ag2O, 5 parts electrolytic MnO2 and 10 parts graphite. The second layer, which contacts the bottom of the element, contains 35 parts Ag2O, 60 parts electrolytic MnO2 and 5 parts graphite. The electrical capacity of the first and second layers is 60 and 40, respectively. The first layer may be discharged with a high current density and the second layer with less current density. The element has high characteristics with comparatively low cost.

  17. Intelligent Elements for ISHM

    Science.gov (United States)

    Schmalzel, John L.; Morris, Jon; Turowski, Mark; Figueroa, Fernando; Oostdyk, Rebecca

    2008-01-01

    There are a number of architecture models for implementing Integrated Systems Health Management (ISHM) capabilities. For example, approaches based on the OSA-CBM and OSA-EAI models, or specific architectures developed in response to local needs. NASA s John C. Stennis Space Center (SSC) has developed one such version of an extensible architecture in support of rocket engine testing that integrates a palette of functions in order to achieve an ISHM capability. Among the functional capabilities that are supported by the framework are: prognostic models, anomaly detection, a data base of supporting health information, root cause analysis, intelligent elements, and integrated awareness. This paper focuses on the role that intelligent elements can play in ISHM architectures. We define an intelligent element as a smart element with sufficient computing capacity to support anomaly detection or other algorithms in support of ISHM functions. A smart element has the capabilities of supporting networked implementations of IEEE 1451.x smart sensor and actuator protocols. The ISHM group at SSC has been actively developing intelligent elements in conjunction with several partners at other Centers, universities, and companies as part of our ISHM approach for better supporting rocket engine testing. We have developed several implementations. Among the key features for these intelligent sensors is support for IEEE 1451.1 and incorporation of a suite of algorithms for determination of sensor health. Regardless of the potential advantages that can be achieved using intelligent sensors, existing large-scale systems are still based on conventional sensors and data acquisition systems. In order to bring the benefits of intelligent sensors to these environments, we have also developed virtual implementations of intelligent sensors.

  18. Elements of social security

    DEFF Research Database (Denmark)

    Hansen, Hans

    (Alte Länder). This is the 9th and last edition of the publication,covering income levels and rules for social security and personal taxation for 1999. Basis for the projections to 1999 income levels is the 1998 data (in some cases 1999 data)for OECD's Taxing Wages as reported by national experts.......Elements of Social Security is a comparative study of important elements of the social security systems in Denmark (DK), Sweden (S), Finland (FIN), Austria (A), Germany (D), the Netherlands (NL), Great Britain (GB) and Canada (CAN). It should be emphasized that Germany is the former West Germany...

  19. Rocket propulsion elements

    CERN Document Server

    Sutton, George P

    2011-01-01

    The definitive text on rocket propulsion-now revised to reflect advancements in the field For sixty years, Sutton's Rocket Propulsion Elements has been regarded as the single most authoritative sourcebook on rocket propulsion technology. As with the previous edition, coauthored with Oscar Biblarz, the Eighth Edition of Rocket Propulsion Elements offers a thorough introduction to basic principles of rocket propulsion for guided missiles, space flight, or satellite flight. It describes the physical mechanisms and designs for various types of rockets' and provides an unders

  20. Finite element analysis

    CERN Document Server

    2010-01-01

    Finite element analysis is an engineering method for the numerical analysis of complex structures. This book provides a bird's eye view on this very broad matter through 27 original and innovative research studies exhibiting various investigation directions. Through its chapters the reader will have access to works related to Biomedical Engineering, Materials Engineering, Process Analysis and Civil Engineering. The text is addressed not only to researchers, but also to professional engineers, engineering lecturers and students seeking to gain a better understanding of where Finite Element Analysis stands today.

  1. Finite elements and approximation

    CERN Document Server

    Zienkiewicz, O C

    2006-01-01

    A powerful tool for the approximate solution of differential equations, the finite element is extensively used in industry and research. This book offers students of engineering and physics a comprehensive view of the principles involved, with numerous illustrative examples and exercises.Starting with continuum boundary value problems and the need for numerical discretization, the text examines finite difference methods, weighted residual methods in the context of continuous trial functions, and piecewise defined trial functions and the finite element method. Additional topics include higher o

  2. Elements of social security

    DEFF Research Database (Denmark)

    Hansen, Hans

    Elements of Social Security is a comparative study of important elements of the social security systems in Denmark (DK), Sweden (S), Finland (FIN), Austria (A), Germany (D), the Netherlands (NL), Great Britain (GB) and Canada (CAN). It should be emphasized that Germany is the former West Germany...... (Alte Länder). This is the 9th and last edition of the publication,covering income levels and rules for social security and personal taxation for 1999. Basis for the projections to 1999 income levels is the 1998 data (in some cases 1999 data)for OECD's Taxing Wages as reported by national experts....

  3. Elements of linear space

    CERN Document Server

    Amir-Moez, A R; Sneddon, I N

    1962-01-01

    Elements of Linear Space is a detailed treatment of the elements of linear spaces, including real spaces with no more than three dimensions and complex n-dimensional spaces. The geometry of conic sections and quadric surfaces is considered, along with algebraic structures, especially vector spaces and transformations. Problems drawn from various branches of geometry are given.Comprised of 12 chapters, this volume begins with an introduction to real Euclidean space, followed by a discussion on linear transformations and matrices. The addition and multiplication of transformations and matrices a

  4. Annihilators of nilpotent elements

    Directory of Open Access Journals (Sweden)

    Abraham A. Klein

    2005-01-01

    Full Text Available Let x be a nilpotent element of an infinite ring R (not necessarily with 1. We prove that A(x—the two-sided annihilator of x—has a large intersection with any infinite ideal I of R in the sense that card(A(x∩I=cardI. In particular, cardA(x=cardR; and this is applied to prove that if N is the set of nilpotent elements of R and R≠N, then card(R\\N≥cardN.

  5. Archaeal extrachromosomal genetic elements

    DEFF Research Database (Denmark)

    Wang, Haina; Peng, Nan; Shah, Shiraz Ali

    2015-01-01

    SUMMARY: Research on archaeal extrachromosomal genetic elements (ECEs) has progressed rapidly in the past decade. To date, over 60 archaeal viruses and 60 plasmids have been isolated. These archaeal viruses exhibit an exceptional diversity in morphology, with a wide array of shapes, such as spind......SUMMARY: Research on archaeal extrachromosomal genetic elements (ECEs) has progressed rapidly in the past decade. To date, over 60 archaeal viruses and 60 plasmids have been isolated. These archaeal viruses exhibit an exceptional diversity in morphology, with a wide array of shapes...

  6. An alkaline element

    Energy Technology Data Exchange (ETDEWEB)

    Obi, F.

    1983-09-29

    A current tap pressed into the anode is installed in the central part of the top of the element. There is an internal top made of plastic under the top. There is a projection in the center of the top with an opening, through which the current tap is passed. The edge of the plastic top serves as an insulation lining between the metallic top and the body, which serves as the current tap for the cathode. A separator is placed between the anode and the cathode. Electrolyte leaks are prevented in the slotted disk elements.

  7. Glucocorticoids regulate the expression of the mouse urocortin II gene: a putative connection between the corticotropin-releasing factor receptor pathways.

    Science.gov (United States)

    Chen, Alon; Vaughan, Joan; Vale, Wylie W

    2003-08-01

    Peptides encoded by the urocortin II (Ucn II) gene were recently identified as new members of the corticotropin-releasing factor (CRF) family. Ucn II is a specific ligand for the type 2 CRF receptor. Using RT-PCR, DNA sequencing, and immunofluorescence staining, we report the expression of Ucn II mRNA in several human and mouse (m) neuronal cell lines. Using these neuronal cell lines, we provide evidence that exposure to glucocorticoid hormones increases mUcn II mRNA expression and promoter activation. The effect of glucocorticoids on mUcn II mRNA expression was tested in the Ucn II/glucocorticoid receptor-positive cell line NG108-15. The results demonstrate that mUcn II mRNA expression is up-regulated by dexamethasone in a dose- and time-dependent fashion. Computer analysis revealed the presence of 14 putative half-palindrome glucocorticoid response element sequences within 1.2 kb of the mUcn II 5' flanking region. Transfections with different fragments of the 5'-flanking region of the mUcn II gene fused to a luciferase reporter gene showed a promoter-dependent expression of the reporter gene and regulation by dexamethasone. Promoter deletion studies clarify the sufficient putative glucocorticoid response element site mediating this effect. The steroid hormone antagonist RU486 blocked the effect of dexamethasone on mUcn II mRNA expression and promoter activation, suggesting a direct glucocorticoid receptor-mediated effect of dexamethasone on mUcn II mRNA expression. Ucn II is expressed in vivo in the hypothalamus, brainstem, olfactory bulb, and pituitary. Low levels were also detected in the mouse cortex, hippocampus, and spinal cord. We demonstrated that mUcn II gene transcription was stimulated by glucocorticoid administration in vivo and inhibited by removal of glucocorticoids by adrenalectomy. Administration of dexamethasone to mice resulted in an increase of mUcn II levels in the hypothalamus and brainstem but not in the olfactory bulb region 12 h following

  8. Localization of putative carbonic anhydrases in two marine diatoms, Phaeodactylum tricornutum and Thalassiosira pseudonana.

    Science.gov (United States)

    Tachibana, Masaaki; Allen, Andrew E; Kikutani, Sae; Endo, Yuri; Bowler, Chris; Matsuda, Yusuke

    2011-09-01

    It is believed that intracellular carbonic anhydrases (CAs) are essential components of carbon concentrating mechanisms in microalgae. In this study, putative CA-encoding genes were identified in the genome sequences of the marine diatoms Phaeodactylum tricornutum and Thalassiosira pseudonana. Subsequently, the subcellular localizations of the encoded proteins were determined. Nine and thirteen CA sequences were found in the genomes of P. tricornutum and T. pseudonana, respectively. Two of the β-CA genes in P. tricornutum corresponded to ptca1 and ptca2 identified previously. Immunostaining transmission electron microscopy of a PtCA1:YFP fusion expressed in the cells of P. tricornutum clearly showed the localization of PtCA1 within the central part of the pyrenoid structure in the chloroplast. Besides these two β-CA genes, P. tricornutum likely contains five α- and two γ-CA genes, whereas T. pseudonana has three α-, five γ-, four δ-, and one ζ-CA genes. Semi-quantitative reverse transcription PCR performed on mRNA from the two diatoms grown in changing light and CO(2) conditions revealed that levels of six putative α- and γ-CA mRNAs in P. tricornutum did not change between cells grown in air-level CO(2) and 5% CO(2). However, mRNA levels of one putative α-CA gene, CA-VII in P. tricornutum, were reduced in the dark compared to that in the light. In T. pseudonana, mRNA accumulation levels of putative α-CA (CA-1), ζ-CA (CA-3) and δ-CA (CA-7) were analyzed and all levels found to be significantly reduced when cells were grown in 0.16% CO(2). Intercellular localizations of eight putative CAs were analyzed by expressing GFP fusion in P. tricornutum and T. pseudonana. In P. tricornutum, CA-I and II localized in the periplastidial compartment, CA-III, VI, VII were found in the chloroplast endoplasmic reticulum, and CA-VIII was localized in the mitochondria. On the other hand, T. pseudonana CA-1 localized in the stroma and CA-3 was found in the periplasm

  9. Genetic organisation, mobility and predicted functions of genes on integrated, mobile genetic elements in sequenced strains of Clostridium difficile.

    Directory of Open Access Journals (Sweden)

    Michael S M Brouwer

    Full Text Available BACKGROUND: Clostridium difficile is the leading cause of hospital-associated diarrhoea in the US and Europe. Recently the incidence of C. difficile-associated disease has risen dramatically and concomitantly with the emergence of 'hypervirulent' strains associated with more severe disease and increased mortality. C. difficile contains numerous mobile genetic elements, resulting in the potential for a highly plastic genome. In the first sequenced strain, 630, there is one proven conjugative transposon (CTn, Tn5397, and six putative CTns (CTn1, CTn2 and CTn4-7, of which, CTn4 and CTn5 were capable of excision. In the second sequenced strain, R20291, two further CTns were described. RESULTS: CTn1, CTn2 CTn4, CTn5 and CTn7 were shown to excise from the genome of strain 630 and transfer to strain CD37. A putative CTn from R20291, misleadingly termed a phage island previously, was shown to excise and to contain three putative mobilisable transposons, one of which was capable of excision. In silico probing of C. difficile genome sequences with recombinase gene fragments identified new putative conjugative and mobilisable transposons related to the elements in strains 630 and R20291. CTn5-like elements were described occupying different insertion sites in different strains, CTn1-like elements that have lost the ability to excise in some ribotype 027 strains were described and one strain was shown to contain CTn5-like and CTn7-like elements arranged in tandem. Additionally, using bioinformatics, we updated previous gene annotations and predicted novel functions for the accessory gene products on these new elements. CONCLUSIONS: The genomes of the C. difficile strains examined contain highly related CTns suggesting recent horizontal gene transfer. Several elements were capable of excision and conjugative transfer. The presence of antibiotic resistance genes and genes predicted to promote adaptation to the intestinal environment suggests that CTns play a

  10. Overexpression of Brucella putative glycosyltransferase WbkA in B. abortus RB51 leads to production of exopolysaccharide

    Directory of Open Access Journals (Sweden)

    Neha eDabral

    2015-06-01

    Full Text Available Brucella spp. are Gram-negative, facultative intracellular bacteria that cause brucellosis in mammals. Brucella strains containing the O-polysaccharide in their cell wall structure exhibit a smooth phenotype whereas the strains devoid of the polysaccharide show rough phenotype. B. abortus strain RB51 is a stable rough attenuated mutant which is used as a licensed live vaccine for bovine brucellosis. Previous studies have shown that the wboA gene, which encodes a glycosyltransferase required for the synthesis of O-polysaccharide, is disrupted in B. abortus RB51 by an IS711 element. Although complementation of strain RB51 with a functional wboA gene results in O-polysaccharide synthesis in the cytoplasm, it does not result in smooth phenotype. The aim of this study was to determine if overexpression of Brucella WbkA or WbkE, two additional putative glycosyltransferases essential for O-polysaccharide synthesis, in strain RB51 would result in the O-polysaccharide synthesis and smooth phenotype. Our results demonstrate that overexpression of wbkA or wbkE gene in RB51 does not result in O-polysaccharide expression as shown by Western blotting with specific antibodies. However, wbkA, but not wbkE, overexpression leads to the development of a clumping phenotype and the production of exopolysaccharide(s containing mannose, galactose, N-acetylglucosamine and N-acetylgalactosamine. Moreover, we found that the clumping recombinant strain displays increased adhesion to polystyrene plates. The recombinant strain was similar to strain RB51 in its attenuation characteristic and in its ability to induce protective immunity against virulent B. abortus challenge in mice.

  11. Overexpression of Brucella putative glycosyltransferase WbkA in B. abortus RB51 leads to production of exopolysaccharide.

    Science.gov (United States)

    Dabral, Neha; Jain-Gupta, Neeta; Seleem, Mohamed N; Sriranganathan, Nammalwar; Vemulapalli, Ramesh

    2015-01-01

    Brucella spp. are Gram-negative, facultative intracellular bacteria that cause brucellosis in mammals. Brucella strains containing the O-polysaccharide in their cell wall structure exhibit a smooth phenotype whereas the strains devoid of the polysaccharide show rough phenotype. B. abortus strain RB51 is a stable rough attenuated mutant which is used as a licensed live vaccine for bovine brucellosis. Previous studies have shown that the wboA gene, which encodes a glycosyltransferase required for the synthesis of O-polysaccharide, is disrupted in B. abortus RB51 by an IS711 element. Although complementation of strain RB51 with a functional wboA gene results in O-polysaccharide synthesis in the cytoplasm, it does not result in smooth phenotype. The aim of this study was to determine if overexpression of Brucella WbkA or WbkE, two additional putative glycosyltransferases essential for O-polysaccharide synthesis, in strain RB51 would result in the O-polysaccharide synthesis and smooth phenotype. Our results demonstrate that overexpression of wbkA or wbkE gene in RB51 does not result in O-polysaccharide expression as shown by Western blotting with specific antibodies. However, wbkA, but not wbkE, overexpression leads to the development of a clumping phenotype and the production of exopolysaccharide(s) containing mannose, galactose, N-acetylglucosamine, and N-acetylgalactosamine. Moreover, we found that the clumping recombinant strain displays increased adhesion to polystyrene plates. The recombinant strain was similar to strain RB51 in its attenuation characteristic and in its ability to induce protective immunity against virulent B. abortus challenge in mice.

  12. Beam transport elements

    CERN Multimedia

    1965-01-01

    Two of the beam transport elements for the slow ejection system. On the left, a quadrupole 1.2 m long with a 5 cm aperture, capable of producing a gradient of 5000 gauss. On the right, a 1 m bending magnet with a 4 cm gap; its field is 20 000 gauss.

  13. Movies and Literary Elements.

    Science.gov (United States)

    Keller, Rodney D.

    1987-01-01

    Recommends using movie clips to stimulate students' interest in reading a novel as well as to teach elements of fiction such as plot, character, setting, symbol, irony, and theme. Describes each clip and provides study questions. Includes a listing of movies made from books. (NH)

  14. Elements of Social Security

    DEFF Research Database (Denmark)

    Hansen, Hans

    Elements of Social Security contains an overview of important benefit schemes in Denmark, Sweden, Finland, Germany, the Netherlands, Great Britain and Canada. The schemes are categorized according to common sets of criteria and compared. Stylized cases illustrate the impact on disposable income...

  15. Light element ceramics

    OpenAIRE

    Rao, KJ; Varma, KBR; Raju, AR

    1988-01-01

    An overview of a few structually important light element ceramics is presented. Included in the overview are silicon nitide, sialon, aluminium nitride, boron carbide and silicon carbide. Methods of preparation, characterization and industrial applications of these ceramics are summarized. Mechanical properties, industrial production techniques and principal uses of these ceramics are emphasized.

  16. Water, the intangible element

    NARCIS (Netherlands)

    Schotting, R.J.

    2009-01-01

    Water is the key to life. No living creature can survive without water. Too much water or polluted water are serious threats to mankind. Managing this intangible element is complex, not only in wet deltaic regions but also in the (semi-)arid regions of the world. Combined efforts of the hydro(geo)lo

  17. CEDS Addresses: Rubric Elements

    Science.gov (United States)

    US Department of Education, 2015

    2015-01-01

    Common Education Data Standards (CEDS) Version 4 introduced a common data vocabulary for defining rubrics in a data system. The CEDS elements support digital representations of both holistic and analytic rubrics. This document shares examples of holistic and analytic project rubrics, available CEDS Connections, and a logical model showing the…

  18. The elements of life

    Directory of Open Access Journals (Sweden)

    Sergey P. Fedotov

    2016-11-01

    Full Text Available Article "Elements of Life" offers a hypothesis about the relationship of the phenomenon of traditional Chinese medicine with the physical laws. It shows the principle of forming a sequence of daily activity of the acupuncture meridians as a consequence of the Doppler effect in the process of flowing around the planet Earth by cosmic wind (by Ether. In accordance with this specification the daily structure of meridians had been built. It is suggested that the essence of the Chinese Qi (Chi are vibrations of a certain range in the medium. Consequently, it became possible to set the interrelation of frequencies of the visible spectrum with certain meridians. It is shown that the topological relationship of ancient (barrier points of the Five Elements (Wu-Shu points are associated with the wave lengths of the so-called Qi. It is shown also that the essence of the Wu-Xing law is based on daily circulation patterns of meridians. The examples of the surrounding world, including pulses processes in the human body, are confirming the above mentioned theses. A correlation diagram between the main elements by Dr. Samohotsky A.S. (dissertation "The experience of the definition of medical laws", 1946 and the Five Elements of traditional Chinese philosophy is established. The above represented hypotheses are yet introduced in practice in form of pulse spectral analysis system.

  19. Elements of Social Security

    DEFF Research Database (Denmark)

    Hansen, Hans

    Elements of Social Security contains an overview of important benefit schemes in Denmark, Sweden, Finland, Germany, Great Britain, the Netherlands and Canada. The schemes are categorized according to common sets of criteria and compared. Stylized cases illustrate the impact on disposable income...

  20. Elements of Social Security

    DEFF Research Database (Denmark)

    Hansen, Hans

    Elements of Social Security contains an overview of important benefit schemes in Denmark, Sweden, Finland, Austria, Germany, the Netherlands, Great Britain and Canada. The schemes are categorized according to common sets of criteria and compared. Stylized cases illustrate the impact on disposable...

  1. Elemental Chem Lab

    Science.gov (United States)

    Franco Mariscal, Antonio Joaquin

    2008-01-01

    This educative material uses the symbols of 45 elements to spell the names of 32 types of laboratory equipment usually found in chemical labs. This teaching material has been divided into three puzzles according to the type of the laboratory equipment: (i) glassware as reaction vessels or containers; (ii) glassware for measuring, addition or…

  2. Reference: -300ELEMENT [PLACE

    Lifescience Database Archive (English)

    Full Text Available -300ELEMENT Kreis M, Williamson MS, Forde J, Schmitz D, Clark J, Buxton B, Pywell J..., Marris C, Henderson J, Harris N, Shewry PR, Forde BG, Miflin BJ Differential gene expression in the developing barley endosperm. Philos Trans R Soc Lond B314:355-365 (1986) ...

  3. Inside finite elements

    CERN Document Server

    Weiser, Martin

    2016-01-01

    All relevant implementation aspects of finite element methods are discussed in this book. The focus is on algorithms and data structures as well as on their concrete implementation. Theory is covered as far as it gives insight into the construction of algorithms. Throughout the exercises a complete FE-solver for scalar 2D problems will be implemented in Matlab/Octave.

  4. Element-topology-independent preconditioners for parallel finite element computations

    Science.gov (United States)

    Park, K. C.; Alexander, Scott

    1992-01-01

    A family of preconditioners for the solution of finite element equations are presented, which are element-topology independent and thus can be applicable to element order-free parallel computations. A key feature of the present preconditioners is the repeated use of element connectivity matrices and their left and right inverses. The properties and performance of the present preconditioners are demonstrated via beam and two-dimensional finite element matrices for implicit time integration computations.

  5. Element-topology-independent preconditioners for parallel finite element computations

    Science.gov (United States)

    Park, K. C.; Alexander, Scott

    1992-01-01

    A family of preconditioners for the solution of finite element equations are presented, which are element-topology independent and thus can be applicable to element order-free parallel computations. A key feature of the present preconditioners is the repeated use of element connectivity matrices and their left and right inverses. The properties and performance of the present preconditioners are demonstrated via beam and two-dimensional finite element matrices for implicit time integration computations.

  6. Conjugative transfer and cis-mobilization of a genomic island by an integrative and conjugative element of Streptococcus agalactiae.

    Science.gov (United States)

    Puymège, Aurore; Bertin, Stéphane; Chuzeville, Sarah; Guédon, Gérard; Payot, Sophie

    2013-03-01

    Putative integrative and conjugative elements (ICEs), i.e., genomic islands which could excise, self-transfer by conjugation, and integrate into the chromosome of the bacterial host strain, were previously identified by in silico analysis in the sequenced genomes of Streptococcus agalactiae (M. Brochet et al., J. Bacteriol. 190:6913-6917, 2008). We investigated here the mobility of the elements integrated into the 3' end of a tRNA(Lys) gene. Three of the four putative ICEs tested were found to excise but only one (ICE_515_tRNA(Lys)) was found to transfer by conjugation not only to S. agalactiae strains but also to a Streptococcus pyogenes strain. Transfer was observed even if recipient cell already carries a related resident ICE or a genomic island flanked by attL and attR recombination sites but devoid of conjugation or recombination genes (CIs-Mobilizable Element [CIME]). The incoming ICE preferentially integrates into the 3' end of the tRNA(Lys) gene (i.e., the attR site of the resident element), leading to a CIME-ICE structure. Transfer of the whole composite element CIME-ICE was obtained, showing that the CIME is mobilizable in cis by the ICE. Therefore, genomic islands carrying putative virulence genes but lacking the mobility gene can be mobilized by a related ICE after site-specific accretion.

  7. A Hypothesis for the Abiotic and Non-Martian Origins of Putative Signs of Ancient Martian Life in ALH84001

    Science.gov (United States)

    Treiman, Allan H.

    2001-01-01

    Putative evidence of martian life in ALH84001 can be explained by abiotic and non-martian processes consistent with the meteorite's geological history. Additional information is contained in the original extended abstract.

  8. Haemolytic and cytotoxic activities of the Tween 80-extracted putative haemolysin of Pasteurella multocida B:2.

    Science.gov (United States)

    Chachra, D; Coote, J G; Parton, R; Jand, S K

    2011-06-02

    The objective of this study was to investigate the haemolytic and cytotoxic activity of Pasteurella multocida B:2 strains, originally from cases of haemorrhagic septicaemia in cattle. All six P. multocida B:2 strains were non-haemolytic on sheep blood agar (SBA) and horse blood agar (HBA) when grown aerobically and on SBA anaerobically but they were haemolytic on HBA when grown anaerobically. No haemolytic activity against horse red blood cells was detected in culture supernates from aerobically or anaerobically grown cultures and only very weak haemolytic activity was obtained in supernates or pellet fractions from sonicated cells. However, after repeated extraction of sonicated cells with Tween 80, haemolytic activity was found in various cell fractions, both Tween-soluble and -insoluble. The Tween-extracted putative haemolysin and other bacterial fractions were also cytotoxic for mouse macrophage-like J774.2 cells. Further characterisation of the putative haemolysin revealed it to be a heat-labile, non-pore-forming protein of molecular weight >10 kDa whose activity was completely destroyed by trypsin and greatly reduced with protease and proteinase K treatment. Congo red also reduced the haemolytic activity. Non-denaturing gel-electrophoresis and RBC agar overlay revealed clear haemolytic zones but suggested that Tween was bound to some component of the P. multocida B:2 fractions and was responsible, to some extent, for the haemolytic activity observed. However, the effect of heat and other reagents on the Tween-extracted fractions and the lack of haemolytic activity in different Tween-extracted cell fractions of organisms other than P. multocida suggested that some proteinaceous component of the organism could indeed act as a haemolysin. This putative haemolysin may be one of the virulence attributes of P. multocida, but its characterisation and role in pathogenesis require further study.

  9. Isolation and characterization of two mitoviruses and a putative alphapartitivirus from Fusarium spp.

    Science.gov (United States)

    Osaki, Hideki; Sasaki, Atsuko; Nomiyama, Koji; Sekiguchi, Hiroyuki; Tomioka, Keisuke; Takehara, Toshiaki

    2015-06-01

    The filamentous fungus Fusarium spp. includes several important plant pathogens. We attempted to reveal presence of double-stranded (ds) RNAs in the genus. Thirty-seven Fusarium spp. at the MAFF collection were analyzed. In the strains of Fusarium coeruleum, Fusarium globosum and Fusarium solani f. sp. pisi, single dsRNA bands were detected. The strains of F. coeruleum and F. solani f. sp. pisi cause potato dry rot and mulberry twig blight, respectively. Sequence analyses revealed that dsRNAs in F. coeruleum and F. globosum consisted of 2423 and 2414 bp, respectively. Using the fungal mitochondrial translation table, the positive strands of these cDNAs were found to contain single open reading frames with the potential to encode a protein of putative 757 and 717 amino acids (molecular mass 88.5 and 84.0 kDa, respectively), similar to RNA-dependent RNA polymerases of members of the genus Mitovirus. These dsRNAs in F. coeruleum and F. globosum were assigned to the genus Mitovirus (family Narnaviridae), and these two mitoviruses were designated as Fusarium coeruleum mitovirus 1 and Fusarium globosum mitovirus 1. On the other hand, a positive strand of cDNA (1950 bp) from dsRNA in F. solani f. sp. pisi contained an ORF potentially encoding a putative RdRp of 608 amino acids (72.0 kDa). The putative RdRp was shown to be related to those of members of the genus of Alphapartitivirus (family Partitiviridae). We coined the name Fusarium solani partitivirus 2 for dsRNA in F. solani f. sp. pisi.

  10. The intracellular mobility of NPY and a putative mitochondrial form of NPY in neuronal cells.

    Science.gov (United States)

    Kaipio, Katja; Pesonen, Ullamari

    2009-01-30

    Preproneuropeptide Y is a precursor peptide to mature neuropeptide Y (NPY), which is a universally expressed peptide in the central and peripheral nervous system. NPY is normally routed to endoplasmic reticulum and secretory vesicles in cells, which secrete NPY. In our previous studies, we found a functional Leucine7 to Proline7 (L7P) polymorphism in the signal peptide sequence of preproNPY. This polymorphism affects the secretion of NPY and causes multiple physiological effects in humans. The sequence of NPY mRNA contains two in frame kozak sequences that allow translation initiation to shift, and translation of two proteins. In addition to mature NPY(1-36) also a putative truncated NPY(17-36) with mitochondrial targeting signal is produced. The purpose of this study was to investigate the protein mobility of the putative mitochondrial fragment and the effect of the L7P polymorphism on the cellular level using GFP tagged constructs. The mobility was studied with fluorescence recovery after photobleaching technique in a neuronal cell line. We found that the mobility of the secretory vesicles with NPY(1-36) in cells with L7P genotype was increased in comparison to vesicle mobility in cells with the more abundant L7L genotype. The mobility in the cells with the putative mitochondrial construct was found to be very low. According to the results of the present study, the mitochondrial truncated peptide stays in the mitochondrion. It can be hypothesized that this could be one of the factors affecting energy balance of the membranes of the mitochondrion.

  11. Identification and characterization of a gene encoding a putative lysophosphatidyl acyltransferase from Arachis hypogaea

    Indian Academy of Sciences (India)

    Si-Long Chen; Jia-Quan Huang; Lei Yong; Yue-Ting Zhang; Xiao-Ping Ren; Yu-Ning Chen; Hui-Fang Jiang; Li-Ying Yan; Yu-Rong Li; Bo-Shou Liao

    2012-12-01

    Lysophosphatidyl acyltransferase (LPAT) is the important enzyme responsible for the acylation of lysophosphatidic acid (LPA), leading to the generation of phosphatidic acid (PA) in plant. Its encoding gene is an essential candidate for oil crops to improve oil composition and increase seed oil content through genetic engineering. In this study, a full-length AhLPAT4 gene was isolated via cDNA library screening and rapid amplification of cDNA ends (RACE); our data demonstrated that AhLPAT4 had 1631 nucleotides, encoding a putative 43.8 kDa protein with 383 amino acid residues. The deduced protein included a conserved acyltransferase domain and four motifs (I–IV) with putative LPA and acyl-CoA catalytic and binding sites. Bioinformatic analysis indicated that AhLPAT4 contained four transmembrane domains (TMDs), localized to the endoplasmic reticulum (ER) membrane; detailed analysis indicated that motif I and motifs II–III in AhLPAT4 were separated by the third TMD, which located on cytosolic and ER luminal side respectively, and hydrophobic residues on the surface of AhLPAT4 protein fold to form a hydrophobic tunnel to accommodate the acyl chain. Subcellular localization analysis confirmed that AhLPAT4 was a cytoplasm protein. Phylogenetic analysis revealed that AhLPAT4 had a high homology (63.7–78.3%) with putative LPAT4 proteins from Glycine max, Arabidopsis thaliana and Ricinus communis. AhLPAT4 was ubiquitously expressed in diverse tissues except in flower, which is almost undetectable. The expression analysis in different developmental stages in peanut seeds indicated that AhLPAT4 did not coincide with oil accumulation.

  12. Identification of putative rhamnogalacturonan-II specific glycosyltransferases in Arabidopsis using a combination of bioinformatics approaches.

    Science.gov (United States)

    Voxeur, Aline; André, Aurélie; Breton, Christelle; Lerouge, Patrice

    2012-01-01

    Rhamnogalacturonan-II (RG-II) is a complex plant cell wall polysaccharide that is composed of an α(1,4)-linked homogalacturonan backbone substituted with four side chains. It exists in the cell wall in the form of a dimer that is cross-linked by a borate di-ester. Despite its highly complex structure, RG-II is evolutionarily conserved in the plant kingdom suggesting that this polymer has fundamental functions in the primary wall organisation. In this study, we have set up a bioinformatics strategy aimed at identifying putative glycosyltransferases (GTs) involved in RG-II biosynthesis. This strategy is based on the selection of candidate genes encoding type II membrane proteins that are tightly coexpressed in both rice and Arabidopsis with previously characterised genes encoding enzymes involved in the synthesis of RG-II and exhibiting an up-regulation upon isoxaben treatment. This study results in the final selection of 26 putative Arabidopsis GTs, including 10 sequences already classified in the CAZy database. Among these CAZy sequences, the screening protocol allowed the selection of α-galacturonosyltransferases involved in the synthesis of α4-GalA oligogalacturonides present in both homogalacturonans and RG-II, and two sialyltransferase-like sequences previously proposed to be involved in the transfer of Kdo and/or Dha on the pectic backbone of RG-II. In addition, 16 non-CAZy GT sequences were retrieved in the present study. Four of them exhibited a GT-A fold. The remaining sequences harbored a GT-B like fold and a fucosyltransferase signature. Based on homologies with glycosyltransferases of known functions, putative roles in the RG-II biosynthesis are proposed for some GT candidates.

  13. A Proteomics Approach to Identify New Putative Cardiac Intercalated Disk Proteins.

    Directory of Open Access Journals (Sweden)

    Siddarth Soni

    Full Text Available Synchronous beating of the heart is dependent on the efficient functioning of the cardiac intercalated disk (ID. The ID is composed of a complex protein network enabling electrical continuity and chemical communication between individual cardiomyocytes. Recently, several different studies have shed light on increasingly prevalent cardiac diseases involving the ID. Insufficient knowledge of its composition makes it difficult to study these disease mechanisms in more detail and therefore here we aim expand the ID proteome. Here, using a combination of general membrane enrichment, in-depth quantitative proteomics and an intracellular location driven bioinformatics approach, we aim to discover new putative ID proteins in rat ventricular tissue.General membrane isolation, enriched amongst others also with ID proteins as based on presence of the established markers connexin-43 and n-cadherin, was performed using centrifugation. By mass spectrometry, we quantitatively evaluated the level of 3455 proteins in the enriched membrane fraction (EMF and its counterpart, the soluble cytoplasmic fraction. These data were stringently filtered to generate a final set of 97 enriched, putative ID proteins. These included Cx43 and n-cadherin, but also many interesting novel candidates. We selected 4 candidates (Flotillin-2 (FLOT2, Nexilin (NEXN, Popeye-domain-containg-protein 2 (POPDC2 and thioredoxin-related-transmembrane-protein 2 (TMX2 and confirmed their co-localization with n-cadherin in the ID of human and rat heart cryo-sections, and isolated dog cardiomyocytes.The presented proteomics dataset of putative new ID proteins is a valuable resource for future research into this important molecular intersection of the heart.

  14. Two putative-aquaporin genes are differentially expressed during arbuscular mycorrhizal symbiosis in Lotus japonicus

    Science.gov (United States)

    2012-01-01

    Background Arbuscular mycorrhizas (AM) are widespread symbioses that provide great advantages to the plant, improving its nutritional status and allowing the fungus to complete its life cycle. Nevertheless, molecular mechanisms that lead to the development of AM symbiosis are not yet fully deciphered. Here, we have focused on two putative aquaporin genes, LjNIP1 and LjXIP1, which resulted to be upregulated in a transcriptomic analysis performed on mycorrhizal roots of Lotus japonicus. Results A phylogenetic analysis has shown that the two putative aquaporins belong to different functional families: NIPs and XIPs. Transcriptomic experiments have shown the independence of their expression from their nutritional status but also a close correlation with mycorrhizal and rhizobial interaction. Further transcript quantification has revealed a good correlation between the expression of one of them, LjNIP1, and LjPT4, the phosphate transporter which is considered a marker gene for mycorrhizal functionality. By using laser microdissection, we have demonstrated that one of the two genes, LjNIP1, is expressed exclusively in arbuscule-containing cells. LjNIP1, in agreement with its putative role as an aquaporin, is capable of transferring water when expressed in yeast protoplasts. Confocal analysis have demonstrated that eGFP-LjNIP1, under its endogenous promoter, accumulates in the inner membrane system of arbusculated cells. Conclusions Overall, the results have shown different functionality and expression specificity of two mycorrhiza-inducible aquaporins in L. japonicus. One of them, LjNIP1 can be considered a novel molecular marker of mycorrhizal status at different developmental stages of the arbuscule. At the same time, LjXIP1 results to be the first XIP family aquaporin to be transcriptionally regulated during symbiosis. PMID:23046713

  15. Enrichment of putative human epidermal stem cells based on cell size and collagen type IV adhesiveness

    Institute of Scientific and Technical Information of China (English)

    Juxue Li; Enkui Duan; Chenglin Miao; Weixiang Guo; Liwei Jia; Jiaxi Zhou; Baohua Ma; Sha Peng; Shuang Liu; Yujing Cao

    2008-01-01

    The enrichment and identification of human epidermal stem cells (EpSCs) are of paramount importance for both basic research and clinical application. Although several approaches for the enrichment of EpSCs have been established, enriching a pure population of viable EpSCs is still a challenging task. An improved approach is worth developing to enhance the purity and viability of EpSCs. Here we report that cell size combined with collagen type IV adhesiveness can be used in an improved approach to enrich pure and viable human EpSCs. We separated the rapidly adherent keratinocytes into three populations that range in size from 5-7 μm (population A), to 7-9 μm (population B), to >9 μm (population C) in diameter, and found that human putative EpSCs could be further enriched in population A with the smallest size. Among the three populations, population A displayed the highest density of βl-integrin receptor, contained the highest percentage of cells in G0/G1 phase, showed the highest nucleus to cytoplasm ratio, and possessed the highest colony formation efficiency (CFE). When injected into murine blastocysts, these cells participated in multi-tissue formation. More significantly, compared with a previous approach that sorted putative EpSCs according to pl-integrin antibody staining, the viability of the EpSCs enriched by the improved approach was significantly enhanced. Our results provide a putative strategy for the enrichment of human EpSCs, and encourage further study into the role of cell size in stem cell biology.

  16. Two putative-aquaporin genes are differentially expressed during arbuscular mycorrhizal symbiosis in Lotus japonicus

    Directory of Open Access Journals (Sweden)

    Giovannetti Marco

    2012-10-01

    Full Text Available Abstract Background Arbuscular mycorrhizas (AM are widespread symbioses that provide great advantages to the plant, improving its nutritional status and allowing the fungus to complete its life cycle. Nevertheless, molecular mechanisms that lead to the development of AM symbiosis are not yet fully deciphered. Here, we have focused on two putative aquaporin genes, LjNIP1 and LjXIP1, which resulted to be upregulated in a transcriptomic analysis performed on mycorrhizal roots of Lotus japonicus. Results A phylogenetic analysis has shown that the two putative aquaporins belong to different functional families: NIPs and XIPs. Transcriptomic experiments have shown the independence of their expression from their nutritional status but also a close correlation with mycorrhizal and rhizobial interaction. Further transcript quantification has revealed a good correlation between the expression of one of them, LjNIP1, and LjPT4, the phosphate transporter which is considered a marker gene for mycorrhizal functionality. By using laser microdissection, we have demonstrated that one of the two genes, LjNIP1, is expressed exclusively in arbuscule-containing cells. LjNIP1, in agreement with its putative role as an aquaporin, is capable of transferring water when expressed in yeast protoplasts. Confocal analysis have demonstrated that eGFP-LjNIP1, under its endogenous promoter, accumulates in the inner membrane system of arbusculated cells. Conclusions Overall, the results have shown different functionality and expression specificity of two mycorrhiza-inducible aquaporins in L. japonicus. One of them, LjNIP1 can be considered a novel molecular marker of mycorrhizal status at different developmental stages of the arbuscule. At the same time, LjXIP1 results to be the first XIP family aquaporin to be transcriptionally regulated during symbiosis.

  17. Temporal dynamics and decay of putatively allochthonous and autochthonous viral genotypes in contrasting freshwater lakes.

    Science.gov (United States)

    Hewson, Ian; Barbosa, Jorge G; Brown, Julia M; Donelan, Ryan P; Eaglesham, James B; Eggleston, Erin M; LaBarre, Brenna A

    2012-09-01

    Aquatic viruses play important roles in the biogeochemistry and ecology of lacustrine ecosystems; however, their composition, dynamics, and interactions with viruses of terrestrial origin are less extensively studied. We used a viral shotgun metagenomic approach to elucidate candidate autochthonous (i.e., produced within the lake) and allochthonous (i.e., washed in from other habitats) viral genotypes for a comparative study of their dynamics in lake waters. Based on shotgun metagenomes prepared from catchment soil and freshwater samples from two contrasting lakes (Cayuga Lake and Fayetteville Green Lake), we selected two putatively autochthonous viral genotypes (phycodnaviruses likely infecting algae and cyanomyoviruses likely infecting picocyanobacteria) and two putatively allochthonous viral genotypes (geminiviruses likely infecting terrestrial plants and circoviruses infecting unknown hosts but common in soil libraries) for analysis by genotype-specific quantitative PCR (TaqMan) applied to DNAs from viruses in the viral size fraction of lake plankton, i.e., 0.2 μm > virus > 0.02 μm. The abundance of autochthonous genotypes largely reflected expected host abundance, while the abundance of allochthonous genotypes corresponded with rainfall and storm events in the respective catchments, suggesting that viruses with these genotypes may have been transported to the lake in runoff. The decay rates of allochthonous and autochthonous genotypes, assessed in incubations where all potential hosts were killed, were generally lower (0.13 to 1.50% h(-1)) than those reported for marine virioplankton but similar to those for freshwater virioplankton. Both allochthonous and autochthonous viral genotypes were detected at higher concentrations in subsurface sediments than at the water-sediment interface. Our data indicate that putatively allochthonous viruses are present in lake plankton and sediments, where their temporal dynamics reflect active transport to the lake during

  18. Further insight into reproductive incompatibility between putative cryptic species of the Bemisia tabaci whitefly complex.

    Science.gov (United States)

    Qin, Li; Pan, Li-Long; Liu, Shu-Sheng

    2016-04-01

    The whitefly Bemisia tabaci (Gennadius) (Hemiptera: Aleyrodidae), with its global distribution and extensive genetic diversity, is now known to be a complex of over 35 cryptic species. However, a satisfactory resolution of the systematics of this species complex is yet to be achieved. Here, we designed experiments to examine reproductive compatibility among species with different levels of mitochondrial cytochrome oxidase I (mtCOI) divergence. The data show that putative species with mtCOI divergence of >8% between them consistently exhibited complete reproductive isolation. However, two of the putative species, Asia II 9 and Asia II 3, with mtCOI divergence of 4.47% between them, exhibited near complete reproductive compatibility in one direction of their cross, and partial reproductive compatibility in the other direction. Together with some recent reports on this topic from the literature, our data indicates that, while divergence in the mtCOI sequences provides a valid molecular marker for species delimitation in most clades, more genetic markers and more sophisticated molecular phylogeny will be required to achieve adequate delimitation of all species in this whitefly complex. While many attempts have been made to examine the reproductive compatibility among genetic groups of the B. tabaci complex, our study represents the first effort to conduct crossing experiments with putative species that were chosen with considerations of their genetic divergence. In light of the new data, we discuss the best strategy and protocols to conduct further molecular phylogenetic analysis and crossing trials, in order to reveal the overall pattern of reproductive incompatibility among species of this whitefly complex.

  19. Elements of quantum optics

    CERN Document Server

    Meystre, Pierre

    2007-01-01

    Elements of Quantum Optics gives a self-contained and broad coverage of the basic elements necessary to understand and carry out research in laser physics and quantum optics, including a review of basic quantum mechanics and pedagogical introductions to system-reservoir interactions and to second quantization. The text reveals the close connection between many seemingly unrelated topics, such as probe absorption, four-wave mixing, optical instabilities, resonance fluorescence and squeezing. It also comprises discussions of cavity quantum electrodynamics and atom optics. The 4th edition includes a new chapter on quantum entanglement and quantum information, as well as added discussions of the quantum beam splitter, electromagnetically induced transparency, slow light, and the input-output formalism needed to understand many problems in quantum optics. It also provides an expanded treatment of the minimum-coupling Hamiltonian and a simple derivation of the Gross-Pitaevskii equation, an important gateway to rese...

  20. Elements of String Cosmology

    Science.gov (United States)

    Gasperini, Maurizio

    2011-03-01

    Preface; Acknowledgements; Notation, units and conventions; 1. A short review of standard and inflationary cosmology; 2. The basic string cosmology equations; 3. Conformal invariance and string effective action; 4. Duality symmetries and cosmological solutions; 5. Inflationary kinematics; 6. The string phase; 7. The cosmic background of relic gravitational waves; 8. Scalar perturbations and the anisotropy of the CMB radiation; 9. Dilaton phenomenology; 10. Elements of brane cosmology; Index.

  1. Quantum theory elements

    CERN Document Server

    Bates, David Robert

    1962-01-01

    Quantum Theory: A Treatise in Three Volumes, I: Elements focuses on the principles, methodologies, and approaches involved in quantum theory, including quantum mechanics, linear combinations, collisions, and transitions. The selection first elaborates on the fundamental principles of quantum mechanics, exactly soluble bound state problems, and continuum. Discussions focus on delta function normalization, spherically symmetric potentials, rectangular potential wells, harmonic oscillators, spherically symmetrical potentials, Coulomb potential, axiomatic basis, consequences of first three postula

  2. Nuclear fuel element

    Science.gov (United States)

    Meadowcroft, Ronald Ross; Bain, Alastair Stewart

    1977-01-01

    A nuclear fuel element wherein a tubular cladding of zirconium or a zirconium alloy has a fission gas plenum chamber which is held against collapse by the loops of a spacer in the form of a tube which has been deformed inwardly at three equally spaced, circumferential positions to provide three loops. A heat resistant disc of, say, graphite separates nuclear fuel pellets within the cladding from the plenum chamber. The spacer is of zirconium or a zirconium alloy.

  3. A sealing element

    Energy Technology Data Exchange (ETDEWEB)

    Obi, F.; Takada, K.

    1983-09-22

    A sealing lining made of a mixture of polyethylene or polypropylene with an additive which prevents destruction of the lining is used in the element. The content of the additive in the mixture is 10 to 30 percent by mass. The additive basically consists of polyethylene. Carboxyl groups are introduced into the polymer as an anion and sodium, potassium and molybdenum are introduced as the cation.

  4. Evaluation of two putative susceptibility loci for oral clefts in the Danish population

    DEFF Research Database (Denmark)

    Mitchell, L E; Murray, J C; O'Brien, S;

    2001-01-01

    Previous studies suggest that the risk of nonsyndromic cleft lip with or without cleft palate (CL+/-P) and isolated cleft palate (CP) is influenced by genetic variation at several loci and that the relation between specific genetic variants and disease risk may be modified by environmental factors....... The present study evaluated potential associations between CL+/-P and CP and two putative clefting susceptibility loci, MSX1 and TGFB3, using data from a nationwide case-control study conducted in Denmark from 1991 to 1994. The potential effects of interactions between these genes and two common environmental...

  5. Role of putative neurotransmitters in the central gastric antisecretory effect of prostaglandin E2 in rats.

    OpenAIRE

    Puurunen, J.

    1985-01-01

    The role of putative neurotransmitters of the central nervous system in the central gastric antisecretory effect of prostaglandin E2 (PGE2) was investigated in pylorus-ligated rats. Pretreatment of the rats with an intracerebroventricular (i.c.v.) injection of 6-hydroxydopamine (6-OHDA) prevented the antisecretory effect of the i.c.v. administration of PGE2, whereas pretreatment with 5,6-dihydroxytryptamine (5,6-DHT) plus p-chlorophenylalanine (PCPA) had no effect. I.c.v.-administered phentol...

  6. A putative, novel coli surface antigen 8B (CS8B) of enterotoxigenic Escherichia coli.

    Science.gov (United States)

    Njoroge, Samuel M; Boinett, Christine J; Madé, Laure F; Ouko, Tom T; Fèvre, Eric M; Thomson, Nicholas R; Kariuki, Samuel

    2015-10-01

    Enterotoxigenic Escherichia coli (ETEC) strains harbor multiple fimbriae and pili to mediate host colonization, including the type IVb pilus, colonization factor antigen III (CFA/III). Not all colonization factors are well characterized or known in toxin positive ETEC isolates, which may have an impact identifying ETEC isolates based on molecular screening of these biomarkers. We describe a novel coli surface antigen (CS) 8 subtype B (CS8B), a family of CFA/III pilus, in a toxin producing ETEC isolate from a Kenyan collection. In highlighting the existence of this putative CS, we provide the sequence and specific primers, which can be used alongside other ETEC primers previously described.

  7. A specimen of Rhamphorhynchus with soft tissue preservation, stomach contents and a putative coprolite

    Directory of Open Access Journals (Sweden)

    David Hone

    2015-08-01

    Full Text Available Despite being known for nearly two centuries, new specimens of the derived non-pterodactyloid pterosaur Rhamphorhynchus continue to be discovered and reveal new information about their anatomy and palaeobiology. Here we describe a specimen held in the collections of the Royal Tyrrell Museum of Palaeontology, Alberta, Canada that shows both preservation and impressions of soft tissues, and also preserves material interpreted as stomach contents of vertebrate remains and, uniquely, a putative coprolite. The specimen also preserves additional evidence for fibers in the uropatagium.

  8. A putative transglycosylase encoded by SCO4132 influences morphological differentiation and actinorhodin production in Streptomyces coelicolor

    Institute of Scientific and Technical Information of China (English)

    Pengfei Xie; Ana Zeng; Xiaoting Lv; Qiuxiang Cheng; Zhongjun Qin

    2013-01-01

    Here we report that tgdA,a novel gene encoding a putative transglycosylase,affects both the morphological differentiation and the yield of blue-pigmented compound actinorhodin in Streptomyces coelicolor.The tgdA null mutant displays sparse aerial hyphae and irregular spore chains frequently lacking chromosomal DNA.Elevated actinorhodin production coincides with the overexpression of actⅡ-orf4 in mutant.tgdA expression is temporally and developmentally regulated.The tgdA orthologs in Streptomyces avermilitis and Streptomyces lividans also affect differentiation.

  9. Prevalence of Clinical Periodontitis and Putative Periodontal Pathogens among South Indian Pregnant Women

    Directory of Open Access Journals (Sweden)

    Chaitanya Tellapragada

    2014-01-01

    Full Text Available In view of recent understanding of the association of periodontal infections and adverse pregnancy outcomes, the present investigation was undertaken to study the periodontal infections among 390 asymptomatic pregnant women and to find an association of bacterial etiologies with the disease. Prevalence of gingivitis was 38% and clinical periodontitis was 10% among the study population. Subgingival plaque specimens were subjected to multiplex PCR targeting ten putative periodontopathogenic bacteria. Among the periodontitis group, high detection rates of Porphyromonas gingivalis (56%, Prevotella nigrescens (44%, Treponema denticola (32%, and Prevotella intermedius (24% were noted along with significant association with the disease (P<0.05.

  10. A specimen of Rhamphorhynchus with soft tissue preservation, stomach contents and a putative coprolite.

    Science.gov (United States)

    Hone, David; Henderson, Donald M; Therrien, François; Habib, Michael B

    2015-01-01

    Despite being known for nearly two centuries, new specimens of the derived non-pterodactyloid pterosaur Rhamphorhynchus continue to be discovered and reveal new information about their anatomy and palaeobiology. Here we describe a specimen held in the collections of the Royal Tyrrell Museum of Palaeontology, Alberta, Canada that shows both preservation and impressions of soft tissues, and also preserves material interpreted as stomach contents of vertebrate remains and, uniquely, a putative coprolite. The specimen also preserves additional evidence for fibers in the uropatagium.

  11. A comparison of the effects of two putative 5-hydroxytryptamine renal prodrugs in normal man.

    OpenAIRE

    LI KAM WA, T. C.; Freestone, S.; Samson, R. R.; JOHNSTON, N. R.; Lee, M.R.

    1993-01-01

    1. The effects of 1 h intravenous infusions of equimolar amounts of two putative 5-hydroxytryptamine (5-HT) renal prodrugs, 5-hydroxy-L-tryptophan (5-HTP, 10 micrograms kg-1 min-1) and gamma-L-glutamyl-5-hydroxy-L-tryptophan (glu-5-HTP, 16.6 micrograms kg-1 min-1) were examined in five healthy male volunteers in a randomised, placebo-controlled, cross-over study. 2. Both compounds increased urinary excretion of 5-HT and there was greater extra-renal formation of 5-HT following 5-HTP administr...

  12. New Aperture Partitioning Element

    Science.gov (United States)

    Griffin, S.; Calef, B.; Williams, S.

    Postprocessing in an optical system can be aided by adding an optical element to partition the pupil into a number of segments. When imaging through the atmosphere, the recorded data are blurred by temperature-induced variations in the index of refraction along the line of sight. Using speckle imaging techniques developed in the astronomy community, this blurring can be corrected to some degree. The effectiveness of these techniques is diminished by redundant baselines in the pupil. Partitioning the pupil reduces the degree of baseline redundancy, and therefore improves the quality of images that can be obtained from the system. It is possible to implement the described approach on an optical system with a segmented primary mirror, but not very practical. This is because most optical systems do not have segmented primary mirrors, and those that do have relatively low bandwidth positioning of segments due to their large mass and inertia. It is much more practical to position an active aperture partitioning element at an aft optics pupil of the optical system. This paper describes the design, implementation and testing of a new aperture partitioning element that is completely reflective and reconfigurable. The device uses four independent, annular segments that can be positioned with a high degree of accuracy without impacting optical wavefront of each segment. This mirror has been produced and is currently deployed and working on the 3.6 m telescope.

  13. Helium the disappearing element

    CERN Document Server

    Sears, Wheeler M

    2015-01-01

    The subject of the book is helium, the element, and its use in myriad applications including MRI machines, particle accelerators, space telescopes, and of course balloons and blimps. It was at the birth of our Universe, or the Big Bang, where the majority of cosmic helium was created; and stellar helium production continues. Although helium is the second most abundant element in the Universe, it is actually quite rare here on Earth and only exists because of radioactive elements deep within the Earth. This book includes a detailed history of the discovery of helium, of the commercial industry built around it, how the helium we actually encounter is produced within the Earth, and the state of the helium industry today. The gas that most people associate with birthday party balloons is running out. “Who cares?” you might ask. Well, without helium, MRI machines could not function, rockets could not go into space, particle accelerators such as those used by CERN could not operate, fiber optic cables would not...

  14. New mobile genetic elements in Cupriavidus metallidurans CH34, their possible roles and occurrence in other bacteria.

    Science.gov (United States)

    Van Houdt, Rob; Monchy, Sébastien; Leys, Natalie; Mergeay, Max

    2009-08-01

    Cupriavidus metallidurans strain CH34 is a beta-Proteobacterium that thrives in low concentrations of heavy metals. The genetic determinants of resistance to heavy metals are located on its two chromosomes, and are particularly abundant in the two megaplasmids, pMOL28 and pMOL30. We explored the involvement of mobile genetic elements in acquiring these and others traits that might be advantageous in this strain using genome comparison of Cupriavidus/Ralstonia strains and related beta-Proteobacteria. At least eleven genomic islands were identified on the main replicon, three on pMOL28 and two on pMOL30. Multiple islands contained genes for heavy metal resistance or other genetic determinants putatively responding to harsh environmental conditions. However, cryptic elements also were noted. New mobile genetic elements (or variations of known ones) were identified through synteny analysis, allowing the detection of mobile genetic elements outside the bias of a selectable marker. Tn4371-like conjugative transposons involved in chemolithotrophy and degradation of aromatic compounds were identified in strain CH34, while similar elements involved in heavy metal resistance were found in Delftia acidovorans SPH-1 and Bordetella petrii DSM12804. We defined new transposons, viz., Tn6048 putatively involved in the response to heavy metals and Tn6050 carrying accessory genes not classically associated with transposons. Syntenic analysis also revealed new transposons carrying metal response genes in Burkholderia xenovorans LB400, and other bacteria. Finally, other putative mobile elements, which were previously unnoticed but apparently common in several bacteria, were also revealed. This was the case for triads of tyrosine-based site-specific recombinases and for an int gene paired with a putative repressor and associated with chromate resistance.

  15. The transuranium elements: From neptunium and plutonium to element 112

    Energy Technology Data Exchange (ETDEWEB)

    Hoffman, D.C. [California Univ., Berkeley, CA (United States)]|[Lawrence Livermore National Lab., CA (United States)

    1996-07-26

    Beginning in the 1930`s, both chemists and physicists became interested in synthesizing new artificial elements. The first transuranium element, Np, was synthesized in 1940. Over the past six decades, 20 transuranium elements have been produced. A review of the synthesis is given. The procedure of naming the heavy elements is also discussed. It appears feasible to produce elements 113 and 114. With the Berkeley Gas-filled Separator, it should be possible to reach the superheavy elements in the region of the spherical Z=114 shell, but with fewer neutrons than the N=184 spherical shell. 57 refs, 6 figs.

  16. Identification of a novel insertion sequence element in Streptococcus agalactiae. bspeller@imib.rwth-aachen.de.

    Science.gov (United States)

    Spellerberg, B; Martin, S; Franken, C; Berner, R; Lütticken, R

    2000-01-04

    Gain and loss of bacterial pathogenicity is often associated with mobile genetic elements. A novel insertion sequence (IS) element designated ISSa4 was identified in Streptococcus agalactiae (group B streptococci). The 963bp IS element is flanked by 25bp perfect inverted repeats and led to the duplication of a 9bp target sequence at the insertion site. ISSa4 contains one open reading frame coding for a putative transposase of 287 aa and exhibits closest similarities to insertion elements of the IS982 family which has previously not been identified in streptococci. Analysis of different S. agalactiae strains showed that the copy number of ISSa4 in S. agalactiae varies significantly between strains. The S. agalactiae strain with the highest copy number of ISSa4 was nonhemolytic and harbored one copy inserted in cylB, which encodes the membrane-spanning domain of the putative hemolysin transporter (Spellerberg et al., 1999. Identification of genetic determinants for the hemolytic activity of Streptococcus agalactiae by ISS1 transposition. J. Bacteriol. 181, 3212-3219). Determination of the distribution of ISSa4 in different S. agalactiae strains revealed that ISSa4 could be detected only in strains isolated after 1996, which might indicate a recent acquisition of this novel insertion element by S. agalactiae.

  17. Constraints on the location of a putative distant massive body in the Solar System and on the External Field Effect of MOND from recent planetary data

    CERN Document Server

    Iorio, Lorenzo

    2011-01-01

    We analytically work out the long-term variations caused on the motion of a planet orbiting a star by a distant, pointlike massive object X (Planet X/Nemesis/Tyche). It turns out that, apart from the semimajor axis $a$, all the other Keplerian orbital elements of the perturbed planet experience long-term variations which are complicated functions of the orbital configurations of both the planet itself and of X. A numerical integration of the equations of motion of the perturbed planet yielding the temporal evolution of all its orbital elements successfully confirms our analytical results. We infer constraints on the minimum distance $d_{\\rm X}$ at which the putative body X can exist by comparing, first, our prediction of the long-term variation of the longitude of the perihelion $\\varpi$ to the latest empirical determinations of the corrections $\\Delta\\dot\\varpi$ to the standard Newtonian/Einsteinian secular precessions of several planets of the solar system recently obtained. Independent teams of astronomers...

  18. 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.

  19. Putative cholesterol-binding sites in human immunodeficiency virus (HIV) coreceptors CXCR4 and CCR5.

    Science.gov (United States)

    Zhukovsky, Mikhail A; Lee, Po-Hsien; Ott, Albrecht; Helms, Volkhard

    2013-04-01

    Using molecular docking, we identified a cholesterol-binding site in the groove between transmembrane helices 1 and 7 near the inner membrane-water interface of the G protein-coupled receptor CXCR4, a coreceptor for HIV entry into cells. In this docking pose, the amino group of lysine K67 establishes a hydrogen bond with the hydroxyl group of cholesterol, whereas tyrosine Y302 stacks with cholesterol by its aromatic side chain, and a number of residues form hydrophobic contacts with cholesterol. Sequence alignment showed that a similar putative cholesterol-binding site is also present in CCR5, another HIV coreceptor. We suggest that the interaction of cholesterol with these putative cholesterol-binding sites in CXCR4 and CCR5 is responsible for the presence of these receptors in lipid rafts, for the effect of cholesterol on their conformational stability and function, and for the role that cell cholesterol plays in the cell entry of HIV strains that use these membrane proteins as coreceptors. We propose that mutations of residues that are involved in cholesterol binding will make CXCR4 and CCR5 insensitive to membrane cholesterol content. Cholesterol-binding sites in HIV coreceptors are potential targets for steroid drugs that bind to CXCR4 and CCR5 with higher binding affinity than cholesterol, but do not stabilize the native conformation of these proteins.

  20. On the Origin of the Supergiant HI Shell and Putative Companion in NGC 6822

    CERN Document Server

    Cannon, John M; Weisz, Daniel R; Skillman, Evan D; Dolphin, Andrew E; Bigiel, Frank; Cole, Andrew A; de Blok, W J G; Walter, Fabian

    2012-01-01

    We present new Hubble Space Telescope Advanced Camera for Surveys imaging of six positions spanning 5.8 kpc of the HI major axis of the Local Group dIrr NGC 6822, including both the putative companion galaxy and the large HI hole. The resulting deep color magnitude diagrams show that NGC 6822 has formed >50% of its stars in the last ~5 Gyr. The star formation histories of all six positions are similar over the most recent 500 Myr, including low-level star formation throughout this interval and a weak increase in star formation rate during the most recent 50 Myr. Stellar feedback can create the giant HI hole, assuming that the lifetime of the structure is longer than 500 Myr; such long-lived structures have now been observed in multiple systems and may be the norm in galaxies with solid-body rotation. The old stellar populations (red giants and red clump stars) of the putative companion are consistent with those of the extended halo of NGC 6822; this argues against the interpretation of this structure as a bon...

  1. ON THE ORIGIN OF THE SUPERGIANT H I SHELL AND PUTATIVE COMPANION IN NGC 6822

    Energy Technology Data Exchange (ETDEWEB)

    Cannon, John M.; O' Leary, Erin M. [Department of Physics and Astronomy, Macalester College, 1600 Grand Avenue, Saint Paul, MN 55105 (United States); Weisz, Daniel R. [Department of Astronomy, University of Washington, Seattle, WA 98195 (United States); Skillman, Evan D. [Minnesota Institute for Astrophysics, University of Minnesota, Minneapolis, MN 55455 (United States); Dolphin, Andrew E. [Raytheon Company, 1151 East Hermans Road, Tucson, AZ 85706 (United States); Bigiel, Frank [Institut fuer Theoretische Astrophysik, Universitaet Heidelberg, Albert-Ueberle-Str. 2, 69120 Heidelberg (Germany); Cole, Andrew A.; Walter, Fabian [School of Mathematics and Physics, University of Tasmania, Private Bag 37, Hobart 7001, Tasmania (Australia); De Blok, W.J.G., E-mail: jcannon@macalester.edu, E-mail: eoleary@macalester.edu, E-mail: dweisz@astro.washington.edu, E-mail: skillman@astro.umn.edu, E-mail: adolphin@raytheon.com, E-mail: bigiel@uni-heidelberg.de, E-mail: andrew.cole@utas.edu.au, E-mail: edeblok@ast.uct.ac.za, E-mail: walter@mpia.de [Department of Astronomy, University of Cape Town, Rondebosch 7700 (South Africa)

    2012-03-10

    We present new Hubble Space Telescope Advanced Camera for Surveys imaging of six positions spanning 5.8 kpc of the H I major axis of the Local Group dIrr NGC 6822, including both the putative companion galaxy and the large H I hole. The resulting deep color-magnitude diagrams show that NGC 6822 has formed >50% of its stars in the last {approx}5 Gyr. The star formation histories of all six positions are similar over the most recent 500 Myr, including low-level star formation throughout this interval and a weak increase in star formation rate during the most recent 50 Myr. Stellar feedback can create the giant H I hole, assuming that the lifetime of the structure is longer than 500 Myr; such long-lived structures have now been observed in multiple systems and may be the norm in galaxies with solid-body rotation. The old stellar populations (red giants and red clump stars) of the putative companion are consistent with those of the extended halo of NGC 6822; this argues against the interpretation of this structure as a bona fide interacting companion galaxy and against its being linked to the formation of the H I hole via an interaction. Since there is no evidence in the stellar population of a companion galaxy, the most likely explanation of the extended H I structure in NGC 6822 is a warped disk inclined to the line of sight.

  2. Identification of putative DnaN-binding motifs in plasmid replication initiation proteins.

    Science.gov (United States)

    Dalrymple, Brian P; Kongsuwan, Kritaya; Wijffels, Gene

    2007-01-01

    Recently the plasmid RK2 replication initiation protein, TrfA, has been shown to bind to the beta subunit of DNA Polymerase III (DnaN) via a short pentapeptide with the consensus QL[S/D]LF. A second consensus peptide, the hexapeptide QLxLxL, has also been demonstrated to mediate binding to DnaN. Here we describe the results of a comprehensive survey of replication initiation proteins encoded by bacterial plasmids to identify putative DnaN-binding sites. Both pentapeptide and hexapeptide motifs have been identified in a number of families of replication initiation proteins. The distribution of sites is sporadic and closely related families of proteins may differ in the presence, location, or type of putative DnaN-binding motif. Neither motif has been identified in replication initiation proteins encoded by plasmids that replicate via rolling circles or strand displacement. The results suggest that the recruitment of DnaN to the origin of replication of a replisome by plasmid replication initiation proteins is not generally required for plasmid replication, but that in some cases it may be beneficial for efficiency of replication initiation.

  3. Uncovering the Putative B-Star Binary Companion of the SN 1993J Progenitor

    CERN Document Server

    Fox, Ori D; Van Dyk, Schuyler D; Filippenko, Alexei V; Fransson, Claes; Matheson, Thomas; Cenko, S Bradley; Chandra, Poonam; Dwarkadas, Vikram; Li, Weidong; Parker, Alex H; Smith, Nathan

    2014-01-01

    The Type IIb supernova (SN) 1993J is one of only a few stripped-envelope supernovae with a progenitor star identified in pre-explosion images. SN IIb models typically invoke H envelope stripping by mass transfer in a binary system. For the case of SN 1993J, the models suggest that the companion grew to 22 M_solar and became a source of ultraviolet (UV) excess. Located in M81, at a distance of only 3.6 Mpc, SN 1993J offers one of the best opportunities to detect the putative companion and test the progenitor model. Previously published near-UV spectra in 2004 showed evidence for absorption lines consistent with a hot (B2 Ia) star, but the field was crowded and dominated by flux from the SN. Here we present Hubble Space Telescope (HST) Cosmic Origins Spectrograph (COS) and Wide-Field Camera 3 (WFC3) observations of SN 1993J from 2012, at which point the flux from the SN had faded sufficiently to potentially measure the UV continuum properties from the putative companion. The resulting UV spectrum is consistent ...

  4. Evolution and structural diversification of PILS putative auxin carriers in plants

    Directory of Open Access Journals (Sweden)

    Elena eFeraru

    2012-10-01

    Full Text Available The phytohormone auxin contributes to virtually every aspect of the plant development. The spatiotemporal distribution of auxin depends on a complex interplay between auxin metabolism and intercellular auxin transport. Intracellular auxin compartmentalization provides another link between auxin transport processes and auxin metabolism. The PIN-LIKES (PILS putative auxin carriers localize to the endoplasmic reticulum (ER and contribute to cellular auxin homeostasis. PILS proteins regulate intracellular auxin accumulation, the rate of auxin conjugation and subsequently, affect nuclear auxin signalling. Here, we investigate sequence diversification of the PILS family in Arabidopsis thaliana and provide insights into the evolution of these novel putative auxin carriers in plants. Our data suggest that PILS proteins are conserved throughout the plant lineage and expanded during higher plant evolution. PILS proteins diversified early during plant evolution into three clades. Besides the ancient Clade I encompassing non-land plant species, PILS proteins evolved into two clades. The diversification of Clade II and Clade III occurred already at the level of non-vascular plant evolution and, hence, both clades contain vascular and non-vascular plant species. Nevertheless, Clade III contains fewer non- and increased numbers of vascular plants, indicating higher importance of Clade III for vascular plant evolution. Notably, PILS proteins are distinct and appear evolutionarily older than the prominent PIN-FORMED auxin carriers. Moreover, we revealed particular PILS sequence divergence in Arabidopsis and assume that these alterations could contribute to distinct gene regulations and protein functions.

  5. Characterization of putative multidrug resistance transporters of the major facilitator-superfamily expressed in Salmonella Typhi.

    Science.gov (United States)

    Shaheen, Aqsa; Ismat, Fouzia; Iqbal, Mazhar; Haque, Abdul; De Zorzi, Rita; Mirza, Osman; Walz, Thomas; Rahman, Moazur

    2015-05-01

    Multidrug resistance mediated by efflux pumps is a well-known phenomenon in infectious bacteria. Although much work has been carried out to characterize multidrug efflux pumps in Gram-negative and Gram-positive bacteria, such information is still lacking for many deadly pathogens. The aim of this study was to gain insight into the substrate specificity of previously uncharacterized transporters of Salmonella Typhi to identify their role in the development of multidrug resistance. S. Typhi genes encoding putative members of the major facilitator superfamily were cloned and expressed in the drug-hypersensitive Escherichia coli strain KAM42, and tested for transport of 25 antibacterial compounds, including representative antibiotics of various classes, antiseptics, dyes and detergents. Of the 15 tested putative transporters, STY0901, STY2458 and STY4874 exhibited a drug-resistance phenotype. Among these, STY4874 conferred resistance to at least ten of the tested antimicrobials: ciprofloxacin, norfloxacin, levofloxacin, kanamycin, streptomycin, gentamycin, nalidixic acid, chloramphenicol, ethidium bromide, and acriflavine, including fluoroquinolone antibiotics, which were drugs of choice to treat S. Typhi infections. Cell-based functional studies using ethidium bromide and acriflavine showed that STY4874 functions as a H(+)-dependent exporter. These results suggest that STY4874 may be an important drug target, which can now be tested by studying the susceptibility of a STY4874-deficient S. Typhi strain to antimicrobials.

  6. Mutation analysis of novel human liver-related putative tumor suppressor gene in hepatocellular carcinoma

    Institute of Scientific and Technical Information of China (English)

    Cheng Liao; Tsai-Ping Li; Mu-Jun Zhao; Jing Zhao; Hai Song; Pascal Pineau; Agnès Marchio; Anne Dejean; Pierre Tiollais; Hong-Yang Wang

    2003-01-01

    AIM: To find the point mutations meaningful for inactivationof liver-related putative tumor suppressor gene (LPTS) gene,a human novel liver-related putative tumor suppressor geneand telomerase inhibitor in hepatocellular carcinoma.METHODS: The entire coding sequence of LPTS genewas examined for mutations by single strand conformationpolymorphism (SSCP) assay and PCR products directsequencing in 56 liver cancer cell lines, 7 ovarian cancerand 7 head & neck tumor cell lines and 70 pairs of HCCtissues samples. The cDNA fragment coding for the mostfrequent mutant protein was subcloned into GST fusionexpression vector. The product was expressed in E. coliand purified by glutathione-agarose column. Telomericrepeat amplification protocol (TRAP) assays wereperformed to study the effect of point mutation totelomerase inhibitory activity.RESULTS: SSCP gels showed the abnormal shifting bandsand DNA sequencing found that there were 5 differentmutations and/or polymorphisms in 12 tumor cell lineslocated at exon2, exon5 and exon7. The main alterationswere A(778)A/G and A(880)T in exon7. The change in siteof 778 could not be found in HCC tissue samples, while themutation in position 880 was seen in 7 (10 %) cases. Themutation in the site of 880 had no effect on telomeraseinhibitory activity.CONCLUSION: Alterations identified in this study arepolymorphisms of LPTS gene. LPTS mutations occur in HCCbut are infrequent and of little effect on the telomeraseinhibitory function of the protein. Epigenetics, such asmethylation, acetylation, may play the key role in inactivationof LPTS.

  7. Expression of putative virulence factors in the potato pathogen Clavibacter michiganensis subsp. sepedonicus during infection.

    Science.gov (United States)

    Holtsmark, Ingrid; Takle, Gunnhild W; Brurberg, May Bente

    2008-02-01

    The Gram-positive bacterium Clavibacter michiganensis subsp. sepedonicus is the causal agent of bacterial wilt and ring rot of potato. So far, only two proteins have been shown to be essential for virulence, namely a plasmid-encoded cellulase CelA and a hypersensitive response-inducing protein. We have examined the relative expression of CelA and eight putative virulence factors during infection of potato and in liquid culture, using quantitative real-time PCR. The examined putative virulence genes were celB, a cellulase-encoding gene and genes encoding a pectate lyase, a xylanase and five homologues of the Clavibacter michiganensis subsp. michiganensis pathogenicity factor Pat-1 thought to encode a serine protease. Six of the nine assayed genes were up-regulated during infection of potato, including celA, celB, the xylanase gene, and two of the pat genes. The pectate lyase gene showed only slightly elevated expression, whereas three of the five examined pat genes were down-regulated during infection in potato. Interestingly, the two up-regulated pat genes showed a noticeable sequence difference compared to the three down-regulated pat genes. These results reveal several new proteins that are likely to be involved in Clavibacter michiganensis subsp. sepedonicus pathogenicity.

  8. Transcriptome of Aphanomyces euteiches: new oomycete putative pathogenicity factors and metabolic pathways.

    Directory of Open Access Journals (Sweden)

    Elodie Gaulin

    Full Text Available Aphanomyces euteiches is an oomycete pathogen that causes seedling blight and root rot of legumes, such as alfalfa and pea. The genus Aphanomyces is phylogenically distinct from well-studied oomycetes such as Phytophthora sp., and contains species pathogenic on plants and aquatic animals. To provide the first foray into gene diversity of A. euteiches, two cDNA libraries were constructed using mRNA extracted from mycelium grown in an artificial liquid medium or in contact to plant roots. A unigene set of 7,977 sequences was obtained from 18,864 high-quality expressed sequenced tags (ESTs and characterized for potential functions. Comparisons with oomycete proteomes revealed major differences between the gene content of A. euteiches and those of Phytophthora species, leading to the identification of biosynthetic pathways absent in Phytophthora, of new putative pathogenicity genes and of expansion of gene families encoding extracellular proteins, notably different classes of proteases. Among the genes specific of A. euteiches are members of a new family of extracellular proteins putatively involved in adhesion, containing up to four protein domains similar to fungal cellulose binding domains. Comparison of A. euteiches sequences with proteomes of fully sequenced eukaryotic pathogens, including fungi, apicomplexa and trypanosomatids, allowed the identification of A. euteiches genes with close orthologs in these microorganisms but absent in other oomycetes sequenced so far, notably transporters and non-ribosomal peptide synthetases, and suggests the presence of a defense mechanism against oxidative stress which was initially characterized in the pathogenic trypanosomatids.

  9. Regulation of Arabidopsis Early Anther Development by Putative Cell-Cell Signaling Molecules and Transcriptional Regulators

    Institute of Scientific and Technical Information of China (English)

    Yu-Jin Sun; Carey LH Hord; Chang-Bin Chen; Hong Ma

    2007-01-01

    Anther development in flowering plants involves the formation of several cell types, including the tapetal and pollen mother cells. The use of genetic and molecular tools has led to the identification and characterization of genes that are critical for normal cell division and differentiation in Arabidopsis early anther development. We review here several recent studies on these genes, including the demonstration that the putative receptor protein kinases BAM1 and BAM2 together play essential roles in the control of early cell division and differentiation. In addition, we discuss the hypothesis that BAM1/2 may form a positive-negative feedback regulatory loop with a previously identified key regulator, SPOROCYTELESS (also called NOZZLE),to control the balance between sporogenous and somatic cell types in the anther. Furthermore, we summarize the isolation and functional analysis of the DYSFUNCTIONAL TAPETUM1 (DYT1) gene in promoting proper tapetal cell differentiation. Our finding that DYT1 encodes a putative transcription factor of the bHLH family, as well as relevant expression analyses, strongly supports a model that DYT1 serves as a critical link between upstream factors and downstream target genes that are critical for normal tapetum development and function. These studies, together with other recently published works, indicate that cell-cell communication and transcriptional control are key processes essential for cell fate specification in anther development.

  10. Identification and site of action of the remaining four putative pseudouridine synthases in Escherichia coli.

    Science.gov (United States)

    Del Campo, M; Kaya, Y; Ofengand, J

    2001-11-01

    There are 10 known putative pseudouridine synthase genes in Escherichia coli. The products of six have been previously assigned, one to formation of the single pseudouridine in 16S RNA, three to the formation of seven pseudouridines in 23S RNA, and three to the formation of three pseudouridines in tRNA (one synthase makes pseudouridine in 23S RNA and tRNA). Here we show that the remaining four putative synthase genes make bona fide pseudouridine synthases and identify which pseudouridines they make. RluB (formerly YciL) and RluE (formerly YmfC) make pseudouridine2605 and pseudouridine2457, respectively, in 23S RNA. RluF (formerly YjbC) makes the newly discovered pseudouridine2604 in 23S RNA, and TruC (formerly YqcB) makes pseudouridine65 in tRNA(Ile1) and tRNA(Asp). Deletion of each of these synthase genes individually had no effect on exponential growth in rich media at 25 degrees C, 37 degrees C, or 42 degrees C. A strain lacking RluB and RluF also showed no growth defect under these conditions. Mutation of a conserved aspartate in a common sequence motif, previously shown to be essential for the other six E. coli pseudouridine synthases and several yeast pseudouridine synthases, also caused a loss of in vivo activity in all four of the synthases studied in this work.

  11. Transcriptome Analysis Reveals Putative Genes Involved in Iridoid Biosynthesis in Rehmannia glutinosa

    Directory of Open Access Journals (Sweden)

    Xianen Li

    2012-10-01

    Full Text Available Rehmannia glutinosa, one of the most widely used herbal medicines in the Orient, is rich in biologically active iridoids. Despite their medicinal importance, no molecular information about the iridoid biosynthesis in this plant is presently available. To explore the transcriptome of R. glutinosa and investigate genes involved in iridoid biosynthesis, we used massively parallel pyrosequencing on the 454 GS FLX Titanium platform to generate a substantial EST dataset. Based on sequence similarity searches against the public sequence databases, the sequences were first annotated and then subjected to Gene Ontology (GO and Kyoto Encyclopedia of Genes and Genomes (KEGG based analysis. Bioinformatic analysis indicated that the 454 assembly contained a set of genes putatively involved in iridoid biosynthesis. Significantly, homologues of the secoiridoid pathway genes that were only identified in terpenoid indole alkaloid producing plants were also identified, whose presence implied that route II iridoids and route I iridoids share common enzyme steps in the early stage of biosynthesis. The gene expression patterns of four prenyltransferase transcripts were analyzed using qRT-PCR, which shed light on their putative functions in tissues of R. glutinosa. The data explored in this study will provide valuable information for further studies concerning iridoid biosynthesis.

  12. Mapping the flow of information within the putative mirror neuron system during gesture observation.

    Science.gov (United States)

    Schippers, Marleen B; Keysers, Christian

    2011-07-01

    The putative mirror neuron system may either function as a strict feed-forward system or as a dynamic control system. A strict feed-forward system would predict that action observation leads to a predominantly temporal→parietal→premotor flow of information in which a visual representation is transformed into motor-programs which contribute to action understanding. Instead, a dynamic feedback control system would predict that the reverse direction of information flow predominates because of a combination of inhibitory forward and excitatory inverse models. Here we test which of these conflicting predictions best matches the information flow within the putative mirror neuron system (pMNS) and between the pMNS and the rest of the brain during the observation of comparatively long naturalistic stretches of communicative gestures. We used Granger causality to test the dominant direction of influence. Our results fit the predictions of the dynamic feedback control system: we found predominantly an information flow within the pMNS from premotor to parietal and middle temporal cortices. This is more pronounced during an active guessing task than while passively reviewing the same gestures. In particular, the ventral premotor cortex sends significantly more information to other pMNS areas than it receives during active guessing than during passive observation.

  13. Identification and functional analysis of Penicillium digitatum genes putatively involved in virulence towards citrus fruit.

    Science.gov (United States)

    López-Pérez, Mario; Ballester, Ana-Rosa; González-Candelas, Luis

    2015-04-01

    The fungus Penicillium digitatum, the causal agent of green mould rot, is the most destructive post-harvest pathogen of citrus fruit in Mediterranean regions. In order to identify P. digitatum genes up-regulated during the infection of oranges that may constitute putative virulence factors, we followed a polymerase chain reaction (PCR)-based suppression subtractive hybridization and cDNA macroarray hybridization approach. The origin of expressed sequence tags (ESTs) was determined by comparison against the available genome sequences of both organisms. Genes coding for fungal proteases and plant cell wall-degrading enzymes represent the largest categories in the subtracted cDNA library. Northern blot analysis of a selection of P. digitatum genes, including those coding for proteases, cell wall-related enzymes, redox homoeostasis and detoxification processes, confirmed their up-regulation at varying time points during the infection process. Agrobacterium tumefaciens-mediated transformation was used to generate knockout mutants for two genes encoding a pectin lyase (Pnl1) and a naphthalene dioxygenase (Ndo1). Two independent P. digitatum Δndo1 mutants were as virulent as the wild-type. However, the two Δpnl1 mutants analysed were less virulent than the parental strain or an ectopic transformant. Together, these results provide a significant advance in our understanding of the putative determinants of the virulence mechanisms of P. digitatum.

  14. Unprecedented diversity of catalytic domains in the first four modules of the putative pederin polyketide synthase.

    Science.gov (United States)

    Piel, Jörn; Wen, Gaiping; Platzer, Matthias; Hui, Dequan

    2004-01-03

    Polyketides of the pederin group are highly potent antitumor compounds found in terrestrial beetles and marine sponges. Pederin is used by beetles of the genera Paederus and Paederidus as a chemical defense. We have recently identified a group of putative pederin biosynthesis genes and localized them to the genome of an as yet unculturable Pseudomonas sp. symbiont, the likely true pederin producer. However, this polyketide synthase cluster lacks several genes expected for pederin production. Here we report an additional polyketide synthase encoded on a separate region of the symbiont genome. It contains at least three novel catalytic domains that are predicted to be involved in pederin chain initiation and the formation of an unusual exomethylene bond. The region is bordered by mobility pseudogenes; this suggests that gene transposition led to the disjointed cluster organization. With this work, all putative pederin genes have been identified. Their heterologous expression in a culturable bacterium will provide important insights into how sustainable sources of invertebrate-derived drug candidates can be created.

  15. Biochemical Characterization of Putative Adenylate Dimethylallyltransferase and Cytokinin Dehydrogenase from Nostoc sp. PCC 7120.

    Science.gov (United States)

    Frébortová, Jitka; Greplová, Marta; Seidl, Michael F; Heyl, Alexander; Frébort, Ivo

    2015-01-01

    Cytokinins, a class of phytohormones, are adenine derivatives common to many different organisms. In plants, these play a crucial role as regulators of plant development and the reaction to abiotic and biotic stress. Key enzymes in the cytokinin synthesis and degradation in modern land plants are the isopentyl transferases and the cytokinin dehydrogenases, respectively. Their encoding genes have been probably introduced into the plant lineage during the primary endosymbiosis. To shed light on the evolution of these proteins, the genes homologous to plant adenylate isopentenyl transferase and cytokinin dehydrogenase were amplified from the genomic DNA of cyanobacterium Nostoc sp. PCC 7120 and expressed in Escherichia coli. The putative isopentenyl transferase was shown to be functional in a biochemical assay. In contrast, no enzymatic activity was detected for the putative cytokinin dehydrogenase, even though the principal domains necessary for its function are present. Several mutant variants, in which conserved amino acids in land plant cytokinin dehydrogenases had been restored, were inactive. A combination of experimental data with phylogenetic analysis indicates that adenylate-type isopentenyl transferases might have evolved several times independently. While the Nostoc genome contains a gene coding for protein with characteristics of cytokinin dehydrogenase, the organism is not able to break down cytokinins in the way shown for land plants.

  16. TiD: Standalone software for mining putative drug targets from bacterial proteome.

    Science.gov (United States)

    Gupta, Reena; Pradhan, Dibyabhaba; Jain, Arun Kumar; Rai, Chandra Shekhar

    2017-01-01

    TiD is a standalone application, which relies on basic assumption that a protein must be essential for pathogens survival and non-homologous with host to qualify as putative target. With an input bacterial proteome, TiD removes paralogous proteins, picks essential ones, and excludes proteins homologous with host organisms. The targets illustrate non-homology with at least 40 out of 84 gut microbes, considered safe for human. TiD classifies proposed targets as known, novel and virulent. Users can perform pathway analysis, choke point analysis, interactome analysis, subcellular localization and functional annotations through web servers cross-referenced with the application. Drug targets identified by TiD for Listeria monocytogenes, Bacillus anthracis and Pseudomonas aeruginosa have revealed significant overlaps with previous studies. TiD takes <2h to scan putative targets from a bacterial proteome with ~5000 proteins; hence, we propose it as a useful tool for rational drug design. TiD is available at http://bmicnip.in/TiD/. Copyright © 2016 Elsevier Inc. All rights reserved.

  17. VenomKB, a new knowledge base for facilitating the validation of putative venom therapies.

    Science.gov (United States)

    Romano, Joseph D; Tatonetti, Nicholas P

    2015-11-24

    Animal venoms have been used for therapeutic purposes since the dawn of recorded history. Only a small fraction, however, have been tested for pharmaceutical utility. Modern computational methods enable the systematic exploration of novel therapeutic uses for venom compounds. Unfortunately, there is currently no comprehensive resource describing the clinical effects of venoms to support this computational analysis. We present VenomKB, a new publicly accessible knowledge base and website that aims to act as a repository for emerging and putative venom therapies. Presently, it consists of three database tables: (1) Manually curated records of putative venom therapies supported by scientific literature, (2) automatically parsed MEDLINE articles describing compounds that may be venom derived, and their effects on the human body, and (3) automatically retrieved records from the new Semantic Medline resource that describe the effects of venom compounds on mammalian anatomy. Data from VenomKB may be selectively retrieved in a variety of popular data formats, are open-source, and will be continually updated as venom therapies become better understood.

  18. Small-fragment genomic libraries for the display of putative epitopes from clinically significant pathogens.

    Science.gov (United States)

    Henics, T; Winkler, B; Pfeifer, U; Gill, S R; Buschle, M; von Gabain, A; Meinke, A L

    2003-07-01

    Taking advantage of whole genome sequences of bacterial pathogens in many thriving diseases with global impact, we developed a comprehensive screening procedure for the identification of putative vaccine candidate antigens. Importantly, this procedure relies on highly representative small-fragment genomic libraries that are expressed to display frame-selected epitope-size peptides on a bacterial cell surface and to interact directly with carefully selected disease-relevant high-titer sera. Here we describe the generation of small-fragment genomic libraries of Gram-positive and Gram-negative clinically significant pathogens, including Staphylococcus aureus and Staphylococcus epidermidis, Streptococcus pyogenes, Streptococcus agalactiae, and Streptococcus pneumoniae, Enterococcus faecalis, Helicobacter pylori, Chlamydia pneumoniae, the enterotoxigenic Escherichia coli, and Campylobacter jejuni. Large-scale sequencing revealed that the libraries, which provide an average of 20-fold coverage, were random and, as demonstrated with two S. aureus libraries, highly representative. Consistent with the comprehensive nature of this approach is the identification of epitopes that reside in both annotated and putatively novel open reading frames. The use of these libraries therefore allows for the rapid and direct identification of immunogenic epitopes with no apparent bias or difficulty that often associate with conventional expression methods.

  19. Five putative nucleoside triphosphate diphosphohydrolase genes are expressed in Trichomonas vaginalis.

    Science.gov (United States)

    Frasson, Amanda Piccoli; Dos Santos, Odelta; Meirelles, Lúcia Collares; Macedo, Alexandre José; Tasca, Tiana

    2016-01-01

    Trichomonas vaginalis is a protozoan that parasitizes the human urogenital tract causing trichomoniasis, the most common non-viral sexually transmitted disease. The parasite has unique genomic characteristics such as a large genome size and expanded gene families. Ectonucleoside triphosphate diphosphohydrolase (E-NTPDase) is an enzyme responsible for hydrolyzing nucleoside tri- and diphosphates and has already been biochemically characterized in T. vaginalis. Considering the important role of this enzyme in the production of extracellular adenosine for parasite uptake, we evaluated the gene expression of five putative NTPDases in T. vaginalis. We showed that all five putative TvNTPDase genes (TvNTPDase1-5) were expressed by both fresh clinical and long-term grown isolates. The amino acid alignment predicted the presence of the five crucial apyrase conserved regions, transmembrane domains, signal peptides, phosphorylation and catalytic sites. Moreover, a phylogenetic analysis showed that TvNTPDase sequences make up a clade with NTPDases intracellularly located. Biochemical NTPDase activity (ATP and ADP hydrolysis) is responsive to the serum-restrictive conditions and the gene expression of TvNTPDases was mostly increased, mainly TvNTPDase2 and TvNTPDase4, although there was not a clear pattern of expression among them. In summary, the present report demonstrates the gene expression patterns of predicted NTPDases in T. vaginalis.

  20. Functional characterization of PaLAX1, a putative auxin permease, in heterologous plant systems.

    Science.gov (United States)

    Hoyerová, Klára; Perry, Lucie; Hand, Paul; Lanková, Martina; Kocábek, Tomás; May, Sean; Kottová, Jana; Paces, Jan; Napier, Richard; Zazímalová, Eva

    2008-03-01

    We have isolated the cDNA of the gene PaLAX1 from a wild cherry tree (Prunus avium). The gene and its product are highly similar in sequences to both the cDNAs and the corresponding protein products of AUX/LAX-type genes, coding for putative auxin influx carriers. We have prepared and characterized transformed Nicotiana tabacum and Arabidopsis thaliana plants carrying the gene PaLAX1. We have proved that constitutive overexpression of PaLAX1 is accompanied by changes in the content and distribution of free indole-3-acetic acid, the major endogenous auxin. The increase in free indole-3-acetic acid content in transgenic plants resulted in various phenotype changes, typical for the auxin-overproducing plants. The uptake of synthetic auxin, 2,4-dichlorophenoxyacetic acid, was 3 times higher in transgenic lines compared to the wild-type lines and the treatment with the auxin uptake inhibitor 1-naphthoxyacetic acid reverted the changes caused by the expression of PaLAX1. Moreover, the agravitropic response could be restored by expression of PaLAX1 in the mutant aux1 plants, which are deficient in auxin influx carrier activity. Based on our data, we have concluded that the product of the gene PaLAX1 promotes the uptake of auxin into cells, and, as a putative auxin influx carrier, it affects the content and distribution of free endogenous auxin in transgenic plants.

  1. Sensitivity analysis of the relationship between disease occurrence and distance from a putative source of pollution

    Directory of Open Access Journals (Sweden)

    Emanuela Dreassi

    2008-05-01

    Full Text Available The relation between disease risk and a point source of pollution is usually investigated using distance from the source as a proxy of exposure. The analysis may be based on case-control data or on aggregated data. The definition of the function relating risk of disease and distance is critical, both in a classical and in a Bayesian framework, because the likelihood is usually very flat, even with large amounts of data. In this paper we investigate how the specification of the function relating risk of disease with distance from the source and of the prior distributions on the parameters of the function affects the results when case-control data and Bayesian methods are used. We consider different popular parametric models for the risk distance function in a Bayesian approach, comparing estimates with those derived by maximum likelihood. As an example we have analyzed the relationship between a putative source of environmental pollution (an asbestos cement plant and the occurrence of pleural malignant mesothelioma in the area of Casale Monferrato (Italy in 1987-1993. Risk of pleural malignant mesothelioma turns out to be strongly related to distance from the asbestos cement plant. However, as the models appeared to be sensitive to modeling choices, we suggest that any analysis of disease risk around a putative source should be integrated with a careful sensitivity analysis and possibly with prior knowledge. The choice of prior distribution is extremely important and should be based on epidemiological considerations.

  2. Putative monofunctional type I polyketide synthase units: a dinoflagellate-specific feature?

    Directory of Open Access Journals (Sweden)

    Karsten Eichholz

    Full Text Available Marine dinoflagellates (alveolata are microalgae of which some cause harmful algal blooms and produce a broad variety of most likely polyketide synthesis derived phycotoxins. Recently, novel polyketide synthesase (PKS transcripts have been described from the Florida red tide dinoflagellate Karenia brevis (gymnodiniales which are evolutionarily related to Type I PKS but were apparently expressed as monofunctional proteins, a feature typical of Type II PKS. Here, we investigated expression units of PKS I-like sequences in Alexandrium ostenfeldii (gonyaulacales and Heterocapsa triquetra (peridiniales at the transcript and protein level. The five full length transcripts we obtained were all characterized by polyadenylation, a 3' UTR and the dinoflagellate specific spliced leader sequence at the 5'end. Each of the five transcripts encoded a single ketoacylsynthase (KS domain showing high similarity to K. brevis KS sequences. The monofunctional structure was also confirmed using dinoflagellate specific KS antibodies in Western Blots. In a maximum likelihood phylogenetic analysis of KS domains from diverse PKSs, dinoflagellate KSs formed a clade placed well within the protist Type I PKS clade between apicomplexa, haptophytes and chlorophytes. These findings indicate that the atypical PKS I structure, i.e., expression as putative monofunctional units, might be a dinoflagellate specific feature. In addition, the sequenced transcripts harbored a previously unknown, apparently dinoflagellate specific conserved N-terminal domain. We discuss the implications of this novel region with regard to the putative monofunctional organization of Type I PKS in dinoflagellates.

  3. Method of securing filter elements

    Energy Technology Data Exchange (ETDEWEB)

    Brown, Erik P.; Haslam, Jeffery L.; Mitchell, Mark A.

    2016-10-04

    A filter securing system including a filter unit body housing; at least one tubular filter element positioned in the filter unit body housing, the tubular filter element having a closed top and an open bottom; a dimple in either the filter unit body housing or the top of the tubular filter element; and a socket in either the filter unit body housing or the top of the tubular filter element that receives the dimple in either the filter unit body housing or the top of the tubular filter element to secure the tubular filter element to the filter unit body housing.

  4. Identification of putative silicon transporters in the low accumulators N. tabacum and S. lycopersicum

    Science.gov (United States)

    Silicon (Si) has long been excluded from the list of essential elements based on a definition developed over 60 years ago. However, how can one deem an element “non-essential” if through millennia of adaptation a fundamental pathway of perception and accumulation leading to molecular changes within...

  5. It is elemental

    DEFF Research Database (Denmark)

    Delgado-Baquerizo, Manuel; Reich, Peter B.; Khachane, Amit N.

    2017-01-01

    It is well established that resource quantity and elemental stoichiometry play major roles in shaping below and aboveground plant biodiversity, but their importance for shaping microbial diversity in soil remains unclear. Here, we used statistical modeling on a regional database covering 179...... diversity and composition were primarily driven by variation in soil resource stoichiometry (total C:N:P ratios), itself linked to different land uses, and secondarily driven by other important biodiversity drivers such as climate, soil spatial heterogeneity, soil pH, root influence (plant-soil microbe...

  6. Elements of energy conversion

    CERN Document Server

    Russell, Charles R

    2013-01-01

    Elements of Energy Conversion brings together scattered information on the subject of energy conversion and presents it in terms of the fundamental thermodynamics that apply to energy conversion by any process. Emphasis is given to the development of the theory of heat engines because these are and will remain most important power sources. Descriptive material is then presented to provide elementary information on all important energy conversion devices. The book contains 10 chapters and opens with a discussion of forms of energy, energy sources and storage, and energy conversion. This is foll

  7. Osteoporosis and trace elements

    DEFF Research Database (Denmark)

    Aaseth, J.; Boivin, G.; Andersen, Ole

    2012-01-01

    More than 200 million people are affected by osteoporosis worldwide, as estimated by 2 million annual hip fractures and other debilitating bone fractures (vertebrae compression and Colles' fractures). Osteoporosis is a multi-factorial disease with potential contributions from genetic, endocrine...... in new bone and results in a net gain in bone mass, but may be associated with a tissue of poor quality. Aluminum induces impairment of bone formation. Gallium and cadmium suppresses bone turnover. However, exact involvements of the trace elements in osteoporosis have not yet been fully clarified...

  8. Elements of probability theory

    CERN Document Server

    Rumshiskii, L Z

    1965-01-01

    Elements of Probability Theory presents the methods of the theory of probability. This book is divided into seven chapters that discuss the general rule for the multiplication of probabilities, the fundamental properties of the subject matter, and the classical definition of probability. The introductory chapters deal with the functions of random variables; continuous random variables; numerical characteristics of probability distributions; center of the probability distribution of a random variable; definition of the law of large numbers; stability of the sample mean and the method of moments

  9. Elements of real analysis

    CERN Document Server

    Sprecher, David A

    2010-01-01

    This classic text in introductory analysis delineates and explores the intermediate steps between the basics of calculus and the ultimate stage of mathematics: abstraction and generalization.Since many abstractions and generalizations originate with the real line, the author has made it the unifying theme of the text, constructing the real number system from the point of view of a Cauchy sequence (a step which Dr. Sprecher feels is essential to learn what the real number system is).The material covered in Elements of Real Analysis should be accessible to those who have completed a course in

  10. Elements of analytical dynamics

    CERN Document Server

    Kurth, Rudolph; Stark, M

    1976-01-01

    Elements of Analytical Dynamics deals with dynamics, which studies the relationship between motion of material bodies and the forces acting on them. This book is a compilation of lectures given by the author at the Georgia and Institute of Technology and formed a part of a course in Topological Dynamics. The book begins by discussing the notions of space and time and their basic properties. It then discusses the Hamilton-Jacobi theory and Hamilton's principle and first integrals. The text concludes with a discussion on Jacobi's geometric interpretation of conservative systems. This book will

  11. Analytical elements of mechanics

    CERN Document Server

    Kane, Thomas R

    2013-01-01

    Analytical Elements of Mechanics, Volume 1, is the first of two volumes intended for use in courses in classical mechanics. The books aim to provide students and teachers with a text consistent in content and format with the author's ideas regarding the subject matter and teaching of mechanics, and to disseminate these ideas. The book opens with a detailed exposition of vector algebra, and no prior knowledge of this subject is required. This is followed by a chapter on the topic of mass centers, which is presented as a logical extension of concepts introduced in connection with centroids. A

  12. [Healthcare marketing elements].

    Science.gov (United States)

    Ameri, Cinzia; Fiorini, Fulvio

    2014-01-01

    Marketing puts its foundation on a few key concepts: need-demand, product-service, satisfaction, exchange, market, or business structure manufacturing / supply. The combination of these elements allows you to build an effective marketing strategy. Crucial in this respect is to remember the Porter matrix, which shows that for a correct analysis of the relevant market is necessary to refer to the "five forces at play", ie: customers, competitors, new entrants and substitutes threat. Another key lever for proper marketing oriented approach is the continuous and constant monitoring of the application, anticipating their dissatisfactions.

  13. Elements of Architecture

    DEFF Research Database (Denmark)

    Elements of Architecture explores new ways of engaging architecture in archaeology. It conceives of architecture both as the physical evidence of past societies and as existing beyond the physical environment, considering how people in the past have not just dwelled in buildings but have existed...... and affective impacts, of these material remains. The contributions in this volume investigate the way time, performance and movement, both physically and emotionally, are central aspects of understanding architectural assemblages. It is a book about the constellations of people, places and things that emerge...

  14. Ring-laser gyroscope system using dispersive element(s)

    Science.gov (United States)

    Smith, David D. (Inventor)

    2010-01-01

    A ring-laser gyroscope system includes a ring-laser gyroscope (RLG) and at least one dispersive element optically coupled to the RLG's ring-shaped optical path. Each dispersive element has a resonant frequency that is approximately equal to the RLG's lasing frequency. A group index of refraction defined collectively by the dispersive element(s) has (i) a real portion that is greater than zero and less than one, and (ii) an imaginary portion that is less than zero.

  15. A novel begomovirus isolated from sida contains putative cis- and trans-acting replication specificity determinants that have evolved independently in several geographical lineages.

    Science.gov (United States)

    Mauricio-Castillo, J A; Torres-Herrera, S I; Cárdenas-Conejo, Y; Pastor-Palacios, G; Méndez-Lozano, J; Argüello-Astorga, G R

    2014-09-01

    A novel begomovirus isolated from a Sida rhombifolia plant collected in Sinaloa, Mexico, was characterized. The genomic components of sida mosaic Sinaloa virus (SiMSinV) shared highest sequence identity with DNA-A and DNA-B components of chino del tomate virus (CdTV), suggesting a vertical evolutionary relationship between these viruses. However, recombination analysis indicated that a short segment of SiMSinV DNA-A encompassing the plus-strand replication origin and the 5´-proximal 43 codons of the Rep gene was derived from tomato mottle Taino virus (ToMoTV). Accordingly, the putative cis- and trans-acting replication specificity determinants of SiMSinV were identical to those of ToMoTV but differed from those of CdTV. Modeling of the SiMSinV and CdTV Rep proteins revealed significant differences in the region comprising the small β1/β5 sheet element, where five putative DNA-binding specificity determinants (SPDs) of Rep (i.e., amino acid residues 5, 8, 10, 69 and 71) were previously identified. Computer-assisted searches of public databases led to identification of 33 begomoviruses from three continents encoding proteins with SPDs identical to those of the Rep encoded by SiMSinV. Sequence analysis of the replication origins demonstrated that all 33 begomoviruses harbor potential Rep-binding sites identical to those of SiMSinV. These data support the hypothesis that the Rep β1/β5 sheet region determines specificity of this protein for DNA replication origin sequences.

  16. Reconstruction of putative DNA virus from endogenous rice tungro bacilliform virus-like sequences in the rice genome: implications for integration and evolution

    Directory of Open Access Journals (Sweden)

    Kishima Yuji

    2004-10-01

    Full Text Available Abstract Background Plant genomes contain various kinds of repetitive sequences such as transposable elements, microsatellites, tandem repeats and virus-like sequences. Most of them, with the exception of virus-like sequences, do not allow us to trace their origins nor to follow the process of their integration into the host genome. Recent discoveries of virus-like sequences in plant genomes led us to set the objective of elucidating the origin of the repetitive sequences. Endogenous rice tungro bacilliform virus (RTBV-like sequences (ERTBVs have been found throughout the rice genome. Here, we reconstructed putative virus structures from RTBV-like sequences in the rice genome and characterized to understand evolutionary implication, integration manner and involvements of endogenous virus segments in the corresponding disease response. Results We have collected ERTBVs from the rice genomes. They contain rearranged structures and no intact ORFs. The identified ERTBV segments were shown to be phylogenetically divided into three clusters. For each phylogenetic cluster, we were able to make a consensus alignment for a circular virus-like structure carrying two complete ORFs. Comparisons of DNA and amino acid sequences suggested the closely relationship between ERTBV and RTBV. The Oryza AA-genome species vary in the ERTBV copy number. The species carrying low-copy-number of ERTBV segments have been reported to be extremely susceptible to RTBV. The DNA methylation state of the ERTBV sequences was correlated with their copy number in the genome. Conclusions These ERTBV segments are unlikely to have functional potential as a virus. However, these sequences facilitate to establish putative virus that provided information underlying virus integration and evolutionary relationship with existing virus. Comparison of ERTBV among the Oryza AA-genome species allowed us to speculate a possible role of endogenous virus segments against its related disease.

  17. Novel non-autonomous transposable elements onWchromosome of the silkworm, Bombyx mori

    Indian Academy of Sciences (India)

    Hiroaki Abe; Tsuguru Fujii; Toru Shimada; Kazuei Mita

    2010-09-01

    The sex chromosomes of the silkworm Bombyx mori are designated ZW(XY) for females and ZZ (XX) for males. Numerous long terminal repeat (LTR) and non-LTR retrotransposons, retroposons and DNA transposons have accumulated as strata on the W chromosome. However, there are nucleotide sequences that do not show the characteristics of typical transposable elements on the W chromosome. To analyse these uncharacterized nucleotide sequences on the W chromosome, we used whole-genome shotgun (WGS) data and assembled data that was obtained using male genome DNA. Through these analyses, we found that almost all of these uncharacterized sequences were non-autonomous transposable elements that do not fit into the conventional classification. It is notable that some of these transposable elements contained the Bombyx short interspersed element (Bm1) sequences in the elements. We designated them as secondary-Bm1 transposable elements (SBTEs). Because putative ancestral SBTE nucleotide sequences without Bm1 do not occur in theWGS data, we suggest that the Bm1 sequences of SBTEs are not carried on each element merely as a package but are components of each element. Therefore, we confirmed that SBTEs should be classified as a new group of transposable elements.

  18. Frequent gain and loss of functional transcription factor binding sites.

    Directory of Open Access Journals (Sweden)

    Scott W Doniger

    2007-05-01

    Full Text Available Cis-regulatory sequences are not always conserved across species. Divergence within cis-regulatory sequences may result from the evolution of species-specific patterns of gene expression or the flexible nature of the cis-regulatory code. The identification of functional divergence in cis-regulatory sequences is therefore important for both understanding the role of gene regulation in evolution and annotating regulatory elements. We have developed an evolutionary model to detect the loss of constraint on individual transcription factor binding sites (TFBSs. We find that a significant fraction of functionally constrained binding sites have been lost in a lineage-specific manner among three closely related yeast species. Binding site loss has previously been explained by turnover, where the concurrent gain and loss of a binding site maintains gene regulation. We estimate that nearly half of all loss events cannot be explained by binding site turnover. Recreating the mutations that led to binding site loss confirms that these sequence changes affect gene expression in some cases. We also estimate that there is a high rate of binding site gain, as more than half of experimentally identified S. cerevisiae binding sites are not conserved across species. The frequent gain and loss of TFBSs implies that cis-regulatory sequences are labile and, in the absence of turnover, may contribute to species-specific patterns of gene expression.

  19. NIH Common Data Elements Repository

    Data.gov (United States)

    U.S. Department of Health & Human Services — The NIH Common Data Elements (CDE) Repository has been designed to provide access to structured human and machine-readable definitions of data elements that have...

  20. An efficient rectangular plate element

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    A new 12-parameter rectangular plate element is presented by useof the double set parameter method. The error in the energy norm is of order O(h2), one order higher than the commonly used Adini nonconforming element.